Protocol. This trial protocol has been provided by the authors to give readers additional information about their work.

Transcription

1 Protocol This trial protocol has been provided by the authors to give readers additional information about their work. Protocol for: Cardoso F, van t Veer LJ, Bogaerts J, et al. 70-Gene signature as an aid to treatment decisions in early-stage breast cancer. N Engl J Med 2016;375: DOI: /NEJMoa

2 This supplement contains the following items: Original protocol (including statistical analysis plan) Final protocol Final statistical analysis plan Summaries of changes (including changes in the statistical plan)

3 BIG 3-04 Intergroup Study (EudraCT number ) (Microarray In Node-negative Disease may Avoid Chemotherapy): A prospective, randomized study comparing the 70-gene signature with the common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in nodenegative breast cancer Coordinating Group: EORTC Breast Cancer Group Collaborative Groups: TRANSBIG Consortium and BIG Groups Study Coordinator: Emiel Rutgers Vice-coordinators: Fatima Cardoso and Martine Piccart Warning: This is an Intergroup study coordinated by the EORTC. The present protocol is written according to the EORTC template and is fully applicable to all participants. The PIS & IC (Appendix F, Appendix G, and Appendix H) are templates to be adapted by National Coordinating Centers / Groups to National and local requirements. Contact addresses for National Coordinating Centers / Groups are provided in the administrative chapter 21. May 10, 2005 PRC outline approval January 23, 2006 Full protocol approval Version 1.0 Version 1.0 / 23 January, 2006 Copyright EORTC 2005

4 Contacts Study coordinators: EORTC (coordinating group): Emiel Rutgers, Fatima Cardoso and Martine Piccart Breast Cancer Group Steering Committee: Core Composition Representatives from EORTC, TRANSBIG, IDDI, NKI, Agendia, IGR, FNCLCC, NCC/Gs and patient s representative organizations. Individuals who may be reappointed Representatives from high recruiting groups e.g. NL, FR, IE, SP & UK Writing Committee: J. Bogaerts, EORTC Data Center, Brussels, Belgium M. Buyse, IDDI, Brussels, Belgium F. Cardoso, Institut Jules Bordet, Brussels, Belgium J. Harrison, EORTC Data Center, Brussels, Belgium M. Piccart, Institut Jules Bordet, Brussels, Belgium E. Rutgers, NKI, Amsterdam, The Netherlands P. Therasse, EORTC Data Center, Brussels, Belgium L. van t Veer, NKI, Amsterdam, The Netherlands Clinical fellows: J. Bines, Institut Jules Bordet, Brussels, Belgium S. Braga, Institut Jules Bordet, Brussels, Belgium G. Demonty, EORTC Data Center, Brussels, Belgium S. Loi, Institut Jules Bordet, Brussels, Belgium Version / January, 2006

5 Table of contents: Protocol summary 9 Trial organization 15 1 Background and introduction General introduction Risk assessment in node-negative breast cancer Node-negative early breast cancer: who can be safely spared adjuvant chemotherapy? Limitations of current prognostic factors Tumor size: women with small (<1cm) node-negative cancers (T1a,b,N0) Histological grade Molecular prognostic markers in breast cancer High throughput gene expression profiling and the development of a multi-gene prognostic signature for early breast cancer The Amsterdam 70-gene prognostic signature Independent validation of the 70-gene signature by the TRANSBIG research consortium The trial: The first trial to evaluate the clinical value of a genomic tool Chemotherapy in the trial: can anthracycline-based chemotherapy be safely replaced by the combination of docetaxel-capecitabine as adjuvant treatment of node-negative breast cancer? Adjuvant Chemotherapy Anthracyclines Taxanes The Combination Docetaxel-Capecitabine Chemotherapy in the trial Endocrine therapy in the trial: is upfront Aromatase Inhibitor (AI) letrozole for 7 years safer and more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? Adjuvant endocrine treatment in breast cancer Use of Aromatase Inhibitors in the adjuvant setting Tolerability profiles Hot flushes and general tolerability Skeletal effects Thromboembolic events and endometrial cancers Lipid metabolism Improving adjuvant therapy in the first 5 years The sequential approach 31 Version / January, 2006

6 1.4.5 Extending adjuvant therapy beyond 5 years Which strategy is best? 32 2 Objectives of the trial Primary objectives Secondary objectives Endpoints Treatment decision randomization (R-T) Chemotherapy randomization (R-C) Endocrine therapy randomization (R-E) 35 3 Patient selection criteria General eligibility criteria for inclusion in the trial Selection criteria for treatment decision randomization (R-T) Selection criteria for chemotherapy randomization (R-C) Selection criteria for endocrine therapy randomization (R-E) 37 4 Trial Design Screening Enrollment Treatment decision randomization for chemotherapy allocation Chemotherapy randomization trial design Endocrine therapy randomization trial design pilot study 41 5 Guidelines for breast cancer surgery and adjuvant radiotherapy Breast cancer surgery Adjuvant radiotherapy 43 6 Chemotherapy: therapeutic regimens, expected toxicity, dose modifications Treatment plan, anthracycline arm Drug information, anthracycline arm Drug administration, anthracycline arm Premedication, anthracycline arm Patient monitoring anthracycline arm Dose adjustments anthracycline arm Treatment plan docetaxel-capecitabine arm Drug information docetaxel-capecitabine arm Docetaxel Capecitabine Docetaxel-capecitabine combination Drug administration docetaxel-capecitabine arm 51 Version / January, 2006

7 6.2.3 Premedication docetaxel-capecitabine arm Patient monitoring docetaxel-capecitabine arm Dose adjustments docetaxel-capecitabine arm Concomitant therapy, docetaxel-capecitabine arm Treatment withdrawal criteria 60 7 Endocrine therapy Trial treatments Side effects of drugs Ovarian function suppression Concomitant treatments 62 8 Clinical evaluation, laboratory tests and follow-up Baseline evaluations Specifically to be done prior to chemotherapy randomization (R-C) Specifically to be done prior to endocrine randomization (R-E) During treatment Chemotherapy Endocrine therapy After the end of treatment (Follow-up) Follow-up for patients receiving neither chemotherapy nor endocrine therapy Chemotherapy follow-up For patients receiving chemotherapy and endocrine therapy For patients receiving only chemotherapy Endocrine therapy follow-up Summary table 66 9 Criteria of evaluation Time to event endpoints Types of recurrence Distant Metastasis Free Survival (DMFS) Disease Free Survival (DFS) Overall Survival (OS) Reporting of results Evaluation of toxicity Reporting of and general evaluation of adverse events Long-term chemotherapy toxicity Serious adverse events Statistical considerations Statistical design 69 Version / January, 2006

8 Sample size Randomization and stratifications Statistical analysis plan Primary and secondary endpoints Analysis populations Timing of analyses Statistical methods Pre-planned sensitivity or exploratory analyses Prognostic factor analyses Data recoding and display End of study Stopping rules and interim analyses Pilot study of the trial Stopping rule after 800 patients enrolled Stopping rule to be applied every year Interim analyses for the chemotherapy and the endocrine questions Data monitoring Translational research Sample collection Central pathology review Proteomics Complex microarrays Biological materials bank storage Publication policy Investigator authorization procedure Patient enrollment procedure Enrollment procedure Access to the interactive randomization program Overview of patient screening and enrollment Patient registration for screening Patient enrollment and randomization for treatment decision making tool (R-T) Randomization of chemotherapy (R-C) Randomization of endocrine therapy (R-E) Forms and procedures for collecting data Electronic case report forms and schedule for completion Data Flow (Remote Data Capture) Reporting of Serious Adverse Events 89 Version / January, 2006

9 17.1 Definitions Reporting procedure Non- serious adverse events and/or non-serious adverse drug reactions Serious adverse events or serious adverse drug reactions Quality assurance Control of data consistency Audits Ethical considerations Patient protection Subject identification Informed consent Sample schedule and trial logistics Frozen tumor sample schedule Paraffin block schedule Serum sample schedule Administrative responsibilities The study coordinator The EORTC Data Center The TRANSBIG Secretariat The EORTC Group National Coordinating Centers/Groups Trial sponsorship and financing Trial insurance 101 Version / January, 2006

10 Table of appendices: Appendix A: References 102 Appendix B: WHO performance status scale 109 Appendix C : Calculation of the glomerular filtration rate (GFR) 110 Appendix D: Common Terminology Criteria for Adverse Events 111 Appendix E: Screening patient information sheet and informed consent document. 112 Appendix F: Patient information sheet and informed consent document for participation in the clinical trial 117 Appendix G: Patient information sheet and informed consent document for the clinical trial chemotherapy section. 127 Appendix H: Patient information sheet and informed consent document for the clinical trial endocrine therapy section. 135 Appendix I: Capecitabine schedule and interruptions to schedule 142 Appendix J: Abbreviations 143 Version / January, 2006

11 Protocol summary Title of the Study EORTC Protocol BIG 3-04 (Microarray In Node-negative Disease may Avoid ChemoTherapy) A prospective, randomized study comparing the Amsterdam 70-gene expression signature (70-gene signature) with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer. Objective(s) PRIMARY 1. The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to node-negative breast cancer patients. 2. The trial will also aim to compare a docetaxel-capecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens. 3. The last primary objective will be to determine the best endocrine treatment strategy i.e. single agent up-front, aromatase inhibitor (AI) (letrozole) for seven years compared to the sequential endocrine strategy of 2 years of tamoxifen followed by 5 years of letrozole. SECONDARY Comparison of the two prognostic methods: To estimate both relative (Hazard Ratio, HR) and absolute (as % at 5 years) efficacy in terms of disease free survival (DFS), distant metastasis free survival (DMFS) and overall survival (OS) of chemotherapy in the two subgroups where the clinical-pathological prognosis and the 70-gene signature molecular prognosis are discordant. To calculate overall estimates of efficacy (DFS, DMFS, OS) for each treatment strategy according to clinical-pathological prognosis and according to molecular prognosis. To estimate the percentage of patients receiving chemotherapy per each prognostic method. Gene profiling: To identify and/or validate predictive gene expression profiles of clinical response/resistance to anthracycline-based and docetaxelcapecitabine chemotherapy. The identification and validation of novel gene expression signatures predicting clinical response to sequential tamoxifen-ai versus AI alone. Chemotherapy: To compare the OS distributions associated with the 2 chemotherapy regimens To determine and compare the early and late toxicities associated with Version / January, 2006

12 each regimen. Endocrine therapy: To evaluate adjuvant endocrine treatment success or failure in the subgroup of postmenopausal patients with endocrine responsive disease. To compare the safety profile of the two endocrine therapy regimens. Long term research objectives: To set up several biological material bank resources (RNA, tumor tissue, serum) for future translational studies in both genomics and proteomics. Methodology Randomization for treatment decision making tool (R-T) of discordant cases (clinical-pathological prognosis using Adjuvant! Online vs. gene expression prognosis using the 70-gene signature), with additional randomizations for chemotherapy (R-C) and endocrine therapy (R-E) comparisons. Number of patients Enrollment of 6,000 patients is planned with those having discordant risk assessments (an estimated 1920 patients) being randomized for treatment decision making. 4,000 patients are planned for the chemotherapy randomization and 3,500 patients planned for the endocrine therapy Diagnosis and main criteria for inclusion randomization. women with histologically proven operable invasive breast cancer and a negative sentinel node or negative axillary clearance (N0, M0). acceptable treatment options are: breast conserving surgery or mastectomy with either a sentinel node procedure or full axillary clearance. radiotherapy is mandatory in the case of breast conserving surgery and will be administered according to local institutional policy after mastectomy. patients with unresectable positive deep margins who receive adjuvant radiotherapy are eligible provided that all other margins are negative. with a clinical tumor classification of T1, T2 or operable T3. the breast tumor must be unilateral, however DCIS or LCIS are allowed if invasive cancer is present. a frozen tumor sample (not fixed in formalin) must be available for inclusion. The tumor sample must be taken from the excised primary tumor (a core punch biopsy). patients should be aged between 18 and 70 years at randomization. (Elderly patients for whom adjuvant treatment is considered could be offered participation in one of the BIG Elderly trials.) have a WHO performance status of 0 or 1. have adequate bone marrow reserves (neutrophil count >1.5 x10 9 /l and platelet count >100 x10 9 /l), adequate renal function (creatinine clearance 50 ml/min (calculated according to Cockroft and Gault, see Appendix C), or serum creatinine 1.5 x upper limit of normal), hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline Version / January, 2006

13 Treatment phosphatase 2.5 x upper limit of normal, total bilirubin 2.0 x upper limit of normal) and normal ECG compatible with chemotherapy administration. have NO serious cardiac illness or medical condition including but not confined to: a history of documented congestive heart failure (CHF), high-risk uncontrolled arrhythmias, angina pectoris requiring antianginal medication, clinically significant valvular heart disease, evidence of transmural infarction on ECG, poorly controlled hypertension (e.g. systolic > 180 mm Hg or diastolic > 100 mm Hg), symptomatic coronary artery disease or a myocardial infarction within the last 12 months or other risk factors that contra-indicate the use of anthracycline-based chemotherapy. NOT have serious uncontrolled intercurrent infections, or other serious uncontrolled concomitant disease. NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any cancer (other than breast cancer) in complete remission for 5 years. NOT have received previous chemotherapy, hormonal therapy or radiotherapy. NOT have participated in any investigational drug study within the 4 weeks preceding the start of treatment. NOT be pregnant or breast-feeding at the time of diagnosis or randomization. A woman of childbearing potential must have a negative pregnancy test. If post-menopausal, the woman must have been amenorrheic for at lest 12 months to be considered of non-childbearing potential while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, injectables, combined oral contraceptives, some IUDs, sexual abstinence or vasectomised partner (Note 3 of the guidance on nonclinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). NOT have any psychological, familial, sociological, geographical or serious uncontrolled medical (e.g. a history of uncontrolled seizures, central nervous system disorders, or psychiatric disability) condition judged by the investigator to be clinically significant which could potentially preclude informed consent or interfere with compliance for oral drug intake or with the study protocol and follow-up schedule. These conditions should be discussed with the patient before enrollment in the trial. signed written informed consent must be given according to ICH GCP, and national/local regulations before enrollment in the trial (both a screening PIS & IC and the PIS & IC 1 must be signed before enrollment). The following women, if eligible, will be offered the chemotherapy randomization (R-C) comparing a commonly used anthracycline-based Version / January, 2006

14 Test product, dose and mode of administration Duration of treatment Reference therapy, dose and mode of administration Criteria for evaluation Efficacy endpoints Safety Statistical methods regimen, selected from the list of acceptable regimens, and the new capecitabine-docetaxel combination: 1. Women that have a high risk according to the clinical-pathological criteria and a high risk according to the 70-gene signature prognosis. 2. Women that have a high risk according to the clinical-pathological criteria and a low risk according to the 70-gene signature, and were randomized (R-T) for chemotherapy treatment decision to use the clinicalpathological criteria. 3. Women that have a low risk according to the clinical-pathological criteria and a high risk according to the 70-gene signature, and were randomized (R-T) to use the 70-gene signature prognosis for chemotherapy decision. Trastuzumab will be allowed and recommended (however not provided free as a trial drug) for women with HER-2 positive tumors if it becomes approved and available for use in the adjuvant setting during the trial. Treatment with trastuzumab is recommended to start after the end of chemotherapy. All postmenopausal endocrine-responsive (i.e. ER and/or PgR positive) patients on the study will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. It will be left to the decision of the investigator whether or not a premenopausal woman can be randomized into this question, with concomitant ovarian function suppression while taking letrozole. Chemotherapy will last for approximately 6 months and endocrine therapy for a maximum of 7 years. For the treatment decision randomization (R-T) the standard arm is risk assessment using clinical-pathological criteria (Adjuvant! Online). For the chemotherapy randomization (R-C) the standard arm is an anthracycline based chemotherapy regimen chosen from the list of allowed regimens. For the endocrine therapy randomization (R-E) the standard arm is sequential 2 years of tamoxifen followed by 5 years of letrozole. For R-T the primary endpoint is DMFS at 5 years. Secondary endpoints are: the proportion of women treated with chemotherapy per treatment decision making tool i.e. clinical prognosis compared to the 70-gene signature prognosis, OS at 5 and 10 years, DFS and safety (both early and late). For R-C and R-E the primary endpoint is DFS at 5 years. Toxicities for R-C and R-E will be recorded and summarized according to the Common Terminology Criteria for Adverse Events v3.0 (CTCAE). Additionally, cardiac toxicity and secondary leukemia, as potential longterm toxicities of chemotherapy will be monitored for a minimum of 15 years in all patients who were randomized for chemotherapy (R-C). Eligible patients with adequate material for microarray analysis and a successful 70-gene signature prognostic test will be enrolled. Risk will be determined using both the 70-gene signature and clinical-pathological criteria. Patients are evaluated as low clinical-pathological risk, if their 10- year disease specific survival (without chemotherapy or endocrine therapy) is estimated by Adjuvant! Online as greater than 88% for endocrine receptor (ER) positive patients, and greater than 92% for ER negative patients. Patients who are high risk for both methods will receive chemotherapy, and patients who are low risk for both methods will not receive chemotherapy. Patients for whom the methods are discordant will Version / January, 2006

15 Translational research Quality of Life Cost effectiveness/ health economic studies be randomized for treatment decision making (R-T) between chemotherapy-decision-making according to clinical-pathological criteria or chemotherapy-decision-making according to the 70-gene signature prognosis. Patients who are candidates for chemotherapy (see also treatment section) will be proposed for randomization R-C: between anthracycline based chemotherapy and docetaxel-capecitabine chemotherapy. All patients who are candidates for endocrine therapy will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. Primary test: In the set of patients who have a low risk 70-gene signature prognosis and high risk clinical-pathological prognosis, and who were randomized (R-T) to use the 70-gene signature prognosis and thus to receive no chemotherapy, a null hypothesis of a 5-year DMFS of 92% will be tested. With 6,000 patients accrued overall, this set has an expected size of 672 patients. With an accrual of 3 years, and a total duration of 6 years (so 3 to 6 years follow up for each patient), a one-sided test at 97.5% confidence level has 80% power to reject this hypothesis if the true 5-year DMFS is 95%. Test of hypothesis for R-T: - In the subgroup of clinical-pathological high risk and 70-gene signature prognosis low risk patients, assuming a 5-year DMFS of 93% without chemotherapy, with an expected size of 1,344 patients, there is 80% power to detect a hazard ratio of 0.47 (or a 5-year DMFS of 96.6% on chemotherapy) at the time of the primary analysis. - The subgroup of clinical-pathological low risk and 70-gene signature prognosis high risk, with an expected size of 576 patients, has low power to detect any realistic chemotherapy effect. Test of hypothesis for R-C: Assuming a control 5-year DFS of 86%, with an expected 4,000 patients in this section of the study, there is 80% power to detect a hazard ratio of 0.76 (or 89% experimental 5-year DFS) at the time of the primary analysis. Test of hypothesis for R-E: With an expected 3,500 patients being randomized into this question, and assuming a DFS rate at 5 years of 86% on the control arm, there is 80% power to detect a hazard ratio of 0.75 at the time of the primary analysis (i.e. a 5-year DFS of 86% vs. 89.3%). Objectives: See primary and secondary objectives. Biological material will be stored for the use in future translational research projects (see rationale & chapter 12). Whole genome complex arrays will be performed for all patients providing a unique opportunity to investigate the relationship between gene expression patterns and disease prognosis and response to treatment. No studies proposed Various sub studies are proposed. Version / January, 2006

16 Delay between surgery and start of treatment ideally not more than 6 weeks, can not be > 8 weeks Screening Visit surgeon (T1, T2 or operable T3 & cn0 & M0) screening PIS & IC, Screening Surgery Tumor sample shipment, RNA extraction, microarray analysis, local pathology, TNM, lymph node status, HR status pn0 Signing of PIS & IC 1 & Enrollment Genomic/Clinical Prognosis review & randomization Discordant Low/Low High/High R-T No CT CT R-C HR - HR - HR + HR + R-E CT Chemotherapy R-T Treatment decision Randomization based on genomic vs clinical prognosis R-C Chemotherapy Randomization of Anthracycline based CT vs Docetaxel/ Capecitabine R-E Endocrine treatment Randomization Letrozole vs Tamoxifen followed by Letrozole HR Hormone receptor PIS & IC Patient information sheet and informed consent For details of timelines and trial logistics please see the Timelines from diagnosis to start of treatment diagram in Chapter 20. Version / January, 2006

17 Trial organization This trial is an Intergroup trial, jointly conducted by several National and International cancer clinical research groups throughout the European Union and the rest of the world under the Breast International Group (BIG) / TRANSBIG umbrella. The EORTC is the coordinating group of this trial and will be the coordinating Data Center. It will centrally manage trial design and center activation, attribution of duties and responsibilities between participating research groups, data collection and quality control of data, statistical analysis and publication. Globally the EORTC will be the Sponsor. Some particular arrangements will be made, exceptionally, due to logistical or legal reasons and these will be communicated to investigators in these countries by their National Coordinating Center/Group (NCC/G). The EORTC will cover insurance in all countries except American and Australian continents. Precise details about insurance and an insurance certificate will be provided to Investigators at site initiation by their NCC/G. NCC/Gs will locally coordinate the conduct of the trial in their respective countries; one of their most important duties will be to manage the notification/ submission of all necessary documents to the Competent Authorities and to obtain the confirmation of the review by Institutional Review Board/local Ethics Committee if applicable following the national law. This protocol is to be followed by all participants. All chapters are fully applicable to all participants. All countries will conform to a standard administrative model where each center will contact their NCC/G-national structure who will provide them with all practical information. Participation in this trial is only possible through a national structure (coordinating group/center, NCC/G or EORTC Data Center) designated for this purpose. For investigators who do not find contact details for a NCC/G in the Administrative Responsibilities chapter (Chapter 21), please contact the EORTC Data Center directly and we will communicate the appropriate NCC/G contact details to you. The patient information sheets and informed consent templates (Appendix F, Appendix G, Appendix H) are provided as a basis for translation/adaptation by NCC/Gs to national/local regulations and sensibilities. Investigators will receive the translated and adapted PIS & ICs directly from their NCC/G or national structure. For the screening phase an example of possible text for the screening PIS & IC (spis & IC) is also provided however although investigators will have to certify the aptness of the PIS & IC to cover the genomic test, this consent is the responsibility of each individual center as it is signed prior to registration for screening. Investigators who are members of several groups participating in the trial should participate through the national structure detailed in the Administrative Responsibilities chapter and include all patients through this national structure. For EORTC members all patients will be counted for membership accrual independently of the group they participate through (see EORTC Policy 10). The investigational drugs (Docetaxel, Capecitabine and Letrozole) will be supplied by the industrial partners. This trial is an academic trial with partial support from the European Commission Framework Programme VI (FP6-LSHC-CT ), industrial partners, the US NCI and charitable grants. Version / January, 2006

18 1 Background and introduction 1.1 General introduction Breast cancer is the most common malignant disease in women, occurring more frequently in developed countries but showing a rising incidence in developing countries. Breast cancer is a public health issue on a global scale. According to estimates in 2002, there were 1,151,298 new cases of breast cancer diagnosed, 410,712 deaths caused by breast cancer and more than 4.4 million women living with breast cancer world wide (Ref. 1). Breast cancer is the second most commonly diagnosed cancer in the European Union (EU). An estimated 245,000 women were diagnosed with breast cancer in the EU in 2000, representing about a quarter of all female cancers diagnosed. Breast cancer is also the most common cause of cancer death in European women with 90,000 deaths in 2000 (Ref. 2-4). In 2004, in all of Europe, there were 371,000 estimated new cases of breast cancer diagnosed and 129,900 breast-cancer-related deaths (Ref. 5). At diagnosis, 90% of patients appear to have operable breast cancer, that is, disease that is confined to the breast and ipsilateral axilla. However up to 50% of these women will develop metastatic disease. Metastatic breast cancer is unfortunately incurable. As a result, since the mid 1980 s randomized trials of adjuvant systemic therapy (both endocrine and cytotoxic) have been conducted in an effort to reduce the rate of recurrence and to prolong the survival of patients with operable disease. Surgery is the main modality of treatment in patients with early breast cancer and, by itself or together with radiotherapy can control local-regional disease in the majority of patients. In spite of these treatments, subclinical tumor cells remaining following surgery may be responsible for the progression/recurrence of the disease at a later date. Adjuvant hormonal or cytotoxic treatments have been increasingly used with the goal of eradicating these remaining foci, in order to achieve longer survival and the eventual cure of the disease. The utilization of systemic treatment following surgery has become the standard of care and current objectives are to better target, more effective and less toxic treatments for breast cancer and to identify those patients who will have a clear benefit from these treatments. Current standard of care for those patients who will receive chemotherapy is 6 cycles of anthracycline based chemotherapy with some experts favoring the incorporation of taxanes for node-positive disease. For patients with endocrine responsive disease, the standard endocrine care consists of 5 years of tamoxifen, with many experts favoring the use of an aromatase inhibitor in sequence with or in replacement of tamoxifen. In the last 20 years, little progress has been made with respect to assisting oncologists in determining which node-negative breast cancer patients require chemotherapy or other systemic therapy and which women can safely be treated with only loco-regional treatment. Great discomfort still persists in selecting those node-negative women who need adjuvant chemotherapy (Ref. 6). 1.2 Risk assessment in node-negative breast cancer Mortality rates from breast cancer rose until 1990 but fell afterwards in a number of European countries. Reasons for this decline include widespread mammographic screening, precise diagnosis and the extensive use of tamoxifen. Due to the widespread adoption of screening mammography in women of 50 years and older in developed countries, a growing proportion of women are being diagnosed with smaller tumors and no axillary node involvement. Although many of these women can enjoy long-term survival, 20-30% will relapse and die from their disease. Distant metastases account for the majority of these deaths and have been the impetus for proposing, in addition to optimal loco-regional treatment (surgery and radiotherapy), adjuvant systemic therapy to all fit Version / January, 2006

19 women considered to be at moderate or high risk of relapse. There is however, much controversy related to the optimal definition of a low/minimal versus a moderate/high risk of relapse for these women with node-negative breast cancer. Therefore, many oncologists rely on the guidelines issued by experts following consensus conferences, such as the 2000 National Institute of Health (NIH) meeting and the 2005 St. Gallen conference (Ref. 7-9). Given the close to 100% death rate from metastatic breast cancer, these guidelines are produced with the goal of avoiding under-treatment of affected women, and they assign only 15% to 20% of them to a low/minimal risk subset for which no adjuvant treatment at all or only adjuvant endocrine treatment will be considered. Nowadays, the great majority of early stage breast cancer patients are therefore offered some type of systemic chemotherapy. Unfortunately, whilst the majority of adjuvant chemotherapy regimens commonly used today are rarely life-threatening, they can have considerable both short and long term morbidity. As a result, many women with early breast cancer in the western world are probably over-treated, a phenomenon that may not only decrease the quality of life of these women but also increases the economic burden of this frequent disease on society. Furthermore, many advances in supportive care in recent years have facilitated the administration of adjuvant chemotherapy regimens to a greater number of patients, while also increasing its costs. Despite nodal status being one of the strongest prognostic factors used for predicting breast cancer clinical outcome, the Early Breast Cancer Trialist Collaborative Group (EBCTCG) overview recently reported an almost equal proportional reduction in recurrence and breast cancer death produced by adjuvant chemotherapy in node-positive and node-negative women less than 50 years old, and only a small difference in women older than 50 years (Ref. 10). Therefore, whereas both tumor size and nodal status are of prognostic significance, neither has substantial influence on the relative benefits of systemic therapy. However, these proportional benefits translate into larger absolute benefits for higher risk patients with node-positive disease than lower risk patients with node-negative disease. Hence, it is generally accepted that the absolute benefit for the higher risk patients is large enough to warrant the risks of adjuvant chemotherapy. For women who have nodenegative breast cancer, it is important to be able to accurately quantify the risk of recurrence and death so that the decisions about adjuvant chemotherapy can be taken appropriately. For these women, it is crucial to determine if there is a subgroup of breast cancers that, due to their excellent long term survival, can be spared adjuvant chemotherapy from which they are unlikely to benefit Node-negative early breast cancer: who can be safely spared adjuvant chemotherapy? Unfortunately little progress has been made with regards to new prognostic markers that can assist oncologists in treatment decision-making for node-negative early stage breast cancer. As a result, considerable differences exist worldwide in the selection of women whose risk of breast cancer recurrence justifies the need for adjuvant chemotherapy. Current breast cancer TNM staging is based on anatomical extent (size, lymph node status) but this classification gives little insight into breast cancer biology. Clinicians have long recognized the heterogeneity of human breast cancers, not only in terms of their diverse natural histories despite identical morphological features, but also in their variation in treatment response. Attempts have been made to identify good and poor prognosis groups based on pathological features such as tumor grade, lymphatic invasion and S- phase fraction (Ref. 11) which may better reflect tumor biology. In recent years there have been numerous reports on molecular prognostic and predictive markers in oncology. These tumor markers have had little impact in routine clinical practice. Often studies are based on small, heterogeneous retrospective series, and have not been reported in a rigorous enough fashion with insufficient information, particularly methodologically, to reproduce results (Ref. 12). Many followup studies have been inconsistent with original results: this has variously been attributed to lack of power, different patient populations and technical limitations. There is also a paucity of well designed, prospective assessments of the clinical value of these tumor markers. As a result, the value of many promising prognostic markers is still uncertain. Version / January, 2006

20 The prognostic factors accepted by the NIH 2000 Consensus Conference on Adjuvant Therapy did not include any molecular markers relevant to breast cancer biology apart from the hormone receptors (Ref. 8). The most recent St Gallen Consensus Panel (2005) (Ref. 9) established three risk categories: minimal, intermediate and high risk. Hormone receptors, tumor size, tumor grade and age remain key discriminating factors, and HER-2 status and lymphatic &/or vascular invasion or both in the primary tumor are newly accepted prognostic factors. In the United Kingdom, the Nottingham Prognostic Index is commonly used to predict clinical outcome; it is based on tumor size, tumor grade and lymph node status, and plays a key role in clinical risk assessment for nodenegative tumors for which chemotherapy may or may not be considered (Ref. 13) Limitations of current prognostic factors Tumor size: women with small (<1cm) node-negative cancers (T1a,b,N0) To date, uncertainty exists about the long term prognosis and treatment of lymph node-negative patients with tumors 1 cm or less. The lack of long term (>15 years) follow-up data has contributed to this ambiguity as many studies do not account for the propensity of breast cancer to relapse after 10 years (Ref. 14;15). Two population-based data bases have reported excellent prognoses for these women. The Breast Cancer Detection Demonstration Project, which included 880 patients with T1N0 tumors, reported a survival rate of 96% at 8 years of follow-up (Ref. 16). Rosen and colleagues reported a 20 year recurrence-free survival of 86% for tumors measuring 1 cm or less (Ref. 17). In contrast, the National Surgical Adjuvant Breast and Bowel Project (NSABP) presented data from women enrolled onto five NSABP trials (B-06, B-13, B-14, B-19 and B-20). There were 235 patients with ER-negative tumors and 1,024 patients with ER-positive tumors 1 cm or less (Ref. 18). For ER-negative tumors, the 8 year relapse free survival for those who had surgery alone or with chemotherapy was 81% and 90% respectively (p=0.06). For the women with ER-positive tumors, these figures were 86% for surgery alone, 93% when tamoxifen was added (p=0.01) and 95% after the addition of both tamoxifen and chemotherapy (p=0.07 when compared with tamoxifen). The authors concluded that chemotherapy and/or tamoxifen should be considered for the treatment of women with tumors of 1 cm or less and negative axillary lymph nodes. A subset of these tumors seems to have poor clinical outcome, despite their small size. In a study of 218 patients with tumors 1 cm or less, poor tumor grade and lymphatic invasion were independent predictors of recurrence. For patients with both these features, the 7-year recurrence free survival rate was 67% compared with 92% observed with one risk factor and 99% observed with neither (p=0.0001) (Ref. 19). A population-based cohort of 430 women with 1 cm or less, node-negative, lymphatic and vascular invasion-negative early breast cancers that had received no systemic adjuvant therapy reported a 10-year relapse free survival of 82%. However, histological grade 3 tumors had a > 25% 10-year risk of relapse and >10% 10-year risk of breast cancer death (Ref. 20). Hence, these studies suggest that tumor size cannot reliably be considered to identify a group of women with excellent long term clinical outcome Histological grade Histological grade is currently not included in the revised American Joint Cancer Committee (AJCC) guidelines on breast cancer TNM staging (Ref. 21). Yet, tumor grade has been recognized since the early 20 th century to provide important prognostic information. The AJCC cited the poor reproducibility of tumor grading due to the variety of approaches used and the inherently subjective nature of tumor grading as the reason for its exclusion from a staging system that needs to be reproducible from institution to institution. Elston and Ellis introduced a new grading system with the aim of making the grading criteria more quantitative (Ref. 22). A modification of the Bloom and Richardson grading system, a numerical score is assigned to three morphologic features and is used in the compilation of an overall grade. This system was named the Nottingham combined Version / January, 2006

21 histological grade. Subsequent studies have shown improved interobserver agreement and it is recommended by the College of American Pathologists. However, whilst the diagnoses of a poor or well differentiated histological grade may influence some treatment decision making, a large proportion of breast cancers are classified as grade II or intermediate grade, for which the prognostic value is not so helpful. As can be seen above, currently used prognostic factors have a poor ability to identify a very good prognostic group within those tumors diagnosed as node-negative Molecular prognostic markers in breast cancer The last ten years have witnessed several attempts to more clearly define groups of women with extremely good prognosis who can be spared adjuvant systemic therapy, or at least adjuvant chemotherapy using molecular markers. The most interesting data in this field have been published in major scientific journals and are summarized in the following table: Table 1 New candidate prognostic tools UPA PAI-1 Cyclin E Gene expression signature Authors Janicke Look Keyomarsi Van de Vijver Reference JNCI, 2001 JNCI, 2002 N Engl J Med, 2002 N Engl. J Med, 2002 Type of study Prospective Metaanalysis Retrospective Retrospective Node-negative (N) 374 3, Pt characteristics median age % ER low % grade 3 % treated with No grading done 50 < adj. therapy Median f.-up (y) Type of assay ELISA ELISA Western Blot RNA Array Minimum amount of tissue 50 mg snap frozen 50 mg snap frozen 100 mg snap frozen 3 µg snap frozen Quality control in multicentre setting Yes Yes No No Prognostic value of tool in multivariate analysis 1) In node-negative Strong, superseded only by tumor grade 2) In node-negative & positive Strong, supersedes all others Very strong, more powerful than nodal status Very strong, more powerful than nodal status High throughput gene expression profiling and the development of a multi-gene prognostic signature for early breast cancer The advent of high throughput array-based technology and the sequencing of the human genome has provided the opportunity to begin comprehensive molecular profiling of cancers. Such efforts have, in recent times given us new insights into breast cancer biology and confirmed that the Version / January, 2006

22 disease is considerably more heterogeneous than can be predicted by traditional histopathological methods. The collective results demonstrate that 1) the estrogen receptor (ER) status of the tumor is the most important discriminator of expression subtypes; 2) breast tumors can be grouped according to at least 3 individual molecular subgroups: basal (almost exclusively ER-negative), HER-2 and luminal (almost exclusively ER-positive) subtypes; and 3) these subgroups seem to have distinct clinical outcomes and may respond differently to various therapies (Ref ). A breast cancer classification system based on molecular expression patterns that defines different breast cancer subtypes, analogous to what exists for lymphoma or leukemia, rather than based on anatomic extent of the disease could emerge and potentially allow clinicians to individualize treatment according to molecular subtype. These results suggest that molecular profiling can substantially improve our understanding of breast cancer biology and therefore improve our management of breast cancer patients The Amsterdam 70-gene prognostic signature New methods of prognostic classification have been developed using array technology. Three years ago, in an attempt to better understand breast cancer biology and to find new clinically relevant molecular makers, the Netherlands Cancer Institute looked at global gene expression levels of 78 tumor samples from untreated node-negative breast cancer patients using oligonucleotide microarray technology. They used a homogenous population from node-negative patients, all under 55 years of age, all treated with loco-regional therapy only, with a median follow-up of 8 years, to derive their prognostic gene signature. In their landmark study, they were able to establish a 70- gene expression profile for the prediction of early distant metastases (recurrence in less than 5 years) (Ref. 28). They then validated the prognostic power of this unique set of genes in 295 patients and confirmed that this gene signature outperformed all the traditional prognostic guidelines, such as those from the NIH and St Gallen, clearly separating a group of patients with an excellent prognosis at 10 years from a group with a high risk of distant recurrence. These results were independent of age, tumor size, number of positive nodes and centrally reviewed histological grade (Ref. 29). Since the publication of these results, a number of authors have underlined critical issues in microarray-derived analysis including the fragility of gene signatures and over-optimistic performance estimation due to model over-fit (Ref ). Gene signatures, promising as they are, still have to make their way to the clinic and are vulnerable to the same problems as single gene tumor markers before them. Furthermore, some authors have expressed doubts that they add prognostic value to that provided by the best risk classifications based on clinical-pathological factors including age, tumor size, tumor grade, nodal involvement and presence of hormonal receptors, though this of course places great faith in the reproducibility of current clinical marker assessments, very few of which undergo critical quality control (Ref. 33). Gene signatures have enormous potential towards better individualization of treatment options in breast cancer therapy if they can reliably identify patients for whom adjuvant chemotherapy is definitely not indicated despite poor clinical risk factors, and patients who need chemotherapy despite good clinical risk factors (Ref. 34). This better selection of patients could avoid both over-treatment, with its potential for severe complications and under-treatment with deleterious consequences for patient survival. Prior to the commencement of the trial, the TRANSBIG research consortium (sister translational research network to the clinical trials network the Breast International Group) undertook an independent and external validation of the Amsterdam-70-gene expression signature (which will be referred to as the 70-gene signature) using prospectively defined criteria, to establish its reproducibility and to confirm its prognostic ability. Version / January, 2006

23 1.2.4 Independent validation of the 70-gene signature by the TRANSBIG research consortium This study was based on frozen archival tumor material of 302 node-negative patients from five non-dutch cancer centers from three different countries (United Kingdom, Sweden, and France). The patients were aged 60 years old or less, were diagnosed before and including 1998 and, as was the case with the Dutch patients, had received only loco-regional therapy. The median follow-up was 13.6 years. Frozen samples were sent to Amsterdam for gene expression profiling using a custom designed chip, which assesses the mrna expression of the 70-genes in triplicate (Mammaprint from Agendia, a spin-off commercial company of the Netherlands Cancer Institute using Agilent technologies). Researchers in Amsterdam had no knowledge of the clinical outcome data. An independent statistical office in Brussels determined the clinical risk groups. Central pathology review of estrogen receptor status and grade could be obtained for nearly 80% of samples. The first results of this validation study were presented during the 2004 San Antonio Breast Cancer Symposium (Ref. 35) and confirmed the previous findings, namely that the 70-gene signature was able to distinguish a significantly decreased time to distant metastases (TDM) and overall survival (OS), outperforming both the Nottingham Prognostic Index, the St-Gallen criteria and the Adjuvant! Online software ( which calculates a 10-year survival probability based on the patient s age, tumor size, grade and estrogen receptor status (Ref. 36). A recent evaluation of the Adjuvant! Online program found that known clinical prognostic factors were able to predict OS, breast cancer-specific survival, and event free survival quite accurately in 4,083 patients diagnosed with breast cancer in British Columbia between 1989 & 1993, with the exception of patients who were diagnosed when they were younger than 35 years (Ref. 37). The 70-gene signature remained a significant prognostic factor for TDM and OS even after adjustment for all clinical-pathological factors known to have prognostic value in this disease. Consensus within the consortium decided that the low clinical risk group consisted of patients with a 10-year survival probability of at least 88% (using Adjuvant! Online) if they were 1% positive for expression of estrogen receptors by immunohistochemistry, and of at least 92% if they were not. These two cut-offs were chosen to reflect the fact that today patients with hormone receptorpositive tumors receive adjuvant endocrine therapy (with an estimated absolute 10-year benefit of about 4% overall), whereas patients in the validation series were all untreated regardless of their hormone receptor status. When adjusted for the clinical risk groups based on 10-year survival probability using the Adjuvant! Online software, the 70-gene signature adjusted hazard ratios were 2.13 (95% CI ) for TDM, 2.66 (95% CI ) for OS, and 1.36 (95% CI ) for DFS. Similar hazard ratios were found in multivariate Cox s regression analysis. These results indicate that the gene signature adds independent prognostic information to that provided by a risk assessment based solely on clinical-pathological factors. Central pathology review of hormone receptor status and tumor grade and an independent source verification of all data by external auditors added significant strength to these findings. Furthermore, when the patients were divided according to their gene signature risk groups, for those patients classified as gene signature low risk, regardless of being classified clinical high or low risk (by Adjuvant! Online), their Kaplan-Meier estimates of 10-year OS were practically identical (88%, 89% respectively), with the same result for the gene signature high risk groups (69% for both). These results reinforced the original evidence and lent strong support to the initiation of the trial. The results of the validation study have been submitted for publication. Since the initial validation, the 70-gene signature has also been validated in a similar population of women aged from years. In summary, the TRANSBIG consortium has completed the first independent and external validation of a gene signature derived from microarray technology. Their analyses confirm that the 70-gene signature adds significant independent prognostic information to that produced by current clinical-pathological factors and provides sufficient evidence to develop the study. The 70-gene signature remains the only prognostic tool derived from microarray technology to have undergone stringent external and independent evaluation. Version / January, 2006

24 1.2.5 The trial: The first trial to evaluate the clinical value of a genomic tool The success of this validation study allowed for the commencement of the large collaborative trial (Microarray In Node-negative Disease may Avoid ChemoTherapy), which will be the first prospective trial to validate a molecular-based signature for the adjuvant treatment of nodenegative early breast cancer patients. The goal of is to prospectively validate the 70- gene signature providing the level-1 evidence of its efficacy that is needed before its wide clinical application. This study aims to give us a definitive answer to the clinical relevance of the 70-gene signature, its performance compared to traditional prognostic factors and its ability to predict response to commonly prescribed adjuvant treatments. will enroll 6,000 patients to investigate the benefit/risk ratio of chemotherapy when the assessment of prognosis based on clinical-pathological features differs from that provided by the gene signature. It will use the 70-gene signature to classify node-negative early stage breast cancer patients into high and low risk of distant relapse and to compare this assessment to the Adjuvant! Online software risk assessment, which is widely used in clinical practice today and which takes into consideration traditional clinical-pathological factors. All 6,000 patients enrolled in the study will have their risk of recurrence assessed by both clinical-pathological and 70-gene signature methods. Those patients for whom both methods classify the patient as high risk will be proposed adjuvant chemotherapy; those patients for whom both methods classify the patient as low risk will not be offered adjuvant chemotherapy, however they will be offered endocrine therapy if their tumors are endocrine responsive. Those patients for whom the two risk assessments are discordant will be randomized for treatment decision making tool to either clinical criteria (using Adjuvant! Online) or genomic prognosis (using the 70-gene signature). Using this new tool, aims to better define patient prognosis and therefore to better select patients who need adjuvant chemotherapy; by doing so it is expected that 10% to 20% of women who would normally receive adjuvant chemotherapy based on their clinical-pathological factors will be spared the inconvenience and morbidity of this therapy, without having any negative impact on their survival. The primary objective of the trial is to confirm that the number of patients that can be spared adjuvant chemotherapy is significantly increased when the decision is based on the gene prognosis signature rather than on the clinical-pathological criteria, while the critical group of patients who have a high risk of recurrence according to the clinical-pathological criteria but a low risk according to the gene prognosis signature is not under-treated. If this is confirmed, it will have immense benefits for women and will markedly improve our ability to predict which patients are likely to experience a distant relapse and who therefore require additional systemic therapy. The economics of using multi-gene assays that are considerably more expensive than currently used prognostic assessments has been appraised for another multi-gene assay derived from high throughput technology (Ref. 38). The Oncotype assay was found to predict more accurately than current guidelines recurrence risk in lymph node-negative, estrogen receptor-positive early-stage breast cancer treated with tamoxifen. Using an economic model that considered survival, quality of life and relevant costs on data from a large multi-centre clinical trial, among a hypothetical cohort of 100 patients, the multi-gene assay was found on average to increase quality-adjusted survival by 8.6 years and reduce overall costs by US$ 202,828, with cost effectiveness most influenced by the propensity to administer less chemotherapy (Ref. 39). Further randomizations, as applicable, will be offered to enrolled patients to answer important questions about adjuvant therapy. For those patients receiving chemotherapy, an optional randomization will compare anthracyline-based regimens to a new docetaxel-capecitabine combination (see section 1.3). A third optional randomization for all patients with endocrine responsive disease will compare 7 years of letrozole to 2 years of tamoxifen followed by 5 years of letrozole (see section 1.4). Version / January, 2006

25 A prospective randomized trial like is essential to determine, in an unbiased way, the true value of gene signatures in clinical practice today. Given the immense therapeutic and emotional repercussions for women, it is essential to know the true meaning of the poor prognosis signature and whether any of our currently prescribed treatments can alter the natural history of this high risk group. Retrospective studies are susceptible to all sorts of bias which may significantly influence results and hence, will be unable to accurately determine these implications. Furthermore, the evaluation of the potential of gene signatures to individualize management, to spare treatment without compromising long term outcome and hence diminish the public health burden is of utmost priority to women and to nations. 1.3 Chemotherapy in the trial: can anthracyclinebased chemotherapy be safely replaced by the combination of docetaxel-capecitabine as adjuvant treatment of node-negative breast cancer? Adjuvant Chemotherapy The Early Breast Cancer Trialists Collaborative Group (EBCTCG) meta-analysis (Ref. 40) showed that combination chemotherapy yields a significant reduction in mortality when compared to no chemotherapy. This benefit occurs in all patients, irrespective of nodal or estrogen-receptor status and the use or not of anti-estrogen therapy (tamoxifen). The benefits of chemotherapy are seen in all age groups analyzed though the absolute benefit is greater for younger women (less than 50 versus 50 to 69 years old) and in node-positive disease as compared to node-negative disease. For women with early-stage breast cancer, adjuvant polychemotherapy reduced the annual risk of death by 30% in women aged 40 to 50 (10% difference in risk of death within 15 years), and by 12% in women aged 50 to 70 (3% difference in risk of death within 15 years). These risk reductions were very similar in node-negative and node-positive patients, but had a larger absolute effect in node-positive patients, because of their higher mortality rate. In node-positive patients, a 37% reduction in the odds of recurrence and a 30% reduction in the odds of death were reported, while for node-negative patients, these numbers were almost the same, being 36% and 29% respectively. The absolute benefit for polychemotherapy versus no adjuvant chemotherapy in women less than 50 years old is twice as great at 15 years (10%) as at 5 years (4.7%) Anthracyclines The overview also highlights an advantage in outcome with the utilization of anthracycline based regimens in the adjuvant setting, when compared to CMF-based regimens, of 16% reduction in the annual risk of death, and 11% reduction in the annual risk of recurrence. These effects seem to be irrespective of age and nodal status. Nevertheless, not all anthracycline regimens can be considered adequate. The most convincing results are derived from treatments that included a three-drug combination, as opposed to a two drug (such as doxorubicin plus cyclophosphamide (AC)). Regimens such as FEC100, CAF/FAC and CEF, for 4 to 6 cycles, are considered the treatment of choice for node-positive and high risk node-negative patients, based on the results of several randomized trials (Ref. 8;41-43) and the EBCTCG Overview. Of concern to women with a medium rather than a high risk of relapse are two possible long-term toxicities of anthracycline-based therapy: secondary leukemia and cardiotoxicity. The cumulative Version / January, 2006

26 risk, at 5 years, of developing anthracycline-related leukemia is around 1% for both doxorubicin and epirubicin and is directly related to the administered dose (Ref. 44;45). Regarding epirubicin, good quality data is available from the Pharmacia Clinical Trials Database, derived from 7,110 patients treated with epirubicin-containing regimens. The cumulative probability of AML/MDS was 0.27% (95% CI, 0.14% to 0.40%) at 3 years, 0.46% (95% CI, 0.28% to 0.65%) at 5 years, and 0.55% (95% CI, 0.33% to 0.78%) at 8 years (Ref. 46). In a recent study, the National Cancer Institute of Canada Clinical Trials Group (NCIC-CTG) analyzed the risk of acute leukemia in 1,463 premenopausal node-positive patients enrolled onto four NCIC-CTG adjuvant therapy trials. The conditional probability at 5 years of developing acute leukemia was 1.5% for all epirubicin-containing regimens, as opposed to 0.3% for non-epirubicin-containing ones (Ref. 47). For doxorubicin, the NSABP group has reviewed the data from all its adjuvant trials that explored the regimens containing doxorubicin given at the fixed dose per cycle of 60 mg/m² and a fixed cumulative dose of 240 mg/m² and cyclophosphamide administered at variable doses per cycle and variable cumulative doses in 8,533 patients. Thirty-nine cases of AML/MDS were reported. The incidence was found to be particularly correlated with cyclophosphamide doseintensity, with a corresponding cumulative incidence of AML/MDS at 5 years of 1.04% (95% CI, 0.64% to 1.67%) for patients receiving more intense regimens (often with prophylactic growth colony-stimulating factor support), compared with 0.22% (95% CI, 0.12% to 0.43%) for those treated with standard AC. Breast radiotherapy also seemed to increase the leukemia risk (relative risk, 2.45; P = 0.007) (Ref. 48). Late cardiac toxicity associated with anthracycline-based chemotherapy is of low frequency (< 1%), but is a potentially serious and even life-threatening event. It varies from an asymptomatic decrease in left ventricular ejection fraction to clinically relevant congestive heart failure. It is directly correlated with the cumulative dose administered, increasing exponentially above 500 mg/m² for doxorubicin and 900 mg/m² for epirubicin. Other associated risk factors include underlying cardiovascular disease, hypertension, advanced age, prior or concomitant radiotherapy to the mediastinal or pericardial areas, and previous therapy with anthracyclines or anthracenediones. A recent publication reported on 355 breast cancer patients who were previously treated in three prospective trials of adjuvant chemotherapy and were still free of disease at 11 years of follow-up. Two hundred of these women had received doxorubicin-containing regimens, whereas 155 were treated with CMF. Both groups were balanced in terms of median age at randomization in the initial trial, median age at recall and percentage of risk factors for cardiovascular disease. The percentage of systolic dysfunction was higher in the doxorubicin group (8% versus 2%), and the cumulative cardiac mortality was 0.6% vs. 0% (Ref. 49). The French Adjuvant Study Group recently reported a very similar rate of systolic dysfunction in breast cancer survivors treated with the FEC (5-FU, epirubicin, cyclophosphamide) regimen (Ref. 50) Taxanes In recent years, the taxanes have been investigated in several trials of adjuvant chemotherapy. The Cancer and Leukemia Group B (CALGB) 9344 trial (Ref. 51) reported a benefit for the addition of four cycles of paclitaxel to a standard regimen of four cycles of doxorubicincyclophosphamide (AC) chemotherapy when compared to AC alone, in node-positive patients. A recent update of that trial, with a median follow-up of 69 months, demonstrated significantly better DFS and OS rates for paclitaxel-treated women, but the advantage seemed confined to the subset of patients with estrogen receptor-negative tumors. The NSABP B28 trial, comparing four postoperative cycles of doxorubicin and cyclophosphamide versus the same combination followed by four cycles of paclitaxel showed a significant reduction in recurrence with paclitaxel, but no difference in OS (Ref. 52). In both studies, the control arm consisted of 4 cycles of AC, considered to be sub-optimal treatment by today s standards. Version / January, 2006

27 A third trial, showed the triple-drug combination of docetaxel, doxorubicin, and cyclophosphamide (TAC) to be superior, in terms of DFS and OS, to standard 5-fluorouracil, doxorubicin, and cyclophosphamide (FAC) chemotherapy in a randomized comparison in node-positive patients. The benefit appeared to be confined to patients with 1 to 3 positive lymph nodes (Ref. 53;54). A French trial, PACS 01, also showed statistically significant improvement in DFS and OS in patients with 1 to 3 positive nodes and women older than 50 years with the sequence of 3 courses of FEC followed by 3 courses of docetaxel in comparison to 6 cycles of FEC (Ref. 55) Hence, the benefit appears to be restricted to post menopausal women. Regimens comprising anthracyclines and taxanes, either concomitantly (e.g. TAC) or in sequence (e.g. EC/EC followed by taxane) are generally reserved for node-positive patients with other features of poor prognosis in view of the significantly higher toxicity of these regimens. Among the multitude of trials attempting to delineate the role of the taxanes in the adjuvant treatment of breast cancer and enrolling more than 30,000 women, only one an American study (N=1,016) has addressed this question in node-negative women. This trial compared TC (docetaxelcyclophosphamide) to AC (doxorubicin-cyclophosphamide): with a median follow-up of 43 months there was no significant difference in either DFS or OS between the two regimens. While the follow-up is still insufficient, preliminary reports have shown TC to be at least as effective as AC (Ref. 56) The Combination Docetaxel-Capecitabine Capecitabine is an oral fluoropyrimidine, a prodrug that generates 5-Fluorouracil (5-FU), preferentially in tumor tissue, thus mimicking continuous-infusion administration. This selectivity is explained by the significantly higher concentration and activity of the enzyme responsible for the conversion, thymidine phosphorylase, in tumor tissue compared with healthy tissues. Preclinical studies in human cancer xenograft models demonstrated that administration of taxanes results in further up-regulation of thymidine phosphorylase in tumor tissue, as shown in studies with preoperative docetaxel. These same studies showed that its administration with capecitabine resulted in synergistic anti-tumor activity (Ref. 57;58). Subsequent clinical studies have shown that single-agent capecitabine is an active and tolerable treatment in metastatic breast cancer that has progressed during or after anthracycline and taxane therapy, achieving response rates of approximately 25% and a median survival in excess of 1 year. Activity in heavily pre-treated patients paved the way to investigate capecitabine activity earlier in the disease course and in combination with other cytotoxic agents. In addition, it proved to be an attractive agent for incorporation into combination regimens because of the low incidence of myelosuppression. A randomized phase III trial in the metastatic setting (Ref. 59), based on the results of clinical and preclinical studies, compared single-agent docetaxel to a docetaxel-capecitabine combination, and could demonstrate the latter to be more effective. The use of the combination resulted in superior response rate, improved DFS and notably a 3-month improvement in median survival. These results not only established this regimen as an important treatment option for patients with anthracyclinepretreated metastatic breast cancer but made it an attractive combination for transfer to the adjuvant setting. The side effect profile of capecitabine-docetaxel therapy was manageable and consistent with the known toxicities of the individual agents. There was a higher incidence of gastrointestinal side effects and hand-foot syndrome in patients receiving the combination therapy; indeed, the aggregate incidence of grade III adverse events was higher in the combination arm, predominantly because of grade III hand-foot syndrome and myalgia. On the other hand, arthralgia and neutropenic fever were more common with single-agent docetaxel (used at the higher dose of 100 mg/m 2 in single agent Version / January, 2006

28 therapy) and the incidence of grade IV adverse events was higher in the single-agent docetaxel arm, primarily because of neutropenic fever. All the side effects associated with the combination were manageable with appropriate medical intervention (e.g., loperamide and rehydration for diarrhea, mouthwash and fluconazole for stomatitis, and oral vitamin B 6 preparations and emollients for hand-foot syndrome) and treatment interruption and/or dose reductions when needed. Dose reductions were required in 65% of patients and capecitabine interruption was required in 34% of cycles. This combination is currently being evaluated in a randomized phase III trial in the adjuvant setting for the treatment of high-risk breast cancer (both node-positive and node-negative patients), run by the US Oncology Group. It compares four cycles of the AC regimen followed by either four cycles of docetaxel alone (at 100 mg/m²) or four cycles of docetaxel (75mg/m²) plus capecitabine (825 mg/m² twice daily - lower than in the pivotal phase III trial). Since in the trial this combination will be given in chemotherapy-naïve patients, i.e., having not previously received AC, the toxicity is expected to be reduced further Chemotherapy in the trial The trial will ask an important question regarding chemotherapy optimization for node-negative breast cancer: can a docetaxel-based regimen safely replace an anthracycline-based one? Docetaxel was the only agent shown to generate higher response rates than doxorubicin in a head to head comparison of single-agent use for the treatment of advanced breast cancer (Ref. 44) and among the many docetaxel-based combinations explored so far in breast cancer, one of the most interesting ones is docetaxel plus capecitabine; aside from the synergistic anti-tumor activity obtained in preclinical models, it showed improved response rate (RR), time to progression (TTP) and OS when compared to docetaxel alone in the previously detailed metastatic breast cancer randomized trial. The combination of docetaxel-capecitabine was also compared to doxorubicin-cyclophosphamide in the neoadjuvant setting; 78 patients with stages II and III breast cancer were randomized between the two treatments and those in the docetaxel-capecitabine combination presented higher clinical response (91% vs 74%) and, importantly, higher pathological complete response (23% vs 6%) (Ref. 60) however we are waiting for later time points to be reported to confirm these results. Although short-term side effects of this combination were significant, there are good reasons to believe that tolerance might be better in early breast cancer, when given upfront, and that, importantly, long-term side effects might be less than what is commonly observed with anthracyclines. Given the high rate of capecitabine dose reductions in the metastatic breast cancer trial, capecitabine will be given at the dose of 1650 mg/m² p.o. d1-14 q21 days (instead of 2500 mg/m²) in combination with docetaxel 75 mg/m² i.v. q21 days (dose unchanged). Since the combination of docetaxel-capecitabine has proved to be both safe and well-tolerated in women with metastatic breast cancer it is an exciting regimen to study in the adjuvant setting. In addition to their activity as single agents, promising preliminary results of their combined activity and the extent of experience with these two agents encourages their further investigation particularly in node-negative disease. This combination, with its different toxicity profile from anthracycline based regimes, will have a much lower incidence of serious long-term side-effects such as cardiotoxicity and leukemia, making it an attractive replacement for anthracycline-based chemotherapy. hopes to validate this new non-anthracycline-based chemotherapy regimen for use in high-risk women with node-negative breast cancer and to generate long-term safety data on this combination. Version / January, 2006

29 1.4 Endocrine therapy in the trial: is upfront Aromatase Inhibitor (AI) letrozole for 7 years safer and more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? For many years the current standard treatment for endocrine responsive breast cancer is 5 years of the antiestrogen tamoxifen; very recently national and international opinion guidelines have advocated the incorporation of an AI in sequence with or as a replacement to tamoxifen Adjuvant endocrine treatment in breast cancer Recently, the available data on the use of tamoxifen in the adjuvant setting of ER-positive breast cancer has been reviewed extensively by the Early Breast Cancer Trialists Collaborative Group (EBCTCG) (Ref. 10). In this meta-analysis, tamoxifen has been shown to reduce the annual odds of breast cancer death by a third and the annual recurrence rate by half in ER-positive patients regardless of progesterone receptor status, age and the use of chemotherapy. In addition, it reduced the odds of developing a contralateral breast cancer by more than 30%. For this reason, endocrine therapy is clearly the most important systemic adjuvant treatment for women with hormone receptor positive breast cancer, and is considered the standard of care. At present, the most important prognostic factor in breast cancer relates to axillary lymph node involvement. Due to the increasing use of screening mammography in women of 50 years of age in developed countries, the prevalence of node-negative breast cancer has been continually increasing. Adjuvant therapy for women with node-negative disease is therefore of prime importance for the treating physician today. Only one large trial, the NSABP Study B-14, has studied tamoxifen in the node-negative setting. In this study, 2,644 patients with ER-positive primary tumors were randomly allocated to receive either tamoxifen for 5 years or placebo. At the end of four years of follow-up the difference in DFS between the tamoxifen and placebo groups was highly statistically significant (p< ) (Ref. 61), confirming the benefit of tamoxifen therapy in this group. Tamoxifen is the trans-isomer of a substituted triphenylethylene with anti-estrogenic activity due to a competitive inhibition of estrogen action. It blocks the binding of (3H)estradiol to the estrogen receptor. Its clinical efficacy and long term side effects have been clearly demonstrated by trials conducted in the 1970 s and the Oxford Overview. A minimum of five years of treatment with tamoxifen is considered standard. Tamoxifen is well tolerated, and adverse effects are mild when compared with cytotoxic chemotherapy. Short term toxicities of hot flushes and vaginal discharge are related to the anti-estrogenic effect of the drug. Tamoxifen has been shown to significantly increase the incidence of endometrial cancer (Ref. 62), and at high doses retinopathy has been observed (Ref. 63). Tamoxifen s estrogenic properties also reduce the mean levels of cholesterol, which is thought to be the cause of the reduction of cardiovascular deaths observed in some studies and a bone protective effect. An association between the use of tamoxifen and increased thromboembolic disease is also firmly established (Ref. 10;62) Use of Aromatase Inhibitors in the adjuvant setting There have been three main strategies in the effort to improve cure rates from breast cancer using third generation AIs in the adjuvant setting: first as initial treatment in direct comparison with tamoxifen, second in sequential use before or after tamoxifen for a total period of five years, and third as extended therapy after five years of tamoxifen. At least 10 adjuvant trials involving over 40,000 women with early stage breast cancer are currently in progress. The results of those trials that have been reported are shown in Table 2. This section will summarize these results and the questions that still remain. Version / January, 2006

30 Table 2: Comparison of current published studies involving adjuvant AIs Trial name: ATAC (anastrozole) BIG 1-98 (letrozole) IES (exemestane) ABCSG/ARNO (anastrozole) ITA (anastrozole) MA-17 (letrozole) Comparison/duration TAM 5yrs vs AI 5yrs TAM 5yrs vs AI 5yrs (core analysis) TAM 5 yrs vs TAM 2yr-AI 3yrs TAM 5yrs vs TAM 2yrs-AI 3yrs TAM 5yrs vs TAM 2yrs-AI 3yrs TAM 5yrs vs TAM 5yrs-AI 5yrs No. of women 6,241 8,028 4,742 3, ,187 randomized Median age Node-positive % HR+ (%) Adj chemotherapy given (%) Follow-up (months) Endocrine-refractory Yes Yes No No No No women included Hazard Ratio for disease free survival P= P= P= ** P< P= Hazard Ratio for disease free survival² (HR+ only) Hazard Ratio for overall survival 0.97 P= P=NS 0.83 P= P=0.16 Not reported 0.61 P=0.04 (node-positive) TAM: tamoxifen 20 mg daily, AI: aromatase inhibitor; HR+: hormone receptor positive; Adj: adjuvant, 1 Disease free survival is defined differently for each trial. Please refer to references for further detail. 2 Relapse free survival defined as locoregional, distant metastases, and contralateral breast cancer recurrences only. Version / January, 2006

31 1.4.3 Tolerability profiles The side effect profile of tamoxifen is well known and the third generation AIs are generally well tolerated. The available data indicates at least equal short term safety including symptoms of menopause and quality of life. However, long term data will not be available for the next 10 years. The main side effects are described below: Hot flushes and general tolerability In the ATAC and IES adjuvant AI studies in which tamoxifen is the standard arm, the short term tolerability of the AI was at least as good as that of tamoxifen. There was significantly less vaginal bleeding in patients receiving anastrozole than tamoxifen in the ATAC study. In the IES study, the frequency of hot flushes was similar in both arms. In the MA-17 study, where letrozole was compared with placebo, there were more hot flushes in the letrozole arm. Otherwise, short term tolerability was excellent. The percentage of women discontinuing treatment because of adverse events was not significantly different between the two arms, in all studies involving AIs Skeletal effects Tamoxifen has been shown to preserve bone mineral density in post menopausal breast cancer patients. AI may reduce bone density as has been suggested by the ATAC study. Women taking anastrozole were more likely to suffer from musculoskeletal disorders (p<0.001), in particular fractures (p<0.001). Letrozole has also been shown to significantly increase bone resorption. In the MA-17 study, more women in the letrozole group compared with placebo suffered from new-onset osteoporosis. There are clinical trials currently assessing bisphosphonates in the prevention and treatment of AI induced bone loss (ZO-FAST, SABRE). In addition, mylagia and arthralgia were reported more frequently for women receiving letrozole (Ref. 64) Thromboembolic events and endometrial cancers In all trials comparing the third generation AIs with tamoxifen, the adverse events caused by tamoxifen s estrogenic properties were significantly less common in the AI groups. This particularly included ischaemic cerebrovascular disease, venous thromboembolic (including deep venous thrombosis) events and endometrial cancer. Compared with placebo, letrozole did not lead to increased rates of any of these serious adverse events in MA-17 and vaginal bleeding occurred more often in the placebo arm (p=0.01) Lipid metabolism Whilst retrospective analyses of the three randomized tamoxifen trials suggested a reduction in coronary heart disease in favor of tamoxifen (Ref. 65), no such benefit was reported in the NSABP- P1 chemoprevention study (Ref. 62). In view of the recent Women s Health Initiative Study (Ref. 66), it is unclear whether the favorable influence of tamoxifen on lipid metabolism translates into a true reduction in coronary heart disease, and whether lipid metabolism is the true underlying reason for this observation, if indeed it is correct. In the ATAC trial, more patients receiving anastrozole were reported to have an elevated cholesterol compared with patients receiving tamoxifen (7% vs 3% respectively) (product information Arimidex, Astra-Zeneca & Whereas in the adjuvant MA.17 study which is the only trial to compare letrozole to placebo, there was no increase in cardiovascular events between the two arms. Results from the lipid substudy suggests that letrozole does not significantly alter serum cholesterol, high density lipoprotein (HDL) cholesterol, low Version / January, 2006

32 density lipoprotein (LDL) cholesterol, triglycerides or Lp(a) in non-hyperlidiemic postmenopausal women with primary breast cancer treated up to 36 months following at least 5 years of adjuvant tamoxifen therapy (Ref. 68). Exemestane may have effects converse to the other AI on lipid metabolism due to it being an androgenic steroid, however the IES study did not specifically address this endpoint. In two small studies looking at the effects of letrozole on lipid metabolism in healthy women, conflicting results were seen. In one study, there was no effect on the serum lipid profile, whilst the other reported significantly increased total and LDL cholesterol levels, as well as an increase in the atherogenic risk ratios of total/hdl and LDL/HDL cholesterol (Ref. 69;70). In summary, the evidence suggests that AI use has benefits in terms of overall tolerability with fewer (but different) toxicities compared with tamoxifen. Women who receive AI have fewer gynecological symptoms, no increase in the incidence of endometrial cancer, fewer thromboembolic events and less hot flushes offset by the chance of more fractures and more arthralgias. The possible toxicities from long term AI use, particularly the possibility of a higher risk of cardiac disease, need to be studied in further detail Improving adjuvant therapy in the first 5 years Two trials have studied the benefit of upfront AI instead of tamoxifen in the first 5 years of adjuvant therapy (also see table 2). ATAC The results of the largest and first published study, Arimidex and Tamoxifen Alone or in Combination (ATAC) (Ref. 71) with 9,366 postmenopausal patients randomized to tamoxifen or anastrozole or to a combination of tamoxifen and anastrozole for a period of five years showed statistically significant reduction in the rate of relapses (local, distant or contralateral disease) with anastrozole as compared to tamoxifen (87% vs 89%, respectively) when it was first reported at median follow-up of 33 months. At this time, this absolute 2% reduction translated into a relative risk reduction of 17% (p=0.013). The results of the first efficacy update, based on a longer median follow-up period of 47 months and a second recent update after 68 months were confirmatory, with the absolute difference in risk reduction now being 3.7% (Ref. 72). As expected the effect was seen only in patients whose tumors were known to be hormone receptor positive but interestingly, the risk reduction was greater for the ER+, PgR- tumors than for the ER+, PgR+ ones (HR: 0.48 vs. 0.82) (Ref. 73). While this retrospective subset analysis needs prospective confirmation, the result might be explained by the fact that ER+, PgR- tumors co-segregate with HER-2 positivity. Interestingly, the combination of tamoxifen and anastrozole was not superior to tamoxifen alone proving that a combination of two efficient drugs is not always better than their single agent use. There was also a lower incidence of contralateral breast cancer with the use of anastrozole (HR: 0.62, p=0.062). To date there is no difference in the rate of breast cancer related deaths or OS. BIG 1-98 In this study, 8,028 patients were randomized to; tamoxifen for 5 years, letrozole for 5 years, tamoxifen for 2 years followed by letrozole for 3 yrs, or the 2 drugs in the reverse sequence. The first results have recently been reported (Ref. 67). The primary core analysis compared only the first two arms and the sequential arms censored at 2 years (prior to the switch). The analysis was based on 8,010 patients with a median follow-up of 26 months. Letrozole was found to significantly prolong DFS compared with tamoxifen for post menopausal women (0.81, p=0.003) and time to recurrence (HR:0.72, p=0.0002) especially in distant sites. This supports the results of the ATAC study. Again, there was no difference in OS seen at this stage. Both of these trials demonstrate that 5 years of upfront AI therapy is a reasonable alternative to the previous standard of 5 years of adjuvant tamoxifen therapy for postmenopausal women with Version / January, 2006

33 endocrine responsive breast cancer. Indeed, for women with a contraindication to treatment with tamoxifen, an AI is the treatment of choice, given the current evidence from both an efficacy and short-to moderate term toxicity stand-point The sequential approach Intergroup Exemestane Study (IES) The Intergroup Exemestane Study (IES, BIG 2-97), investigated the concept of sequential therapy with AI after tamoxifen. After two to three years of tamoxifen, women were randomized to switch to exemestane or to continue with tamoxifen for the remainder of the five-year treatment programme. After a median follow-up of 37.6 months results of the second interim analysis indicated a significantly higher DFS (HR 0.73, p<0.001), corresponding to an absolute benefit in terms of DFS of 4.7% at three years after randomization and breast cancer free survival benefit (HR:0.7; , p=0.0005) in favor of sequential therapy (Ref. 74;75). Noteworthy, distant metastases-free survival (DMFS) and time to contralateral breast cancer were also significantly better with sequential therapy (HR of 0.63 and 0.44, respectively with p values of <0.001 and <0.004). So far no difference in OS has been reported. The ABCSG Trial 8 and ARNO 95 trial The efficacy of anastrozole after two to three years of tamoxifen was assessed in two other phase III trials (ABCSG Trial 8, ARNO 95) which were similar in design and prospectively planned for combined analysis by the Austrian and German Breast Cancer Clinical Trial Groups. The recently presented results also confirm the benefit of sequential AI therapy after tamoxifen (Ref. 76). In these trials 3,123 women were randomized to receive anastrozole after two years of tamoxifen or continue with tamoxifen for a total duration of 5 years. At a median 26 months of follow-up and after 143 events (local, metastatic or contralateral) the HR for risk reduction with anastrozole versus tamoxifen is 0.59; (p<0.0018). Italian Tamoxifen and Anastrozole (ITA) trial A substantially smaller study of 448 node-positive patients, with a similar design to the above trials has also been reported after a median follow-up of 36 months (Ref. 77). There were 45 events reported in the tamoxifen arm, and 17 events in the sequential arm (p<0.0002). DFS and local recurrence free survival were also significantly longer in the anastrozole arm (HR: 0.35, 95% CI: , p<0.001 and HR 0.15; 95% CI: ). Overall survival is not different between the arms (HR: 0.52, 95% CI , p=ns) and updated efficacy data after a median follow-up of 52 months are confirmatory (Ref. 78). Hence, these four trials demonstrate that the sequential strategy of tamoxifen followed by a switch to an AI after 2 to 3 years for 5 years total duration is superior for reducing the risk of breast cancer recurrence to the previous standard of 5 years of tamoxifen. However, it is unclear if for women who do not have a contraindication to tamoxifen, if treatment with an upfront AI is superior, equivalent, or inferior to a planned cross over from tamoxifen to an AI at some fixed point in time. Furthermore, these trials arbitrarily selected five years as the total duration of endocrine therapy, and it is not known if longer treatment involving a strategy of an early switch would be of sustained clinical benefit Extending adjuvant therapy beyond 5 years Approximately half of all breast cancer relapses in women receiving 5 years of adjuvant tamoxifen occur between 5 and 15 years after surgery and that risk of recurrence appears to continue indefinitely (Ref. 10). The average hazard of recurrence between years 5 and 12 after primary breast surgery for 2,257 ER-positive women was around 5% per year (Ref. 79). Based on this rationale, the NCIC-CTG MA.17 / BIG 1-97 study evaluated the strategy of extending adjuvant endocrine Version / January, 2006

34 therapy beyond 5 years of tamoxifen with an AI (Ref. 64). After completing 5 years of tamoxifen, 5,187 women were randomly assigned to receive letrozole 2.5 mg or placebo. The results of the first interim analysis, conducted after a median follow-up of 2.4 years, indicated a significantly higher DFS in the letrozole group than in the placebo group (HR for a recurrence or a new contralateral breast cancer 0.57, p<0.001). This translated into an absolute difference of 2.2% in the rate of breast cancer events, including distant metastases, ipsilateral recurrences and contralateral breast cancers, with an actuarial projection of an absolute difference of 6% in the rate of events over a follow-up of four years. The effect of letrozole was equally distributed between women with node-negative and node-positive disease. The robust difference in the rate of events led to the early reporting of the study results at a time where less than 1% of the participating women had completed their randomly assigned therapy. So in reality, the benefit was seen when no women had completed the assigned 10 years of therapy (median of 7.4 years in total). The benefit in survival recently reported has been seen only in the node-positive subset (p=0.04) (Ref. 80). This trial established that a strategy using AI in the extended adjuvant setting (longer than 5 years) could improve recurrence rates from breast cancer above that obtained from 5 years of tamoxifen treatment alone. At present, it seems that a minimum of 2.4 years of extended therapy can be recommended based on the median follow-up from MA.17. Since all patients on placebo were offered the option of beginning letrozole, the optimal extended duration will not be provided by the long term data from this trial. However, the survival advantage in the node-positive subset is noteworthy and it is clear that extended therapy (more than 5 years of adjuvant treatment) has advantages for certain women. This trial led to the early closure of the similarly designed National Surgical Adjuvant Breast and Bowel Project Study (B33) comparing exemestane and placebo after 5 years of tamoxifen. Seemingly confirming the benefit of extended duration adjuvant endocrine therapy are the results of the ABSCG Trial 6a. In this trial, 856 postmenopausal women who had previously received 5 years of tamoxifen on a previous phase III trial were randomized to receive anastrozole for 2-3 years or placebo (Ref. 81). Reported after a median follow-up of 60 months, the risk of recurrence was reduced (HR: 0.64; 95% CI , p=0.0477) with no OS advantage seen as yet Which strategy is best? At present, it is difficult to know how to compare results across these trials to determine which particular agent or strategy is superior for an individual breast cancer patient. Based on the published results of these studies, the American Society of Clinical Oncology and the St Gallen consensus panel in 2005 concluded that the optimal adjuvant endocrine treatment for post menopausal women with HR-positive breast cancer today should include an AI as initial therapy or sequential therapy consisting of tamoxifen followed by an AI (Ref. 82). As a result, for postmenopausal women diagnosed today with hormone responsive early breast cancer there are many possibilities for her adjuvant endocrine therapy. The optimal option is uncertain: if treatment upfront with an AI is superior, inferior or equivalent to a switch from tamoxifen to an AI, or if tamoxifen after an AI switch could be advantageous or detrimental. The optimal duration is also unclear: 5 years or extended treatment however there is increasing evidence supporting extended adjuvant endocrine therapy. It is also unknown if the use of an AI with ovarian function suppression can be extrapolated into the premenopausal setting. It is unknown which of these treatments or combinations is most effective and for which individuals/subgroups of patients. It is also possible that some individuals will respond better to one treatment compared to another treatment or due to the characteristics of their disease one strategy may be more appropriate than another. The BIG 1-98 four arm study, where women are randomized to receive either 5 years of tamoxifen, 5 years of letrozole, 2 years of tamoxifen followed by 3 years of letrozole or 2 years of letrozole followed by 3 years of tamoxifen, will help us answer some of these questions in a few years time. It will not help us in delineating the benefits of extended therapy however, which clearly has Version / January, 2006

35 advantages for certain women. At present, we can conclude that the group of women who recur early (in the first 2-3years) gain an advantage from upfront AI however, unfortunately we have not been able to identify the characteristics of this group of patients yet. In trials of switching strategies we see a greater treatment effect for sequential AI, than when AI is compared as an initial therapy, this maybe due to the initial period of tamoxifen treatment selecting a population that respond better to endocrine therapy than the starting population do. Furthermore, the full long term health costs (particularly skeletal and cardiovascular problems) of AI use will not be known for another 10 years when toxicity reporting is complete for these trials. From these data, it is clear that AI have a role in the adjuvant setting. However, the optimal sequence and duration of the AI and tamoxifen combination and its performance in comparison to a strategy of upfront AI is unknown. Also, it is clear that extended therapy with endocrine treatments has definite merit. Whilst part of these questions may be addressed by the mature results of the BIG 1-98 trial, it has also become evident that the one shoe fits all approach is unsuitable for all HRpositive breast cancers. This trial will therefore evaluate the following comparison: 2 years of tamoxifen followed by 5 years of letrozole versus 7 years of upfront letrozole. Seven years has been chosen to replicate the MA.17 data where the median duration of treatment was 2.4 years after 5 years of tamoxifen. Whilst this duration of adjuvant endocrine therapy is not currently standard, it is clear that extended therapy is beneficial and should be explored in a clinical trial setting. will have the huge advantage over all other clinical trials involving AIs: it will secure the collection of frozen and paraffin-embedded tumor tissue as well as serum for all participating women. In this way, it has great potential for the discovery or validation of molecular signatures indicating which women are candidates for an AI upfront and which women may be best served by a sequence of tamoxifen followed by AI. 2 Objectives of the trial 2.1 Primary objectives The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to node-negative breast cancer patients. The trial will also aim to compare a docetaxel-capecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens. The last primary objective will be to determine the best endocrine treatment strategy i.e. single agent up-front, AI (letrozole) for seven years compared to the sequential endocrine strategy of 2 years of tamoxifen followed by 5 years of letrozole. 2.2 Secondary objectives The trial has several secondary objectives: Comparison of the two prognostic methods: To estimate both relative (Hazard Ratio, HR) and absolute (as % at 5 years) efficacy in terms of disease free survival (DFS), distant metastasis free survival (DMFS) and overall survival (OS) Version / January, 2006

36 of chemotherapy in the two subgroups where the clinical-pathological prognosis and the 70- gene signature molecular prognosis are discordant. To calculate overall estimates of efficacy (DFS, DMFS, OS) for each treatment strategy according to clinical-pathological prognosis and according to molecular prognosis. To estimate the percentage of patients receiving chemotherapy per each prognostic method. Gene profiling: To identify and/or validate predictive gene expression profiles of clinic al response/resistance to anthracycline-based and docetaxel-capecitabine chemotherapy. To identify and/or validate novel gene expression signatures predicting clinical response to sequential tamoxifen-ai versus AI alone. Chemotherapy: To compare the OS distributions associated with the 2 chemotherapy regimens. To determine and compare the early and late toxicities associated with each regimen. Endocrine therapy: To evaluate adjuvant endocrine treatment success or failure in the subgroup of postmenopausal patients with endocrine responsive disease. To compare the safety profile of the two endocrine therapy regimens. Long term research objectives: To set up several biological material bank resources (RNA, tumor tissue, serum) for future translation studies in both genomics and proteomics. 2.3 Endpoints Treatment decision randomization (R-T) The primary endpoint for the treatment decision randomization is distant metastasis free survival (DMFS) at 5 years. Secondary endpoints are: the proportion of women treated with chemotherapy per treatment decision making tool i.e. clinical prognosis compared to 70-gene signature prognosis overall survival at 5 years DFS at 5 years Chemotherapy randomization (R-C) The primary endpoint for the chemotherapy randomization is disease free survival (DFS). Secondary endpoints are: overall survival at 5 years safety both early and late DMFS at 5 years Version / January, 2006

37 2.3.3 Endocrine therapy randomization (R-E) The primary endpoint for the endocrine therapy randomization is disease free survival (DFS). Secondary endpoints are: overall survival at 5 years DMFS at 5 years 3 Patient selection criteria 3.1 General eligibility criteria for inclusion in the trial To be eligible for the trial patients will have to comply with the following eligibility criteria: women with histologically proven operable invasive breast cancer and a negative sentinel node or negative axillary clearance (N0, M0). Acceptable primary treatment options are: breast conserving surgery or mastectomy with either a sentinel node procedure or full axillary clearance. radiotherapy is mandatory in the case of breast conserving surgery and will be administered according to local institutional policy after mastectomy. patients with unresectable positive deep margins who receive adjuvant radiotherapy are eligible provided that all other margins are negative. a tumor clinical classification of T1, T2 or operable T3. the breast tumor must be unilateral, however DCIS or LCIS are allowed if invasive cancer is present. a frozen tumor sample (not fixed in formalin) must be available for inclusion. The tumor sample must be taken from the excised primary tumor (a core punch biopsy). patients should: be aged between 18 and 70 years at randomization. (Elderly patients for whom adjuvant treatment is considered could be offered participation in one of the BIG Elderly trials.) have a WHO performance status of 0 or 1. have adequate bone marrow reserves (neutrophil count >1.5 x10 9 /l and platelet count >100 x10 9 ), adequate renal function (creatinine clearance 50 ml/min (calculated according to Cockroft and Gault, see Appendix C), or serum creatinine 1.5 x upper limit of normal), hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline phosphatase 2.5 x upper limit of normal, total bilirubin 2.0 x upper limit of normal) and normal ECG compatible with chemotherapy administration. have NO serious cardiac illness or medical condition including but not confined to: a history of documented congestive heart failure (CHF), high-risk uncontrolled arrhythmias, angina pectoris requiring antianginal medication, clinically significant valvular heart disease, evidence of transmural infarction on ECG, poorly controlled hypertension (e.g. systolic > 180 mm Hg or diastolic > 100 mm Hg), symptomatic coronary artery disease or a myocardial infarction within the last 12 months or other risk factors that contra-indicate the use of anthracycline-based chemotherapy. Version / January, 2006

38 NOT have serious uncontrolled intercurrent infections, or other serious uncontrolled concomitant disease. NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any cancer (other than breast cancer) in complete remission for 5 years. NOT have received previous chemotherapy, hormonal therapy or radiotherapy. NOT have participated in any investigational drug study within the 4 weeks preceding the start of treatment. NOT be pregnant or breast-feeding at the time of diagnosis or randomization. A woman of childbearing potential must have a negative pregnancy test. If post-menopausal, the woman must have been amenorrheic for at lest 12 months to be considered of non-childbearing potential while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, injectables, combined oral contraceptives, some IUDs, sexual abstinence or vasectomised partner (Note 3 of the guidance on non-clinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). NOT have any psychological, familial, sociological, geographical or serious uncontrolled medical (e.g. a history of uncontrolled seizures, central nervous system disorders, or psychiatric disability) condition judged by the investigator to be clinically significant which could potentially preclude informed consent or interfere with compliance for oral drug intake or with the study protocol and follow-up schedule. These conditions should be discussed with the patient before enrollment in the trial. signed written informed consent must be given according to ICH GCP, and national/local regulations before enrollment in the trial (Both a screening PIS & IC and the PIS & IC 1 must be signed before enrollment). 3.2 Selection criteria for treatment decision randomization (R-T) Patients who meet all the general eligibility criteria and for whom the genomic and clinical risk assessments are discordant will be randomized between genomic-treatment decision making and clinical-treatment decision making. (details of stratification factors are given in section ) 3.3 Selection criteria for chemotherapy randomization (R-C) Patients will be eligible for inclusion in the chemotherapy randomization (R-C) if they meet all of the general eligibility criteria AND the following criteria: (details of stratification factors are given in section ) women who have a high risk of recurrence according to both the clinical-pathological criteria and the 70-gene signature. OR women who have a high risk according to the clinical-pathological criteria and a low-risk according to the 70-gene signature, and were randomized (R-T) to use the clinical-pathological criteria for chemotherapy decision. OR Version / January, 2006

39 women who have a low-risk according to the clinical-pathological criteria and a high-risk according to the 70-gene signature and were randomized (R-T) to use the 70-gene signature for chemotherapy decision. AND WHO status 0 or 1 (see Appendix B) have cardiac function (left ventricular ejection fraction (LVEF) within the normal limits of each institution, measured either by echocardiography or by radionuclide angiocardiography (MUGA)) according to the standard of care. written informed consent (PIS & IC 2) prior to beginning specific protocol chemotherapy procedures, to be given and documented according to ICH GCP and national/local regulatory requirements. interval between definitive surgery and start of chemotherapy should ideally be no more than 6 weeks but can not exceed 8 weeks. NOT have had any organ allografts requiring immunosuppressive therapy. NOT have a requirement for concurrent use of the antiviral agent sorivudine (antiviral) or chemically related analogues, such as brivudine. NOT have a history of severe hypersensitivity reaction to drugs formulated with Polysorbate 80. have physical integrity of the upper gastrointestinal tract, ability to swallow tablets, and NO malabsorption syndrome. 3.4 Selection criteria for endocrine therapy randomization (R-E) Patients who meet all the general eligibility criteria (see section 3.1) and who have endocrine responsive disease AND meet the following criteria will be offered the endocrine therapy randomization (R-E). (details of stratification factors are given in section ) are hormone receptor positive, either ER, PgR or both. have given written informed consent (PIS & IC 3) prior to beginning specific protocol endocrine therapy procedures, according to ICH GCP and national/local regulatory requirements. NOT have a prior history of thromboembolic disorder, DVT or pulmonary emboli. premenopausal women must have ovarian function suppression during endocrine therapy with aromatase inhibitor (see 7.3 for details). patients must not have previously received high dose systemic corticosteroids (except as part of their anti-emetic treatment), immunotherapy or biological response modifiers (e.g.: interferon). hormone replacement therapy (HRT) must be stopped at least 4 weeks before trial endocrine treatment is initiated. patients who have received adjuvant anti-estrogen therapy for more than 1 month immediately following surgery, radiotherapy, and/or chemotherapy may NOT be proposed enrollment in the endocrine question of the trial. women with a history of breast cancer chemoprevention with anti-estrogens can be included if at least 18 months has elapsed between discontinuation of chemoprevention and clinical or radiological diagnosis of breast cancer. Version / January, 2006

40 4 Trial Design is a phase III randomized superiority trial with three randomizations to compare clinical-pathological risk assessment to gene expression risk assessment using the 70-gene signature. It will also assess the efficacy of commonly used anthracycline-based chemotherapy regimens, selected from a list of acceptable regimens, to the new capecitabine-docetaxel combination. In addition the long term safety and efficacy of 2 years of tamoxifen followed by 5 years of letrozole versus 7 years of letrozole will be investigated. Node-negative women, will be offered those randomizations for which they are eligible, as shown below. Eligibility will be determined depending on an individual s clinical and genomic risk assessments, the consequent decision to administer chemotherapy, and their eligibility for endocrine therapy. Delay between surgery and start of treatment ideally not more than 6 weeks, can not be > 8 weeks Screening Visit surgeon (T1, T2 or operable T3 & cn0 & M0) screening PIS & IC, Screening Surgery Tumor sample shipment, RNA extraction, microarray analysis, local pathology, TNM, lymph node status, HR status pn0 Signing of PIS & IC 1 & Enrollment Genomic/Clinical Prognosis review & randomization Discordant Low/Low High/High R-T No CT CT R-C HR - HR - HR + HR + R-E CT Chemotherapy R-T Treatment decision Randomization based on genomic vs clinical prognosis R-C Chemotherapy Randomization of Anthracycline based CT vs Docetaxel/ Capecitabine R-E Endocrine treatment Randomization Letrozole vs Tamoxifen followed by Letrozole HR Hormone receptor PIS & IC Patient information sheet and informed consent Version / January, 2006

41 4.1 Screening Prior to entering into the trial, at the time of surgery, when patient eligibility is not known, all patients who sign a screening PIS & IC to consent to the donation of a tumor sample, will be registered for screening. 4.2 Enrollment The date when the patient signs the PIS & IC 1 (after both the genomic and clinical risk assessments have been performed) is the date the patient enters the trial. This date will be referred to as the ENROLLMENT DATE. There are 3 possible randomizations in this trial and each of the 3 will only randomize a sub-set of the enrolled patients. Eligible patients with adequate tumor tissue for microarray analysis will be enrolled. Genomic risk using the 70-gene signature prognosis and clinical-pathological risk using Adjuvant! Online will be determined. Patients are evaluated as low clinical-pathological risk, if their 10-year disease specific survival (without chemotherapy or endocrine therapy) is estimated by Adjuvant! Online (Ref. 36) as greater than 88% for ER-positive patients, and greater than 92% for ER-negative patients (details given in section ). Adjuvant! Online The current standard version of Adjuvant! Online is version 8.0, as new standard (but not genomic) versions become available the new standard version will be incorporated into the web based platform and used to determine clinical risk for the trial. Local pathology categorization of hormone receptor status for stratification For stratification at the time of randomization, the local immunohistochemistry results will be used. Investigators will report positive, negative or unknown for both estrogen receptor and progesterone receptor according to local policies. Some guidelines are given below on levels for receptor positivity using various methods. Local pathology can report estrogen and progesterone receptor levels either as a percentage of immunoreactive cells, as a combination of percentage of immunoreactive cells and staining intensity using the Allred scale or as a biochemical concentration. If a percentage of immunoreactive cells is reported, the absence of immunoreactive cells is negative, 1% immunoreactive cells is positive. Using the Allred scale below 3 is deemed negative and 3-8 positive. If a biochemical concentration is reported, < 10 fmol receptor per mg cytosolic protein is negative and 10 fmol receptor protein/mg cytosolic protein is positive. Guidelines for reporting local ER/PgR positivity & negativity Positive: ER 1% immunoreactive cells, Allred score > 2 or 10 fmol/mg PgR 1% immunoreactive cells, Allred score > 2 or 10 fmol/mg Negative: ER < 1% immunoreactive cells, Allred score < 3 or < 10 fmol/mg PgR < 1% immunoreactive cells, Allred score < 3 or < 10 fmol/mg Local pathology categorization of HER-2 status for stratification For stratification at the time of randomization, the local pathology results will be used. Investigators will report positive, negative or unknown for HER-2 status according to local policies. Immediately after enrollment the web based system will inform the Investigator if the patient will receive chemotherapy or not. The web based platform will calculate both clinical and genomic risk and randomize those patients who are discordant for the two methods. For all enrolled patients the Version / January, 2006

42 web based platform will specify the individual genomic and clinical test results and the outcome of the randomization if the patient is discordant. 4.3 Treatment decision randomization for chemotherapy allocation Patients who are high risk for both prognostic methods will receive chemotherapy, and patients who are low risk for both prognostic methods will not receive chemotherapy. Patients for whom both methods are discordant will be randomized (R-T) between chemotherapy-decision-making according to clinical criteria (using Adjuvant! Online) or chemotherapy-decision-making according to genomic prognosis using the 70-gene signature. Use Clin-Path risk to decide Chemo or not Use 70-gene signature risk to decide Chemo or not Clin-Path High 70-gene signature Low: CT 70% pts Clin-Path High 70-gene signature Low: no CT 70% pts Clin-Path Low 70-gene signature High: no CT 30% pts Clin-Path Low 70-gene signature High: CT 30% pts As a result of this randomization, as a whole, half of the patients with discordant risks will be proposed chemotherapy and half will not be proposed chemotherapy. Also, for half of these patients chemotherapy will be assigned according to clinical-pathological risk, and for the other half chemotherapy will be assigned according to the 70-gene signature risk. 4.4 Chemotherapy randomization trial design The chemotherapy question is a randomized, two-arm, prospective, non-blinded, multi-center phase III study. Patients who are candidates for chemotherapy (also see treatment section) will be proposed randomization R-C between anthracycline based chemotherapy or docetaxel-capecitabine chemotherapy. The following patients will be offered the chemotherapy randomization (R-C): Women who have a high risk of recurrence according to both the clinical-pathological criteria and the 70-gene signature. Women who have a high risk according to the clinical-pathological criteria and a low-risk according to the 70-gene signature, and were randomized (R-T) to use the clinical-pathological criteria for chemotherapy decision. Women who have a low-risk according to the clinical-pathological criteria and a high-risk according to the 70-gene signature and were randomized (R-T) to use the gene signature for chemotherapy decision. The objective is to evaluate if the docetaxel-capecitabine regimen is a more effective and less toxic alternative to an anthracycline-based regimen, for high-risk node-negative patients. Those patients who should be proposed chemotherapy but who do not wish to participate in the chemotherapy randomization (R-C) should receive the hospital s standard care chemotherapy. They remain on protocol and should be offered the endocrine therapy randomization if applicable. Trastuzumab will be allowed and recommended (however not provided free as a trial drug) for women with HER-2 positive tumors if it becomes approved and available for use in the adjuvant Version / January, 2006

43 setting during the trial. Treatment with trastuzumab is recommended to start after the end of chemotherapy. Hormone therapy treatment should be started only after the completion of chemotherapy and according to the endocrine therapy randomization (R-E) for consenting patients. 4.5 Endocrine therapy randomization trial design All post-menopausal patients who are candidates for endocrine therapy will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. Pre-menopausal patients may be offered this randomization at the discretion of their treating physician. For these patients, the suppression of ovarian function (temporarily or definitively) is mandatory (until the patient is > 50 years for temporary suppression). GnRH will not be provided. Endocrine therapy should start after surgery for those patients who did not receive chemotherapy or after chemotherapy for those patients who received chemotherapy (R-C). Those patients who have endocrine responsive tumors but who do not wish to participate in the endocrine therapy randomization (R-E) should receive the hospital s standard care endocrine therapy and remain on protocol. 4.6 pilot study The first 800 women randomized into the trial will form a pilot study to ensure that the clinical trial population is not biased and that key assumptions hold. They will also be used to check that the trial logistics function correctly and that physicians/patients comply with the allocated randomizations. The trial will not be interrupted after the first 800 patients unless the assumptions prove to be incorrect. (details of which can be found in the IDMC section, chapter 11) Note: For details of statistical testing refer to the statistics section (chapter 10). 5 Guidelines for breast cancer surgery and adjuvant radiotherapy The following information is given as recommendations but it is not an integral part of protocol investigations. 5.1 Breast cancer surgery Patients should receive optimal loco-regional treatment. The surgical part of the breast cancer treatment may consist of breast conservation surgery (wide local excision of the primary tumor) or mastectomy. The surgical part of the axilla encompasses the sentinel node procedure (SNP) for lymphatic staging or axillary lymph node dissection (ALND) for the treatment of lymph node metastases or staging procedure. Since the SNP can be considered as standard of care for lymphatic staging, SNP is preferred over ALND for this purpose. To ensure as much as possible uniformity in surgical techniques, the operations are described in more detail. It is recommended to follow these evidence based guidelines (Ref. 83;84) for surgery as much as possible. Wide local excision The main aim is to achieve free tumor margins. To achieve this aim, the surgeon should try to remove at least a 1 cm rim of healthy breast tissue. Particularly in large tumors, substantial parts of the breast have to be removed and special attention should be made to breast reconstruction and Version / January, 2006

44 cosmesis. The so called segment (or preferably sector resection, which is the proper mathematical denominator for a piece of pie excision) resection is one of the better techniques. Breast tissue is removed from the skin (either with or without a skin island for cosmetic reasons) down to the basal fascia. The advantage of this technique is that margin assessment is facilitated (only the lateral margins, periphery and nipple side are relevant; not skin and fascia side), as is the possibility to reconstruct the breast tissue. To find and excise non-palpable cancers, a number of aids are available and the guide wire is the most widely applied method. Gamma probe directed excision after intra-lesional radioisotope injection and ultrasound directed excisional biopsy appears to be helpful to achieve free margins, compared to guidewire directed wide local excisions. If cancer is diagnosed with minimal invasive techniques, it should be possible to achieve free margins in over 80% of patients at the first operation. The complication rate should be as low as 5%. For invasive ductal cancer, after complete excision and radiotherapy to the breast, the local relapse rate should be less than 1% per year of follow-up. Cosmetic outcome should be acceptable and good in over 70% of patients. Simple mastectomy The aim of simple mastectomy is the removal of preferably all breast tissue. Removal of the entire breast with its skin and the nipple-areola complex, with the pectoralis major muscle fascia, but without axillary node dissection. There is no axillary clearance involved. However, if the axillary tail is removed frequently some lower axillary nodes are included in the resection. Skin-sparing mastectomy Removal of the entire breast with preservation of its skin, except the nipple and the areola to facilitate immediate reconstruction. Skin sparing mastectomy should only be performed in the context of immediate reconstruction. When patient desires breast reconstruction, immediate reconstruction has advantages over delay: less procedures, better cosmesis and better patient satisfaction. Lymphatic mapping by sentinel node procedure SN biopsy should only be performed in patients with a clinically negative axilla. It has been shown convincingly that sentinel node can stage the axilla with high accuracy. Consensus on the sentinel node procedure states the following quality issues. The procedure should be performed within a team of surgeons, pathologists and nuclear physicians dedicated to breast pathology. The team should go through a learning phase of at least 20 procedures, including backup ALND. Newly trained surgical members of such a team can master the procedure under supervision without backup ALND. The sentinel node should be identified in over 90% of the patients. The long-term false negative rate should be < 5%. The preferred surgical technique is as follows. Just before sterile draping the blue dye is injected (1-4 cc; in or around the tumor, intracutaneously) The location of the SN is identified by scanning with the probe over the site where the SN's are suspected (and preferably indicated by the nuclear physician according to the lymphoscintigraphy) After massaging the breast (a few minutes), the incision is placed over the location were the sentinel node is suspected. If the sentinel node is in the axilla, the incision is usually placed behind the free edge of the major pectoral muscle at the lower border of the hair implant. The blue lymphatics can usually be Version / January, 2006

45 identified behind Scarpas fascia. The dissection should be meticulous, not to sever the lymphatics. If no blue lymphatics are identified, the probe can be used to identify the radioactive nodes. Radioactive and/or blue nodes are removed and afferent or efferent lymphatics secured. Internal mammary chain nodes can be approached either by a separate incision or through the same incision for mastectomy or wide local excision. The identification and removal of the SN is at the medial site of the intercostal space along the internal mammary artery. It requires careful dissection, but it is not a difficult technique. All nodes that are possibly SN are removed (if lymphoscintigraphy shows unequivocally non SN or second echelon SN, these hot nodes are not to be removed). Axillary Lymph Node Dissection (ALND) Traditionally, the indication for ALND is either to provide prognostic information on the basis of the nodal status and provide excellent regional control. If ALND is applied for this purpose, at least 10 lymph nodes should be retrieved and examined. For staging the axilla, the SN procedure appears to be equal to ALND. All incisions are acceptable, providing there has been sufficient exposure of the entire axillary content. In breast conservation cases the clearance should preferably be performed discontinuously from the breast excision using transverse incisions. All axillary fat from at least levels I and II should be excised in one specimen; The medial border is formed by a curved plane ranging from the muscles to the vessels and nerves going to the pectoral muscles (in the interpectoral area, the interpectoral fat, which might contain nodes, should be dissected carefully from this neurovascular bundles). The medial line is further indicated by a sagittal plane through the medial border of the m. pectoralis minor (patient in supine position). The cranial border is formed by plexus and axillary vein; The lateral border is from the white tendon downward to the m. latissimus dorsi; The dorsal border is the fascia of subscapular muscles; the n. thoracodorsalis and vessels may be spared, but should be resected in cases with suspicious nodes fixed to these structures; Caudally the upper-outer quadrant of the breast; care is given to resect the fat between thoracic wall and upper-outer quadrant of the breast; resection of parts of breast tissue might therefore be necessary; Level III axillary fat (apical, subclavicular) may be resected, en bloc with the specimen or separately for staging purposes. 5.2 Adjuvant radiotherapy Studies have reported similar rates of loco-regional recurrence in patients undergoing mastectomy without radiotherapy and patients undergoing complete tumor excision without mastectomy followed by radiotherapy to the breast. Radiotherapy to the breast is mandatory in all patients whose primary tumor surgery comprises a complete local excision without mastectomy. Radiotherapy is not mandatory for patients who have undergone a mastectomy. Tumor bed irradiation is mandatory for patients with non-resectable deep margin invasion. In order to minimize cardiac irradiation, all patients who receive internal mammary node (IMN) irradiation must have the radiotherapy planned using a technique that enables minimization of cardiac irradiation, preferably 3-D conformational CAT scan. This radiotherapy field is mandatory in patients with confirmed or suspicious IMN spread (by sentinel node technique) and optional in Version / January, 2006

46 patients with tumors judged to be at high risk of IMN spread (such as central and medial hemisphere tumors). 6 Chemotherapy: therapeutic regimens, expected toxicity, dose modifications 6.1 Treatment plan, anthracycline arm The anthracycline arm will include a standard three or four-drug anthracycline-based regimen, with either doxorubicin or epirubicin, chosen from the list of permitted anthracycline-based regimes (see below) and provided according to standard of care. Cycles should be given every 3 to 4 weeks, depending on the choice of treatment. The following are accepted regimens: CAF (d1+8), FAC (cyclophosphamide, doxorubicin and 5- fluorouracil), FEC (cyclophosphamide, epirubicin and 5-fluorouracil), CEF (d1+8) or four cycles of full-dose, single-agent epirubicin followed by 4 cycles of cyclophosphamide, methotrexate, and fluorouracil (CMF) called E-CMF. Despite their differences, these regimens have in common a high and adequate dose-intensity of anthracycline and enable various collaborating groups with different policies to participate in the trial Drug information, anthracycline arm Common toxic effects of the above mentioned regimens include nausea, vomiting, myelosuppression, febrile neutropenia, hemorrhagic cystitis (in schemes using cyclophosphamide), fever and infection, alopecia, mucositis, fatigue, local reaction (in the site of injection), anorexia, diarrhea, amenorrhea (in premenopausal women) and serum biochemistry abnormalities. More serious longer term side effects are cardiotoxicity and leukemia. For cardiotoxicity, the clinical presentation can vary from an asymptomatic decrease in the left ventricular ejection fraction to clinically overt congestive heart failure. It is directly related to the cumulative dose administered, with an incidence of 1-2% at a cumulative dose of 400 mg/m² of doxorubicin, rising exponentially above a total dose of mg/m², when the incidence may be as high as 26%. Epirubicin is associated with a lower frequency of cardiotoxicity at therapeutic dosages in comparison with doxorubicin (i.e., a reported incidence of 2-3% at 900 mg/m² epirubicin). Associated risk factors include underlying cardiac disease, prior or concomitant radiotherapy to the mediastinal area and advanced age. The utilization of modern techniques of radiotherapy seems not to add to this risk substantially. The incidence of secondary acute leukemia, another potentially serious long-term side effect, is also uncommon and ranges from 1-2% at 5 to 10 years of follow-up. It should be noted that the dose of anthracyclines used in the trial is kept well below the maximal tolerated doses around 360 mg/m² for doxorubicin and 600 mg/m² for epirubicin Drug administration, anthracycline arm The following are the accepted regimens and doses that can be used in the trial: FEC 100: Fluorouracil 500 mg/m², Epirubicin 100 mg/m², Cyclophosphamide 500 mg/m², all i.v. every 3 weeks for 6 cycles (Ref. 85) Canadian CEF: Cyclophosphamide orally 75 mg/m² d1-d14, Epirubicin 60 mg/m² i.v. d1 and d8, Fluorouracil 500 mg/m² iv d1 and d8, every 4 weeks for 6 cycles; if oral cyclophosphamide is not tolerated, it may be switched to i.v. doses of 370 mg/m² on days 1 and 8 (Ref. 41) Version / January, 2006

47 FAC: Cyclophosphamide 500 mg/m² d1, Doxorubicin 50 mg/m² i.v. d1, Fluorouracil 500 mg/m² iv d1, every 3 weeks for 6 cycles (Ref. 86) CAF: Cyclophosphamide 600 mg/m² d1, Doxorubicin 60 mg/m² i.v. d1, Fluorouracil 600 mg/m² iv d1 & d8, every 4 weeks for 6 cycles (Ref. 87) E CMF: Epirubicin 100 mg/m² d1 every 3 weeks for 4 cycles followed by either classical CMF (Cyclophosphamide orally 100 mg/m² d1-d14, methotrexate 40mg/m² i.v. d1 and 8 and 5- fluorouracill 600 mg/m² i.v. d1 and 8) every 4 weeks for 4 cycles or i.v. dose modified CMF (750 mg/m², 50 mg/m², 600 mg/m², days 1) every 3 weeks for 4 cycles (Ref. 88) Each treatment will be given as scheduled (3 or 4 weeks) ± 3 days. 5-Fluorouracil may be administered as an i.v. bolus over 15 minutes or less; Epirubicin as an i.v. infusion over 1 hour; Doxorubicin as an i.v. bolus in 10 to 15 minutes, Cyclophosphamide as an i.v. infusion over 1 hour or less and Methotrexate as an i.v. bolus. The first cycle of chemotherapy should be administered within 4 weeks following the randomization R-C date and ideally no more than 6 weeks after surgery. If treatment has not started within 8 weeks of surgery the patient becomes ineligible. The chemotherapy doses will be calculated according to baseline body surface area (BSA) for all cycles. If there is a 10% or greater decrease in body weight compared to baseline, the BSA will be recalculated. No ideal body weight should be used for the calculation of BSA. Dose reduction and/or treatment delay and treatment discontinuation are planned for the 2 arms in case of severe hematological and/or non-hematological toxicities. Duration of therapy: Chemotherapy will be given for 6 cycles or 8 cycles for E-CMF, which roughly corresponds to 5 to 6 months of treatment, depending on the chemotherapy scheme to be used. Anthracycline based Regimens FEC cycles Day Day 1 8 FEC Day 15 Day 22 CAF 6 cycles Day Day 1 8 CAF F Day 15 Day 22 Day 29 1 Cycle = 21 days 1 Cycle = 28 days Canadian CEF 6 cycles Day Day Day EF EF C C Day 22 Day 29 FAC 6 cycles Day Day 1 8 FAC Day 15 Day 22 1 Cycle = 28 days 1 Cycle = 21 days E CMF 8 cycles (4 x A followed by 4 x B) Day Day Day Day Day MF E x4 C Day 8 MF C Day 15 Day 22 Day 29 (1 Cycle A = 21 days) x 4 F = 5 Fluorouracil E = Epirubicin C = Cyclophosphamide A = Doxorubicin M = Methotrexate (1 Cycle B = 28 days) x 4 OR Day Day Day Day CMF (1 Cycle B = 21 days) x 4 Version / January, 2006

48 6.1.3 Premedication, anthracycline arm A prophylactic antiemetic treatment is highly recommended and should be given according to standard of care in each center. No prophylactic antibiotics are recommended (except with Canadian CEF). No primary prophylaxis with G-CSF/GM-CSF/PEG-filgrastim is allowed Patient monitoring anthracycline arm The usual pattern of care is required during infusion, including careful attention to reactions and side effects Dose adjustments anthracycline arm Full blood count will be performed on day 1 of the cycle and dose adjustments will be based on these results (and also based on clinical adverse events or findings observed during the study). Doses will be reduced for hematological and other adverse events (see table 3 & 4 below). Adverse events will be graded using the NCI Common Terminology Criteria for Adverse Events Version 3.0 (CTCAE) (see Appendix D). No dose modifications should be done for patients who experience grade I toxicities or other toxicities considered unlikely to become serious or life threatening. Dose modifications should be done based on the initial doses used and according to the greatest toxicity observed. Once done, they cannot be re-escalated. Treatment should be interrupted (for a maximum of 2 weeks) and the doses of drugs reduced by 25% in patients who experience a second occurrence of a given grade II toxicity (except alopecia) or any grade III toxicity. For patients who have received no trial drugs for 3 weeks due to toxicity, trial chemotherapy treatment should be stopped. Version / January, 2006

49 Table 3 Dose modification for Hematological Toxicities Anthracycline-based regimens Toxicity Treatment of Modification of chemotherapy toxicity ANC < 1.0x10 9 /l on the day of treatment None Wait until ANC > 1.0x10 9 /l (maximum time 3 weeks), then use full dose; G-CSF or GM-CSF or PEG-Filgrastim may be used in subsequent cycles if allowed by the countries registration/reimbursement rules Febrile Neutropenia According to standard care in institute G-CSF or GM-CSF or PEG- Filgrastim with subsequent cycles 2 nd episode of Febrile Neutropenia According to standard care in institute Reduce dose of anthracycline (and accompanying drugs) by 25% 3 rd episode of Febrile According to standard Discontinue chemotherapy treatment Neutropenia Platelets < 75x10 9 /l on the day of treatment Platelets < 50x10 9 /l on the day of treatment care in institute None Wait until platelet count > 75x10 9 /l and reduce dose of anthracycline (and accompanying drugs) by 25% None Wait until platelet count > 75x10 9 /l and reduce dose of anthracycline (and accompanying drugs) by 25% Platelet transfusion Wait until platelet count > 75x10 9 /l and reduce dose of anthracycline (and accompanying drugs) by 25% Version / January, 2006

50 Table 4 Dose modification of Anthracycline-Regimens for Non-Hematological Toxicities Toxicity Grade Treatment of toxicity Change to chemotherapy Nausea Any Local guidelines antiemetics and/or corticoid Vomiting Any Local guidelines antiemetics and/or corticoid Diarrhea III-IV Stop treatment until grade I, then change dose Mucositis III-IV Stop treatment until grade I, then change dose Dysphagia III-IV Stop treatment until grade I, then change dose Cystitis (If using Cyclophosphamide) Other I-II III-IV III IV Careful follow-up plus orientation Stop treatment until grade I, then change dose Stop treatment until grade I, then change dose Stop treatment until grade I, then change dose No modification No modification 25% dose reduction (Anthracycline) 1st occurrence- 25% dose reduction (anthracycline) 2nd occurrence discontinue chemotherapy treatment 25% dose reduction (Anthracycline) No change 1st occurrence- 25% dose reduction (cyclophosphamide) 2nd occurrence discontinue chemotherapy treatment Reduce 25% (anthracycline and accompanying drugs) 1st occurrence- 25% dose reduction (anthracycline) 2nd occurrence discontinue chemotherapy treatment Cardiotoxicity Patients should be followed according to local practice for determining and following any cardiac toxicity. Cardiac toxicity events will be recorded according to the CTCAE guidelines. Version / January, 2006

51 6.2 Treatment plan docetaxel-capecitabine arm Drug information docetaxel-capecitabine arm Docetaxel When used as a single agent, the major toxic effect of docetaxel is neutropenia. Bone marrow suppression is dose-dependent and is the dose-limiting toxicity. It may be associated with fever (febrile neutropenia). Other toxic effects include anaphylactoid type reactions and cutaneous reactions, nausea, vomiting, oral mucositis, diarrhea, reversible paresthesia, alopecia, asthenia, and mild local venous reactions (phlebitis) at site of injection and fluid retention/edema. Reversible skin reactions characterized by localized erythema of the extremities followed by desquamation may occur within 1 week of the infusion but are rarely severe. Their incidence is almost negligible with the proposed premedication. Severe nail reactions, such as onycholysis, occur in less than 1% of patients. Alopecia occurs in approximately 40-80% of patients. Edema may occur, with several forms of presentations and varying degrees of intensity (mild to poorly tolerated peripheral edema, generalized edema, pleural effusion, cardiac tamponade, or pronounced abdominal distention due to ascites). It usually starts in the lower extremities and may ascend to become generalized. This severe form of presentation may occur in up to 6% of the patients and the treatment consists of salt restriction and diuretics if needed. The fluid overload is reversible. Neurosensory symptoms consist mainly of distal paresthesia, dysesthesia and/or pain and may be severe in approximately 5% of the patients; the dosage of the drug must be adjusted when these symptoms appear and if they continue or worsen, the treatment must be interrupted. Peripheral motor neuropathy occurs in less than 5% of the patients and manifests as distal extremity weakness. Anaphylaxis and severe hypersensitivity reactions characterized by bronchospasm/dyspnea and/or hypotension requiring treatment, angioedema, and generalized urticaria are the most troublesome adverse effects and have occurred in 2% of patients treated in previous large clinical trials. Although variable and possibly affecting any organ system, these toxicities usually do not cause a large number of patients to cease treatment. The drug is contraindicated in patients who have a history of severe hypersensitivity reactions to drugs formulated with polysorbate 80 and should be used according to the guidelines in patients with elevated baseline liver function tests. Non-compliance with these guidelines puts the patient at risk for docetaxel induced toxic death Capecitabine Capecitabine is foreseen as an outpatient treatment, and in certain circumstances adverse events that could occur, such as diarrhea, can rapidly become serious. In the case where a patient experiences any toxicity in between scheduled visits, the patient should be encouraged to contact the clinic as soon as is practical, for further directions or for treatment. It is essential that the patients are informed to interrupt capecitabine treatment as soon as a grade II toxicity occurs, therefore the patients will need specific explanations what to do in the case of the occurrence of the most frequent toxicities (diarrhea, hand-foot syndrome and stomatitis). Renal Impairment: Capecitabine is contraindicated in patients with moderate or severe renal impairment (creatinine clearance below 50 ml/min [Cockroft and Gault]); treatment should not be started or continued in patients with severe renal impairment. Patients with moderate or severe renal impairment at baseline will be ineligible for the study, as per the exclusion criteria. The incidence of Version / January, 2006

52 grade III-IV adverse events in capecitabine (1250 mg/m 2 twice-daily, intermittent therapy) treated patients with moderate renal impairment (creatinine clearance ml/min [Cockroft and Gault]) was increased compared to the overall treated population. Careful monitoring and prompt treatment interruption is recommended if patients develop grade II, III, or IV adverse events with subsequent dose adjustments as outlined in the does modification table 8. Coagulopathy: Patients receiving concomitant capecitabine and oral coumarin-derived anticoagulants should have their anticoagulant response (INR or prothrombin time) monitored frequently in order to adjust the anticoagulant dose accordingly. A clinically important capecitabine-warfarin drug interaction was demonstrated in a clinical pharmacology trial. Altered coagulation parameters and/or bleeding, including death, have been reported in patients taking capecitabine concomitantly with coumarin-derived anticoagulants such as warfarin and phenprocoumon. Postmarketing reports have shown clinically significant increases in prothrombin time (PT) and INR in patients who were stabilized on anticoagulants at the time capecitabine was introduced. These events occurred within several days and up to several months after initiating capecitabine therapy and, in a few cases, within one month after stopping capecitabine. These events occurred in patients with and without liver metastases. Age greater than 60 and a diagnosis of cancer independently predispose patients to an increased risk of coagulopathy. The use of low molecular weight heparin instead of coumarin is at the discretion of the investigator. Diarrhea: Capecitabine can induce diarrhea. Patients with severe diarrhea should be carefully monitored and given fluid and electrolyte replacement. If grade II, III or IV diarrhea occurs, administration of capecitabine should be immediately interrupted until the diarrhea resolves or decreases in intensity to grade 1 (and see guidelines in section 6.2.5). Hand-Foot Syndrome: Capecitabine has been shown to cause hand-foot syndrome (palmar-plantar erythrodysesthesia or chemotherapy-induced acral erythema-erythema) (Ref. 89;90) which is characterized by varying degrees of redness, tenderness, pain and desquamation of the palms of the hands and the soles of the feet; numbness and tingling may also appear and in its most severe form it can interfere with daily activities. Nevertheless, it is reversible and responds to the diminution of the dose or the interruption of the treatment. If grade II or III hand-foot syndrome occurs, administration of capecitabine should be interrupted until the event resolves or decreases in intensity to < grade I. Subsequent doses of capecitabine should be administered as per the does modification table 8. Use of vitamin B6 pyridoxine ( mg bid) is permitted for symptomatic or secondary prophylactic treatment of hand-foot syndrome. Cardiotoxicity: There has been cardiotoxicity associated with fluorinated pyrimidine therapy (including capecitabine) including myocardial infarction, angina, dysrhythmias, cardiogenic shock, sudden death and electrocardiograph changes. These adverse events may be more common in patients with a prior history of coronary artery disease. Hyperbilirubinemia: Administration of capecitabine should be interrupted if treatment-related elevations in bilirubin of 3.0 x upper limit of the normal (ULN) or treatment-related elevations in hepatic aminotransferases (ALAT, ASAT) of 2.5 x ULN occur. Treatment may be resumed when bilirubin decreases to 3.0 x ULN or hepatic aminotransferases decrease to 2.5 x ULN. Pregnancy: Female patients must be instructed to stop taking capecitabine if they become pregnant during the study, and immediately inform the Investigator. The Investigator should counsel the patient, and discuss the risk of continuing with the pregnancy and the possible effects on the fetus. Monitoring of the patient should continue until conclusion of the pregnancy. The Investigator should report all pregnancies in the study to the Sponsor. Contraindications: Capecitabine is contraindicated in patients with severe leukopenia, neutropenia, or thrombocytopenia, severe hepatic impairment, or severe renal impairment (creatinine clearance below 30 ml/min). Use of capecitabine is contraindicated during pregnancy Version / January, 2006

53 and lactation, and concomitantly with sorivudine or its chemically related analogues, such as brivudine. Capecitabine is generally well tolerated. The most common side effects are hand-foot syndrome and diarrhea which generally respond to changes in the treatment dose or schedule. Other side effects associated with the use of capecitabine are: nausea, vomiting, stomatitis, dyspepsia/abdominal pain, fatigue, anorexia, myalgia, insomnia, fever, myelosuppression, paresthesia, eye irritation and serum biochemistry alterations. Other skin toxicities apart from the hand-foot syndrome include rash, dry and/or itchy skin and are mainly grades I and II Docetaxel-capecitabine combination When used in combination with docetaxel, the incidence of neutropenia/neutropenic fever, gastrointestinal side effects and hand-foot syndrome may be higher, but still manageable with appropriate medical intervention, dose reduction and/or treatment interruption, as seen in the already mentioned phase III trial in the metastatic setting. In spite of the apparent increased toxicity, the measured Quality of Life (QoL) was not adversely affected. Docetaxel Capecitabine Regimen 6 cycles Docetaxel Day 1 Doc Day 8 Day 15 Day 22 Capecitabine Day Day Day Cap Cap Rest Day 22 1 Cycle = 21 days Drug administration docetaxel-capecitabine arm The combination docetaxel-capecitabine will be given for 6 cycles which will roughly correspond to 5 months of treatment. Docetaxel 75 mg/m² should be administered as a 1-hour i.v. infusion, on day 1. Capecitabine 825 mg/m² will be given p.o., twice a day (total daily dose 1650 mg/m²) from days 1 to 14. Treatment will be given every 3 weeks (± 3 days) for a total of 6 cycles. For capecitabine, the daily dose will be derived by determining the body surface area (BSA). Use the tables of capecitabine doses in Table 5 to determine the appropriate dose for a given BSA. If body weight varies during the study, it is assumed that the body surface area will remain approximately constant (i.e. no dose adjustments for changes in body weight will be done). Capecitabine (Xeloda ) will be provided as film-coated 150 mg and 500 mg tablets, packed in polyethylene bottles containing 56 or 120 tablets. Tablets are not scored and should not be split. Capecitabine will be taken as 825 mg/m 2 doses twice-daily (total daily dose of 1650 mg/m 2 ) morning and evening (3-week cycles consisting of 2 weeks of capecitabine treatment followed by 1 week without capecitabine treatment). At the discretion of the patient and the Investigator, one of two schedules can be used. The first schedule allows for the first dose of each cycle to be administered in the morning of day 1 and the last dose of each cycle to be administered in the evening of day 14, followed by a seven day rest period. This provides for a total of 28 single doses per cycle over 14 calendar days. The second schedule allows for the first dose of each cycle to be administered in the evening on day 1 and the last dose of each cycle to be administered on the morning of day 15, followed by a 6.5 day rest period. This provides for a total of 28 single doses Version / January, 2006

54 per cycle over 15 calendar days. Either schedule may be used as long as the patient takes the required 28 single doses over consecutive days. The patient does not need to use the same schedule for all cycles, but may choose the other schedule for the next cycle. For example, on cycle 1, the patient visits the clinic in the morning and starts capecitabine in the evening. The patient's last dose is on the morning of day 15. For cycle 2, the patient takes a dose in the morning of day 1 and takes the last dose on day 14 in the evening. The morning and evening dose should be given approximately 12 hours apart, and taken within 30 minutes after the ingestion of food with approximately 200 ml of water, ideally after the breakfast and evening meal. The patient will be given a sufficient supply of study medication to accommodate 14 days (28 doses) of capecitabine treatment every 3 weeks. Table 5 Pill combinations to be taken to deliver the correct dose per ranges of BSA based on a daily posology of 1650 mg/m 2 100% Dose Level Number of tablets to be taken in the = twice daily 825 mg/m 2 Morning Evening Surface Area (m 2 Total Dose per ) Administration (mg) 150 mg 500 mg 150 mg 500 mg < Premedication docetaxel-capecitabine arm Following the standard of care, prophylactic steroids should be given before docetaxel infusion; a scheme with oral steroids such as dexamethasone 16 mg per day (e.g., 8 mg BID) is suggested, for 3 days starting the day before docetaxel administration, in order to reduce the incidence and severity of fluid retention as well as the severity of hypersensitivity reactions. The following schedule is recommended; dexamethasone 8 mg p.o. for 6 doses given: the night before chemotherapy, the morning of chemotherapy, 1 hour before docetaxel infusion (this dose may be given intravenously), the night of chemotherapy, the morning of the day after chemotherapy and the night of the day after chemotherapy. If the patient does not take the 8 mg p.o. dexamethasone the morning before and/or the night before the scheduled dose of docetaxel, she can not be treated as scheduled. Table 6 Prophylactic regimen for docetaxel-capecitabine arm 3 day schedule Steroid: Dexamethasone 8 mg i.v. or i.m. or orally, or Methylprednisolone 40 mg i.v. or i.m. or 32 mg orally, or Prednisone 50 mg i.v. or i.m. or orally, or Prednisolone 50 mg i.v. or i.m. or other equivalent For 6 doses 1. night before chemotherapy (- 12 h) 2. the morning of chemotherapy (- 2 h) 3. 1 hour before docetaxel infusion (- 30 min) Additional 3 doses (+12h, +24h, +36h) Observation: It is advisable not to use doses of dexamethasone (or its equivalents) greater than 30 mg/day but additional oral doses of dexamethasone may be used according to local policy. No prophylactic antibiotics are allowed. Hematopoietic growth factors (i.e., G- or GM-CSF) may be used according to institutional guidelines to treat febrile neutropenia, but should not be used as primary prophylaxis. Use of any Version / January, 2006

55 growth factor support must be documented in the patient records. Growth factors must be discontinued at least 48 hours prior to initiation of the next capecitabine cycle Patient monitoring docetaxel-capecitabine arm No special monitoring, standard care, as with patients on the anthracycline arm. Attention must be given to the possible occurrence of hypersensitivity reactions Dose adjustments docetaxel-capecitabine arm Full blood count will be performed on day 1 of the cycle and adjustments will be based on these results (and also based on the guidelines in tables 7 & 8 below). No changes should be done if patients experience grade I toxicities. Dose modifications should be done based on the initial doses used and according to the greatest toxicity observed. Once done, they cannot be re-escalated. Treatment should be interrupted (for a maximum of 2 weeks) and the doses of drugs reduced by 25% in patients who experience a second occurrence of a given grade II toxicity (except alopecia) or any grade III toxicity. Docetaxel therapy should be discontinued if toxicities do not resolve to grade 0 or I within 3 weeks; in that case the patient should be withdrawn from the chemotherapy part of the study and be treated at the discretion of the attending physician. Capecitabine should be resumed at 75% of the starting dose on resolution of the toxicity to grade 0 or I. Capecitabine dose reduction should be adapted following the recommendations in table 9. If the calculated creatinine clearance decreases during treatment to a value of 50 ml/min (due to an increase in serum creatinine or decrease in body weight), this change is not, by itself, a reason for a dose reduction. Dose reductions during treatment should be based on adverse events. For any adverse event apparent at baseline, the dose modifications will apply according to the corresponding shift in toxicity grade, if the Investigator feels it is appropriate. For example, if a patient has grade I asthenia at baseline which increases to grade II during treatment, this will be considered a shift of 1 grade and treated as a grade I toxicity for dose modification purposes however this is limited to changes up to a maximum of grade II. For toxicities which are considered by the Investigator unlikely to develop into serious or life threatening events (e.g. alopecia, altered taste etc.), treatment will be continued at the same dose without reduction or interruption. In addition, no dose reductions or interruptions will be required for anemia (non-hemolytic) as it can be satisfactorily managed by transfusions. If any grade I toxicity or any grade of alopecia occurs, treatment will be continued at the original dose without interruption. If in the opinion of the Investigator, a toxicity is considered due solely to one drug (e.g. hand-foot syndrome caused by capecitabine) the dose of the other drug does not require modification. Capecitabine treatment interruptions are regarded as lost treatment days and the planned treatment schedule should be maintained. Missed doses due to treatment interruptions should not be replaced (see Appendix I, Capecitabine schedule and interruptions to capecitabine schedule). If toxicity requires a dosing delay or interruption of both drugs of more than three weeks, the patient will be withdrawn from the chemotherapy part of the study for toxicity reasons. Where several toxicities with different grades or severity occur at the same time, the dose modifications applied should be the greatest reduction applicable. Once a dose has been reduced it should not be increased at a later time. Version / January, 2006

56 If both docetaxel treatment delay or capecitabine treatment interruption is indicated, then the treatment should be delayed or interrupted. Table 7 Dose modification for Hematological Toxicities - Docetaxel-Capecitabine combination Toxicity Treatment of toxicity Modification of chemotherapy ANC < 1.0x10 9 /l on day of infusion (day 1 of cycle) None Wait until ANC > 1.0x10 9 /l (maximum time 3 weeks), then use full dose G-CSF or GM-CSF or PEG-Filgrastim may be used in subsequent cycles Febrile Neutropenia 2 nd episode of Febrile Neutropenia 3 rd episode of Febrile Neutropenia Platelets < 75x10 9 /l on the day of treatment Platelets < 50x10 9 /l on the day of treatment Platelet transfusion According to standard care in institute According to standard care in institute According to standard care in institute None None G-CSF or GM-CSF or PEG-Filgrastim with subsequent cycles Reduce dose of docetaxel & capecitabine by 25% Discontinue chemotherapy treatment Wait until platelet count > 75x10 9 /l and reduce dose of docetaxel & capecitabine by 25% Wait until platelet count > 75x10 9 /l and reduce dose of docetaxel and capecitabine by 25% Wait until platelet count > 75x10 9 /l and reduce dose of docetaxel & capecitabine by 25% Version / January, 2006

57 Table 8 Dose modification of docetaxel-capecitabine for non-hematological toxicities Toxicity Grade Treatment of toxicity Change to chemotherapy Bilirubin I Wait until within normal limits, then change dose Reduce docetaxel by 25% Alkaline III-IV Wait until Grade II, then Reduce docetaxel by 25% Phosphatase change dose ASAT or ALAT III-IV Wait until Grade II, then Reduce docetaxel by 25% change dose Alkaline II Change dose Reduce docetaxel by 25% Phosphatase and ASAT or ALAT Edema Neurologic Any II-III No dose change; further treatment at physician discretion Wait until subsequent cycle; if I, no change If there is no resolution Reduce docetaxel by 25% IV Discontinue chemotherapy treatment Arthralgia/Myalgia Any Symptomatic treatment Skin rash Any Symptomatic treatment Asthenia III Symptomatic treatment 1 st occurrence- reduce docetaxel by 25% 2 nd occurrence reduce capecitabine by 25% Nausea Any Local guidelines No modification antiemetics and/or corticoid Vomiting Any Local guidelines antiemetics and/or corticoid No modification Hand-foot syndrome II-III Stop capecitabine until grade I, then change dose 1 st occurrence reduce capecitabine by 25% 2 nd occurrence reduce capecitabine by a further 25% (to 50% of initial dose) 3 rd occurrence - stop docetaxel & capecitabine IV Same as for grades II-III 1 st occurrence reduce capecitabine by 50% 2 nd occurrence stop docetaxel & capecitabine Version / January, 2006

58 Table 8 - Dose modification of docetaxel-capecitabine for Non-Hematological Toxicities Continued Toxicity Grade Treatment Change Diarrhea I None Mucositis Dysphagia II-III IV I-II III-IV III IV Stop capecitabine until grade I, then change dose and treat diarrhea Stop capecitabine until grade I, then change dose and treat diarrhea Stop capecitabine until grade I, then change dose Stop capecitabine until grade 1, then change dose Stop capecitabine until grade 1, then change dose Stop capecitabine until grade 1, then change dose Creatinine II No treatment; perform a sensitive test to ascertain the real value Other 1 st occurrence reduce capecitabine by 25% 2 nd occurrence reduce capecitabine by a further 25% (i.e. to 50% of initial dose) 3 rd occurrence - stop chemotherapy treatment 1 st occurrence reduce capecitabine and docetaxel by 25% 2 nd occurrence stop chemotherapy treatment 1 st occurrence reduce capecitabine by 25% 2 nd occurrence reduce capecitabine by a further 25% (i.e. to 50% of initial dose) 3 rd occurrence - stop chemotherapy treatment 1 st occurrence reduce capecitabine and docetaxel by 25% 2 nd occurrence stop chemotherapy treatment Reduce capecitabine by 25% Reduce docetaxel and capecitabine by 25% Stop capecitabine if Cr clearance 30 ml/min; if it increases to 30 ml/min, then resume it III Wait until I, then change dose Reduce capecitabine and docetaxel by 25% IV Stop treatment until grade I, then change dose 1 st occurrence- 25% dose reduction 2 nd occurrence stop chemotherapy treatment Version / January, 2006

59 Table 9 Pill combinations to be taken, twice daily, to deliver the correct dose reductions per ranges of BSA based on a starting daily posology of 1650 mg/m 2 75% Dose Level Number of tablets to be taken in the Morning Evening Surface Area (m 2 Total Dose per ) Administration (mg) 150 mg 500 mg 150 mg 500 mg < % Dose Level Number of tablets to be taken in the Morning Evening Surface Area (m 2 Total Dose per ) Administration (mg) 150 mg 500 mg 150 mg 500 mg < Recommendations for the management and treatment of delayed diarrhea The use of drugs with laxative properties should be avoided because of the potential for exacerbation of diarrhea. Patients should be advised to contact their physician to discuss any laxative use. Capecitabine can induce diarrhea, which can sometimes be severe. In patients receiving capecitabine monotherapy, the median time to first occurrence of grade II-IV diarrhea was 31 days, and median duration of grade III or IV diarrhea was 4.5 days. Patients with severe diarrhea should be carefully monitored and, if they become dehydrated, should be given fluid and electrolyte replacement. Prophylactic treatment No prophylaxis should be given, most of all no loperamide should be administered prophylactically. Loperamide and a fluoroquinolone have to be prescribed to the patients when they leave hospital. Adequate oral and written information about their use and about the management of diarrhea has to be properly given at this time, including the necessity of oral rehydration when diarrhea occurs. Patients should stop any laxative treatment and avoid food and beverages which might accelerate intestinal transit. Curative treatment If grade II, III or IV diarrhea occurs, administration of capecitabine should be immediately interrupted until the diarrhea resolves or decreases in intensity to grade I. Following grade II or higher diarrhea, subsequent doses of capecitabine should be decreased (see dose modification table for non-hematological toxicities table 8). As soon as the first liquid stool or abnormal bowel movement occurs, the patient must immediately start loperamide, two capsules, p.o., then one capsule, p.o. every two hours for at least 12 hours and up to 12 hours after the last liquid stool without exceeding a total treatment duration of 48 hours. Oral rehydration with large volumes of water and electrolytes is to be prescribed during the entire episode of diarrhea. If diarrhea persists for more than 48 hours despite the recommended loperamide treatment, the patient must be hospitalized for parenteral support. A 7-day prophylactic oral course of a fluoroquinolone antibiotic is to be started after medical advice. Loperamide will be replaced by Version / January, 2006

60 another antidiarrheal treatment (e.g. Octreotide 0.1 mg s.c. x 2/d for 3 days (see (Ref. 91) for further information) Patients experiencing any grade IV diarrhea, or diarrhea with grade III-IV neutropenia, or with fever or with vomiting should be hospitalized quickly for parenteral hydration and any other required treatment after adapted work-up. Capecitabine cannot be re-started until diarrhea has resolved to grade 0 I with the last loperamide dose given at least 24 hours beforehand. Anaphylactoid type reactions and hypersensitivity reactions If a hypersensitivity reaction occurs despite premedication, it is very likely to occur within a few minutes of the start of the first or the second infusion of docetaxel. Therefore, during the 1 st and the 2 nd infusions, the infusion must be given drop by drop for the first 5 minutes, and a careful evaluation of general sense of well being and, whenever possible, blood pressure and heart rate monitoring must be performed so that immediate intervention can occur in response to symptoms of an untoward reaction. Facilities and equipment for resuscitation must be immediately available: antihistamine, corticosteroids, aminophylline, epinephrine. If a reaction occurs, the specific treatment that is medically indicated for a given symptom (e.g. epinephrine in case of anaphylactic shock, aminophylline in case of bronchospasm, etc.) must be instituted. In addition, the following measures are recommended (Table 10): Table 10 Recommendations for management of hypersensitivity reaction Mild symptoms: Localized cutaneous reaction, such as pruritus, flushing, rash Moderate symptoms: any symptom not listed above (mildsymptoms) or below (severe symptoms), such as generalized pruritus, flushing, rash; dyspnea, hypotension with systolic B.P. > 80 mm Hg Severe symptoms: such as: bronchospasm, generalized urticaria, hypotension with systolic B.P. 80 mm Hg, angioedema Anaphylaxis (CTCAEv3 grade IV reaction) - consider decreasing the rate of infusion until recovery of symptoms, stay at bedside - then, complete docetaxel infusion at the initial planned rate - At subsequent cycles, use the same premedication outlined in section stop docetaxel infusion. - give i.v. dexamethasone 10 mg (or equivalent) and i.v. diphehydramine 50 mg (or equivalent). - resume docetaxel infusion after recovery of symptoms. - At subsequent cycles, give i.v. dexamethasone 20 mg (or equivalent) and i.v. diphehydramine 50 mg (or equivalent) one hour before infusion, in addition to the premedication schedule. - Immediately stop infusion - Give i.v.diphenydramine 50 mg with or without dexamethasone and/or epinephrine as needed; monitor patient until resolution of symptoms - Resume docetaxel infusion after recovery of symptoms. - At subsequent cycles, give i.v. dexamethasone 20 mg (or equivalent) and i.v. diphehydramine 50 mg (or equivalent) one hour before infusion, in addition to the premedication schedule. Patients must be monitored carefully and preferably in an intensive care setting. OFF CHEMOTHERAPY TREATMENT Version / January, 2006

61 Fluid retention In case fluid retention occurs during treatment with docetaxel, the signs and symptoms should be graded according to CTC criteria. The weight will be recorded and followed as frequently as possible to document any weight gain, which could be related to edema. Recommended curative treatment for fluid retention Curative treatment should commence when signs and/or symptoms of fluid retention are observed, including weight gain from baseline grade I not otherwise explained and should be given according to standard care. The following treatment is recommended: Furosemide 20 mg p.o. once a day; potassium-sparing diuretics alone or in combination are also allowed. If the symptoms cannot be controlled adequately, i.e. worsening of the fluid retention or spread to another area, the dose of Furosemide should be increased to 40 mg. The clinical tolerance of the patient and the medical judgment of the Investigator will determine if it is in the patients best interest to continue or to discontinue the study chemotherapy drug. It is recommended, however, that patients with severe fluid retention should be withdrawn from chemotherapy. When it is unclear whether an effusion is disease-related or study-drug related, treatment should be continued until progressive disease is clearly documented, unless it becomes symptomatic Concomitant therapy, docetaxel-capecitabine arm The usual pattern of care is required during infusion, including careful attention to reactions and side effects. In addition, some guidelines regarding drug interactions should be observed: Capecitabine: Patients receiving concomitant capecitabine and oral coumarin-derivative anticoagulant therapy should have their anticoagulant response (INR or prothrombin time) monitored frequently in order to adjust the anticoagulant dose accordingly. Altered coagulation parameters and/or bleeding, including death, have been reported in patients taking capecitabine concomitantly with coumarinderivative anticoagulants such as warfarin and phenprocoumon. A PK interaction has been observed. The use of low molecular weight heparin instead of coumarin is at the discretion of the Investigator. Phenytoin Increased phenytoin plasma concentrations have been reported during concomitant use of capecitabine with phenytoin. Formal drug-drug interaction studies with phenytoin have not been conducted. Patients taking phenytoin concomitantly with capecitabine should be regularly monitored for increased phenytoin plasma concentrations and associated clinical symptoms. Allopurinol Interactions with allopurinol have been observed for 5-FU; with possible decreased efficacy of 5- FU. Concomitant use of allopurinol with capecitabine should be avoided. Antivirals and Antiprotozoals Capecitabine or 5-FU should not be administered together with the antiviral drug sorivudine or its chemically related analogues, such as brivudine. A clinically significant drug-drug interaction between sorivudine and 5-FU, resulting from the inhibition of DPD by sorivudine, has been described in the literature. This interaction, which leads to increased fluoropyrimidine toxicity, is potentially fatal. Metronidazole increased the toxicity of fluorouracil in patients with colorectal cancer, apparently by reducing the clearance of the antineoplastic. As it has been described in the literature, caution should be exercised. Version / January, 2006

62 Gastrointestinal Drugs Pretreatment with cimetidine for 4 weeks led to increased plasma concentrations of fluorouracil following intravenous and oral administration in six patients. The effect was probably due to a combination of hepatic enzyme inhibition and reduced hepatic blood flow. No such effect was seen following single doses of cimetidine in five patients or pretreatment for just one week in six. Care is required in patients taking both drugs simultaneously. Hematopoietic growth factors Hematopoietic growth factors (i.e., G- or GM-CSF) may be used according to institutional guidelines to treat febrile neutropenia, but should not be used as primary prophylaxis. Use of any growth factor support must be documented in the patient record. Growth factors must be discontinued at least 48 hours prior to initiation of the next capecitabine cycle. Bisphosphonates Patients receiving bisphosphonates are eligible for this study but should have bone scans (and X- rays of areas of enhanced uptake indicative of bone metastasis) at baseline. Those patients starting bisphosphonates during the study but without other clear evidence of disease progression will not be diagnosed as having progressive disease on that evidence alone. Docetaxel: In vitro studies have shown that the metabolism of docetaxel may be modified by the concomitant administration of compounds that induce, inhibit, or are metabolized by cytochrome P450 3A4, such as cyclosporine, terfenadine, ketoconazole, erythromycin, and troleandomycin. Caution should be exercised with these drugs when treating patients with docetaxel as there is a potential for a significant interaction. 6.3 Treatment withdrawal criteria The treatment should be withdrawn if: The patient, at any time, withdraws consent to participate. The Investigator judges that the decision is in the best interest of the patient The treatment must be interrupted for more than 3 weeks There is evidence of disease recurrence/relapse The patient becomes pregnant The patient is lost to follow-up The patient will then be considered off-protocol chemotherapy treatment, but should be followed according to the protocol 7 Endocrine therapy 7.1 Trial treatments Tamoxifen will be taken according to the standard of care, consisting of a 20 mg tablet orally once daily for 2 years from date of endocrine therapy randomization (R-E) for all patients randomized to receive tamoxifen. Letrozole will be provided free of charge for treatment within the study protocol by Novartis. It is to be taken as a 2.5 mg tablet, orally once daily for 7 years from date of endocrine randomization (R- E) for all patients randomized to the letrozole arm or for 5 years after an initial 2 years of tamoxifen for patients randomized to the tamoxifen arm. Version / January, 2006

63 7.2 Side effects of drugs Tamoxifen: The most common side effects reported are hot flushes, nausea, vaginal discharge or abnormal vaginal bleeding. Hypercalcemia, bone pain, and retinopathy are rare. Long term beneficial effects on lipid and bone metabolism have been reported in post menopausal women, but there may be an increased risk of endometrial cancer, polyps and hyperplasia associated with the estrogen agonist action of tamoxifen. In premenopausal women, bone mineral density loss may occur. Fluid retention and skin rash have been reported. Tamoxifen is known to increase the risk of thromboembolic disease, and a short period of cessation of treatment may be required should some women need to undergo minor or major surgery. Modification of tamoxifen dosage is rarely indicated. No standard dosage modifications are prescribed. Letrozole: In clinical trials, adverse events (AEs) were generally mild to moderate and rarely severe enough to require discontinuation of treatment. AEs that have been reported include: fatigue, chest pain, peripheral edema (swelling), high blood pressure, nausea, vomiting, constipation, diarrhea, abdominal pain, anorexia, viral infection, musculoskeletal pain (including arm pain, back pain, leg pain, skeletal pain), arthralgia, headache, dyspnea, coughing, hot flushes, and rash. Less frequent (< 5%) AEs reported include hypercalcemia, fracture, depression, anxiety, pleural effusion, hair thinning, increased sweating and vertigo. Other possibly drug-related AEs were: vaginal bleeding, leukorrhea, weight loss, thrombophlebitis, generalized edema, dizziness, changes in lymphocyte counts and increase in aminotransferase. Special emphasis should be given to problems associated with lipid and bone metabolism such as fractures, osteoporosis and hypercholesterolemia. Modification of letrozole dosage is rarely indicated. No standard dosage modifications are prescribed. 7.3 Ovarian function suppression All premenopausal women (until the age of 50 years) at diagnosis who decide to participate in the endocrine randomization in the trial must have adequate ovarian function suppression while taking letrozole. Each centre will decide, upfront, their policy regarding whether they will also provide ovarian function suppression for all of their premenopausal patients during the initial 2 years of tamoxifen treatment. Menopausal status should be confirmed by dosing estradiol, FSH and LH, or E2 if appropriate, in the case of a GnRH or bilateral ovarian irradiation. Ovarian function suppression can be achieved in several ways: The use of a GnRH analogue such as goserelin (Zoladex TM ), taken as a 3.6 mg subcutaneous injection every 28 days. NOTE that this product will NOT be supplied by the EORTC. GnRH analogues administered at 3 monthly intervals are not approved due to uncertainty concerning the duration of ovarian function suppression in women with this schedule. Bilateral surgical oopherectomy. Bilateral ovarian irradiation. Radiation should be given according to accepted guidelines. Biochemical verification of ovarian function suppression is required by 2 months using measurement of estradiol, FSH and LH. There are cases of failure of ovarian function suppression under GnRH analogues. In this case, ovarian function suppression should be achieved by other means, provided the patient accepts an alternative method. For women reaching the age of fifty while on GnRH there is the possibility, at the treating physician s discretion, of stopping the AI and the GnRH for at least three months and confirming menopausal status by dosing estradiol, FSH and LH. If menopausal status is confirmed endocrine treatment can be resumed with the AI alone. Version / January, 2006

64 7.4 Concomitant treatments Restriction of concomitant treatments: the following concomitant treatments are NOT ALLOWED during the endocrine therapy part of the trial: Any other anti-cancer therapy (anti-estrogens, aromatase inhibitors, chemotherapy) other than the primary endocrine treatment Other investigational agents whilst on study. Raloxifene or other SERMS Systemic HRT with or without progestins. A patient should go off trial if she receives more than 3 months of HRT. Hormonal contraceptives (including depot injections and implants) However intrauterine devices, including those progesterone coated ARE ALLOWED Any other hormonal treatments (either oral of transdermal) including estrogen, progesterone, androgens, and aromatase inhibitors. Topical estrogens are DISCOURAGED during the endocrine therapy part of the trial. However, for women with severe vaginal dryness/dyspareunia that does not respond to non-hormonal treatments, then a local vaginal treatment with minimal systemic absorption is allowed. Concomitant treatment with bisphosphonates IS ALLOWED (including on other clinical trial protocols) and should be recorded during the trial. This should be prescribed as recommended by the ASCO guidelines on the role of bisphosphonates and the use of AI (Ref. 92). The administration of vitamin D3 and calcium supplements is recommended. Considering the potential increased risk of osteoporosis in women in this study, patients should be advised about regular calcium intake and weight bearing exercise (see ASCO guidelines on bone care (Ref. 92)). 8 Clinical evaluation, laboratory tests and followup 8.1 Baseline evaluations Baseline evaluations that must be carried out prior to enrollment: Chest x-ray Bilateral mammography ECG (the pre-surgical ECG can be used provided it was normal) and within 21 days of the start of treatment for women receiving either chemotherapy or endocrine therapy: Clinical evaluations (physical examination, medical history & weight). Any abnormal or existing clinical condition should be reported on the baseline AE case report form. WHO performance status Routine Hematology (WBC, absolute neutrophils, hemoglobin, platelets) Routine blood chemistry (glucose, calcium, creatinine, total bilirubin, alkaline phosphatase, ALAT & ASAT) Version / January, 2006

65 CA 15-3 if standard of care in local institution For women receiving neither chemotherapy nor endocrine therapy the baseline analyses will be performed once and they will be followed as described in section Specifically to be done prior to chemotherapy randomization (R-C) To be performed within 21 days of randomization R-C and before the start of chemotherapy treatment. (see the summary table 11 below) Cardiac function (left ventricular ejection fraction (LVEF) within the normal limits of each institution, measured either by echocardiography or by radionuclide angiocardiography (MUGA)) according to standard of care in the local institution Specifically to be done prior to endocrine randomization (R-E) Patients should be fit for endocrine therapy, for patients who do not receive chemotherapy, baseline evaluations need only be done once prior to enrollment. For patients receiving adjuvant chemotherapy and going on to receive endocrine treatment in R-E, some baseline evaluations need to be repeated i.e. once at enrollment/r-c and then again for entry into R-E (details are given below and in summary table 11). To be performed within 21 days of randomization R-E and before the start of endocrine therapy treatment. Repeated tests for those patients who received chemotherapy: Clinical examination (a thorough clinical history should be taken, especially history of previous thromboembolism, cardiovascular or cerebrovascular events) Routine hematology (WBC, absolute neutrophils, hemoglobin, platelets) Routine blood chemistry (glucose, calcium, creatinine, total bilirubin, alkaline phosphatase, ALAT & ASAT), LH/FSH & E2 if appropriate (menopausal status should be determined on plasma estrogen levels) for women of childbearing potential. Baseline radiological evaluations should be repeated if clinically indicated, or in the case of liver and bone scan imaging, if biochemical blood tests are persistently abnormal after completion of chemotherapy. A gynecological examination is recommended at baseline in case of pelvic complaints. This may include a Pap smear, bimanual and recto-vaginal examination or vaginal ultrasound. A bone mineral density study (DEXA) should be performed at baseline for all women randomized in R-E if this is standard care in the local institution. Non-fasting cholesterol should be taken and recorded in the patients file if this is standard care in the local institution. 8.2 During treatment Chemotherapy The following evaluations will be performed on Day 1 (or within 3 days prior to the start) of each chemotherapy cycle. Pre-enrollment assessments will be used for Cycle 1. A brief physical examination, including vital signs, and weight. Version / January, 2006

66 WHO performance status. Routine hematology (WBC, absolute neutrophils, hemoglobin, platelets) Routine blood chemistry (glucose, creatinine, alkaline phosphatase, total bilirubin, and ALAT and ASAT) An adverse event assessment will be performed according to the NCI common terminology criteria (CTCAE) version 3.0 (see Appendix D) Endocrine therapy Visits should be every 3 months for the first two years, and every 6 months for years 3-7 and then once a year (note: laboratory tests should be adapted to standard of care in local institutions if one of the tests recommended is not routine practice) First 2 years every 3 months physical examination and medical history. Routine hematology, liver function test and CA 15-3 if standard of care in the local institution. every 6 months non-fasting cholesterol if standard of care in the institute every 12 months mammography. Chest X ray, gynecological assessment, bone mineral density study (DEXA) if standard of care in the institute Years 3-7 every 6 months physical exam and medical history. Routine hematology & liver function if standard of care in the institute tests every 12 months mammography. CA 15-3 if standard of care in the local institution, chest X ray (at the discretion of the investigator), gynecological assessment (at the discretion of the investigator), Bone mineral density study (DEXA) and nonfasting cholesterol if standard of care in the institute After Year 7 Every 12 months physical examination, medical history, and mammography. Routine hematology, liver function tests and CA 15-3 if standard of care in the local institution, chest X ray (at the discretion of the investigator), gynecological assessment (at the discretion of the investigator), bone mineral density study (DEXA) and non-fasting cholesterol if standard of care in the institute At each visit Dispense medication Adverse Event and follow-up forms should be completed and submitted. Report bisphosphonate treatment for osteoporosis, medication for hypercholesterolemia, and development of cardiac or cerebrovascular events for each visit. Other diagnostic tests (i.e.: abdominal ultrasound and/or CT scan, bone scan, MRI) should be performed only in the presence of signs and/or symptoms and/or blood test abnormalities suggestive of cancer recurrence. 8.3 After the end of treatment (Follow-up) The following follow schedules apply to all patients except those patients who have progressive disease with distant metastasis who will then be followed yearly for survival. Version / January, 2006

67 8.3.1 Follow-up for patients receiving neither chemotherapy nor endocrine therapy Follow-up for those patients who have received neither chemotherapy nor endocrine therapy will be done according to standard of care and consist of the following examinations to ideally be done every 3 months for the first 2 years then every 6 months for years 3-7 and every 12 months for subsequent years. Clinical evaluations (physical examination, medical history & weight) WHO performance status Routine Hematology, if standard of care in the institute Routine blood and biochemistry, if standard of care in the institute Yearly mammography Other procedures will be done as clinically indicated Chemotherapy follow-up All patients will be followed as described in section After local recurrence patients will continue to be followed for recurrence at other sites For patients receiving chemotherapy and endocrine therapy Follow-up will be done according to standard care in the local institute and will follow the scheme for endocrine therapy follow-up as described in section & summary table For patients receiving only chemotherapy Follow-up visits will be done according to standard care in the local institute and ideally every 3 months during the first 2 years, every 6 months during years 3-7 and then every 12 months. The assessments will be done according to standard care in the local institute and should follow the schedule described in section and summary table Endocrine therapy follow-up After the first seven years, disease status and survival data will be collected yearly for all patients. Annual follow-up to report disease status and survival will be requested for at least a minimum of 15 years. Survival data will be collected for all patients who have progressed with distant metastasis during or after trial treatment until death. Version / January, 2006

68 8.3.4 Summary table Table 11 - Schedule of Assessments Required Investigations Informed Consent Inclusion criteria Assess quality of tumor samples Demographics, medical history Pregnancy test 1 Physical examination/ Performance Status Prior to enrollment/ randomization Every chemotherapy cycle End of treatment Followup, Years 1 and 2 a a a a a Every 3 months Routine hematology a a a Biochemistry (glucose, calcium, creatinine, total Bilirubin, alkaline phosphatase, ALAT, ASAT) LH/FSH & E2 if appropriate CA Non fasting total cholesterol 3 a 2 Optional Optional Followup, Years 3 to 7 Every 6 months Chest X-ray a If clinically indicated Bone Scan a If clinically indicated Mammography a Once a year DEXA scan Following standard care ECG If clinically indicated Echo/MUGA scan a If clinically indicated Other Investigations As needed or clinically indicated (CT, PET scan, liver echo) Adverse Events Throughout the study Serious adverse events (SAE) 1 Only for patients of childbearing potential ² For all pre-menopausal women 3 If standard of care in the institute Legend to table Follow-up, Years 7 to 10 Yearly All endocrine therapy baseline assessments must be performed after the end of chemotherapy and before starting endocrine treatment. Non-fasting total cholesterol should be collected according to standard care at local institutions and preferably at baseline, every 6 months during the first 2 years, and yearly for the following 5 years. An additional sample should be collected one year after completion of the 7 year trial treatment period, at any time of premature trial treatment discontinuation and once more at the next scheduled 6-monthly visit following premature trial treatment discontinuation. If premature trial treatment discontinuation occurred within the 6 months prior to completion of the 7 year trial treatment period, the additional sample should be collected at the next scheduled yearly visit. Version / January, 2006

69 Radiological assessments Bone mineral density studies should be performed according to standard care at local institutions and preferably at baseline and yearly whilst on AI treatment. A bilateral mammography must be taken within 3 months prior to surgery. A mammography (and/or ultrasound) of the conserved and contra lateral breast should be performed at yearly intervals or should be done according to national standards or hospital specific requirements. A Chest X-ray should be performed according to standard of care in the institute and preferably yearly. Posterioranterior (PA) and lateral views are preferred however a PA alone will be accepted subject to local practice. A bone scan should be taken according to standard of care in the institute or if medically indicated. If the bone scan showed areas suspicious for tumor then these areas should be assessed by X-ray, CT or MRI. Metastatic disease should be confirmed by a second bone scan at least three months later that shows progressive changes. Abdominal ultrasound or liver scan or CT abdomen is required if medically indicated. It is mandatory in all patients with liver function tests or ALP that are above the limit of normal prior to randomization, after completion of any chemotherapy treatment. If after randomization, liver enzymes become abnormal, further staging is required to exclude metastatic disease (ie: Chest XR, liver imaging, bone scan, etc). Other procedures In the event of a pelvic complaint patients should have a gynecological examination because of increased risk of uterine cancer in patients receiving tamoxifen. Otherwise patients should be followed according to standard of care in the institute. Note: Any woman with abnormal/unexplained vaginal bleeding should have an endometrial biopsy and the results must be recorded on the Adverse Event Form during protocol therapy Note: Patients who discontinued trial treatment for any reason before the planned treatment duration of 7 years must continue to be followed every six months for assessment of disease status, cardiovascular, skeletal, and urogenital events, and for survival data collection. After 7 years all patients will be followed yearly for disease status, cardiovascular, skeletal, and urogenital events and survival status for a minimum of 15 years. 9 Criteria of evaluation 9.1 Time to event endpoints Special note: In this complex trial the date of enrollment/randomization used below in definitions of time to event endpoints (overall survival, disease free survival, distant metastasis free survival) depends upon the analysis and the dataset in which it is being analyzed. This is clarified in the statistical section Types of recurrence Recurrence will be categorized as local, regional or distant. Local (Breast) Recurrence is defined as recurrence within the breast. Local (Chest Wall) Recurrence is defined as recurrent cutaneous or subcutaneous tumor occurring in an area bounded superiorly by the clavicle, inferiorly by a horizontal line at the level of the xiphisternum, medially by the midline and laterally by the posterior axillary line. Regional (Nodal) Recurrence is defined as recurrent tumor in the lymph nodes in the homolateral axilla, homolateral supraclavicular fossa or ipsilateral internal mammary chain. Distant Metastatic Recurrence is defined as spread of disease beyond the limits specified above as local or regional recurrence. Contralateral breast cancer and secondary cancers are not recurrence, but will be considered as events for the endpoint of DFS, see section Version / January, 2006

70 9.1.2 Distant Metastasis Free Survival (DMFS) For the endpoint of DMFS only distant metastatic recurrences as defined above and deaths (any cause) will be taken as events. Contralateral breast cancer and secondary cancers will not be taken into account. DMFS is calculated as the time from enrollment/randomization to either the first date of distant metastatic recurrence or the date of death. The date of first documented distant metastasis (if applicable) will be used as the event date. Patients alive with no evidence of metastasis at the time of their last visit are censored at the time of the last examination Disease Free Survival (DFS) Definition of date of disease recurrence: date on which a clinically suspicious lesion is first recorded in the patient file provided action is taken as result of which the diagnosis of any type of recurrence is confirmed. Recurrence of disease can be a loco-regional recurrence, a distant (metastatic) recurrence or a second primary. As documented in Section 9.2.1, contralateral breast cancer and second malignancies will be considered to be events. Disease Free Survival (DFS) is calculated as the time from enrollment/randomization to either the date of disease progression or the date of death. The date of first documented disease recurrence (if applicable) will be used as the date of event. Patients alive with no evidence of disease recurrence at the time of their last visit are censored at the time of the last examination Overall Survival (OS) Overall Survival (OS) is calculated as the time from enrollment/randomization to the date of death (any cause). Patients still alive at the time of analysis are censored at the last time they are known to be alive. 9.2 Reporting of results All patients enrolled in the study must be followed for progression and survival, according to intent to treat, even if there are major protocol or treatment deviations or if they are found to be ineligible. 9.3 Evaluation of toxicity Reporting of and general evaluation of adverse events All adverse events will be recorded on the applicable case report forms This study will use the Common Terminology Criteria for Adverse Events v3.0 (CTCAE). A copy of the CTCAE can be accessed from the CTEP home page ( A link to this page is provided on the EORTC web site Investigators who do not have access to Internet can contact the EORTC Data Center to receive a copy by post. The actual laboratory values will be collected and grading according to CTCAE will be centrally applied for reporting Long-term chemotherapy toxicity The following potential long-term toxicities of chemotherapy will be monitored for a minimum of 15 years in all patients who were randomized for chemotherapy (R-Chemotherapy), cardiac toxicity and secondary leukemias. Version / January, 2006

71 9.3.3 Serious adverse events Serious adverse events are defined by the Good Clinical Practice Guideline. SERIOUS ADVERSE EVENTS SHOULD BE IMMEDIATELY REPORTED ACCORDING TO THE PROCEDURE DETAILED IN THIS PROTOCOL (see chapter on Reporting adverse events) 10 Statistical considerations 10.1 Statistical design Sample size In the set of patients who have a low risk gene prognosis signature using the 70-gene signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature prognosis and thus receive no chemotherapy, a null hypothesis of a 5-year distant metastasis free survival (DMFS) of 92% will be tested. With 6,000 patients accrued overall, this set has an expected size of 672 patients. With an accrual of 3 years, and a total duration of 6 years (so 3 to 6 years follow up for each patient), a one-sided test at 97.5% confidence level has 80% power to reject this hypothesis if the true 5-year DMFS is 95%. The primary test will be performed when three conditions are met: the standard error of DMFS at 5 years (obtained by estimating the standard error of log(-log(s(t)), and back-transformation) is 0.01 or less, at least one third of the patients in the above dataset have been followed for five years, and the number of events for the primary test for R-C has been reached. To perform the test, a 95% twosided confidence interval will be constructed for the DMFS at 5 years, using the log-log transform formula (Ref. 93), page 18). The test will be significant if the confidence interval does not include the value of The primary test for R-C is to compare DFS between the two chemotherapy regimens randomized. With an expected 4,000 patients in this section of the study, there is 80% power to detect a hazard ratio of 0.76 (for example a control 5-year DFS of 86% versus a 89.2% experimental 5-year DFS) with at least 422 events (deaths or recurrences among the patients randomized for R-C). The comparisons for R-C will be performed after at least 422 events have been observed for this comparison. After 211 events have been observed, an interim test will be performed for this comparison, using a rho spending function (Ref. 94) with rho=2.5, which is somewhat less conservative than an O Brien-Fleming spending function. If the number of events at the time of the interim test is exactly 211, the tests at interim and at final analysis will have nominal alpha values of and , respectively. If the true hazard ratio is 0.6, this interim test has 85% power of rejecting the null hypothesis. The primary test for R-E is to compare DFS between the two sequences of endocrine therapy randomized. With an expected 3,500 patients being randomized into this question, there is 80% power to detect a hazard ratio of 0.75 (for example a DFS rate at 5 years of 86% on the control arm versus an experimental 5-year DFS of 89.3%) with at least 382 events. After 191 events have been observed, an interim test will be performed for this comparison, also using a rho spending function with rho=2.5. If the number of events at the time of the interim test is exactly 191, the tests at interim and at final analysis will have nominal alpha values of and , respectively. If the true hazard ratio is 0.6, this interim test has 81% power. Version / January, 2006

72 Randomization and stratifications For eligible patients with adequate material for microarray analysis, both the 70-gene signature and clinical-pathological risk will be assessed, after which they can be enrolled. Patients will be centrally enrolled and randomized as applicable for each of the three possible randomizations (R-T, R-C, R-E, for practical details, see chapter on enrollment / randomization procedure). Patients are evaluated as low clinical-pathological risk, if their 10-year disease specific survival (without chemotherapy or endocrine therapy) is estimated by Adjuvant! Online (Ref. 95) as greater than 88% for ER-positive patients, and greater than 92% for ER-negative patients. Patients who are at high risk according to both methods will receive chemotherapy, and patients who are at low risk according to both methods will not receive chemotherapy. As new versions of Adjuvant! Online (Standard version) become available, these will be implemented. The version at the time of the writing of this protocol is Standard Version 8.0. R-T: Patients for whom both methods are discordant will be randomized between chemotherapydecision-making according to clinical criteria and chemotherapy-decision-making according to 70- gene signature prognosis. For R-T, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genetic/low risk clinical vs. low risk genetic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), age (<50 vs. >=50), HER-2 (Positive vs. Negative vs. Unknown), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). Patients who are candidates for chemotherapy (see also treatment section) will be proposed for R-C: randomize to anthracycline based chemotherapy or taxane/capecitabine chemotherapy. For R-C, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genetic/low risk clinical vs. low risk genetic/high risk clinical vs. high risk genetic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), age (<50 vs. >=50), HER-2 (Positive vs. Negative vs. Unknown (at the time of R-C)), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). All patients who are candidates for endocrine therapy will be offered randomization (R-E) to 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. For R-E, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genetic/high risk clinical vs. high risk genetic/low risk clinical vs. low risk genetic/high risk clinical vs. low risk genetic/low risk clinical), chemotherapy (no chemotherapy, chemotherapy without R-C, R-C arm A, R-C arm B), type of endocrine sensitivity (both ER and PgR positive vs. either ER or PgR positive), age (<50 vs. >=50), HER-2/neu (Positive vs. Negative vs. Unknown (at the time of R)), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). The three randomizations will be open-label Statistical analysis plan Primary and secondary endpoints In this complex protocol, time to event endpoints (DMFS, OS, and DFS) will start at different dates depending on the analysis: For all overall analyses, and analyses related to R-T, time to event endpoints will start at the date of enrollment of the patient. These are the analyses on the data sets AP, PT, ITT1 (see section ). Version / January, 2006

73 For all analyses related to R-C, time to event endpoints will start at the date of R-C. These are the analyses on the data set ITT2 (see section ). For all analyses related to R-E, time to event endpoints will start at the date of R-E. These are the analyses on the data set ITT3 (see section ). Using these conventions for starting dates, the definitions in section 9.2 will apply Analysis populations In this study, the following analysis datasets will be used. Because of the complexity of the study, there are many datasets, and so a code is added between brackets for ease of reference. All patients dataset (AP): All patients who were enrolled into the study. Primary test dataset (PT): The set of patients who have a low risk gene prognosis signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy. This dataset is used for the primary test. Intention-to-treat population for R-T (ITT1): All randomized patients for R-T will be analyzed in the arm they were allocated by randomization. Per protocol population for R-T (PP1): All patients who are eligible and randomized for R-T and have started their allocated treatment (at least one dose of chemotherapy if so randomized, respectively no chemotherapy). Intention-to-treat population for R-C (ITT2): All randomized patients for R-C will be analyzed in the arm they were allocated by randomization. Intention-to-treat population for R-E (ITT3): All randomized patients for R-E will be analyzed in the arm they were allocated by randomization. Safety population for R-C (SP2): All patients who were randomized for R-C and who have started their allocated chemotherapy (at least one dose of the study drug(s)). Safety population for R-E (SP3): All patients who were randomized for R-E and who have started their allocated endocrine treatment (at least one dose of the study drug(s)). A patient will be considered to be eligible if she did not have any major deviations from the patient entry criteria listed in chapter 3. Eligibility will be assessed by the Study Coordinator based on the review of each patient file Timing of analyses Interim analysis on the ITT2, SP2 datasets will be performed at the time at least 211 events for DFS have been observed on the ITT2 dataset. If that analysis is significant as described above, this will be the final analysis for the chemotherapy comparison. If not, the final analysis on the ITT2, SP2 datasets will be performed at the time at least 422 events for DFS have been observed on the ITT2 dataset. All analyses on the AP, PT, ITT1, PP1 datasets will be performed when the three conditions in section have been met. Interim analysis on the ITT3, SP3 datasets will be performed at the time at least 191 events for DFS have been observed on the ITT3 dataset. If that analysis is significant as described above, this will be the final analysis for the endocrine comparison. If not, the final analysis on the ITT3, SP3 datasets will be performed at the time at least 382 events for DFS have been observed on the ITT3 dataset. Version / January, 2006

74 All analyses are driven on the number of events observed, and so the timing depends on accrual, relative size of the appropriate analysis set, event rates, and loss to follow-up. Under the assumption of a homogeneous 3 year accrual, and under the alternative hypotheses stated above for the respective tests, the expected timing of the analyses is as follows: - interim test for chemotherapy (ITT2): 4 years after start of accrual - interim test for endocrine therapy (ITT3): 4.5 years after start of accrual - primary test on AP, analyses for ITT1, final test for chemotherapy (ITT2): 6.3 years after start of accrual - primary test for endocrine therapy (ITT3): 7 years after start of accrual Statistical methods General All statistical tests and confidence intervals will be two-sided. All statistical tests will be tests for superiority (difference). For tests of primary endpoints, the nominal alpha level will be as defined by the interim analysis procedure (see Section for an overview table). All secondary endpoints will have a 95% confidence level at the final analysis. All comparisons for time to event endpoints described below will be adjusted Logrank tests, as performed by estimating a Cox model with the effect under consideration, as well as the stratification factors used for the applicable randomization (with the exception of site). As there is no a priori reason to believe that the stratification factors used will exhibit non-proportional hazards, preference is given to adjusting for them by using them as covariates (not by stratified Cox model). Corresponding hazard ratios and confidence intervals will be calculated by the same Cox regression. Time to event comparisons will be accompanied by Kaplan-Meier curves. Baseline characteristics will include age, menopausal status, ER/PgR status, HER-2 status, tumor size, differentiation, WHO performance status, eligibility, method of axillary evaluation (sentinel only, dissection), type of surgery (mastectomy, quadrantectomy/tumorectomy). Analyses on AP dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable), once split by clinical pathological risk, and once by 70-gene signature risk. A separate table will be made to describe the types of chemotherapy and/or endocrine therapy, as well as radiotherapy administration (yes/no) patients received during the course of the trial, to show treatment of those patients who did not consent to or were not eligible for R-C and/or R-E, but still were given some treatment. This table will be on the AP dataset, with subcategories according to R- C (arm 1, arm 2, not done) and R-E (arm 1, arm 2, not done). McNemar s test will be applied to the 2x2 table of clinical-pathological (high/low) by 70-gene signature (high/low) risk to test whether there is a difference in assignment percentages to chemotherapy. Kaplan-Meier curves for OS, DFS and DMFS will be calculated for the following six subgroups: high risk for both risk assessments, low risk for both risk assessments, high risk for clinical-pathological criteria and low risk for 70-gene signature and randomized (R-T) to use clinical pathological assessment of risk Version / January, 2006

75 high risk for clinical-pathological criteria and low risk for 70-gene signature and randomized (R-T) to use 70-gene signature assessment of risk low risk for clinical-pathological criteria and high risk for 70-gene signature and randomized (R-T) to use clinical pathological assessment of risk low risk for clinical-pathological criteria and high risk for 70-gene signature and randomized (R-T) to use 70-gene signature assessment of risk These six curves will be put on the same plot, and their qualitative interpretation will be of great importance for the evaluation of the two assessment methods. However no formal statistical testing will be performed on this plot other than those described for the ITT1 dataset. Analyses on PT dataset The analysis of DMFS at 5 years on the PT dataset is described in the sample size section. Analyses on ITT1 dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable) in the ITT1 dataset, split by R-T. In this dataset special attention will be given to the compliance to chemotherapy administration as assigned by R-T. In the subgroup of clinical-pathological high risk and 70-gene signature low risk, DMFS, OS and DFS will be compared. As these comparisons are low-powered, primary attention should go to the confidence intervals. In the subgroup of clinical-pathological low risk and 70-gene signature high risk, DMFS, OS and DFS will be compared. Again, as these comparisons are low-powered, primary attention should go to the confidence intervals. The interaction between the effect of chemotherapy and the subgroup (clinical-pathological high risk and 70-gene signature low risk versus clinical-pathological low risk and 70-gene signature high risk) will also be tested. Just as for the subset tests, the interaction test will be low-powered, so it will be informative only if there is a large difference in the effect of chemotherapy between the subsets, a situation that is unlikely to occur. Analyses on ITT2 dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable) in the ITT2 dataset, split by R-C. The primary comparison of R-C will be for DFS. Secondary comparisons are for DMFS and OS. For these three time-to-event endpoints, the analyses described above (under general ) will be performed. Analyses on SP2 dataset Chemotherapy administration will be summarized by number of cycles, dose intensity of the respective drugs, dose reductions, and cycle delays. Adverse events and laboratory abnormalities during chemotherapy will be tabulated (worst CTC grade per patient) by treatment arm. No formal toxicity analyses with p values will be carried out. Separate tables and listings will be made for late toxicity ascribed to chemotherapy (cardiac toxicity, secondary leukemia). Version / January, 2006

76 Analyses on ITT3 dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable) in the ITT3 dataset, split by R-E. The timing of this randomization, relative to initial enrollment into the study will be summarized The primary comparison of R-E will be for DFS. Secondary comparisons are for DMFS and OS. For these three time-to-event endpoints, the analyses described above (under general ) will be performed. Analyses on SP3 dataset Duration of endocrine therapy will be described by R-E-arm by Kaplan-Meier curves. A table will specify numbers of patients who started endocrine treatment, numbers who discontinued, reasons for discontinuation and the number of patients who switched from tamoxifen to letrozole (in the appropriate arm). The timing of the switch to letrozole will be summarized. Adverse events and laboratory abnormalities during endocrine therapy will be tabulated (worst CTC grade per patient) by treatment arm. No formal toxicity analyses with p values will be carried out Pre-planned sensitivity or exploratory analyses As discussed in the section on randomization and stratifications (ref ), the clinicalpathological risk evaluation will be modified as new (Standard) versions of Adjuvant! Online become available. A table will be constructed (on the AP dataset) to reflect the impact of these changes, showing how the distribution of risk would have been if the latest version (as at time of analysis) had been applied to all patients. If discrepancies exceed 10% of all patients, sensitivity analyses will be performed to assess the study results under the assumptions that all patients had been evaluated as per the latest version of Adjuvant! Online. To do this, all patients who were categorized differently during the trial will be removed from the respective datasets, and the three primary efficacy tests (respectively on the PT, ITT2 and ITT3 datasets) will be repeated. This sensitivity analysis is not a priory unbiased, because it removes borderline cases, but may give reassurance about the effect of updates of Adjuvant! Online. On the PP1 dataset: if there are more than 10% ineligible patients in the ITT1 dataset, the analyses discussed under PT and ITT1 will be repeated as sensitivity analyses on the PP1 dataset (and its corresponding subset for PT). If a large effect of the type of chemotherapy (R-C) is observed, further subgroup analyses of the analyses on the ITT1 dataset may be performed by type of chemotherapy, to generate confidence intervals of effects per chemotherapy type. Sensitivity analyses will be performed for overall survival analyses on the AP, PT, ITT1, ITT2, ITT3 datasets, by taking only cancer related deaths into account. These analyses will use competing risk methods to describe cumulative incidence of cancer related deaths. In this analysis deaths due to treatment toxicity or other treatment related deaths will be interpreted as cancer related. Similarly, sensitivity analyses will be performed for DMFS analyses on the AP, PT, ITT1, ITT2, ITT3 datasets, using time to distant metastasis, by taking only metastasis as an event, and not deaths. These analyses will also use competing risk methods to describe cumulative incidence of distant metastasis. Version / January, 2006

77 Prognostic factor analyses Prognosis is a key element of the trial. However, evaluation of prognostic factors is confounded by chemotherapy administration (yes, no), because key prognostic factors were used to decide on chemotherapy. Therefore, chemotherapy will be included in all models below, in an attempt to adjust for the effect. If the chemotherapy comparison on ITT2 is significant, the variable will have three levels (no, chemo arm A, chemo arm B). Endocrine therapy will not start at the time of entry into the trial, and will not be taken into account as a covariate. The effect of factors defining endocrine therapy will thus include endocrine therapy use. On the AP dataset, the three time-to-event endpoints (DMFS, OS, DFS) will be subjected to a prognostic factor analysis, using the following conventions: For each variable a first analysis will be performed using a Cox proportional hazards model with the variable, and chemotherapy as covariates. Variables with p<0.10 will be selected for potential inclusion into the final model. If there is significant (p<0.05) interaction with chemotherapy, the interaction will be included. If interaction is included, interaction and primary effect should be moved in or out of the model jointly. All selected variables will be put into a multivariate Cox proportional hazards model, together with chemotherapy, and the final model will be built using backward selection (until all p<0.05), while keeping chemotherapy in the model. The final model will be further evaluated by performing a confirmatory forward selection technique. Variables will be included or excluded with their full categorization, so no regrouping or selection of categories is allowed. The focus of this analysis will be on determining the extent to which 70-gene signature risk assessment may or may not replace other risk factors. After determining a final model, it will be evaluated for calibration and discrimination ability, using bootstrapping. The stability of the selected covariates in the final model will also be evaluated by the bootstrap (Ref. 96). The variables to be included, and their categories, are: Age (<40, 40-49, 50-59, 60) Baseline WHO performance status (WHO 0, WHO >0) Type of surgery (breast conserving surgery, radical mastectomy) Type of node evaluation (SNB, full axillary clearance) Tumor size (0-1 cm, >1-2 cm, >2-3 cm, >3-4 cm, >4-5 cm) Differentiation (grade I well differentiated, grade II moderately differentiated, grade III poorly differentiated, unknown) HER-2 status (positive, negative, unknown) ER-PgR status (Positive (ER and/or PgR), Negative (both)) Clinical-pathological risk assessment (low, high) 70-gene signature risk assessment (low, high) Additional prognostic and predictive analyses are very likely, either to develop additional or improve existing prognostic gene signatures on the basis of microarray data for 6,000 patients, or to develop or validate predictive signatures for endocrine therapy or chemotherapy response. The strategy to perform these will be described in ad-hoc research protocols. Version / January, 2006

78 Prognostic factor analysis For prognostic factor analysis, the central pathology results will be used. Central pathology review will repeat immunohistochemistry for estrogen and progesterone receptors to give standardized levels of expression which will be expressed as a percentage of immunoreactive cells. Categories used will be: positive: ER and/or PgR > 10% low positive (not positive) ER and/or PgR 1-10% negative: both ER and PgR = 0% There are no cut offs in positive i.e. when the expression is above 10% there are no further stratification levels. Further analysis will look into the potential further splitting of the positive category and determining whether expression levels (within positive ) may be an indicator of disease severity Data recoding and display Frequency tables will be tabulated (by treatment group or otherwise) for all categorical variables by the levels of the variables as they appear on the CRF (with %). Categories with a text field specification will be tabulated as categories and then supplemented by a listing with the following information for the patients fulfilling the condition for the specification (patient id, institution, treatment group, value of the item and text field contents). Dates relating to events prior to entry will be presented as the delay in days (or weeks, months, or years) between the past event and the date of entry (date of randomization date of past event + 1) and presented using the median and range. For example, on the randomization checklist, the date of last administration of prior treatment (or the date of first diagnosis of the cancer) will be presented as the time elapsed (in days, weeks, months or years, as appropriate) since the day of the last administration and the date of entry on study (date of randomization last administration/diagnosis +1). Other delays (e.g. re-treatment delays) are presented as continuous variables using the median and range. Safety and toxicity counts will be tabulated as worst grade per patient, within CTC term, or laboratory test. Continuous variables for which a coding system exists (such as for laboratory data) will be recoded into categories (for adverse events, the grading scale specified in the protocol will be used). Whenever no specific scale exists, lab data will be categorized based on the normal range: for example, below the lower normal limit (when appropriate), within the normal range, above the upper normal limit (UNL) and the degree to which it is above the UNL (for example > 2.5 x UNL, > 5 x UNL, > 10 x UNL). For laboratory data, the nadir is generally displayed. The nadir in a given cycle is the lowest laboratory value in that cycle; the overall nadir for a patient is the lowest laboratory value among all cycles. Other continuous variables (for example age, dose ) are presented using the median and range (minimum, maximum). If appropriate, continuous data may also be presented in categories (for example, age may also be grouped in decades). Version / January, 2006

79 10.3 End of study End of study occurs when all of the following criteria have been satisfied: 1. Thirty days after all patients have stopped protocol treatment 2. The trial is mature for the analyses of the three primary endpoints (for R-T, R-C and R-E) as defined in the protocol 3. The database has been fully cleaned and frozen for this analysis 10.4 Stopping rules and interim analyses Pilot study of the trial The trial, a multi-center, prospective, phase III randomized trial that will include approximately 6,000 patients, will start as a randomized study of 800 women, called pilot study, which should ensure the following: The logistics of the trial, as described in chapter 20, prove to be feasible from the point of view of patients, physicians and laboratories involved; The patient population recruited upfront for the trial is not biased : clinicians are entering in the trial a reasonable spectrum of node-negative disease and not just very low risk patients. This will be observed from the actual distribution of high risk vs. low risk patients as compared to expected numbers; Clinicians comply with the protocol in the 70-gene signature /genomic arm of the study: in other words, a good 70-gene signature prognosis signature combined with bad clinicalpathological characteristics leads to a no chemotherapy decision if the patient is randomized to the genomic arm (R-T) and a poor gene prognosis signature combined with good clinical-pathological characteristics leads to a chemotherapy decision if the patient is randomized to the genomic arm (R-T); Among all patients who are candidates for adjuvant chemotherapy (see section 4.3), there are at least 66% who are undergoing the second randomization (R-C); A statistically significant difference is observed between the percentages of patients that have a high clinical-pathological risk (pc) and a high genomic risk (pg) (thus reflecting a likewise reduction in chemotherapy administration). With 800 patients in the pilot study and the following assumptions: the null hypothesis is H0: pc - pg = 0% the type I error rate is set to 5% then the pilot study has 80% power to reject the null hypothesis of no difference if the true difference is at least 7%. Monitoring of pc and pg (as well as of other trial data, especially crossovers between the chemotherapy and the no chemotherapy groups) should be the responsibility of an Independent Data Monitoring Committee Stopping rule after 800 patients enrolled If the data contradict the assumption of less patients receiving chemotherapy in the genomic arm in the first cohort of node-negative, breast cancer patients (N=800) then the prospective, randomized study will be put on hold until the reason for the deviation has been resolved; the same remark applies in case major issues regarding the logistics of the trial are identified. Version / January, 2006

80 Stopping rule to be applied every year In this study, the group of patients who have a low risk genomic prognosis signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy, are potentially at risk of being under treated. In analogy to the primary test of the study, a rule to be used in this particular subset of patients, to trigger review by the EORTC IDMC of the available efficacy data will be as follows: if the lower one-sided 95% confidence limit of the (exponential) hazard of distant metastasis or death exceeds /year (corresponding to a 5% event rate for DMFS at 5 years). This is equivalent to an unadjusted test at 95% confidence level rejecting a hypothesis of a 95% 5-year DMFS. This test will be performed every year. The test is not the same as the primary test, because it uses a different null hypothesis, and because it will rely on early data. Thus it assumes a constant hazard. As the test does not correct for multiplicity, it should be used as a trigger for review. The EORTC IDMC will review the relapse data in case the rule described above triggers a review Interim analyses for the chemotherapy and the endocrine questions An interim analysis of the primary comparison for R-C will be performed as described in section This interim analysis will be reviewed by the EORTC-IDMC. It will be restricted to the ITT2 dataset, and only consider the comparison of the two randomized regimens of chemotherapy. A significant outcome may lead to early publication of results, restricted to this particular comparison. An interim analysis of the primary comparison for R-E will be performed as described in section This interim analysis will also be reviewed by the EORTC-IDMC. It will be restricted to the ITT3 dataset, and only consider the comparison of the two randomized sequences of endocrine therapy. A significant outcome may lead to early publication of results, restricted to this particular comparison. The following table gives an overview of the analyses planned for each of the three primary endpoints. Table 12: Overview of interim and final analyses of primary endpoints Primary endpoint for Endpoint Analysis population Alpha spending function Analysis Timing Nominal 2-sided alpha level R-T DMFS PT NA Final See R-C DFS ITT2 Rho = 2.5 R-E DFS ITT3 Rho = 2.5 Interim 211 events * Final 422 events * Interim 191 events * Final 382 events * * These are the nominal levels if the interim analysis is at exactly half the planned number of events. If the number of events at interim is slightly more, the correct values according to the spending function will be used. These are very close to the tabulated values. Version / January, 2006

81 11 Data monitoring A Data and Safety Monitoring Board (DSMB) will monitor the recruitment, the reported adverse events and the data quality at least twice a year. Arising problems will be discussed with the Study Coordinator who will take appropriate measures. Relevant information (including relevant safety data) will be included in the study status reports serving as a basis of discussion during EORTC Group meetings. These reports will be made available to investigators participating in the study and to the EORTC Independent Data Monitoring Committee (IDMC) when interim analyses (planned or unplanned, see below) are carried out. When interim analyses are conducted for this study (see chapter on statistics), monitoring of efficacy and safety data will be performed according to the statistical design of the study (see chapter on statistics) and the EORTC policy on Independent Data Monitoring Committee and Interim Analyses. Results of interim analyses are confidential and will be discussed by the EORTC IDMC that will subsequently advise the Group on eventual changes to be brought to the study. No efficacy results will be presented at Groups meetings before the trial is closed to recruitment and data are mature for the analysis of the primary endpoint, unless recommended otherwise by the IDMC. 12 Translational research 12.1 Sample collection The following material should be obtained from each patient: 1(2) punch biopsy sample(s) from the breast cancer tissue for microarray analysis, biological materials bank storage and proteomics analysis. A representative paraffin tissue block from the breast cancer tissue for central histopathology review and for the production of tissue microarrays (TMAs). 1 optional blood sample (for serum storage) for proteomics analysis and biological materials bank storage to be taken between enrollment and the start of treatment. For details of sampling see chapter 20, sample schedule and trial logistics. Objectives: Samples will be collected to perform genomic prognosis (by microarray analysis) and central pathology review. will establish a central TRANSBIG biological materials bank for research described in the protocol and future research. The research will be evaluated and agreed upon by the appropriate research/transbig committees Central pathology review Central pathology review will assess tumor grade and identify cases with discordant diagnosis between local and central pathology review. It will also generate feed-back information for local pathologists following confirmation of the local diagnoses. Since several studies have observed a high level of discordance between the analysis of histopathology (e.g. levels of hormone receptor and HER-2 expression) in peripheral pathology Version / January, 2006

82 laboratories and central more experienced laboratories, a central review of the main standard histopathology will be performed. This will also enable the assessment of the prognostic value of level of expression of ER and PgR rather than the current negative, low positive, positive scaling. For these purposes a representative diagnostic paraffin tissue block of each tumor will be sent from each participating center to IEO (Instituto Europeo di Oncologia, Milan, Italy), attention of Prof Guiseppe Viale. The sending of these paraffin blocks to Milan will be at least once every 6 months. Tissue microarray production To evaluate hundreds of tumors in a high-throughput manner, tissue microarrays (TMA) will be made. The TMAs remaining after central histopathology analysis will be stored in the TRANSBIG biological materials bank and made available for research. The individual paraffin blocks will be returned to the participating centers after the tissue microarray of each sample has been made. Objectives: Central review of the main standard pathology in order to determine cases where local and central pathology are discordant and to generate feedback information for the local pathologist. To prepare a database of histopathology of all of the samples. To perform immunostaining for ER, PgR, Ki-67 and HER-2. HER-2 status will be confirmed by FISH. To determine the clinical relevance of molecular markers. To validate the clinical significance of clusters of genes stemming from gene expression profile. To establish a permanent tissue microarrays (TMA) tumor bank without the need for retaining the original donor paraffin blocks. Methods Standard histopathology and immunhistochemistry techniques will be used in combination with TMA technology. For TMA construction, two sections from each paraffin tissue block will be stained with HE to determine representative tumor regions. Tissue cores will be punched from the primary paraffin tumor blocks (the donor blocks) and subsequently brought into one empty recipient block. Numerous sections can be cut from the array blocks. Sections from these array blocks will be transferred to glass slides Proteomics Proteomics can provide insight in the post-transcriptional gene function and elucidate post translational pathway not obtainable by microarrays. This research area can make the connection between genes and biological function. Therefore, one of the side studies of the trial comprise of proteomics research. The initial proteomics studies will be performed at the proteomics facility in Swansea, UK. Therefore, at the end of accrual serum samples will be shipped from each participating centre (on dry ice) to the TRANSBIG biological materials bank in Brussels; from there one of the 0.5 ml serum aliquots will be dispatched to Southwest Wales Cancer Institute, Swansea UK for analysis. Objectives To standardize methods for the extraction of proteins and preparation for analysis from sera and tissue from breast cancer patients. To identify new biomarkers (tumor specific proteins and peptides) in tissue and sera allowing early detection of breast cancer. Version / January, 2006

83 To discover clinically relevant protein profiles from sera and tissue from breast cancer patients. To identify novel prognostic markers for breast cancer. To identify novel predictive markers for breast cancer. To correlate the expression of these markers with tissue histopathology and with clinical outcome. To link gene expression profiles to biological function. To identify tumor specific proteins or peptides which might be used as new pharmacological targets. Methods The primary analytical tools used for the proteomics will be SELDI, MALDI-TOF mass spectrometry, immunochemistry and high throughput 2D gel electrophoreses Complex microarrays In addition to the 70-gene signature, the complex microarrays used will yield information on gene expression of genes (i.e., oligonucleotides that represent thousands of genes on a microarray). For these complex microarrays, RNA isolated from the fresh frozen punch biopsy samples will be used. These complex microarrays will be performed at Agendia, Amsterdam, The Netherlands. Objectives will measure RNA expression levels using microarray technology and determine whether expression signatures can give prognostic information (i.e., 70-gene signature). To determine the overall gene expression pattern on the genomic array in order to identify new prognostic gene expression profiles. To determine the overall gene expression pattern on the genomic array in order to identify new predictive gene expression profiles. To validate previously discovered gene profiles. Methods Briefly, one section will be stained with HE to determine the percentage tumor cells in the frozen biopsy sample. When the tumor cell percentage is sufficient (> 50%), RNA will be isolated from the sample. Then the isolated RNA is amplified and labeled. The labeled sample RNA and labeled control oligonucleotides are combined and placed in a hybridization chamber in contact with the complex array. After hybridization slides were washed and scanned using a confocal laser scanner. Fluorescence intensities on scanned images were corrected for background noise and normalized. For detailed methods see (Ref. 28) Biological materials bank storage Objectives To establish an early breast cancer biological materials bank with fresh frozen tumor tissue samples, RNA, serum and tissue microarrays. To establish a resource for future research. Version / January, 2006

84 RNA will be isolated from the frozen tumor biopsy samples to perform the complex microarray. Remaining RNA will be transferred to and stored in the TRANSBIG biological materials bank. When the microarray analysis is performed successfully, the remaining frozen biopsy tissue will be transferred to the TRANSBIG biological materials bank for use in future research and proteomics analysis. These RNA and tissue samples will be shipped on dry ice from Agendia to the TRANSBIG biological materials bank every 6 months. The serum samples will be shipped to the TRANSBIG biological materials bank at the end of accrual and then an aliquot transferred to Swansea for proteomics analysis. The biological material will be stored under the guardianship of the TRANSBIG consortium. The biological materials bank will be in Brussels Belgium in a facility run by World Courier. The rules and priorities for access to the biological material are decided on scientific merit of the proposed projects and set out in a separate document. 13 Publication policy It is the EORTC s policy not to release trial results before data maturity has been reached for the primary endpoint(s) of the trial unless the publication is authorized by an Independent Data Monitoring Committee. If the group wishes to publish or present study data before the publication of the primary trial endpoint, this may be authorized under the conditions specified in the EORTC Policy 009 Release of Results and Authorship Policy available from or authorized by an Independent Data Monitoring Committee and with the approval of the TRANSBIG Steering Committee. The final publication of the trial results will be written by the Study Coordinators from the EORTC Breast Cancer Group on the basis of the final analysis performed at the EORTC Data Center and then approved by the EORTC Data Center. The manuscript will then be approved by all co-authors prior to being circulated to the steering committees of the EORTC Breast Group and TRANSBIG, in parallel, for comments and approval. Authorship on publications and presentations shall be based on academic standards and customs (i.e. those investigators who have included a substantial number of patients or who have contributed considerably to the development, conduct and analysis of the trial). All public presentations or publications shall also give appropriate acknowledgement to the EORTC and TRANSBIG consortium, including various sources of funding received, such as support from the European Commission Framework Programme VI (FP6-LSHC-CT ) and industrial partners. After revision by the EORTC Data Center and other co-authors (and the industrial partners) the manuscript will be sent to a major scientific journal. All abstracts or manuscripts as well as detailed descriptions of any planned oral presentations must be submitted to the EORTC and to the TRANSBIG Steering Committee for approval and transbig@bordet.be) at the earliest practicable time, but at least within four weeks prior to any proposed submission for publication of any manuscript or any presentation or other public disclosure. Abstracts must be sent to the EORTC and TRANSBIG Steering Committee at the earliest practicable time, but at least within ten days before submission. The EORTC Group Chairman, Study Coordinators, EORTC Data Center Team, the TRANSBIG Steering Committee or any of the industrial partners may comment upon, but may not change, the conclusions and content of any such publication or presentation. Authors are entitled to withhold any proprietary confidential information from publications / presentations. The EORTC, the TRANSBIG Steering Committee or any of the industrial partners may also, under exceptional circumstances, object to or request a delay before publication / presentation. In such cases, the EORTC, the TRANSBIG Steering Committee or any of the industrial partners must provide in writing its objections and recommendations within ten days for abstracts and within twenty days for all other publications / presentations. To allow for protection, after any objection, planned Version / January, 2006

85 publications / presentations may be suspended until the end of a consultation period and up to a maximum of sixty days. This is applicable to any individual patient enrolled/randomized in the trial, or any subgroup of these. Such publications must comply with the terms specified in the EORTC Policy 009 Release of Results and Publication Policy, the Consortium Agreement and with Contract Agreements with the companies involved. Therefore, such a publication cannot include any comparisons between any of the randomized treatment arms or an analysis of any of the study endpoints unless the final results of the trial have already been published. 14 Investigator authorization procedure Investigators will be authorized to register for screening, enroll, or randomize patients in this trial only when they have returned to their National Coordinating Center/Group (NCC/NCG) or the EORTC: The updated signed and dated Curriculum Vitae of the Principle Investigator. The (updated) list of the normal ranges, in their own institution, of all laboratory data required by the protocol, preferably signed and dated by the head of the laboratory. A commitment statement / study acknowledgment form, indicating that they will fully comply with the protocol, to include an estimate of their yearly accrual and if any conflict of interest may arise due to their participation in the trial. A signed conflict of interest disclosure form: this document will be required only if a possible conflict is declared on the commitment form. A copy of the favorable opinion of their local or national (whichever is applicable) ethics committee mentioning the documents that have been reviewed (incl. version number and date of documents) and indicating the list of the ethics committee members. A copy of the translated and adapted (according to all national requirements), Patient Information / Informed Consent sheets, clearly mentioning the version number and the date. The signature log-list of the staff members with a sample of each authorized signature and the indication of the level of delegations. (This will be used for allocating usernames and passwords for remote data capture) The coordinates of the pharmacist who will be responsible for the trial medication (this will be filled in on the signature log). The accreditation letter for the laboratory. (if available for your center and/or applicable by your national law) A signed document to confirm receipt of the Investigator s brochure. A signed initial Material Transfer Agreement (MTA) to be sent to TRANSBIG. (Confirm MTA online at the end of accrual listing all sample SeqIDs) The center specific applicable list of required documents will be included in the site initiation package, with proper instructions as required by this protocol, your group and/or the applicable national law The new investigator will be added to the authorization list, and will be allowed to register/enroll/randomize patients in the trial as soon as All the above mentioned documents are available at their NCC/G or EORTC Data Center Version / January, 2006

86 All applicable national legal and regulatory requirements are fulfilled Patient registration for screening/enrollment/randomization from centers not (yet) included on the authorization list will not be accepted. 15 Patient enrollment procedure 15.1 Enrollment procedure Patient enrollment/randomization will only be accepted from authorized investigators (see "Authorization procedure") Access to the interactive randomization program All trial authorized investigators can enroll/randomize patients directly on the website, 24 hours a day, 7 days a week ( To access the website containing the interactive randomization program, investigators need a username and a password. All authorized investigators will be provided with a username and password prior to site initiation. EORTC participants can use their ORTA username and password to access the website and if they have forgotten their ORTA password, they can interactively request a password reminder from the EORTC ORTA website ( Overview of patient screening and enrollment Steps to be taken for patient registration for screening (also see diagram chapter 20) identify patients who may be eligible give information and ask patient to sign screening PIS & IC register patient for screening on web page and receive patient identification number (SeqID) breast cancer surgery, freeze tumor biopsy & register tumor sample as available for collection on website give tumor biopsy to courier & confirm sample pick-up as soon as lymph node status is confirmed by local pathology this information must be recorded on the website (then the genomic test will be performed for eligible patients) once the genomic prognosis has been successfully performed, the PI will receive an notification that patient is eligible for entry/enrollment in the trial inform eligible patient about (general information) Steps to be taken for patient enrollment: timing and logistics for first post-operative visit A flow diagram similar to the one in chapter 20 will be provided to and adapted by each center to reflect local procedures and detail the logistics of the screening/enrollment process a paper enrollment checklist (detailing the eligibility criteria) will be provided and should be filled in before connecting to the website to enroll a patient inform the patient about, signing of PIS & IC 1 Version / January, 2006

87 connect to website, input SeqID & confirm date of birth then input clinical data and date of signing of PIS & IC 1 to enroll the patient. Comparison of clinical and genomic risk assessments and receive the treatment allocation (results of clinical and genomic prognostic tests) Inform patient of proposed treatment Discuss further randomizations as applicable, give patient additional PIS & ICs Sign and date the enrollment checklist and send to NCC/G Patient registration for screening Patients will be registered for screening before or at the time of surgery and tumor sample availability. When a patient is registered for screening some clinical information will be requested and the web based platform will generate a unique sequential identification number (SeqID) for each patient which will also serve to identify the patient during the trial if the patient is eligible and enrolled. This SeqID must be quoted at all subsequent sessions on the website. The SeqID number attributed to the patient at the end of the registration for screening procedure must be recorded on the enrollment checklist, and must be reported on all case report forms Patient enrollment and randomization for treatment decision making tool (R-T) A patient can be enrolled and randomized only after verification of eligibility. This must be done before the start of the protocol treatment. An exhaustive list of questions to be answered during the enrollment/randomization procedure is included in the enrollment checklist, which is part of the case report forms. This checklist should be completed by the responsible investigator before the patient is enrolled. In order to enroll a patient in the trial, the interactive randomization program needs to be accessed via the website and the following questions answered: provide SeqID confirm Date Of Birth (DOB) (day/month/year) patient's code (maximum 4 letters or digits chosen at the discretion of the investigator) hospital patient's chart number (if available) Note this item is optional. date of written informed consent Eligibility will be confirmed by collection of all the data requested on the enrollment checklist. Risk group assignment The computer will calculate a 10-year cancer specific survival probability, using the adjuvant! online software. The risk according to clinical-pathological criteria will be calculated by the Adjuvant! Online software incorporated into the web based platform. The patient s age, ER status, tumor grade and tumor size will be passed on to Adjuvant! Online by the web based platform. Comorbidity will be defaulted to Minor problems and positive nodes to 0. The resulting 10 year cancer specific death risk will then be used to define the risk. If Adjuvant! Online is upgraded during the trial the new version will be used. If a new variable is added (for example HER-2 status) Version / January, 2006

88 the new variable will be incorporated into the clinical prognosis. The standard version will always be used, not the genomic version. The computer will display all known information entered for this patient up to the time of enrollment. The computer will allow changes to any incorrect information (except the patient identification). After all information is confirmed as being correct the patient will be enrolled and risk assessments compared. For concordant patients: Clinical high risk-genomic high risk patients will receive chemotherapy Clinical low risk-genomic low risk patients will not receive chemotherapy For discordant patients the computer will randomized between the 2 decision making tools and, therefore allocate each patient to either: 1. decision according to clinical prognosis: with no chemotherapy if patient has good clinical prognosis, and chemotherapy if patient has poor clinical prognosis 2. decision according to 70-gene signature prognosis: with no chemotherapy if patient has good 70-gene signature prognosis, and chemotherapy if patient has poor 70-gene signature prognosis This completes enrollment. The risk assessment results and allocated prognostic tool, where applicable, must be recorded on the enrollment checklist which will be signed and dated by the Principal Investigator and sent to the NCC/G. The Principal Investigator, NCC/G where applicable and the EORTC Data Center will receive an confirming patient enrollment with details of both the genomic and the clinical risk assessment results and the result of randomization for all discordant patients Randomization of chemotherapy (R-C) Once a patient has been assigned to receive chemotherapy, she can be offered the randomization to anthracycline based chemotherapy or docetaxel-capecitabine chemotherapy. This randomization can be done immediately after the assignment to chemotherapy, or later. If the chemotherapy randomization (R-C) is done later, the interactive randomization program needs to be accessed and the sequential identification number attributed to the patient provided. The computer will display all known information entered for this patient, including whether this patient has been allocated to receive chemotherapy. Fill in the chemotherapy randomization checklist and then log-on to the website to randomize the patient If the patient agrees to the chemotherapy randomization, fill in all the information from the chemotherapy randomization checklist and the computer will allocate her to 1. anthracycline based chemotherapy 2. docetaxel-capecitabine chemotherapy This completes the chemotherapy randomization. The allocated treatment must be recorded on the randomization checklist which will be signed and dated by the Principal Investigator and sent to the NCC/G. The Principal Investigator, NCC/G if applicable and the EORTC Data Center will receive an confirming the chemotherapy treatment allocation. If a patient who should be proposed chemotherapy does not wish to participate in the chemotherapy randomization (R-C), she should receive the hospital s standard care chemotherapy. The patient Version / January, 2006

89 remains on protocol and a summary of the chemotherapy treatment given should be recorded on the non-randomized chemotherapy CRF Randomization of endocrine therapy (R-E) All hormone receptor positive patients (i.e. ER &/or PgR positive) can be offered the endocrine therapy randomization to 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. In order to randomize a patient, complete the endocrine therapy randomization checklist, then connect to the website. Provide the SeqID attributed to the patient and confirm her date of birth then the computer will display all known information entered for this patient, including whether this patient is eligible to receive endocrine therapy. If the patient agrees to the endocrine therapy randomization, fill in all the information from the endocrine therapy randomization checklist and the computer will allocate her to 1. 2 years of tamoxifen followed by 5 years of letrozole 2. 7 years of letrozole This completes the endocrine therapy randomization. The allocated treatment must be recorded on the randomization checklist which will be signed and dated by the Principal Investigator and sent to the NCC/G. The Principal Investigator, NCC/G if applicable and the EORTC Data Center will receive an confirming the endocrine therapy treatment allocation. If a patient with an endocrine responsive tumor does not wish to participate in the endocrine therapy randomization (R-E), she should receive the hospital s standard care endocrine therapy. The patient remains on protocol and the endocrine therapy treatment given should be recorded on the adjuvant endocrine therapy CRF. 16 Forms and procedures for collecting data 16.1 Electronic case report forms and schedule for completion All further data will be reported on electronic forms which will be completed and submitted using Remote Data capture (RDC). Guidelines on how to use RDC will be provided to all centers by the EORTC. All RDC forms will be specifically designed by the EORTC Data Center for this study. These electronic forms will be used by all participants. Queries will be generated by the EORTC and sent (as.pdf) either to the National Coordinating Centers/Groups (NCC/G) to distribute or to the centers directly to centers in countries where the EORTC is the NCC/G. The query.pdf forms will be printed and filled in on paper by the PI (and anyone authorized to sign forms by the PI and included on the signature log) and then signed and returned to their NCC/G or directly to the EORTC data center in countries where the EORTC is the NCC/G. Forms to be sent to the EORTC Data Center directly should be sent to: Breast cancer group Data Manager EORTC Data Center Avenue Emmanuel Mounier, 83, bte 11 B-1200 Brussels, Belgium Version / January, 2006

90 A. Before the adjuvant treatment starts: the patient must be enrolled/randomized through the web based platform a signed and dated hardcopy/paper copy of the enrollment check list, must be sent either to your NCC/G or directly to the EORTC Data Center (if the EORTC is your NCC/G). The optimal way to work is to complete the enrollment check-list first and to enroll/randomize the patient as soon as it is completed. The date of registration and patient sequential identification number are then completed on the check-list, and this form can be sent to the Data Center. Further randomization checklist(s) (chemotherapy R-C & endocrine therapy R-E as applicable), must also be sent either to your NCC/G or directly to the EORTC Data Center (if the EORTC is your NCC/G). B. Upon occurrence of a Pregnancy: Any pregnancy in a female subject diagnosed during the treatment period or within 30 days after last study treatment administration must be reported to the EORTC Pharmacovigilance Unit. This must be reported on a Pregnancy Notification Form/Fax within 24 hours of first becoming aware of the event and sent by fax to the allocated NCC who will in turn forward it to the EORTC Pharmacovigilance Unit. If a Serious Adverse Event (SAE) occurs in conjunction with the pregnancy, please also complete an SAE form as explained in the SAE chapter Upon notification of a pregnancy, the EORTC will coordinate the follow up, the development and outcome of the pregnancy. C. Upon occurrence of a Serious Adverse Event (SAE): SAEs occurring from the time a subject is enrolled until 30 days after last protocol treatment must be promptly reported. Any SAE occurring after the 30-days period and considered to be reasonably related to the investigational product or study participation, also have to be promptly notified All these events must be reported by fax to the allocated NCC/G who will in turn forward it to the EORTC Pharmacovigilance Unit on a Serious Adverse Event Form within 24 hours of the initial observation. A completed SAE-form must be sent back within 10 calendar days of the initial observation of the Serious Adverse Event. ALL Forms must be dated and signed by the responsible investigator or one of his/her authorized staff members 16.2 Data Flow (Remote Data Capture) All forms must be electronically completed according to the schedule defined in the CRF guidelines through the EORTC web based Remote Data Capture (RDC) system. The list of staff members authorized to enter forms must be identified on the signature log and sent to the Data Center by the responsible investigator before the start of the study. All staff need ORTA usernames and passwords to use the EORTC web based RDC system. Version / January, 2006

91 In all cases, it remains the responsibility of the investigator to check that data are entered on the RDC electronic forms as soon as possible and that they are completely and correctly filled out and submitted to the EORTC database. On the data received, the EORTC Data Center will perform extensive consistency checks and issue queries in case of inconsistent data. Queries will be sent by (PDF), and must be filled out on the printed paper copy. A copy should be kept on site, the original must be sent by regular mail to your NCC/G or to the EORTC Data Center. The EORTC data manager will subsequently apply the corrections into the database. If an investigator (or an authorized staff member) needs to modify a CRF after the form has been sent to the EORTC Data Center, he/she should use a Data Correction Sheet and sent it to their NCC/G or the EORTC Data Center. 17 Reporting of Serious Adverse Events 17.1 Definitions AE: An Adverse Event is defined as any untoward medical occurrence in a patient or clinical trial subject administered a medicinal product and which may not necessarily have a causal relationship with this treatment. An adverse event can therefore be any unfavorable and unintended signs (such as rash or enlarged liver), symptoms (such as nausea or chest pain), an abnormal laboratory finding (including blood tests, x-rays or scans) or a disease temporarily associated with the use of the protocol treatment, whether or not considered related to the investigational medicinal product. AR: An Adverse reaction of an investigational medicinal product is any untoward and unintended responses to an investigational medicinal product related to any dose administered. All adverse events judged by either the reporting investigator or the sponsor as having a reasonable causal relationship to a medicinal product qualify as adverse reactions. The expression reasonable causal relationship means to convey in general that there is evidence or argument to suggest a causal relationship. UAR: An Unexpected Adverse Reaction is any adverse reaction, the nature, or severity of which is not consistent with the applicable product information (e.g. investigator's brochure for an unapproved investigational product or summary of product characteristics (SmPC) for an authorized product). When the outcome of the adverse reaction is not consistent with the applicable product information this adverse reaction should be considered as unexpected. Severity: The term severe is often used to describe the intensity (severity) of a specific event. This is not the same as serious, which is based on patient/event outcome or action criteria. SAE: A Serious Adverse Event is defined as any undesirable experience occurring to a patient, whether or not considered related to the protocol treatment. SAR: A Serious Adverse Event (SAE) which is considered related to the protocol treatment is defined as a Serious Adverse Reaction An Adverse Event or Adverse Reaction which is considered as serious: results in death, is life-threatening (i.e. an event in which the subject was at risk of death at the time of event; it does not refer to an event which hypothetically might have caused death if it were more severe) Version / January, 2006

92 requires hospitalization or prolongation of existing inpatients hospitalization, results in persistent or significant disability or incapacity, is a congenital anomaly or birth defect. results in any other medically important condition (i.e. important adverse reactions that are not immediately life threatening or do not result in death or hospitalization but may jeopardize the patient or may require intervention to prevent one of the other outcomes listed above). SUSAR: Suspected Unexpected Serious Adverse Reactions 17.2 Reporting procedure Non- serious adverse events and/or non-serious adverse drug reactions Adverse Events (AE) and /or Adverse Reactions (AR) must be recorded as indicated in the protocol Serious adverse events or serious adverse drug reactions All Serious Adverse Events (SAE) occurring from the time a subject is enrolled until 30 days after last protocol treatment, must be reported to the EORTC Pharmacovigilance Unit within 24 hours. (Ref. SAE s will either be sent to the NCC/G (who will forward to the EORTC) or directly to the EORTC depending on the organization in the country. All SAEs that are simply signs and symptoms of the disease being studied do NOT need to be collected! Examples of SAEs that do not need to be reported: Medical or surgical procedure (e.g., endoscopy, appendectomy); the condition that leads to the procedure is an AE Situations where an untoward medical occurrence did not occur (social and/or convenience admission to a hospital) Anticipated day-to-day fluctuations of pre-existing disease(s) or condition(s) present or detected at the start of the study that do not worsen The disease/disorder being studied, or expected progression, signs, or symptoms of the disease/disorder being studied, unless more severe than expected for the subject's condition A hospitalization which was planned before the patient consented for study participation and where admission did not take longer than anticipated. Any SAE that occurs outside of the SAE detection period (after the 30-days period), considered to be reasonably related to the investigational product or study participation, have to be promptly notified to the EORTC Pharmacovigilance Unit, either via the NCC/G (who will forward to the EORTC) or directly to the EORTC depending on the organization in the country This must be done by fax within 24 hours of the initial observation of the event. The principal investigator will decide if these events are related to the protocol treatment (i.e. unrelated, likely related, and not assessable) and the decision will be recorded on the Serious Adverse Event form, if necessary with the reasoning of the principal investigator. The investigator is obligated to assess the relationship between investigational product and the occurrence of each SAE. A "reasonable possibility" is meant to convey that there are facts/evidence or arguments to suggest a causal relationship, rather than a relationship cannot be ruled out. The Version / January, 2006

93 investigator will use clinical judgement to determine the relationship. Alternative causes, such as natural history of the underlying diseases, concomitant therapy, other risk factors, and the temporal relationship of the event to the investigational product will be considered and investigated. The product reference documents are For marketed products: Summary Of Product Characteristics which can be found on For non-marketed products: Current version of the Investigators Brochure For the causality assessment, the following definitions must be used: Relationship to the Description protocol treatment UNRELATED LIKELY RELATED NOT ASSESSABLE There is no evidence of any causal relationship to the protocol treatment There is (some) evidence to suggest a causal relationship to the protocol treatment and influence of other factors is unlikely or absent. There is insufficient or incomplete evidence to make a clinical judgment of the causal relationship to the protocol treatment. Details should be documented on the specified Serious Adverse Event Form. All Investigators participating through their National structure /NCC/G should follow recommendations below: PLEASE FAX THE REPORT TO: Your NCC/G (details can be found in the Administrative responsibilities chapter 21) (who will forward to the EORTC) or directly to the EORTC DC if they are your NCC. NCC/Gs will forward to: Pharmacovigilance Unit fax no The EORTC Pharmacovigilance Unit will forward all Serious Adverse Event reports within 24 hours of receipt to all appropriate persons (See Administrative chapter). To enable the EORTC Pharmacovigilance Unit/sponsor to comply with regulatory reporting requirements, completed documentation of any reported serious adverse events or serious adverse reactions must be returned within 10 calendar days of the initial report. If the completed form is not received within this deadline, the Pharmacovigilance Unit will make a written request to the investigator. It should be recognized that Serious Adverse Reactions (SAR) which have not been previously documented in the Investigators Brochure, or which occur in a more severe form than anticipated (i.e. they are unexpected by nature or severity), are subject to rapid reporting to the Regulatory Authorities. Version / January, 2006

94 ANY QUESTION CONCERNING SAE OR SAR REPORTING CAN BE DIRECTED TO: EORTC Pharmacovigilance Unit phone: fax: pharmacovigilance@eortc.be ALL FORMS MUST BE DATED AND SIGNED BY THE RESPONSIBLE INVESTIGATOR OR ONE OF HIS/HER AUTHORIZED STAFF MEMBERS. 18 Quality assurance 18.1 Control of data consistency Data forms will be entered in the database of the EORTC Data Center using the RDC system. Computerized and manual consistency checks will be performed on newly entered forms; queries will be issued in case of inconsistencies. Consistent forms will be validated by the Data Manager to be entered on the master database. Inconsistent forms will be kept "pending" until resolution of the inconsistencies Audits To ensure quality of data, study integrity, and compliance with the protocol and the various applicable regulations and guidelines, the EORTC Quality Assurance Unit regularly conducts site visits to institutions participating to EORTC protocols. The investigator, by accepting to participate to this protocol, agrees to co-operate fully with any quality assurance visit undertaken by third parties, including representatives from the EORTC, national and/or foreign regulatory authorities or company supplying the product under investigation, as well as to allow direct access to documentation pertaining to the clinical trial (including CRFs, source documents, hospital patient charts and other study files) to these authorized individuals. The investigator must inform the EORTC immediately in case a regulatory authority inspection would be scheduled. This procedure does not apply to non-eortc investigators who should contact their NCC/G for the information on eventual audits performed by their group. 19 Ethical considerations 19.1 Patient protection The responsible investigator will ensure that this study is conducted in agreement with either the Declaration of Helsinki (Tokyo, Venice, Hong Kong, Somerset West and Edinburgh amendments) or the laws and regulations of the country, whichever provides the greatest protection of the patient. The protocol has been written, and the study will be conducted according to the ICH Harmonized Tripartite Guideline for Good Clinical Practice (ref: Version / January, 2006

95 The protocol will be approved by the Local, Regional or National Ethics Committees Subject identification The name of the patient will not be asked for nor recorded at the Data Center. A sequential identification number will be automatically attributed to each patient enrolled in the trial. This number will identify the patient and must be included on all case report forms. In order to avoid identification errors, patient s code (maximum of 4 letters), date of birth and local chart number (if available) will also be reported on the case report forms Informed consent All patients will be informed of the aims of the study, the possible adverse events, the procedures and possible hazards to which she will be exposed, and the mechanism of treatment allocation. They will be informed as to the strict confidentiality of their patient data, but that their medical records may be reviewed for trial purposes by authorized individuals other than their treating physician. An example of the patient informed consent statements are given as an appendix to this protocol. Investigators will receive the translated PIS & ICs from their NCC/G. For countries where there is not an NCC/G the PIS & ICs will be translated by the national coordinator and will be sent to all centers by the EORTC. All PIS & ICs must be version controlled and dated and this information must always be stated in any communication with ethics committees. The translated informed consent form is part of the documents to be submitted to the ethics committee for approval. The competent ethics committee for each institution must validate local informed consent documents before the center can join the study. It is the responsibility of the Local Ethical Committee to guarantee that the translation is conforming to the ICH-GCP guidelines. It will be emphasized that the participation is voluntary and that the patient is allowed to refuse further participation in the protocol whenever she wants. This will not prejudice the patient s subsequent care. Documented informed consent must be obtained for all patients included in the study before they are enrolled or randomized at the EORTC Data Center. This must be done in accordance with the national and local regulatory requirements. For European Union member states, the informed consent procedure must conform to the ICH guidelines on Good Clinical Practice. This implies that the written informed consent form should be signed and personally dated by the patient or by the patient s legally acceptable representative. 20 Sample schedule and trial logistics Detailed instructions describing the precautions involved in obtaining the tumor biopsy and tissue handling are described in a separate brochure and in SOPs & 15. SOPs 00-04, SOP 15 and the tissue handling brochure will be provided along with the tissue sample kit which will contain blood, serum and tumor tubes, a biopsy punch and coded stickers Frozen tumor sample schedule For each patient a tumor biopsy is a prerequisite for inclusion in the trial. RNA will be extracted from part of the tumor biopsy and used for microarray analysis to determine the genomic prognosis. As RNA is very unstable special precautions need to be taken when taking and freezing the biopsy. These are summarized below and detailed in SOPs Version / January, 2006

96 Immediately after surgery, the excised breast tumor tissue must be transported to the place where the biopsy will be taken and frozen, usually the pathology department. The tissue must be transported in a dry tumor container without fixatives as detailed in SOP 00. To decide the best region of the tumor from which the punch biopsy should be taken, obtain a good overview of the different sections of the tumor. The centre of the tumor is often necrotic, while the edge of the tumor often contains many stromal cells, which in both cases will lead to percentage of tumor cells less than 50% and therefore not enough to isolate tumor RNA from. Take a representative biopsy, from the periphery of the tumor thereby optimizing the chance of a high proportion of tumor cells and a good yield of tumor RNA. Tumor biopsy cellularity will be determined by Agendia/NKI along with other quality checks. One, or if feasible 2, punch biopsy samples must be obtained using the biopsy punch provided. A 6 mm biopsy is preferred however for small tumors (e.g. of <1 cm) biopsies of 3 mm are acceptable. The punch biopsy samples must be taken and frozen in liquid nitrogen within 1 hour of surgery, as detailed in SOP 01. Immediately after taken the punch biopsy sample place it in the labeled tube provided, firmly close the tube and submerge the tube in liquid nitrogen. After 2 minutes the biopsy tube can be transferred to a -80 C freezer (being transported on dry ice if necessary to prevent thawing). The availability of the tumor biopsy should then be registered on the web site (as detailed in SOP 02) and collected by a courier (as detailed in SOP 03). The frozen biopsy will be used for microarray analysis only when a patient is confirmed to be lymph node-negative. Therefore it is essential that the results of local pathology determination of lymph node status are promptly recorded on the web page for both lymph nodenegative and node-positive individuals Paraffin block schedule The remaining breast cancer tissue will be fixed according to each center s policy (with at least one paraffin block) and analyzed by the institution s pathologist, who will determine the histopathology. A representative paraffin block for each patient enrolled in must be sent for central pathology review by post, at least once every 6 months to: Dr. Giuseppe VIALE, ISTITUTO EUROPEO DI ONCOLOGIA Via Ripamonti, 435 IT MILAN Italy Tel.: Fax: Giuseppe.Viale@ieo.it Blocks will be returned by post to each center once the central pathology review has been performed and TMAs produced. This will usually be within 2-3 weeks Serum sample schedule For those patients who consent to the optional blood sampling for serum storage for proteomics analysis, blood should be taken after the patient has signed the PIS & IC 1 and before the start of any protocol treatment. Serum should be separated and frozen according to SOP 15 and stored at - 80 C until the end of accrual when it should be sent to the TRANSBIG biological materials bank in Belgium. Version / January, 2006

97 study timeline guidelines from diagnosis to start of treatment max 8 weeks max 21 days max 21 days Signing of spis & IC Patient receives general information surgeon / research nurse A. B. T1 T2 T3a T3b T4 Diagnosis Hospitalization Surgery Surgical Oncologist post-op Start post-op visit visit Treatment Lymph node status known -ve Cancer Institute Key T = Hospital visit / Time point R = randomization PIS-IC = Patient information & Informed Consent C. 1. A. Tumor Diagnosis B. Patient signs screening PIS & IC C. Tumor sample sent to Cancer Institute D. Local pathology, Lymph node status determination E. RNA quantity & quality good, for LN ve PI told patient eligible & microarray analysis performed F. Genomic prognostic test performed, result communicated to PI after signing of PIS & IC 1 G. Patient returns home with R-C / R-E information to decide on participation. H. Patient signs PIS & IC 2 or 3 and brings/posts it to Center/signs at the start of treatment I. Patient inclusion in and randomization for discordant patients D. G. F. E. Discussion RNA QC good + Proposal for Randomizations microarray according to risk Chemotherapy and Hormonal therapy Information & signing of PIS & IC 1 (Discordant patients are randomized) This is a guideline for centers who will adapt this scheme to their local procedures R I. F. YES/NO Feedback from patient H. Signing of PIS & IC 2 or 3 Bring to Center during LVEF visit or post to Center V 3p Administrative responsibilities 21.1 The study coordinator The Study Coordinator (in cooperation with the Data Center) will be responsible for writing the protocol, reviewing all clinical data entered onto electronic case report forms and documenting his/her review on evaluation forms, discussing the contents of the reports with the Data Manager and the Statistician, and for publishing the study results. He will also generally be responsible for answering all clinical questions concerning eligibility, treatment, and the evaluation of the patients. Study coordinator: Dr. Emiel Rutgers address: The Netherlands Cancer Institute-Antoni Van Leeuwenhoekziekenhuis Plesmanlaan 121 NL 1066 CX Amsterdam The Netherlands Phone: Fax: E.Rutgers@nki.nl Version / January, 2006

98 Study co-coordinator: Dr. Fatima Cardoso address: Institut Jules Bordet 121 Blvd de Waterloo, B-1000 Brussels, Belgium Phone: Fax: fatima.cardoso@bordet.be Study co-coordinator: Dr. Martine Piccart address: Institut Jules Bordet 121 Blvd de Waterloo, B-1000 Brussels, Belgium Phone: Fax: martine.piccart@bordet.be 21.2 The EORTC Data Center The EORTC Data Center will be responsible for reviewing the protocol, collecting electronic case report forms, controlling the quality of the reported data, and generating reports and analyses in cooperation with the Study Coordinator. All methodological questions should be addressed to the EORTC Data Center. EORTC DATA CENTER 83, avenue Emmanuel Mounier, Bte 11 B-1200 Brussels, Belgium Fax: Registration of patients: Statistician: Jan Bogaerts Phone: Fax: jan.bogaerts@eortc.be Data Manager: Breast Group Data Manager (to be appointed) Fax: Project Managers: Frédéric Hénot Phone: Fax: frederic.henot@eortc.be Jillian Harrison Phone: Fax: jillian.harrison@eortc.be Version / January, 2006

99 Coordinating Physician: Dr. Patrick Therasse Phone: Fax: patrick.therasse@eortc.be Pharmacovigilance Unit: Nathalie Dubois Phone: Fax: pharmacovigilance@eortc.be The EORTC Pharmacovigilance Unit will forward all SAE within 24 hours of receipt to the EORTC Study Coordinator, the EORTC Data Manager and the contact persons at the pharmaceutical companies. All SUSARs will additionally be notified to all NCC/G s who will in turn inform all participating investigators and EC s from their group. The EORTC Pharmacovigilance Unit will take in charge the expedited reporting to the Competent Authorities, unless otherwise specified. The EORTC Pharmacovigilance Unit will prepare the Annual Safety report and distribute it to the Competent Authorities and the NCC/G s Pharmaceutical company contact person details Novartis Pharma AG Nora Schenk, Pharm D Novartis Pharmaceuticals Corporation Oncology Business Unit One Health Plaza, Blg 105 East Hanover, NJ , USA Tel: F. Hoffmann La Roche Cristella LaRosa Clinical Trial Manager-Consultant Pharma Business, Hoffman-LaRoche Inc. 340 Kingsland Street Nutley, NJ Tel: Fax: Sanofi-Aventis: Medical Sanofi-Aventis: Operational & Financial Isabelle Tabah-Fisch, MD Blandine Boussard Head of Medical Affairs Oncology, GMA ClinOps-Study Operations, Trial Manager Corporate Oncology Franchise Oncology Sanofi-Aventis Sanofi-Aventis 46, Quai de la Rapée, Paris, France 46 Quai de la Rapée, Paris cedex 12, France Tel Tel: + 33-(0) Fax: Fax: + 33-(0) Version / January, 2006

100 21.3 The TRANSBIG Secretariat Breast International Group (BIG)-aisbl Institut Jules Bordet / 121 Blvd de Waterloo, 7th floor B-1000 Brussels, Belgium Tel: Fax: transbig@bordet.be The EORTC Group All questions concerning membership in the group should be addressed to the chairman and/or secretary of the group. EORTC Breast Cancer Group Chairman: Dr. Emiel Rutgers Phone: Fax: E.Rutgers@nki.nl Secretary: Dr. Hervé Bonnefoi Phone: Fax: // herve.bonnefoi@hcuge.ch 21.5 National Coordinating Centers/Groups Country Austria Belgium Cyprus Czech Republic National structure contact details Medical University of Vienna - Department of Surgery A-1090 Wien, Waehringer Guertel 18-20, AUSTRIA Tel Fax michael.gnant@meduniwien.ac.at EORTC Data Center 83, avenue Emmanuel Mounier, Bte 11, B-1200 Brussels, Belgium Tel: /1655 Fax: Bank of Cyprus Oncology Centre- (BOCOC) Medical Oncology, 32 Acropoleos Avenue, 2006, Nicosia, Cyprus Tel Fax yiola.marcou@bococ.org.cy Charles University Hospital Oncology dept., U nemocnice 2, Prague 2, , Czech Republic. Tel Fax petruzelka.lubos@seznam.cz Version / January, 2006

101 France Pharmacovigilance & mailbox functions Germany Ireland Italy Luxembourg Poland Scandinavia (Sweden, Denmark, Finland & Norway) Serbia Montenegro Institut Gustave Roussy (IGR) 39, rue Camille Desmoulins, Villejuif cedex, France Tel: Fax: pelissier@igr.fr Fédération Nationale des Centres de Lutte Contre le Cancer (FNCLCC) 101, rue de Tolbiac, PARIS cedex 13, FRANCE Tel : Fax : al-martin@fnclcc.fr West German Study Group (WSG) gynaecological hospital - university clinics Duesseldorf, Mooresnstraße 5, Duesseldorf , Germany Tel / Fax / nitzu@uni-duesseldorf.de Irish Clinical Oncology Research Group (ICORG) 120 Pembroke Road, Ballsbridge, Dublin 4, Ireland Tel: Fax: brian.moulton@icorg.ie Italian oncology Group for clinical research (GOIRC) Medicina e Oncologia Medica, Azienda Ospedaliera di Cremona, Viale Concordia 1, Cremona, Italy. Tel Fax passalacqua.aioc@e-cremona.it Centre Hospitalier Luxembourg (CHL) Rue Barble, 4, L-1210 Luxembourg, Luxembourg Tel Fax Duhem.Caroline@chl.lu Medical University of Gdansk (AMG) Department of Oncology and Radiotherapy, Debinki 7, Gdansk, , Poland Tel Fax jjassem@amg.gda.pl Karolinska Institute (KI) Department of Oncology, Karolinska University Hospital Solna, SE Stockholm, Sweden Tel Fax nils.wilking@karolinska.se EORTC Data Center 83, avenue Emmanuel Mounier, Bte 11, B-1200 Brussels, Belgium Tel: /1655 Fax: Version / January, 2006

102 Slovenia Spain The Netherlands Pharmacovigilance & mailbox functions Turkey UK Institute of Oncology Ljubljana (IOL) Medical Oncology, Zaloska 2, 1000 Slovenia Tel Fax tcufer@onko-i.si SOLTI SANT LLORENÇ, 23-1º, 08202, Spain Tel: ; Fax: josep.vazquez@gruposolti.org The Netherlands Cancer Institute-Antoni van Leeuwenhoekziekenhuis Plesmanlann 121, NL 1066 CX Amsterdam, The Netherlands. Tel Fax E.Rutgers@nki.nl University Hospital Maastricht (UM) Internal medicine, PO Box 5800, Maastricht, 6202 AZ, The Netherlands Tel Fax PHU@SINT.AZM.NL Marmara Universitesi Hastanesi Tıbbi Onkoloji Bölümü Tophanelioglu Cad No:13-15 Altunizade Altunizade Istanbul Tel: Fax gulbasaran@superonline.com South West Wales Cancer Institute Singleton Hospital Sketty Lane Swansea SA2 8QA Tel: Fax: robert@swwci.vianw.co.uk 22 Trial sponsorship and financing EORTC is the legal European Sponsor unless otherwise specified for legal or logistical reasons. The Director General of the EORTC is: Professor Françoise Meunier EORTC Central Office Avenue Mounier 83, Bte 11 B-1200 Brussels, Belgium Tel: Fax: francoise.meunier@eortc.be Version / January, 2006

103 Financial support has primarily been provided by: Industrial partners Novartis F. Hoffman La Roche Sanofi-Aventis The European Commission Framework Programme VI (FP6-LSHC-CT ), the NCI (through the EORTC) and Charitable trusts including the Breast Cancer Research Foundation, the Jacqueline Seroussi Memorial Foundation and the Prix Mois du Cancer du Sein. Indirect financial support has been provided by Agendia (via reduced cost for performing microarray analysis). 23 Trial insurance A clinical trial insurance has been taken according to the laws of the countries where the study will be conducted. An insurance certificate will be made available to the participating sites at the time of study initiation. Clinical trial insurance is only valid if the treatment is given in a center authorized by the EORTC Data Center and which has obtained Ethical Committee approval (individually or centrally depending on the national regulations applicable). Therefore all centers will have to declare to their NCC/G, who will inform the EORTC Data Center, of details of other satellite institutions which may be responsible for providing protocol treatment to the patients. Details on these satellite institutions including CV for the local investigator, laboratory normal ranges and the Ethical Approval, will have to be transmitted to the NCC/G. Correspondence on study issues and data collection will however only be performed with the primary institution which will assume all responsibilities and liabilities issues with the principle investigator. Version / January, 2006

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108 Ref 61. Fisher, B. et al. Treatment of lymph-node-negative, oestrogen-receptor-positive breast cancer: long-term findings from National Surgical Adjuvant Breast and Bowel Project randomised clinical trials. Lancet 364, (2004). Ref 62. Fisher, B. et al. Tamoxifen for prevention of breast cancer: report of the National Surgical Adjuvant Breast and Bowel Project P-1 Study. J Natl Cancer Inst 90, (1998). Ref 63. Gorin, M.B. et al. Long-term tamoxifen citrate use and potential ocular toxicity. Am J Ophthalmol 125, (1998). Ref 64. Goss, P.E. et al. A randomized trial of letrozole in postmenopausal women after five years of tamoxifen therapy for early-stage breast cancer. N Engl J Med 349, (2003). Ref 65. Elisaf, M. et al. The beneficial effect of tamoxifen on serum lipoprotein-a levels: an additional anti-atherogenic property. Anticancer Res 16, (1996). Ref 66. Rossouw, J.E. et al. Risks and benefits of estrogen plus progestin in healthy postmenopausal women: principal results From the Women's Health Initiative randomized controlled trial. Jama 288, (2002). Ref 67. Thurlimann, B.J. et al. BIG 1-98: Randomized double-blind phase III study to evaluate letrozole (L) vs. tamoxifen (T) as adjuvant endocrine therapy for postmenopausal women with receptor-positive breast cancer. Proc Am Soc Clin Oncol 23, (2005). Ref 68. Wasan, K.M. et al. The influence of letrozole on serum lipid concentrations in postmenopausal women with primary breast cancer who have completed 5 years of adjuvant tamoxifen (NCIC CTG MA.17L). Ann Oncol 16, (2005). Ref 69. Harper-Wynne, C. et al. Effects of the aromatase inhibitor letrozole on normal breast epithelial cell proliferation and metabolic indices in postmenopausal women: a pilot study for breast cancer prevention. Cancer Epidemiol Biomarkers Prev 11, (2002). Ref 70. Elisaf, M.S. et al. Effect of letrozole on the lipid profile in postmenopausal women with breast cancer. Eur J Cancer 37, (2001). Ref 71. Baum, M. et al. Anastrozole alone or in combination with tamoxifen versus tamoxifen alone for adjuvant treatment of postmenopausal women with early-stage breast cancer: results of the ATAC (Arimidex, Tamoxifen Alone or in Combination) trial efficacy and safety update analyses. Cancer 98, (2003). Ref 72. Howell, A. et al. Results of the ATAC (Arimidex, Tamoxifen, Alone or in Combination) trial after completion of 5 years' adjuvant treatment for breast cancer. Lancet 365, 60-2 (2005). Ref 73. Dowsett, M. ATAC trialists group: Analysis of time to recurrence in the ATAC trial according to estrogen receptor and progesterone receptor status. Breast Cancer Res Treat 82, S7 (2003). Ref 74. Coombes, R.C. et al. A randomized trial of exemestane after two to three years of tamoxifen therapy in postmenopausal women with primary breast cancer. N Engl J Med 350, (2004). Version / January, 2006

109 Ref 75. Coombes, R.C., Hall, E., and Snowden, C. The Intergroup Exemestane study: a randomized trial in postmenopausal patients with early breast cancer who remain disease free after two to three years of tamoxifen- updated survival analysis. Br Cancer Res Treat 88, (2004). Ref 76. Jackesz, R., Kaufman, M., and Gnant, M. Benefits of switching postmenopausal women with hormone -sensitive disease to anastrozole after 2 years of adjuvant tamoxifen. Breast Cancer Res Treat 88, S7 (2004). Ref 77. Boccardo F. M., Rubagotti A., Puntoni M., et al. Switching to anastrozole (ANA) vs continued tamoxifen (TAM) treatment of early breast cancer (EBC). Updated results of the Italian tamoxifen anastrozole (ITA) trial. ASCO 2005, abstract 526, (2005). Ref 78. Boccardo, F. et al. Switching to Anastrozole Versus Continued Tamoxifen Treatment of Early Breast Cancer: Preliminary Results of the Italian Tamoxifen Anastrozole Trial. J Clin Oncol, 23, (2005). Ref 79. Saphner, T., Tormey, D.C., and Gray, R. Annual hazard rates of recurrence for breast cancer after primary therapy. J Clin Oncol 14, (1996). Ref 80. Goss, P.E. et al. Randomized trial of letrozole following tamoxifen as extended adjuvant therapy in receptor-positive breast cancer: updated findings from NCIC CTG MA.17. J Natl Cancer Inst 97, (2005). Ref 81. Jakesz, R., Samonigg, R., Griel, R., et al. Extending adjuvant treatment with anastrozole: Results from the Austrian Breast and Colorectal Cancer Study Group Trial 6a (ABCSG-6a). American Society of Clinical Oncology, abstract 527 (2005). Ref 82. Winer, E.P. et al. American Society of Clinical Oncology technology assessment on the use of aromatase inhibitors as adjuvant therapy for postmenopausal women with hormone receptor-positive breast cancer: status report J Clin Oncol 23, (1920). Ref 83. Rutgers, E.J. Guidelines to assure quality in breast cancer surgery. Eur J Surg Oncol 31, (2005). Ref 84. EORTC Breast Cancer Group. Manual for clinical research and treatment in breast cancer. (2005). Ref 85. Benefit of a high-dose epirubicin regimen in adjuvant chemotherapy for node-positive breast cancer patients with poor prognostic factors: 5-year follow-up results of French Adjuvant Study Group 05 randomized trial. J Clin Oncol 19, (2001). Ref 86. Stewart, D.J. et al. Cyclophosphamide and fluorouracil combined with mitoxantrone versus doxorubicin for breast cancer: superiority of doxorubicin. J Clin Oncol 15, (1997). Ref 87. Budman, D.R. et al. Dose and dose intensity as determinants of outcome in the adjuvant treatment of breast cancer. The Cancer and Leukemia Group B. J Natl Cancer Inst 90, (1998). Ref 88. Poole C. J., Earl H. M., Dunn J. A., et al. NEAT (National Epirubicin Adjuvant Trial) and SCTBG BR9601 (Scottish Cancer Trials Breast Group) phase III adjuvant breast trials show a significant relapse-free and overall survival advantage for sequential ECMF. Proc Am Soc Clin Oncol 22, 4, abstract 13. (2003). Version / January, 2006

110 Ref 89. Vukelja, S.J., Lombardo, F.A., James, W.D., and Weiss, R.B. Pyridoxine for the palmarplantar erythrodysesthesia syndrome. Ann Intern Med 111, (1989). Ref 90. Vukelja, S.J., Baker, W.J., Burris, H.A. 3rd, et al. Pyridoxine therapy for palmar-plantar erythrodysesthesia associated with taxotere. J Natl Cancer Inst 85, (1993). Ref 91. Cascinu, S., Fedeli, A., Fedeli, S.L., and Catalano, G. Octreotide versus loperamide in the treatment of fluorouracil-induced diarrhea: a randomized trial. J Clin Oncol 11, (1993). Ref 92. Hillner, B.E. et al. American Society of Clinical Oncology 2003 update on the role of bisphosphonates and bone health issues in women with breast cancer. J Clin Oncol 21, (2003). Ref 93. Kalbfleisch J.D., & Prentice R. L. The statistical analysis of failure time data. Wiley. (1980). Ref 94. Kim K. and DeMets D.L. Design and analysis of group sequential tests based on the type I error spending rate function. Biometrika 74, (1987). Ref 95. Baum, M. and Ravdin, P.M. Decision-making in early breast cancer: guidelines and decision tools. Eur J Cancer 38, (2002). Ref 96. Harrell, F.E. Jr, Lee, K.L., & Mark, D.B. Multivariable prognostic models: issues in developing models, evaluating assumptions and adequacy, and measuring and reducing errors. Stat Med 15, (1996). Version / January, 2006

111 Appendix B: WHO performance status scale Grade Performance scale 0 Able to carry out all normal activity without restriction 1 Restricted in physically strenuous activity but ambulatory and able to carry out light work. 2 Ambulatory and capable of all self-care but unable to carry out any work; up and about more than 50% of waking hours. 3 Capable of only limited self-care; confined to bed or chair more than 50% of waking hours 4 Completely disabled; cannot carry on any self-care; totally confined to bed or chair. Version / January, 2006

112 Appendix C : Calculation of the glomerular filtration rate (GFR) COCKCROFT AND GAULT FORMULA For the calculation of GFR age is measured in years and weight is measured in kilograms. If serum creatinine is measured in µmol/l, the following formula applies: In males: GFR[ml/min] = 1.23 x (140 age) x weight serum creatinine In females: GFR[ml/min] = 1.05 x (140 age) x weight serum creatinine If serum creatinine is measured in mg/dl, the following formula applies: In males: GFR[ml/min] = ( 140 age) 72 x serum x weight creatinine In females: GFR[ml/min] = 0.85 x (140 age) x weight 72 x serum creatinine Version / January, 2006

113 Appendix D: Common Terminology Criteria for Adverse Events In the present study, adverse events and/or adverse drug reactions will be recorded according to the Common Terminology Criteria for Adverse Events (CTCAE), version 3.0. At the time this protocol was issued, the full CTC document was available on the NCI web site, at the following address: The EORTC Data Center web site provides a link to the appropriate CTC web site. This link will be updated if the CTC address is changed. Investigators who do not have access to Internet can contact the Data Center to receive a hard copy of this document by mail. Version / January, 2006

114 Appendix E: Screening patient information sheet and informed consent document. Screening The screening patient information sheet & informed consent (spis & IC) gives consent for the donation of a piece of tumor tissue and for its being sent to a specialized cancer center and for the microarray analysis to be performed. We have enclosed an example template for this consent and each centre can choose whether they would like to use/adapt this template or they can use their center s standard PIS & IC if they are confident that it covers the sending of tissue and the microarray analysis. Each center will decide on a single spis & IC which will be used for all potential patients following local policies. Each center will sign a certificate to certify the content of the PIS & IC in use for screening, on which will specify that it is either: a faithful translation of the spis & IC template provided a modified translation of the template provided their own PIS & IC In the case of a center using a modified template or their own PIS & IC that it covers: the sending of tissue abroad the microarray analysis and does not state that an EORTC clinical trials insurance is in place for the screening period and also that a blank copy of the PIS & IC in use for screening is filed locally in the master file There will be a maximum of 3 PIS & ICs for this study. The first one, the PIS & IC 1, will be signed by all eligible patients before the two methods of risk assessment are compared and patients are enrolled/ randomized for treatment decision making tool. Patients who will receive chemotherapy and who agree to be randomized for chemotherapy regimen will also sign the PIS & IC 2 and patients who are eligible for endocrine therapy and who agree to be randomized for endocrine therapy regimen will sign the PIS & IC 3. Version / January, 2006

115 Patient Information Sheet Request for consent to send tumor tissue to another institution for specialized analysis Tumor removal, examination and breast cancer prognosis You have been diagnosed with breast cancer and will soon undergo an operation to remove your tumor. During the surgical procedure, lymph nodes located under the arm (called axillary lymph nodes) will also be removed. Lymph nodes work like filters in your body to catch things like germs or cancer cells. After surgery, the axillary lymph nodes, together with tissue from your tumor, will be examined closely under a microscope (pathological examination). The results of the pathological examination will help us determine the specific characteristics of your disease. These results will also give us some information about the prognosis of your breast cancer. In other words, the results will give us some information about whether it is likely that your breast cancer will come back - in which case additional treatment will be needed - or whether your breast cancer can be safely managed with local treatment alone (surgery with or without radiotherapy). The pathological examination of your tissue can be performed in this hospital and will take a maximum of 2 weeks. A new test for breast cancer prognosis We know that cancer can appear when some genes are damaged and some of the body s cells no longer function normally. Consequently, many scientists now think that by looking at the genes of tumor cells we will get even more information about a particular tumor than through pathological examinations. In other words, the genes of tumor cells may give us more accurate information about whether a woman s breast cancer is likely to be aggressive. Patients with aggressive tumors need more than local treatment and are given chemotherapy or hormonal therapy (also called systemic or adjuvant therapy). This hospital is involved in a research study that will look at a new and possibly more accurate test for determining breast cancer prognosis than traditional pathological examinations. The new test uses information about gene activity in tumor cells, and it was developed by researchers from the Netherlands Cancer Institute (Nederlands Kanker Instituut) in Amsterdam. The test is based on a technology called microarray (also called genomics), which determines which genes are active ( gene expression ) in a tumor, much like taking a fingerprint of it. The purpose of the research study this hospital is involved in is to find out if this new test is really better than traditional pathological examinations of tumor tissue for those patients who do not have cancer cells in the lymph nodes (called lymph node-negative breast cancer). If the new test is eventually shown to be more accurate than traditional pathological examinations, it will be easier for doctors to know which women really need systemic cancer treatment like chemotherapy and which women are likely to be cured by local treatment alone. Where can the new prognostic test be done? The new prognostic test is not available in this hospital. To have the gene expression test done, we would need to send a piece of your tumor tissue to the Netherlands where the technique was developed. After your surgery, and depending on the results of the pathological examination of your axillary lymph nodes, we may invite you to participate in the research study investigating the new test, and we will provide you with detailed information at that time. However, because of the way this new prognostic test is designed and to avoid any delay in deciding if you need additional treatment beyond surgery, we have to send your tissue to the Netherlands shortly after your operation. Researchers there will perform the test and send the results to your treating physician. Version / January, 2006

116 It is important to note that this prognostic test is not a test to determine whether breast cancer runs in your family, since it is not your own genes that will be examined but those of your tumor. The test will simply determine the pattern of gene expression in your tumor tissue (the fingerprint ) by comparing it to the expression patterns that the Dutch researchers have found to be good (the tumor is unlikely to come back and may not need further treatment) and those they have found to be aggressive (the tumor is likely to need further treatment to prevent it from coming back). This is why this is not a genetic or hereditary test but a genomic one (analysis of the genes of the tumor). Once the results of the test are reported to your treating physician, he or she will then discuss with you how this information could be used to deliver the best treatment for you. The purpose of this document is therefore to ask you if you agree to have a piece of your tumor, which will be routinely removed during your surgery, sent to the Netherlands in order for its gene expression pattern to be analyzed. Enough of your tumor tissue will also remain at your hospital for any additional tests you may need in the future. What will happen after the test? Once the results of both the pathological examination and gene expression test are known, your treating physician will discuss the following options with you: 1. If there are no tumor cells found in your axillary lymph nodes you will be offered the opportunity to participate in the research study mentioned above. a) you may decide to participate in the research study. b) you may decide not to participate in this research study in which case you may ask for the piece of your tumor tissue that was sent to the Netherlands to be destroyed. c) you may decide not to participate in this research study but wish to donate that piece of your tumor tissue for other research studies not defined at the present time. 2. If tumor cells have been identified in one or more of your axillary lymph nodes the research study is not appropriate to your situation. You may decide to participate in a different study, appropriate for your situation, if there is such an opportunity in this hospital or elsewhere. Concerning the piece of your tumor tissue that was sent to the Netherlands: a) you may prefer that the piece of your tumor tissue that was sent to the Netherlands is destroyed. b) you may decide to donate that piece of your tumor tissue for other research studies not defined at the present time. What are my rights? Your privacy will be protected at all times and your tumor tissue will not be used to test whether your breast cancer is hereditary (that is, breast cancer runs in your family). Your tumor tissue will not be used for any other purpose unless you are contacted again with more information and a new request for your consent. There are no additional examinations, medical procedures, discomfort, risks or costs to you if you decide to give permission for a piece of the tumor that is removed during your operation to be sent for analysis to the Netherlands. Regardless of whether you decide to have a piece of your tumor sent to the Netherlands or not, your relationship with your physician will in no way be affected. Version / January, 2006

117 After you have read this information carefully and you have discussed it with your treating physician, we kindly ask you to complete the enclosed informed consent form if you decide to have a piece of your tumor sent to the Netherlands. We thank you warmly for your attention. Version / January, 2006

118 Informed consent form (Consent to send tumor tissue to the Netherlands for specialized analysis) I have received a copy of the Patient Information Sheet and have understood the information. I accept that a part of my tumor will be sent to the Netherlands, where it will be analyzed using the new genomic prognostic test. The options that will be available to me after the test results are known have been clearly explained. If tumor cells are found in my axillary lymph nodes and the new genomic test is not appropriate for me, I would like to donate the piece of my tumor tissue to research studies not defined at the present time. I have had enough time to take my decision. My consent to send tumor tissue to the Netherlands is completely voluntary, and it will not affect my relationship with my treating doctor. The data collected on my behalf will be strictly confidential and treated according to the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the local applicable laws My consent does not discharge the organizers of the research from their responsibilities, and I keep all my rights guaranteed by the law. Patient's name: Patient's signature: Date: Person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Investigator's name: Title/Position: Investigator's Signature: Date: This document has been prepared taking the following documents into account: - World Medical Association Declaration of Helsinki, adopted by the 18 th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( - ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, 2000 Version / January, 2006

119 Appendix F: Patient information sheet and informed consent document for participation in the clinical trial INFORMATION FOR THE NATIONAL COORDINATING CENTER/GROUP OR NATIONAL COORDINATOR: This document is an English version of the patient information sheet & informed consent 1 (PIS & IC 1) for participation in the clinical trial. The translation and national regulatory submission process of this document is the responsibility of the National Coordinating Center/Group or the National Coordinator for this trial. The EORTC would like the NCC/G to: - translate the patient information sheet and informed consent in preparation for the submission of the dossier to the ethics committee (the submission may be the responsibility of the NCC or the EORTC depending on the local regulations) - We would like you to adapt the text to national guidelines and sensibilities however you must tell the EORTC what you have changed and why and have our agreement on the new text. - send a copy of the approved translated document to the EORTC Data Center who will then incorporate it into the initiation package which will be sent to you to distribute to other national participating investigators. (When each center has changed the header to that of their individual hospital you will have to collect a paper version from each participating center and send these to the EORTC, i.e. a paper copy of each PIS & IC document that is in use for the trial in it s final form.) - the final translated and approved PIS & IC 1 must have a version number and date which must be quoted in all correspondence with local and national ethics committees. Version / January, 2006

120 Patient Information Sheet - Comparison of risk assessment methods in the trial 1. Introduction You have previously agreed for a part of your breast tumor to be sent to the Netherlands to look at the genes in your tumor (gene expression test). This was done after you signed a similar document (consent form) before your surgery. The gene expression test analyzed which genes were active or not in your tumor, and this gives us information about your prognosis (that is, the likelihood of your cancer coming back). Now we would like to invite you to participate in the clinical trial (a research study) called (Microarray In Node-negative Disease may Avoid ChemoTherapy). This study involves comparing the results of your gene expression test with the results obtained by the traditional method of looking at clinical and pathological information about your cancer. This includes your age, the size of your tumor, the way your cells look under the microscope (called grade ), and hormone receptors, which are like ears on cells that respond to hormones. The results of both the gene expression and the clinical-pathological tests are important because they will be used to decide if you will be proposed chemotherapy or not. The main objective of the trial is to determine which of the two types of tests (both called prognostic tests) gives us the best information necessary to decide which patients should receive chemotherapy. 2. Invitation to participate in the study Researchers in COUNTRY are initiating a clinical trial within the framework of an International research network* coordinated by NATIONAL GROUP/The EORTC* with patients that have a disease similar to yours. You are invited to take part in this clinical research project after having received full information about the study. (* details are provided in section 15) You are now being asked to consider taking part in this trial. You should make your decision after reading carefully the detailed information below and discussing any questions you have with your doctor or research nurse. This is so that you understand this trial and can choose what is best for you. If you decide to take part in this trial, you will need to sign the informed consent form which is included after the explanation of the trial. 3. How doctors decide what treatment is necessary for patients after surgery After breast cancer surgery, when the breast tumor and one or more of the lymph nodes from under your arm have been removed, doctors evaluate the risk of your cancer coming back. Traditionally, doctors do this by analyzing the clinical-pathological information already mentioned above: If the analysis of this information shows that your cancer has a high risk of coming back, you will be proposed chemotherapy. If your tumor has hormone receptors you will also be proposed endocrine therapy, often with tamoxifen (a treatment that stops the activity of certain hormones). Finally, if you had breast-conserving surgery (meaning your breast was not removed) you will also be proposed radiotherapy. These treatments - chemotherapy, endocrine therapy and radiotherapy - are Version / January, 2006

121 called adjuvant treatments, adjuvant because they are additional treatments after surgery that can prevent cancer from coming back. Chemotherapy, as you have probably heard, is a treatment which can have unpleasant side effects. Some of these side effects are short term, but others can be long term. In recent years our understanding of the biology of breast cancer has improved. We have learned that chemotherapy is only necessary and effective for breast cancer patients that have tumors with a high risk of coming back. Unfortunately, up to now, we have not always been able to identify these aggressive tumors. Because of this uncertainty, doctors tend to treat most patients with chemotherapy in order to not to risk missing someone who really will benefit from it. However, this leads to over-treatment. In other words, at the moment, some women receive chemotherapy without needing it because in reality their tumors are less aggressive than we think. The most important goal of the trial is to be able to more precisely identify which tumors are aggressive. By doing so, in the future only women who will really benefit from chemotherapy will receive it; and women who do not need this therapy can avoid its unpleasant and sometimes long term side effects. 4. A new way of making decisions about chemotherapy: a new prognostic test Tests that doctors use to get information about tumor aggressiveness are called prognostic tests or tools because they help make predictions about how likely it is that a cancer will come back. In the sections above, the traditional prognostic test, looking at clinical-pathological information has been described. Recently, scientists have developed a new and different way to analyze tumor aggressiveness and make predictions about prognosis. This new prognostic test is not based on the traditional clinical-pathological factors already described, but looks at the activity of 70 specific genes ( gene expression ) in your tumor using a technology called microarray technology. The trial will examine this new prognostic test. However, in addition to looking at the 70 genes that we think may give us the most information about your tumor, we will also look at the expression of all the other genes in your tumor. This is like having a fingerprint or an identity card of your tumor, and this may help us get additional information about how aggressive the cancer might be. It is important to understand that this new prognostic test, also called a genomic test, will look only at the genes in your tumor. It will not look at your own genes, that is, the ones that you pass on to your children. A test of your own genes is called a genetic test and provides hereditary information, for example, whether cancer (or other diseases) runs in your family. Again, the new prognostic test in is not a genetic test, but a genomic one. In using the new genomic prognostic test we hope to be able to better identify which women really need chemotherapy and which women can safely avoid this treatment without any threat to their long-term survival. We also hope to be able to better identify the minority of women who would not normally receive chemotherapy based on their clinical-pathological prognosis but for whom this treatment may be necessary to prevent their tumor from coming back. 5. Description of the clinical trial will test whether the new prognostic tool can help better identify which women need chemotherapy. To do so, the new genomic test must be compared to the traditional way of assessing the risk of the cancer coming back. In other words, if you participate in this study, your tumor s aggressiveness will be assessed by using both the common clinical-pathological criteria and the genomic test just described above. The results from the two types of tests will then be compared. For the vast majority of women (estimated to be two thirds) the two tests are expected to show the same results, meaning that the test results would agree about whether you should be proposed chemotherapy or not. Women whose tumors are shown to be aggressive and have a high risk of Version / January, 2006

122 recurrence by both tests will be proposed chemotherapy (see diagram 1 below, number 1) and women whose tumors are shown to be less aggressive and are at low risk of recurrence by both methods will not be proposed chemotherapy (see diagram 1 below, number 2). For a minority of women (estimated to be one third), we expect that the results of the two types of tests will not agree, that is, they will be discordant (see diagram 1 below, number 3). Diagram 1. Comparison of the two risk assessment methods Node-negative breast cancer (1) (3) Genomic : Clinical High : High risk risk Discordant Genomic : Clinical High : Low risk risk (2) Low : Low risk risk Genomic : Clinical Low : High risk risk Genomic : Clinical Patients in for whom the test results are discordant will participate in the part of the study comparing the two methods. These women with discordant risk assessment results will have which of the two methods used for determining their treatment chosen by chance (randomly) by a computer, this is called randomization. Neither you nor your doctor can choose which of the two treatments you will receive. By participating in this randomization, it means that you might be proposed chemotherapy according to your clinical-pathological test, even if the genomic test results would not recommend that you are proposed chemotherapy. Or the opposite might happen. You might not be proposed chemotherapy according to the clinical-pathological test, although the genomic test suggests you should have it. The various possibilities are summarized in Diagram 2 below. Your doctor or research nurse can help explain this to you. Diagram 2. Randomization of discordant cases Decide with clinical tool High clinical risk (Low genomic risk) - Chemotherapy Low clinical risk (High genomic risk) - No chemotherapy R High genomic risk (Low clinical risk) - Chemotherapy Decide with genomic tool Low genomic risk (High clinical risk) - No chemotherapy If the genomic tool is proven to be better than the traditional clinical pathological methods of risk assessment, we estimate that 10-20% fewer women will receive chemotherapy. So in this study Version / January, 2006

123 approximately one in ten women who would normally receive chemotherapy will be safely spared this treatment. In addition, we believe that the new test can also identify those women who have a low clinical risk but who would nevertheless require chemotherapy so one in twenty women who would normally not receive chemotherapy will receive it in order to prevent their breast cancer from coming back. The trial will include 6,000 women with similar disease to yours over a period of about 3 years or slightly more. The total time that you will receive treatment, depends on what is decided to be best for you. If you receive only chemotherapy, it will last for about 6 months. If you receive endocrine therapy, which has fewer side effects than chemotherapy and is a different type of treatment, this will last for 7 years. It is also possible that you will receive both types of therapy. As a patient in the trial you will also have regular visits with your study doctor or nurse for at least 7 years. All women who participate in the trial should be under effective contraception while taking trial medication. Your doctor will discuss with you which methods constitute effective contraception. 6. Description of the ultimate goal of the clinical trial The main goal of is to show that the new genomic prognostic test can help better identify which women require chemotherapy than the traditional clinical-pathological method without having a negative effect on a patient s long-term survival. It also aims to study the relationship between gene expression patterns and the way breast cancer develops. Understanding this relationship will help us to be better able to give treatments designed specifically for each individual patient s breast cancer. There are additional, longer-term research objectives that depend on the collection and banking of tumor tissue and blood samples from all participants in. 7. Description of foreseeable risks We do not know if the new prognostic test is better than the clinical-pathological method that we currently use - and that s why we are doing this trial. On the one hand, because your treatment was decided using the new genomic test, there might be less chance of your cancer coming back, you might not be treated with chemotherapy unnecessarily and you may live longer. On the other hand, might show that the new tool is less good than the current clinical-pathological method and, as a result, you may end up having received insufficient treatment. Although we are fairly confident that this is not the case, we are not completely sure. So there is a risk that by decreasing the amount of chemotherapy prescribed, some women may be under-treated, therefore increasing the chance that their cancer will come back. In order to quickly detect if this happens, the group of women who will not receive chemotherapy according to the genomic test but who would normally have received it according to the clinical-pathological test (an estimated 672 women of the total 6,000 participants) will be very closely monitored. However, as mentioned above, we feel confident that the genomic tool is quite accurate in identifying high and low risk patients. This test has been pre-tested in small studies using tumors from women that had their tumors removed about ten years ago. The results of these studies involving women from several different countries showed that the genomic test did predict which cancers would come back and which would not. Please consider that even if you receive all possible treatments, there is always a chance that your tumor may come back. If your cancer does come back at any time, regardless of whether you received chemotherapy or not, you will be given all necessary treatment according to the standard care of your hospital. This may include chemotherapy. Version / January, 2006

124 8. Expected benefits If you decide to participate in this trial there will be no financial or material compensation to you. No one knows if you will benefit personally from taking part in this study. However, your participation is very important to create knowledge of how to treat breast cancer. We have learned a lot about how to treat breast cancer because of the millions of women who have volunteered to participate in breast cancer clinical trials all over the world through the years. New relevant information that directly concerns your future health will be made available from your treating doctor at the institution where you have received treatment in this study. 9. Research on biological material - objectives and description You have already donated a piece of your tumor and consented for the new genomic prognostic test to be performed. In addition to this tumor donation, if you participate in this study, you will be asked to donate an optional blood sample before the start of treatment and perhaps another optional blood sample at the end of chemotherapy. These samples will be stored in what we call a biological materials bank. These samples will be used for measuring the types and amounts of proteins in your blood. This type of research is called proteomics. In the future, we hope that proteomics will help us predict if or when tumors will come back. Because proteomics is still a new type of research, your blood sample will not immediately be useful for seeing when and if your cancer may come back. However, it will help us to further develop this new technology, which will help other breast cancer patients in the future. The blood sample donation is optional; if you decide to donate 1 or maybe 2, 10 ml (1 tablespoon) blood samples, they will be stored anonymously in a biological materials bank. The biological materials bank will be under the guardianship of the international, academic, non-profit organization the Breast International Group (BIG*). (*see section 15 for details) Blood samples from the biological materials bank will be made available to scientific researchers in the future who want to improve the understanding and treatment of breast cancer. However, any researchers who want to use the blood samples for research will first have to have their project approved by a committee of academic researchers representing the groups participating in. All research projects that use samples from the biological materials bank will also have to be approved by local and national ethics committees. Your blood sample will not be used for any genetic tests (for example, hereditary tests to see if cancer runs in your family), nor to do research on any disease other than breast cancer. 10. Voluntary participation Your participation in the trial is entirely voluntary and you will be given enough time to make your decision. Your donation of a blood sample for future research is also entirely voluntary. In other words, you can decide to participate in without giving a sample of your blood. You are free to decide at anytime that you no longer wish to participate in the trial. You do not have to give any reasons for doing so. Leaving the trial will not affect your future treatment. It will also not have any negative effect on your relationship with your treating doctor or the hospital staff. The only information that we will keep for research and analysis is what has been collected about you until the time you decide to stop participating in the trial. If you wish you can ask for any of your tissue that has not been used to be destroyed. This patient information document and consent form invites you to participate in the main part of. It specifically asks you to participate in the comparison of the genomic and clinicalpathological risk assessment methods. Version / January, 2006

125 In the near future, we may invite you to participate in one or two other parts of the trial, depending on the treatment that is appropriate for you. If you are proposed chemotherapy, we will invite you to participate in a part of that compares two different types of chemotherapy. If your tumor cells contain hormone receptors, we will invite you to participate in the part of that compares two different endocrine therapies. The details of these other parts of and any potential benefits and risks, will be explained to you at the appropriate time by your doctor or research nurse. At that time, you will also receive separate patient information sheets and informed consent forms. We recognize that this is a long and possibly confusing process, but it is necessary in order to be sure that every patient entering the trial does so on the basis of having received proper information about every aspect of the study. 11. Data (information) and legal protection If you decide to participate in this part of, your privacy will be protected at all times. All information (personal, clinical, economic, and data from research on tumor and blood samples material) collected because of your participation in the trial will be treated according to all national applicable laws. However, your consent does allow for the information in your medical records to be linked to data coming from other sources such as cancer registries. Cancer registries are a way for countries to collect information about cancer in general, & to help to better understand, prevent and treat cancer. It is very important that all information collected in this trial is accurate. Therefore, from time to time, this collected information may be checked against your medical records. Only authorized persons (EORTC research staff, national and/or foreign health authority representatives) may have access to your medical records. Except for access to your information by such authorized persons, all information about you will be strictly confidential. Tumor and blood samples (biological material) Some of the pathological information about your tumor determined by the doctors in your hospital will also be analyzed by other pathologists who are experts in breast cancer. In, this group of experts will review a small fragment of some of the tumors of the 6,000 women participating in this trial. This is called a central pathology review and will be done in Milan, Italy. The purpose of this review is to ensure that is run according to the highest scientific standards possible. The handling of tumor and blood samples in will always be done in such a way that scientists analyzing it for research purposes will not be able to know your name or other personal information about you. The results of the research on biological material are unlikely to be available in the immediate future. This is because research can take a long time and tissue and blood samples and information must be collected from many patients before the results are known. Collaboration with third parties The mission of the EORTC and BIG, being non-profit research organizations, is to do everything they can in the best interests of cancer patients. Collaboration with third parties, including commercial companies, may be necessary for the EORTC and BIG to develop more effective anticancer treatments. This may lead to a company owning a discovery made using the biological materials bank. If the EORTC or BIG ever receive any money as a result of such a discovery, it will be reinvested in non-commercial cancer research to improve cancer care. The EORTC has asked your treating doctor to disclose any existing conflict of interest he/she may have as a result of his/her activities related to this trial. The EORTC has set up procedures to ensure the integrity of this process. Version / January, 2006

126 12. Insurance The EORTC has obtained clinical trial insurance in accordance with all applicable laws. If you need to undergo other medical treatment, we advise you to inform the doctor treating you in to ensure that the other medical treatment will not have any effect on your participation in the trial. Everything has been done and will continue to be done to prevent additional health problems occurring as a result of your taking part in this trial. (Specific clause per country following national law) 13. Ethics Committee A complete description of the research (the research protocol for ) has been submitted to the responsible ethics committee. The mission of this committee is to verify that all conditions with respect to your safety and rights are respected. Approval for this research has been given by the Ethics Committee of on 14. Contact persons In case of any problem or question, your doctor will be pleased to answer any further questions and may be contacted as follows: Name of the doctor: Hospital: Telephone: If you give your consent to join this trial, you will be given a telephone number of the hospital that you can contact at any time if you feel unwell or have further questions. With your agreement, if you have a family doctor we will also inform him or her about your taking part in this trial and what is involved Please take your time to read and think about this information, and do not hesitate to ask further questions to your doctor or research nurse if anything is unclear. You should keep a copy of this document after you and your doctor have signed it. 15. Responsibilities The clinical trial will be managed, coordinated and analyzed by the European Organization for Research and Treatment of Cancer (EORTC) which will act as the European legal sponsor of the trial. The EORTC Breast Group (a group of European cancer specialists interested in improving breast cancer treatment) is the coordinating group however in (COUNTRY) a group of breast cancer specialists from (NATIONAL GROUP/EORTC) will be responsible for the management the study. The EORTC is itself part of the non-profit, international academic network for breast cancer research called the Breast International Group (BIG), which has a laboratory research arm called TRANSBIG. The trial is being run as a joint project of the EORTC, BIG / TRANSBIG, and the participating national groups. This means that is being conducted as an international collaborative study under the legal framework of the EORTC and BIG. This research receives support through financial and other contributions from the European Commission s Framework Programme VI; the companies Novartis, F. Hoffmann-La Roche, Sanofi-Aventis, Agendia; and national charities. Version / January, 2006

127 INFORMED CONSENT FORM (Consent to participate in comparison of risk assessment methods in the trial) I have received a copy of the patient information and have understood the information. All my rights have been clearly explained to me. I have had enough time to take my decision. I agree to participate in the clinical trial called ( Microarray in Node-negative Disease may Avoid Chemo Therapy. A prospective, randomized study comparing the 70-gene signature with common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer), EORTC study number and BIG study number BIG My participation is completely voluntary. I accept that data resulting from this study can be linked to other sources (such as cancer registries) for research purposes. I understand that I have the right to withdraw my consent at any time without explanation. I also understand that if I withdraw my consent, it will not affect my relationship with my treating doctor. I understand that all the information collected on my behalf will be strictly confidential and treated according to the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the local applicable laws. I accept that future cancer research studies may be conducted on the biological material (tumor or blood) that I provide and that any data resulting from these studies can, in the future, be linked with other resources for research purposes. I agree to donate 1 or possibly 2, 10 ml sample(s) of my blood (1 tablespoon) for research purposes only. In case of the EORTC or the BIG / TRANSBIG research consortium agreeing to a collaboration with a third party (not already foreseen in the protocol), I agree that my biological material can be used by: Another academic institution/organization A commercial company (i.e. one of the pharmaceutical companies supporting ) My consent does not discharge the organizers of the research from their responsibilities and I keep all my rights guaranteed by the law. Version / January, 2006

128 Patient's name: Patient's signature: Date: Person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Investigator's name: Title/Position: Investigator's Signature: Date: This document has been prepared taking the following documents into account: World Medical Association Declaration of Helsinki, adopted by the 18 th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, Version / January, 2006

129 Appendix G: Patient information sheet and informed consent document for the clinical trial chemotherapy section. INFORMATION FOR THE NATIONAL COORDINATING CENTER/GROUP OR NATIONAL COORDINATOR: This document is an English version of the patient information sheet & informed consent 2 (PIS & IC 2) for the chemotherapy therapy study in the clinical trial. The translation and national regulatory submission process for this document is the responsibility of the National Coordinating Center/Group or the National Coordinator for this trial. The EORTC would like the NCC/G to: - translate the patient information sheet and informed consent in preparation for the submission of the dossier to the ethics committee (the submission may be the responsibility of the NCC or the EORTC depending on the local regulations) - There are sections which you must NOT change the meaning of the text in the translation; these are sections 1, 2, There are certain sections where you can adapt the text to national guidelines and sensibilities however you must tell the EORTC what you have changed and have our agreement on the new text. The sections that can be adapted are send a copy of the approved translated document to the EORTC Data Center who will then incorporate it into the initiation package which will be sent to you to distribute to other national participating investigators. (When each center has changed the header to that of their individual hospital you will have to collect a paper version from each participating center and send these to the EORTC, i.e. a paper copy of each PIS & IC document that is in use for the trial in it s final form.) - the final translated and approved PIS & IC 2 must have a version number and date which must be quoted in all correspondence with local and national ethics committees. Version / January, 2006

130 Chemotherapy question in trial (Randomization-Chemotherapy) You have previously agreed for a part of your breast tumor to be sent for special analysis (screening informed consent) and consented to be included in the clinical trial called where the new test has been compared to current clinical risk assessment (informed consent 1). In accordance with the results of your risk assessments you will shortly receive chemotherapy. We would like to invite you to participate in the chemotherapy question of the study, described below. 1. Title of the research protocol As part of the - Microarray In Node-negative Disease may Avoid ChemoTherapy - trial (A prospective, randomised study comparing the 70-gene signature with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer) the following chemotherapy therapy question will be asked. Can anthracycline-based chemotherapy be safely replaced by the combination of Docetaxel- Capecitabine as adjuvant treatment of node-negative breast cancer? 2. Invitation to participate in the study The EORTC Breast Group (a group of cancer specialists interested in improving breast cancer treatment) is initiating a clinical trial with patients that have a disease similar to yours. The study will be conducted at the International level under the supervision of doctors recognized as experts in this field of medicine. You are invited to take part in this clinical research project after having received full information about the study and the implications for you as a patient. 3. Introduction Your breast tumor was removed by surgery however there may be a possibility that it may come back, this is called relapse. Therefore current medical opinion recommends the use of chemotherapy (called adjuvant chemotherapy) to try to prevent your cancer from coming back. Current standard care consists of a combination of several chemotherapy drugs, which constitutes a regimen, and almost always contains a drug called anthracycline. This is the most active drug of this combination but it is also quite toxic and can cause not only short term toxicity during treatment but also long term side effects that may only appear in the future such as leukemia and heart disease. Although this risk is small it exists and these diseases are serious. New drugs active against breast cancer have recently been developed and tested in other studies showing apparently fewer long term side effects. The present study has been designed to test a combination of two of these drugs (docetaxel and capecitabine) by comparing the new combination to the current standard treatment and to detect if it has an additional benefit in terms of efficacy (increase duration of life without the cancer coming back) and safety (fewer long term side effects). All women who participate in the trial should be under effective contraception for the entire duration of the study. Your doctor will discuss with you which methods constitute effective contraception. Version / January, 2006

131 4. Description of the research This research will involve approximately 4,000 patients in several countries throughout the world but principally in Europe. You will receive either docetaxel-capecitabine chemotherapy (chemotherapy A the new regimen) or an anthracycline based chemotherapy (chemotherapy B the standard regimen) at doses that have been well studied in other research studies, and that are commonly used internationally. The main objective of this study is to compare the efficacy and safety of chemotherapy A to the efficacy and safety of chemotherapy B (by counting and comparing the number of relapses in patients treated with each chemotherapy regimen). The duration of treatment A and B will be similar, i.e. six or eight cycles with a total duration of approximately five months. The side effects are different and are detailed below. Both chemotherapy treatments are effective against breast cancer, but we don t know if one is better than the other in your particular situation. Treatment Selection No one knows which treatment is better for you therefore which of the two treatments you will receive will be decided by chance. This is called randomization. Your doctor will call a central statistical office, which will assign one of the two treatments to you. Your chances of getting either the new chemotherapy or the standard chemotherapy are equal (50% either way) neither you nor your doctor can choose the treatment. Chemotherapy A Docetaxel is a very active drug in breast cancer either alone or in combination with Capecitabine. You will receive 6 cycles of Docetaxel in combination with Capecitabine. Docetaxel will be given intravenously in the day care unit once every 3 weeks (infusion in your veins over one-hour in the day care unit). Capecitabine will be given in pill form, which must be taken twice a day for 14 consecutive days followed by 7 days with no medication (rest period). This set of 21 days forms one cycle. Both Docetaxel and Capecitabine will be provided free of charge. Chemotherapy B The four drugs that can be used in different combinations are 5-Fluorouracil (5-FU), Cyclophosphamide, Epirubicin and methotrexate. Sometimes, Doxorubicin is used instead of Epirubicin. The mode of delivery of this chemotherapy regimen may vary slightly from one hospital to the other according to the standard of care applied in each hospital. The drugs will be given intravenously in the day care unit usually once every 3 or 4 weeksweeks (it will take about 2 hours in the day care unit). In some regimens the treatment is given for 2 consecutive weeks followed by a two week rest period. We plan to give you 6 or 8 cycles (one cycle is 3 or 4 weeks) of this chemotherapy regimen. If you participate in this study you will not have to make more visits to hospital, nor undergo more examinations or blood samples during the chemotherapy period and after completion of the study than patients treated for the same disease who are not taking part in the study. Results of the study will not be available until after the study is finished (recruitment of patient is expected to finish in 2009 and the study to have initial results 2012), but an individual patient will know on a regular basis the effectiveness and outcome of her treatment. Version / January, 2006

132 5. Description of foreseeable risks and discomforts As with most forms of chemotherapy it is likely that you will experience at least some side-effects, although the extent of these varies considerably from patient to patient. During your treatment, you are at a slightly higher risk of developing an infection. If at any time during chemotherapy you develop a fever ( 38 C when taken under the arm), you must contact your doctor (see phone number at the end of this sheet). During this treatment, you may feel tired. You may need to adjust your daily activities to help cope with this. Before having chemotherapy, you should let your physician know all of the medications you are taking, including over-the-counter medicines and any herbal supplements. Chemotherapy A Side effects you might experience with docetaxel When used as a single agent, the major toxic effect of docetaxel is neutropenia (decrease in the numbered white blood cells). It may be associated with fever (called febrile neutropenia) and infection. Other toxic effects may include allergic type reactions and skin reactions, nausea, vomiting, oral mucositis (inflammation of a mucous membrane), diarrhea, reversible paresthesia (skin sensation, such as burning, prickling, itching, or tingling), alopecia (loss of hair), fatigue, neurosensory symptoms, mild local venous reactions (phlebitis) at site of injection and fluid retention/edema. During these 6 cycles a treatment with oral corticosteroids will be given before each infusion of docetaxel and continued daily for 2 more days, as it has been shown to decrease the risk of allergy and fluid retention (edema) due to docetaxel. All these side effects are manageable with appropriate medical intervention, dose reduction and/or treatment interruption. Side effects you might experience with capecitabine Capecitabine is generally well tolerated. The most common side effects are hand-foot syndrome and diarrhea. Hand-foot syndrome is characterized by varying degrees of redness, tenderness, pain and desquamation (shedding/peeling off of skin) of the palms of the hands and the soles of the feet; numbness and tingling may also appear and in its most severe form it can interfere with daily activities. Nevertheless, it is reversible and responds to the diminution of the dose or the interruption of the treatment. If hand-foot syndrome occurs it is helpful to use a moisturizing cream and sometimes vitamin B6 supplements; your doctor will give you specific details if necessary. Diarrhea may also be an important side effect and like dermatitis, responds to changes in the treatment dose or schedule. Other side effects that may be associated with the use of capecitabine are: nausea, vomiting, stomatitis (inflammation of the mucous tissue of the mouth), dyspepsia (indigestion)/abdominal pain, fatigue, anorexia, myalgia (muscular pain or tenderness), insomnia, fever, myelosuppression (decrease in white blood cells and platelets), paresthesia (skin sensation, such as burning, prickling, itching, or tingling), eye irritation and serum (blood) biochemistry alterations. Other skin toxicities apart from the hand-foot syndrome include rash, dry and/or itchy skin. Before taking capecitabine, tell your doctor if you are allergic to capecitabine, fluorouracil, or any other drugs, what prescription and nonprescription medications you are taking, especially anticoagulants, antacids, folic acid, leucovorin, phenytoin (Dilantin), asprin and vitamins. Also do not have any vaccinations (e.g., measles or flu shots) without talking to your doctor. Version / January, 2006

133 Side effects you might experience with the combination capecitabine-docetaxel When capecitabine is used in combination with docetaxel, the incidence of neutropenia/neutropenic fever, gastrointestinal side effects and hand-foot syndrome may be higher, but still manageable with appropriate medical intervention, dose reduction and/or treatment interruption. Chemotherapy B General Side-effects with the drugs combination This combination of drugs has been used in breast cancer treatment for many years and short term and long term side effects are well known and documented. Side effects you might experience with anthracyclines (Epirubicin / Doxorubicin) The most common side effects are hair loss (alopecia), nausea, vomiting, myelosuppression (increasing the risk of infection, bruising, bleeding or anemia), sore mouth or taste change, discolored urine, darken skin, sensitivity to the sun, tiredness. All these side effects are still manageable with appropriate medical intervention, dose reduction and/or treatment interruption. Sometimes anthracyclines also have deleterious effects on the heart muscle decreasing the capacity of the muscle to contract correctly. Accordingly, this treatment is only delivered to women with no cardiovascular problems and your cardiac condition will be evaluated before the treatment. For your information, scalp cooling at the time of treatment can reduce hair loss in some cases. This may be available to you. However, you must expect some loss of hair. In case of severe hair loss a wig will be provided. It is important to note that your hair will grow back after your treatment is finished. Side effects you might experience with cyclophosphamide This drug is responsible for a transitory myelosuppression, with a decrease in white blood cells and platelets (increasing the risk of infection, bruising or bleeding). You will be carefully monitored with regular blood test and your treatment will be modified if necessary. Side effects you might experience with the combination of anthracyclines & cyclophosphimide This combination of drugs is also known to provoke secondary leukemia occurring usually 3 to 4 years after the end of the treatment. This is a rare side effect and globally (all patients taken together) the benefit of administering this drug is largely superior to the risk undertaken. Side effects you might experience with 5-Fluorouracil (5-FU) Following intravenous infusion of this drug, diarrhea, stomatitis (inflammation of the mucous tissue of the mouth) and conjunctivitis (inflammation of the conjunctiva of the eye) may occur. These side-effects are usually mild to moderate. In case of a severe side-effect your doctor will reduce the dose of this drug for following cycles. While using this medicine, and for 1 or 2 months after you stop using it, your skin may become more sensitive to sunlight than usual and too much sunlight may increase the effect of the drug. During this period of time: - Stay out of direct sunlight, especially between the hours of 10:00 a.m. and 3:00 p.m., if possible. - Wear protective clothing, including a hat and sunglasses. - Apply a sun block product that has a skin protection factor (SPF) of at least 15. Some patients may require a product with a higher SPF number, especially if they have a fair complexion. If you have any questions about this, check with your health care professional. - Do not use a sunlamp or tanning bed or booth. Version / January, 2006

134 Side effects you might experience with methotrexate When taking methotrexate, the following side-effects may be observed : bruising or bleeding, anaemia (low number of red blood cells), sore mouth and taste change, diarrhea, tiredness and a general feeling of weakness, skin changes (your skin may darken, due to excess production of pigment; this usually returns to normal a few months after the treatment finishes), gritty eyes. All drugs in this chemotherapy combination will be prescribed as they would be normally by your doctor following the standard of care applicable in the hospital. 6. Expected benefits If you accept to participate in this study we cannot be sure that there will be a direct benefit for you. However you will receive either the standard chemotherapy treatment or the new regimen which has already been extensively tested in the metastatic setting. If your disease progresses during the treatment allocated to you by randomization there is a high probability that you will be treated with the drugs of the other arm of this study (if there is no medical reason why you should not be) but outside the context of this clinical trial. If new information about the novel drug combination (docetaxel & capecitabine) becomes available it will be communicated to you by your treating doctor. As will any information that might affect your willingness to continue participation in the trial. We hope this research will teach us more about cancer. This will enable us to improve the standard of treatment and so help other patients with cancer in the future. However, nobody can predict whether you will directly benefit from participating in this clinical study. The anticipated benefits of this study for the breast cancer community include a better understanding of cancer and finding a medication regimen that has as good or better ability to control breast cancer with lower side effects than the commonly used regimen. 7. Voluntary participation Your participation in this clinical trial is entirely voluntary and you will be given sufficient time to decide whether or not you wish to participate. You are free to decide at any time without giving any reason that you no longer wish to participate in the trial. Such decision will not affect your subsequent treatment or relationship with your treating doctor or the hospital staff in any way. Medical data collected during your participation to the clinical trial as well as follow up data which will still be collected afterwards will be kept for research and analysis unless you specify otherwise. If you consent to join this trial, you will be given a telephone number at the hospital that you can contact at any time if you feel unwell or have further questions. We encourage you to take time to consider participation in this trial and to discuss it with your family, friends and your family doctor. 8 Data protection Your consent for participation in this part of the trial also includes your consent to allow the use of the data in your medical/clinical record to be used for research purposes. Your consent also includes allowing this data to be linked to data coming from other sources (such as cancer registries, medical/clinical records, ). All data (personal, clinical, economic and data coming from research on biological material) collected on your behalf will be treated in compliance with the national applicable laws. It is very important that the information collected is accurate and therefore, from time to time, this collected information may be checked against your medical records. Duly authorized persons (EORTC research staff, national and/or foreign health authority representatives) may have access to your medical records. With the exception of access by the duly authorized persons to your personal data on your medical record, all information will be strictly confidential. Version / January, 2006

135 This clinical research project is conducted under the legal framework of the EORTC with partial financial contribution of the European Commission, national charities and pharmaceutical companies. The EORTC, which is responsible for the conduct of this trial, has asked your treating doctor to disclose any existing conflict of interest he/she may have as a result of his/her activities related to this trial. The EORTC has set up procedures to ensure the integrity of this process 9. Insurance The EORTC who act as legal sponsor of the Study has obtained a clinical trial insurance in accordance with the applicable legislation. If you need to undergo another medical treatment, we advise you to inform the study doctor to ensure this will not have any effect on your participation to the trial. Everything has been done and will continue to be done to prevent additional health problems occurring as a result of your taking part in this trial. 10. Ethics Committee This research protocol has been submitted to the ethics committee whose mission is to verify that all conditions with respect to your safety and rights are respected. Approval to this research has been given by the Ethics Committee of on 11. Contact persons In case of any problem or question, your doctor will be pleased to answer any further questions and may be contacted as follows: Name of the doctor: Hospital: Telephone: If you consent to join this trial, you will be given a telephone number of the hospital that you can contact at any time if you feel unwell or have further questions. With your agreement, your family doctor (if applicable) will also be informed about your taking part in this trial and what is involved, if you agree. Please take your time to consider this information and do not hesitate to ask further questions to your doctor if anything is unclear. You are entitled to keep a copy of this document after you and your doctor have signed it. Version / January, 2006

136 Informed consent I have been properly informed about the clinical research study and have been given sufficient time to consider my participation. I have received a copy of the patient information sheet. All my rights have been clearly explained to me. I agree to participate in the clinical research study entitled - Microarray In Nodenegative Disease may Avoid ChemoTherapy in the chemotherapy therapy question titled Can anthracycline-based chemotherapy be safely replaced by the combination of Docetaxel- Capecitabine as adjuvant treatment of node-negative breast cancer? and to be enrolled in EORTC study number BIG I accept that any data resulting from this clinical research study can be linked with other resources for research purposes. My participation is completely voluntary and I have the possibility to withdraw my consent at any time without explanation. This will not affect my relationship with my treating doctor. The data collected on my behalf will be strictly confidential and treated according to the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the local applicable laws. I have been informed that the data (personal, clinical and biological material) collected may be used in the future for cancer scientific research purposes while confidentiality will be ensured. All data (personal, clinical and research on biological material) collected on my behalf will be treated in compliance with the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the national applicable laws. My consent does not discharge the organizers of the research from their responsibilities and I keep all my rights guaranteed by the law. Patient's name: Patient's signature: Date: Person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Investigator's name: Title/Position: Investigator's Signature: Date: This document has been prepared taking the following documents into account: World Medical Association Declaration of Helsinki, adopted by the 18 th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, Version / January, 2006

137 Appendix H: Patient information sheet and informed consent document for the clinical trial endocrine therapy section. INFORMATION FOR THE NATIONAL COORDINATING CENTER/GROUP OR NATIONAL COORDINATOR: This document is an English version of the patient information sheet & informed consent 3 (PIS & IC 3) for the endocrine therapy study in the clinical trial. The translation and national regulatory submission process of this document is the responsibility of the National Coordinating Center/Group or the National Coordinator for this trial. The EORTC would like the NCC/G to: - translate the patient information sheet and informed consent in preparation for the submission of the dossier to the ethics committee (the submission may be the responsibility of the NCC or the EORTC depending on the local regulations) - There are sections which you must NOT change the meaning of the text in the translation; these are sections 1, 2, There are certain sections where you can adapt the text to national guidelines and sensibilities however you must tell the EORTC what you have changed and have our agreement on the new text. The sections that can be adapted are send a copy of the approved translated document to the EORTC Data Center who will then incorporate it into the initiation package which will be sent to you to distribute to other national participating investigators. (When each center has changed the header to that of their individual hospital you will have to collect a paper version from each participating center and send these to the EORTC, i.e. a paper copy of each PIS & IC document that is in use for the trial in it s final form.) - the final translated and approved PIS & IC 3 must have a version number and date which must be quoted in all correspondence with local and national ethics committees. Version / January, 2006

138 Endocrine question in (Randomization endocrine-therapy) You have previously agreed for a part of your breast tumor to be sent for special analysis (screening informed consent) and consented to be included in the clinical trial called where the new test has been compared to current clinical risk assessment (informed consent 1). In accordance with the results of your risk assessments you may or may not have received chemotherapy. Your tumor expressed hormone receptors and therefore current clinical practice recommends that you receive endocrine therapy. We would like to invite you to participate in the endocrine therapy question of the study, described below. 1. Title of the research protocol As part of the - Microarray In Node-negative Disease may Avoid ChemoTherapy - trial (A prospective, randomised study comparing the 70-gene signature with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer). Is 7 years of upfront aromatase inhibitor, letrozole, more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? 2. Invitation to participate in the endocrine therapy section of the study The EORTC Breast Group (a group of cancer specialists interested in improving breast cancer treatment) is initiating a clinical trial with patients that have a disease similar to yours. The study will be conducted at the International level under the supervision of doctors recognized as experts in this field of medicine. You are invited to take part in this clinical research project after having received full information about the study and the implications for you as a patient. 3. Introduction Even though your breast cancer was removed by surgery there is a chance that it may come back (called relapse). Your tumor expressed (produced) hormone receptors which capture hormones produced by your body and thereby can stimulate the growth of any remaining tumor cells. Therefore current medical opinion recommends that you receive hormone therapy (which reduces or stops the signal between your body s hormones and their receptors) to help prevent your cancer from coming back. The old standard treatment for you would be 5-years of hormonal/endocrine therapy with a drug called tamoxifen. Recent results suggest that a new class of hormone therapy drugs (Aromatase Inhibitors) may be more effective than tamoxifen at preventing cancers, like yours, from coming back after surgery (treatment after surgery is called adjuvant therapy). However the ideal length of treatment or whether these drugs should be given following tamoxifen or instead of tamoxifen is not yet known. One of these new aromatase inhibitor drugs is called letrozole. As a part of the trial your doctor would like to ask you to participate in a study that will test whether 7 years of letrozole is more effective and safe than 2 years of tamoxifen followed by 5 years of letrozole as adjuvant treatment of breast cancer. Aromatase inhibitors are better studied in post-menopausal women with a disease similar to yours. However pre-menopausal women can also be invited to participate in this study at their doctor s discretion and as long as they agree to have the production of estrogen by the ovaries suppressed Version / January, 2006

139 (ovarian function suppression) while they take the hormonal therapy i.e. for a maximum of 7 years. Ovarian function suppression can be achieved either reversibly or irreversibly and your doctor will discuss the various options with you. All women who participate in the trial should be under effective contraception for the entire duration of the study. Your doctor will discuss with you which methods constitute effective contraception. 4. Description of the research The endocrine therapy part of this clinical trial will test a new regimen of adjuvant hormonal therapy which is given after surgery for breast cancer and is aimed at preventing any tumor cells still left in your body from developing and growing in your breast or in another part of your body. In addition to adjuvant hormone therapy you may also receive (or have already received) chemotherapy or radiotherapy. You have been asked to participate in a research study involving about 3,500 women throughout the world but predominantly in Europe. The purpose of this study is to find out whether it is better to receive a new drug, letrozole (registered as Femara ), for 7 years or tamoxifen for 2 years followed by letrozole for 5 years. To determine which treatment is better, half of the patients in the study will receive letrozole for 7 years and the other half will receive 2 years of tamoxifen followed by 5 years of letrozole. Treatment Selection No one knows which treatment is better for you therefore which of the two treatments you will receive will be decided by chance. This is called randomization. Your doctor will call a central statistical office, which will assign one of the two treatments to you. Your chances of getting either 7 years of letrozole or 2 years of tamoxifen followed by 5 years of letrozole are equal (50% either way), neither you nor your doctor can choose the treatment. Description of Treatment If you agree to take part in this study, you will get one of the following treatments: -7 years of letrozole (2.5 mg). This is a tablet that you will take once a day, by mouth, every day for seven years. - 2 years of tamoxifen (20 mg) followed by 5 years letrozole (2.5 mg). You will take a tablet of tamoxifen (20 mg) once a day, by mouth, for the first 2 years and after that a tablet of letrozole (2.5 mg) once a day, by mouth, every day for five years. Letrozole will be provided free of charge and tamoxifen will be prescribed by your doctor as it would normally be following the standard of care. Other Treatment Choices If you do not take part in this study, other choices are available. Alternatives to participating in this study would be to receive standard care, which at the present time is still 5 years of tamoxifen in some countries, 5 years of an aromatase inhibitor in others or a sequence of 2 to 3 years of tamoxifen followed by 2 to 3 years of an aromatase inhibitor. Your doctor will explain the standard treatment in your country and hospital. If you participate in this study you will not have to make more visits to hospital, nor undergo more examinations or blood samples during the endocrine therapy period and after completion of the study than patients being treated for the same disease who are not taking part in the study. Results of the study will not be available until after the study is finished, however an individual patient will know on a regular basis if the medication is working against her cancer. Version / January, 2006

140 Length of Study It is expected that you will take study medication for 7 years. If an examination shows that your cancer has come back, you will be told to stop taking the study medication. Treatment with letrozole or tamoxifen followed by letrozole will also be stopped if: - the side effects of this treatment become intolerable - new information indicates that this treatment is not in your best interest - you have taken this treatment for 7 years After you stop taking letrozole or letrozole followed by tamoxifen, your progress will be followed every 12 months. If new side effects or information about your disease or treatment are discovered during the study, you will be told. 5. Description of foreseeable risks and discomforts Possible Side Effects and Risks Generally, the observed adverse reactions with hormonal therapy are mainly mild to moderate in nature. Several of these side effects can be attributed to the normal effects of estrogen removal or blocking of the estrogen signal (are similar to symptoms of the menopause). Side effects you might experience with letrozole are: hot flushes (11% of patients) nausea (7% of patients) fatigue (5% of patients) headache/dizziness (7% of patients) edema (6% of patients) self reported osteoporosis, (7% of patients) Other side effects affecting between 1-10% of patients are: anorexia or appetite increase, vomiting, dyspepsia, diarrhea, hair loss, increased sweating, rash, myalgia (muscle pain), bone pain, athralgia (joint pain) and arthritis. Less frequent side effects affecting less than 1% of patients are: tumor pain, leucopenia, hypercholestrolaemia (high blood cholesterol), general oedema (swelling), depression, anxiety, somnolence, insomnia, memory impairment, dysaesthesia (unpleasant abnormal sensations), taste disturbance, cataract, eye irritation, blurred vision, palpitations, tachycardia (rapid beating of the heart), thrombophlebitis (inflammation of a vein forming a clot), hypertension, dyspnoea (shortness of breath), abdominal pain, stomatitis (swelling of soft tissues of the mouth), dry mouth, increased hepatic enzymes, pruritus (itching), dry skin, urticaria (hives), increased urinary frequency, vaginal bleeding, vaginal discharge, vaginal dryness, breast pain, pyrexia, mucosal dryness and thirst. These side effects are generally mild to moderate in nature, although unlikely, unforeseeable or unexpected risk(s)/side effects may occur. Letrozole will strongly decrease the body s estrogen levels. The effect of this lowering of estrogen may increase your chances of osteoporosis (thinning of the bones). These risks will be closely monitored. Side effects you might experience with tamoxifen are: hot flushes (up to 25% of patients) nausea and/or vomiting (up to 25% of patients) vaginal bleeding, vaginal discharge, menstrual irregularities and skin rash (less frequent) an increase in the risk of developing endometrial cancer Version / January, 2006

141 Some of the beneficial side effects of tamoxifen are: strengthening of bones and lowering of blood cholesterol The side effects documented for letrozole and tamoxifen are symptoms that may occur. It is unlikely, however, that a patient would experience all, or even most of them. You should immediately report any discomfort or problem to your doctor. The effectiveness of this new study medication may be endangered by the use of hormone replacement therapy drugs (oral, patch, vaginal creams). Additionally the majority of hormone replacement therapies may increase the risk of breast cancer coming back. Therefore you must inform the study personnel of any hormone replacement medication that you are taking and it may be necessary to discontinue the use of these drugs. 6. Description of the ultimate goal of the clinical research project The main objective of the endocrine part of the trial is to determine if 7 years of letrozole is more effective and safer than 2 years of tamoxifen followed by 5 years of letrozole. The trial will measure which treatment prevents your cancer from coming back for longer (longer disease free survival). It will also measure the short and long term side effects for each treatment to determine which is less toxic. 7. Expected benefits Possible Benefits If you agree to participate in this study we cannot be sure that you will benefit personally. However you will receive treatment that is standard of care in many European countries or an extended treatment regimen with aromatase inhibitors which are extensively used in cases where tamoxifen is no longer effective. There might be less chance of your cancer coming back, you might live longer. The quality of your life might be better. These things cannot be predicted for any individual patient, however we hope that information from this study will help cancer patients in the future. Early results from trials testing letrozole have shown it to be well tolerated and effective in advanced breast cancer (where the cancer has spread). Also some early results show letrozole to be better than tamoxifen in early breast cancer when taken for 5 years. However the long term efficacy and safety results have not been reported yet. Both tamoxifen and letrozole have relatively mild side effects as described above. If new information about letrozole or the sequential treatment of 5 years of letrozole after an initial 2 years of tamoxifen becomes available it will be communicated to you by your treating doctor, as will any information that may be relevant to your willingness to continue participation in the trial. 8. Voluntary participation Your participation in this clinical trial is entirely voluntary and you will be given sufficient time to decide whether or not you wish to participate. You are free to decide at any time, without giving any reason, that you no longer wish to participate in the trial. Such decision will not affect your subsequent treatment or relationship with your treating doctor or the hospital staff in any way. Medical data collected during your participation to the clinical trial as well as follow up data which will still be prospectively collected will be kept for research and analysis unless you specify otherwise. If you consent to join this trial, you will be given a telephone number at the hospital that you can contact at any time if you feel unwell or have further questions. We encourage you to take time to consider participation in this trial and to discuss it with your family, friends and your family doctor. Version / January, 2006

142 9. Data protection Your consent for participation in this part of the trial also includes your consent to allow the use of the data in your medical/clinical record to be used for research purposes. Your consent also includes allowing this data to be linked to data coming from other sources (such as cancer registries, medical/clinical records, ). All data (personal, clinical, economic and data coming from research on biological material) collected on your behalf will be treated in compliance with the national applicable laws. It is very important that the information collected is accurate and therefore, from time to time, this collected information may be checked against your medical records. Duly authorized persons (EORTC research staff, national and/or foreign health authority representatives) may have access to your medical records. With the exception of access by the duly authorized persons to your personal data on your medical record, all information will be strictly confidential. This clinical research project is conducted under the legal framework of the EORTC with partial financial contribution of the European Commission, national charities and pharmaceutical companies. The EORTC, which is responsible for the conduct of this trial, has asked your treating doctor to disclose any existing conflict of interest he/she may have as a result of his/her activities related to this trial. The EORTC has set up procedures to ensure the integrity of this process 11. Insurance The EORTC who act as legal sponsor of the Study has obtained a clinical trial insurance in accordance with the applicable legislation. If you need to undergo another medical treatment, we advise you to inform the study doctor to ensure this will not have any effect on your participation to the trial. Everything has been done and will continue to be done to prevent additional health problems occurring as a result of your taking part in this trial. 12. Ethics Committee This research protocol has been submitted to the ethics committee whose mission is to verify that all conditions with respect to your safety and rights are respected. Approval to this research has been given by the Ethics Committee of your hospital on 13. Contact persons In case of any problem or question, your doctor will be pleased to answer any further questions and may be contacted as follows: Name of the doctor: Hospital: Telephone: If you consent to join this trial, you will be given a telephone number of the hospital that you can contact at any time if you feel unwell or have further questions. With your agreement, your family doctor (if applicable) will also be informed about your taking part in this trial and what is involved, if you agree. Please take your time to consider this information and do not hesitate to ask further questions to your doctor if anything is unclear. You are entitled to keep a copy of this document after you and your doctor have signed it. Version / January, 2006

143 Informed consent I have received a copy of the patient information sheet. I have been properly informed about the clinical trial and have been given sufficient time to consider my participation. All my rights have been clearly explained to me. I agree to participate in the clinical research study entitled Microarray In Nodenegative Disease may Avoid ChemoTherapy in the endocrine therapy question titled: Is 7 years of upfront aromatase inhibitor, letrozole, safer and more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? and to be enrolled in EORTC study number BIG I accept that any data resulting from this clinical research study can be linked with other resources for cancer research purposes. My participation is completely voluntary and I have the possibility to withdraw my consent at any time without explanation. This will not affect my relationship with my treating doctor. The data collected on my behalf will be strictly confidential and treated according to the European and national applicable laws I have been informed that the data (personal, clinical and biological material) collected may be used in the future for cancer scientific research purposes while confidentiality will be ensured. All data (personal, clinical and research on biological material) collected on my behalf will be treated in compliance with the European and national applicable laws. My consent does not discharge the organizers of the research from their responsibilities and I keep all my rights guaranteed by the law. Patient's name: Patient's signature: Date: Person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Investigator's name: Title/Position: Investigator's Signature: Date: This document has been prepared taking the following documents into account: World Medical Association Declaration of Helsinki, adopted by the 18 th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, European Union Directive on the protection of individuals with regard to the processing of personal data (Dir/95/46/EC) Version / January, 2006

144 Appendix I: Capecitabine schedule and interruptions to schedule Normal Treatment: Day Day 1 8 Day 15 Day 22 Day 29 Day 36 Day 43 Day 50 Interruptions During Treatment: Interruptions are regarded as lost treatment days and the planned treatment schedule should be maintained. Version / January, 2006

145 ABCGS AJCC AI AML ARNO ASAT ASCO ATAC bid BIG BSA BSO CA 15-3 CAF CALGB CBC CEF CHF CMF CT CTCAE DCIS DEXA DFS DMFS DVT EBCTCG EGFR ER ET FAC FEC100 FISH FSH Appendix J: Abbreviations Austrian breast cancer study group American Joint Cancer Committee aromatase inhibitor acute myeloid leukemia German Adjuvant Breast Cancer Group alanine aminotransferase American Society of Clinical Oncology Arimidex and Tamoxifen Alone or in Combination bis in die Breast International Group Body surface area bilateral salpingo-oophorectomy cancer antigen 15-3 (protein product of the MUC1 gene) Cyclophosphamide Adriamycin 5-Fluorouracil the Cancer and Leukemia Group B complete blood count Cyclophosphamide Epirubicin 5-Fluorouracil congestive heart failure Cyclophosphamide Methotrexate 5-Fluorouracil Computed Tomography scan common terminology criteria for adverse advents ductal carcinoma in situ Dual Energy X-ray Absorptiometry disease free survival distant metastasis free survival deep vein thrombosis The Early Breast Cancer Trialists Collaborative Group epidermal growth factor receptor estrogen receptor endocrine therapy 5-Fluorouracil Adriamycin Cyclophosphamide 5-Fluorouracil (500 mg/m²) Epirubicin (100 mg/m²) and Cyclophosphamide (500 mg/m²) q3w 6 cycles Fluorescent in situ hybridization follicle stimulating hormone Version / January, 2006

146 5-FU GnRH HE HER-2 HRT ICH-GCP IES ITA LCIS LH LVEF MDS MRI MUGA NCC/G NCIC-CTG NSABP OS p.o. PET PgR PIS & IC pts QoL R-C R-E R-T SAE SERMS TMA TRANSBIG UNL WHO 5-FluoroUracil gonadotrophin releasing hormone hematoxylin and eosin Human Epidermal growth factor Receptor (also HER-2/neu) hormone replacement therapy International Conference on Harmonization-Good Clinical Practice Intergroup Exemestane Study Italian tamoxifen and anastrozole trial lobular carcinoma in situ luteinising hormone left ventricle ejection fraction myelodysplastic syndromes Microarray In Node-negative Disease may Avoid ChemoTherapy magnetic resonance imaging MUltiple Gate Acquisition scan national coordinating center/group National Cancer Institute of Canada Clinical Trials Group National Surgical Adjuvant Breast and Bowel Project overall survival per os position emission tomography progesterone receptor patient information & informed consent patients quality of life randomization for chemotherapy regimen randomization for endocrine therapy regimen randomization for treatment decision serious adverse event selective estrogen receptor modulators tissue microarray Translational research arm of BIG upper normal limit World Health Organization Version / January, 2006

147 EORTC Avenue E. Mounierlaan 83/11 Brussel 1200 Bruxelles België Belgique Tel: eortc@eortc.be EORTC BIG 3-04 Intergroup Study (EudraCT number ) (NCT ) (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy): A prospective, randomized study comparing the 70-gene signature with the common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0 to 3 positive nodes. Coordinating Group: EORTC Breast Cancer Group Collaborative Groups: TRANSBIG Consortium and BIG Groups (BOOG, UNICANCER, GOIRC, NCRI Breast Cancer Group, SOLTI, WSG, SI_IOL) Study Coordinator: Emiel Rutgers Co-coordinators: Fatima Cardoso and Martine Piccart Warning: This is an Intergroup study coordinated by the EORTC. The present protocol is written according to the EORTC template and is fully applicable to all participants. The PIS & IC (Appendix E, Appendix F, Appendix G, and Appendix H) are templates to be adapted by National Coordinating Centers / Groups to National and local requirements. Contact addresses for National Coordinating Centers / Groups are provided in the administrative chapter 21. May 10, 2005 PRC outline approval January 23, 2006 Full protocol approval Version 1.0 February 08, 2006 First administrative change Version 1.1 April 25, 2008 First amendment (substantial) Version 2.0 May 31, 2011 Second amendment (scientific) Version 3.0 May 21, 2014 Third amendment (scientific) Version 4.0 Version 4.0 / May 21, 2014 Copyright EORTC 2014

148 Contacts Study coordinators: EORTC (coordinating group): Steering Committee: Emiel Rutgers, Fatima Cardoso and Martine Piccart Breast Cancer Group Representatives from the EORTC, TRANSBIG, IDDI, NKI, Agendia, NCC/Gs, patient s organizations and representatives from participating groups and countries not already represented. Writing Committee: Clinical fellows: J. Bogaerts, EORTC Headquarters, Brussels, Belgium M. Buyse, IDDI, Brussels, Belgium F. Cardoso, Champalimaud Cancer Center, Lisbon, Portugal J. Harrison, EORTC Headquarters, Brussels, Belgium M. Piccart, Institut Jules Bordet, Brussels, Belgium E. Rutgers, NKI, Amsterdam, The Netherlands P. Therasse, EORTC Headquarters, Brussels, Belgium L. van t Veer, NKI, Amsterdam, The Netherlands J. Bines, Institut Jules Bordet, Brussels, Belgium S. Braga, Institut Jules Bordet, Brussels, Belgium G. Demonty, EORTC Headquarters, Brussels, Belgium S. Loi, Institut Jules Bordet, Brussels, Belgium C. Moulin, EORTC Headquarters, Brussels, Belgium G. Werutsky, EORTC Headquarters, Brussels, Belgium Collaborative groups: BOOG Borstkanker Onderzoeks Groep Nederland CEEOG Central and Eastern European Oncology Group FNCLCC Fédération nationale des centres de lute contre le cancer GOIRC Italian oncology group for clinical research IBCSG International Breast Cancer Study Group NCRI Breast Cancer Group National Cancer Research Institute Breast Cancer Group SAKK Swiss Group for Clinical Cancer Research SOLTI Grupo Español de Estudio y Tratamiento de Intensificación en Tumores Sólidos WSG Westdeutsche Studien Gruppe Version / 152 May 21, 2014

149

150 Table of contents: Protocol summary 10 Trial organization 17 1 Background and introduction General introduction Risk assessment in early breast cancer Early breast cancer: who can be safely spared adjuvant chemotherapy? Limitations of current prognostic factors Tumor size: women with small (<1cm) node-negative cancers (T1a,b,N0) Histological grade Patients with 1 to 3 positive lymph nodes: Do all of them benefit from chemotherapy? Molecular prognostic markers in breast cancer High throughput gene expression profiling and the development of a multi-gene prognostic signature for early breast cancer The Amsterdam 70-gene prognostic signature Independent validation of the 70-gene signature by the TRANSBIG research consortium Initial validation in lymph node negative patients Subsequent validation in lymph node-positive patients The trial: The first trial to evaluate the clinical value of a genomic tool Chemotherapy in the trial: can anthracycline-based chemotherapy be safely replaced by the combination of docetaxel-capecitabine as adjuvant treatment of early breast cancer? Adjuvant Chemotherapy Anthracyclines Taxanes The Combination Docetaxel-Capecitabine Chemotherapy in the trial Endocrine therapy in the trial: is upfront Aromatase Inhibitor (AI) letrozole for 7 years safer and more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? Adjuvant endocrine treatment in breast cancer Use of Aromatase Inhibitors in the adjuvant setting Tolerability profiles Hot flushes and general tolerability Skeletal effects Thromboembolic events and endometrial cancers Lipid metabolism 34 Version / 152 May 21, 2014

151 1.4.4 Improving adjuvant therapy in the first 5 years The sequential approach Extending adjuvant therapy beyond 5 years Which strategy is best? 37 2 Objectives of the trial Primary objectives Secondary objectives Endpoints Treatment decision randomization (R-T) Chemotherapy randomization (R-C) Endocrine therapy randomization (R-E) 40 3 Patient selection criteria General eligibility criteria for inclusion in the trial Selection criteria for treatment decision randomization (R-T) Selection criteria for chemotherapy randomization (R-C) Selection criteria for endocrine therapy randomization (R-E) 43 4 Trial Design Screening Enrollment Treatment decision randomization for chemotherapy allocation Chemotherapy randomization trial design Endocrine therapy randomization trial design pilot study 47 5 Guidelines for breast cancer surgery and adjuvant radiotherapy Breast cancer surgery Adjuvant radiotherapy 50 6 Chemotherapy: therapeutic regimens, expected toxicity, dose modifications Treatment plan, anthracycline arm Drug information, anthracycline arm Drug administration, anthracycline arm Premedication, anthracycline arm Patient monitoring anthracycline arm Dose adjustments anthracycline arm Treatment plan docetaxel-capecitabine arm Drug information docetaxel-capecitabine arm Docetaxel Capecitabine 58 Version / 152 May 21, 2014

152 Docetaxel-capecitabine combination Drug administration docetaxel-capecitabine arm Premedication docetaxel-capecitabine arm Patient monitoring docetaxel-capecitabine arm Dose adjustments docetaxel-capecitabine arm Concomitant therapy, docetaxel-capecitabine arm Treatment withdrawal criteria 69 7 Endocrine therapy Trial treatments Side effects of drugs Ovarian function suppression Concomitant treatments Treatment withdrawal criteria 72 8 Clinical evaluation, laboratory tests and follow-up Baseline evaluations Specifically to be done prior to chemotherapy randomization (R-C) Specifically to be done prior to endocrine randomization (R-E) During treatment Chemotherapy Endocrine therapy After the end of treatment (Follow-up) Follow-up for patients receiving neither chemotherapy nor endocrine therapy Chemotherapy follow-up For patients receiving chemotherapy and endocrine therapy For patients receiving only chemotherapy Endocrine therapy follow-up Summary table 76 9 Criteria of evaluation Time to event endpoints Types of recurrence Distant Metastasis Free Survival (DMFS) Disease Free Survival (DFS) Overall Survival (OS) Reporting of results Evaluation of toxicity Reporting of and general evaluation of adverse events Long-term chemotherapy toxicity 79 Version / 152 May 21, 2014

153 9.3.3 Serious adverse events Statistical considerations Statistical design Sample size Randomization and stratifications Statistical analysis plan Primary and secondary endpoints Analysis populations Timing of analyses Statistical methods Pre-planned sensitivity or exploratory analyses Prognostic factor analyses Data recoding and display End of study Stopping rules and interim analyses Pilot study of the trial Stopping rule after 800 patients enrolled Stopping rule to be applied every year Data monitoring Translational research Sample collection Central pathology review Proteomics Complex microarrays Biological materials bank storage Publication policy Investigator authorization procedure Patient enrollment procedure Enrollment procedure Access to RDC for registration/enrollment/randomization Overview of patient screening and enrollment Patient enrollment and randomization for treatment decision making tool (R-T) Randomization of chemotherapy (R-C) Randomization of endocrine therapy (R-E) Forms and procedures for collecting data Electronic case report forms and schedule for completion Data Flow (Remote Data Capture) 96 Version / 152 May 21, 2014

154 17 Reporting of Serious Adverse Events Definitions Reporting procedure Non- serious adverse events and/or non-serious adverse drug reactions Serious adverse events or serious adverse drug reactions Quality assurance Control of data consistency Quality Assurance and Quality Control Program Audits Ethical considerations Patient protection Subject identification Informed consent Sample schedule and trial logistics Frozen tumor sample schedule Paraffin block schedule Blood and serum sample schedule Trial logistics Administrative responsibilities The study coordinator The EORTC Headquarters The TRANSBIG Secretariat The EORTC Group National Coordinating Centers/Groups Trial sponsorship and financing Trial insurance 110 Version / 152 May 21, 2014

155 Table of appendices: Appendix A: References Appendix B: WHO performance status scale Appendix C: Common Terminology Criteria for Adverse Events Appendix D: World Medical Association Declaration of Helsinki Appendix E: Screening patient information sheet and informed consent document Appendix F: Patient information sheet and informed consent document for participation in the clinical trial Appendix G: Patient information sheet and informed consent document for the clinical trial chemotherapy section Appendix H: Patient information sheet and informed consent document for the clinical trial endocrine therapy section Appendix I: Capecitabine schedule and interruptions to schedule Appendix J: Abbreviations Version / 152 May 21, 2014

156 Protocol summary Title of the Study EORTC Protocol BIG 3-04 Objective(s) (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy) A prospective, randomized study comparing the Amsterdam 70-gene expression signature (70-gene signature) with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0 to 3 positive nodes. PRIMARY 1. The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to breast cancer patients with 0 to 3 positive lymph nodes. 2. The trial will also aim to compare a docetaxel-capecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens (anthracyclines followed by docetaxel for node positive). 3. The last primary objective will be to determine the best endocrine treatment strategy i.e. single agent up-front, aromatase inhibitor (AI) (letrozole) for seven years compared to the sequential endocrine strategy of 2 years of tamoxifen followed by 5 years of letrozole. SECONDARY Comparison of the two prognostic methods: To estimate both relative (Hazard Ratio, HR) and absolute (as % at 5 years) efficacy in terms of disease free survival (DFS), distant metastasis free survival (DMFS) and overall survival (OS) of chemotherapy in the two subgroups where the clinical-pathological prognosis and the 70-gene signature molecular prognosis are discordant. To calculate overall estimates of efficacy (DFS, DMFS, OS) for each treatment strategy according to clinical-pathological prognosis and according to molecular prognosis. To estimate the percentage of patients receiving chemotherapy per each prognostic method. Gene profiling: To identify and/or validate predictive gene expression profiles of clinical response/resistance to anthracycline-based and docetaxelcapecitabine chemotherapy. The identification and validation of novel gene expression signatures predicting clinical response to sequential tamoxifen-ai versus AI alone. Chemotherapy: To compare the OS distributions associated with the 2 chemotherapy arms To determine and compare the early and late toxicities associated with Version / 152 May 21, 2014

157 Methodology Number of patients Diagnosis and main criteria for inclusion each arm. Endocrine therapy: To evaluate adjuvant endocrine treatment success or failure in the subgroup of postmenopausal patients with endocrine responsive disease. To compare the safety profile of the two endocrine therapy regimens. Long term research objectives: To set up several biological material bank resources (RNA, tumor tissue, blood & serum) for future translational studies in both genomics and proteomics. Randomization for treatment decision making tool (R-T) of discordant cases (clinical-pathological prognosis using Adjuvant! Online vs. gene expression prognosis using the 70-gene signature), with additional randomizations for chemotherapy (R-C) and endocrine therapy (R-E) comparisons. Enrollment of 6,600 patients is planned with those having discordant risk assessments being randomized for treatment decision making. Up to 4,000 patients are planned for the chemotherapy randomization and 3,500 patients are planned for the endocrine therapy randomization. women with histologically proven operable invasive breast cancer, with 0 to 3 positive lymph nodes and no distant metastases. acceptable treatment options are: breast conserving surgery or mastectomy with either a sentinel node procedure or full axillary clearance. radiotherapy is mandatory in the case of breast conserving surgery and will be administered according to local institutional policy after mastectomy. patients with unresectable positive deep margins who will receive adjuvant radiotherapy are eligible provided that all other margins are negative. with a clinical tumor classification of T1, T2 or operable T3. the breast tumor must be unilateral. Multi focal tumors are allowed provided that they have identical histology.dcis or LCIS are allowed if invasive cancer is present. a frozen tumor sample (not fixed in formalin) must be available for inclusion. The tumor sample must be taken from the excised primary tumor (a core punch biopsy). patients should be aged 18 years at randomization. have a WHO performance status of 0 or 1. have adequate bone marrow reserves (neutrophil count >1.5 x109 /l and platelet count >100 x109/l), adequate renal function (serum creatinine 1.5 x upper limit of normal) and hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline phosphatase 2.5 x upper limit of normal and total bilirubin 2.0 x upper limit of normal). Version / 152 May 21, 2014

158 have NO serious cardiac illness or medical condition including but not confined to: a history of documented congestive heart failure (CHF), high-risk uncontrolled arrhythmias, angina pectoris requiring antianginal medication, clinically significant valvular heart disease, evidence of transmural infarction on ECG, poorly controlled hypertension (e.g. systolic > 180 mm Hg or diastolic > 100 mm Hg), symptomatic coronary artery disease or a myocardial infarction within the last 12 months or other risk factors that contra-indicate the use of anthracycline-based chemotherapy. NOT have serious uncontrolled intercurrent infections, or other serious uncontrolled concomitant disease. NOT have previous or concurrent cancer, possible exceptions are: adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, lobular or ductal carcinoma in situ of the breast and any invasive cancer (other than breast cancer) in complete remission for 5 years. NOT have received (neo) adjuvant chemotherapy, (neo) adjuvant endocrine therapy or radiotherapy for the primary breast cancer considered for this trial. NOT have participated in any investigational drug study within the 4 weeks preceding the start of treatment. NOT be pregnant or breast-feeding at the time of diagnosis or randomization. A woman of childbearing potential must have a negative pregnancy test. If post-menopausal, the woman must have been amenorrheic for at lest 12 months to be considered of non-childbearing potential while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, some IUDs, condoms, sexual abstinence or vasectomised partner (Note 3 of the guidance on non-clinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). NOT have any psychological, familial, sociological, geographical or serious uncontrolled medical (e.g. a history of uncontrolled seizures, central nervous system disorders, or psychiatric disability) condition judged by the investigator to be clinically significant which could potentially preclude informed consent or interfere with compliance for oral drug intake or with the study protocol and follow-up schedule. These conditions should be discussed with the patient before enrollment in the trial. signed written informed consent must be given according to ICH GCP, and national/local regulations before enrollment in the trial (both a screening PIS & IC and the PIS & IC 1 must be signed before enrollment). Version / 152 May 21, 2014

159 Treatment Test product, dose and mode of administration Duration of treatment Reference therapy, dose and mode of administration Criteria for evaluation Efficacy endpoints Safety The following women, if eligible, will be offered the chemotherapy randomization (R-C) comparing a commonly used anthracycline-based regimen (anthracyclines followed by docetaxel for node positive), selected from the list of acceptable regimens, and the new capecitabine-docetaxel combination: 1. Women that have a high risk according to the clinical-pathological criteria and a high risk according to the 70-gene signature prognosis. 2. Women that have a high risk according to the clinical-pathological criteria and a low risk according to the 70-gene signature, and were randomized (R-T) for chemotherapy treatment decision to use the clinicalpathological criteria. 3. Women that have a low risk according to the clinical-pathological criteria and a high risk according to the 70-gene signature, and were randomized (R-T) to use the 70-gene signature prognosis for chemotherapy decision. Trastuzumab will be allowed and recommended (however not provided free as a trial drug) for women with HER-2 positive tumors if it becomes approved and available for use in the adjuvant setting during the trial. Treatment with trastuzumab can be proposed either sequential or concomitant with chemotherapy (including patients in capectibainedocetaxel arm). All postmenopausal endocrine-responsive (i.e. ER and/or PgR positive) patients on the study will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. It will be left to the decision of the investigator whether or not a premenopausal woman can be randomized into this question, with concomitant ovarian function suppression (surgery, LHRH agonist or ovarian radiotherapy) during the complete duration of endocrine therapy. Chemotherapy will last for approximately 6 months and endocrine therapy for a maximum of 7 years. For the treatment decision randomization (R-T) the standard arm is risk assessment using clinical-pathological criteria (Adjuvant! Online). For the chemotherapy randomization (R-C) the standard arm is an anthracycline based chemotherapy regimen (anthracyclines followed by docetaxel for node positive) chosen from the list of allowed regimens. For the endocrine therapy randomization (R-E) the standard arm is sequential 2 years of tamoxifen followed by 5 years of letrozole. For R-T the primary endpoint is DMFS at 5 years. Secondary endpoints are: the proportion of women treated with chemotherapy per treatment decision making tool i.e. clinical prognosis compared to the 70-gene signature prognosis, OS at 5 and 10 years, DFS and safety (both early and late). For R-C and R-E the primary endpoint is DFS at 5 years. Toxicities for R-C and R-E will be recorded and summarized according to the Common Terminology Criteria for Adverse Events v3.0 (CTCAE). Additionally, cardiac toxicity and secondary leukemia, as potential longterm toxicities of chemotherapy will be monitored for a minimum of 15 years in all patients who were randomized for chemotherapy (R-C). Version / 152 May 21, 2014

160 Statistical methods Eligible patients with adequate material for microarray analysis and a successful 70-gene signature prognostic test will be enrolled. Risk will be determined using both the 70-gene signature and clinical-pathological criteria. Patients are evaluated as low clinical-pathological risk, if their 10- year disease specific survival (without chemotherapy or endocrine therapy) is estimated by Adjuvant! Online as greater than 88% for endocrine receptor (ER) positive patients, and greater than 92% for ER negative patients. Patients who are high risk for both methods will receive chemotherapy, and patients who are low risk for both methods will not receive chemotherapy. Patients for whom the methods are discordant will be randomized for treatment decision making (R-T) between chemotherapy-decision-making according to clinical-pathological criteria or chemotherapy-decision-making according to the 70-gene signature prognosis. Patients who are candidates for chemotherapy (see also treatment section) will be proposed for randomization R-C: between anthracycline based chemotherapy (anthracyclines followed by docetaxel for node positive) and docetaxel-capecitabine chemotherapy. All patients who are candidates for endocrine therapy will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. Primary test: In the set of patients who have a low risk 70-gene signature prognosis and high risk clinical-pathological prognosis, and who were randomized (R-T) to use the 70-gene signature prognosis and thus to receive no chemotherapy, a null hypothesis of a 5-year DMFS of 92% will be tested. With 6,600 patients accrued overall, this set has an expected size of 672 patients. The primary test will be a one-sided test at 97.5% confidence level. Test of hypothesis for R-T: - In the subgroup of clinical-pathological high risk and 70-gene signature prognosis low risk patients, assuming a 5-year DMFS of 93% without chemotherapy, with an expected size of 1,344 patients, there is 80% power to detect a hazard ratio of 0.47 (or a 5-year DMFS of 96.6% on chemotherapy) at the time of the primary analysis. - The subgroup of clinical-pathological low risk and 70-gene signature prognosis high risk, with an expected size of 576 patients, has low power to detect any realistic chemotherapy effect. Test of hypothesis for R-C: Assuming a control 5-year DFS of 86%, due to an insufficient number of patients randomized into this part of the study, there will be less than 80% power to detect the original alternative hypothesis of a hazard ratio of 0.76 (or 89% experimental 5-year DFS) at the time of the primary analysis. Test of hypothesis for R-E: Due to insufficient number of patients being randomized into this question, and assuming a DFS rate at 5 years of 86% on the control arm, there will be less than 80% power to detect the original alternative hypothesis of a hazard ratio of 0.75 (i.e. a 5-year DFS of 86% vs. 89.3%). The final analysis of the endocrine comparison will be performed when 379 events have been observed among the patients randomized for the endocrine question, yielding 80% power to detect a hazard ration of This is projected to occur at a median follow up of 7.9 years. Version / 152 May 21, 2014

161 Translational research Quality of Life Cost effectiveness/ health economic studies Objectives: See primary and secondary objectives. Biological material will be stored for the use in future translational research projects (see rationale & chapter 12). Whole genome complex arrays will be performed for all patients providing a unique opportunity to investigate the relationship between gene expression patterns and disease prognosis and response to treatment. No studies proposed Various sub studies are proposed. Version / 152 May 21, 2014

162 Screening Visit surgeon (T1, T2 or operable T3 & M0) screening PIS & IC, Registration Surgery* Tumor sample shipment, RNA extraction, microarray analysis, local pathology, TNM, lymph node status, HR status 0 to 3 positive nodes Signing of PIS & IC 1 & Enrollment Genomic/Clinical Prognosis review & randomization Low/Low Discordant R-T High/High No CT CT R -C HR - HR + HR + HR - R-E CT Chemotherapy R -T Treatment decision Randomization based on genomic vs clinical prognosis R -C Chemotherapy Randomization of Anthracycline based CT (FEC D for N+) vs Docetaxel /Capecitabine R -E Endocrine treatment Randomization Letrozole vs Tamoxifen followed by Letrozole HR Hormone receptor PIS & IC Patient information sheet and informed consent * Delay between surgery and start of systemic treatment ideally not more than 8 weeks and it cannot be > 120 days For details of timelines and trial logistics please see the Timelines from diagnosis to start of treatment diagram in Chapter 20. Version / 152 May 21, 2014

163 Trial organization This trial is an Intergroup trial, jointly conducted by several National and International cancer clinical research groups throughout the European Union and the rest of the world under the Breast International Group (BIG) / TRANSBIG umbrella. The EORTC is the coordinating group of this trial and will be the coordinating Data Center. It will centrally manage trial design and center activation, attribution of duties and responsibilities between participating research groups, data collection and quality control of data, statistical analysis and publication. Globally the EORTC will be the Sponsor. Some particular arrangements will be made, exceptionally, due to logistical or legal reasons and these will be communicated to investigators in these countries by their National Coordinating Center/Group (NCC/G). The EORTC will cover insurance in all countries except American and Australian continents. Precise details about insurance and an insurance certificate will be provided to Investigators at site initiation by their NCC/G. NCC/Gs will locally coordinate the conduct of the trial in their respective countries; one of their most important duties will be to manage the notification/ submission of all necessary documents to the Competent Authorities and to obtain the confirmation of the review by Institutional Review Board/local Ethics Committee if applicable following the national law. This protocol is to be followed by all participants. All chapters are fully applicable to all participants. All countries will conform to a standard administrative model where each center will contact their NCC/G-national structure who will provide them with all practical information. Participation in this trial is only possible through a national structure (coordinating group/center, NCC/G or EORTC Headquarters) designated for this purpose. For investigators who do not find contact details for a NCC/G in the Administrative Responsibilities chapter (Chapter 21), please contact the EORTC Headquarters directly and we will communicate the appropriate NCC/G contact details to you. The patient information sheets and informed consent templates (Appendix E, Appendix F, Appendix G, Appendix H) are provided as a basis for translation/adaptation by NCC/Gs to national/local regulations and sensibilities. Investigators will receive the translated and adapted PIS & ICs directly from their NCC/G or national structure. For the screening phase an example of possible text for the screening PIS & IC (spis & IC) is also provided however although investigators will have to certify the aptness of the PIS & IC to cover the genomic test, this consent is the responsibility of each individual center as it is signed prior to registration for screening. Investigators who are members of several groups participating in the trial should participate through the national structure detailed in the Administrative Responsibilities chapter and include all patients through this national structure. For EORTC members all patients will be counted for membership accrual independently of the group they participate through (see EORTC Policy 10). The investigational drugs (Docetaxel*, Capecitabine and Letrozole) will be supplied by the industrial partners. This trial is an academic trial with partial support from the European Commission Framework Programme VI (FP6-LSHC-CT ), industrial partners, the US NCI and charitable grants. * Footnote the docetaxel in the standard arm FEC D for lymph node positive patients will not be provided by the industrial partners Version / 152 May 21, 2014

164 1 Background and introduction 1.1 General introduction Breast cancer is the most common malignant disease in women, occurring more frequently in developed countries but showing a rising incidence in developing countries. Breast cancer is a public health issue on a global scale. According to estimates in 2002, there were 1,151,298 new cases of breast cancer diagnosed, 410,712 deaths caused by breast cancer and more than 4.4 million women living with breast cancer world wide (Ref. 1). Breast cancer is the second most commonly diagnosed cancer in the European Union (EU). An estimated 245,000 women were diagnosed with breast cancer in the EU in 2000, representing about a quarter of all female cancers diagnosed. Breast cancer is also the most common cause of cancer death in European women with 90,000 deaths in 2000 (Ref. 2, Ref. 3, Ref. 4). In 2004, in all of Europe, there were 371,000 estimated new cases of breast cancer diagnosed and 129,900 breast-cancer-related deaths (Ref. 5). At diagnosis, 90% of patients appear to have operable breast cancer, that is, disease that is confined to the breast and ipsilateral axilla. However up to 50% of these women will develop metastatic disease. Metastatic breast cancer is unfortunately incurable. As a result, since the mid 1980 s randomized trials of adjuvant systemic therapy (both endocrine and cytotoxic) have been conducted in an effort to reduce the rate of recurrence and to prolong the survival of patients with operable disease. Surgery is the main modality of treatment in patients with early breast cancer and, by itself or together with radiotherapy can control local-regional disease in the majority of patients. In spite of these treatments, subclinical tumor cells remaining following surgery may be responsible for the progression/recurrence of the disease at a later date. Adjuvant hormonal or cytotoxic treatments have been increasingly used with the goal of eradicating these remaining foci, in order to achieve longer survival and the eventual cure of the disease. The utilization of systemic treatment following surgery has become the standard of care and current objectives are to better target, more effective and less toxic treatments for breast cancer and to identify those patients who will have a clear benefit from these treatments. Current standard of care for node negative patients for those patients who will receive chemotherapy is 6 cycles of anthracycline based chemotherapy with the incorporation of taxanes for node-positive disease. For patients with endocrine responsive disease, the standard endocrine care consists of 5 years of tamoxifen, with many experts favoring the use of an aromatase inhibitor in sequence with or in replacement of tamoxifen. In the last 20 years, little progress has been made with respect to assisting oncologists in determining which early breast cancer patients require chemotherapy or other systemic therapy and which women can safely be treated with only loco-regional treatment. Great discomfort still persists in selecting those node-negative women and women with 1-3 positive nodes who need adjuvant chemotherapy (Ref. 6). 1.2 Risk assessment in early breast cancer Mortality rates from breast cancer rose until 1990 but fell afterwards in a number of European countries. Reasons for this decline include widespread mammographic screening, precise diagnosis and the extensive use of tamoxifen. Due to the widespread adoption of screening mammography in women of 50 years and older in developed countries, a growing proportion of women are being diagnosed with smaller tumors and no axillary node involvement. Although many of these women can enjoy long-term survival, 20-30% will relapse and die from their disease. Distant metastases account for the majority of these deaths and have been the impetus for proposing, in addition to optimal loco-regional treatment (surgery and radiotherapy), adjuvant systemic therapy to all fit women considered to be at moderate or high risk of relapse. There is however, much controversy Version / 152 May 21, 2014

165 related to the optimal definition of a low/minimal versus a moderate/high risk of relapse for these women with early breast cancer. Therefore, many oncologists rely on the guidelines issued by experts following consensus conferences, such as the 2000 National Institute of Health (NIH) meeting and the 2005 St. Gallen conference (Ref. 7, Ref. 8, Ref. 9). Given the close to 100% death rate from metastatic breast cancer, these guidelines are produced with the goal of avoiding undertreatment of affected women, and they assign only 15% to 20% of them to a low/minimal risk subset for which no adjuvant treatment at all or only adjuvant endocrine treatment will be considered. Nowadays, the great majority of early stage breast cancer patients are therefore offered some type of systemic chemotherapy. Unfortunately, whilst the majority of adjuvant chemotherapy regimens commonly used today are rarely life-threatening, they can have considerable both short and long term morbidity. As a result, many women with early breast cancer in the western world are probably over-treated, a phenomenon that may not only decrease the quality of life of these women but also increases the economic burden of this frequent disease on society. Furthermore, many advances in supportive care in recent years have facilitated the administration of adjuvant chemotherapy regimens to a greater number of patients, while also increasing its costs. Despite nodal status being historically one of the strongest prognostic factors used for predicting breast cancer clinical outcome, the Early Breast Cancer Trialist Collaborative Group (EBCTCG) overview recently reported an almost equal proportional reduction in recurrence and breast cancer death produced by adjuvant chemotherapy in node-positive and node-negative women less than 50 years old, and only a small difference in women older than 50 years (Ref. 10). Therefore, whereas both tumor size and nodal status are of prognostic significance, neither has substantial influence on the relative benefits of systemic therapy. However, these proportional benefits translate into larger absolute benefits for higher risk patients with node-positive disease than lower risk patients with node-negative disease. Hence, it is generally accepted that the absolute benefit for the higher risk patients is large enough to warrant the risks of adjuvant chemotherapy. For women who have early breast cancer, it is important to be able to accurately quantify the risk of recurrence and death so that the decisions about adjuvant chemotherapy can be taken appropriately. For these women, it is crucial to determine if there is a subgroup of breast cancers that, due to their excellent long term survival, can be spared adjuvant chemotherapy from which they are unlikely to benefit Early breast cancer: who can be safely spared adjuvant chemotherapy? Unfortunately little progress has been made with regards to new prognostic markers that can assist oncologists in treatment decision-making for early stage breast cancer. As a result, considerable differences exist worldwide in the selection of women whose risk of breast cancer recurrence justifies the need for adjuvant chemotherapy. Current breast cancer TNM staging is based on anatomical extent (size, lymph node status) but this classification gives little insight into breast cancer biology. Clinicians have long recognized the heterogeneity of human breast cancers, not only in terms of their diverse natural histories despite identical morphological features, but also in their variation in treatment response. Attempts have been made to identify good and poor prognosis groups based on pathological features such as tumor grade, lymphatic invasion and S-phase fraction (Ref. 11) which may better reflect tumor biology. In recent years there have been numerous reports on molecular prognostic and predictive markers in oncology. These tumor markers have had little impact in routine clinical practice. Often studies are based on small, heterogeneous retrospective series, and have not been reported in a rigorous enough fashion with insufficient information, particularly methodologically, to reproduce results (Ref. 12). Many follow-up studies have been inconsistent with original results: this has variously been attributed to lack of power, different patient populations and technical limitations. There is also a paucity of well designed, prospective assessments of the clinical value of these tumor markers. As a result, the value of many promising prognostic markers is still uncertain. Version / 152 May 21, 2014

166 The prognostic factors accepted by the NIH 2000 Consensus Conference on Adjuvant Therapy did not include any molecular markers relevant to breast cancer biology apart from the hormone receptors (Ref. 8). The most recent St Gallen Consensus Panel (2005) (Ref. 9) established three risk categories: minimal, intermediate and high risk. Hormone receptors, tumor size, tumor grade and age remain key discriminating factors, and HER-2 status and lymphatic &/or vascular invasion or both in the primary tumor are newly accepted prognostic factors. In the United Kingdom, the Nottingham Prognostic Index is commonly used to predict clinical outcome; it is based on tumor size, tumor grade and lymph node status, and plays a key role in clinical risk assessment for nodenegative tumors for which chemotherapy may or may not be considered (Ref. 13) Limitations of current prognostic factors Tumor size: women with small (<1cm) node-negative cancers (T1a,b,N0) To date, uncertainty exists about the long term prognosis and treatment of lymph node-negative patients with tumors 1 cm or less. The lack of long term (>15 years) follow-up data has contributed to this ambiguity as many studies do not account for the propensity of breast cancer to relapse after 10 years (Ref. 14, Ref. 15). Two population-based data bases have reported excellent prognoses for these women. The Breast Cancer Detection Demonstration Project, which included 880 patients with T1N0 tumors, reported a survival rate of 96% at 8 years of follow-up (Ref. 16). Rosen and colleagues reported a 20 year recurrence-free survival of 86% for tumors measuring 1 cm or less (Ref. 17). In contrast, the National Surgical Adjuvant Breast and Bowel Project (NSABP) presented data from women enrolled onto five NSABP trials (B-06, B-13, B-14, B-19 and B-20). There were 235 patients with ER-negative tumors and 1,024 patients with ER-positive tumors 1 cm or less (Ref. 18). For ER-negative tumors, the 8 year relapse free survival for those who had surgery alone or with chemotherapy was 81% and 90% respectively (p=0.06). For the women with ER-positive tumors, these figures were 86% for surgery alone, 93% when tamoxifen was added (p=0.01) and 95% after the addition of both tamoxifen and chemotherapy (p=0.07 when compared with tamoxifen). The authors concluded that chemotherapy and/or tamoxifen should be considered for the treatment of women with tumors of 1 cm or less and negative axillary lymph nodes. A subset of these tumors seems to have poor clinical outcome, despite their small size. In a study of 218 patients with tumors 1 cm or less, poor tumor grade and lymphatic invasion were independent predictors of recurrence. For patients with both these features, the 7-year recurrence free survival rate was 67% compared with 92% observed with one risk factor and 99% observed with neither (p=0.0001) (Ref. 19). A population-based cohort of 430 women with 1 cm or less, node-negative, lymphatic and vascular invasion-negative early breast cancers that had received no systemic adjuvant therapy reported a 10-year relapse free survival of 82%. However, histological grade 3 tumors had a > 25% 10-year risk of relapse and >10% 10-year risk of breast cancer death (Ref. 20). Hence, these studies suggest that tumor size cannot reliably be considered to identify a group of women with excellent long term clinical outcome Histological grade Histological grade is currently not included in the revised American Joint Cancer Committee (AJCC) guidelines on breast cancer TNM staging (Ref. 21). Yet, tumor grade has been recognized since the early 20th century to provide important prognostic information. The AJCC cited the poor reproducibility of tumor grading due to the variety of approaches used and the inherently subjective nature of tumor grading as the reason for its exclusion from a staging system that needs to be reproducible from institution to institution. Elston and Ellis introduced a new grading system with the aim of making the grading criteria more quantitative (Ref. 22). A modification of the Bloom and Richardson grading system, a numerical score is assigned to three morphologic features and is used in the compilation of an overall grade. This system was named the Nottingham combined histological grade. Subsequent studies have shown improved interobserver agreement and it is Version / 152 May 21, 2014

167 recommended by the College of American Pathologists. However, whilst the diagnoses of a poor or well differentiated histological grade may influence some treatment decision making, a large proportion of breast cancers are classified as grade II or intermediate grade, for which the prognostic value is not so helpful. As can be seen above, currently used prognostic factors have a poor ability to identify a very good prognostic group within those tumors diagnosed as node-negative Patients with 1 to 3 positive lymph nodes: Do all of them benefit from chemotherapy? The Oxford Overview published in 2005 suggested that the addition of chemotherapy to tamoxifen was beneficial for patients with hormone receptor positive cancer (Ref. 23). But despite the clear difference it is evident that not all the patients obtain the same benefit. For example the benefit provided by chemotherapy on top of endocrine therapy varies according to patient s age, and there may be other situations where the addition of chemotherapy may not be translated in clinical benefit. An example of this can be found in an exploratory analysis of the SWOG 8814 trial performed by Albain et al (Ref. 24). This retrospective analysis suggested that the subgroup of patients with ER positive, HER-2 negative and 1-3 positive lymph nodes did not obtain any benefit from the addition of chemotherapy to endocrine therapy. Other reports also suggest that not all patients with 1-3 positive lymph nodes may require chemotherapy (Ref. 25). This data, in addition to the ongoing research on Luminal A type tumors, warrants further investigations aimed to the identification of a subgroup of patients with 1-3 positive lymph nodes who may safely be spared chemotherapy. The St Gallen Consensus Guidelines consider 1-3 positive nodes as an intermediate risk group which, depending on other biological characteristics, may not necessarily need adjuvant chemotherapy (Ref. 26) particularly in the case of endocrine responsive disease Molecular prognostic markers in breast cancer The last ten years have witnessed several attempts to more clearly define groups of women with extremely good prognosis who can be spared adjuvant systemic therapy, or at least adjuvant chemotherapy using molecular markers. The most interesting data in this field have been published in major scientific journals and are summarized in the following table: Table 1 New candidate prognostic tools UPA PAI-1 Cyclin E Gene expression signature Authors Janicke Look Keyomarsi Van de Vijver Reference JNCI, 2001 JNCI, 2002 N Engl J Med, 2002 Type of study Prospective Metaanalysis Retrospective Node-negative (N) 374 3, Pt characteristics median age % ER low % grade 3 % treated with adj No grading done 50 N Engl. J Med, 2002 Retrospective < Version / 152 May 21, 2014

168 therapy UPA PAI-1 Cyclin E Gene expression signature Median f.-up (y) Type of assay ELISA ELISA Western Blot RNA Array Minimum amount of tissue Quality control in multicentre setting 50 mg snap frozen 50 mg snap frozen 100 mg snap frozen 3 µg Yes Yes No No Prognostic value of tool in multivariate analysis 1) In node-negative Strong, superseded only by tumor grade 2) In node-negative & positive Strong, supersedes all others Very strong, more powerful than nodal status snap frozen Very strong, more powerful than nodal status High throughput gene expression profiling and the development of a multi-gene prognostic signature for early breast cancer The advent of high throughput array-based technology and the sequencing of the human genome has provided the opportunity to begin comprehensive molecular profiling of cancers. Such efforts have, in recent times given us new insights into breast cancer biology and confirmed that the disease is considerably more heterogeneous than can be predicted by traditional histopathological methods. The collective results demonstrate that 1) the estrogen receptor (ER) status of the tumor is the most important discriminator of expression subtypes; 2) breast tumors can be grouped according to at least 3 individual molecular subgroups: basal (almost exclusively ER-negative), HER-2 and luminal (almost exclusively ER-positive) subtypes; and 3) these subgroups seem to have distinct clinical outcomes and may respond differently to various therapies (Ref. 27, Ref. 28, Ref. 29, Ref. 30, Ref. 31). A breast cancer classification system based on molecular expression patterns that defines different breast cancer subtypes, analogous to what exists for lymphoma or leukemia, rather than based on anatomic extent of the disease could emerge and potentially allow clinicians to individualize treatment according to molecular subtype. These results suggest that molecular profiling can substantially improve our understanding of breast cancer biology and therefore improve our management of breast cancer patients The Amsterdam 70-gene prognostic signature New methods of prognostic classification have been developed using array technology. Six years ago, in an attempt to better understand breast cancer biology and to find new clinically relevant molecular makers, the Netherlands Cancer Institute looked at global gene expression levels of 78 tumor samples from untreated node-negative breast cancer patients using oligonucleotide microarray technology. They used a homogenous population from node-negative patients, all under 55 years of age, all treated with loco-regional therapy only, with a median follow-up of 8 years, to derive their prognostic gene signature. In their landmark study, they were able to establish a 70- gene expression profile for the prediction of early distant metastases (recurrence in less than 5 years) (Ref. 32). They then validated the prognostic power of this unique set of genes in 295 patients and confirmed that this gene signature outperformed all the traditional prognostic Version / 152 May 21, 2014

169 guidelines, such as those from the NIH and St Gallen, clearly separating a group of patients with an excellent prognosis at 10 years from a group with a high risk of distant recurrence. These results were independent of age, tumor size, number of positive nodes and centrally reviewed histological grade (Ref. 33). Since the publication of these results, a number of authors have underlined critical issues in microarray-derived analysis including the fragility of gene signatures and over-optimistic performance estimation due to model over-fit (Ref. 34, Ref. 35, Ref. 36). Gene signatures, promising as they are, still have to make their way to the clinic and are vulnerable to the same problems as single gene tumor markers before them. Furthermore, some authors have expressed doubts that they add prognostic value to that provided by the best risk classifications based on clinical-pathological factors including age, tumor size, tumor grade, nodal involvement and presence of hormonal receptors, though this of course places great faith in the reproducibility of current clinical marker assessments, very few of which undergo critical quality control (Ref. 37). Gene signatures have enormous potential towards better individualization of treatment options in breast cancer therapy if they can reliably identify patients for whom adjuvant chemotherapy is definitely not indicated despite poor clinical risk factors, and patients who need chemotherapy despite good clinical risk factors (Ref. 38). This better selection of patients could avoid both overtreatment, with its potential for severe complications and under-treatment with deleterious consequences for patient survival. Prior to the commencement of the trial, the TRANSBIG research consortium (sister translational research network to the clinical trials network the Breast International Group) undertook an independent and external validation of the Amsterdam-70-gene expression signature (which will be referred to as the 70-gene signature) using prospectively defined criteria, to establish its reproducibility and to confirm its prognostic ability Independent validation of the 70-gene signature by the TRANSBIG research consortium Initial validation in lymph node negative patients This study was based on frozen archival tumor material of 302 node-negative patients from five non-dutch cancer centers from three different countries (United Kingdom, Sweden, and France). The patients were aged 60 years old or less, were diagnosed before and including 1998 and, as was the case with the Dutch patients, had received only loco-regional therapy. The median follow-up was 13.6 years. Frozen samples were sent to Amsterdam for gene expression profiling using a custom designed chip, which assesses the mrna expression of the 70-genes in triplicate (Mammaprint from Agendia, a spin-off commercial company of the Netherlands Cancer Institute using Agilent technologies). Researchers in Amsterdam had no knowledge of the clinical outcome data. An independent statistical office in Brussels determined the clinical risk groups. Central pathology review of estrogen receptor status and grade could be obtained for nearly 80% of samples. The first results of this validation study were presented during the 2004 San Antonio Breast Cancer Symposium (Ref. 39) and confirmed the previous findings, namely that the 70-gene signature was able to distinguish a significantly decreased time to distant metastases (TDM) and overall survival (OS), outperforming both the Nottingham Prognostic Index, the St-Gallen criteria and the Adjuvant! Online software ( which calculates a 10-year survival probability based on the patient s age, tumor size, grade and estrogen receptor status (Ref. 40). A recent evaluation of the Adjuvant! Online program found that known clinical prognostic factors were able to predict OS, breast cancer-specific survival, and event free survival quite accurately in 4,083 patients diagnosed with breast cancer in British Columbia between 1989 & 1993, with the exception of patients who were diagnosed when they were younger than 35 years (Ref. 41). The 70-gene signature remained a significant prognostic factor for TDM and OS even after adjustment for all clinical-pathological factors known to have prognostic value in this disease. Version / 152 May 21, 2014

170 Consensus within the consortium decided that the low clinical risk group consisted of patients with a 10-year survival probability of at least 88% (using Adjuvant! Online) if they were 1% positive for expression of estrogen receptors by immunohistochemistry, and of at least 92% if they were not. These two cut-offs were chosen to reflect the fact that today patients with hormone receptorpositive tumors receive adjuvant endocrine therapy (with an estimated absolute 10-year benefit of about 4% overall), whereas patients in the validation series were all untreated regardless of their hormone receptor status. When adjusted for the clinical risk groups based on 10-year survival probability using the Adjuvant! Online software, the 70-gene signature adjusted hazard ratios were 2.13 (95% CI ) for TDM, 2.66 (95% CI ) for OS, and 1.36 (95% CI ) for DFS. Similar hazard ratios were found in multivariate Cox s regression analysis. These results indicate that the gene signature adds independent prognostic information to that provided by a risk assessment based solely on clinical-pathological factors. Central pathology review of hormone receptor status and tumor grade and an independent source verification of all data by external auditors added significant strength to these findings. Furthermore, when the patients were divided according to their gene signature risk groups, for those patients classified as gene signature low risk, regardless of being classified clinical high or low risk (by Adjuvant! Online), their Kaplan-Meier estimates of 10-year OS were practically identical (88%, 89% respectively), with the same result for the gene signature high risk groups (69% for both). These results reinforced the original evidence and lent strong support to the initiation of the trial. The results of the validation study have been published (Ref. 42). Since the initial validation, the 70-gene signature has also been validated in a similar population of women aged from years Subsequent validation in lymph node-positive patients In the first retrospective validation study, by van de Vijver et al., the 70-gene profile was shown to be prognostic in lymph node-positive patients, with a hazard ratio for distant metastases of 5.5 (95% CI ; p<0.001) (Ref. 33). To further substantiate these results the TRANSBIG consortium conducted a validation study in breast cancer patients with 1-3 positive lymph nodes. This study was based on 241 archival tumor samples from breast cancer patients with 1-3 positive lymph nodes. Patients had a unilateral T1, T2 or operable T3 tumors and were treated with breast conserving therapy or mastectomy with axillary lymph node dissection. One-hundred-seventy-four out of the 241 patients were diagnosed between 1996 and 2001, treated in the Netherlands Cancer Institute, Amsterdam and were up to 71 years of age at diagnosis. The remaining 67 patients were diagnosed between 1994 and 1998, treated at the European Institute of Oncology, Milan and were under the age of 61 years at diagnosis. Twenty patients (8.3%) received no adjuvant systemic therapy, 55 patients (22.8%) were treated with chemotherapy only, 81 patients (33.6%) were treated with endocrine therapy only and 73 patients (30.3%) received both chemo- and endocrine therapy. Frozen tumor samples were evaluated by gene expression profiling for the 70-gene profile using a custom designed microarray measuring the 70 genes in triplicate (MammaPrint). Patients were classified as genomic high-risk or genomic low-risk. Among the 241 patients, 99 (41%) were classified as genomic low-risk and 142 (59%) as genomic high-risk. At 5-years, the overall survival probability was 98% (SE 0.01) for the genomic low-risk group and 88% (SE 0.03) for the genomic high-risk group, respectively. Patients were also classified by Adjuvant! Online, using clinical-pathological criteria. The low clinical risk group was defined as a 10-year breast-cancer specific survival probability of more than 88% and more than 92% for ER positive and ER negative tumors, respectively (see paragraph ) The estimated hazard ratio (HR) for the genomic risk was 5.4 (95%CI ; p<0.001) and 4.1 (95%CI ; p=0.002) for OS and distant metastases as first event, respectively, and 4.5 (95%CI ; p=0.002) and 3.4 (95%CI ; p=0.007) when adjusted for the clinical risk as defined by Adjuvant!. These results indicate that the 70-gene profile adds prognostic information to the risk assessment based on the clinical-pathological criteria. Version / 152 May 21, 2014

171 The 70-gene profile seemed to perform equally well in both the group of patients treated and those untreated with chemotherapy. In a multivariate analyses, the 70-gene profile, number of positive lymph nodes (3 versus 1) and adjuvant endocrine therapy were the only significant independent factors for prediction of OS with HRs of 5.3 (95%CI ; p=0.006), 3.6 (95%CI ; p=0.001) and 0.3 (95%CI ; p=0.01), respectively. In summary these results show that the 70-gene profile is a strong prognostic marker in breast cancer patients with 1-3 positive lymph nodes and adds prognostic information to the clinicalpathological criteria (paper in preparation). Moreover, the 70-gene profile can accurately identify a group of patients with 1-3 positive lymph nodes with an excellent survival who can be safely spared chemotherapy. These data provide sufficient evidence to enlarge the inclusion criteria of to include patients with 1-3 positive lymph nodes (Ref. 43). In summary, the TRANSBIG consortium has completed the first independent and external validation of a gene signature derived from microarray technology. Their analyses confirm that the 70-gene signature adds significant independent prognostic information to that produced by current clinical-pathological factors and provides sufficient evidence to develop the study. The 70-gene signature remains the only prognostic tool derived from microarray technology to have undergone stringent external and independent evaluation The trial: The first trial to evaluate the clinical value of a genomic tool The success of these validation studies allowed for the commencement of the large collaborative trial (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy), which will be the first prospective trial to validate a molecular-based signature for the adjuvant treatment of early breast cancer patients with 0 to 3 positive nodes. The goal of is to prospectively validate the 70-gene signature providing the level-1 evidence of its efficacy that is needed before its wide clinical application. This study aims to give us a definitive answer to the clinical relevance of the 70-gene signature, its performance compared to traditional prognostic factors and its ability to predict response to commonly prescribed adjuvant treatments. will enroll 6,600 patients to investigate the benefit/risk ratio of chemotherapy when the assessment of prognosis based on clinical-pathological features differs from that provided by the gene signature. It will use the 70-gene signature to classify early stage breast cancer patients into high and low risk of distant relapse and to compare this assessment to the Adjuvant! Online software risk assessment, which is widely used in clinical practice today and which takes into consideration traditional clinical-pathological factors. All 6,600 patients enrolled in the study will have their risk of recurrence assessed by both clinical-pathological and 70-gene signature methods. Those patients for whom both methods classify the patient as high risk will be proposed adjuvant chemotherapy; those patients for whom both methods classify the patient as low risk will not be offered adjuvant chemotherapy, however they will be offered endocrine therapy if their tumors are endocrine responsive. Those patients for whom the two risk assessments are discordant will be randomized for treatment decision making tool to either clinical criteria (using Adjuvant! Online) or genomic prognosis (using the 70-gene signature). Using this new tool, aims to better define patient prognosis and therefore to better select patients who need adjuvant chemotherapy; by doing so it is expected that 10% to 20% of women who would normally receive adjuvant chemotherapy based on their clinical-pathological factors will be spared the inconvenience and morbidity of this therapy, without having any negative impact on their survival. The primary objective of the trial is to confirm that the number of patients that can be spared adjuvant chemotherapy is significantly increased when the decision is based on the gene prognosis signature rather than on the clinical-pathological criteria, while the critical group of patients who have a high risk of recurrence according to the clinical-pathological criteria but a low risk according to the gene prognosis signature is not under-treated. If this is confirmed, it will have Version / 152 May 21, 2014

172 immense benefits for women and will markedly improve our ability to predict which patients are likely to experience a distant relapse and who therefore require additional systemic therapy. The economics of using multi-gene assays that are considerably more expensive than currently used prognostic assessments has been appraised for another multi-gene assay derived from high throughput technology (Ref. 44). The Oncotype assay was found to predict more accurately than current guidelines recurrence risk in lymph node-negative, estrogen receptor-positive early-stage breast cancer treated with tamoxifen. Using an economic model that considered survival, quality of life and relevant costs on data from a large multi-centre clinical trial, among a hypothetical cohort of 100 patients, the multi-gene assay was found on average to increase quality-adjusted survival by 8.6 years and reduce overall costs by US$ 202,828, with cost effectiveness most influenced by the propensity to administer less chemotherapy (Ref. 45). Further randomizations, as applicable, will be offered to enrolled patients to answer important questions about adjuvant therapy. For those patients receiving chemotherapy, an optional randomization will compare anthracycline-based regimens (anthracyclines followed by docetaxel for node positive) to a new docetaxel-capecitabine combination (see section 1.3). A third optional randomization for all patients with endocrine responsive disease will compare 7 years of letrozole to 2 years of tamoxifen followed by 5 years of letrozole (see section 1.4). A prospective randomized trial like is essential to determine, in an unbiased way, the true value of gene signatures in clinical practice today. Given the immense therapeutic and emotional repercussions for women, it is essential to know the true meaning of the poor prognosis signature and whether any of our currently prescribed treatments can alter the natural history of this high risk group. Retrospective studies are susceptible to all sorts of bias which may significantly influence results and hence, will be unable to accurately determine these implications. Furthermore, the evaluation of the potential of gene signatures to individualize management, to spare treatment without compromising long term outcome and hence diminish the public health burden is of utmost priority to women and to nations. 1.3 Chemotherapy in the trial: can anthracyclinebased chemotherapy be safely replaced by the combination of docetaxel-capecitabine as adjuvant treatment of early breast cancer? Adjuvant Chemotherapy The Early Breast Cancer Trialists Collaborative Group (EBCTCG) meta-analysis (Ref. 46) showed that combination chemotherapy yields a significant reduction in mortality when compared to no chemotherapy. This benefit occurs in all patients, irrespective of nodal or estrogen-receptor status and the use or not of anti-estrogen therapy (tamoxifen). The benefits of chemotherapy are seen in all age groups analyzed though the absolute benefit is greater for younger women (less than 50 versus 50 to 69 years old) and in node-positive disease as compared to node-negative disease. For women with early-stage breast cancer, adjuvant polychemotherapy reduced the annual risk of death by 30% in women aged 40 to 50 (10% difference in risk of death within 15 years), and by 12% in women aged 50 to 70 (3% difference in risk of death within 15 years). These risk reductions were very similar in node-negative and node-positive patients, but had a larger absolute effect in node-positive patients, because of their higher mortality rate. In node-positive patients, a 37% reduction in the odds of recurrence and a 30% reduction in the odds of death were reported, while for node-negative patients, these numbers were almost the same, being 36% and 29% respectively. Version / 152 May 21, 2014

173 The absolute benefit for polychemotherapy versus no adjuvant chemotherapy in women less than 50 years old is twice as great at 15 years (10%) as at 5 years (4.7%) Anthracyclines The overview also highlights an advantage in outcome with the utilization of anthracycline based regimens in the adjuvant setting, when compared to CMF-based regimens, of 16% reduction in the annual risk of death, and 11% reduction in the annual risk of recurrence. These effects seem to be irrespective of age and nodal status. Nevertheless, not all anthracycline regimens can be considered adequate. The most convincing results are derived from treatments that included a three-drug combination, as opposed to a two drug (such as doxorubicin plus cyclophosphamide (AC)). Regimens such as FEC100, CAF/FAC and CEF, for 4 to 6 cycles, are considered the treatment of choice for high risk node-negative patients, based on the results of several randomized trials (Ref. 8, Ref. 47, Ref. 48, Ref. 49) and the EBCTCG Overview. Of concern to women with a medium rather than a high risk of relapse are two possible long-term toxicities of anthracycline-based therapy: secondary leukemia and cardiotoxicity. The cumulative risk, at 5 years, of developing anthracycline-related leukemia is around 1% for both doxorubicin and epirubicin and is directly related to the administered dose (Ref. 50, Ref. 51). Regarding epirubicin, good quality data is available from the Pharmacia Clinical Trials Database, derived from 7,110 patients treated with epirubicin-containing regimens. The cumulative probability of AML/MDS was 0.27% (95% CI, 0.14% to 0.40%) at 3 years, 0.46% (95% CI, 0.28% to 0.65%) at 5 years, and 0.55% (95% CI, 0.33% to 0.78%) at 8 years (Ref. 52). In a recent study, the National Cancer Institute of Canada Clinical Trials Group (NCIC-CTG) analyzed the risk of acute leukemia in 1,463 premenopausal node-positive patients enrolled onto four NCIC-CTG adjuvant therapy trials. The conditional probability at 5 years of developing acute leukemia was 1.5% for all epirubicin-containing regimens, as opposed to 0.3% for non-epirubicin-containing ones (Ref. 47). For doxorubicin, the NSABP group has reviewed the data from all its adjuvant trials that explored the regimens containing doxorubicin given at the fixed dose per cycle of 60 mg/m² and a fixed cumulative dose of 240 mg/m² and cyclophosphamide administered at variable doses per cycle and variable cumulative doses in 8,533 patients. Thirty-nine cases of AML/MDS were reported. The incidence was found to be particularly correlated with cyclophosphamide doseintensity, with a corresponding cumulative incidence of AML/MDS at 5 years of 1.04% (95% CI, 0.64% to 1.67%) for patients receiving more intense regimens (often with prophylactic growth colony-stimulating factor support), compared with 0.22% (95% CI, 0.12% to 0.43%) for those treated with standard AC. Breast radiotherapy also seemed to increase the leukemia risk (relative risk, 2.45; P = 0.007) (Ref. 54). Late cardiac toxicity associated with anthracycline-based chemotherapy is of low frequency (< 1%), but is a potentially serious and even life-threatening event. It varies from an asymptomatic decrease in left ventricular ejection fraction to clinically relevant congestive heart failure. It is directly correlated with the cumulative dose administered, increasing exponentially above 500 mg/m² for doxorubicin and 900 mg/m² for epirubicin. Other associated risk factors include underlying cardiovascular disease, hypertension, advanced age, prior or concomitant radiotherapy to the mediastinal or pericardial areas, and previous therapy with anthracyclines or anthracenediones. A recent publication reported on 355 breast cancer patients who were previously treated in three prospective trials of adjuvant chemotherapy and were still free of disease at 11 years of follow-up. Two hundred of these women had received doxorubicin-containing regimens, whereas 155 were treated with CMF. Both groups were balanced in terms of median age at randomization in the initial trial, median age at recall and percentage of risk factors for cardiovascular disease. The percentage of systolic dysfunction was higher in the doxorubicin group (8% versus 2%), and the cumulative cardiac mortality was 0.6% vs. 0% (Ref. 55). The French Adjuvant Study Group recently reported a Version / 152 May 21, 2014

174 very similar rate of systolic dysfunction in breast cancer survivors treated with the FEC (5-FU, epirubicin, cyclophosphamide) regimen (Ref. 56) Taxanes In recent years, the taxanes have been investigated in several trials of adjuvant chemotherapy. The Cancer and Leukemia Group B (CALGB) 9344 trial (Ref. 57) reported a benefit for the addition of four cycles of paclitaxel to a standard regimen of four cycles of doxorubicincyclophosphamide (AC) chemotherapy when compared to AC alone, in node-positive patients. A recent update of that trial, with a median follow-up of 69 months, demonstrated significantly better DFS and OS rates for paclitaxel-treated women, but the advantage seemed confined to the subset of patients with estrogen receptor-negative tumors. The NSABP B28 trial, comparing four postoperative cycles of doxorubicin and cyclophosphamide versus the same combination followed by four cycles of paclitaxel showed a significant reduction in recurrence with paclitaxel, but no difference in OS (Ref. 58). In both studies, the control arm consisted of 4 cycles of AC, considered to be sub-optimal treatment by today s standards. A third trial, showed the triple-drug combination of docetaxel, doxorubicin, and cyclophosphamide (TAC) to be superior, in terms of DFS and OS, to standard 5-fluorouracil, doxorubicin, and cyclophosphamide (FAC) chemotherapy in a randomized comparison in node-positive patients. The benefit appeared to be confined to patients with 1 to 3 positive lymph nodes (Ref. 59, Ref. 60). A French trial, PACS 01, also showed statistically significant improvement in DFS and OS in patients with 1 to 3 positive nodes and women older than 50 years with the sequence of 3 courses of FEC followed by 3 courses of docetaxel in comparison to 6 cycles of FEC (Ref. 61) Hence, the benefit appears to be restricted to post menopausal women. Regimens comprising anthracyclines and taxanes, either concomitantly (e.g. TAC) or in sequence (e.g. EC/FEC followed by taxane) are generally reserved for node-positive patients with other features of poor prognosis in view of the significantly higher toxicity of these regimens. Among the multitude of trials attempting to delineate the role of the taxanes in the adjuvant treatment of breast cancer and enrolling more than 30,000 women, only one an American study (N=1,016) has addressed this question in node-negative women. This trial compared TC (docetaxelcyclophosphamide) to AC (doxorubicin-cyclophosphamide): with a median follow-up of 43 months there was no significant difference in either DFS or OS between the two regimens. While the follow-up is still insufficient, preliminary reports have shown TC to be at least as effective as AC (Ref. 62) The Combination Docetaxel-Capecitabine Capecitabine is an oral fluoropyrimidine, a prodrug that generates 5-Fluorouracil (5-FU), preferentially in tumor tissue, thus mimicking continuous-infusion administration. This selectivity is explained by the significantly higher concentration and activity of the enzyme responsible for the conversion, thymidine phosphorylase, in tumor tissue compared with healthy tissues. Preclinical studies in human cancer xenograft models demonstrated that administration of taxanes results in further up-regulation of thymidine phosphorylase in tumor tissue, as shown in studies with preoperative docetaxel. These same studies showed that its administration with capecitabine resulted in synergistic anti-tumor activity (Ref. 63, Ref. 64). Subsequent clinical studies have shown that single-agent capecitabine is an active and tolerable treatment in metastatic breast cancer that has progressed during or after anthracycline and taxane therapy, achieving response rates of approximately 25% and a median survival in excess of 1 year. Activity in heavily pre-treated patients paved the way to investigate capecitabine activity earlier in the disease course and in combination with other cytotoxic agents. In addition, it proved to be an Version / 152 May 21, 2014

175 attractive agent for incorporation into combination regimens because of the low incidence of myelosuppression. A randomized phase III trial in the metastatic setting (Ref. 65), based on the results of clinical and preclinical studies, compared single-agent docetaxel to a docetaxel-capecitabine combination, and could demonstrate the latter to be more effective. The use of the combination resulted in superior response rate, improved DFS and notably a 3-month improvement in median survival. These results not only established this regimen as an important treatment option for patients with anthracyclinepretreated metastatic breast cancer but made it an attractive combination for transfer to the adjuvant setting. The side effect profile of capecitabine-docetaxel therapy was manageable and consistent with the known toxicities of the individual agents. There was a higher incidence of gastrointestinal side effects and hand-foot syndrome in patients receiving the combination therapy; indeed, the aggregate incidence of grade III adverse events was higher in the combination arm, predominantly because of grade III hand-foot syndrome and myalgia. On the other hand, arthralgia and neutropenic fever were more common with single-agent docetaxel (used at the higher dose of 100 mg/m2 in single agent therapy) and the incidence of grade IV adverse events was higher in the single-agent docetaxel arm, primarily because of neutropenic fever. All the side effects associated with the combination were manageable with appropriate medical intervention (e.g., loperamide and rehydration for diarrhea, mouthwash and fluconazole for stomatitis, and oral vitamin B6 preparations and emollients for hand-foot syndrome) and treatment interruption and/or dose reductions when needed. Dose reductions were required in 65% of patients and capecitabine interruption was required in 34% of cycles. This combination is currently being evaluated in a randomized phase III trial in the adjuvant setting for the treatment of high-risk breast cancer (both node-positive and node-negative patients), run by the US Oncology Group. It compares four cycles of the AC regimen followed by either four cycles of docetaxel alone (at 100 mg/m²) or four cycles of docetaxel (75mg/m²) plus capecitabine (825 mg/m² twice daily - lower than in the pivotal phase III trial). Since in the trial this combination will be given in chemotherapy-naïve patients, i.e., having not previously received AC, the toxicity is expected to be reduced further Chemotherapy in the trial The trial will ask an important question regarding chemotherapy optimization for early breast cancer: can a docetaxel-based regimen safely replace an anthracycline-based one? Docetaxel was the only agent shown to generate higher response rates than doxorubicin in a head to head comparison of single-agent use for the treatment of advanced breast cancer (Ref. 50) and among the many docetaxel-based combinations explored so far in breast cancer, one of the most interesting ones is docetaxel plus capecitabine; aside from the synergistic anti-tumor activity obtained in preclinical models, it showed improved response rate (RR), time to progression (TTP) and OS when compared to docetaxel alone in the previously detailed metastatic breast cancer randomized trial. The combination of docetaxel-capecitabine was also compared to doxorubicin-cyclophosphamide in the neoadjuvant setting; 78 patients with stages II and III breast cancer were randomized between the two treatments and those in the docetaxel-capecitabine combination presented higher clinical response (91% vs 74%) and, importantly, higher pathological complete response (23% vs 6%) (Ref. 66) however we are waiting for later time points to be reported to confirm these results. Although short-term side effects of this combination were significant, there are good reasons to believe that tolerance might be better in early breast cancer, when given upfront, and that, importantly, long-term side effects might be less than what is commonly observed with anthracyclines. Version / 152 May 21, 2014

176 Given the high rate of capecitabine dose reductions in the metastatic breast cancer trial, capecitabine will be given at the dose of 1650 mg/m² p.o. d1-14 q21 days (instead of 2500 mg/m²) in combination with docetaxel 75 mg/m² i.v. q21 days (dose unchanged). Since the combination of docetaxel-capecitabine has proved to be both safe and well-tolerated in women with metastatic breast cancer it is an exciting regimen to study in the adjuvant setting. In addition to their activity as single agents, promising preliminary results of their combined activity and the extent of experience with these two agents encourages their further investigation. This combination, with its different toxicity profile from anthracycline based regimes, will have a much lower incidence of serious long-term side-effects such as cardiotoxicity and leukemia, making it an attractive replacement for anthracycline-based chemotherapy. hopes to validate this new non-anthracycline-based chemotherapy regimen for use in high-risk women with early breast cancer and to generate long-term safety data on this combination. 1.4 Endocrine therapy in the trial: is upfront Aromatase Inhibitor (AI) letrozole for 7 years safer and more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? For many years the current standard treatment for endocrine responsive breast cancer is 5 years of the antiestrogen tamoxifen; very recently national and international opinion guidelines have advocated the incorporation of an AI in sequence with or as a replacement to tamoxifen Adjuvant endocrine treatment in breast cancer Recently, the available data on the use of tamoxifen in the adjuvant setting of ER-positive breast cancer has been reviewed extensively by the Early Breast Cancer Trialists Collaborative Group (EBCTCG) (Ref. 10). In this meta-analysis, tamoxifen has been shown to reduce the annual odds of breast cancer death by a third and the annual recurrence rate by half in ER-positive patients regardless of progesterone receptor status, age and the use of chemotherapy. In addition, it reduced the odds of developing a contralateral breast cancer by more than 30%. For this reason, endocrine therapy is clearly the most important systemic adjuvant treatment for women with hormone receptor positive breast cancer, and is considered the standard of care. At present, the most important prognostic factor in breast cancer relates to axillary lymph node involvement. Due to the increasing use of screening mammography in women of 50 years of age in developed countries, the prevalence of node-negative breast cancer or with few positive nodes has been continually increasing. Adjuvant therapy for these women is therefore of prime importance for the treating physician today. Only one large trial, the NSABP Study B-14, has studied tamoxifen in the node-negative setting. In this study, 2,644 patients with ER-positive primary tumors were randomly allocated to receive either tamoxifen for 5 years or placebo. At the end of four years of follow-up the difference in DFS between the tamoxifen and placebo groups was highly statistically significant (p< ) (Ref. 67), confirming the benefit of tamoxifen therapy in this group. Tamoxifen is the trans-isomer of a substituted triphenylethylene with anti-estrogenic activity due to a competitive inhibition of estrogen action. It blocks the binding of (3H)estradiol to the estrogen receptor. Its clinical efficacy and long term side effects have been clearly demonstrated by trials conducted in the 1970 s and the Oxford Overview. A minimum of five years of treatment with tamoxifen is considered standard. Tamoxifen is well tolerated, and adverse effects are mild when compared with cytotoxic chemotherapy. Short term toxicities of hot flushes and vaginal discharge are related to the anti-estrogenic effect of the drug. Tamoxifen has been shown to significantly increase the incidence of endometrial cancer (Ref. 68), and at high doses retinopathy has been observed (Ref. 69). Tamoxifen s estrogenic properties also reduce the mean levels of cholesterol, which is thought to be the cause of the reduction of cardiovascular deaths observed in some studies Version / 152 May 21, 2014

177 and a bone protective effect. An association between the use of tamoxifen and increased thromboembolic disease is also firmly established (Ref. 10, Ref. 68) Use of Aromatase Inhibitors in the adjuvant setting There have been three main strategies in the effort to improve cure rates from breast cancer using third generation AIs in the adjuvant setting: first as initial treatment in direct comparison with tamoxifen, second in sequential use before or after tamoxifen for a total period of five years, and third as extended therapy after five years of tamoxifen. At least 10 adjuvant trials involving over 40,000 women with early stage breast cancer are currently in progress. The results of those trials that have been reported are shown in Table 2. This section will summarize these results and the questions that still remain. Version / 152 May 21, 2014

178 Table 2: Comparison of current published studies involving adjuvant AIs Trial name: ATAC (anastrozole) Comparison/duration TAM 5yrs vs No. of women randomized AI 5yrs BIG 1-98 (letrozole) TAM 5yrs vs AI 5yrs (core analysis) Version / 152 May 21, 2014 IES (exemestane) TAM 5 yrs vs TAM 2yr-AI 3yrs ABCSG/ARNO (anastrozole) TAM 5yrs vs TAM 2yrs-AI 3yrs ITA (anastrozole) TAM 5yrs vs TAM 2yrs-AI 3yrs MA-17 (letrozole) 6,241 8,028 4,742 3, ,187 Median age Node-positive % HR+ (%) Adj chemotherapy given (%) Follow-up (months) Endocrine-refractory women included Hazard Ratio for disease free survival1 Hazard Ratio for disease free survival² Hazard Ratio for overall survival Yes Yes No No No No 0.87 P= (HR+ only) 0.97 P= P= P= ** P< P=NS 0.83 P= P=0.16 TAM: tamoxifen 20 mg daily, AI: aromatase inhibitor; HR+: hormone receptor positive; Adj: adjuvant, Not reported 0.61 TAM 5yrs vs TAM 5yrs-AI 5yrs P= P=0.04 (node-positive)

179 1Disease free survival is defined differently for each trial. Please refer to references for further detail. 2Disease free survival defined as locoregional, distant metastases, and contralateral breast cancer recurrences only. Version / 152 May 21, 2014

180 1.4.3 Tolerability profiles The side effect profile of tamoxifen is well known and the third generation AIs are generally well tolerated. The available data indicates at least equal short term safety including symptoms of menopause and quality of life. However, long term data will not be available for the next 10 years. The main side effects are described below: Hot flushes and general tolerability In the ATAC and IES adjuvant AI studies in which tamoxifen is the standard arm, the short term tolerability of the AI was at least as good as that of tamoxifen. There was significantly less vaginal bleeding in patients receiving anastrozole than tamoxifen in the ATAC study. In the IES study, the frequency of hot flushes was similar in both arms. In the MA-17 study, where letrozole was compared with placebo, there were more hot flushes in the letrozole arm. Otherwise, short term tolerability was excellent. The percentage of women discontinuing treatment because of adverse events was not significantly different between the two arms, in all studies involving AIs Skeletal effects Tamoxifen has been shown to preserve bone mineral density in post menopausal breast cancer patients. AI may reduce bone density as has been suggested by the ATAC study. Women taking anastrozole were more likely to suffer from musculoskeletal disorders (p<0.001), in particular fractures (p<0.001). Letrozole has also been shown to significantly increase bone resorption. In the MA-17 study, more women in the letrozole group compared with placebo suffered from new-onset osteoporosis. There are clinical trials currently assessing bisphosphonates in the prevention and treatment of AI induced bone loss (ZO-FAST, SABRE). In addition, mylagia and arthralgia were reported more frequently for women receiving letrozole (Ref. 70) Thromboembolic events and endometrial cancers In all trials comparing the third generation AIs with tamoxifen, the adverse events caused by tamoxifen s estrogenic properties were significantly less common in the AI groups. This particularly included ischaemic cerebrovascular disease, venous thromboembolic (including deep venous thrombosis) events and endometrial cancer. Compared with placebo, letrozole did not lead to increased rates of any of these serious adverse events in MA-17 and vaginal bleeding occurred more often in the placebo arm (p=0.01) Lipid metabolism Whilst retrospective analyses of the three randomized tamoxifen trials suggested a reduction in coronary heart disease in favor of tamoxifen (Ref. 71), no such benefit was reported in the NSABP- P1 chemoprevention study (Ref. 68). In view of the recent Women s Health Initiative Study (Ref. 72), it is unclear whether the favorable influence of tamoxifen on lipid metabolism translates into a true reduction in coronary heart disease, and whether lipid metabolism is the true underlying reason for this observation, if indeed it is correct. In the ATAC trial, more patients receiving anastrozole were reported to have an elevated cholesterol compared with patients receiving tamoxifen (7% vs 3% respectively) (product information Arimidex, Astra-Zeneca & Whereas in the adjuvant MA.17 study which is the only trial to compare letrozole to placebo, there was no increase in cardiovascular events between the two arms. Results from the lipid substudy suggests that letrozole does not significantly alter serum cholesterol, high density lipoprotein (HDL) cholesterol, low density lipoprotein (LDL) cholesterol, triglycerides or Lp(a) in non-hyperlidiemic postmenopausal women with primary breast cancer treated up to 36 months following at least 5 years of adjuvant tamoxifen therapy (Ref. 74). Exemestane may have effects converse to the other AI on lipid Version / 152 May 21, 2014

181 metabolism due to it being an androgenic steroid, however the IES study did not specifically address this endpoint. In two small studies looking at the effects of letrozole on lipid metabolism in healthy women, conflicting results were seen. In one study, there was no effect on the serum lipid profile, whilst the other reported significantly increased total and LDL cholesterol levels, as well as an increase in the atherogenic risk ratios of total/hdl and LDL/HDL cholesterol (Ref. 75, Ref. 76). In summary, the evidence suggests that AI use has benefits in terms of overall tolerability with fewer (but different) toxicities compared with tamoxifen. Women who receive AI have fewer gynecological symptoms, no increase in the incidence of endometrial cancer, fewer thromboembolic events and less hot flushes offset by the chance of more fractures and more arthralgias. The possible toxicities from long term AI use, particularly the possibility of a higher risk of cardiac disease, need to be studied in further detail Improving adjuvant therapy in the first 5 years Two trials have studied the benefit of upfront AI instead of tamoxifen in the first 5 years of adjuvant therapy (also see table 2). ATAC The results of the largest and first published study, Arimidex and Tamoxifen Alone or in Combination (ATAC) (Ref. 77) with 9,366 postmenopausal patients randomized to tamoxifen or anastrozole or to a combination of tamoxifen and anastrozole for a period of five years showed statistically significant reduction in the rate of relapses (local, distant or contralateral disease) with anastrozole as compared to tamoxifen (87% vs 89%, respectively) when it was first reported at median follow-up of 33 months. At this time, this absolute 2% reduction translated into a relative risk reduction of 17% (p=0.013). The results of the first efficacy update, based on a longer median follow-up period of 47 months and a second recent update after 68 months were confirmatory, with the absolute difference in risk reduction now being 3.7% (Ref. 78). As expected the effect was seen only in patients whose tumors were known to be hormone receptor positive but interestingly, the risk reduction was greater for the ER+, PgR- tumors than for the ER+, PgR+ ones (HR: 0.48 vs. 0.82) (Ref. 79). While this retrospective subset analysis needs prospective confirmation, the result might be explained by the fact that ER+, PgR- tumors co-segregate with HER-2 positivity. Interestingly, the combination of tamoxifen and anastrozole was not superior to tamoxifen alone proving that a combination of two efficient drugs is not always better than their single agent use. There was also a lower incidence of contralateral breast cancer with the use of anastrozole (HR: 0.62, p=0.062). To date there is no difference in the rate of breast cancer related deaths or OS. BIG 1-98 In this study, 8,028 patients were randomized to; tamoxifen for 5 years, letrozole for 5 years, tamoxifen for 2 years followed by letrozole for 3 yrs, or the 2 drugs in the reverse sequence. The first results have recently been reported (Ref. 73). The primary core analysis compared only the first two arms and the sequential arms censored at 2 years (prior to the switch). The analysis was based on 8,010 patients with a median follow-up of 26 months. Letrozole was found to significantly prolong DFS compared with tamoxifen for post menopausal women (0.81, p=0.003) and time to recurrence (HR:0.72, p=0.0002) especially in distant sites. This supports the results of the ATAC study. Again, there was no difference in OS seen at this stage. Both of these trials demonstrate that 5 years of upfront AI therapy is a reasonable alternative to the previous standard of 5 years of adjuvant tamoxifen therapy for postmenopausal women with endocrine responsive breast cancer. Indeed, for women with a contraindication to treatment with tamoxifen, an AI is the treatment of choice, given the current evidence from both an efficacy and short-to moderate term toxicity stand-point. Version / 152 May 21, 2014

182 The sequential approach Intergroup Exemestane Study (IES) The Intergroup Exemestane Study (IES, BIG 2-97), investigated the concept of sequential therapy with AI after tamoxifen. After two to three years of tamoxifen, women were randomized to switch to exemestane or to continue with tamoxifen for the remainder of the five-year treatment programme. After a median follow-up of 37.6 months results of the second interim analysis indicated a significantly higher DFS (HR 0.73, p<0.001), corresponding to an absolute benefit in terms of DFS of 4.7% at three years after randomization and breast cancer free survival benefit (HR:0.7; , p=0.0005) in favor of sequential therapy (Ref. 80, Ref. 81). Noteworthy, distant metastases-free survival (DMFS) and time to contralateral breast cancer were also significantly better with sequential therapy (HR of 0.63 and 0.44, respectively with p values of <0.001 and <0.004). So far no difference in OS has been reported. The ABCSG Trial 8 and ARNO 95 trial The efficacy of anastrozole after two to three years of tamoxifen was assessed in two other phase III trials (ABCSG Trial 8, ARNO 95) which were similar in design and prospectively planned for combined analysis by the Austrian and German Breast Cancer Clinical Trial Groups. The recently presented results also confirm the benefit of sequential AI therapy after tamoxifen (Ref. 82). In these trials 3,123 women were randomized to receive anastrozole after two years of tamoxifen or continue with tamoxifen for a total duration of 5 years. At a median 26 months of follow-up and after 143 events (local, metastatic or contralateral) the HR for risk reduction with anastrozole versus tamoxifen is 0.59; (p<0.0018). Italian Tamoxifen and Anastrozole (ITA) trial A substantially smaller study of 448 node-positive patients, with a similar design to the above trials has also been reported after a median follow-up of 36 months (Ref. 83). There were 45 events reported in the tamoxifen arm, and 17 events in the sequential arm (p<0.0002). DFS and local recurrence free survival were also significantly longer in the anastrozole arm (HR: 0.35, 95% CI: , p<0.001 and HR 0.15; 95% CI: ). Overall survival is not different between the arms (HR: 0.52, 95% CI , p=ns) and updated efficacy data after a median follow-up of 52 months are confirmatory (Ref. 84). Hence, these four trials demonstrate that the sequential strategy of tamoxifen followed by a switch to an AI after 2 to 3 years for 5 years total duration is superior for reducing the risk of breast cancer recurrence to the previous standard of 5 years of tamoxifen. However, it is unclear if for women who do not have a contraindication to tamoxifen, if treatment with an upfront AI is superior, equivalent, or inferior to a planned cross over from tamoxifen to an AI at some fixed point in time. Furthermore, these trials arbitrarily selected five years as the total duration of endocrine therapy, and it is not known if longer treatment involving a strategy of an early switch would be of sustained clinical benefit Extending adjuvant therapy beyond 5 years Approximately half of all breast cancer relapses in women receiving 5 years of adjuvant tamoxifen occur between 5 and 15 years after surgery and that risk of recurrence appears to continue indefinitely (Ref. 10). The average hazard of recurrence between years 5 and 12 after primary breast surgery for 2,257 ER-positive women was around 5% per year (Ref. 85). Based on this rationale, the NCIC-CTG MA.17 / BIG 1-97 study evaluated the strategy of extending adjuvant endocrine therapy beyond 5 years of tamoxifen with an AI (Ref. 70). After completing 5 years of tamoxifen, 5,187 women were randomly assigned to receive letrozole 2.5 mg or placebo. The results of the first interim analysis, conducted after a median follow-up of 2.4 years, indicated a significantly higher DFS in the letrozole group than in the placebo group (HR for a recurrence or a new contralateral breast cancer 0.57, p<0.001). This translated into an absolute difference of 2.2% in the rate of breast Version / 152 May 21, 2014

183 cancer events, including distant metastases, ipsilateral recurrences and contralateral breast cancers, with an actuarial projection of an absolute difference of 6% in the rate of events over a follow-up of four years. The effect of letrozole was equally distributed between women with node-negative and node-positive disease. The robust difference in the rate of events led to the early reporting of the study results at a time where less than 1% of the participating women had completed their randomly assigned therapy. So in reality, the benefit was seen when no women had completed the assigned 10 years of therapy (median of 7.4 years in total). The benefit in survival recently reported has been seen only in the node-positive subset (p=0.04) (Ref. 86). This trial established that a strategy using AI in the extended adjuvant setting (longer than 5 years) could improve recurrence rates from breast cancer above that obtained from 5 years of tamoxifen treatment alone. At present, it seems that a minimum of 2.4 years of extended therapy can be recommended based on the median follow-up from MA.17. Since all patients on placebo were offered the option of beginning letrozole, the optimal extended duration will not be provided by the long term data from this trial. However, the survival advantage in the node-positive subset is noteworthy and it is clear that extended therapy (more than 5 years of adjuvant treatment) has advantages for certain women. This trial led to the early closure of the similarly designed National Surgical Adjuvant Breast and Bowel Project Study (B33) comparing exemestane and placebo after 5 years of tamoxifen. Seemingly confirming the benefit of extended duration adjuvant endocrine therapy are the results of the ABSCG Trial 6a. In this trial, 856 postmenopausal women who had previously received 5 years of tamoxifen on a previous phase III trial were randomized to receive anastrozole for 2-3 years or placebo (Ref. 87). Reported after a median follow-up of 60 months, the risk of recurrence was reduced (HR: 0.64; 95% CI , p=0.0477) with no OS advantage seen as yet Which strategy is best? At present, it is difficult to know how to compare results across these trials to determine which particular agent or strategy is superior for an individual breast cancer patient. Based on the published results of these studies, the American Society of Clinical Oncology and the St Gallen consensus panel in 2005 concluded that the optimal adjuvant endocrine treatment for post menopausal women with HR-positive breast cancer today should include an AI as initial therapy or sequential therapy consisting of tamoxifen followed by an AI (Ref. 88). As a result, for postmenopausal women diagnosed today with hormone responsive early breast cancer there are many possibilities for her adjuvant endocrine therapy. The optimal option is uncertain: if treatment upfront with an AI is superior, inferior or equivalent to a switch from tamoxifen to an AI, or if tamoxifen after an AI switch could be advantageous or detrimental. The optimal duration is also unclear: 5 years or extended treatment however there is increasing evidence supporting extended adjuvant endocrine therapy. It is also unknown if the use of an AI with ovarian function suppression can be extrapolated into the premenopausal setting. It is unknown which of these treatments or combinations is most effective and for which individuals/subgroups of patients. It is also possible that some individuals will respond better to one treatment compared to another treatment or due to the characteristics of their disease one strategy may be more appropriate than another. The BIG 1-98 four arm study, where women are randomized to receive either 5 years of tamoxifen, 5 years of letrozole, 2 years of tamoxifen followed by 3 years of letrozole or 2 years of letrozole followed by 3 years of tamoxifen, will help us answer some of these questions in a few years time. It will not help us in delineating the benefits of extended therapy however, which clearly has advantages for certain women. At present, we can conclude that the group of women who recur early (in the first 2-3years) gain an advantage from upfront AI however, unfortunately we have not been able to identify the characteristics of this group of patients yet. In trials of switching strategies we see a greater treatment effect for sequential AI, than when AI is compared as an initial therapy, this maybe due to the initial period of tamoxifen treatment selecting Version / 152 May 21, 2014

184 a population that respond better to endocrine therapy than the starting population do. Furthermore, the full long term health costs (particularly skeletal and cardiovascular problems) of AI use will not be known for another 10 years when toxicity reporting is complete for these trials. From these data, it is clear that AI have a role in the adjuvant setting. However, the optimal sequence and duration of the AI and tamoxifen combination and its performance in comparison to a strategy of upfront AI is unknown. Also, it is clear that extended therapy with endocrine treatments has definite merit. Whilst part of these questions may be addressed by the mature results of the BIG 1-98 trial, it has also become evident that the one shoe fits all approach is unsuitable for all HRpositive breast cancers. This trial will therefore evaluate the following comparison: 2 years of tamoxifen followed by 5 years of letrozole versus 7 years of upfront letrozole. Seven years has been chosen to replicate the MA.17 data where the median duration of treatment was 2.4 years after 5 years of tamoxifen. Whilst this duration of adjuvant endocrine therapy is not currently standard, it is clear that extended therapy is beneficial and should be explored in a clinical trial setting. will have the huge advantage over all other clinical trials involving AIs: it will secure the collection of frozen and paraffin-embedded tumor tissue as well as serum for all participating women. In this way, it has great potential for the discovery or validation of molecular signatures indicating which women are candidates for an AI upfront and which women may be best served by a sequence of tamoxifen followed by AI. 2 Objectives of the trial 2.1 Primary objectives The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to breast cancer patients with 0 to 3 positive nodes. The trial will also aim to compare a docetaxel-capecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens (anthracyclines followed by docetaxel for node positive). The last primary objective will be to determine the best endocrine treatment strategy i.e. single agent up-front, AI (letrozole) for seven years compared to the sequential endocrine strategy of 2 years of tamoxifen followed by 5 years of letrozole. Version / 152 May 21, 2014

185 2.2 Secondary objectives The trial has several secondary objectives: Comparison of the two prognostic methods: To estimate both relative (Hazard Ratio, HR) and absolute (as % at 5 years) efficacy in terms of disease free survival (DFS), distant metastasis free survival (DMFS) and overall survival (OS) of chemotherapy in the two subgroups where the clinical-pathological prognosis and the 70- gene signature molecular prognosis are discordant. To calculate overall estimates of efficacy (DFS, DMFS, OS) for each treatment strategy according to clinical-pathological prognosis and according to molecular prognosis. To estimate the percentage of patients receiving chemotherapy per each prognostic method. Gene profiling: To identify and/or validate predictive gene expression profiles of clinic al response/resistance to anthracycline-based and docetaxel-capecitabine chemotherapy. To identify and/or validate novel gene expression signatures predicting clinical response to sequential tamoxifen-ai versus AI alone. Chemotherapy: To compare the OS distributions associated with the 2 chemotherapy arms. To determine and compare the early and late toxicities associated with each regimen. Endocrine therapy: To evaluate adjuvant endocrine treatment success or failure in the subgroup of postmenopausal patients with endocrine responsive disease. To compare the safety profile of the two endocrine therapy regimens. Long term research objectives: To set up several biological material bank resources (RNA, tumor tissue, blood & serum) for future translation studies in both genomics and proteomics. 2.3 Endpoints Treatment decision randomization (R-T) The primary endpoint for the treatment decision randomization is distant metastasis free survival (DMFS) at 5 years. Secondary endpoints are: the proportion of women treated with chemotherapy per treatment decision making tool i.e. clinical prognosis compared to 70-gene signature prognosis overall survival at 5 years DFS at 5 years Version / 152 May 21, 2014

186 2.3.2 Chemotherapy randomization (R-C) The primary endpoint for the chemotherapy randomization is disease free survival (DFS). Secondary endpoints are: overall survival at 5 years safety both early and late DMFS at 5 years Endocrine therapy randomization (R-E) The primary endpoint for the endocrine therapy randomization is disease free survival (DFS). Secondary endpoints are: overall survival at 5 years DMFS at 5 years 3 Patient selection criteria 3.1 General eligibility criteria for inclusion in the trial To be eligible for the trial patients will have to comply with the following eligibility criteria: women with histologically proven operable invasive breast cancer, 0 to 3 positive nodes and no distant metastases. Acceptable primary treatment options are: breast conserving surgery or mastectomy with either a sentinel node procedure or full axillary clearance. radiotherapy is mandatory in the case of breast conserving surgery and will be administered according to local institutional policy after mastectomy. patients with unresectable positive deep margins who will receive adjuvant radiotherapy are eligible provided that all other margins are negative. a tumor clinical classification of T1, T2 or operable T3. the breast tumor must be unilateral. Multi focal tumors are allowed provided that they have identical histology. DCIS or LCIS are allowed if invasive cancer is present. a frozen tumor sample (not fixed in formalin) must be available for inclusion. The tumor sample must be taken from the excised primary tumor (a core punch biopsy). patients should be aged 18 years at randomization. have a WHO performance status of 0 or 1. have adequate bone marrow reserves (neutrophil count >1.5 x109 /l and platelet count >100 x109), adequate renal function (serum creatinine 1.5 x upper limit of normal) and hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline phosphatase 2.5 x upper limit of normal and total bilirubin 2.0 x upper limit of normal). have NO serious cardiac illness or medical condition including but not confined to: a history of documented congestive heart failure (CHF), high-risk uncontrolled arrhythmias, angina pectoris requiring antianginal medication, clinically significant valvular heart disease, evidence of Version / 152 May 21, 2014

187 transmural infarction on ECG, poorly controlled hypertension (e.g. systolic > 180 mm Hg or diastolic > 100 mm Hg), symptomatic coronary artery disease or a myocardial infarction within the last 12 months or other risk factors that contra-indicate the use of anthracycline-based chemotherapy. NOT have serious uncontrolled intercurrent infections, or other serious uncontrolled concomitant disease. NOT have previous or concurrent cancer, possible exceptions are: adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, lobular or ductal carcinoma in situ of the breast and any invasive cancer (other than breast cancer) in complete remission for 5 years. NOT have received (neo) adjuvant chemotherapy, (neo) adjuvant endocrine therapy or radiotherapy for the primary breast cancer considered for this trial. NOT have participated in any investigational drug study within the 4 weeks preceding the start of treatment. NOT be pregnant or breast-feeding at the time of diagnosis or randomization. A woman of childbearing potential must have a negative pregnancy test. If post-menopausal, the woman must have been amenorrheic for at lest 12 months to be considered of non-childbearing potential while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, some IUDs, condoms, sexual abstinence or vasectomised partner (Note 3 of the guidance on non-clinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). NOT have any psychological, familial, sociological, geographical or serious uncontrolled medical (e.g. a history of uncontrolled seizures, central nervous system disorders, or psychiatric disability) condition judged by the investigator to be clinically significant which could potentially preclude informed consent or interfere with compliance for oral drug intake or with the study protocol and follow-up schedule. These conditions should be discussed with the patient before enrollment in the trial. signed written informed consent must be given according to ICH GCP, and national/local regulations before enrollment in the trial (Both a screening PIS & IC and the PIS & IC 1 must be signed before enrollment). 3.2 Selection criteria for treatment decision randomization (R-T) Patients who meet all the general eligibility criteria and for whom the genomic and clinical risk assessments are discordant will be randomized between genomic-treatment decision making and clinical-treatment decision making. (details of stratification factors are given in section ) Version / 152 May 21, 2014

188 3.3 Selection criteria for chemotherapy randomization (R-C) Patients will be eligible for inclusion in the chemotherapy randomization (R-C) if they meet all of the general eligibility criteria AND the following criteria: (details of stratification factors are given in section ) women who have a high risk of recurrence according to both the clinical-pathological criteria and the 70-gene signature. OR women who have a high risk according to the clinical-pathological criteria and a low-risk according to the 70-gene signature, and were randomized (R-T) to use the clinical-pathological criteria for chemotherapy decision. OR women who have a low-risk according to the clinical-pathological criteria and a high-risk according to the 70-gene signature and were randomized (R-T) to use the 70-gene signature for chemotherapy decision. AND WHO status 0 or 1 (see Appendix B) normal ECG compatible with chemotherapy administration have cardiac function (left ventricular ejection fraction (LVEF) within the normal limits of each institution, measured either by echocardiography or by radionuclide angiocardiography (MUGA)) according to the standard of care. written informed consent (PIS & IC 2) prior to beginning specific protocol chemotherapy procedures, to be given and documented according to ICH GCP and national/local regulatory requirements. interval between definitive surgery (second surgery, if applicable) and start of chemotherapy should ideally be not more than 8 weeks but cannot exceed 120 days. NOT have had any organ allografts requiring immunosuppressive therapy. NOT have a requirement for concurrent use of the antiviral agent sorivudine (antiviral) or chemically related analogues, such as brivudine. NOT have a history of severe hypersensitivity reaction to drugs formulated with Polysorbate 80. have physical integrity of the upper gastrointestinal tract, ability to swallow tablets, and NO malabsorption syndrome. Version / 152 May 21, 2014

189 3.4 Selection criteria for endocrine therapy randomization (R-E) Patients who meet all the general eligibility criteria (see section 3.1) and who have endocrine responsive disease AND meet the following criteria will be offered the endocrine therapy randomization (R-E). (details of stratification factors are given in section ) are hormone receptor positive, either ER, PgR or both. have given written informed consent (PIS & IC 3) prior to beginning specific protocol endocrine therapy procedures, according to ICH GCP and national/local regulatory requirements. NOT have a prior history of thromboembolic disorder, DVT or pulmonary emboli. premenopausal women must have ovarian function suppression during the whole of the duration of protocol endocrine therapy (see 7.3 for details). patients must not have previously received high dose systemic corticosteroids (except as part of their anti-emetic treatment), immunotherapy or biological response modifiers (e.g.: interferon). hormone replacement therapy (HRT) must be stopped at least 4 weeks before trial endocrine treatment is initiated. patients who have received adjuvant anti-estrogen therapy for more than 1 month immediately following surgery, radiotherapy, and/or chemotherapy may NOT be proposed enrollment in the endocrine question of the trial. women with a history of breast cancer chemoprevention or osteoporosis treatment with antiestrogens (e.g. tamoxifen, raloxifen) can be included if at least 18 months has elapsed between anti-estrogens discontinuation and clinical or radiological diagnosis of breast cancer. 4 Trial Design is a phase III randomized superiority trial with three randomizations to compare clinical-pathological risk assessment to gene expression risk assessment using the 70-gene signature. It will also assess the efficacy of commonly used anthracycline-based chemotherapy regimens (anthracyclines followed by docetaxel for node positive), selected from a list of acceptable regimens, to the new capecitabine-docetaxel combination. In addition the long term safety and efficacy of 2 years of tamoxifen followed by 5 years of letrozole versus 7 years of letrozole will be investigated. Women with 0 to 3 positive lymph nodes will be offered those randomizations for which they are eligible, as shown below. Eligibility will be determined depending on an individual s clinical and genomic risk assessments, the consequent decision to administer chemotherapy, and their eligibility for endocrine therapy. Version / 152 May 21, 2014

190 Screening Visit surgeon (T1, T2 or operable T3 & M0) screening PIS & IC, Registration Surgery* Tumor sample shipment, RNA extraction, microarray analysis, local pathology, TNM, lymph node status, HR status 0 to 3 positive nodes Signing of PIS & IC 1 & Enrollment Genomic/Clinical Prognosis review & randomization Low/Low Discordant R-T High/High No CT CT R -C HR - HR + HR + HR - R-E CT Chemotherapy R - T Treatment decision Randomization based on genomic vs clinical prognosis R - C Chemotherapy Randomization of Anthracycline based CT (FEC D for N+) vs Docetaxel /Capecitabine R - E Endocrine treatment Randomization Letrozole vs Tamoxifen followed by Letrozole HR Hormone receptor PIS & IC Patient information sheet and informed consent * Delay between surgery and start of systemic treatment ideally not more than 8 weeks and it cannot be > 120 days Version / 152 May 21, 2014

191 4.1 Screening Prior to entering into the trial, at the time of surgery, when patient eligibility is not known, all patients who sign a screening PIS & IC to consent to the donation of a tumor and (optional) blood sample will be registered for screening. 4.2 Enrollment The date when the patient signs the PIS & IC 1 (after both the genomic and clinical risk assessments have been performed) will be referred to as the ENROLLMENT DATE. There are 3 possible randomizations in this trial and each of the 3 will only randomize a sub-set of the enrolled patients. Eligible patients with adequate tumor tissue for microarray analysis will be enrolled. Genomic risk using the 70-gene signature prognosis and clinical-pathological risk using Adjuvant! Online will be determined. Patients are evaluated as low clinical-pathological risk, if their 10-year disease specific survival (without chemotherapy or endocrine therapy) is estimated by Adjuvant! Online (Ref. 40) as greater than 88% for ER-positive patients, and greater than 92% for ER-negative patients (details given in section ). Adjuvant! Online The current standard version of Adjuvant! Online is version 8.0, as new standard (but not genomic) versions become available the new standard version will be incorporated into the web based platform and used to determine clinical risk for the trial. Local pathology categorization of hormone receptor status for stratification For stratification at the time of randomization, the local immunohistochemistry results will be used. Investigators will report positive, negative or unknown for both estrogen receptor and progesterone receptor according to local policies. Some guidelines are given below on levels for receptor positivity using various methods. Local pathology can report estrogen and progesterone receptor levels either as a percentage of immunoreactive cells, as a combination of percentage of immunoreactive cells and staining intensity using the Allred scale or as a biochemical concentration. If a percentage of immunoreactive cells is reported, the absence of immunoreactive cells is negative, 1% immunoreactive cells is positive. Using the Allred scale below 3 is deemed negative and 3-8 positive. If a biochemical concentration is reported, < 10 fmol receptor per mg cytosolic protein is negative and 10 fmol receptor protein/mg cytosolic protein is positive. Guidelines for reporting local ER/PgR positivity & negativity Positive: Negative: ER 1% immunoreactive cells, Allred score > 2 or 10 fmol/mg PgR 1% immunoreactive cells, Allred score > 2 or 10 fmol/mg ER < 1% immunoreactive cells, Allred score < 3 or < 10 fmol/mg PgR < 1% immunoreactive cells, Allred score < 3 or < 10 fmol/mg Local pathology categorization of HER-2 status for stratification For stratification at the time of randomization, the local pathology results will be used. Investigators will report positive, negative or unknown for HER-2 status according to local policies. Immediately after enrollment the web based system will inform the Investigator if the patient will receive chemotherapy or not. The web based platform will calculate both clinical and genomic risk and randomize those patients who are discordant for the two methods. For all enrolled patients the web based platform will specify the individual genomic and clinical test results and the outcome of the randomization if the patient is discordant. Version / 152 May 21, 2014

192 4.3 Treatment decision randomization for chemotherapy allocation Patients who are high risk for both prognostic methods will receive chemotherapy, and patients who are low risk for both prognostic methods will not receive chemotherapy. Patients for whom both methods are discordant will be randomized (R-T) between chemotherapy-decision-making according to clinical criteria (using Adjuvant! Online) or chemotherapy-decision-making according to genomic prognosis using the 70-gene signature. Use Clin-Path risk to decide Chemo or not Use 70-gene signature risk to decide Chemo or not Clin-Path High 70-gene signature Low: CT 70% pts Clin-Path High 70-gene signature Low: no CT 70% pts Clin-Path Low 70-gene signature High: no CT 30% pts Clin-Path Low 70-gene signature High: CT 30% pts As a result of this randomization, as a whole, half of the patients with discordant risks will be proposed chemotherapy and half will not be proposed chemotherapy. Also, for half of these patients chemotherapy will be assigned according to clinical-pathological risk, and for the other half chemotherapy will be assigned according to the 70-gene signature risk. Patients with HER-2 positive tumors which have both methods discordant and were randomized to no chemotherapy, can receive adjuvant trastuzumab alone or with adjuvant endocrine therapy (if hormonal receptor positive), if decided by the treating physician. Patients with HER-2 positive tumors that are classified low risk by both methods can receive adjuvant trastuzumab alone or with adjuvant endocrine therapy (if hormonal receptor positive), if decided by the treating physician and if no issues for trastuzumab reimbursement exist in the investigator s country. 4.4 Chemotherapy randomization trial design The chemotherapy question is a randomized, two-arm, prospective, non-blinded, multi-center phase III study. Patients who are candidates for chemotherapy (also see treatment section) will be proposed randomization R-C between anthracycline based chemotherapy (anthracyclines followed by docetaxel for node positive) or docetaxel-capecitabine chemotherapy. The following patients will be offered the chemotherapy randomization (R-C): Women who have a high risk of recurrence according to both the clinical-pathological criteria and the 70-gene signature. Women who have a high risk according to the clinical-pathological criteria and a low-risk according to the 70-gene signature, and were randomized (R-T) to use the clinical-pathological criteria for chemotherapy decision. Women who have a low-risk according to the clinical-pathological criteria and a high-risk according to the 70-gene signature and were randomized (R-T) to use the gene signature for chemotherapy decision. The objective is to evaluate if the docetaxel-capecitabine regimen is a more effective and less toxic alternative to an anthracycline-based regimen, for high-risk patients with 0 to 3 positive lymph nodes. Version / 152 May 21, 2014

193 Those patients who should be proposed chemotherapy but who do not wish to participate in the chemotherapy randomization (R-C) should receive the hospital s standard chemotherapy. They remain on protocol and should be offered the endocrine therapy randomization if applicable. Trastuzumab will be allowed and recommended (however not provided free as a trial drug) for women with HER-2 positive tumors if it becomes approved and available for use in the adjuvant setting during the trial. Treatment with trastuzumab can be proposed either sequential or concomitant with chemotherapy (including patients in capectibaine-docetaxel arm). Endocrine therapy treatment should be started only after the completion of chemotherapy and according to the endocrine therapy randomization (R-E) for consenting patients. 4.5 Endocrine therapy randomization trial design All post-menopausal patients who are candidates for endocrine therapy will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. Pre-menopausal patients may be offered this randomization at the discretion of their treating physician. For these patients, the suppression of ovarian function (temporarily or definitively) is mandatory during the whole duration of the protocol endocrine therapy. GnRH will not be provided. Endocrine therapy should start after surgery for those patients who did not receive chemotherapy or after chemotherapy for those patients who received chemotherapy (R-C). Those patients who have endocrine responsive tumors but who do not wish to participate in the endocrine therapy randomization (R-E) should receive the hospital s standard endocrine therapy and remain on protocol. 4.6 pilot study The first 800 women randomized into the trial will form a pilot study to ensure that the clinical trial population is not biased and that key assumptions hold. They will also be used to check that the trial logistics function correctly and that physicians/patients comply with the allocated randomizations. The trial will not be interrupted after the first 800 patients unless the assumptions prove to be incorrect. (details of which can be found in the IDMC section, chapter 11) Note: For details of statistical testing refer to the statistics section (chapter 10). 5 Guidelines for breast cancer surgery and adjuvant radiotherapy The following information is given as recommendations but it is not an integral part of protocol investigations. 5.1 Breast cancer surgery Patients should receive optimal loco-regional treatment. The surgical part of the breast cancer treatment may consist of breast conservation surgery (wide local excision of the primary tumor) or mastectomy. The surgical part of the axilla encompasses the sentinel node procedure (SNP) for lymphatic staging or axillary lymph node dissection (ALND) for the treatment of lymph node metastases or staging procedure. Since the SNP can be considered as standard of care for lymphatic staging, SNP is preferred over ALND for this purpose. To ensure as much as possible uniformity in surgical techniques, the operations are described in more detail. It is recommended to follow these evidence based guidelines (Ref. 89, Ref. 90) for surgery as much as possible. Version / 152 May 21, 2014

194 Wide local excision The main aim is to achieve free tumor margins. To achieve this aim, the surgeon should try to remove at least a 1 cm rim of healthy breast tissue. Particularly in large tumors, substantial parts of the breast have to be removed and special attention should be made to breast reconstruction and cosmesis. The so called segment (or preferably sector resection, which is the proper mathematical denominator for a piece of pie excision) resection is one of the better techniques. Breast tissue is removed from the skin (either with or without a skin island for cosmetic reasons) down to the basal fascia. The advantage of this technique is that margin assessment is facilitated (only the lateral margins, periphery and nipple side are relevant; not skin and fascia side), as is the possibility to reconstruct the breast tissue. To find and excise non-palpable cancers, a number of aids are available and the guide wire is the most widely applied method. Gamma probe directed excision after intra-lesional radioisotope injection and ultrasound directed excisional biopsy appears to be helpful to achieve free margins, compared to guidewire directed wide local excisions. If cancer is diagnosed with minimal invasive techniques, it should be possible to achieve free margins in over 80% of patients at the first operation. The complication rate should be as low as 5%. For invasive ductal cancer, after complete excision and radiotherapy to the breast, the local relapse rate should be less than 1% per year of follow-up. Cosmetic outcome should be acceptable and good in over 70% of patients. Simple mastectomy The aim of simple mastectomy is the removal of preferably all breast tissue. Removal of the entire breast with its skin and the nipple-areola complex, with the pectoralis major muscle fascia, but without axillary node dissection. There is no axillary clearance involved. However, if the axillary tail is removed frequently some lower axillary nodes are included in the resection. Skin-sparing mastectomy Removal of the entire breast with preservation of its skin, except the nipple and the areola to facilitate immediate reconstruction. Skin sparing mastectomy should only be performed in the context of immediate reconstruction. When patient desires breast reconstruction, immediate reconstruction has advantages over delay: less procedures, better cosmesis and better patient satisfaction. Lymphatic mapping by sentinel node procedure SN biopsy should only be performed in patients with a clinically negative axilla. It has been shown convincingly that sentinel node can stage the axilla with high accuracy. Consensus on the sentinel node procedure states the following quality issues. The procedure should be performed within a team of surgeons, pathologists and nuclear physicians dedicated to breast pathology. The team should go through a learning phase of at least 20 procedures, including backup ALND. Newly trained surgical members of such a team can master the procedure under supervision without backup ALND. The sentinel node should be identified in over 90% of the patients. The long-term false negative rate should be < 5%. The preferred surgical technique is as follows. Just before sterile draping the blue dye is injected (1-4 cc; in or around the tumor, intracutaneously) The location of the SN is identified by scanning with the probe over the site where the SN's are suspected (and preferably indicated by the nuclear physician according to the lymphoscintigraphy) After massaging the breast (a few minutes), the incision is placed over the location were the sentinel node is suspected. Version / 152 May 21, 2014

195 If the sentinel node is in the axilla, the incision is usually placed behind the free edge of the major pectoral muscle at the lower border of the hair implant. The blue lymphatics can usually be identified behind Scarpas fascia. The dissection should be meticulous, not to sever the lymphatics. If no blue lymphatics are identified, the probe can be used to identify the radioactive nodes. Radioactive and/or blue nodes are removed and afferent or efferent lymphatics secured. Internal mammary chain nodes can be approached either by a separate incision or through the same incision for mastectomy or wide local excision. The identification and removal of the SN is at the medial site of the intercostal space along the internal mammary artery. It requires careful dissection, but it is not a difficult technique. All nodes that are possibly SN are removed (if lymphoscintigraphy shows unequivocally non SN or second echelon SN, these hot nodes are not to be removed). Local pathology categorization of lymph node status micromets (0.2mm - 2mm) pn1mi are considered as LN positive submicromets (<0.2mm) pn0(i+) are considered as LN negative Management of sentinel lymph node micrometastasis Patients with micrometastasis (0.2 to 2 mm) in a SLN should be considered as LN positive and undergo a complete axillary dissection unless they are randomized in a clinical trial evaluating treatment options for this patient population. In such case, patients can follow the allocated treatment. Axillary Lymph Node Dissection (ALND) Traditionally, the indication for ALND is either to provide prognostic information on the basis of the nodal status and provide excellent regional control. If ALND is applied for this purpose, at least 10 lymph nodes should be retrieved and examined. For staging the axilla, the SN procedure appears to be equal to ALND. All incisions are acceptable, providing there has been sufficient exposure of the entire axillary content. In breast conservation cases the clearance should preferably be performed discontinuously from the breast excision using transverse incisions. All axillary fat from at least levels I and II should be excised in one specimen; The medial border is formed by a curved plane ranging from the muscles to the vessels and nerves going to the pectoral muscles (in the interpectoral area, the interpectoral fat, which might contain nodes, should be dissected carefully from this neurovascular bundles). The medial line is further indicated by a sagittal plane through the medial border of the m. pectoralis minor (patient in supine position). The cranial border is formed by plexus and axillary vein; The lateral border is from the white tendon downward to the m. latissimus dorsi; The dorsal border is the fascia of subscapular muscles; the n. thoracodorsalis and vessels may be spared, but should be resected in cases with suspicious nodes fixed to these structures; Caudally the upper-outer quadrant of the breast; care is given to resect the fat between thoracic wall and upper-outer quadrant of the breast; resection of parts of breast tissue might therefore be necessary; Level III axillary fat (apical, subclavicular) may be resected, en bloc with the specimen or separately for staging purposes. Version / 152 May 21, 2014

196 5.2 Adjuvant radiotherapy Radiotherapy to the breast is mandatory for all patients who have undergone breast-conserving surgery. The use of a boost dose to the tumor bed should be considered, especially for patients 60 years of age or less, and is mandatory for patients with histologically incomplete tumor resection margins. Chest wall radiotherapy after mastectomy is optional, but mandatory for patients with incomplete tumor resection margins. Regional nodal irradiation (axillary, peri-clavicular and /or internal mammary chain) should be given according to local or national guidelines. For all patients, care should be taken to minimize exposure to heart and lung tissue. To achieve this, the use of a computer tomography (CT) based treatment simulation and planning is recommended, and is mandatory if the internal mammary lymph node chain is part of the target volume. 6 Chemotherapy: therapeutic regimens, expected toxicity, dose modifications The first cycle of chemotherapy will be administered within 4 weeks following the randomization R-C date and ideally not more than 8 weeks after the definitive surgery. If chemotherapy has not started within 120 days of the definitive surgery the patient becomes ineligible for R-C. 6.1 Treatment plan, anthracycline arm The Anthracycline arm will be different for patients with lymph-node negative breast cancer and for patients with 1 to 3 positive lymph nodes, as the current standard therapies are different for each subgroup. Lymph node negative The anthracycline arm will include a standard three or four-drug anthracycline-based regimen, with either doxorubicin or epirubicin, chosen from the list of permitted anthracycline-based regimes (see below) and provided according to standard of care. Cycles should be given every 3 to 4 weeks, depending on the choice of treatment. The following are accepted regimens: CAF (d1+8), FAC (cyclophosphamide, doxorubicin and 5- fluorouracil), FEC (cyclophosphamide, epirubicin and 5-fluorouracil), CEF (d1+8) or four cycles of full-dose, single-agent epirubicin followed by 4 cycles of cyclophosphamide, methotrexate, and fluorouracil (CMF) called E-CMF. Despite their differences, these regimens have in common a high and adequate dose-intensity of anthracycline and enable various collaborating groups with different policies to participate in the trial. One to 3 positive lymph nodes The anthracycline arm will include one chemotherapy regimen, consisting of the sequence of 3 cycles of FEC 100 followed by 3 cycles of docetaxel. Cycles should be given every 3 weeks. This regimen is currently perceived as standard treatment in several EU countries for this subpopulation, and therefore it will be the standard anthracycline-based treatment for patients with 1-3 positive nodes in the chemotherapy question. Version / 152 May 21, 2014

197 6.1.1 Drug information, anthracycline arm Common toxic effects of the above mentioned regimens include nausea, vomiting, myelosuppression, febrile neutropenia, hemorrhagic cystitis (in schemes using cyclophosphamide), fever and infection, alopecia, mucositis, fatigue, local reaction (in the site of injection), anorexia, diarrhea, amenorrhea (in premenopausal women) and serum biochemistry abnormalities. More serious longer term side effects are cardiotoxicity and leukemia. For cardiotoxicity, the clinical presentation can vary from an asymptomatic decrease in the left ventricular ejection fraction to clinically overt congestive heart failure. It is directly related to the cumulative dose administered, with an incidence of 1-2% at a cumulative dose of 400 mg/m² of doxorubicin, rising exponentially above a total dose of mg/m², when the incidence may be as high as 26%. Epirubicin is associated with a lower frequency of cardiotoxicity at therapeutic dosages in comparison with doxorubicin (i.e., a reported incidence of 2-3% at 900 mg/m² epirubicin). Associated risk factors include underlying cardiac disease, prior or concomitant radiotherapy to the mediastinal area and advanced age. The utilization of modern techniques of radiotherapy seems not to add to this risk substantially. The incidence of secondary acute leukemia, another potentially serious long-term side effect, is also uncommon and ranges from 1-2% at 5 to 10 years of follow-up. It should be noted that the dose of anthracyclines used in the trial is kept well below the maximal tolerated doses around 360 mg/m² for doxorubicin and 600 mg/m² for epirubicin. For toxicity information regarding docetaxel, used in the FEC D regimen for lymph node positive disease, please refer to section Drug administration, anthracycline arm The following are the accepted regimens and doses that can be used in the trial: Options for node negative patients FEC 100: Fluorouracil 500 mg/m², Epirubicin 100 mg/m², Cyclophosphamide 500 mg/m², all i.v. every 3 weeks for 6 cycles (Ref. 91) Canadian CEF: Cyclophosphamide orally 75 mg/m² d1-d14, Epirubicin 60 mg/m² i.v. d1 and d8, Fluorouracil 500 mg/m² iv d1 and d8, every 4 weeks for 6 cycles; if oral cyclophosphamide is not tolerated, it may be switched to i.v. doses of 370 mg/m² on days 1 and 8 (Ref. 47) FAC: Cyclophosphamide 500 mg/m² d1, Doxorubicin 50 mg/m² i.v. d1, Fluorouracil 500 mg/m² iv d1, every 3 weeks for 6 cycles (Ref. 92) CAF: Cyclophosphamide 600 mg/m² d1, Doxorubicin 60 mg/m² i.v. d1, Fluorouracil 600 mg/m² iv d1 & d8, every 4 weeks for 6 cycles (Ref. 93) E CMF: Epirubicin 100 mg/m² d1 every 3 weeks for 4 cycles followed by either classical CMF (Cyclophosphamide orally 100 mg/m² d1-d14, methotrexate 40mg/m² i.v. d1 and 8 and 5- fluorouracill 600 mg/m² i.v. d1 and 8) every 4 weeks for 4 cycles or i.v. dose modified CMF (750 mg/m², 50 mg/m², 600 mg/m², days 1) every 3 weeks for 4 cycles (Ref. 94) Options for node positive patients FEC D: Fluorouracil 500 mg/m2, Epirubicin 100 mg/m², and Cyclophosphamide 500 mg/m2 (FEC) intravenously on day 1 every 3 weeks for 3 cycles followed by docetaxel 100 mg/m2 intravenously on day 1 every 3 weeks for 3 cycles (Ref. 61) Version / 152 May 21, 2014

198 Each treatment will be given as scheduled (3 or 4 weeks) ± 3 days. 5-Fluorouracil may be administered as an i.v. bolus over 15 minutes or less; Epirubicin as an i.v. infusion over 1 hour; Doxorubicin as an i.v. bolus in 10 to 15 minutes, Cyclophosphamide as an i.v. infusion over 1 hour or less and Methotrexate as an i.v. bolus. The chemotherapy doses will be calculated according to baseline body surface area (BSA) for all cycles. If there is a 10% or greater change in body weight compared to baseline, the BSA will be recalculated. If the calculated BSA of the patient is >2.2 m2, it is recommended to calculate the dose to be given using BSA=2.2m2. No ideal body weight should be used for the calculation of BSA. Dose reduction and/or treatment delay and treatment discontinuation are planned for the 2 arms in case of severe hematological and/or non-hematological toxicities. Duration of therapy: Chemotherapy will be given for 6 cycles or 8 cycles for E-CMF, which roughly corresponds to 5 to 6 months of treatment, depending on the chemotherapy scheme to be used. Version / 152 May 21, 2014

199 Lymph node negative Anthracycline based Regimens FEC cycles Day Day 1 8 FEC Day 15 Day 22 CAF 6 cycles Day 1 Day 8 CAF F Day 15 Day 22 Day 29 1 Cycle = 21 days 1 Cycle = 28 days Canadian CEF 6 cycles Day 1 Day 8 Day 15 EF EF C C Day 22 Day 29 FAC 6 cycles Day Day 1 8 FAC Day 15 Day 22 1 Cycle = 28 days 1 Cycle = 21 days E CMF 8 cycles (4 x A followed by 4 x B) Day Day Day Day Day MF E x4 C Day 8 MF C Day 15 Day 22 Day 29 (1 Cycle A = 21 days) x 4 F = 5 Fluorouracil E = Epirubicin C = Cyclophosphamide A = Doxorubicin M = Methotrexate D - Docetaxel (1 Cycle B = 28 days) x 4 OR Day Day Day Day CMF (1 Cycle B = 21 days) x 4 Lymph node positive FEC D 6 cycles (3x FEC100 followed by 3 x Docetaxel) Day 1 FEC Day 8 Day 15 Day 22 x3 Day 1 D Day 8 Day 15 Day 22 x3 1 Cycle = 21 days 1 Cycle = 21 days Version / 152 May 21, 2014

200 6.1.3 Premedication, anthracycline arm A prophylactic antiemetic treatment is highly recommended and should be given according to standard of care in each center. No prophylactic antibiotics are recommended (except with Canadian CEF). No primary prophylaxis with G-CSF/GM-CSF/PEG-filgrastim is allowed. Pre-medication for docetaxel infusions is described in Patient monitoring anthracycline arm The usual pattern of care is required during infusion, including careful attention to reactions and side effects Dose adjustments anthracycline arm Full blood count will be performed on day 1 of the cycle and dose adjustments will be based on these results (and also based on clinical adverse events or findings observed during the study). Rounding of dosage is allowed if the rounding is within 5% of the calculated dose. Doses will be reduced for hematological and other adverse events (see table 3 & 4 below). Adverse events will be graded using the NCI Common Terminology Criteria for Adverse Events Version 3.0 (CTCAE) (see Appendix C). No dose modifications should be done for patients who experience grade I toxicities or other toxicities considered unlikely to become serious or life threatening. Dose modifications should be done based on the initial doses used and according to the greatest toxicity observed. Once done, they cannot be re-escalated. Treatment should be interrupted (for a maximum of 2 weeks) and the doses of drugs reduced by 25% in patients who experience a second occurrence of a given grade II toxicity (except alopecia) or any grade III toxicity. For patients who have received no trial drugs for 3 weeks due to toxicity, trial chemotherapy treatment should be stopped. Version / 152 May 21, 2014

201 Table 3 Dose modification for Hematological Toxicities Anthracycline-based regimens Toxicity Treatment of toxicity Modification of chemotherapy ANC < 1.0x109/l on the day of treatment Febrile Neutropenia 2nd episode of Febrile Neutropenia 3rd episode of Febrile Neutropenia Platelets < 75x109/l on the day of treatment Platelets < 50x109/l on the day of treatment Platelet transfusion None According to standard care in institute According to standard care in institute According to standard care in institute None None Wait until ANC > 1.0x109/l (maximum time up to 3 weeks), then use full dose; G-CSF or GM-CSF or PEG-Filgrastim may be used in subsequent cycles if allowed by the countries registration/reimbursement rules G-CSF or GM-CSF or PEG- Filgrastim with subsequent cycles Reduce dose of anthracycline (and accompanying drugs) or docetaxel for N+ subgroup by 25% Discontinue chemotherapy treatment Wait until platelet count > 75x109/l and reduce dose of anthracycline (and accompanying drugs) or docetaxel for N+ subgroup by 25% Wait until platelet count > 75x109/l and reduce dose of anthracycline (and accompanying drugs) or docetaxel for N+ subgroup by 25% Wait until platelet count > 75x109/l and reduce dose of anthracycline (and accompanying drugs) or docetaxel for N+ subgroup by 25% Version / 152 May 21, 2014

202 Table 4 Dose modification of Anthracycline-Regimens for Non-Hematological Toxicities Toxicity Grade Treatment of toxicity Nausea Any Local guidelines antiemetics and/or corticoid Vomiting Any Local guidelines antiemetics and/or corticoid Diarrhea III-IV Stop treatment until grade I, then change dose Mucositis III-IV Stop treatment until grade I, then change dose Dysphagia III-IV Stop treatment until grade I, then change dose Cystitis (If using Cyclophosphamide) I-II III-IV Careful follow-up plus orientation Stop treatment until grade I, then change dose Bilirubin I Wait until within normal limits, then change dose Alkaline Phosphatase III-IV Wait until Grade II, then change dose ASAT or ALAT III-IV Wait until Grade II, then change dose Alkaline Phosphatase and ASAT or ALAT Change to chemotherapy No modification No modification 25% dose reduction (Anthracycline) 1st occurrence- 25% dose reduction (anthracycline or docetaxel for N+ subgroup if applicable) 2nd occurrence discontinue chemotherapy treatment 25% dose reduction (Anthracycline or docetaxel for N+ subgroup if applicable) No change 1st occurrence- 25% dose reduction (cyclophosphamide) 2nd occurrence discontinue chemotherapy treatment Reduce docetaxel by 25% Reduce docetaxel by 25% Reduce docetaxel by 25% II Change dose Reduce docetaxel by 25% Edema Any No dose change; further treatment at physician s discretion Neurologic II Wait until subsequent cycle; if I, no change Version / 152 May 21, 2014

203 Toxicity Grade Treatment of toxicity III If there is no resolution Arthralgia/Myalgia Any Symptomatic treatment Skin rash Any Symptomatic treatment Asthenia III Symptomatic treatment Other III Stop treatment until grade I, then change dose IV Stop treatment until grade I, then change dose Change to chemotherapy Reduce docetaxel by 25% If persisting after dose reduction, discontinue docetaxel Discontinue docetaxel 1st occurrence- reduce docetaxel by 25% 2nd occurrence reduce docetaxel by 25% Reduce 25% (anthracycline and accompanying drugs or docetaxel for N+ subgroup if applicable) 1st occurrence- 25% dose reduction (anthracycline and accompanying drugs or docetaxel for N+ subgroup if applicable) 2nd occurrence discontinue chemotherapy treatment Cardiotoxicity Patients should be followed according to local practice for determining and following any cardiac toxicity. Cardiac toxicity events will be recorded according to the CTCAE guidelines. 6.2 Treatment plan docetaxel-capecitabine arm Drug information docetaxel-capecitabine arm Docetaxel When used as a single agent, the major toxic effect of docetaxel is neutropenia. Bone marrow suppression is dose-dependent and is the dose-limiting toxicity. It may be associated with fever (febrile neutropenia). Other toxic effects include anaphylactoid type reactions and cutaneous reactions, nausea, vomiting, oral mucositis, diarrhea, reversible paresthesia, alopecia, asthenia, and mild local venous reactions (phlebitis) at site of injection and fluid retention/edema. Reversible skin reactions characterized by localized erythema of the extremities followed by desquamation may occur within 1 week of the infusion but are rarely severe. Their incidence is almost negligible with the proposed premedication. Severe nail reactions, such as onycholysis, occur in less than 1% of patients. Alopecia occurs in approximately 40-80% of patients. Edema may occur, with several forms of presentations and varying degrees of intensity (mild to poorly tolerated peripheral edema, generalized edema, pleural effusion, cardiac tamponade, or pronounced abdominal distention due to ascites). It usually starts in the lower extremities and may ascend to become generalized. This severe form of presentation may occur in up to 6% of the Version / 152 May 21, 2014

204 patients and the treatment consists of salt restriction and diuretics if needed. The fluid overload is reversible. Neurosensory symptoms consist mainly of distal paresthesia, dysesthesia and/or pain and may be severe in approximately 5% of the patients; the dosage of the drug must be adjusted when these symptoms appear and if they continue or worsen, the treatment must be interrupted. Peripheral motor neuropathy occurs in less than 5% of the patients and manifests as distal extremity weakness. Anaphylaxis and severe hypersensitivity reactions characterized by bronchospasm/dyspnea and/or hypotension requiring treatment, angioedema, and generalized urticaria are the most troublesome adverse effects and have occurred in 2% of patients treated in previous large clinical trials. Although variable and possibly affecting any organ system, these toxicities usually do not cause a large number of patients to cease treatment. The drug is contraindicated in patients who have a history of severe hypersensitivity reactions to drugs formulated with polysorbate 80 and should be used according to the guidelines in patients with elevated baseline liver function tests. Non-compliance with these guidelines puts the patient at risk for docetaxel induced toxic death Capecitabine Capecitabine is foreseen as an outpatient treatment, and in certain circumstances adverse events that could occur, such as diarrhea, can rapidly become serious. In the case where a patient experiences any toxicity in between scheduled visits, the patient should be encouraged to contact the clinic as soon as is practical, for further directions or for treatment. It is essential that the patients are informed to interrupt capecitabine treatment as soon as a grade II toxicity occurs, therefore the patients will need specific explanations what to do in the case of the occurrence of the most frequent toxicities (diarrhea, hand-foot syndrome and stomatitis). Renal Impairment: Capecitabine is contraindicated in patients with moderate or severe renal impairment (creatinine clearance below 50 ml/min [Cockroft and Gault]); treatment should not be started or continued in patients with severe renal impairment. Patients with moderate or severe renal impairment at baseline will be ineligible for the study, as per the exclusion criteria. The incidence of grade III-IV adverse events in capecitabine (1250 mg/m2 twice-daily, intermittent therapy) treated patients with moderate renal impairment (creatinine clearance ml/min [Cockroft and Gault]) was increased compared to the overall treated population. Careful monitoring and prompt treatment interruption is recommended if patients develop grade II, III, or IV adverse events with subsequent dose adjustments as outlined in the does modification table 8. Coagulopathy: Patients receiving concomitant capecitabine and oral coumarin-derived anticoagulants should have their anticoagulant response (INR or prothrombin time) monitored frequently in order to adjust the anticoagulant dose accordingly. A clinically important capecitabine-warfarin drug interaction was demonstrated in a clinical pharmacology trial. Altered coagulation parameters and/or bleeding, including death, have been reported in patients taking capecitabine concomitantly with coumarin-derived anticoagulants such as warfarin and phenprocoumon. Postmarketing reports have shown clinically significant increases in prothrombin time (PT) and INR in patients who were stabilized on anticoagulants at the time capecitabine was introduced. These events occurred within several days and up to several months after initiating capecitabine therapy and, in a few cases, within one month after stopping capecitabine. These events occurred in patients with and without liver metastases. Age greater than 60 and a diagnosis of cancer independently predispose patients to an increased risk of coagulopathy. The use of low molecular weight heparin instead of coumarin is at the discretion of the investigator. Diarrhea: Capecitabine can induce diarrhea. Patients with severe diarrhea should be carefully monitored and given fluid and electrolyte replacement. If grade II, III or IV diarrhea occurs, Version / 152 May 21, 2014

205 administration of capecitabine should be immediately interrupted until the diarrhea resolves or decreases in intensity to grade 1 (and see guidelines in section 6.2.5). Hand-Foot Syndrome: Capecitabine has been shown to cause hand-foot syndrome (palmar-plantar erythrodysesthesia or chemotherapy-induced acral erythema-erythema) (Ref. 95, Ref. 96) which is characterized by varying degrees of redness, tenderness, pain and desquamation of the palms of the hands and the soles of the feet; numbness and tingling may also appear and in its most severe form it can interfere with daily activities. Nevertheless, it is reversible and responds to the diminution of the dose or the interruption of the treatment. If grade II or III hand-foot syndrome occurs, administration of capecitabine should be interrupted until the event resolves or decreases in intensity to < grade I. Subsequent doses of capecitabine should be administered as per the does modification table 8. Use of vitamin B6 pyridoxine ( mg bid) is permitted for symptomatic or secondary prophylactic treatment of hand-foot syndrome. Cardiotoxicity: There has been cardiotoxicity associated with fluorinated pyrimidine therapy (including capecitabine) including myocardial infarction, angina, dysrhythmias, cardiogenic shock, sudden death and electrocardiograph changes. These adverse events may be more common in patients with a prior history of coronary artery disease. Hyperbilirubinemia: Administration of capecitabine should be interrupted if treatment-related elevations in bilirubin of 3.0 x upper limit of the normal (ULN) or treatment-related elevations in hepatic aminotransferases (ALAT, ASAT) of 2.5 x ULN occur. Treatment may be resumed when bilirubin decreases to 3.0 x ULN or hepatic aminotransferases decrease to 2.5 x ULN. Pregnancy: Female patients must be instructed to stop taking capecitabine if they become pregnant during the study, and immediately inform the Investigator. The Investigator should counsel the patient, and discuss the risk of continuing with the pregnancy and the possible effects on the fetus. Monitoring of the patient should continue until conclusion of the pregnancy. The Investigator should report all pregnancies in the study to the Sponsor. Contraindications: Capecitabine is contraindicated in patients with severe leukopenia, neutropenia, or thrombocytopenia, severe hepatic impairment, or severe renal impairment (creatinine clearance below 30 ml/min). Use of capecitabine is contraindicated during pregnancy and lactation, and concomitantly with sorivudine or its chemically related analogues, such as brivudine. Capecitabine is generally well tolerated. The most common side effects are hand-foot syndrome and diarrhea which generally respond to changes in the treatment dose or schedule. Other side effects associated with the use of capecitabine are: nausea, vomiting, stomatitis, dyspepsia/abdominal pain, fatigue, anorexia, myalgia, insomnia, fever, myelosuppression, paresthesia, eye irritation and serum biochemistry alterations. Other skin toxicities apart from the hand-foot syndrome include rash, dry and/or itchy skin and are mainly grades I and II Docetaxel-capecitabine combination When used in combination with docetaxel, the incidence of neutropenia/neutropenic fever, gastrointestinal side effects and hand-foot syndrome may be higher, but still manageable with appropriate medical intervention, dose reduction and/or treatment interruption, as seen in the already mentioned phase III trial in the metastatic setting. In spite of the apparent increased toxicity, the measured Quality of Life (QoL) was not adversely affected. Version / 152 May 21, 2014

206 Docetaxel Capecitabine Regimen 6 cycles Docetaxel Day 1 Doc Day 8 Day 15 Day 22 Capecitabine Day Day Day Cap Cap Rest Day 22 1 Cycle = 21 days Drug administration docetaxel-capecitabine arm The combination docetaxel-capecitabine will be given for 6 cycles which will roughly correspond to 5 months of treatment. Docetaxel 75 mg/m² should be administered as a 1-hour i.v. infusion, on day 1. If the calculated BSA of the patient is >2.2 m², the dose to be given to the patient will be calculated according to BSA = 2.2m². No ideal body weight should be used for the calculation of BSA. Capecitabine 825 mg/m² will be given p.o., twice a day (total daily dose 1650 mg/m²) from days 1 to 14. Treatment will be given every 3 weeks (± 3 days) for a total of 6 cycles. For capecitabine, the daily dose will be derived by determining the body surface area (BSA). Use the tables of capecitabine doses in Table 5 to determine the appropriate dose for a given BSA. If the calculated BSA of the patient is >2.2 m², the dose to be given to the patient will be calculated according to BSA = 2.2m². If there is a 10% or more change in body weight between cycles, the BSA will be recalculated and capecitabine dose should be adjusted, according to table 5. Capecitabine (Xeloda ) will be provided as film-coated 150 mg and 500 mg tablets. Tablets are not scored and should not be split. Capecitabine will be taken as 825 mg/m2 doses twice-daily (total daily dose of 1650 mg/m2) morning and evening (3-week cycles consisting of 2 weeks of capecitabine treatment followed by 1 week without capecitabine treatment). At the discretion of the patient and the Investigator, one of two schedules can be used. The first schedule allows for the first dose of each cycle to be administered in the morning of day 1 and the last dose of each cycle to be administered in the evening of day 14, followed by a seven day rest period. This provides for a total of 28 single doses per cycle over 14 calendar days. The second schedule allows for the first dose of each cycle to be administered in the evening on day 1 and the last dose of each cycle to be administered on the morning of day 15, followed by a 6.5 day rest period. This provides for a total of 28 single doses per cycle over 15 calendar days. Either schedule may be used as long as the patient takes the required 28 single doses over consecutive days. The patient does not need to use the same schedule for all cycles, but may choose the other schedule for the next cycle. For example, on cycle 1, the patient visits the clinic in the morning and starts capecitabine in the evening. The patient's last dose is on the morning of day 15. For cycle 2, the patient takes a dose in the morning of day 1 and takes the last dose on day 14 in the evening. The morning and evening dose should be given approximately 12 hours apart, and taken within 30 minutes after the ingestion of food with approximately 200 ml of water, ideally after the breakfast and evening meal. The patient will be given a sufficient supply of study medication to accommodate 14 days (28 doses) of capecitabine treatment every 3 weeks. Version / 152 May 21, 2014

207 Table 5 Pill combinations to be taken to deliver the correct dose per ranges of BSA based on a daily posology of 1650 mg/m2 100% Dose Level = twice daily 825 mg/m2 Surface Area (m2) Total Dose per Administration (mg) Number of tablets to be taken in the Morning Evening 150 mg 500 mg 150 mg 500 mg < Premedication docetaxel-capecitabine arm Following the standard of care, prophylactic steroids should be given before docetaxel infusion; a scheme with oral steroids such as dexamethasone 16 mg per day (e.g., 8 mg BID) is suggested, for 3 days starting the day before docetaxel administration, in order to reduce the incidence and severity of fluid retention as well as the severity of hypersensitivity reactions. The following schedule is recommended; dexamethasone 8 mg p.o. for 6 doses given: the night before chemotherapy, the morning of chemotherapy, 1 hour before docetaxel infusion (this dose may be given intravenously), the night of chemotherapy, the morning of the day after chemotherapy and the night of the day after chemotherapy. If the patient does not take the 8 mg p.o. dexamethasone the morning before and/or the night before the scheduled dose of docetaxel, she can not be treated as scheduled. Table 6 Prophylactic regimen for docetaxel-capecitabine arm 3 day schedule Steroid: Dexamethasone 8 mg i.v. or i.m. or orally, or Methylprednisolone 40 mg i.v. or i.m. or 32 mg orally, or Prednisone 50 mg i.v. or i.m. or orally, or Prednisolone 50 mg i.v. or i.m. For 6 doses 1. night before chemotherapy (- 12 h) 2. the morning of chemotherapy (- 2 h) 3. 1 hour before docetaxel infusion (- 30 min) or other equivalent Additional 3 doses (+12h, +24h, +36h) Observation: It is advisable not to use doses of dexamethasone (or its equivalents) greater than 30 mg/day but additional oral doses of dexamethasone may be used according to local policy. No prophylactic antibiotics are allowed. Hematopoietic growth factors (i.e., G- or GM-CSF) may be used according to institutional guidelines to treat febrile neutropenia, but should not be used as primary prophylaxis. Use of any growth factor support must be documented in the patient records. Growth factors must be discontinued at least 48 hours prior to initiation of the next capecitabine cycle Patient monitoring docetaxel-capecitabine arm No special monitoring, standard care, as with patients on the anthracycline arm. Attention must be given to the possible occurrence of hypersensitivity reactions. Version / 152 May 21, 2014

208 6.2.5 Dose adjustments docetaxel-capecitabine arm The full blood counts are to be performed on day 1 of each cycle and/or if symptoms are present. In case there is an isolated grade 3 or 4 neutropenia/granulocytopenia without fever during the cycle, capecitabine will not be interrupted. No changes should be done if patients experience grade I toxicities. Dose modifications should be done based on the initial doses used and according to the greatest toxicity observed. Once done, they cannot be re-escalated. Treatment should be interrupted (for a maximum of 2 weeks) and the doses of drugs reduced by 25% in patients who experience a second occurrence of a given grade II toxicity (except alopecia) or any grade III toxicity. If Docetaxel and/or Capecitabine therapy should be discontinued, due to toxicities and/or interruption lasts for more than 3 weeks, the patient should be withdrawn from the chemotherapy part of the study and be treated at the discretion of the attending physician. However, if the patient needs discontinuation of one of the drugs due to toxicity, it is at the investigators discretion to continue with the remaining drug or to stop overall. Capecitabine dose reduction should be adapted following the recommendations in table 9. In case blood count test is performed during the 14 days of Capecitabine administration, treatment should only be interrupted if a non-hematological adverse event (grade 2) coincides with a grade 3 or 4 neutropenia. In that case, one should wait until resolution of both adverse events (including the neutropenia) to a grade 0 or I. It is recommended to use G-CSF or PEG-filgrastim in subsequent cycles. The doses for toxicity grade II and III occurrence should be adjusted as mentioned above. If the calculated creatinine clearance decreases during treatment to a value of 50 ml/min (due to an increase in serum creatinine or decrease in body weight), this change is not, by itself, a reason for a dose reduction. Dose reductions during treatment should be based on adverse events. For any adverse event apparent at baseline, the dose modifications will apply according to the corresponding shift in toxicity grade, if the Investigator feels it is appropriate. For example, if a patient has grade I asthenia at baseline which increases to grade II during treatment, this will be considered a shift of 1 grade and treated as a grade I toxicity for dose modification purposes however this is limited to changes up to a maximum of grade II. For toxicities which are considered by the Investigator unlikely to develop into serious or life threatening events (e.g. alopecia, altered taste etc.), treatment will be continued at the same dose without reduction or interruption. In addition, no dose reductions or interruptions will be required for anemia (non-hemolytic) as it can be satisfactorily managed by transfusions. If any grade I toxicity or any grade of alopecia occurs, treatment will be continued at the original dose without interruption. If in the opinion of the Investigator, a toxicity is considered due solely to one drug (e.g. hand-foot syndrome caused by capecitabine) the dose of the other drug does not require modification. Capecitabine treatment interruptions are regarded as lost treatment days and the planned treatment schedule should be maintained. Missed doses due to treatment interruptions should not be replaced (see Appendix I, Capecitabine schedule and interruptions to capecitabine schedule). If toxicity requires a dosing delay or interruption of both drugs of more than three weeks, the patient will be withdrawn from the chemotherapy part of the study for toxicity reasons. Where several toxicities with different grades or severity occur at the same time, the dose modifications applied should be the greatest reduction applicable. Once a dose has been reduced it should not be increased at a later time. Version / 152 May 21, 2014

209 If both docetaxel treatment delay or capecitabine treatment interruption is indicated, then the treatment should be delayed or interrupted. Table 7 Dose modification for Hematological Toxicities - Docetaxel-Capecitabine combination Toxicity ANC < 1.0x109/l on day of infusion (day 1 of cycle) Febrile Neutropenia or Neutrophil Counts 0.5x 109/L (grade 4) for more than 7 days. 2nd episode of Febrile Neutropenia or Neutrophil Counts 0.5x 109/L (grade 4) for more than 7 days. 3rd episode of Febrile Neutropenia or Neutrophil Counts 0.5x 109/L (grade 4) for more than 7 days. Platelets < 75x109/l on the day of treatment Platelets < 50x109/l on the day of treatment Platelet transfusion Treatment of toxicity Modification of chemotherapy None Wait until ANC > 1.0x109/l (maximum time up to 3 weeks), then use full dose. According to standard care in institute According to standard care in institute According to standard care in institute None None G-CSF or PEG-Filgrastim may be used in subsequent cycles. Wait until ANC > 1.0x109/l (maximum time up to 3 weeks), then use full dose. G-CSF or PEG-Filgrastim may be used in subsequent cycles. Reduce dose of docetaxel and capecitabine by 25% Discontinue chemotherapy treatment Wait until platelet count > 75x109/l and reduce dose of docetaxel and capecitabine by 25% Wait until platelet count > 75x109/l and reduce dose of docetaxel and capecitabine by 25% Wait until platelet count > 75x109/l and reduce dose of docetaxel and capecitabine by 25% Version / 152 May 21, 2014

210 Table 8 Dose modification of docetaxel-capecitabine for non-hematological toxicities Toxicity Grade Treatment of toxicity Change to chemotherapy Bilirubin I Wait until within normal limits, then change dose Alkaline Phosphatase III-IV Wait until Grade II, then change dose ASAT or ALAT III-IV Wait until Grade II, then change dose Alkaline Phosphatase and ASAT or ALAT Reduce docetaxel by 25% Reduce docetaxel by 25% Reduce docetaxel by 25% II Change dose Reduce docetaxel by 25% Edema Any No dose change; further treatment at physician discretion Neurologic II Wait until subsequent cycle; if I, no change III Arthralgia/Myalgia Any If there is no resolution Reduce docetaxel by 25% Symptomatic treatment Skin rash Any Symptomatic treatment If persisting after dose reduction, discontinue docetaxel Discontinue chemotherapy treatment Asthenia III Symptomatic treatment 1st occurrence- reduce docetaxel by 25% Nausea Any Local guidelines antiemetics and/or corticoid Vomiting Any Local guidelines antiemetics and/or corticoid Hand-foot syndrome II-III Stop capecitabine until grade I, then change dose 2nd occurrence reduce capecitabine by 25% No modification No modification 1st occurrence reduce capecitabine by 25% 2nd occurrence reduce capecitabine by a further 25% (to 50% of initial dose) 3rd occurrence - stop docetaxel and capecitabine Version / 152 May 21, 2014

211 Table 8 - Dose modification of docetaxel-capecitabine for Non-Hematological Toxicities Continued Toxicity Grade Treatment Change Diarrhea I None II-III IV Stop capecitabine until grade I, then change dose and treat diarrhea Stop capecitabine until grade I, then change dose and treat diarrhea Mucositis II Stop capecitabine until grade III-IV I, then change dose Stop capecitabine until grade 1, then change dose Dysphagia III Stop capecitabine until grade IV 1, then change dose Stop capecitabine until grade 1, then change dose Creatinine II No treatment; perform a sensitive test to ascertain the real value 1st occurrence reduce capecitabine by 25% 2nd occurrence reduce capecitabine by a further 25% (i.e. to 50% of initial dose) 3rd occurrence - stop chemotherapy treatment 1st occurrence reduce capecitabine and docetaxel by 25% 2nd occurrence stop chemotherapy treatment 1st occurrence reduce capecitabine by 25% 2nd occurrence reduce capecitabine by a further 25% (i.e. to 50% of initial dose) 3rd occurrence - stop chemotherapy treatment 1st occurrence reduce capecitabine and docetaxel by 25% 2nd occurrence stop chemotherapy treatment Reduce capecitabine by 25% Reduce docetaxel and capecitabine by 25% Stop capecitabine if Cr clearance 30 ml/min; if it increases to 30 ml/min, then resume it Other III Wait until I, then change dose Reduce capecitabine and docetaxel by 25% IV Stop treatment until grade I, then change dose 1st occurrence- 25% dose reduction 2nd occurrence stop chemotherapy treatment Version / 152 May 21, 2014

212 Table 9 Pill combinations to be taken, twice daily, to deliver the correct dose reductions per ranges of BSA based on a starting daily posology of 1650 mg/m2 75% Dose Level Number of tablets to be taken in the Surface Area (m2) Total Dose per Administration (mg) Morning Evening 150 mg 500 mg 150 mg 500 mg < % Dose Level Number of tablets to be taken in the Surface Area (m2) Total Dose per Administration (mg) Morning Evening 150 mg 500 mg 150 mg 500 mg < Recommendations for the management and treatment of delayed diarrhea The use of drugs with laxative properties should be avoided because of the potential for exacerbation of diarrhea. Patients should be advised to contact their physician to discuss any laxative use. Capecitabine can induce diarrhea, which can sometimes be severe. In patients receiving capecitabine monotherapy, the median time to first occurrence of grade II-IV diarrhea was 31 days, and median duration of grade III or IV diarrhea was 4.5 days. Patients with severe diarrhea should be carefully monitored and, if they become dehydrated, should be given fluid and electrolyte replacement. Prophylactic treatment No prophylaxis should be given, most of all no loperamide should be administered prophylactically. Loperamide and a fluoroquinolone have to be prescribed to the patients when they leave hospital. Adequate oral and written information about their use and about the management of diarrhea has to be properly given at this time, including the necessity of oral rehydration when diarrhea occurs. Patients should stop any laxative treatment and avoid food and beverages which might accelerate intestinal transit. Curative treatment If grade II, III or IV diarrhea occurs, administration of capecitabine should be immediately interrupted until the diarrhea resolves or decreases in intensity to grade I. Following grade II or higher diarrhea, subsequent doses of capecitabine should be decreased (see dose modification table for non-hematological toxicities table 8). As soon as the first liquid stool or abnormal bowel movement occurs, the patient must immediately start loperamide, two capsules, p.o., then one capsule, p.o. every two hours for at least 12 hours and up to 12 hours after the last liquid stool without exceeding a total treatment duration of 48 hours. Oral rehydration with large volumes of water and electrolytes is to be prescribed during the entire episode of diarrhea. Version / 152 May 21, 2014

213 If diarrhea persists for more than 48 hours despite the recommended loperamide treatment, the patient must be hospitalized for parenteral support. A 7-day prophylactic oral course of a fluoroquinolone antibiotic is to be started after medical advice. Loperamide will be replaced by another antidiarrheal treatment (e.g. Octreotide 0.1 mg s.c. x 2/d for 3 days (see (Ref. 97) for further information). Patients experiencing any grade IV diarrhea, or diarrhea with grade III-IV neutropenia, or with fever or with vomiting should be hospitalized quickly for parenteral hydration and any other required treatment after adapted work-up. Capecitabine cannot be re-started until diarrhea has resolved to grade 0 I with the last loperamide dose given at least 24 hours beforehand. Anaphylactoid type reactions and hypersensitivity reactions If a hypersensitivity reaction occurs despite premedication, it is very likely to occur within a few minutes of the start of the first or the second infusion of docetaxel. Therefore, during the 1st and the 2nd infusions, the infusion must be given drop by drop for the first 5 minutes, and a careful evaluation of general sense of well being and, whenever possible, blood pressure and heart rate monitoring must be performed so that immediate intervention can occur in response to symptoms of an untoward reaction. Facilities and equipment for resuscitation must be immediately available: antihistamine, corticosteroids, aminophylline, epinephrine. If a reaction occurs, the specific treatment that is medically indicated for a given symptom (e.g. epinephrine in case of anaphylactic shock, aminophylline in case of bronchospasm, etc.) must be instituted. In addition, the following measures are recommended (Table 10): Table 10 Recommendations for management of hypersensitivity reaction Mild symptoms: Localized cutaneous reaction, such as pruritus, flushing, rash Moderate symptoms: any symptom not listed above (mild symptoms) or below (severe symptoms), such as generalized pruritus, flushing, rash; dyspnea, hypotension with systolic B.P. > 80 mm Hg Severe symptoms: such as: bronchospasm, generalized urticaria, hypotension with systolic B.P. 80 mm Hg, angioedema - consider decreasing the rate of infusion until recovery of symptoms, stay at bedside - then, complete docetaxel infusion at the initial planned rate - At subsequent cycles, use the same premedication outlined in section stop docetaxel infusion. - give i.v. dexamethasone 10 mg (or equivalent) and i.v. diphehydramine 50 mg (or equivalent). - resume docetaxel infusion after recovery of symptoms. - At subsequent cycles, give i.v. dexamethasone 20 mg (or equivalent) and i.v. diphehydramine 50 mg (or equivalent) one hour before infusion, in addition to the premedication schedule. - Immediately stop infusion - Give i.v.diphenydramine 50 mg with or without dexamethasone and/or epinephrine as needed; monitor patient until resolution of symptoms - Resume docetaxel infusion after recovery of symptoms. - At subsequent cycles, give i.v. dexamethasone 20 mg (or equivalent) and i.v. diphehydramine 50 mg (or equivalent) one hour before infusion, in addition to the premedication schedule. Patients must be monitored carefully and preferably in an intensive care setting. Version / 152 May 21, 2014

214 Anaphylaxis (CTCAEv3 grade IV reaction) Fluid retention OFF CHEMOTHERAPY TREATMENT In case fluid retention occurs during treatment with docetaxel, the signs and symptoms should be graded according to CTC criteria. The weight will be recorded and followed as frequently as possible to document any weight gain, which could be related to edema. Recommended curative treatment for fluid retention Curative treatment should commence when signs and/or symptoms of fluid retention are observed, including weight gain from baseline grade I not otherwise explained and should be given according to standard care. The following treatment is recommended: Furosemide 20 mg p.o. once a day; potassium-sparing diuretics alone or in combination are also allowed. If the symptoms cannot be controlled adequately, i.e. worsening of the fluid retention or spread to another area, the dose of Furosemide should be increased to 40 mg. The clinical tolerance of the patient and the medical judgment of the Investigator will determine if it is in the patients best interest to continue or to discontinue the study chemotherapy drug. It is recommended, however, that patients with severe fluid retention should be withdrawn from chemotherapy. When it is unclear whether an effusion is disease-related or study-drug related, treatment should be continued until progressive disease is clearly documented, unless it becomes symptomatic Concomitant therapy, docetaxel-capecitabine arm The usual pattern of care is required during infusion, including careful attention to reactions and side effects. In addition, some guidelines regarding drug interactions should be observed: Capecitabine: Patients receiving concomitant capecitabine and oral coumarin-derivative anticoagulant therapy should have their anticoagulant response (INR or prothrombin time) monitored frequently in order to adjust the anticoagulant dose accordingly. Altered coagulation parameters and/or bleeding, including death, have been reported in patients taking capecitabine concomitantly with coumarinderivative anticoagulants such as warfarin and phenprocoumon. A PK interaction has been observed. The use of low molecular weight heparin instead of coumarin is at the discretion of the Investigator. Phenytoin Increased phenytoin plasma concentrations have been reported during concomitant use of capecitabine with phenytoin. Formal drug-drug interaction studies with phenytoin have not been conducted. Patients taking phenytoin concomitantly with capecitabine should be regularly monitored for increased phenytoin plasma concentrations and associated clinical symptoms. Allopurinol Interactions with allopurinol have been observed for 5-FU; with possible decreased efficacy of 5- FU. Concomitant use of allopurinol with capecitabine should be avoided. Antivirals and Antiprotozoals Capecitabine or 5-FU should not be administered together with the antiviral drug sorivudine or its chemically related analogues, such as brivudine. A clinically significant drug-drug interaction between sorivudine and 5-FU, resulting from the inhibition of DPD by sorivudine, has been described in the literature. This interaction, which leads to increased fluoropyrimidine toxicity, is potentially fatal. Version / 152 May 21, 2014

215 Metronidazole increased the toxicity of fluorouracil in patients with colorectal cancer, apparently by reducing the clearance of the antineoplastic. As it has been described in the literature, caution should be exercised. Gastrointestinal Drugs Pretreatment with cimetidine for 4 weeks led to increased plasma concentrations of fluorouracil following intravenous and oral administration in six patients. The effect was probably due to a combination of hepatic enzyme inhibition and reduced hepatic blood flow. No such effect was seen following single doses of cimetidine in five patients or pretreatment for just one week in six. Care is required in patients taking both drugs simultaneously. Hematopoietic growth factors Hematopoietic growth factors (i.e., G- or GM-CSF) may be used according to institutional guidelines to treat febrile neutropenia, but should not be used as primary prophylaxis. Use of any growth factor support must be documented in the patient record. Growth factors must be discontinued at least 48 hours prior to initiation of the next capecitabine cycle. Bisphosphonates Patients receiving bisphosphonates are eligible for this study but should have bone scans (and X- rays of areas of enhanced uptake indicative of bone metastasis) at baseline. Those patients starting bisphosphonates during the study but without other clear evidence of disease progression will not be diagnosed as having progressive disease on that evidence alone. Docetaxel In vitro studies have shown that the metabolism of docetaxel may be modified by the concomitant administration of compounds that induce, inhibit, or are metabolized by cytochrome P450 3A4, such as cyclosporine, terfenadine, ketoconazole, erythromycin, and troleandomycin. Caution should be exercised with these drugs when treating patients with docetaxel as there is a potential for a significant interaction. 6.3 Treatment withdrawal criteria The treatment should be withdrawn if: The patient, at any time, withdraws consent to participate. The Investigator judges that the decision is in the best interest of the patient The treatment must be interrupted for more than 3 weeks There is evidence of disease recurrence/relapse The patient becomes pregnant The patient is lost to follow-up The patient will then be considered off-protocol randomized chemotherapy treatment, however the patient remains eligible for R-E and should be followed according to the protocol. 7 Endocrine therapy The first dose of endocrine therapy should be administered within 4 weeks following the randomization (R-E) date. If treatment has not started within 300 days after definitive surgery, the patient becomes ineligible for randomized endocrine therapy. Version / 152 May 21, 2014

216 7.1 Trial treatments Tamoxifen will be taken according to the standard of care, consisting of a 20 mg tablet orally once daily for 2 years from date of endocrine therapy randomization (R-E) for all patients randomized to receive tamoxifen. Letrozole will be provided free of charge for treatment within the study protocol by Novartis. It is to be taken as a 2.5 mg tablet, orally once daily for 7 years from date of endocrine randomization (R- E) for all patients randomized to the letrozole arm or for 5 years after an initial 2 years of tamoxifen for patients randomized to the tamoxifen arm. 7.2 Side effects of drugs Tamoxifen: The most common side effects reported are hot flushes, nausea, vaginal discharge or abnormal vaginal bleeding. Hypercalcemia, bone pain, and retinopathy are rare. Long term beneficial effects on lipid and bone metabolism have been reported in post menopausal women, but there may be an increased risk of endometrial cancer, polyps and hyperplasia associated with the estrogen agonist action of tamoxifen. In premenopausal women, bone mineral density loss may occur. Fluid retention and skin rash have been reported. Tamoxifen is known to increase the risk of thromboembolic disease, and a short period of cessation of treatment may be required should some women need to undergo minor or major surgery. Modification of tamoxifen dosage is rarely indicated. No standard dosage modifications are prescribed. Letrozole: In clinical trials, adverse events (AEs) were generally mild to moderate and rarely severe enough to require discontinuation of treatment. AEs that have been reported include: fatigue, chest pain, peripheral edema (swelling), high blood pressure, nausea, vomiting, constipation, diarrhea, abdominal pain, anorexia, viral infection, musculoskeletal pain (including arm pain, back pain, leg pain, skeletal pain), arthralgia, headache, dyspnea, coughing, hot flushes, and rash. Less frequent (< 5%) AEs reported include hypercalcemia, fracture, depression, anxiety, pleural effusion, hair thinning, increased sweating and vertigo. Other possibly drug-related AEs were: vaginal bleeding, leukorrhea, weight loss, thrombophlebitis, generalized edema, dizziness, changes in lymphocyte counts and increase in aminotransferase. Special emphasis should be given to problems associated with lipid and bone metabolism such as fractures, osteoporosis and hypercholesterolemia. Modification of letrozole dosage is rarely indicated. No standard dosage modifications are prescribed. Patients in the Tamoxifen-Letrozole arm, who are under tamoxifen treatment and need to discontinue tamoxifen due to toxicity, can switch early to letrozole and complete the 7 years of treatment per protocol. Patients who discontinue the treatment are considered off randomized endocrine therapy and should be treated at the discretion of treating physician. All patients should be followed up according to study protocol. Missing doses due to treatment interruption should not be replaced. Treatment cannot be interrupted for more than 90 consecutive days (see treatment withdrawal criteria). 7.3 Ovarian function suppression All premenopausal women (until the age of 50 years) at diagnosis who decide to participate in the endocrine randomization in the trial must have adequate ovarian function suppression while or ablation for the whole duration of randomized endocrine therapy treatment in both arms. Menopausal status should be confirmed by dosing estradiol, FSH and LH, or E2 if appropriate, in the case of a GnRH or bilateral ovarian irradiation. Ovarian function suppression can be achieved in several ways: Version / 152 May 21, 2014

217 The use of a GnRH analogue such as goserelin (Zoladex TM), taken as a 3.6 mg subcutaneous injection every 28 days. NOTE that this product will NOT be supplied by the EORTC. GnRH analogues administered at 3 monthly intervals are not approved due to uncertainty concerning the duration of ovarian function suppression in women with this schedule. Bilateral surgical oophorectomy. Bilateral ovarian irradiation. Radiation should be given according to accepted guidelines. Biochemical verification of ovarian function suppression is required after 2 months using measurement of estradiol, FSH and LH. There are cases of failure of ovarian function suppression under GnRH analogues. In this case, ovarian function suppression should be achieved by other means, provided the patient accepts an alternative method. For women reaching the age of fifty while on GnRH there is the possibility, at the treating physician s discretion, of stopping the AI and the GnRH for at least three months and confirming menopausal status by dosing estradiol, FSH and LH. If menopausal status is confirmed endocrine treatment can be resumed with the AI alone. 7.4 Concomitant treatments Restriction of concomitant treatments: the following concomitant treatments are NOT ALLOWED during the endocrine therapy part of the trial: Any other anti-cancer therapy (anti-estrogens, aromatase inhibitors, chemotherapy) other than the primary endocrine treatment Other investigational agents whilst on study. Raloxifene or other SERMS Systemic HRT with or without progestins. A patient should go off trial if she receives more than 3 months of HRT. Hormonal contraceptives (including depot injections and implants) However intrauterine devices, including those progesterone coated ARE ALLOWED Any other hormonal treatments (either oral of transdermal) including estrogen, progesterone, androgens, and aromatase inhibitors. Topical estrogens are DISCOURAGED during the endocrine therapy part of the trial. However, for women with severe vaginal dryness/dyspareunia that does not respond to non-hormonal treatments, then a local vaginal treatment with minimal systemic absorption is allowed. Concomitant treatment with bisphosphonates IS ALLOWED (including on other clinical trial protocols) and should be recorded during the trial. This should be prescribed as recommended by the ASCO guidelines on the role of bisphosphonates and the use of AI (Ref. 98). The administration of vitamin D3 and calcium supplements is recommended. Considering the potential increased risk of osteoporosis in women in this study, patients should be advised about regular calcium intake and weight bearing exercise (see ASCO guidelines on bone care (Ref. 98)). Version / 152 May 21, 2014

218 7.5 Treatment withdrawal criteria The treatment should be withdrawn if: The patient, at any time, withdraws consent to participate The Investigator judges that the decision is in the best interest of the patient The treatment must be interrupted for more than 90 consecutive days There is evidence of disease recurrence/relapse The patient becomes pregnant The patient is lost to follow-up The patient will then be followed-up according to the protocol. 8 Clinical evaluation, laboratory tests and followup 8.1 Baseline evaluations Baseline evaluations that must be carried out prior to enrollment: Chest x-ray or Thoracic CT scan Bilateral mammography and within a clinically acceptable period before the start of treatment for women receiving either chemotherapy or endocrine therapy: Clinical evaluations (physical examination, medical history & weight). Any abnormal or existing clinical condition should be reported on the baseline AE case report form. WHO performance status Routine Hematology (WBC, absolute neutrophils, hemoglobin, platelets) Routine biochemistry CA 15-3 if standard of care in local institution For women receiving neither chemotherapy nor endocrine therapy the baseline analyses will be performed once and they will be followed as described in section Specifically to be done prior to chemotherapy randomization (R-C) To be performed within a clinically acceptable period before randomization R-C and before the start of chemotherapy treatment. (see the summary table 11 below) ECG (the pre-surgical ECG can be used provided it was normal) Cardiac function (left ventricular ejection fraction (LVEF) within the normal limits of each institution, measured either by echocardiography or by radionuclide angiocardiography (MUGA)) according to standard of care in the local institution. Version / 152 May 21, 2014

219 8.1.2 Specifically to be done prior to endocrine randomization (R-E) Patients should be fit for endocrine therapy, for patients who do not receive chemotherapy, baseline evaluations need only be done once prior to enrollment. For patients receiving adjuvant chemotherapy and going on to receive endocrine treatment in R-E, some baseline evaluations need to be repeated i.e. once at enrollment/r-c and then again for entry into R-E (details are given below and in summary table 11). To be performed within a clinically acceptable period before randomization R-E and before the start of endocrine therapy treatment. Repeated tests for those patients who received chemotherapy: Clinical examination (a thorough clinical history should be taken, especially history of previous thromboembolism, cardiovascular or cerebrovascular events) Routine hematology (WBC, absolute neutrophils, hemoglobin, platelets) Routine biochemistry (if standard of care in the institute), LH/FSH & E2 if appropriate (menopausal status should be determined on plasma estrogen levels) for women of childbearing potential. Baseline radiological evaluations should be repeated if clinically indicated, or in the case of liver and bone scan imaging, if biochemical blood tests are persistently abnormal after completion of chemotherapy. A gynecological examination is recommended at baseline in case of pelvic complaints. This may include a Pap smear, bimanual and recto-vaginal examination or vaginal ultrasound. A bone mineral density study (DEXA) should be performed at baseline for all women randomized in R-E if this is standard care in the local institution. Non-fasting cholesterol should be taken and recorded in the patients file if this is standard care in the local institution. 8.2 During treatment Chemotherapy The following evaluations will be performed on Day 1 (or within 3 days prior to the start) of each chemotherapy cycle. Pre-enrollment assessments will be used for Cycle 1. A brief physical examination, including vital signs, and weight. WHO performance status. Routine hematology (WBC, absolute neutrophils, hemoglobin, platelets) Routine biochemistry (if standard of care in the institute) An adverse event assessment will be performed according to the NCI common terminology criteria (CTCAE) version 3.0 (see Appendix C) Version / 152 May 21, 2014

220 8.2.2 Endocrine therapy Visits should be every 3 months for the first two years and then once a year (note: laboratory tests should be adapted to standard of care in local institutions if one of the tests recommended is not routine practice). First 2 years every 3 months physical examination and medical history. Routine hematology, liver function test and CA 15-3 if standard of care in the local institution. every 6 months non-fasting cholesterol if standard of care in the institute every 12 months mammography. Chest X ray, gynecological assessment, bone mineral density study (DEXA) if standard of care in the institute Years 3-7 every 12 months physical exam and medical history. Routine hematology & liver function if standard of care in the institute, mammography. CA 15-3 if standard of care in the local institution, chest X ray (at the discretion of the investigator), gynecological assessment (at the discretion of the investigator), Bone mineral density study (DEXA) and non-fasting cholesterol if standard of care in the institute After Year 7 Every 12 months physical examination, medical history, and mammography. Routine hematology, liver function tests and CA 15-3 if standard of care in the local institution, chest X ray (at the discretion of the investigator), gynecological assessment (at the discretion of the investigator), bone mineral density study (DEXA) and non-fasting cholesterol if standard of care in the institute At each visit Dispense medication Adverse Event and follow-up forms should be completed and submitted. Report bisphosphonate treatment for osteoporosis, medication for hypercholesterolemia, and development of cardiac or cerebrovascular events for each visit. Other diagnostic tests (i.e.: abdominal ultrasound and/or CT scan, bone scan, MRI) should be performed only in the presence of signs and/or symptoms and/or blood test abnormalities suggestive of cancer recurrence. 8.3 After the end of treatment (Follow-up) The following follow schedules apply to all patients except those patients who have progressive disease with distant metastasis who will then be followed yearly for survival. Version / 152 May 21, 2014

221 8.3.1 Follow-up for patients receiving neither chemotherapy nor endocrine therapy Follow-up for those patients who have received neither chemotherapy nor endocrine therapy will be done according to standard of care and consist of the following examinations to ideally be done every 3 months for the first 2 years then yearly for subsequent years. Clinical evaluations (physical examination, medical history & weight) WHO performance status Routine Hematology, if standard of care in the institute Routine blood and biochemistry, if standard of care in the institute Yearly mammography Other procedures will be done as clinically indicated Chemotherapy follow-up All patients will be followed as described in section After local recurrence patients will continue to be followed for recurrence at other sites For patients receiving chemotherapy and endocrine therapy Follow-up will be done according to standard care in the local institute and will follow the scheme for endocrine therapy follow-up as described in section & summary table For patients receiving only chemotherapy Follow-up visits will be done according to standard care in the local institute and ideally every 3 months during the first 2 years, and then every 12 months. The assessments will be done according to standard care in the local institute and should follow the schedule described in section and summary table Endocrine therapy follow-up After the first seven years, disease status and survival data will be collected yearly for all patients. Annual follow-up to report disease status and survival will be requested for at least a minimum of 15 years. Survival data will be collected for all patients who have progressed with distant metastasis during or after trial treatment until death. Version / 152 May 21, 2014

222 8.3.4 Summary table Table 11 - Schedule of Assessments Required Investigations Informed Consent Inclusion criteria Assess quality of tumor samples Demographics, medical history Pregnancy test 1 Physical examination/ Performance Status Routine hematology Routine Biochemistry LH/FSH & E2 if appropriate CA Non fasting total cholesterol3 Prior to enrollment/ randomization Every chemotherapy cycle End of treatment Followup, Years 1 and 2 Every 3 months Followup, Years 3 to 7 Yearly Optional Optional Chest X-ray ³ If clinically indicated Bone Scan ³ If clinically indicated Mammography Once a year DEXA scan Following standard care ECG 4 If clinically indicated Echo/MUGA scan Other Investigations (CT, PET scan, liver echo) Adverse Events Serious adverse events (SAE) 4 If clinically indicated As needed or clinically indicated Throughout the study Followup, Years 7 to 10 Yearly Version / 152 May 21, 2014

223 1 Only for patients of childbearing potential ² For all pre-menopausal women 3 If standard of care in the institute (thoracic CT allowed) 4 An ECG and Echo/MUGA scan should be performed prior to randomization in the chemotherapy randomization Legend to table All endocrine therapy baseline assessments must be performed after the end of chemotherapy and before starting endocrine treatment. Non-fasting total cholesterol should be collected according to standard care at local institutions and preferably at baseline, and yearly for the following 5 years. An additional sample should be collected one year after completion of the 7 year trial treatment period, at any time of premature trial treatment discontinuation and once more at the next scheduled 6-monthly visit following premature trial treatment discontinuation. If premature trial treatment discontinuation occurred within the 6 months prior to completion of the 7 year trial treatment period, the additional sample should be collected at the next scheduled yearly visit. Radiological assessments Bone mineral density studies should be performed according to standard care at local institutions and preferably at baseline and yearly whilst on AI treatment. A bilateral mammography must be taken within 3 months prior to surgery. A mammography (and/or ultrasound) of the conserved and contra lateral breast should be performed at yearly intervals or should be done according to national standards or hospital specific requirements. A Chest X-ray or Thoracic CT scan should be performed prior to enrollment (baseline evaluation) according to standard of care in the institution and if clinically indicated during follow-up. For chest X-ray posterior- anterior (PA) and lateral views are preferred however a PA alone will be accepted subject to local practice. A bone scan should be taken according to standard of care in the institute or if medically indicated. If the bone scan showed areas suspicious for tumor then these areas should be assessed by X-ray, CT or MRI. Metastatic disease should be confirmed by a second bone scan at least three months later that shows progressive changes. Abdominal ultrasound or liver scan or CT abdomen is required if medically indicated. It is mandatory in all patients with liver function tests or ALP that are above the limit of normal prior to randomization, after completion of any chemotherapy treatment. If after randomization, liver enzymes become abnormal, further staging is required to exclude metastatic disease (ie: Chest XR, liver imaging, bone scan, etc). Other procedures In the event of a pelvic complaint patients should have a gynecological examination because of increased risk of uterine cancer in patients receiving tamoxifen. Otherwise patients should be followed according to standard of care in the institute. Note: Any woman with abnormal/unexplained vaginal bleeding should have an endometrial biopsy and the results must be recorded on the Adverse Event Form during protocol therapy Note: Patients who discontinued trial treatment for any reason before the planned treatment duration of 7 years must continue to be followed every six months for the first two years for assessment of disease status, cardiovascular, skeletal, and urogenital events, and for survival data collection. After 3 years all patients will be followed yearly for disease status, cardiovascular, skeletal, and urogenital events and survival status for a minimum of 15 years. Version / 152 May 21, 2014

224 9 Criteria of evaluation 9.1 Time to event endpoints Special note: In this complex trial the date of enrollment/randomization used below in definitions of time to event endpoints (overall survival, disease free survival, distant metastasis free survival) depends upon the analysis and the dataset in which it is being analyzed. This is clarified in the statistical section Types of recurrence Recurrence will be categorized as local, regional or distant. Local (Breast) Recurrence is defined as recurrence within the breast. Local (Chest Wall) Recurrence is defined as recurrent cutaneous or subcutaneous tumor occurring in an area bounded superiorly by the clavicle, inferiorly by a horizontal line at the level of the xiphisternum, medially by the midline and laterally by the posterior axillary line. Regional (Nodal) Recurrence is defined as recurrent tumor in the lymph nodes in the homolateral axilla, homolateral supraclavicular fossa or ipsilateral internal mammary chain. Distant Metastatic Recurrence is defined as spread of disease beyond the limits specified above as local or regional recurrence. Contralateral breast cancer and secondary cancers are not recurrence, but will be considered as events for the endpoint of DFS, see section Distant Metastasis Free Survival (DMFS) For the endpoint of DMFS only distant metastatic recurrences as defined above and deaths (any cause) will be taken as events. Contralateral breast cancer and secondary cancers will not be taken into account. DMFS is calculated as the time from enrollment/randomization to either the first date of distant metastatic recurrence or the date of death. The date of first documented distant metastasis (if applicable) will be used as the event date. Patients alive with no evidence of metastasis at the time of their last visit are censored at the time of the last examination Disease Free Survival (DFS) Definition of date of disease recurrence: date on which a clinically suspicious lesion is first recorded in the patient file provided action is taken as result of which the diagnosis of any type of recurrence is confirmed. Recurrence of disease can be a loco-regional recurrence, a distant (metastatic) recurrence or a second primary. As documented in Section 9.2.1, contralateral breast cancer and second malignancies will be considered to be events. Disease Free Survival (DFS) is calculated as the time from enrollment/randomization to either the date of disease progression or the date of death. The date of first documented disease recurrence (if applicable) will be used as the date of event. Patients alive with no evidence of disease recurrence at the time of their last visit are censored at the time of the last examination Overall Survival (OS) Overall Survival (OS) is calculated as the time from enrollment/randomization to the date of death (any cause). Patients still alive at the time of analysis are censored at the last time they are known to be alive. Early death will be defined as death occurring within 30 days of last protocol treatment or within 90 days from enrollment/randomization. Version / 152 May 21, 2014

225 9.2 Reporting of results All patients enrolled in the study must be followed for progression and survival, according to intent to treat, even if there are major protocol or treatment deviations or if they are found to be ineligible. 9.3 Evaluation of toxicity Reporting of and general evaluation of adverse events All adverse events will be recorded on the applicable case report forms This study will use the Common Terminology Criteria for Adverse Events v3.0 (CTCAE). A copy of the CTCAE can be accessed from the CTEP home page ( A link to this page is provided on the EORTC web site Investigators who do not have access to Internet can contact the EORTC Headquarters to receive a copy by post. The actual laboratory values will be collected and grading according to CTCAE will be centrally applied for reporting Long-term chemotherapy toxicity The following potential long-term toxicities of chemotherapy will be monitored for a minimum of 15 years in all patients who were randomized for chemotherapy (R-Chemotherapy), cardiac toxicity and secondary leukemias Serious adverse events Serious adverse events are defined by the Good Clinical Practice Guideline. Serious adverse events should be immediately reported according to the procedure detailed in this PROTOCOL (see chapter on Reporting adverse events) 10 Statistical considerations 10.1 Statistical design Sample size In the set of patients who have a low risk gene prognosis signature using the 70-gene signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature prognosis and thus received no chemotherapy (i.e. they truly did not receive chemotherapy), a null hypothesis of a 5-year distant metastasis free survival (DMFS) of 92% will be tested. With 6,600 patients accrued overall, this set has an expected size of 672 patients. The primary test will be a one-sided test at 97.5% confidence level. This primary test will be performed when two conditions are met: the standard error of DMFS at 5 years (obtained by estimating the standard error of log(-log(s(t)), and back-transformation) is 0.01 or less and at least one third of the patients in the above dataset have been followed for five years. To perform the test, a 95% two-sided confidence interval will be constructed for the DMFS at 5 years, using the log-log transform formula (Ref. 99), page 18). The test will be significant if the confidence interval does not include the value of Using these conditions, this test has 80% power to reject the null hypothesis if the true 5-year DMFS is 95%. The primary test for R-C is to compare DFS between the two chemotherapy arms randomized. Due to insufficient numbers of patients randomized into this part of the study, there will be less than Version / 152 May 21, 2014

226 80% power to detect the original alternative hypothesis of hazard ratio of 0.76 (for example a control 5-year DFS of 86% versus a 89.2% experimental 5-year DFS). The comparisons for R-C will be performed at the time of the primary test (as described above) at 5% alpha and an a posteriori power calculation will be performed. The primary test for R-E is to compare DFS between the two sequences of endocrine therapy randomized. Due to insufficient numbers of patients being randomized into this question, there will be less than 80% power to detect the original alternative hypothesis of hazard ratio of 0.75 (for example a DFS rate at 5 years of 86% on the control arm versus an experimental 5-year DFS of 89.3%). The primary analysis of the endocrine comparison will be performed when 379 events have been observed among the patients randomized for the endocrine question, yielding 80% power to detect a hazard ratio of This is projected to occur at a median follow up of 7.9 years Randomization and stratifications For eligible patients with adequate material for microarray analysis, both the 70-gene signature and clinical-pathological risk will be assessed, after which they can be enrolled. Patients will be centrally enrolled and randomized as applicable for each of the three possible randomizations (R-T, R-C, R-E, for practical details, see chapter on enrollment / randomization procedure). Patients are evaluated as low clinical-pathological risk, if their 10-year disease specific survival (without chemotherapy or endocrine therapy) is estimated by Adjuvant! Online (Ref. 101) as greater than 88% for ER-positive patients, and greater than 92% for ER-negative patients. Patients who are at high risk according to both methods will receive chemotherapy, and patients who are at low risk according to both methods will not receive chemotherapy. As new versions of Adjuvant! Online (Standard version) become available, these will be implemented. The version at the time of the writing of this protocol is Standard Version 8.0. R-T: Patients for whom both methods are discordant will be randomized between chemotherapydecision-making according to clinical criteria and chemotherapy-decision-making according to 70- gene signature prognosis. For R-T, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/low risk clinical vs. low risk genomic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), nodal involvement (yes, no), age (<50 vs. >=50), HER-2 (Positive vs. Negative vs. Unknown), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). In case PgR is unknown, the patient will be stratified to the HR negative group if ER is negative, and to the HR positive group if ER is positive. Patients who are candidates for chemotherapy (see also treatment section) will be proposed for R-C: randomize to anthracycline based chemotherapy or taxane/capecitabine chemotherapy. For R-C, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/low risk clinical vs. low risk genomic/high risk clinical vs. high risk genomic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), age (<50 vs. >=50), HER-2 (Positive vs. Negative vs. Unknown (at the time of R-C)), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). In case PgR is unknown, the patient will be stratified to the HR negative group if ER is negative, and to the HR positive group if ER is positive. All patients who are candidates for endocrine therapy will be offered randomization (R-E) to 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. For R-E, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/high risk clinical vs. high risk genomic/low risk clinical vs. low risk genomic/high risk clinical vs. low risk genomic/low risk clinical), chemotherapy (no chemotherapy, chemotherapy without R-C, R-C arm A, R-C arm B), type of endocrine sensitivity (both ER and PgR positive vs. either ER or PgR positive), age (<50 vs. >=50), HER-2/neu (Positive vs. Negative vs. Unknown (at Version / 152 May 21, 2014

227 the time of R)), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). The three randomizations will be open-label Statistical analysis plan Primary and secondary endpoints In this complex protocol, time to event endpoints (DMFS, OS, and DFS) will start at different dates depending on the analysis: For all overall analyses, and analyses related to R-T, time to event endpoints will start at the date of enrollment of the patient. These are the analyses on the data sets AP, PT, ITT1 (see section ). For all analyses related to R-C, time to event endpoints will start at the date of R-C. These are the analyses on the data set ITT2 (see section ). For all analyses related to R-E, time to event endpoints will start at the date of R-E. These are the analyses on the data set ITT3 (see section ). Using these conventions for starting dates, the definitions in section 9.2 will apply Analysis populations In this study, the following analysis datasets will be used. Because of the complexity of the study, there are many datasets, and so a code is added between brackets for ease of reference. All patients dataset (AP): All patients who were enrolled into the study. Primary test dataset (PT): The set of patients who have a low risk gene prognosis signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy and did not deviate from this: no change in risk status post enrollment and no chemotherapy received. This dataset is used for the primary test. Primary test dataset as enrolled (PTE): The set of patients who have a low risk gene prognosis signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy, regardless of compliance to the no chemotherapy assignment. Intention-to-treat population for R-T (ITT1): All randomized patients for R-T will be analyzed in the arm they were allocated by randomization. Per protocol population for R-T (PP1): All patients who are eligible and randomized for R-T and have started their allocated treatment (at least one dose of chemotherapy if so randomized, respectively no chemotherapy). Intention-to-treat population for R-C (ITT2): All randomized patients for R-C will be analyzed in the arm they were allocated by randomization. Intention-to-treat population for R-E (ITT3): All randomized patients for R-E will be analyzed in the arm they were allocated by randomization. Safety population for R-C (SP2): All patients who were randomized for R-C and who have started their allocated chemotherapy (at least one dose of the study drug(s)). Safety population for R-E (SP3): All patients who were randomized for R-E and who have started their allocated endocrine treatment (at least one dose of the study drug(s)). A patient will be considered to be eligible if she did not have any deviations from the patient entry criteria listed in chapter 3 of the protocol. Potential eligibility problems will be assessed by the Version / 152 May 21, 2014

228 Clinical Research Physician (CRP) at time of medical review. If needed the CRP can ask the help of the Study Coordinators Timing of analyses The final analysis on the ITT2, SP2 datasets will be performed at the time of the primary analysis on the PT dataset. All analyses on the AP, PT, PTE, ITT1, PP1 datasets will be performed when the two conditions in section have been met. The final analysis on the ITT3, SP3 datasets will be performed when 379 events have been observed on the ITT3 dataset. This is projected to occur at a median follow up of 7.9 years Statistical methods Full analysis plan The outline of analysis given in this section of the protocol will be further detailed in a separate analysis plan that will be finalized prior to any analyses being performed. General All statistical tests and confidence intervals will be two-sided. All statistical tests will be tests for superiority (difference). For tests of primary endpoints, the nominal alpha level will be as defined by the interim analysis procedure (see Section for an overview table). All secondary endpoints will have a 95% confidence level at the final analysis. All comparisons for time to event endpoints described below will be adjusted Logrank tests, as performed by estimating a Cox model with the effect under consideration, as well as the stratification factors used for the applicable randomization (with the exception of site). As there is no a priori reason to believe that the stratification factors used will exhibit non-proportional hazards, preference is given to adjusting for them by using them as covariates (not by stratified Cox model). Corresponding hazard ratios and confidence intervals will be calculated by the same Cox regression. Time to event comparisons will be accompanied by Kaplan-Meier curves. Baseline characteristics will include age, menopausal status, ER/PgR status, HER-2 status, nodal involvement, tumor size, differentiation, WHO performance status, eligibility, method of axillary evaluation (sentinel only, dissection), type of surgery (mastectomy, quadrantectomy/tumorectomy). Analyses on AP dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable), once split by clinical pathological risk, and once by 70-gene signature risk. A separate table will be made to describe the types of chemotherapy and/or endocrine therapy, as well as radiotherapy administration (yes/no) patients received during the course of the trial, to show treatment of those patients who did not consent to or were not eligible for R-C and/or R-E, but still were given some treatment. This table will be on the AP dataset, with subcategories according to R- C (arm 1, arm 2, not done) and R-E (arm 1, arm 2, not done). McNemar s test will be applied to the 2x2 table of clinical-pathological (high/low) by 70-gene signature (high/low) risk to test whether there is a difference in assignment percentages to chemotherapy. Kaplan-Meier curves for OS, DFS and DMFS will be calculated for the following six subgroups: high risk for both risk assessments, low risk for both risk assessments, Version / 152 May 21, 2014

229 high risk for clinical-pathological criteria and low risk for 70-gene signature and randomized (R-T) to use clinical pathological assessment of risk high risk for clinical-pathological criteria and low risk for 70-gene signature and randomized (R-T) to use 70-gene signature assessment of risk low risk for clinical-pathological criteria and high risk for 70-gene signature and randomized (R-T) to use clinical pathological assessment of risk low risk for clinical-pathological criteria and high risk for 70-gene signature and randomized (R-T) to use 70-gene signature assessment of risk These six curves will be put on the same plot, and their qualitative interpretation will be of great importance for the evaluation of the two assessment methods. However no formal statistical testing will be performed on this plot other than those described for the ITT1 dataset. Analyses on PT dataset The analysis of DMFS at 5 years on the PT dataset is described in the sample size section. Analyses on ITT1 dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable) in the ITT1 dataset, split by R-T. In this dataset special attention will be given to the compliance to chemotherapy administration as assigned by R-T. In the subgroup of clinical-pathological high risk and 70-gene signature low risk, DMFS, OS and DFS will be compared. As these comparisons are low-powered, primary attention should go to the confidence intervals. In the subgroup of clinical-pathological low risk and 70-gene signature high risk, DMFS, OS and DFS will be compared. Again, as these comparisons are low-powered, primary attention should go to the confidence intervals. The interaction between the effect of chemotherapy and the subgroup (clinical-pathological high risk and 70-gene signature low risk versus clinical-pathological low risk and 70-gene signature high risk) will also be tested. Just as for the subset tests, the interaction test will be low-powered, so it will be informative only if there is a large difference in the effect of chemotherapy between the subsets, a situation that is unlikely to occur. Analyses on ITT2 dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable) in the ITT2 dataset, split by R-C. The primary comparison of R-C will be for DFS. Secondary comparisons are for DMFS and OS. For these three time-to-event endpoints, the analyses described above (under general ) will be performed. Analyses on SP2 dataset Chemotherapy administration will be summarized by number of cycles, dose intensity of the respective drugs, dose reductions, and cycle delays. Adverse events and laboratory abnormalities during chemotherapy will be tabulated (worst CTC grade per patient) by treatment arm. No formal toxicity analyses with p values will be carried out. Separate tables and listings will be made for late toxicity ascribed to chemotherapy (cardiac toxicity, secondary leukemia). Version / 152 May 21, 2014

230 Analyses on ITT3 dataset Baseline characteristics will be summarized (either by incidence table, or by summary statistics, as applicable) in the ITT3 dataset, split by R-E. The timing of this randomization, relative to initial enrollment into the study will be summarized The primary comparison of R-E will be for DFS. Secondary comparisons are for DMFS and OS. For these three time-to-event endpoints, the analyses described above (under general ) will be performed. Analyses on SP3 dataset Duration of endocrine therapy will be described by R-E-arm by Kaplan-Meier curves. A table will specify numbers of patients who started endocrine treatment, numbers who discontinued, reasons for discontinuation and the number of patients who switched from tamoxifen to letrozole (in the appropriate arm). The timing of the switch to letrozole will be summarized. Adverse events and laboratory abnormalities during endocrine therapy will be tabulated (worst CTC grade per patient) by treatment arm. No formal toxicity analyses with p values will be carried out Pre-planned sensitivity or exploratory analyses As discussed in the section on randomization and stratifications (ref ), the clinicalpathological risk evaluation will be modified as new (Standard) versions of Adjuvant! Online become available. A table will be constructed (on the AP dataset) to reflect the impact of these changes, showing how the distribution of risk would have been if the latest version (as at time of analysis) had been applied to all patients. If discrepancies exceed 10% of all patients, sensitivity analyses will be performed to assess the study results under the assumptions that all patients had been evaluated as per the latest version of Adjuvant! Online. To do this, all patients who were categorized differently during the trial will be removed from the respective datasets, and the three primary efficacy tests (respectively on the PT, ITT2 and ITT3 datasets) will be repeated. This sensitivity analysis is not a priory unbiased, because it removes borderline cases, but may give reassurance about the effect of updates of Adjuvant! Online. On the PP1 dataset: if there are more than 10% ineligible patients in the ITT1 dataset, the analyses discussed under PT and ITT1 will be repeated as sensitivity analyses on the PP1 dataset (and its corresponding subset for PT). If a large effect of the type of chemotherapy (R-C) is observed, further subgroup analyses of the analyses on the ITT1 dataset may be performed by type of chemotherapy, to generate confidence intervals of effects per chemotherapy type. Sensitivity analyses will be performed for overall survival analyses on the AP, PT, ITT1, ITT2, ITT3 datasets, by taking only cancer related deaths into account. These analyses will use competing risk methods to describe cumulative incidence of cancer related deaths. In this analysis deaths due to treatment toxicity or other treatment related deaths will be interpreted as cancer related. Similarly, sensitivity analyses will be performed for DMFS analyses on the AP, PT, ITT1, ITT2, ITT3 datasets, using time to distant metastasis, by taking only metastasis as an event, and not deaths. These analyses will also use competing risk methods to describe cumulative incidence of distant metastasis Prognostic factor analyses Prognosis is a key element of the trial. However, evaluation of prognostic factors is confounded by chemotherapy administration (yes, no), because key prognostic factors were used to decide on chemotherapy. Therefore, chemotherapy will be included in all models below, in an attempt to adjust for the effect. If the chemotherapy comparison on ITT2 is significant, the variable Version / 152 May 21, 2014

231 will have three levels (no, chemo arm A, chemo arm B). Endocrine therapy will not start at the time of entry into the trial, and will not be taken into account as a covariate. The effect of factors defining endocrine therapy will thus include endocrine therapy use. On the AP dataset, the three time-to-event endpoints (DMFS, OS, DFS) will be subjected to a prognostic factor analysis, using the following conventions: For each variable a first analysis will be performed using a Cox proportional hazards model with the variable, and chemotherapy as covariates. Variables with p<0.10 will be selected for potential inclusion into the final model. If there is significant (p<0.05) interaction with chemotherapy, the interaction will be included. If interaction is included, interaction and primary effect should be moved in or out of the model jointly. All selected variables will be put into a multivariate Cox proportional hazards model, together with chemotherapy, and the final model will be built using backward selection (until all p<0.05), while keeping chemotherapy in the model. The final model will be further evaluated by performing a confirmatory forward selection technique. Variables will be included or excluded with their full categorization, so no regrouping or selection of categories is allowed. The focus of this analysis will be on determining the extent to which 70-gene signature risk assessment may or may not replace other risk factors. After determining a final model, it will be evaluated for calibration and discrimination ability, using bootstrapping. The stability of the selected covariates in the final model will also be evaluated by the bootstrap (Ref. 102). The variables to be included, and their categories, are: Age (<40, 40-49, 50-59, 60) Baseline WHO performance status (WHO 0, WHO >0) Type of surgery (breast conserving surgery, radical mastectomy) Type of node evaluation (SNB, full axillary clearance) Tumor size (0-1 cm, >1-2 cm, >2-3 cm, >3-4 cm, >4-5 cm) Differentiation (grade I well differentiated, grade II moderately differentiated, grade III poorly differentiated, unknown) Nodal involvement (yes, no) HER-2 status (positive, negative, unknown) ER-PgR status (Positive (ER and/or PgR), Negative (both)) Clinical-pathological risk assessment (low, high) 70-gene signature risk assessment (low, high) Additional prognostic and predictive analyses are very likely, either to develop additional or improve existing prognostic gene signatures on the basis of microarray data for 6,600 patients, or to develop or validate predictive signatures for endocrine therapy or chemotherapy response. The strategy to perform these will be described in ad-hoc research protocols. Prognostic factor analysis For prognostic factor analysis, the central pathology results will be used. Central pathology review will repeat immunohistochemistry for estrogen and progesterone receptors to give standardized levels of expression which will be expressed as a percentage of immunoreactive cells. Version / 152 May 21, 2014

232 Categories used will be: positive: ER and/or PgR > 10% low positive (not positive) ER and/or PgR 1-10% negative: both ER and PgR = 0% There are no cut offs in positive i.e. when the expression is above 10% there are no further stratification levels. Further analysis will look into the potential further splitting of the positive category and determining whether expression levels (within positive ) may be an indicator of disease severity Data recoding and display Frequency tables will be tabulated (by treatment group or otherwise) for all categorical variables by the levels of the variables as they appear on the CRF (with %). Categories with a text field specification will be tabulated as categories and then supplemented by a listing with the following information for the patients fulfilling the condition for the specification (patient id, institution, treatment group, value of the item and text field contents). Dates relating to events prior to entry will be presented as the delay in days (or weeks, months, or years) between the past event and the date of entry (date of randomization date of past event + 1) and presented using the median and range. For example, on the randomization checklist, the date of last administration of prior treatment (or the date of first diagnosis of the cancer) will be presented as the time elapsed (in days, weeks, months or years, as appropriate) since the day of the last administration and the date of entry on study (date of randomization last administration/diagnosis +1). Other delays (e.g. re-treatment delays) are presented as continuous variables using the median and range. Safety and toxicity counts will be tabulated as worst grade per patient, within CTC term, or laboratory test. Continuous variables for which a coding system exists (such as for laboratory data) will be recoded into categories (for adverse events, the grading scale specified in the protocol will be used). Whenever no specific scale exists, lab data will be categorized based on the normal range: for example, below the lower normal limit (when appropriate), within the normal range, above the upper normal limit (UNL) and the degree to which it is above the UNL (for example > 2.5 x UNL, > 5 x UNL, > 10 x UNL). For laboratory data, the nadir is generally displayed. The nadir in a given cycle is the lowest laboratory value in that cycle; the overall nadir for a patient is the lowest laboratory value among all cycles. Other continuous variables (for example age, dose ) are presented using the median and range (minimum, maximum). If appropriate, continuous data may also be presented in categories (for example, age may also be grouped in decades) End of study End of study occurs when all of the following criteria have been satisfied: 1. Thirty days after all patients have stopped protocol treatment 2. The trial is mature for the analyses of the three primary endpoints (for R-T, R-C and R-E) as defined in the protocol 3. The database has been fully cleaned and frozen for these analyses Version / 152 May 21, 2014

233 10.4 Stopping rules and interim analyses Pilot study of the trial The trial, a multi-center, prospective, phase III randomized trial that will include approximately 6,600 patients, will start as a randomized study of 800 women, called pilot study, which should ensure the following: The logistics of the trial, as described in chapter 20, prove to be feasible from the point of view of patients, physicians and laboratories involved; The patient population recruited upfront for the trial is not biased : clinicians are entering in the trial a reasonable spectrum of node-negative and 1-3 positive node disease and not just very low risk patients. This will be observed from the actual distribution of high risk vs. low risk patients as compared to expected numbers; Clinicians comply with the protocol in the 70-gene signature /genomic arm of the study: in other words, a good 70-gene signature prognosis signature combined with bad clinicalpathological characteristics leads to a no chemotherapy decision if the patient is randomized to the genomic arm (R-T) and a poor gene prognosis signature combined with good clinical-pathological characteristics leads to a chemotherapy decision if the patient is randomized to the genomic arm (R-T); Among all patients who are candidates for adjuvant chemotherapy (see section 4.3), there are at least 66% who are undergoing the second randomization (R-C); A statistically significant difference is observed between the percentages of patients that have a high clinical-pathological risk (pc) and a high genomic risk (pg) (thus reflecting a likewise reduction in chemotherapy administration). With 800 patients in the pilot study and the following assumptions: the null hypothesis is H0: pc - pg = 0% the type I error rate is set to 5% then the pilot study has 80% power to reject the null hypothesis of no difference if the true difference is at least 7%. Monitoring of pc and pg (as well as of other trial data, especially crossovers between the chemotherapy and the no chemotherapy groups) should be the responsibility of an Independent Data Monitoring Committee Stopping rule after 800 patients enrolled If the data contradict the assumption of less patients receiving chemotherapy in the genomic arm in the first cohort of node-negative, breast cancer patients (N=800) then the prospective, randomized study will be put on hold until the reason for the deviation has been resolved; the same remark applies in case major issues regarding the logistics of the trial are identified Stopping rule to be applied every year In this study, the group of patients who have a low risk genomic prognosis signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy, are potentially at risk of being under treated. In analogy to the primary test of the study, a rule to be used in this particular subset of patients, to trigger review by the EORTC IDMC of the available efficacy data will be as follows: if the lower one-sided 95% confidence limit of the (exponential) hazard of distant metastasis or death exceeds /year (corresponding to a 5% event rate for DMFS at 5 years). This is equivalent to an unadjusted test at 95% confidence level rejecting a hypothesis of a 95% 5-year DMFS. This test will be performed every year. The test is not the same as the primary test, because it uses a different null hypothesis, and because it will rely on early data. Thus it assumes a constant hazard. As the test does not correct for multiplicity, it should be used as a trigger for review. Version / 152 May 21, 2014

234 The EORTC IDMC will review the relapse data in case the rule described above triggers a review. 11 Data monitoring A Data and Safety Monitoring Board (DSMB) will monitor the recruitment, the reported adverse events and the data quality at least twice a year. Problems which are identified will be discussed with the Study Coordinator who will take appropriate measures. Relevant information (including relevant safety data) will be included in the study status reports which serve as a basis of discussion during EORTC Group meetings. These reports will be made available to investigators participating in the study and to the EORTC Independent Data Monitoring Committee (IDMC) if interim analyses (planned or not planned) are carried out. If interim analyses are carried out, the interim monitoring of efficacy and safety data will be performed according to the Statistical Considerations chapter and the EORTC policy on Independent Data Monitoring Committees and Interim Analyses. The results of the interim analyses are confidential and are discussed by the EORTC IDMC. The IDMC will subsequently recommend to the EORTC Group whether any changes should be made to the study. No efficacy results will be presented at EORTC Group meetings or elsewhere before the trial is closed to recruitment and the data are mature for the analysis of the primary endpoint, unless recommended otherwise by the EORTC IDMC. 12 Translational research 12.1 Sample collection The following material should be obtained from each patient: 1(2) punch biopsy sample(s) from the breast cancer tissue for microarray analysis, biological materials bank storage and proteomics analysis. A representative paraffin tissue block from the breast cancer tissue for central histopathology review and for the production of tissue microarrays (TMAs). 1 optional blood sample (for whole blood and serum storage) for genetics and proteomics analyses and biological materials bank storage for future research to be taken during the study (preferably before surgery). For details of sampling see chapter 20, sample schedule and trial logistics. Objectives: Samples will be collected to perform genomic prognosis (by microarray analysis) and central pathology review. will establish a central TRANSBIG biological materials bank for research described in the protocol and future research. The research will be evaluated and agreed upon by the appropriate research/transbig committees Central pathology review Central pathology review will assess tumor grade and identify cases with discordant diagnosis between local and central pathology review. It will also generate feed-back information for local pathologists following confirmation of the local diagnoses. Since several studies have observed a high level of discordance between the analysis of histopathology (e.g. levels of hormone receptor and HER-2 expression) in peripheral pathology Version / 152 May 21, 2014

235 laboratories and central more experienced laboratories, a central review of the main standard histopathology will be performed. This will also enable the assessment of the prognostic value of level of expression of ER and PgR rather than the current negative, low positive, positive scaling. For these purposes a representative diagnostic paraffin tissue block of each tumor will be sent from each participating center to IEO (Instituto Europeo di Oncologia, Milan, Italy), attention of Prof Guiseppe Viale. The sending of these paraffin blocks to Milan will be at least once every 6 months. Tissue microarray production To evaluate hundreds of tumors in a high-throughput manner, tissue microarrays (TMA) will be made. The TMAs remaining after central histopathology analysis will be stored in the TRANSBIG biological materials bank and made available for research. The individual paraffin blocks will be returned to the participating centers after the tissue microarray of each sample has been made. Objectives: Central review of the main standard pathology in order to determine cases where local and central pathology are discordant and to generate feedback information for the local pathologist. To prepare a database of histopathology of all of the samples. To perform immunostaining for ER, PgR, Ki-67 and HER-2. HER-2 status will be confirmed by FISH. To determine the clinical relevance of molecular markers. To validate the clinical significance of clusters of genes stemming from gene expression profile. To establish a permanent tissue microarrays (TMA) tumor bank without the need for retaining the original donor paraffin blocks. Methods Standard histopathology and immunhistochemistry techniques will be used in combination with TMA technology. For TMA construction, two sections from each paraffin tissue block will be stained with HE to determine representative tumor regions. Tissue cores will be punched from the primary paraffin tumor blocks (the donor blocks) and subsequently brought into one empty recipient block. Numerous sections can be cut from the array blocks. Sections from these array blocks will be transferred to glass slides Proteomics Proteomics can provide insight in the post-transcriptional gene function and elucidate post translational pathway not obtainable by microarrays. This research area can make the connection between genes and biological function. Therefore, one of the side studies of the trial comprise of proteomics research. The initial proteomics studies will be performed as agreed by the TRANSBIG and Steering Committees. Therefore, at the end of accrual (or sooner if required) serum samples will be shipped from each participating centre (on dry ice) to the TRANSBIG biological materials bank in Brussels. From there 0.5 ml aliquots will be transferred to the research laboratories for analysis. Objectives To standardize methods for the extraction of proteins and preparation for analysis from sera and tissue from breast cancer patients. To identify new biomarkers (tumor specific proteins and peptides) in tissue and sera allowing early detection of breast cancer. To discover clinically relevant protein profiles from sera and tissue from breast cancer patients. Version / 152 May 21, 2014

236 To identify novel prognostic markers for breast cancer. To identify novel predictive markers for breast cancer. To correlate the expression of these markers with tissue histopathology and with clinical outcome. To link gene expression profiles to biological function. To identify tumor specific proteins or peptides which might be used as new pharmacological targets. Methods The primary analytical tools used for the proteomics will be SELDI, MALDI-TOF mass spectrometry, immunochemistry and high throughput 2D gel electrophoreses Complex microarrays In addition to the 70-gene signature, the complex microarrays used will yield information on gene expression of genes (i.e., oligonucleotides that represent thousands of genes on a microarray). For these complex microarrays, RNA isolated from the fresh frozen punch biopsy samples will be used. These complex microarrays will be performed at Agendia, Amsterdam, The Netherlands. Objectives will measure RNA expression levels using microarray technology and determine whether expression signatures can give prognostic information (i.e., 70-gene signature). To determine the overall gene expression pattern on the genomic array in order to identify new prognostic gene expression profiles. To determine the overall gene expression pattern on the genomic array in order to identify new predictive gene expression profiles. To validate previously discovered gene profiles. Methods Briefly, one section will be stained with HE to determine the percentage tumor cells in the frozen biopsy sample. When the tumor cell percentage is sufficient (> 30%), RNA will be isolated from the sample. Then the isolated RNA is amplified and labeled. The labeled sample RNA and labeled control oligonucleotides are combined and placed in a hybridization chamber in contact with the complex array. After hybridization slides were washed and scanned using a confocal laser scanner. Fluorescence intensities on scanned images were corrected for background noise and normalized. For detailed methods see (Ref. 32) Biological materials bank storage Objectives To establish an early breast cancer biological materials bank with fresh frozen tumor tissue samples, RNA, whole blood, serum and tissue microarrays. To establish a resource for future research. RNA will be isolated from the frozen tumor biopsy samples to perform the complex microarray. Remaining RNA will be transferred to and stored in the TRANSBIG biological materials bank. When the microarray analysis is performed successfully, the remaining frozen biopsy tissue will be transferred to the TRANSBIG biological materials bank for use in future research and proteomics analysis. These RNA and tissue samples will be shipped on dry ice from Agendia to the Version / 152 May 21, 2014

237 TRANSBIG biological materials bank every 6 months. The whole blood and serum samples will be shipped to the TRANSBIG biological materials bank at the end of accrual, or sooner if required. The biological material will be stored under the guardianship of the TRANSBIG consortium. The biological materials bank will be in Brussels Belgium in a facility run by World Courier. The rules and priorities for access to the biological material are decided on scientific merit of the proposed projects and set out in a separate document. 13 Publication policy It is the EORTC s policy not to release trial results before data maturity has been reached for the primary endpoint(s) of the trial unless the publication is authorized by an Independent Data Monitoring Committee. If the group wishes to publish or present study data before the publication of the primary trial endpoint, this may be authorized under the conditions specified in the EORTC Policy 009 Release of Results and Authorship Policy available from or authorized by an Independent Data Monitoring Committee and with the approval of the TRANSBIG Steering Committee. The final publication of the trial results will be written by the Study Coordinators from the EORTC Breast Cancer Group on the basis of the final analysis performed at the EORTC Heasquarters and then approved by the EORTC Headquarters. The manuscript will then be approved by all coauthors prior to being circulated to the steering committees of the EORTC Breast Group and TRANSBIG, in parallel, for comments and approval. Authorship on publications and presentations shall be based on academic standards and customs (i.e. those investigators who have included a substantial number of patients or who have contributed considerably to the development, conduct and analysis of the trial). All public presentations or publications shall also give appropriate acknowledgement to the EORTC and TRANSBIG consortium, including various sources of funding received, such as support from the European Commission Framework Programme VI (FP6-LSHC- CT ) and industrial partners. After revision by the EORTC Headquarters and other coauthors (and the industrial partners) the manuscript will be sent to a major scientific journal. All abstracts or manuscripts as well as detailed descriptions of any planned oral presentations must be submitted to the EORTC and to the and TRANSBIG Steering Committees for approval and transbig@bordet.be) at the earliest practicable time. For publication of any manuscript or any presentation or other public disclosure must be at least within four weeks prior to any proposed submission. Abstracts must be sent at least within ten days before submission. Research projects must follow the application policy approved by the TRANSBIG and Steering Committees, information available via the same addresses. The EORTC Group Chairman, Study Coordinators, EORTC Headquarters Team, the TRANSBIG Steering Committee or any of the industrial partners may comment upon, but may not change, the conclusions and content of any such publication or presentation. Authors are entitled to withhold any proprietary confidential information from publications / presentations. The EORTC, the TRANSBIG Steering Committee or any of the industrial partners may also, under exceptional circumstances, object to or request a delay before publication / presentation. In such cases, the EORTC, the TRANSBIG Steering Committee or any of the industrial partners must provide in writing its objections and recommendations within ten days for abstracts and within twenty days for all other publications / presentations. To allow for protection, after any objection, planned publications / presentations may be suspended until the end of a consultation period and up to a maximum of sixty days. This is applicable to any individual patient enrolled/randomized in the trial, or any subgroup of these. Such publications must comply with the terms specified in the EORTC Policy 009 Release of Results and Publication Policy, the Consortium Agreement and with Contract Agreements with the companies involved. Therefore, such a publication cannot include any comparisons between any Version / 152 May 21, 2014

238 of the randomized treatment arms or an analysis of any of the study endpoints unless the final results of the trial have already been published. 14 Investigator authorization procedure Investigators will be authorized to register for screening, enroll, or randomize patients in this trial only when they have returned to their National Coordinating Center/Group (NCC/NCG) or the EORTC: The updated signed and dated Curriculum Vitae of the Principle Investigator. The (updated) list of the normal ranges, in their own institution, of all laboratory data required by the protocol, preferably signed and dated by the head of the laboratory. A commitment statement / study acknowledgment form, indicating that they will fully comply with the protocol, to include an estimate of their yearly accrual and if any conflict of interest may arise due to their participation in the trial. A signed conflict of interest disclosure form: this document will be required only if a possible conflict is declared on the commitment form. A copy of the favorable opinion of their local or national (whichever is applicable) ethics committee mentioning the documents that have been reviewed (incl. version number and date of documents) and indicating the list of the ethics committee members. A copy of the translated and adapted (according to all national requirements), Patient Information / Informed Consent sheets, clearly mentioning the version number and the date. The signature log-list of the staff members with a sample of each authorized signature and the indication of the level of delegations. (This will be used for allocating usernames and passwords for remote data capture) The coordinates of the pharmacist who will be responsible for the trial medication (this will be filled in on the signature log). The accreditation letter for the laboratory. (if available for your center and/or applicable by your national law) A signed document to confirm receipt of the Investigator s brochure. A signed initial Acknowledgement of Transfer of biological Materials (ATM) to be sent to TRANSBIG. (Confirm ATM online at the end of accrual listing all sample SeqIDs) The center specific applicable list of required documents will be included in the site initiation package, with proper instructions as required by this protocol, your group and/or the applicable national law The new investigator will be added to the authorization list, and will be allowed to register/enroll/randomize patients in the trial as soon as All the above mentioned documents are available at their NCC/G or EORTC Headquarters All applicable national legal and regulatory requirements are fulfilled Patient registration for screening/enrollment/randomization from centers not (yet) included on the authorization list will not be accepted. Version / 152 May 21, 2014

239 15 Patient enrollment procedure 15.1 Enrollment procedure Patient enrollment/randomization will only be accepted from authorized investigators (see "Authorization procedure") Access to RDC for registration/enrollment/randomization Patients are registered, enrolled and randomized directly on RDC (RDC= Remote Data Capture), accessible 24 hours a day, 7 days a week ( Guidelines on how to register, enroll and randomize patients in RDC are provided to all centers by the EORTC. To access RDC, investigators need a username and a password. All authorized investigators will be provided with a username and password prior to site initiation. EORTC participants can use their ORTA username and password to access RDC. If an ORTA password has been forgotten, it can interactively be requested by a password reminder through the EORTC ORTA website ( Overview of patient screening and enrollment Steps to be taken for patient registration for screening (also see diagram chapter 20) identify patients who may be eligible. give information and ask patient to sign screening PIS & IC. register patient for screening on web page and receive patient identification number (SeqID). take blood sample for future research (optional for patient). Collected during the study, preferably before surgery. Date of breast cancer surgery, freeze tumor biopsy & register tumor sample as available for collection on RDC and Samples website. give tumor biopsy to courier & confirm sample pick-up. as soon as lymph node status is confirmed by local pathology this information must be recorded on the log lymph node form (RG3) on RDC (then the genomic test will be performed for eligible patients). Note that, in case of reoperation, to achieve axillary clearance (e.g. axillary lymph node dissection due to positive sentinel node biopsy) the lymph node status must be confirmed only after the definitive surgery. once the genomic prognosis has been successfully performed, the PI will receive an notification that patient is eligible for entry/enrollment in the trial. inform eligible patient about (general information). Steps to be taken for patient enrollment: timing and logistics for first post-operative visit A flow diagram similar to the one in chapter 20 will be provided to and adapted by each center to reflect local procedures and detail the logistics of the screening/enrollment process inform the patient about, signing of PIS & IC 1. connect to RDC, select the correct SeqID, create the enrollment checklist (RG5) and input clinical data and date of signing of PIS & IC 1 to enroll the patient. After the previous forms and the enrollment checklist (RG5) have been submitted, the results form (RG5trt) will be available on RDC. After completing Rg5trt, clinical and genomic risk result and treatment decision outcome will be available on this results form (RG5trt). Version / 152 May 21, 2014

240 Inform patient of proposed treatment. Discuss further randomizations as applicable, give patient additional PIS & ICs Patient enrollment and randomization for treatment decision making tool (R-T) A patient can be enrolled and randomized only after verification of eligibility. This must be done before the start of the protocol treatment. An exhaustive list of questions to be answered during the enrollment/randomization procedure is included in the enrollment checklist (RG5), which is part of the case report forms. This checklist should be completed on RDC by the responsible investigator before the patient is enrolled. Eligibility will be confirmed by collection of all the data requested on the enrollment checklist. Risk group assignment The computer will calculate a 10-year cancer specific survival probability, using the adjuvant! online software. The risk according to clinical-pathological criteria will be calculated by the Adjuvant! Online software incorporated into RDC. The patient s age, ER status, HER2 status, tumor grade, tumor size and nodal status will be passed on to Adjuvant! Online by RDC. Co-morbidity will be defaulted to Minor problems. The resulting 10 year cancer specific death risk will then be used to define the risk. If Adjuvant! Online is upgraded during the trial the new version will be used. If a new variable is added the new variable will be incorporated into the clinical prognosis. The standard version will always be used, not the genomic version. The computer will display all known information entered for this patient up to the time of enrollment. The computer will allow changes to any incorrect information (except the patient identification). After all information is confirmed as being correct the patient will be enrolled and risk assessments compared. For concordant patients: Clinical high risk-genomic high risk patients will receive chemotherapy Clinical low risk-genomic low risk patients will not receive chemotherapy For discordant patients the computer will randomized between the 2 decision making tools and, therefore allocate each patient to either: decision according to clinical prognosis: with no chemotherapy if patient has good clinical prognosis, and chemotherapy if patient has poor clinical prognosis decision according to 70-gene signature prognosis: with no chemotherapy if patient has good 70- gene signature prognosis, and chemotherapy if patient has poor 70-gene signature prognosis All resulting outcomes (clinical risk, genomic risk and treatment decision outcome) are visible on the results form (RG5trt) which the Principal Investigator has to create after the enrollment checklist (RG5) is sent. Version / 152 May 21, 2014

241 Randomization of chemotherapy (R-C) Once a patient has been assigned to receive chemotherapy, she can be offered the randomization to anthracycline based chemotherapy or docetaxel-capecitabine chemotherapy. This randomization can be done immediately after the assignment to chemotherapy, or later. In case the treatment decision outcome is chemotherapy, the chemotherapy checklist (RG6) has to be completed on RDC. If the patient agrees to the chemotherapy randomization, complete the section about chemotherapy randomization on the chemotherapy randomization checklist (RG6) and the randomization algorithm will allocate her to 1. anthracycline based chemotherapy (FEC D for N+) 2. docetaxel-capecitabine chemotherapy After sending the chemotherapy checklist (RG6), the chemotherapy treatment allocation will be visible on the chemotherapy checklist (RG6) on RDC. If a patient does not wish to participate in the chemotherapy randomization (R-C), she should receive the hospital s standard chemotherapy. The Principal Investigator will still need to complete the first 2 questions and section 1 of the chemotherapy checklist on RDC. The patient remains on protocol follow-up and the patient will still be a candidate to participate to the endocrine therapy randomization Randomization of endocrine therapy (R-E) All hormone receptor positive patients (i.e. ER &/or PgR positive) can be offered the endocrine therapy randomization to 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. In order to randomize a patient, complete the endocrine therapy checklist (RG8) on RDC. If the patient agrees to the endocrine therapy randomization, complete the section about endocrine therapy randomization on the endocrine therapy randomization checklist (RG8) and the randomization algorithm will allocate her to 1. 2 years of tamoxifen followed by 5 years of letrozole 2. 7 years of letrozole After sending the endocrine therapy checklist (RG8), the endocrine treatment allocation will be visible on the endocrine therapy checklist (RG8) on RDC. If a patient with an endocrine responsive tumor does not wish to participate in the endocrine therapy randomization (R-E), she should receive the hospital s standard endocrine therapy. The Principal Investigator will still need to complete the first 2 questions and section 1 of the endocrine therapy checklist on RDC. Version / 152 May 21, 2014

242 16 Forms and procedures for collecting data 16.1 Electronic case report forms and schedule for completion Once the results form (RG5trt) is completed and the patient is enrolled, all case report forms will be visible and available for completion on RDC. Completion guidelines for all forms are provided to all centers by the EORTC. All RDC forms are specifically designed by the EORTC Headquarters for this study. These electronic forms will be used by all participants. Electronic queries will be generated by the EORTC. These queries will be displayed in RDC and should be answered in RDC. A. Upon occurrence of a Pregnancy: Any pregnancy in a female subject diagnosed during the treatment period or within 30 days after last study treatment administration must be reported to the EORTC Pharmacovigilance Unit. This must be reported on a Pregnancy Notification Form/Fax within 24 hours of first becoming aware of the event and sent by fax to the allocated NCC who will in turn forward it to the EORTC Pharmacovigilance Unit. If a Serious Adverse Event (SAE) occurs in conjunction with the pregnancy, please also complete an SAE form as explained in the SAE chapter Upon notification of a pregnancy, the EORTC will coordinate the follow up, the development and outcome of the pregnancy. B. Upon occurrence of a Serious Adverse Event (SAE): SAEs occurring from the time a subject is enrolled until 30 days after last protocol treatment must be promptly reported. Any SAE occurring after the 30-days period and considered to be reasonably related to the investigational product or study participation, also have to be promptly notified All these events must be reported by fax to the allocated NCC/G who will in turn forward it to the EORTC Pharmacovigilance Unit on a Serious Adverse Event Form within 24 hours of the initial observation. A completed SAE-form must be sent back within 10 calendar days of the initial observation of the Serious Adverse Event. ALL Forms must be dated and signed by the responsible investigator or one of his/her authorized staff members 16.2 Data Flow (Remote Data Capture) The forms must be completed electronically according to the schedule defined in the guidelines. The list of site staff members, authorized to enter data, must be identified on the signature log and sent to the Headquarters by the responsible investigator before the start of the study. All staff need ORTA usernames and passwords to use the EORTC web based RDC system. In all cases, it remains the responsibility of the principal investigator to check that data are entered on the RDC electronic forms as soon as possible and that they are completely and correctly filled out and submitted to the EORTC database. The EORTC Headquarters will perform extensive consistency checks on the received data and will issue queries in case of inconsistent data. These queries will appear in the RDC system and must be answered there directly. Version / 152 May 21, 2014

243 The EORTC data manager will subsequently apply the corrections into the database. If an investigator (or an authorized staff member) needs to modify a CRF after the form has been electronically sent to the EORTC Headquarters, he/she should create a request for data correction in the RDC system. 17 Reporting of Serious Adverse Events 17.1 Definitions AE: An Adverse Event is defined as any untoward medical occurrence in a patient or clinical trial subject administered a medicinal product and which may not necessarily have a causal relationship with this treatment. An adverse event can therefore be any unfavorable and unintended signs (such as rash or enlarged liver), symptoms (such as nausea or chest pain), an abnormal laboratory finding (including blood tests, x-rays or scans) or a disease temporarily associated with the use of the protocol treatment, whether or not considered related to the investigational medicinal product. AR: An Adverse reaction of an investigational medicinal product is any untoward and unintended responses to an investigational medicinal product related to any dose administered. All adverse events judged by either the reporting investigator or the sponsor as having a reasonable causal relationship to a medicinal product qualify as adverse reactions. The expression reasonable causal relationship means to convey in general that there is evidence or argument to suggest a causal relationship. UAR: An Unexpected Adverse Reaction is any adverse reaction, the nature, or severity of which is not consistent with the applicable product information (e.g. investigator's brochure for an unapproved investigational product or summary of product characteristics (SmPC) for an authorized product). When the outcome of the adverse reaction is not consistent with the applicable product information this adverse reaction should be considered as unexpected. Severity: The term severe is often used to describe the intensity (severity) of a specific event. This is not the same as serious, which is based on patient/event outcome or action criteria. SAE: A Serious Adverse Event is defined as any undesirable experience occurring to a patient, whether or not considered related to the protocol treatment. SAR: A Serious Adverse Event (SAE) which is considered related to the protocol treatment is defined as a Serious Adverse Reaction An Adverse Event or Adverse Reaction which is considered as serious: results in death, is life-threatening (i.e. an event in which the subject was at risk of death at the time of event; it does not refer to an event which hypothetically might have caused death if it were more severe) requires hospitalization or prolongation of existing inpatients hospitalization, results in persistent or significant disability or incapacity, is a congenital anomaly or birth defect. results in any other medically important condition (i.e. important adverse reactions that are not immediately life threatening or do not result in death or hospitalization but may jeopardize the patient or may require intervention to prevent one of the other outcomes listed above), e.g. AE as a result of a secondary malignancy. NOTE: Any second primary malignancy will be monitored and must be reported as serious adverse event regardless of the treatment arm the subject is in and regardless of causal relationship with study treatment (randomized/non- Version / 152 May 21, 2014

244 randomized chemotherapy/endocrine therapy). This includes any second primary malignancy occurring at any time for the duration of the study. Documentation on the diagnosis of the second primary malignancy must be provided at the time of reporting as a serious adverse event (e.g., any confirmatory histology or cytology results, X-rays, CT scans, radiology reports with the conclusions summary in English, etc.). Additionally, second primary malignancy should also be reported on the regular CRF. SUSAR: Suspected Unexpected Serious Adverse Reactions 17.2 Reporting procedure Non- serious adverse events and/or non-serious adverse drug reactions Adverse Events (AE) and /or Adverse Reactions (AR) must be recorded as indicated in the protocol Serious adverse events or serious adverse drug reactions All Serious Adverse Events (SAE) occurring from the time a subject is enrolled until 30 days after last protocol treatment (including both randomized and non-randomized chemotherapy and endocrine therapy), must be reported to the EORTC Pharmacovigilance Unit within 24 hours. (Ref. SAE s will either be sent to the NCC/G (who will forward to the EORTC) or directly to the EORTC depending on the organization in the country. All SAEs that are simply signs and symptoms of the disease being studied do NOT need to be collected! All SAEs that are related to standard primary breast cancer surgery and adjuvant radiotherapy do NOT need to be reported. Examples of SAEs that do not need to be reported: Medical or surgical procedure (e.g., endoscopy, appendectomy); the condition that leads to the procedure is an AE Situations where an untoward medical occurrence did not occur (social and/or convenience admission to a hospital) Anticipated day-to-day fluctuations of pre-existing disease(s) or condition(s) present or detected at the start of the study that do not worsen The disease/disorder being studied, or expected progression, signs, or symptoms of the disease/disorder being studied, unless more severe than expected for the subject's condition A hospitalization which was planned before the patient consented for study participation and where admission did not take longer than anticipated. Any SAE that occurs outside of the SAE detection period (after the 30-days period), considered to be reasonably related to the investigational product or study participation, have to be promptly notified to the EORTC Pharmacovigilance Unit, either via the NCC/G (who will forward to the EORTC) or directly to the EORTC depending on the organization in the country This must be done by fax within 24 hours of the initial observation of the event. The principal investigator will decide if these events are related to the protocol treatment (i.e. unrelated, likely related, and not assessable) and the decision will be recorded on the Serious Adverse Event form, if necessary with the reasoning of the principal investigator. The investigator is obligated to assess the relationship between investigational product and the occurrence of each SAE. A "reasonable possibility" is meant to convey that there are facts/evidence or arguments to suggest a causal relationship, rather than a relationship cannot be ruled out. The Version / 152 May 21, 2014

245 investigator will use clinical judgement to determine the relationship. Alternative causes, such as natural history of the underlying diseases, concomitant therapy, other risk factors, and the temporal relationship of the event to the investigational product will be considered and investigated. The product reference documents are For Capecitabine and Letrozole, the Investigator Brochures will be the reference documents. For Docetaxel the reference document will be SmPC. For the other drugs the SmPCs will be the reference documents. For the causality assessment, the following definitions must be used: Relationship to the protocol treatment UNRELATED LIKELY RELATED NOT ASSESSABLE Description There is no evidence of any causal relationship to the protocol treatment There is (some) evidence to suggest a causal relationship to the protocol treatment and influence of other factors is unlikely or absent. There is insufficient or incomplete evidence to make a clinical judgment of the causal relationship to the protocol treatment. Details should be documented on the specified Serious Adverse Event Form. All Investigators participating through their National structure /NCC/G should follow recommendations below: PLEASE FAX THE REPORT TO: Your NCC/G (details can be found in the Administrative responsibilities chapter 21) (who will forward to the EORTC) or directly to the EORTC Headquarters if they are your NCC. NCC/Gs will forward to: Pharmacovigilance Unit fax no The EORTC Pharmacovigilance Unit will forward all Serious Adverse Event reports within 24 hours of receipt to all appropriate persons (See Administrative chapter). To enable the EORTC Pharmacovigilance Unit/sponsor to comply with regulatory reporting requirements, completed documentation of any reported serious adverse events or serious adverse reactions must be returned within 10 calendar days of the initial report. If the completed form is not received within this deadline, the Pharmacovigilance Unit will make a written request to the investigator. It should be recognized that Serious Adverse Reactions (SAR) which have not been previously documented in the applicable Reference Document, or which occur in a more severe form than anticipated (i.e. they are unexpected by nature or severity), are subject to rapid reporting to the Regulatory Authorities. Any question concerning SAE or SAR reporting can be directed to: EORTC Pharmacovigilance Unit phone: fax: pharmacovigilance@eortc.be Version / 152 May 21, 2014

246 ALL FORMS MUST BE DATED AND SIGNED BY THE RESPONSIBLE INVESTIGATOR OR ONE OF HIS/HER AUTHORIZED STAFF MEMBERS. 18 Quality assurance 18.1 Control of data consistency Data forms will be electronically sent to the database of the EORTC Headquarters by the RDC (Remote Data Capture) system. Computerized and manual consistency checks will be performed on newly received forms; queries will be issued in case of inconsistencies. Consistent forms will be validated by the data manager(s) to be entered in the master database. Inconsistent forms will be kept "pending" until resolution of the inconsistencies Quality Assurance and Quality Control Program An integrated Quality Assurance and Quality Control (QA & QC) Program that encompasses the usual EORTC extensive central monitoring checks, data quality checks and data timeliness and a quality check visit for selected sites will be carried out in collaboration between the EORTC and the NCC/Gs. This will insure that we can demonstrate a high level of quality throughout the trial. The QA & QC Program also details the audit plan for the study. Details of the Program and responsibilities can be found in the QA & QC Program and Intergroup agreements. Practically it will mean one 1 day visit for selected centers, with a potential additional visit for high recruiting centers, performed either by the EORTC and/or the NCC/G. The aim of these visits is to identify any areas where sites may require additional help or explanation and to demonstrate compliance to the protocol Audits To ensure quality of data, study integrity, and compliance with the protocol and the various applicable regulations and guidelines, the EORTC Quality Assurance Unit regularly conducts site visits to institutions participating to EORTC protocols. The investigator, by accepting to participate to this protocol, agrees to co-operate fully with any quality assurance visit undertaken by third parties, including representatives from the EORTC, national and/or foreign regulatory authorities or company supplying the product under investigation, as well as to allow direct access to documentation pertaining to the clinical trial (including CRFs, source documents, hospital patient charts and other study files) to these authorized individuals. The investigator must inform the EORTC immediately in case a regulatory authority inspection would be scheduled. Whether other groups center's will participate in the audit part of the QA & QC Program or in their own Group's audit programs will be agreed between the Collaborative Group and the EORTC. For details please contact your Group. 19 Ethical considerations 19.1 Patient protection The responsible investigator will ensure that this study is conducted in agreement with either the Declaration of Helsinki (available on the World Medical Association website ( and/or the laws and regulations of the country, whichever provides the greatest protection of the patient. Version / 152 May 21, 2014

247 The protocol has been written, and the study will be conducted according to the ICH Harmonized Tripartite Guideline for Good Clinical Practice (ICH-GCP, available online at The protocol must be approved by the competent ethics committee(s) as required by the applicable national legislation Subject identification The name of the patient will not be asked for nor recorded at the Headquarters. A sequential identification number will be automatically attributed to each patient enrolled in the trial. This number will identify the patient and must be included on all case report forms. In order to avoid identification errors, patient s code (maximum of 4 letters or numbers) and date of birth will also be reported on the case report forms Informed consent All patients will be informed of the aims of the study, the possible adverse events, the procedures and possible hazards to which she will be exposed, and the mechanism of treatment allocation. They will be informed as to the strict confidentiality of their patient data, but that their medical records may be reviewed for trial purposes by authorized individuals other than their treating physician. An example of the patient informed consent statements are given as an appendix to this protocol. Investigators will receive the translated PIS & ICs from their NCC/G. For countries where there is not an NCC/G the PIS & ICs will be translated by the national coordinator and will be sent to all centers by the EORTC. All PIS & ICs must be version controlled and dated and this information must always be stated in any communication with ethics committees. The translated informed consent form is part of the documents to be submitted to the ethics committee for approval. The competent ethics committee for each institution must validate local informed consent documents before the center can join the study. It is the responsibility of the Local Ethical Committee to guarantee that the translation is conforming to the ICH-GCP guidelines. It will be emphasized that the participation is voluntary and that the patient is allowed to refuse further participation in the protocol whenever she wants. This will not prejudice the patient s subsequent care. Documented informed consent must be obtained for all patients included in the study before they are enrolled or randomized at the EORTC Headquarters. This must be done in accordance with the national and local regulatory requirements. For European Union member states, the informed consent procedure must conform to the ICH guidelines on Good Clinical Practice. This implies that the written informed consent form should be signed and personally dated by the patient or by the patient s legally acceptable representative. 20 Sample schedule and trial logistics Detailed instructions describing the precautions involved in obtaining the tumor biopsy and tissue handling are described in a separate brochure and in SOPs & 15. SOPs 00-04, SOP 15 and the tissue handling brochure will be provided along with the tissue sample kit which will contain blood, serum and tumor tubes, a biopsy punch and coded stickers. Version / 152 May 21, 2014

248 20.1 Frozen tumor sample schedule For each patient a tumor biopsy is a prerequisite for inclusion in the trial. RNA will be extracted from part of the tumor biopsy and used for microarray analysis to determine the genomic prognosis. As RNA is very unstable special precautions need to be taken when taking and freezing the biopsy. These are summarized below and detailed in SOPs Immediately after surgery, the excised breast tumor tissue must be transported to the place where the biopsy will be taken and frozen, usually the pathology department. The tissue must be transported in a dry tumor container without fixatives as detailed in SOP 00. To decide the best region of the tumor from which the punch biopsy should be taken, obtain a good overview of the different sections of the tumor. The centre of the tumor is often necrotic, while the edge of the tumor often contains many stromal cells, which in both cases will lead to percentage of tumor cells less than 30% and therefore not enough to isolate tumor RNA from. Take a representative biopsy, from the periphery of the tumor thereby optimizing the chance of a high proportion of tumor cells and a good yield of tumor RNA. Tumor biopsy cellularity will be determined by Agendia along with other quality checks. One, or if feasible 2, punch biopsy samples must be obtained using the biopsy punch provided. A 6 mm biopsy is preferred however for small tumors (e.g. of <1 cm) biopsies of 3 mm are acceptable. In case of multifocal tumors; if the histopathological profile of all foci are similar (histopathological type, tumor grade, HR, HER2 status), the biopsy for the genomic assessment should be taken from the largest focus. The punch biopsy samples must be taken and frozen in liquid nitrogen within 1 hour of surgery, as detailed in SOP 01. Immediately after taking the punch biopsy sample place it in the labeled tube provided, firmly close the tube and submerge the tube in liquid nitrogen. After 2 minutes the biopsy tube can be transferred to a -80 C freezer (being transported on dry ice if necessary to prevent thawing). The availability of the tumor biopsy should then be registered on the web site (as detailed in SOP 02) and collected by a courier (as detailed in SOP 03). The frozen biopsy will be used for microarray analysis only once local lymph node status is confirmed and the patient is perceived likely to participate. Therefore it is essential that the results of local pathology determination of lymph node status are promptly recorded on the web page for both lymph node-negative and node-positive individuals Paraffin block schedule The remaining breast cancer tissue will be fixed according to each center s policy (with at least one paraffin block) and analyzed by the institution s pathologist, who will determine the histopathology. A representative paraffin block for each patient enrolled in must be sent for central pathology review by post, at least once every 6 months to: Dr. Giuseppe VIALE, ISTITUTO EUROPEO DI ONCOLOGIA Via Ripamonti, 435 IT MILAN Italy Tel: Fax: Blocks will be returned by post to each center once the central pathology review has been performed and TMAs produced. This will usually be within 2-3 weeks. Version / 152 May 21, 2014

249 20.3 Blood and serum sample schedule For those patients who consent to the optional blood sampling for whole blood and serum storage for proteomics analysis, blood should be taken after the patient has signed the screening PIS & IC but can be collected during the study (preferably before surgery). For whole blood, the sample should be taken into an EDTA blood tube. The whole blood should be aliquoted in 0.5 ml aliquots into cryovials pre-labeled with coded stickers according to the instructions provided in SOP MA 15. For serum, the sample should be taken into a serum blood tube with a gel separation system. Serum should be separated and frozen according to SOP 15 and stored at -80 C until the end of accrual when it should be sent to the TRANSBIG biological materials bank in Belgium Trial logistics study timelines guidelines, from diagnosis to start of treatment max 84 days max 56 days max 120 days Signing of spis & IC Patient receives general information surgeon / research nurse A. B. T1 T2 T3a T3b T4 Diagnosis Hospitalization Surgery* Surgical Oncologist post -op Start post-op visit visit Treatment C. D. G. F. E. Discussion Lymph node RNA QC good + status known Proposal for YES/NO Randomizations Feedback Cancer according to risk from patient 0-3 +ve LN Key Institute microarray T = Hospital visit / Time point Chemotherapy R = randomization and Hormonal PIS - IC = Patient information & Informed Consent therapy H. Signing of A. PIS & IC 2 or 3 A. Tumor Diagnosis Bring to Center B. Patient signs screening PIS & IC F. during LVEF visit C. Tumor sample sent to Cancer Institute or post to Center D. Local pathology, Lymph node status determination Information & signing E. RNA quantity & quality good, for 0-3 +ve LN PI told patient eligible & microarray of PIS & IC 1 (Discordant analysis performed patients are randomized) F. Genomic prognostic test performed, result communicated to PI after signing of PIS & IC 1 V 5p G. Patient returns home with R-C / R-E information to decide on participation H. Patient signs PIS & IC 2 or 3 and brings/posts it to Center/signs at the start of Treatment I. Patient inclusion in and randomization for discordant patients R I. * For patients who underwent more than1 surgery, the timelines start from date of last (definitive) surgery. This is a guideline for centers who will adapt this scheme to their local procedures Version / 152 May 21, 2014

250 21 Administrative responsibilities 21.1 The study coordinator The Study Coordinator(s) (in cooperation with the EORTC Headquarters) will be responsible for writing the protocol, contributing to the medical review, discussing the contents of the reports with the data manager(s) and the statistician, and for publishing the study results. He/she will assist the Clinical Research Physician for answering some clinical questions concerning eligibility, treatment, and the medical review of the patients. Study coordinator: Dr. Emiel Rutgers address: The Netherlands Cancer Institute-Antoni Van Leeuwenhoekziekenhuis Plesmanlaan 121 NL 1066 CX Amsterdam The Netherlands Phone: Fax: E.Rutgers@nki.nl Study co-coordinator: Dr. Fatima Cardoso address: Champalimaud Cancer Center Av. De Brasília - Doca de Pedrouços Lisbon, Portugal Phone: +351 (201) Fax: +351 (213) fatimacardoso@fundacaochampalimaud.pt Study co-coordinator: Dr. Martine Piccart address: Institut Jules Bordet 121 Blvd de Waterloo, B-1000 Brussels, Belgium Phone: Fax: martine.piccart@bordet.be 21.2 The EORTC Headquarters The EORTC Headquarters will be responsible for writing the protocol and PIS/IC, reviewing the protocol, setting up the trial, collecting case report forms, controlling the quality of the reported data, organizing the medical review and generating reports and analyses in cooperation with the Study Coordinator. All methodological questions should be addressed to the EORTC Headquarters. EORTC HEADQUARTERS 83, avenue Emmanuel Mounier, Bte 11 B-1200 Brussels, Belgium Fax: Registration of patients: Version / 152 May 21, 2014

251 Statisticians: Jan Bogaerts Phone: jan.bogaerts@eortc.be Leen Slaets Phone: leen.slaets@eortc.be Data Managers: Britt De Jongh Phone: britt.dejongh@eortc.be Roel Goossens Phone: Fax: roel.goossens@eortc.be Project Manager: Mélanie Beauvois Phone: melanie.beauvois@eortc.be Clinical Research Physician: Carlo Messina Phone: carlo.messina@eortc.be Pharmacovigilance Unit: Phone: Fax: pharmacovigilance@eortc.be The EORTC Pharmacovigilance Unit will forward all SAE within 24 hours of receipt to the EORTC Study Coordinator, the EORTC Data Manager and the contact persons at the pharmaceutical companies. All SUSARs will additionally be notified to all NCC/G s who will in turn inform all participating investigators and EC s from their group. The EORTC Pharmacovigilance Unit will take in charge the expedited reporting to the Competent Authorities, unless otherwise specified. The EORTC Pharmacovigilance Unit will prepare the Annual Safety report and distribute it to the Competent Authorities and the NCC/G s Pharmaceutical company contact person details Novartis Pharma AG Nora Schenk, Pharm D Novartis Pharmaceuticals Corporation Oncology Business Unit One Health Plaza, Blg 105 East Hanover, NJ , USA Tel: Version / 152 May 21, 2014

252 F. Hoffmann La Roche Séverine Wollenschneider Ph.D. Clinical Trial Manager XELODA F. Hoffmann-La Roche Ltd Grenzacherstrasse 124 Pharmaceuticals Division Pharma Business Medical Bldg. 074/4W.219 CH-4070 Basel Phone: Fax: Sanofi-Aventis: Medical Isabelle Tabah-Fisch, MD Head of Medical Affairs Oncology, Corporate Oncology Franchise Sanofi-Aventis Sanofi-Aventis: Operational & Financial Blandine Boussard GMA ClinOps-Study Operations, Trial Manager Oncology Sanofi-Aventis 46, Quai de la Rapée, Paris, France 46 Quai de la Rapée, Paris cedex 12, France Tel: Tel: +33-(0) Fax: Fax: +33-(0) The TRANSBIG Secretariat Breast International Group (BIG)-aisbl Institut Jules Bordet / 121 Blvd de Waterloo, 7th floor B-1000 Brussels, Belgium Tel: Fax: transbig@bordet.be The EORTC Group All questions concerning membership in the group should be addressed to the chairman and/or secretary of the group. EORTC Breast Cancer Group Chairman: Dr. Hervé Bonnefoi Phone: Fax: Bonnefoi@bergonie.org Secretary: Dr. David Cameron Phone: Fax: d.cameron@ncrn.org.uk Version / 152 May 21, 2014

253 21.5 National Coordinating Centers/Groups Country Austria Belgium Cyprus Czech Republic France Pharmacovigilance & local trial management Germany National structure contact details Medical University of Vienna - Department of Surgery A-1090 Wien, Waehringer Guertel 18-20, AUSTRIA Tel Fax michael.gnant@meduniwien.ac.at EORTC Headquarters 83, avenue Emmanuel Mounier, Bte 11, B-1200 Brussels, Belgium Tel: /1061 Fax: Bank of Cyprus Oncology Centre- (BOCOC) Medical Oncology, 32 Acropoleos Avenue, 2006, Nicosia, Cyprus Tel Fax yiola.marcou@bococ.org.cy Charles University Hospital Oncology dept., U nemocnice 2, Prague 2, , Czech Republic. Tel Fax petruzelka.lubos@seznam.cz Institut Gustave Roussy (IGR) 39, rue Camille Desmoulins, Villejuif cedex, France Tel: Fax: pelissier@igr.fr Fédération Nationale des Centres de Lutte Contre le Cancer (FNCLCC) 101, rue de Tolbiac, PARIS cedex 13, FRANCE Tel : Fax : West German Study Group (WSG) ggmbh Ludwig-Weber-Strasse Mönchengladbach Tel / Fax / corinna.buehne@wsg-online.com Version / 152 May 21, 2014

254 Country Italy Luxembourg Poland Scandinavia (Sweden, Denmark, Finland & Norway) Serbia Montenegro Slovenia Spain National structure contact details Italian oncology Group for clinical research (GOIRC) Medicina e Oncologia Medica, Azienda Ospedaliera di Cremona, Viale Concordia 1, Cremona, Italy. Tel Fax passalacqua.aioc@e-cremona.it Centre Hospitalier Luxembourg (CHL) Rue Barble, 4, L-1210 Luxembourg, Luxembourg Tel Fax Duhem.Caroline@chl.lu Medical University of Gdansk (AMG) Department of Oncology and Radiotherapy, Debinki 7, Gdansk, , Poland Tel Fax jjassem@amg.gda.pl Karolinska Institute (KI) Department of Oncology, Karolinska University Hospital Solna, SE Stockholm, Sweden Tel Fax nils.wilking@karolinska.se EORTC Headquarters 83, avenue Emmanuel Mounier, Bte 11, B-1200 Brussels, Belgium Tel: /1655 Fax: Institute of Oncology Ljubljana (IOL) Medical Oncology, Zaloska 2, 1000 Slovenia Tel Fax tcufer@onko-i.si SOLTI SANT LLORENÇ, 23 bajos, Barcelona, 08202, Spain Tel: Fax: josep.vazquez@gruposolti.org Version / 152 May 21, 2014

255 Country The Netherlands Turkey UK Pharmacovigilance & local trial management National structure contact details The Netherlands Cancer Institute-Antoni van Leeuwenhoekziekenhuis Plesmanlann 121, NL 1066 CX Amsterdam, The Netherlands. Tel Fax Marmara Universitesi Hastanesi Tıbbi Onkoloji Bölümü Tophanelioglu Cad No:13-15 Altunizade Altunizade Istanbul Tel: Fax gulbasaran@superonline.com ISD Cancer Clinical Trials Team (Partner in CaCTUS - Cancer Clinical Trials Unit Scotland) Open Plan Area 159C, 1st Floor Gyle Square 1 South Gyle Crescent Edinburgh, EH12 9EB Scotland Tel: Fax: kirsten.murray@isd.csa.scot.nhs.uk University of Dundee Ninewells Hospital and Medical School Prof A Thompson Tel: Fax: a.m.thompson@dundee.ac.uk 22 Trial sponsorship and financing EORTC is the legal European Sponsor unless otherwise specified for legal or logistical reasons. The Director General of the EORTC is: Professor Françoise Meunier EORTC Avenue Mounier 83, Bte 11 B-1200 Brussels, Belgium Tel: Fax: francoise.meunier@eortc.be Version / 152 May 21, 2014

256 Financial support has primarily been provided by: Industrial partners Novartis F. Hoffman La Roche Sanofi-Aventis The European Commission Framework Programme VI (FP6-LSHC-CT ), the NCI (through the EORTC) and Charitable trusts including the Breast Cancer Research Foundation, the Jacqueline Seroussi Memorial Foundation and the Prix Mois du Cancer du Sein. Indirect financial support has been provided by Agendia (via reduced cost for performing microarray analysis). 23 Trial insurance A clinical trial insurance has been taken according to the laws of the countries where the study will be conducted. An insurance certificate will be made available to the participating sites at the time of study initiation. Clinical trial insurance is only valid if the treatment is given in a center authorized by the EORTC Headquarters and which has obtained Ethical Committee approval (individually or centrally depending on the national regulations applicable). Therefore all centers will have to declare to their NCC/G, who will inform the EORTC Headquarters, of details of other satellite institutions which may be responsible for providing protocol treatment to the patients. Details on these satellite institutions including CV for the local investigator, laboratory normal ranges and the Ethical Approval, will have to be transmitted to the NCC/G. Correspondence on study issues and data collection will however only be performed with the primary institution which will assume all responsibilities and liabilities issues with the principle investigator. Version / 152 May 21, 2014

257 Appendix A: References Ref. 1 Veronesi, U., Boyle, P., Goldhirsch, A., et al. Breast cancer. Lancet 365, (2005). Ref. 2 Becker, N. Cancer mortality and prevention in the European Union. Eur J Surg Oncol 24, (1998). Ref. 3 Ref. 4 Ferlay J., Bray F., Pisani P. Globocan 2002: cancer incidence, mortality and prevalence worldwide. IARC CancerBase No 5, version 2 0. Parkin, D.M., Bray, F., Ferlay, J., and Pisani, P. Global cancer statistics, CA Cancer J Clin 55, (2005). Ref. 5 Boyle, P. and Ferlay, J. Cancer incidence and mortality in Europe, Ann Oncol 16, (2005). Ref. 6 Ref. 7 Ref. 8 Ref. 9 Ryan, P.D., Kopans, D.B., and Sgroi, D.C. Case records of the Massachusetts General Hospital. Case A 58-year-old woman with early-stage estrogen-receptor-positive breast cancer. N Engl J Med 353, (2005). Goldhirsch, A. et al. Meeting highlights: updated international expert consensus on the primary therapy of early breast cancer. J Clin Oncol 21, (2003). Eifel, P. et al. National Institutes of Health Consensus Development Conference Statement: adjuvant therapy for breast cancer, November 1-3, J Natl Cancer Inst 93, (2001). Goldhirsch, A. et al. Meeting Highlights: International Expert Consensus on the Primary Therapy of Early Breast Cancer Ann Oncol (2005). Ref. 10 Early Breast Cancer Trialists' Collaborative Group (EBCTCG). Effects of chemotherapy and hormonal therapy for early breast cancer on recurrence and 15-year survival: an overview of the randomised trials. Lancet 365, (2005). Ref. 11 Hutchins L., Green S., Ravdin P. et al. CMF versus CAF with and without Tamoxifen in high-risk node-negative breast cancer patients and a natural history follow-up study in lowrisk node-negative patients: First results of Intergroup trial INT 0102 (Meeting abstract). Proc Am Soc Clin Oncol 17, 1a, abstract 2. (1998). Ref. 12 McShane, L.M. et al. REporting recommendations for tumour MARKer prognostic studies. Eur J Cancer 41, (2005). Ref. 13 Haybittle, J.L. et al. A prognostic index in primary breast cancer. Br J Cancer 45, (1982). Ref. 14 Joensuu, H., Pylkkanen, L., and Toikkanen, S. Late mortality from pt1n0m0 breast carcinoma. Cancer 85, (1999). Ref. 15 Quiet, C.A., Ferguson, D.J., Weichselbaum, R.R., & Hellman, S. Natural history of nodenegative breast cancer: a study of 826 patients with long-term follow-up. J Clin Oncol 13, (1995). Ref. 16 Seidman, H., Gelb, S.K., Silverberg, et al. Survival experience in the Breast Cancer Detection Demonstration Project. CA Cancer J Clin 37, (1987). Ref. 17 Rosen, P.R., Groshen, S., Saigo, P.E., et al. A long-term follow-up study of survival in stage I (T1N0M0) and stage II (T1N1M0) breast carcinoma. J Clin Oncol 7, (1989). Ref. 18 Fisher, B. et al. Prognosis and treatment of patients with breast tumors of one centimeter or less and negative axillary lymph nodes. J Natl Cancer Inst 93, (2001). Version / 152 May 21, 2014

258 Ref. 19 Leitner, S.P., Swern, A.S., Weinberger, et al. Predictors of recurrence for patients with small (one centimeter or less) localized breast cancer (T1a,b N0 M0). Cancer 76, (1995). Ref. 20 Chia, S.K., Speers, C.H., Bryce, C.J., et al. Ten-year outcomes in a population-based cohort of node-negative, lymphatic, and vascular invasion-negative early breast cancers without adjuvant systemic therapies. J Clin Oncol 22, (2004). Ref. 21 Singletary, S.E. et al. Revision of the American Joint Committee on Cancer staging system for breast cancer. J Clin Oncol 20, (2002). Ref. 22 Elston, C.W. and Ellis, I.O. Pathological prognostic factors in breast cancer. I. The value of histological grade in breast cancer: experience from a large study with long-term followup. Histopathology 19, (1991). Ref. 23 Early Breast Cancer Trialists Collaborative Group. Effects of chemotherapy and hormonal therapy for early breast cancer on recurrence and 15-year survival: an overview of the randomized trials. Lancet. 2005;365: Ref. 24 Albain K, Barlow W, O Malley F, et al. Concurrent (CAFT) versus sequential (CAF-T) chemohormonal therapy (cyclophosphamide, doxorubicin, 5-fluorouracil, tamoxifen) versus T alone for postmenopausal node-positive, estrogen (ER) and/or progesterone (PgR) receptor-positive breast cancer: mature outcomes and new biologic correlates on phase III intergroup trial 0100 (SWOG 8814). Breast Cancer Res Treat. 2005;90:115. Ref. 25 Harbeck N, Kates RE, Look MP, Meijer-van Gelder ME, Klijn JGM, Jänicke F, Krüger A, Kiechle M, Schmitt M, Foekens JA. Enhanced benefit from adjuvant systemic chemotherapy in breast cancer patients classified high-risk according to upa and PAI-1 (n=3,424). Cancer Res 62 (16): , 2002 Ref. 26 Goldhirsch A, Wood WC, Gelber RD, Coates AS et al 2007, Progress and promise: highlights of the international expert consensus on the primary therapy of early breast cancer Annals of Oncology 18: , 2007 Ref. 27 Sotiriou C., Neo S. Y., McShane L; M., et al. Breast cancer classification and prognosis based on gene expression profiles from a population-based study. Proc Natl Acad Sci U S A 100, (2003). Ref. 28 Sorlie, T. et al. Repeated observation of breast tumor subtypes in independent gene expression data sets. Proc Natl Acad Sci U S A 100, (2003). Ref. 29 Sorlie, T. et al. Gene expression patterns of breast carcinomas distinguish tumour subclasses with clinical implications. Proc Natl Acad Science 98, (2001). Ref. 30 Perou, C.M. et al. Molecular portraits of human breast tumours. Nature 406, (2000). Ref. 31 West, M. et al. Predicting the clinical status of human breast cancer by using gene expression profiles. Proc Natl Acad Sci U S A 98, (2001). Ref. 32 van 't Veer, L.J. et al. Gene expression profiling predicts clinical outcome of breast cancer. Nature 415, (2002). Ref. 33 van de Vijver, M.J. et al. A gene-expression signature as a predictor of survival in breast cancer. N Engl J Med 347, (2002). Ref. 34 Simon, R., Radmacher, M.D., Dobbin, K., and McShane, L.M. Pitfalls in the use of DNA microarray data for diagnostic and prognostic classification. J Natl Cancer Inst 95, 14-8 (2003). Version / 152 May 21, 2014

259 Ref. 35 Ransohoff, D.F. Rules of evidence for cancer molecular-marker discovery and validation. Nat Rev Cancer 4, (2004). Ref. 36 Michiels, S., Koscielny, S., and Hill, C. Prediction of cancer outcome with microarrays. Lancet 365, (2005). Ref. 37 Eden, P., Ritz, C., Rose, C., et al. "Good Old" clinical markers have similar power in breast cancer prognosis as microarray gene expression profilers. Eur J Cancer 40, (2004). Ref. 38 Cardoso, F. Microarray technology and its effect on breast cancer (re)classification and prediction of outcome. Breast Cancer Res 5, (2003). Ref. 39 Piccart, M. Loi S. Van tveer L. et al. o. b. o. t. T. C. In San Antonio Breast Cancer Symposium abstract 38. (2004). Ref. 40 Ravdin, P.M. et al. Computer program to assist in making decisions about adjuvant therapy for women with early breast cancer. J Clin Oncol 19, (2001). Ref. 41 Olivotto, I.A. et al. Population-based validation of the prognostic model ADJUVANT! for early breast cancer. J Clin Oncol 23, (2005). Ref. 42 Buyse M, Loi S, van't Veer L et al Validation and clinical utility of a 70-gene prognostic signature for women with node-negative breast cancer. J Natl Cancer Inst Sep 6;98(17): Ref. 43 Mook S, Schmidt MK, Viale G, Pruneri G, Eekhout I, Piccart MJ, Cardoso F, Rutgers EJ and Van t Veer LJ, on behalf of the TRANSBIG consortium. Breast cancer patients with 1-3 positive lymph nodes and a low risk 70-gene profile have an excellent survival. Breast Cancer Research and Treatment 106 (supp 1): late breaking abstract #50 Ref. 44 Paik, S. et al. A multigene assay to predict recurrence of tamoxifen-treated, node-negative breast cancer. N Engl J Med 351, (2004). Ref. 45 Hornberger, J., Cosler, L.E., and Lyman, G.H. Economic analysis of targeting chemotherapy using a 21-gene RT-PCR assay in lymph-node-negative, estrogen-receptorpositive, early-stage breast cancer. Am J Manag Care 11, (2005). Ref. 46 Early Breast Cancer Trialists' Collaborative Group. Polychemotherapy for early breast cancer: an overview of the randomised trials. Lancet 352, (1998). Ref. 47 Levine, M.N. et al. Randomized trial of intensive cyclophosphamide, epirubicin, and fluorouracil chemotherapy compared with cyclophosphamide, methotrexate, and fluorouracil in premenopausal women with node-positive breast cancer. National Cancer Institute of Canada Clinical Trials Group. J Clin Oncol 16, (1998). Ref. 48 Coombes, R.C. et al. Adjuvant cyclophosphamide, methotrexate, and fluorouracil versus fluorouracil, epirubicin, and cyclophosphamide chemotherapy in premenopausal women with axillary node-positive operable breast cancer: results of a randomized trial. The International Collaborative Cancer Group. J Clin Oncol 14, (1996). Ref. 49 Misset, J.L. et al. Adjuvant treatment of node-positive breast cancer with cyclophosphamide, doxorubicin, fluorouracil, and vincristine versus cyclophosphamide, methotrexate, and fluorouracil: final report after a 16-year median follow-up duration. J Clin Oncol 14, (1996). Ref. 50 Tucker, M.A. et al. Leukemia after therapy with alkylating agents for childhood cancer. J Natl Cancer Inst 78, (1987). Ref. 51 Chambers, S.K., Chopyk, R.L., Chambers, J.T., et al. Development of leukemia after doxorubicin and cisplatin treatment for ovarian cancer. Cancer 64, (1989). Version / 152 May 21, 2014

260 Ref. 52 Praga, C. et al. Risk of acute myeloid leukemia and myelodysplastic syndrome in trials of adjuvant epirubicin for early breast cancer: correlation with doses of epirubicin and cyclophosphamide. J Clin Oncol 23, (2005). Ref. 53 Crump M, Tu D & Shepherd L. Risk of acute leukemia following adjuvant epirubicinbased adjuvant chemotherapy for breast cancer. Breast Cancer Res Treat 69, 247, abstract 243. (2001). Ref. 54 Smith RE, Bryant J & DeCellis A. Acute myeloid leukemia and myelodysplastic syndrome following doxorubicin-cyclophosphamide adjuvant therapy for operable breast cancer: the NSABP experience. Breast Can. Res. Treat. 69: 209 (abst 2), (2001). Ref. 55 Zambetti, M. et al. Long-term cardiac sequelae in operable breast cancer patients given adjuvant chemotherapy with or without doxorubicin and breast irradiation. J Clin Oncol 19, (2001). Ref. 56 Bonneterre, J. et al. Epirubicin increases long-term survival in adjuvant chemotherapy of patients with poor-prognosis, node-positive, early breast cancer: 10-year follow-up results of the French Adjuvant Study Group 05 randomized trial. J Clin Oncol 23, (2005). Ref. 57 Henderson, I.C. et al. Improved outcomes from adding sequential Paclitaxel but not from escalating Doxorubicin dose in an adjuvant chemotherapy regimen for patients with nodepositive primary breast cancer. J Clin Oncol 21, (2003). Ref. 58 Mamounas E. P., Bryant J., Lembersky B. C. et al. Paclitaxel (T) following doxorubicin/cyclophosphamide (AC) as adjuvant chemotherapy for node-positive breast cancer: Results from NSABP B-28. Proc Am Soc Clin Oncol 22, 4, (abstr 12). (2003). Ref. 59 Nabholtz J M., Pienkowski T., Mackey J. et al. Phase III trial comparing TAC (docetaxel, doxorubicin, cyclophosphamide) with FAC (5-fluorouracil, doxorubicin, cyclophosphamide) in the adjuvant treatment of node positive breast cancer (BC) patients: interim analysis of the BCIRG 001 study. Proc Am Soc Clin Oncol 21. (2002). Ref. 60 Martin, M. et al. Adjuvant docetaxel for node-positive breast cancer. N Engl J Med 352, (2005). Ref. 61 Roché H, Fumoleau P, Spielmann M et al. Sequential adjuvant epirubicin-based and docetaxel chemotherapy for node-positive breast cancer patients: the FNCLCC PACS 01 Trial. J Clin Oncol Dec 20;24(36): Ref. 62 Jones S. E., Savin M. A., Asmar L. et al. Three year results of a prospective randomized trial of adjuvant chemotherapy for patients (pts) with stage I-III operable, invasive breast cancer comparing 4 courses of doxorubicin/cyclophosphamide (AC) to 4 courses of docetaxel/cyclophosphamide (TC). Proc ASCO, Abstract 59, (2003). Ref. 63 Miwa, M. et al. Design of a novel oral fluoropyrimidine carbamate, capecitabine, which generates 5-fluorouracil selectively in tumours by enzymes concentrated in human liver and cancer tissue. Eur J Cancer 34, (1998). Ref. 64 Schuller, J. et al. Preferential activation of capecitabine in tumor following oral administration to colorectal cancer patients. Cancer Chemother Pharmacol 45, (2000). Ref. 65 O'Shaughnessy, J. et al. Superior survival with capecitabine plus docetaxel combination therapy in anthracycline-pretreated patients with advanced breast cancer: phase III trial results. J Clin Oncol 20, (2002). Ref. 66 Lee H. G., Lee J. J., Jung K. H. et al. Phase III randomized trial of primary chemotherapy with doxorubicin/cyclophosphamide (AC) vs docetaxel/capecitabine (TX) for stage II/III Version / 152 May 21, 2014

261 breast cancer (BC): Interim analysis. J Clin Oncol ASCO Annual Meeting Proceedings. 22, abstract 607. (2004). Ref. 67 Fisher, B. et al. Treatment of lymph-node-negative, oestrogen-receptor-positive breast cancer: long-term findings from National Surgical Adjuvant Breast and Bowel Project randomised clinical trials. Lancet 364, (2004). Ref. 68 Fisher, B. et al. Tamoxifen for prevention of breast cancer: report of the National Surgical Adjuvant Breast and Bowel Project P-1 Study. J Natl Cancer Inst 90, (1998). Ref. 69 Gorin, M.B. et al. Long-term tamoxifen citrate use and potential ocular toxicity. Am J Ophthalmol 125, (1998). Ref. 70 Goss, P.E. et al. A randomized trial of letrozole in postmenopausal women after five years of tamoxifen therapy for early-stage breast cancer. N Engl J Med 349, (2003). Ref. 71 Elisaf, M. et al. The beneficial effect of tamoxifen on serum lipoprotein-a levels: an additional anti-atherogenic property. Anticancer Res 16, (1996). Ref. 72 Rossouw, J.E. et al. Risks and benefits of estrogen plus progestin in healthy postmenopausal women: principal results From the Women's Health Initiative randomized controlled trial. Jama 288, (2002). Ref. 73 Thurlimann, B.J. et al. BIG 1-98: Randomized double-blind phase III study to evaluate letrozole (L) vs. tamoxifen (T) as adjuvant endocrine therapy for postmenopausal women with receptor-positive breast cancer. Proc Am Soc Clin Oncol 23, (2005). Ref. 74 Wasan, K.M. et al. The influence of letrozole on serum lipid concentrations in postmenopausal women with primary breast cancer who have completed 5 years of adjuvant tamoxifen (NCIC CTG MA.17L). Ann Oncol 16, (2005). Ref. 75 Harper-Wynne, C. et al. Effects of the aromatase inhibitor letrozole on normal breast epithelial cell proliferation and metabolic indices in postmenopausal women: a pilot study for breast cancer prevention. Cancer Epidemiol Biomarkers Prev 11, (2002). Ref. 76 Elisaf, M.S. et al. Effect of letrozole on the lipid profile in postmenopausal women with breast cancer. Eur J Cancer 37, (2001). Ref. 77 Baum, M. et al. Anastrozole alone or in combination with tamoxifen versus tamoxifen alone for adjuvant treatment of postmenopausal women with early-stage breast cancer: results of the ATAC (Arimidex, Tamoxifen Alone or in Combination) trial efficacy and safety update analyses. Cancer 98, (2003). Ref. 78 Howell, A. et al. Results of the ATAC (Arimidex, Tamoxifen, Alone or in Combination) trial after completion of 5 years' adjuvant treatment for breast cancer. Lancet 365, 60-2 (2005). Ref. 79 Dowsett, M. ATAC trialists group: Analysis of time to recurrence in the ATAC trial according to estrogen receptor and progesterone receptor status. Breast Cancer Res Treat 82, S7 (2003). Ref. 80 Coombes, R.C. et al. A randomized trial of exemestane after two to three years of tamoxifen therapy in postmenopausal women with primary breast cancer. N Engl J Med 350, (2004). Ref. 81 Coombes, R.C., Hall, E., and Snowden, C. The Intergroup Exemestane study: a randomized trial in postmenopausal patients with early breast cancer who remain disease free after two to three years of tamoxifen- updated survival analysis. Br Cancer Res Treat 88, (2004). Version / 152 May 21, 2014

262 Ref. 82 Jackesz, R., Kaufman, M., and Gnant, M. Benefits of switching postmenopausal women with hormone -sensitive disease to anastrozole after 2 years of adjuvant tamoxifen. Breast Cancer Res Treat 88, S7 (2004). Ref. 83 Boccardo F. M., Rubagotti A., Puntoni M., et al. Switching to anastrozole (ANA) vs continued tamoxifen (TAM) treatment of early breast cancer (EBC). Updated results of the Italian tamoxifen anastrozole (ITA) trial. ASCO 2005, abstract 526, (2005). Ref. 84 Boccardo, F. et al. Switching to Anastrozole Versus Continued Tamoxifen Treatment of Early Breast Cancer: Preliminary Results of the Italian Tamoxifen Anastrozole Trial. J Clin Oncol, 23, (2005). Ref. 85 Saphner, T., Tormey, D.C., and Gray, R. Annual hazard rates of recurrence for breast cancer after primary therapy. J Clin Oncol 14, (1996). Ref. 86 Goss, P.E. et al. Randomized trial of letrozole following tamoxifen as extended adjuvant therapy in receptor-positive breast cancer: updated findings from NCIC CTG MA.17. J Natl Cancer Inst 97, (2005). Ref. 87 Jakesz, R., Samonigg, R., Griel, R., et al. Extending adjuvant treatment with anastrozole: Results from the Austrian Breast and Colorectal Cancer Study Group Trial 6a (ABCSG- 6a). American Society of Clinical Oncology, abstract 527 (2005). Ref. 88 Winer, E.P. et al. American Society of Clinical Oncology technology assessment on the use of aromatase inhibitors as adjuvant therapy for postmenopausal women with hormone receptor-positive breast cancer: status report J Clin Oncol 23, (1920). Ref. 89 Rutgers, E.J. Guidelines to assure quality in breast cancer surgery. Eur J Surg Oncol 31, (2005). Ref. 90 EORTC Breast Cancer Group. Manual for clinical research and treatment in breast cancer. (2005). Ref. 91 Benefit of a high-dose epirubicin regimen in adjuvant chemotherapy for node-positive breast cancer patients with poor prognostic factors: 5-year follow-up results of French Adjuvant Study Group 05 randomized trial. J Clin Oncol 19, (2001). Ref. 92 Stewart, D.J. et al. Cyclophosphamide and fluorouracil combined with mitoxantrone versus doxorubicin for breast cancer: superiority of doxorubicin. J Clin Oncol 15, (1997). Ref. 93 Budman, D.R. et al. Dose and dose intensity as determinants of outcome in the adjuvant treatment of breast cancer. The Cancer and Leukemia Group B. J Natl Cancer Inst 90, (1998). Ref. 94 Poole C. J., Earl H. M., Dunn J. A., et al. NEAT (National Epirubicin Adjuvant Trial) and SCTBG BR9601 (Scottish Cancer Trials Breast Group) phase III adjuvant breast trials show a significant relapse-free and overall survival advantage for sequential ECMF. Proc Am Soc Clin Oncol 22, 4, abstract 13. (2003). Ref. 95 Vukelja, S.J., Lombardo, F.A., James, W.D., and Weiss, R.B. Pyridoxine for the palmarplantar erythrodysesthesia syndrome. Ann Intern Med 111, (1989). Ref. 96 Vukelja, S.J., Baker, W.J., Burris, H.A. 3rd, et al. Pyridoxine therapy for palmar-plantar erythrodysesthesia associated with taxotere. J Natl Cancer Inst 85, (1993). Ref. 97 Cascinu, S., Fedeli, A., Fedeli, S.L., and Catalano, G. Octreotide versus loperamide in the treatment of fluorouracil-induced diarrhea: a randomized trial. J Clin Oncol 11, (1993). Version / 152 May 21, 2014

263 Ref. 98 Hillner, B.E. et al. American Society of Clinical Oncology 2003 update on the role of bisphosphonates and bone health issues in women with breast cancer. J Clin Oncol 21, (2003). Ref. 99 Kalbfleisch J.D., & Prentice R. L. The statistical analysis of failure time data. Wiley. (1980). Ref. 100 Kim K. and DeMets D.L. Design and analysis of group sequential tests based on the type I error spending rate function. Biometrika 74, (1987). Ref. 101 Baum, M. and Ravdin, P.M. Decision-making in early breast cancer: guidelines and decision tools. Eur J Cancer 38, (2002). Ref. 102 Harrell, F.E. Jr, Lee, K.L., & Mark, D.B. Multivariable prognostic models: issues in developing models, evaluating assumptions and adequacy, and measuring and reducing errors. Stat Med 15, (1996). Version / 152 May 21, 2014

264 Appendix B: WHO performance status scale Grade Performance scale 0 Able to carry out all normal activity without restriction 1 Restricted in physically strenuous activity but ambulatory and able to carry out light work. 2 Ambulatory and capable of all self-care but unable to carry out any work; up and about more than 50% of waking hours. 3 Capable of only limited self-care; confined to bed or chair more than 50% of waking hours 4 Completely disabled; cannot carry on any self-care; totally confined to bed or chair. Version / 152 May 21, 2014

265 Appendix C: Common Terminology Criteria for Adverse Events In the present study, adverse events and/or adverse drug reactions will be recorded according to the Common Terminology Criteria for Adverse Events (CTCAE), version 4.0. At the time this protocol was issued, the full CTC document was available on the NCI web site, at the following address: The EORTC Headquarters web site provides a link to the appropriate CTC web site. This link will be updated if the CTC address is changed. Version / 152 May 21, 2014

266 Appendix D: World Medical Association Declaration of Helsinki The current Declaration of Helsinki can be found on the World Medical Association web page via the link provided below: Version / 152 May 21, 2014

267 Appendix E: Screening patient information sheet and informed consent document. Screening The screening patient information sheet & informed consent (spis & IC) gives consent for the donation of a piece of tumor tissue and for its being sent to a specialized cancer center and for the microarray analysis to be performed. Also for the donation of a blood sample. We have enclosed an example template for this consent and each centre can choose whether they would like to use/adapt this template or they can use their center s standard PIS & IC if they are confident that it covers the sending of tissue and the microarray analysis and the donation of a blood sample. Each center will decide on a single spis & IC which will be used for all potential patients following local policies. Each center will sign a certificate to certify the content of the PIS & IC in use for screening, on which will specify that it is either: a faithful translation of the spis & IC template provided a modified translation of the template provided their own PIS & IC In the case of a center using a modified template or their own PIS & IC that it covers: the sending of tissue abroad the microarray analysis donation of a blood sample and also that a blank copy of the PIS & IC in use for screening is filed locally in the master file There will be a maximum of 4 PIS & ICs for this study. All screened patients will sign the spis & IC. The PIS & IC 1, will be signed by all eligible patients before the two methods of risk assessment are compared and patients are enrolled/ randomized for treatment decision making tool. Patients who will receive chemotherapy and who agree to be randomized for chemotherapy regimen will also sign the PIS & IC 2 and patients who are eligible for endocrine therapy and who agree to be randomized for endocrine therapy regimen will sign the PIS & IC 3. Version / 152 May 21, 2014

268 Patient Information Sheet Request for consent to send tumor tissue to another institution for specialized analysis Tumor removal, examination and breast cancer prognosis You have been diagnosed with breast cancer and will soon undergo an operation to remove your tumor. During the surgical procedure, lymph nodes located under the arm (called axillary lymph nodes) will also be removed. Lymph nodes work like filters in your body to catch things like germs or cancer cells. After surgery, the axillary lymph nodes, together with tissue from your tumor, will be examined closely under a microscope (pathological examination). The results of the pathological examination will help us determine the specific characteristics of your disease. These results will also give us some information about the prognosis of your breast cancer. In other words, the results will give us some information about whether it is likely that your breast cancer will come back - in which case additional treatment will be needed - or whether your breast cancer can be safely managed with local treatment alone (surgery with or without radiotherapy). The pathological examination of your tissue can be performed in this hospital and will take a maximum of 2 weeks. A new test for breast cancer prognosis We know that cancer can appear when some genes are damaged and some of the body s cells no longer function normally. Consequently, many scientists now think that by looking at the genes of tumor cells, we will get even more information about a particular tumor than through pathological examinations. In other words, the genes of tumor cells may give us more accurate information about whether a woman s breast cancer is likely to be aggressive. Patients with aggressive tumors need more than local treatment and are given chemotherapy and/or hormonal therapy (also called systemic or adjuvant therapy). This hospital is involved in a research study that will look at a new and possibly more accurate test for determining breast cancer prognosis than traditional pathological examinations. The new test uses information about gene activity in tumor cells, and it was developed by researchers from the Netherlands Cancer Institute (Nederlands Kanker Instituut) in Amsterdam. The test is based on a technology called microarray (also called genomics), which determines which genes are active ( gene expression ) in a tumor, much like taking a fingerprint of it. The purpose of the research study this hospital is involved in is to find out if this new test is really better than traditional pathological examinations of tumor tissue. The new test has been developed for those patients who do not have cancer cells in the lymph nodes (called lymph node-negative breast cancer) and later for those patients with tumor cells found in 1 to 3 of their lymph nodes. If the new test is eventually shown to be more accurate than traditional pathological examinations, it will be easier for doctors to know which women really need systemic cancer treatment like chemotherapy and which women are likely to be cured by local treatment alone. Where can the new prognostic test be done? The new prognostic test is not available in this hospital. To have the gene expression test done, we would need to send a piece of your left over tumor tissue to the Netherlands where the technique was developed. After your surgery, and depending on the results of the pathological examination of your axillary lymph nodes, we may invite you to participate in the research study investigating the new test, and we will provide you with detailed information at that time. However, because of the way this new prognostic test is designed and to avoid any delay in deciding if you need additional treatment beyond surgery, we have to send your tissue to the Netherlands shortly after your operation. Researchers there will perform the test and send the results to your treating physician. It is important to note that this prognostic test is not a test to determine whether breast cancer runs in your family, since it is not your own genes that will be examined but those of your tumor. The test will simply determine the pattern of gene expression in your tumor tissue (the fingerprint ) by comparing it to the expression patterns that the Dutch researchers have found to be good (the Version / 152 May 21, 2014

269 tumor is unlikely to come back and may not need further treatment) and those they have found to be aggressive (the tumor is likely to need further treatment to prevent it from coming back). This is why this is not a genetic or hereditary test but a genomic one (analysis of the genes of the tumor). Once the results of the test are reported to your treating physician, he or she will then discuss with you how this information could be used to deliver the best treatment for you. The purpose of this document is therefore to ask you if you agree to have a piece of your tumor, which will be routinely removed during your surgery, sent to the Netherlands in order for its gene expression pattern to be analyzed. Enough of your tumor tissue will also remain at your hospital for any additional tests you may need in the future. What will happen after the test? Once the results of both the pathological examination and gene expression test are known, your treating physician will discuss the following options with you: 1. If there are no tumor cells found in your axillary lymph nodes or if there are tumor cells in 1 to 3 lymph nodes, you will be offered the opportunity to participate in the research study mentioned above. a) you may decide to participate in the research study. b) you may decide not to participate in this research study in which case you may ask for the piece of your tumor tissue that was sent to the Netherlands to be destroyed or returned. c) you may decide not to participate in this research study but wish to donate that piece of your tumor tissue for other research studies not defined at the present time. 2. If tumor cells have been identified in more than 3 of your axillary lymph nodes the research study is not appropriate to your situation. You may decide to participate in a different study, appropriate for your situation, if there is such an opportunity in this hospital or elsewhere. Concerning the piece of your tumor tissue that was sent to the Netherlands: a) you may prefer that the piece of your tumor tissue that was sent to the Netherlands is destroyed or returned. b) you may decide to donate that piece of your tumor tissue for other research studies not defined at the present time. Donation of a blood sample for future research As well as deciding whether you would like to donate any remaining tumor tissue after the performance of the prognostic test for future research, we would like to ask you to donate a 20 ml (2 table spoons) blood sample. This blood sample would be taken during the study (preferably before surgery). It will be stored and used for future research projects to increase our understanding of cancer with a view to improving patient care. The blood sample will be used for both serum storage for proteomics analysis (looking at proteins) and whole blood storage for potential genetic research (looking at how your genes may have information about cancer risk). In the future, we hope that proteomics will help us predict if or when tumors will come back. Because proteomics is still a new type of research, your blood sample will not immediately be useful for seeing when and if your cancer may come back. However, it will help us to further develop this new technology, which will help other breast cancer patients in the future. Genetic research is also developing rapidly so we don t yet know the kind of research projects that may be possible. Your blood sample will be stored without personal information so that researchers will not know who you are and will be analyzed with many other samples to increase our understanding of cancer. Version / 152 May 21, 2014

270 The biological samples (blood and tumor), will be stored in a biological materials bank. The biological materials bank will be under the guardianship of an international, academic, non-profit organization. Please be informed that all future research projects must be approved by the relevant research and ethics committees before the samples can be used. What are my rights? If you consent to have a piece of your tumour tissue sent to the Netherlands, your privacy will be protected at all times. In agreeing to the genomic prognostic test your tumor tissue will not be used to test whether your breast cancer is hereditary (that is, breast cancer runs in your family). If you agree to an additional blood sample for research or to donate the remainder of your tumor tissue for future research, this may involve genetic research. There are no additional examinations, medical procedures, discomfort, risks or costs to you if you decide to give permission for a piece of the tumor that is removed during your operation to be sent for analysis to the Netherlands. If you agree to donate a blood sample collected for future research, this will probably be taken at the same time as a routine blood samples, in this case you will not need an additional needle prick/puncture. Regardless of whether you decide to have a piece of your left over tumor sent to the Netherlands or not, your relationship with your physician will in no way be affected. Consent is voluntary and can be withdrawn at anytime without you having to give a reason and without having any affect on your relationship with your treating physician and clinical team. If new information becomes available about the new test, you will be informed of this by your treating physician. After you have read this information carefully and you have discussed it with your treating physician, we kindly ask you to complete the enclosed informed consent form if you decide to have a piece of your tumor sent to the Netherlands. We thank you warmly for your attention. Version / 152 May 21, 2014

271 Informed consent form (Consent to send tumor tissue to the Netherlands for specialized analysis) I have received a copy of the Patient Information Sheet and have understood the information. I accept that a part of my left over tumor tissue will be sent to the Netherlands, where it will be analyzed using the new genomic prognostic test. The options that will be available to me after the test results are known have been clearly explained. I have had enough time to take my decision. My consent to send tumor tissue to the Netherlands is completely voluntary, and it will not affect my relationship with my treating doctor. The data collected on my behalf will be strictly confidential and treated according to the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the local applicable laws Please tick as appropriate: I would like to donate any remaining tumor tissue after the performance of the new test for future research. If the new genomic test is not appropriate for me or I decide not to participate in the proposed study, I would still like to donate the piece of my tumor tissue to research studies not defined at the present time. I accept to donate a blood sample for future research. If the new genomic test is not appropriate for me or I decide not to participate in the proposed study, I prefer that my tumor tissue that has been send to the Netherlands will be destroyed or returned. My consent does not discharge the organizers of the research from their responsibilities, and I keep all my rights guaranteed by the law. Patient's name: Patient's signature: Date: Investigator or person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Title/Position: This document has been prepared taking the following documents into account: World Medical Association Declaration of Helsinki, adopted by the 18th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, 2000 Version / 152 May 21, 2014

272 Appendix F: Patient information sheet and informed consent document for participation in the clinical trial INFORMATION FOR THE NATIONAL COORDINATING CENTER/GROUP OR NATIONAL COORDINATOR: This document is an English version of the patient information sheet & informed consent 1 (PIS & IC 1) for participation in the clinical trial. The translation and national regulatory submission process of this document is the responsibility of the National Coordinating Center/Group or the National Coordinator for this trial. The EORTC would like the NCC/G to: translate the patient information sheet and informed consent in preparation for the submission of the dossier to the ethics committee (the submission may be the responsibility of the NCC or the EORTC depending on the local regulations) - We would like you to adapt the text to national guidelines and sensibilities however you must tell the EORTC what you have changed and why and have our agreement on the new text. send a copy of the approved translated document to the EORTC Headquarters who will then incorporate it into the initiation package which will be sent to you to distribute to other national participating investigators. (When each center has changed the header to that of their individual hospital you will have to collect a paper version from each participating center and send these to the EORTC, i.e. a paper copy of each PIS & IC document that is in use for the trial in it s final form.) the final translated and approved PIS & IC 1 must have a version number and date which must be quoted in all correspondence with local and national ethics committees. Version / 152 May 21, 2014

273 EORTC Avenue E. Mounierlaan 83/11 Brussel 1200 Bruxelles België Belgique Tel: eortc@eortc.be Patient Information Sheet - Comparison of risk assessment methods in the trial 1. Introduction You have previously agreed for a part of your breast tumor to be sent to the Netherlands to look at the genes in your tumor (gene expression test). This was done after you signed a similar document (consent form) before your surgery. The gene expression test analyzed which genes were active or not in your tumor, and this gives us information about your prognosis (that is, the likelihood of your cancer coming back). Now we would like to invite you to participate in the clinical trial (a research study) called (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy). This study involves comparing the results of your gene expression test with the results obtained by the traditional method of looking at clinical and pathological information about your cancer. This includes your age, the size of your tumor, the way your cells look under the microscope (called grade ), and hormone receptors, which are like ears on cells that respond to hormones. The results of both the gene expression and the clinical-pathological tests are important because they will be used to decide if you will be proposed chemotherapy or not. The main objective of the trial is to determine which of the two types of tests (both called prognostic tests) gives us the best information necessary to decide which patients should receive chemotherapy. 2. Invitation to participate in the study Researchers in COUNTRY are initiating a clinical trial within the framework of an International research network* coordinated by NATIONAL GROUP/The EORTC* with patients that have a disease similar to yours. You are invited to take part in this clinical research project after having received full information about the study. (* details are provided in section 14) You are now being asked to consider taking part in this trial. You should make your decision after reading carefully the detailed information below and discussing any questions you have with your doctor or research nurse. This is so that you understand this trial and can choose what is best for you. If you decide to take part in this trial, you will need to sign the informed consent form which is included after the explanation of the trial. 3. How doctors decide what treatment is necessary for patients after surgery After breast cancer surgery, when the breast tumor and one or more of the lymph nodes from under your arm have been removed, doctors evaluate the risk of your cancer coming back. Traditionally, doctors do this by analyzing the clinical-pathological information already mentioned above: If the analysis of this information shows that your cancer has a high risk of coming back, you will be proposed chemotherapy. If your tumor has hormone receptors you will also be proposed endocrine therapy, often with tamoxifen (a treatment that stops the activity of certain hormones). Finally, if you had breast-conserving surgery (meaning your breast was not removed) you will also be proposed radiotherapy. These treatments - chemotherapy, endocrine therapy and radiotherapy - are called adjuvant treatments, adjuvant because they are additional treatments after surgery that can prevent cancer from coming back. Chemotherapy, as you have probably heard, is a treatment which Version / 152 May 21, 2014

274 can have unpleasant side effects. Some of these side effects are short term, but others can be long term. In recent years our understanding of the biology of breast cancer has improved. We have learned that chemotherapy is only necessary and effective for breast cancer patients that have tumors with a high risk of coming back. Unfortunately, up to now, we have not always been able to identify these aggressive tumors. Because of this uncertainty, doctors tend to treat most patients with chemotherapy in order to not to risk missing someone who really will benefit from it. However, this leads to over-treatment. In other words, at the moment, some women receive chemotherapy without needing it because in reality their tumors are less aggressive than we think. The most important goal of the trial is to be able to more precisely identify which tumors are aggressive. By doing so, in the future only women who will really benefit from chemotherapy will receive it; and women who do not need this therapy can avoid its unpleasant and sometimes long term side effects. 4. A new way of making decisions about chemotherapy: a new prognostic test Tests that doctors use to get information about tumor aggressiveness are called prognostic tests or tools because they help make predictions about how likely it is that a cancer will come back. In the sections above, the traditional prognostic test, looking at clinical-pathological information has been described. Recently, scientists have developed a new and different way to analyze tumor aggressiveness and make predictions about prognosis. This new prognostic test is not based on the traditional clinical-pathological factors already described, but looks at the activity of 70 specific genes ( gene expression ) in your tumor using a technology called microarray technology. The trial will examine this new prognostic test. However, in addition to looking at the 70 genes that we think may give us the most information about your tumor, we will also look at the expression of all the other genes in your tumor. This is like having a fingerprint or an identity card of your tumor, and this may help us get additional information about how aggressive the cancer might be. It is important to understand that this new prognostic test, also called a genomic test, will look only at the genes in your tumor. It will not look at your own genes, that is, the ones that you pass on to your children. A test of your own genes is called a genetic test and provides hereditary information, for example, whether cancer (or other diseases) runs in your family. Again, the new prognostic test in is not a genetic test, but a genomic one. In using the new genomic prognostic test we hope to be able to better identify which women really need chemotherapy and which women can safely avoid this treatment without any threat to their long-term survival. We also hope to be able to better identify the minority of women who would not normally receive chemotherapy based on their clinical-pathological prognosis but for whom this treatment may be necessary to prevent their tumor from coming back. 5. Description of the clinical trial will test whether the new prognostic tool can help better identify which women need chemotherapy. To do so, the new genomic test must be compared to the traditional way of assessing the risk of the cancer coming back. In other words, if you participate in this study, your tumor s aggressiveness will be assessed by using both the common clinical-pathological criteria and the genomic test just described above. The results from the two types of tests will then be compared. For the vast majority of women (estimated to be two thirds) the two tests are expected to show the same results, meaning that the test results would agree about whether you should be proposed chemotherapy or not. Women whose tumors are shown to be aggressive and have a high risk of recurrence by both tests will be proposed chemotherapy (see diagram 1 below, number 1) and women whose tumors are shown to be less aggressive and are at low risk of recurrence by both methods will not be proposed chemotherapy (see diagram 1 below, number 2). For a minority of Version / 152 May 21, 2014

275 women (estimated to be one third), we expect that the results of the two types of tests will not agree, that is, they will be discordant (see diagram 1 below, number 3). Diagram 1. Comparison of the two risk assessment methods Node negative and 1-3 positive lymph node breast cancer (1) (3) Genomic : Clinical (2) High : High risk risk Discordant Low : Low risk risk Genomic : Clinical Genomic : Clinical High : Low risk risk Low : High risk risk Genomic : Clinical Patients in for whom the test results are discordant will participate in the part of the study comparing the two methods. These women with discordant risk assessment results will have which of the two methods used for determining their treatment chosen by chance (randomly) by a computer, this is called randomization. Neither you nor your doctor can choose which of the two treatments you will receive. By participating in this randomization, it means that you might be proposed chemotherapy according to your clinical-pathological test, even if the genomic test results would not recommend that you are proposed chemotherapy. Or the opposite might happen. You might not be proposed chemotherapy according to the clinical-pathological test, although the genomic test suggests you should have it. The various possibilities are summarized in Diagram 2 below. Your doctor or research nurse can help explain this to you. Diagram 2. Decide with clinical tool Randomization of discordant cases High clinical risk (Low genomic risk) - Chemotherapy Low clinical risk (High genomic risk) - No chemotherapy R High genomic risk (Low clinical risk) - Chemotherapy Decide with genomic tool Low genomic risk (High clinical risk) - No chemotherapy If the genomic tool is proven to be better than the traditional clinical pathological methods of risk assessment, we estimate that 10-20% fewer women will receive chemotherapy. So in this study approximately one in ten women who would normally receive chemotherapy will be safely spared this treatment. In addition, we believe that the new test can also identify those women who have a low clinical risk but who would nevertheless require chemotherapy so one in twenty women who Version / 152 May 21, 2014

276 would normally not receive chemotherapy will receive it in order to prevent their breast cancer from coming back. The trial will include 6,600 women with a similar disease to yours over a period of about 5 years or slightly more. The total time that you will receive treatment, depends on what is decided to be best for you. If you receive only chemotherapy, it will last for about 6 months. If you receive endocrine therapy, which has fewer side effects than chemotherapy and is a different type of treatment, this will last for 7 years. It is also possible that you will receive both types of therapy. As a patient in the trial you will also have regular visits with your study doctor or nurse for at least 7 years. All women who participate in the trial should be under effective contraception while taking trial medication. Your doctor will discuss with you which methods constitute effective contraception. 6. Description of the ultimate goal of the clinical trial The main goal of is to show that the new genomic prognostic test can help better identify which women require chemotherapy than the traditional clinical-pathological method without having a negative effect on a patient s long-term survival. It also aims to study the relationship between gene expression patterns and the way breast cancer develops. Understanding this relationship will help us to be better able to give treatments designed specifically for each individual patient s breast cancer. There are additional, longer-term research objectives that depend on the collection and banking of tumor tissue and blood samples from all participants in. 7. Description of foreseeable risks We do not know if the new prognostic test is better than the clinical-pathological method that we currently use - and that s why we are doing this trial. On the one hand, because your treatment was decided using the new genomic test, there might be less chance of your cancer coming back, you might not be treated with chemotherapy unnecessarily and you may live longer. On the other hand, might show that the new tool is less good than the current clinical-pathological method and, as a result, you may end up having received insufficient treatment. Although we are fairly confident that this is not the case, we are not completely sure. So there is a risk that by decreasing the amount of chemotherapy prescribed, some women may be under-treated, therefore increasing the chance that their cancer will come back. In order to quickly detect if this happens, the group of women who will not receive chemotherapy according to the genomic test but who would normally have received it according to the clinical-pathological test (an estimated 672 women of the total 6,600 participants) will be very closely monitored. However, as mentioned above, we feel confident that the genomic tool is quite accurate in identifying high and low risk patients. This test has been pre-tested in small studies using tumors from women that had their tumors removed about ten years ago. The results of these studies involving women from several different countries showed that the genomic test did predict which cancers would come back and which would not. Please consider that even if you receive all possible treatments, there is always a chance that your tumor may come back. If your cancer does come back at any time, regardless of whether you received chemotherapy or not, you will be given all necessary treatment according to the standard care of your hospital. This may include chemotherapy. Version / 152 May 21, 2014

277 8. Expected benefits If you decide to participate in this trial there will be no financial or material compensation to you. No one knows if you will benefit personally from taking part in this study. However, your participation is very important to create knowledge of how to treat breast cancer. We have learned a lot about how to treat breast cancer because of the millions of women who have volunteered to participate in breast cancer clinical trials all over the world through the years. New relevant information that directly concerns your future health will be made available from your treating doctor at the institution where you have received treatment in this study. 9. Voluntary participation Your participation in the trial is entirely voluntary and you will be given enough time to make your decision. You are free to decide at anytime that you no longer wish to participate in the trial. You do not have to give any reasons for doing so. Leaving the trial will not affect your future treatment. It will also not have any negative effect on your relationship with your treating doctor or the hospital staff. The only information that we will keep for research and analysis is what has been collected about you until the time you decide to stop participating in the trial. If you wish you can ask for any of your tissue that has not been used to be destroyed. This patient information document and consent form invites you to participate in the main part of. It specifically asks you to participate in the comparison of the genomic and clinicalpathological risk assessment methods. In the near future, we may invite you to participate in one or two other parts of the trial, depending on the treatment that is appropriate for you. If you are proposed chemotherapy, we will invite you to participate in a part of that compares two different types of chemotherapy. If your tumor cells contain hormone receptors, we will invite you to participate in the part of that compares two different endocrine therapies. The details of these other parts of and any potential benefits and risks, will be explained to you at the appropriate time by your doctor or research nurse. At that time, you will also receive separate patient information sheets and informed consent forms. We recognize that this is a long and possibly confusing process, but it is necessary in order to be sure that every patient entering the trial does so on the basis of having received proper information about every aspect of the study. 10. Data (information) and legal protection If you decide to participate in this part of, your privacy will be protected at all times. All information (personal, clinical, economic, and data from research on tumor material and blood samples) collected because of your participation in the trial will be treated according to all national applicable laws. However, your consent does allow for the information in your medical records to be linked to data coming from other sources such as cancer registries. Cancer registries are a way for countries to collect information about cancer in general, & to help to better understand, prevent and treat cancer. It is very important that all information collected in this trial is accurate. Therefore, from time to time, this collected information may be checked against your medical records. Only authorized persons (EORTC research staff and their representatives, national and/or foreign health authority representatives) may have access to your medical records. Except for access to your information by such authorized persons, all information about you will be strictly confidential. Tumor and blood samples (biological material) Some of the pathological information about your tumor determined by the doctors in your hospital will also be analyzed by other pathologists who are experts in breast cancer. In, this Version / 152 May 21, 2014

278 group of experts will review a small fragment of some of the tumors of the 6,600 women participating in this trial. This is called a central pathology review and will be done in Milan, Italy. The purpose of this review is to ensure that is run according to the highest scientific standards possible. The handling of tumor and blood samples in will always be done in such a way that scientists analyzing it for research purposes will not be able to know your name or other personal information about you. The results of the research on biological material are unlikely to be available in the immediate future. This is because research can take a long time and tissue and blood samples and information must be collected from many patients before the results are known. Collaboration with third parties The mission of the EORTC and BIG, being non-profit research organizations, is to do everything they can in the best interests of cancer patients. Collaboration with third parties, including commercial companies, may be necessary for the EORTC and BIG to develop more effective anticancer treatments. This may lead to a company owning a discovery made using the biological materials bank. If the EORTC or BIG ever receive any money as a result of such a discovery, it will be reinvested in non-commercial cancer research to improve cancer care. The EORTC has asked your treating doctor to disclose any existing conflict of interest he/she may have as a result of his/her activities related to this trial. The EORTC has set up procedures to ensure the integrity of this process. 11. Insurance The EORTC has obtained clinical trial insurance in accordance with all applicable laws. If you need to undergo other medical treatment, we advise you to inform the doctor treating you in to ensure that the other medical treatment will not have any effect on your participation in the trial. Everything has been done and will continue to be done to prevent additional health problems occurring as a result of your taking part in this trial. (Specific clause per country following national law) 12. Ethics Committee A complete description of the research (the research protocol for ) has been submitted to the responsible ethics committee. The mission of this committee is to verify that all conditions with respect to your safety and rights are respected. Approval for this research has been given by the Ethics Committee of on Version / 152 May 21, 2014

279 13. Contact persons In case of any problem or question, your doctor will be pleased to answer any further questions and may be contacted as follows: Name of the doctor: Hospital: Telephone: If you give your consent to join this trial, you will be given a telephone number of the hospital that you can contact at any time if you feel unwell or have further questions. With your agreement, if you have a family doctor we will also inform him or her about your taking part in this trial and what is involved Please take your time to read and think about this information, and do not hesitate to ask further questions to your doctor or research nurse if anything is unclear. You should keep a copy of this document after you and your doctor have signed it. 14. Responsibilities The clinical trial will be managed, coordinated and analyzed by the European Organisation for Research and Treatment of Cancer (EORTC) which will act as the European legal sponsor of the trial. The EORTC Breast Group (a group of European cancer specialists interested in improving breast cancer treatment) is the coordinating group however in (COUNTRY) a group of breast cancer specialists from (NATIONAL GROUP/EORTC) will be responsible for the management the study. The EORTC is itself part of the non-profit, international academic network for breast cancer research called the Breast International Group (BIG), which has a laboratory research arm called TRANSBIG. The trial is being run as a joint project of the EORTC, BIG / TRANSBIG, and the participating national groups. This means that is being conducted as an international collaborative study under the legal framework of the EORTC and BIG. This research receives support through financial and other contributions from the European Commission s Framework Programme VI; the companies Novartis, F. Hoffmann-La Roche, Sanofi-Aventis, Agendia; and national charities. Version / 152 May 21, 2014

280 INFORMED CONSENT FORM (Consent to participate in comparison of risk assessment methods in the trial) I have received a copy of the patient information and have understood the information. All my rights have been clearly explained to me. I have had enough time to take my decision. I agree to participate in the clinical trial called ( Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid Chemo Therapy. A prospective, randomized study comparing the 70-gene signature with common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0-3 positive nodes), EORTC study number and BIG study number BIG My participation is completely voluntary. I accept that data resulting from this study can be linked to other sources (such as cancer registries) for research purposes. I understand that I have the right to withdraw my consent at any time without explanation. I also understand that if I withdraw my consent, it will not affect my relationship with my treating doctor. I understand that all the information collected on my behalf will be strictly confidential and treated according to the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the local applicable laws Please tick as appropriate: I accept that future cancer research studies may be conducted on the biological material (tumor or blood) that I provide and that any data resulting from these studies can, in the future, be linked with other resources for research purposes. My consent does not discharge the organizers of the research from their responsibilities and I keep all my rights guaranteed by the law. Patient's name: Patient's signature: Date: Investigator or person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Title/Position: This document has been prepared taking the following documents into account: World Medical Association Declaration of Helsinki, adopted by the 18th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, Version / 152 May 21, 2014

281 Appendix G: Patient information sheet and informed consent document for the clinical trial chemotherapy section. INFORMATION FOR THE NATIONAL COORDINATING CENTER/GROUP OR NATIONAL COORDINATOR: This document is an English version of the patient information sheet & informed consent 2 (PIS & IC 2) for the chemotherapy therapy study in the clinical trial. The translation and national regulatory submission process for this document is the responsibility of the National Coordinating Center/Group or the National Coordinator for this trial. The EORTC would like the NCC/G to: translate the patient information sheet and informed consent in preparation for the submission of the dossier to the ethics committee (the submission may be the responsibility of the NCC or the EORTC depending on the local regulations) - There are sections which you must NOT change the meaning of the text in the translation; these are sections 1, 2, There are certain sections where you can adapt the text to national guidelines and sensibilities however you must tell the EORTC what you have changed and have our agreement on the new text. The sections that can be adapted are 3-7. send a copy of the approved translated document to the EORTC Headquarters who will then incorporate it into the initiation package which will be sent to you to distribute to other national participating investigators. (When each center has changed the header to that of their individual hospital you will have to collect a paper version from each participating center and send these to the EORTC, i.e. a paper copy of each PIS & IC document that is in use for the trial in it s final form.) the final translated and approved PIS & IC 2 must have a version number and date which must be quoted in all correspondence with local and national ethics committees. Version / 152 May 21, 2014

282 EORTC Avenue E. Mounierlaan 83/11 Brussel 1200 Bruxelles België Belgique Tel: eortc@eortc.be Chemotherapy question in trial (Randomization-Chemotherapy) You have previously agreed for a part of your breast tumor to be sent for special analysis (screening informed consent) and consented to be included in the clinical trial called where the new test has been compared to current clinical risk assessment (informed consent 1). In accordance with the results of your risk assessments you will shortly receive chemotherapy. We would like to invite you to participate in the chemotherapy question of the study, described below. 1. Title of the research protocol As part of the - Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy - trial (A prospective, randomised study comparing the 70-gene signature with common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0-3 positive nodes) the following chemotherapy therapy question will be asked. Can anthracycline-based chemotherapy be safely replaced by the combination of Docetaxel- Capecitabine as adjuvant treatment of early breast cancer? 2. Invitation to participate in the study The EORTC Breast Group (a group of cancer specialists interested in improving breast cancer treatment) is initiating a clinical trial with patients that have a disease similar to yours. The study will be conducted at the International level under the supervision of doctors recognized as experts in this field of medicine. You are invited to take part in this clinical research project after having received full information about the study and the implications for you as a patient. 3. Introduction Your breast tumor was removed by surgery however there may be a possibility that it may come back, this is called relapse. Therefore current medical opinion recommends the use of chemotherapy (called adjuvant chemotherapy) to try to prevent your cancer from coming back. Current standard care consists of a combination of several chemotherapy drugs, which constitutes a regimen, and almost always contains a drug called anthracycline. This is the most active drug of this combination but it is also quite toxic and can cause not only short term toxicity during treatment but also long term side effects that may only appear in the future such as leukemia and heart disease. Although this risk is small it exists and these diseases are serious. New drugs active against breast cancer have recently been developed and tested in other studies showing apparently fewer long term side effects. The present study has been designed to test a combination of two of these drugs (docetaxel and capecitabine) by comparing the new combination to the current standard treatment and to detect if it has an additional benefit in terms of efficacy (increase duration of life without the cancer coming back) and safety (fewer long term side effects). All women who participate in the trial should be under effective contraception for the entire duration of the study. Your doctor will discuss with you which methods constitute effective contraception. Version / 152 May 21, 2014

283 4. Description of the research This research will involve approximately 4,000 patients in several countries throughout the world but principally in Europe. You will receive either docetaxel-capecitabine chemotherapy (chemotherapy A the new regimen) or an anthracycline based chemotherapy (chemotherapy B the standard regimen) at doses that have been well studied in other research studies, and that are commonly used internationally. The main objective of this study is to compare the efficacy and safety of chemotherapy A to the efficacy and safety of chemotherapy B (by counting and comparing the number of relapses in patients treated with each chemotherapy regimen). The duration of treatment A and B will be similar, i.e. six or eight cycles with a total duration of approximately five months. The side effects are different and are detailed below. Both chemotherapy treatments are effective against breast cancer, but we don t know if one is better than the other in your particular situation. Treatment Selection No one knows which treatment is better for you therefore which of the two treatments you will receive will be decided by chance. This is called randomization. Your doctor will contact a central statistical office, which will assign one of the two treatments to you. Your chances of getting either the new chemotherapy or the standard chemotherapy are equal (50% either way) neither you nor your doctor can choose the treatment. Chemotherapy A Docetaxel is a very active drug in breast cancer either alone or in combination with Capecitabine. You will receive 6 cycles of Docetaxel in combination with Capecitabine. Docetaxel will be given intravenously in the day care unit once every 3 weeks (infusion in your veins over one-hour in the day care unit). Capecitabine will be given in pill form, which must be taken twice a day for 14 consecutive days followed by 7 days with no medication (rest period). This set of 21 days forms one cycle. Both Docetaxel and Capecitabine will be provided free of charge. Chemotherapy B Depending whether your breast cancer was lymph node-negative or -positive, the chemotherapy options are different. As it is considered that patients needing chemotherapy may be at a higher risk when they are node-positive, the chemotherapy for these patients is more aggressive. The chemotherapy options in this arm are well know and are the standard treatment for patients in your situation. Chemotherapy B for node-negative patients The four drugs that can be used in different combinations are 5-Fluorouracil (5-FU), Cyclophosphamide, Epirubicin and methotrexate. Sometimes, Doxorubicin is used instead of Epirubicin. The mode of delivery of this chemotherapy regimen may vary slightly from one hospital to the other according to the standard of care applied in each hospital. The drugs will be given intravenously in the day care unit usually once every 3 or 4 weeks (it will take about 2 hours in the day care unit). In some regimens the treatment is given for 2 consecutive weeks followed by a two week rest period. We plan to give you 6 or 8 cycles (one cycle is 3 or 4 weeks) of this chemotherapy regimen. Version / 152 May 21, 2014

284 Chemotherapy B for node-positive patients. The option here is a chemotherapy (called FEC D) consisting of 3 cycles of a combination of fluorouracil, cyclophosphamide and epirubicin, followed by 3 cycles of docetaxel. This chemotherapy is given on one day every 3 weeks, up to a total of 6 cycles. If you participate in this study you will not have to make more visits to hospital, nor undergo more examinations or blood samples during the chemotherapy period and after completion of the study than patients treated for the same disease who are not taking part in the study. Results of the study will not be available until after the study is finished (recruitment of patient is expected to finish in 2010 and the study to have initial results 2013), but an individual patient will know on a regular basis the effectiveness and outcome of her treatment. 5. Description of foreseeable risks and discomforts As with most forms of chemotherapy it is likely that you will experience at least some side-effects, although the extent of these varies considerably from patient to patient. During your treatment, you are at a slightly higher risk of developing an infection. If at any time during chemotherapy you develop a fever ( 38 C when taken under the arm), you must contact your doctor (see phone number at the end of this sheet). During this treatment, you may feel tired. You may need to adjust your daily activities to help cope with this. Before having chemotherapy, you should let your physician know all of the medications you are taking, including over-the-counter medicines and any herbal supplements. Chemotherapy A Side effects you might experience with docetaxel When used as a single agent, the major toxic effect of docetaxel is neutropenia (decrease in the numbered white blood cells). It may be associated with fever (called febrile neutropenia) and infection. Other toxic effects may include allergic type reactions and skin reactions, nausea, vomiting, oral mucositis (inflammation of a mucous membrane), diarrhea, reversible paresthesia (skin sensation, such as burning, prickling, itching, or tingling), alopecia (loss of hair), fatigue, neurosensory symptoms, mild local venous reactions (phlebitis) at site of injection and fluid retention/edema. During these 6 cycles a treatment with oral corticosteroids will be given before each infusion of docetaxel and continued daily for 2 more days, as it has been shown to decrease the risk of allergy and fluid retention (edema) due to docetaxel. All these side effects are manageable with appropriate medical intervention, dose reduction and/or treatment interruption. Side effects you might experience with capecitabine Capecitabine is generally well tolerated. The most common side effects are hand-foot syndrome and diarrhea. Hand-foot syndrome is characterized by varying degrees of redness, tenderness, pain and desquamation (shedding/peeling off of skin) of the palms of the hands and the soles of the feet; numbness and tingling may also appear and in its most severe form it can interfere with daily activities. Nevertheless, it is reversible and responds to the diminution of the dose or the interruption of the treatment. If hand-foot syndrome occurs it is helpful to use a moisturizing cream and sometimes vitamin B6 supplements; your doctor will give you specific details if necessary. Diarrhea may also be an important side effect and like dermatitis, responds to changes in the treatment dose or schedule. Other side effects that may be associated with the use of capecitabine are: nausea, vomiting, stomatitis (inflammation of the mucous tissue of the mouth), dyspepsia (indigestion)/abdominal pain, fatigue, anorexia, myalgia (muscular pain or tenderness), insomnia, fever, myelosuppression Version / 152 May 21, 2014

285 (decrease in white blood cells and platelets), paresthesia (skin sensation, such as burning, prickling, itching, or tingling), eye irritation and serum (blood) biochemistry alterations. Other skin toxicities apart from the hand-foot syndrome include rash, dry and/or itchy skin. Before taking capecitabine, tell your doctor if you are allergic to capecitabine, fluorouracil, or any other drugs, what prescription and nonprescription medications you are taking, especially anticoagulants, antacids, folic acid, leucovorin, phenytoin (Dilantin), aspirin and vitamins. Also do not have any vaccinations (e.g., measles or flu shots) without talking to your doctor. Side effects you might experience with the combination capecitabine-docetaxel When capecitabine is used in combination with docetaxel, the incidence of neutropenia/neutropenic fever, gastrointestinal side effects and hand-foot syndrome may be higher, but still manageable with appropriate medical intervention, dose reduction and/or treatment interruption. Chemotherapy B Chemotherapy B for node-negative patients General Side-effects with the drugs combination This combination of drugs has been used in breast cancer treatment for many years and short term and long term side effects are well known and documented. Side effects you might experience with anthracyclines (Epirubicin / Doxorubicin) The most common side effects are hair loss (alopecia), nausea, vomiting, myelosuppression (increasing the risk of infection, bruising, bleeding or anemia), sore mouth or taste change, discolored urine, darken skin, sensitivity to the sun, tiredness. All these side effects are still manageable with appropriate medical intervention, dose reduction and/or treatment interruption. Sometimes anthracyclines also have deleterious effects on the heart muscle decreasing the capacity of the muscle to contract correctly. Accordingly, this treatment is only delivered to women with no cardiovascular problems and your cardiac condition will be evaluated before the treatment. For your information, scalp cooling at the time of treatment can reduce hair loss in some cases. This may be available to you. However, you must expect some loss of hair. In case of severe hair loss a wig will be provided. It is important to note that your hair will grow back after your treatment is finished. Side effects you might experience with cyclophosphamide This drug is responsible for a transitory myelosuppression, with a decrease in white blood cells and platelets (increasing the risk of infection, bruising or bleeding). You will be carefully monitored with regular blood test and your treatment will be modified if necessary. Side effects you might experience with the combination of anthracyclines & cyclophosphimide This combination of drugs is also known to provoke secondary leukemia occurring usually 3 to 4 years after the end of the treatment. This is a rare side effect and globally (all patients taken together) the benefit of administering this drug is largely superior to the risk undertaken. Side effects you might experience with 5-Fluorouracil (5-FU) Following intravenous infusion of this drug, diarrhea, stomatitis (inflammation of the mucous tissue of the mouth) and conjunctivitis (inflammation of the conjunctiva of the eye) may occur. These side-effects are usually mild to moderate. In case of a severe side-effect your doctor will reduce the dose of this drug for following cycles. While using this medicine, and for 1 or 2 months after you stop using it, your skin may become more sensitive to sunlight than usual and too much sunlight may increase the effect of the drug. During this period of time: Version / 152 May 21, 2014

286 - Stay out of direct sunlight, especially between the hours of 10:00 a.m. and 3:00 p.m., if possible. - Wear protective clothing, including a hat and sunglasses. - Apply a sun block product that has a skin protection factor (SPF) of at least 15. Some patients may require a product with a higher SPF number, especially if they have a fair complexion. If you have any questions about this, check with your health care professional. - Do not use a sunlamp or tanning bed or booth. Side effects you might experience with methotrexate When taking methotrexate, the following side-effects may be observed : bruising or bleeding, anaemia (low number of red blood cells), sore mouth and taste change, diarrhea, tiredness and a general feeling of weakness, skin changes (your skin may darken, due to excess production of pigment; this usually returns to normal a few months after the treatment finishes), gritty eyes. All drugs in this chemotherapy combination will be prescribed as they would be normally by your doctor following the standard of care applicable in the hospital. Chemotherapy B for node-positive patients. For the first 3 cycles of FEC, the side effects are described above. For the following 3 cycles of docetaxel, please refer to the section describing docetaxel toxicity in Chemotherapy A. 6. Expected benefits If you accept to participate in this study we cannot be sure that there will be a direct benefit for you. However you will receive either the standard chemotherapy treatment or the new regimen which has already been extensively tested in the metastatic setting. If your disease progresses during the treatment allocated to you by randomization there is a high probability that you will be treated with the drugs of the other arm of this study (if there is no medical reason why you should not be) but outside the context of this clinical trial. If new information about the novel drug combination (docetaxel & capecitabine) becomes available it will be communicated to you by your treating doctor. As will any information that might affect your willingness to continue participation in the trial. We hope this research will teach us more about cancer. This will enable us to improve the standard of treatment and so help other patients with cancer in the future. However, nobody can predict whether you will directly benefit from participating in this clinical study. The anticipated benefits of this study for the breast cancer community include a better understanding of cancer and finding a medication regimen that has as good or better ability to control breast cancer with lower side effects than the commonly used regimen. 7. Voluntary participation Your participation in this clinical trial is entirely voluntary and you will be given sufficient time to decide whether or not you wish to participate. You are free to decide at any time without giving any reason that you no longer wish to participate in the trial. Such decision will not affect your subsequent treatment or relationship with your treating doctor or the hospital staff in any way. Medical data collected during your participation to the clinical trial as well as follow up data which will still be collected afterwards will be kept for research and analysis unless you specify otherwise. If you consent to join this trial, you will be given a telephone number at the hospital that you can contact at any time if you feel unwell or have further questions. We encourage you to take time to consider participation in this trial and to discuss it with your family, friends and your family doctor. 8 Data protection Your consent for participation in this part of the trial also includes your consent to allow the use of the data in your medical/clinical record to be used for research purposes. Your Version / 152 May 21, 2014

287 consent also includes allowing this data to be linked to data coming from other sources (such as cancer registries, medical/clinical records, ). All data (personal, clinical, economic and data coming from research on biological material) collected on your behalf will be treated in compliance with the national applicable laws. It is very important that the information collected is accurate and therefore, from time to time, this collected information may be checked against your medical records. Duly authorized persons (EORTC research staff and their representatives, national and/or foreign health authority representatives) may have access to your medical records. With the exception of access by the duly authorized persons to your personal data on your medical record, all information will be strictly confidential. This clinical research project is conducted under the legal framework of the EORTC with partial financial contribution of the European Commission, national charities and pharmaceutical companies. The EORTC, which is responsible for the conduct of this trial, has asked your treating doctor to disclose any existing conflict of interest he/she may have as a result of his/her activities related to this trial. The EORTC has set up procedures to ensure the integrity of this process 9. Insurance The EORTC who act as legal sponsor of the Study has obtained a clinical trial insurance in accordance with the applicable legislation. If you need to undergo another medical treatment, we advise you to inform the study doctor to ensure this will not have any effect on your participation to the trial. Everything has been done and will continue to be done to prevent additional health problems occurring as a result of your taking part in this trial. 10. Ethics Committee This research protocol has been submitted to the ethics committee whose mission is to verify that all conditions with respect to your safety and rights are respected. Approval to this research has been given by the Ethics Committee of on 11. Contact persons In case of any problem or question, your doctor will be pleased to answer any further questions and may be contacted as follows: Name of the doctor: Hospital: Telephone: If you consent to join this trial, you will be given a telephone number of the hospital that you can contact at any time if you feel unwell or have further questions. With your agreement, your family doctor (if applicable) will also be informed about your taking part in this trial and what is involved, if you agree. Please take your time to consider this information and do not hesitate to ask further questions to your doctor if anything is unclear. You are entitled to keep a copy of this document after you and your doctor have signed it. Version / 152 May 21, 2014

288 Informed consent I have been properly informed about the clinical research study and have been given sufficient time to consider my participation. I have received a copy of the patient information sheet. All my rights have been clearly explained to me. I agree to participate in the clinical research study entitled - Microarray In Nodenegative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy in the chemotherapy therapy question titled Can anthracycline-based chemotherapy be safely replaced by the combination of Docetaxel-Capecitabine as adjuvant treatment of breast cancer with 0-3 positive nodes? and to be enrolled in EORTC study number BIG I accept that any data resulting from this clinical research study can be linked with other resources for research purposes. My participation is completely voluntary and I have the possibility to withdraw my consent at any time without explanation. This will not affect my relationship with my treating doctor. The data collected on my behalf will be strictly confidential and treated according to the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the local applicable laws. I have been informed that the data (personal, clinical and biological material) collected may be used in the future for cancer scientific research purposes while confidentiality will be ensured. All data (personal, clinical and research on biological material) collected on my behalf will be treated in compliance with the "European Union Directive (Dir/95/46/EC) on the protection of individuals with regard to the processing of personal data and the national applicable laws. My consent does not discharge the organizers of the research from their responsibilities and I keep all my rights guaranteed by the law. Patient's name: Patient's signature: Date: Investigator or person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Title/Position: This document has been prepared taking the following documents into account: World Medical Association Declaration of Helsinki, adopted by the 18th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, Version / 152 May 21, 2014

289 Appendix H: Patient information sheet and informed consent document for the clinical trial endocrine therapy section. INFORMATION FOR THE NATIONAL COORDINATING CENTER/GROUP OR NATIONAL COORDINATOR: This document is an English version of the patient information sheet & informed consent 3 (PIS & IC 3) for the endocrine therapy study in the clinical trial. The translation and national regulatory submission process of this document is the responsibility of the National Coordinating Center/Group or the National Coordinator for this trial. The EORTC would like the NCC/G to: translate the patient information sheet and informed consent in preparation for the submission of the dossier to the ethics committee (the submission may be the responsibility of the NCC or the EORTC depending on the local regulations) There are sections which you must NOT change the meaning of the text in the translation; these are sections 1, 2, There are certain sections where you can adapt the text to national guidelines and sensibilities however you must tell the EORTC what you have changed and have our agreement on the new text. The sections that can be adapted are 3-7. send a copy of the approved translated document to the EORTC Headquarters who will then incorporate it into the initiation package which will be sent to you to distribute to other national participating investigators. (When each center has changed the header to that of their individual hospital you will have to collect a paper version from each participating center and send these to the EORTC, i.e. a paper copy of each PIS & IC document that is in use for the trial in it s final form.) the final translated and approved PIS & IC 3 must have a version number and date which must be quoted in all correspondence with local and national ethics committees. Version / 152 May 21, 2014

290 EORTC Avenue E. Mounierlaan 83/11 Brussel 1200 Bruxelles België Belgique Tel: eortc@eortc.be Endocrine question in (Randomization endocrine-therapy) You have previously agreed for a part of your breast tumor to be sent for special analysis (screening informed consent) and consented to be included in the clinical trial called where the new test has been compared to current clinical risk assessment (informed consent 1). In accordance with the results of your risk assessments you may or may not have received chemotherapy. Your tumor expressed hormone receptors and therefore current clinical practice recommends that you receive endocrine therapy. We would like to invite you to participate in the endocrine therapy question of the study, described below. 1. Title of the research protocol As part of the - Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy - trial (A prospective, randomised study comparing the 70-gene signature with common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0-3 positive nodes). Is 7 years of upfront aromatase inhibitor, letrozole, more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? 2. Invitation to participate in the endocrine therapy section of the study The EORTC Breast Group (a group of cancer specialists interested in improving breast cancer treatment) is initiating a clinical trial with patients that have a disease similar to yours. The study will be conducted at the International level under the supervision of doctors recognized as experts in this field of medicine. You are invited to take part in this clinical research project after having received full information about the study and the implications for you as a patient. 3. Introduction Even though your breast cancer was removed by surgery there is a chance that it may come back (called relapse). Your tumor expressed (produced) hormone receptors which capture hormones produced by your body and thereby can stimulate the growth of any remaining tumor cells. Therefore current medical opinion recommends that you receive hormone therapy (which reduces or stops the signal between your body s hormones and their receptors) to help prevent your cancer from coming back. The old standard treatment for you would be 5-years of hormonal/endocrine therapy with a drug called tamoxifen. Recent results suggest that a new class of hormone therapy drugs (Aromatase Inhibitors) may be more effective than tamoxifen at preventing cancers, like yours, from coming back after surgery (treatment after surgery is called adjuvant therapy). However the ideal length of treatment or whether these drugs should be given following tamoxifen or instead of tamoxifen is not yet known. One of these new aromatase inhibitor drugs is called letrozole. As a part of the trial your doctor would like to ask you to participate in a study that will test whether 7 years of letrozole is more effective and safe than 2 years of tamoxifen followed by 5 years of letrozole as adjuvant treatment of breast cancer. Aromatase inhibitors are better studied in post-menopausal women with a disease similar to yours. However pre-menopausal women can also be invited to participate in this study at their doctor s discretion and as long as they agree to have the production of estrogen by the ovaries suppressed (ovarian function suppression) while they take the hormonal therapy i.e. for a maximum of 7 years. Version / 152 May 21, 2014

291 Ovarian function suppression can be achieved either reversibly or irreversibly and your doctor will discuss the various options with you. All women who participate in the trial should be under effective contraception for the entire duration of the study. Your doctor will discuss with you which methods constitute effective contraception. 4. Description of the research The endocrine therapy part of this clinical trial will test a new regimen of adjuvant hormonal therapy which is given after surgery for breast cancer and is aimed at preventing any tumor cells still left in your body from developing and growing in your breast or in another part of your body. In addition to adjuvant hormone therapy you may also receive (or have already received) chemotherapy or radiotherapy. You have been asked to participate in a research study involving about 3,500 women throughout the world but predominantly in Europe. The purpose of this study is to find out whether it is better to receive a new drug, letrozole (registered as Femara ), for 7 years or tamoxifen for 2 years followed by letrozole for 5 years. To determine which treatment is better, half of the patients in the study will receive letrozole for 7 years and the other half will receive 2 years of tamoxifen followed by 5 years of letrozole. Treatment Selection No one knows which treatment is better for you therefore which of the two treatments you will receive will be decided by chance. This is called randomization. Your doctor will contact a central statistical office, which will assign one of the two treatments to you. Your chances of getting either 7 years of letrozole or 2 years of tamoxifen followed by 5 years of letrozole are equal (50% either way), neither you nor your doctor can choose the treatment. Description of Treatment If you agree to take part in this study, you will get one of the following treatments: -7 years of letrozole (2.5 mg). This is a tablet that you will take once a day, by mouth, every day for seven years. - 2 years of tamoxifen (20 mg) followed by 5 years letrozole (2.5 mg). You will take a tablet of tamoxifen (20 mg) once a day, by mouth, for the first 2 years and after that a tablet of letrozole (2.5 mg) once a day, by mouth, every day for five years. Letrozole will be provided free of charge and tamoxifen will be prescribed by your doctor as it would normally be following the standard of care. Other Treatment Choices If you do not take part in this study, other choices are available. Alternatives to participating in this study would be to receive standard care, which at the present time is still 5 years of tamoxifen in some countries, 5 years of an aromatase inhibitor in others or a sequence of 2 to 3 years of tamoxifen followed by 2 to 3 years of an aromatase inhibitor. Your doctor will explain the standard treatment in your country and hospital. If you participate in this study you will not have to make more visits to hospital, nor undergo more examinations or blood samples during the endocrine therapy period and after completion of the study than patients being treated for the same disease who are not taking part in the study. Results of the study will not be available until after the study is finished, however an individual patient will know on a regular basis if the medication is working against her cancer. Version / 152 May 21, 2014

292 Length of Study It is expected that you will take study medication for 7 years. If an examination shows that your cancer has come back, you will be told to stop taking the study medication. Treatment with letrozole or tamoxifen followed by letrozole will also be stopped if: - the side effects of this treatment become intolerable - new information indicates that this treatment is not in your best interest - you have taken this treatment for 7 years After you stop taking letrozole or letrozole followed by tamoxifen, your progress will be followed every 12 months. If new side effects or information about your disease or treatment are discovered during the study, you will be told. 5. Description of foreseeable risks and discomforts Possible Side Effects and Risks Generally, the observed adverse reactions with hormonal therapy are mainly mild to moderate in nature. Several of these side effects can be attributed to the normal effects of estrogen removal or blocking of the estrogen signal (are similar to symptoms of the menopause). Side effects you might experience with letrozole are: hot flushes (11% of patients) nausea (7% of patients) fatigue (5% of patients) headache/dizziness (7% of patients) edema (6% of patients) self reported osteoporosis, (7% of patients) Other side effects affecting between 1-10% of patients are: anorexia or appetite increase, vomiting, dyspepsia, diarrhea, hair loss, increased sweating, rash, myalgia (muscle pain), bone pain, athralgia (joint pain) and arthritis. Less frequent side effects affecting less than 1% of patients are: leucopenia, hypercholestrolaemia (high blood cholesterol), general oedema (swelling), depression, anxiety, somnolence, insomnia, memory impairment, dysaesthesia (unpleasant abnormal sensations), taste disturbance, cataract, eye irritation, blurred vision, palpitations, tachycardia (rapid beating of the heart), thrombophlebitis (inflammation of a vein forming a clot), hypertension, dyspnoea (shortness of breath), abdominal pain, stomatitis (swelling of soft tissues of the mouth), dry mouth, increased hepatic enzymes, pruritus (itching), dry skin, urticaria (hives), increased urinary frequency, vaginal bleeding, vaginal discharge, vaginal dryness, breast pain, pyrexia, mucosal dryness and thirst. These side effects are generally mild to moderate in nature, although unlikely, unforeseeable or unexpected risk(s)/side effects may occur. Letrozole will strongly decrease the body s estrogen levels. The effect of this lowering of estrogen may increase your chances of osteoporosis (thinning of the bones). These risks will be closely monitored. Side effects you might experience with tamoxifen are: hot flushes (up to 25% of patients) nausea and/or vomiting (up to 25% of patients) vaginal bleeding, vaginal discharge, menstrual irregularities and skin rash (less frequent) an increase in the risk of developing endometrial cancer Version / 152 May 21, 2014

293 Some of the beneficial side effects of tamoxifen are: strengthening of bones and lowering of blood cholesterol The side effects documented for letrozole and tamoxifen are symptoms that may occur. It is unlikely, however, that a patient would experience all, or even most of them. You should immediately report any discomfort or problem to your doctor. The effectiveness of this new study medication may be endangered by the use of hormone replacement therapy drugs (oral, patch, vaginal creams). Additionally the majority of hormone replacement therapies may increase the risk of breast cancer coming back. Therefore you must inform the study personnel of any hormone replacement medication that you are taking and it may be necessary to discontinue the use of these drugs. 6. Description of the ultimate goal of the clinical research project The main objective of the endocrine part of the trial is to determine if 7 years of letrozole is more effective and safer than 2 years of tamoxifen followed by 5 years of letrozole. The trial will measure which treatment prevents your cancer from coming back for longer (longer disease free survival). It will also measure the short and long term side effects for each treatment to determine which is less toxic. 7. Expected benefits Possible Benefits If you agree to participate in this study we cannot be sure that you will benefit personally. However you will receive treatment that is standard of care in many European countries or an extended treatment regimen with aromatase inhibitors which are extensively used in cases where tamoxifen is no longer effective. There might be less chance of your cancer coming back, you might live longer. The quality of your life might be better. These things cannot be predicted for any individual patient, however we hope that information from this study will help cancer patients in the future. Early results from trials testing letrozole have shown it to be well tolerated and effective in advanced breast cancer (where the cancer has spread). Also some early results show letrozole to be better than tamoxifen in early breast cancer when taken for 5 years. However the long term efficacy and safety results have not been reported yet. Both tamoxifen and letrozole have relatively mild side effects as described above. If new information about letrozole or the sequential treatment of 5 years of letrozole after an initial 2 years of tamoxifen becomes available it will be communicated to you by your treating doctor, as will any information that may be relevant to your willingness to continue participation in the trial. 8. Voluntary participation Your participation in this clinical trial is entirely voluntary and you will be given sufficient time to decide whether or not you wish to participate. You are free to decide at any time, without giving any reason, that you no longer wish to participate in the trial. Such decision will not affect your subsequent treatment or relationship with your treating doctor or the hospital staff in any way. Medical data collected during your participation to the clinical trial as well as follow up data which will still be prospectively collected will be kept for research and analysis unless you specify otherwise. If you consent to join this trial, you will be given a telephone number at the hospital that you can contact at any time if you feel unwell or have further questions. We encourage you to take time to consider participation in this trial and to discuss it with your family, friends and your family doctor. Version / 152 May 21, 2014

294 9. Data protection Your consent for participation in this part of the trial also includes your consent to allow the use of the data in your medical/clinical record to be used for research purposes. Your consent also includes allowing this data to be linked to data coming from other sources (such as cancer registries, medical/clinical records, ). All data (personal, clinical, economic and data coming from research on biological material) collected on your behalf will be treated in compliance with the national applicable laws. It is very important that the information collected is accurate and therefore, from time to time, this collected information may be checked against your medical records. Duly authorized persons (EORTC research staff and their representatives, national and/or foreign health authority representatives) may have access to your medical records. With the exception of access by the duly authorized persons to your personal data on your medical record, all information will be strictly confidential. This clinical research project is conducted under the legal framework of the EORTC with partial financial contribution of the European Commission, national charities and pharmaceutical companies. The EORTC, which is responsible for the conduct of this trial, has asked your treating doctor to disclose any existing conflict of interest he/she may have as a result of his/her activities related to this trial. The EORTC has set up procedures to ensure the integrity of this process 11. Insurance The EORTC who act as legal sponsor of the Study has obtained a clinical trial insurance in accordance with the applicable legislation. If you need to undergo another medical treatment, we advise you to inform the study doctor to ensure this will not have any effect on your participation to the trial. Everything has been done and will continue to be done to prevent additional health problems occurring as a result of your taking part in this trial. 12. Ethics Committee This research protocol has been submitted to the ethics committee whose mission is to verify that all conditions with respect to your safety and rights are respected. Approval to this research has been given by the Ethics Committee of your hospital on 13. Contact persons In case of any problem or question, your doctor will be pleased to answer any further questions and may be contacted as follows: Name of the doctor: Hospital: Telephone: With your agreement, your family doctor (if applicable) will also be informed about your taking part in this trial and what is involved, if you agree. Please take your time to consider this information and do not hesitate to ask further questions to your doctor if anything is unclear. You are entitled to keep a copy of this document after you and your doctor have signed it. Version / 152 May 21, 2014

295 Informed consent I have received a copy of the patient information sheet. I have been properly informed about the clinical trial and have been given sufficient time to consider my participation. All my rights have been clearly explained to me. I agree to participate in the clinical research study entitled Microarray In Nodenegative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy in the endocrine therapy question titled: Is 7 years of upfront aromatase inhibitor, letrozole, safer and more effective than a sequential strategy of 2 years of tamoxifen followed by 5 years of letrozole? and to be enrolled in EORTC study number BIG I accept that any data resulting from this clinical research study can be linked with other resources for cancer research purposes. My participation is completely voluntary and I have the possibility to withdraw my consent at any time without explanation. This will not affect my relationship with my treating doctor. The data collected on my behalf will be strictly confidential and treated according to the European and national applicable laws I have been informed that the data (personal, clinical and biological material) collected may be used in the future for cancer scientific research purposes while confidentiality will be ensured. All data (personal, clinical and research on biological material) collected on my behalf will be treated in compliance with the European and national applicable laws. My consent does not discharge the organizers of the research from their responsibilities and I keep all my rights guaranteed by the law. Patient's name: Patient's signature: Date: Investigator or person designated by the investigator to participate in the informed consent process: Name: Signature: Date: Title/Position: This document has been prepared taking the following documents into account: World Medical Association Declaration of Helsinki, adopted by the 18th World Medical Assembly, Helsinki, Finland June Revised 1975, 1983, 1989, 1996 and on October 6, 2000 in Edinburgh, Scotland ( ICH-GCP Guidelines; Note for Guidance on Good Clinical Practice (CPMP/ICH/135/95), Sept International Ethical Guidelines for Biomedical Research involving Human Subjects, Council for International Organizations of Medical Sciences (CIOMS), Geneva WHO: Operating Guidelines for Ethics Committee that Review Biomedical Research, Geneva, European Union Directive on the protection of individuals with regard to the processing of personal data (Dir/95/46/EC) Version / 152 May 21, 2014

296 Appendix I: Capecitabine schedule and interruptions to schedule Normal Treatment: Day Day 1 8 Day 15 Day 22 Day 29 Day 36 Day 43 Day 50 Interruptions During Treatment: Interruptions are regarded as lost treatment days and the planned treatment schedule should be maintained. Version / 152 May 21, 2014

297 ABCGS AJCC AI AML ARNO ASAT ASCO ATAC bid BIG BSA BSO CA 15-3 CAF CALGB CBC CEF CHF CMF CT CTCAE DCIS DEXA DFS DMFS DVT EBCTCG EGFR ER ET FAC FEC100 FISH FSH Appendix J: Abbreviations Austrian breast cancer study group American Joint Cancer Committee aromatase inhibitor acute myeloid leukemia German Adjuvant Breast Cancer Group alanine aminotransferase American Society of Clinical Oncology Arimidex and Tamoxifen Alone or in Combination bis in die Breast International Group Body surface area bilateral salpingo-oophorectomy cancer antigen 15-3 (protein product of the MUC1 gene) Cyclophosphamide Adriamycin 5-Fluorouracil the Cancer and Leukemia Group B complete blood count Cyclophosphamide Epirubicin 5-Fluorouracil congestive heart failure Cyclophosphamide Methotrexate 5-Fluorouracil Computed Tomography scan common terminology criteria for adverse advents ductal carcinoma in situ Dual Energy X-ray Absorptiometry disease free survival distant metastasis free survival deep vein thrombosis The Early Breast Cancer Trialists Collaborative Group epidermal growth factor receptor estrogen receptor endocrine therapy 5-Fluorouracil Adriamycin Cyclophosphamide 5-Fluorouracil (500 mg/m²) Epirubicin (100 mg/m²) and Cyclophosphamide (500 mg/m²) q3w 6 cycles Fluorescent in situ hybridization follicle stimulating hormone Version / 152 May 21, 2014

298 5-FU GnRH HE HER-2 HRT ICH-GCP IES ITA IRC LCIS LH LVEF MDS MRI MUGA NCC/G NCIC-CTG NSABP OS p.o. PET PgR PIS & IC pts QoL R-C R-E R-T SAE SERMS TMA TRANSBIG UNL WHO 5-FluoroUracil gonadotrophin releasing hormone hematoxylin and eosin Human Epidermal growth factor Receptor (also HER-2/neu) hormone replacement therapy International Conference on Harmonization-Good Clinical Practice Intergroup Exemestane Study Italian tamoxifen and anastrozole trial Independent Review Committee lobular carcinoma in situ luteinising hormone left ventricle ejection fraction myelodysplastic syndromes Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy magnetic resonance imaging MUltiple Gate Acquisition scan national coordinating center/group National Cancer Institute of Canada Clinical Trials Group National Surgical Adjuvant Breast and Bowel Project overall survival per os position emission tomography progesterone receptor patient information & informed consent patients quality of life randomization for chemotherapy regimen randomization for endocrine therapy regimen randomization for treatment decision serious adverse event selective estrogen receptor modulators tissue microarray Translational research arm of BIG upper normal limit World Health Organization Version / 152 May 21, 2014

299 STATISTICAL ANALYSIS PLAN EORTC Study BIG Study 3-04 A prospective randomized study comparing the 70-gene signature with the common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0-3 positive nodes [Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy: study] Version: FINAL 1.0 Authors: Dr. Leen Slaets, Biostatistician, EORTC Headquarters Dr. Jan Bogaerts, Methodology director, EORTC Headquarters Reviewers: Prof. Emiel Rutgers, Study Coordinator, Amsterdam, Netherlands Dr. Fatima Cardoso, Study Co-Coordinator, Lisbon, Portugal Prof. Martine Piccart, Study Co-Coordinator, Brussels, Belgium Dr. Jan Bogaerts, Methodology director, EORTC Headquarters Dr. Konstantinos Tryfonidis, Clinical Research Physician, Breast Group, EORTC Headquarters Registered Office: avenue E Mounier 83 Bte Brussels Belgium Phone: Fax: eortc@eortc.be

300 EORTC Headquarters March 15, 2016 TABLE OF CONTENTS 1. INTRODUCTION Setting Scope 4 2. DATA SETS, CONVENTIONS Data sets Analysis populations General conventions for statistical analyses Data recoding and display Conventions for statistical analysis and reporting Shift in 70-gene risk caused by a change in RNA extraction solution 7 3. TIMING OF FINAL ANALYSES AS PLANNED IN THE PROTOCOL General conventions for all databases Treatment decision randomization (R-T) Chemotherapy randomization (R-C) Endocrine therapy randomization (R-E) 8 4. ACCRUAL, ELIGIBILITY, BASELINE CHARACTERISTICS Accrual by center and treatment group Eligibility For the trial For the R-C randomization Patient and disease characteristics All patients dataset (AP) R-T randomized dataset (ITT1) R-C randomized dataset (ITT2) Central pathology TREATMENT Exposure to treatment All patients dataset (AP) Chemotherapy For the R-C Randomization safety population (SP2) COMPLIANCE For the R-T Randomization (AP) For the R-C Randomization (ITT2) For the R-C Randomization (SP2) 12 EORTC study 10041: analysis plan Version 1.0 2/21

301 EORTC Headquarters March 15, SAFETY Conventions for safety analyses AE and laboratory abnormalities tables Randomized chemotherapy safety population (SP2) Serious Adverse Events Toxic deaths Adverse events in safety population REASONS OFF TREATMENT Reason off chemotherapy (AP dataset) EFFICACY Endpoints Endpoint definitions Distant Metastasis Free Survival (DMFS) Disease Free Survival (DFS) Overall Survival General note to the above definitions Efficacy analyses for R-T Primary analysis for R-T Analyses on ITT1 dataset Analyses on AP dataset Efficacy analyses for R-C Pre-planned exploratory analyses per protocol or SAP Subgroup analyses for R-T Subgroup analyses for R-C Competing risk analyses Assessment of the prognostic value of the 70-gene signature compared to existing clinical methods Assessment of the clinical utility of the 70-gene signature Assessment of the predictive effect of the 70-gene signature will be investigated Multivariate analyses APPENDICES Recommendations from the EORTC IDMC Protocol texts Sample size Randomization and stratifications Timing of analyses Table of acronyms used in this analysis plan 19 EORTC study 10041: analysis plan Version 1.0 3/21

302 EORTC Headquarters March 15, Introduction 1.1. Setting This document contains a description of how the analyses for the Intergroup Phase III study EORTC / BIG 3-04 will be performed. All presentations and publications for the primary results will be based on the analyses described here. This document is to be used in conjunction with the protocol which contains a detailed description of the clinical aspects of this study, EORTC ST-005-SOP Statistical Analyses, related EORTC Work Instructions, the consecutive recommendations from the EORTC IDMC and the overview of Mindact topline results. For ease of reference, some key texts from the protocol are reproduced in Appendix 10 below. While every care is being taken to avoid contradictions, whenever there should be apparent contradictions, the following order of priority is followed (from high priority to low priority): - Protocol (apart from the changes mentioned below as SAP deviations) The current analysis plan - The IDMC recommendations - EORTC SOPs - EORTC WINs SAP Deviations with respect to the protocol: The multivariate analyses in chapter 9.5, will use a different model building approach than the one mentioned in the protocol. The approach as currently described in the SAP follows more recent guidelines on the development of prognostic models 1. The changes are summarized below: - Using statistical significance in univariate analysis as a pre-screening test to select variables for inclusion is no longer recommended. Therefore, this process had been dropped as the first step of model building. - It is recommended that prognostic risk factors which have clinical credibility and are already well established in the literature are retained in models. Therefore, some factors are retained in all models. - If automated variable selection methods are to be used, then backward elimination is preferred to forward selection. Therefore, we will not further evaluate the final model by performing a confirmatory forward selection technique. The biomarkers ER, PgR and HER2 are categorized according to recent guidelines, as previously analysed and published 2 1. S. Mallet,P. Royston,S. Dutton,R. Waters,D.G. Altman (2010). Reporting methods in studies developing prognostic models in cancer: a review. BMC Medecine 8 (20). 2. G. Viale1, L. Slaets, J. Bogaerts, L. van t Veer, E. Rutgers, M. J. Piccart-Gebhart, F. A. de Snoo, L. Stork-Sloots, L. Russo, P. Dell Orto, J. van den Akker, A. Glas, F. Cardoso on behalf of the TRANSBIG Consortium & the Investigators. (2014) High concordance of protein (by IHC), gene (by FISH; HER2 only), and microarray readout (by TargetPrint) of ER, PgR, and HER2: results from the EORTC 10041/BIG trial. Annals of Oncology 25: Scope This plan covers the final analyses for the primary endpoint as they relate to the objectives stated in the protocol, including the chemotherapy randomization R-C (see section 3). for the It does not deal with the analyses to be performed in side studies to EORTC study or the final analysis of the endocrine randomization. Per the protocol (section ), the primary analysis of the endocrine comparison will be EORTC study 10041: analysis plan Version 1.0 4/21

303 EORTC Headquarters March 15, 2016 performed when 379 DFS events have been observed among the patients randomized for the endocrine question. All additional translational research subprojects will need to be described in separate analysis plans prior to being performed. 2. Data sets, conventions 2.1. Data sets Analysis populations At the various time points of analysis, (some of) the following analysis datasets will be used. Because of the complexity of the study, there are many datasets, and so a code is added between brackets for ease of reference. Texts in italics are additions and clarifications not referenced in the protocol. All registered patients (RP): All patients who were entered into the preliminary registration for the study. These patients consented to sending their tumor sample to Agendia. However many of them were never enrolled into the study itself. All patients dataset (AP): All patients who were enrolled into the study after receipt of the microarray analysis. Unless mentioned otherwise, tables and analyses reported in this population will be done per corrected clinical and/or genomic risk (in case of changes in risk post-enrollment), as shown below. The discordant (corrected) risk groups will be further split into columns according to the randomization to chemo vs no chemo. Corrected* risk combination variable clgl clgh chgl chgh Randomized to no chemo Randomized to chemo Randomized to no chemo Randomized to chemo TOTAL N (%) N (%) N (%) N (%) N (%) N (%) N (%) * Corrected means that, in case an error in risk calculation was made at enrollment, patients are classified according to the actual risk of the patient. For most patients, the corrected risk is identical to the risk at recorded at enrollment.the terminology used to indicate the corrected risk, might still be changed All patients safety dataset (APS): All patients who were enrolled into the study after receipt of the microarray analysis and who followed the allocated therapeutic strategy (chemo versus no chemo). Per protocol primary test dataset (PT): The set of patients who have a low risk gene prognosis signature and high risk clinical-pathological criteria at enrollment, and who were randomized (R-T) to use the 70- gene signature risk and thus receive no chemotherapy, and did not deviate from this: no change in risk status post enrollment and no adjuvant chemotherapy received. This dataset is used for the primary test. Primary test dataset as enrolled (PTE): The set of patients who have a low risk gene prognosis signature and high risk clinical-pathological criteria at the time of enrollment, and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy. This dataset is used for a first sensitivity test of the primary test. Patients for whom it is unknown whether they received chemo (e.g. the patient withdrew consent), will be excluded from this population. EORTC study 10041: analysis plan Version 1.0 5/21

304 EORTC Headquarters March 15, 2016 Per protocol primary test dataset sensitivity (PTS): The subset of patients in the PPT dataset excluding those patients whose samples were received by Agendia between May and January (RNA extraction solution problem, see section 2.3). This dataset is used for a second sensitivity analysis for the primary test. See also the IDMC recommendations in section Intention-to-treat population for R-T (ITT1): All patients randomized for treatment decision (R-T) will be analyzed in the arm they were allocated by randomization. This is the dataset of all patients with discordant risk assessments at the time of enrollment, because these are the patients who were randomized by the platform at that time. Per protocol population for R-T (PP1): All patients who are eligible and randomized for treatment decision (R-T) and have started their allocated treatment (at least one dose of chemotherapy if so randomized, respectively no chemotherapy). The following patients will be considered excluded from the for PP1 analysis: - Patients who were post enrollment found to have an error in either one or both of the risk assessements. While they may have received a treatment that would have been the correct outcome (in the logic of the trial), they should not have been randomized. - Patients found to be ineligible for the trial (protocol chapter 3.1) as per the clinical review (done by CRP or SC). This does not include ineligibility for R-C or R-E. Note that patients who overstepped the timelines from diagnosis to surgery or surgery to enrollment or diagnosis to enrollment (with or without waiver), will not be considered ineligible according to the above definition. Per protocol population for R-T sensitivity (PPS): The subset of patients in the PP1 dataset excluding those patients whose samples were received by Agendia between may and January (RNA extraction solution problem, see section 2.3). See also the IDMC recommendations in section Intention-to-treat population for R-C (ITT2): All patients randomized for R-C will be analyzed in the arm they were allocated by randomization. Safety population for R-C (SP2): All patients who were randomized for R-C and who have started their allocated chemotherapy (at least one dose of the study drug(s)). Intention-to-treat population for R-E (ITT3): All patients randomized for R-E will be analyzed in the arm they were allocated by randomization. Safety population for R-E (SP3): All patients who were randomized for R-E and who have started their allocated endocrine treatment (at least one dose of the study drug(s)). A patient will be considered to be eligible if she did not have any major deviations from the patient entry criteria listed in chapter 3. Eligibility will be assessed by the CRP based on the review of patient files. A review by the CRP was requested by the data manager for those patients for whom there was a potential violation of eligibility, based on checks on the reported variables in the database. The ITT3 and SP3 population will not be part of the current analysis plan General conventions for statistical analyses Data recoding and display Frequency tables will be tabulated (by treatment group or otherwise) for all categorical variables by the levels of the variables as they appear on the CRF (with %). Some categorical variables may be pooled as deemed appropriate (e.g. missing data may be pooled with unknown ). Some labels may be renamed to enhance clarity. Categories with a text field specification will be tabulated as categories and then supplemented by a listing with the following information for the patients fulfilling the condition for the specification (patient id, institution, treatment group, value of the item and text field contents). An EORTC data convention exists whereby multiple reasons are coded as other with additional coding conventions given in the text field, enabling computerized identification of the multiple reasons. If in the EORTC study 10041: analysis plan Version 1.0 6/21

305 EORTC Headquarters March 15, 2016 plan below, a priority list is given, then the reasons as decoded from this method will be prioritized, and the patient will be tabulated in the highest priority class. Example: in the off chemotherapy treatment reason table, the highest priority reason is toxicity. Therefore, any other with documented multiple reasons, including toxicity, will be tabulated under toxicity. Dates relating to events prior to entry will be presented as the delay in days (or weeks, months, or years) between the past event and the date of entry (date of entry date of past event + 1) and presented using the median and range. For the appropriate date of entry per analysis, refer to Section 9.1. For example, on the randomization checklist, the date of last administration of prior treatment (or the date of first diagnosis of the cancer) will be presented as the time elapsed (in days, weeks, months or years, as appropriate) since the day of the last administration and the date of entry on study. Other delays (e.g. re-treatment delays) are presented as continuous variables using the median and range. Safety and toxicity counts will be tabulated as worst grade per patient, within CTC term, or laboratory test. Continuous variables for which a coding system exists (such as for laboratory data) will be recoded into categories (for adverse events, the grading scale specified in the protocol will be used). Whenever no specific scale exists, lab data will be categorized based on the normal range: for example, below the lower normal limit (when appropriate), within the normal range, above the upper normal limit (UNL) and the degree to which it is above the UNL (for example > 2.5 x UNL, > 5 x UNL, > 10 x UNL). For laboratory data, the nadir is generally displayed. The nadir in a given cycle is the lowest laboratory value in that cycle; the overall nadir for a patient is the lowest laboratory value among all cycles. Other continuous variables (for example age, dose ) are presented using the median and range (minimum, maximum). Where appropriate, continuous data may also be presented in categories (for example, age may also be grouped in decades) Conventions for statistical analysis and reporting All statistical tests and confidence intervals will be two-sided at 5% significance level, with the exception of the primary test for R-T, which is one-sided at 97.5% confidence. All statistical tests will be tests for superiority (difference). All secondary endpoints will use a 95% confidence level at the final analysis. All comparisons for time to event endpoints described below will be adjusted Logrank tests, as performed by the Wald test in a Cox model with the effect under consideration, as well as the stratification factors used for the applicable randomization (with the exception of institution). As there is no a priori reason to believe that the stratification factors used will exhibit non-proportional hazards, preference is given to adjusting for them by using them as covariates (not by stratified Cox model). Corresponding hazard ratios and confidence intervals (Wald method) will be calculated by the same Cox regression. Time to event endpoints comparisons will be accompanied by Kaplan-Meier curves Shift in 70-gene risk caused by a change in RNA extraction solution A change in the RNA extraction solution that was not communicated by the manufacturer caused a temporary shift in the 70-gene risk (from approximately end of May 2009 to end of January 2010), until the issue was discovered and rectified. The result of the shift was that a number of patients were identified as being 70-gene high risk, where by an unbiased (correct) test they would have been 70-gene low risk. The retro-actively recalibrated results were communicated to investigators and entered into the system, but for many affected patients this was too late to adjust their (initial) treatment. For the analysis populations (PTE, PT, PTS, PPS) mentioned in this section, refer to section Impact on the primary test: The problem caused 70 patients who should have been ch/gl to be assessed as ch/gh. They thus were not randomized, but allocated to chemotherapy. Since patients with a higher correlation (closest to the cut-off low vs high) were wrongly classified as 70-gene high risk, it can be argued that the patients who ended up in the ch/gl group during the occurrence of the RNA extraction solution problem were at a better risk from the 70-gene risk point of view (as compared to the rest of the enrollment EORTC study 10041: analysis plan Version 1.0 7/21

306 EORTC Headquarters March 15, 2016 period), thereby biasing the final test. However, the actual value of the correlation showed poor prognostic power in the development and validation sets. It is only the division in two groups at a cutpoint of 0.4 correlation that seems to have good prognostic characteristics. The latter argument is in favor of keeping patients randomized during this period in the analysis population for the primary test. Therefore, the impact of the RNA extraction issue will be assessed in a sensitivity analysis and these patients are not excluded from the primary test population. The population PTS was created for this purpose. See also the IDMC recommendations in section Impact on the other risk groups: Since all risk groups as enrolled are somewhat biased due to incorrect risk assessment in the full period from end of May 2009 to January 2010, an additional patient population is defined in the analysis plan for the R-T analyses. This population (PPS) excludes all patients enrolled during this period and will be used in a sensitivity analysis for the R-T analyses. See also the IDMC recommendations in section Timing of final analyses as planned in the protocol 3.1. General conventions for all databases All analyses described in this analysis plan will be performed on databases that were locked for the purpose of performing stated analysis, and as extracted at the time of the database lock. The data will be restricted by the analyzing statistician to include only data points pertaining to patient experiences up to the clinical cutoff date defined for the particular lock Treatment decision randomization (R-T) The protocol (see appendix in this document for the applicable text) calls to do the primary test at the time when two conditions are met: 1. the standard error of DMFS at 5 years (obtained by estimating the standard error of log(- log(s(t)), and back-transformation) is 0.01 or less 2. at least one third of the patients in the primary dataset (PT) have follow-up info for five years Condition 1 aims to ensure the power of the procedure. Condition 2 aims to ensure a valid estimate at 5 years. The analyses on the PTS dataset will be performed at time of primary analysis and when the two conditions in section have been met in the PTS dataset Chemotherapy randomization (R-C) Per the protocol (section ), the primary comparison for R-C for DFS is to be performed at the time of the primary test of the study (see section 3.2). Due to insufficient numbers of patients randomized into this part of the study, there will be less 80% power to detect the original alternative hypothesis of hazard ratio of 0.76 (for example a control 5-year DFS of 86% versus a 89.2% experimental 5-year DFS). The a posteriori power will be calculated Endocrine therapy randomization (R-E) Per the protocol (section ), the primary analysis of the endocrine comparison will be performed when 379 DFS events have been observed among the patients randomized for the endocrine question, yielding 80% power to detect a hazard ratio of EORTC study 10041: analysis plan Version 1.0 8/21

307 EORTC Headquarters March 15, 2016 Since the number of patients randomized for endocrine therapy was smaller than anticipated, it might become impossible to observe the number of required events within a reasonable time. Under the latter scenario, the IDMC will be consulted at around a median follow-up of 8 years, to discuss the release of the R-E data. 4. Accrual, Eligibility, Baseline characteristics 4.1. Accrual by center and treatment group Accrual tables will use the RP dataset, identifying the AP dataset within: 4.2. Eligibility The number of patients enrolled out of those registered in the different centers will be presented in a table. A second table will provide the total number of patients enrolled out of those registered each quarter. Corresponding accrual curves of the RP and AP dataset will be drawn. The eligibility status of the patients as assessed by the Clinical Research Physician and the StudyCoordinator (Review Form) will be summarized as follows. Additionally, lists of the ineligible patients for each of the below subsections will be presented For the trial AP dataset. Eligibility and reason for ineligibility at the trial level will be tabulated. See section 3.1 in the protocol. AP dataset. Eligibility and reason for ineligibility at the R-T randomization level will be tabulated. Of note, this essentially means tabulation of post-enrollment changes in one (or both) of the two risk assessments. For patients who were not randomized, this will usually mean that without the error they would have been randomized, and vice versa. Tabulation will be by risk combination at enrollment. Further tabulation will show the type of change in risk, and the cause of the change For the R-C randomization ITT2 dataset. A few criteria are checked at the time of chemotherapy randomization. Ineligibility for these criteria will be summarized by R-C treatment arm. See section 3.3 in the protocol Patient and disease characteristics The list of patient/disease characteristics to be tabulated (either by incidence table, or by summary statistics, as applicable) will at least consist of the following Age (<35, 35-49, 50-70) Menopausal status Baseline WHO performance status Type of surgery (breast conserving surgery, radical mastectomy) Type of node evaluation (SNB, full axillary clearance) Tumor size (0-1 cm, >1-2 cm, >2-5 cm, >5 cm) Differentiation (grade I well differentiated, grade II moderately differentiated, grade III poorly differentiated, unknown) EORTC study 10041: analysis plan Version 1.0 9/21

308 EORTC Headquarters March 15, 2016 Nodal involvement TNM stage Time from surgery to enrollment HER-2 status (positive, negative, unknown) ER-PgR status (individual, and combined) Clinical-pathological risk assessment (low= cl, high= ch ) 70-gene signature risk assessment (low= gl, high= gh ) For the date of surgery, in case of a second surgery the date of definite surgery will be used when calculating the timelines related to date of surgery. Refer to IDMC recommendations in section All patients dataset (AP) Baseline characteristics will be summarized in the AP dataset. McNemar s test will be applied to the 2x2 table of clinical-pathological (high/low) by 70-gene signature (high/low) risk to test whether there is a difference in assignment percentages to chemotherapy. The difference in high risk percentages will be calculated and a 95% two-sided confidence interval will be calculated (for the paired data) R-T randomized dataset (ITT1) Baseline characteristics will be summarized in the ITT1 dataset, split by type of discordance (at enrollment) and by R-T randomization outcome (chemo/no chemo), thus in 4 columns R-C randomized dataset (ITT2) Baseline characteristics will be summarized in the ITT2 dataset by R-C randomization outcome (chemo type outcome). Also, the timing of this randomization, relative to initial enrollment into the study will be summarized. Proposed timeline categories to be summarized: 1. Enrollment to R-C: 1-4 weeks and > 4 weeks. 2. Surgery to start of R-C: 1-8 weeks, 8 weeks 120 days, > 120 days 4.4. Central pathology In the AP dataset, the results of the central pathology assessment will be reported. This includes: Er, PgR and HER2 status, KI67, histological type and grade, vascular invasion. 5. Treatment 5.1. Exposure to treatment All patients dataset (AP) In the AP dataset, a table will describe administration of chemotherapy (yes/no), the types of chemotherapy, nb of cycles, reason for discontinuation, radiotherapy administration (yes/no) and endocrine therapy (yes/no, type of endocrine therapy and duration). A second table will be made, also on the AP dataset, but with a focus on the R-C randomization within ITT2 (different columns than the first tables). Four columns will be shown as given below: variable Enrolled in R-C randomization chemo received No chemo TOTAL EORTC study 10041: analysis plan Version /21

309 EORTC Headquarters March 15, 2016 (ITT2) (outside of R-C) received Randomized to Arm 1 Randomized to Arm 2 N (%) N (%) N (%) N (%) N (%) In a sub-group analysis we will clarify which type of chemotherapy was received (random vs. non random) per risk assessment, lymph node status, HER2 status to describe any difference between these groups (which can exclude/suggest patient selection) Chemotherapy For randomized chemotherapy, data collection of chemotherapy administration was different between the two arms. In the experimental arm, more detailed records of actual doses and more specific reasons for changes were collected. For the anthracycline arm, only the type of chemotherapy, the number of cycles administered the last administration date, the dose modifications, treatment delays and discontiunations and the reasons thereof are collected. Other variables mentioned below will only be reported in the docetaxel arm For the R-C Randomization safety population (SP2) Chemotherapy administration will be summarized by chemotherapy arm: The number of patients receiving each of the regimens constituting the control arm, split by nodal status. Number of cycles received. Dose intensity and relative dose intensity (<85%, %, 100%-115%,>115%) of docetaxelcapecitabine combination and for each drug individually in the experimental arm. Calculation rules will be as per EORTC work procedures. Body surface areas will be recalculated on the basis of height and last recorded weight at the time of administration. However, as there was no clear guidance on this, if the calculated BSA exceeded a value of 2.2 m 2, the value will be cut at 2.2 m 2 as almost all sites did (protocol page 55). The duration of treatment (in days) for a patient will be calculated as the difference between the start of the last administered cycle + 3 weeks and the first date of treatment. The distribution of the duration of treatment along with the reason for stopping treatment will be tabulated by treatment group. Dose modifications (as calculated being 10% below the per protocol dose) and discontinuations (as calculated), and reasons for modifications. Maximum cycle delay by patient, and by cycle (0-3 days, 4-7 days, 8-14 days, > 14 days), and reasons for delay. For the anthacycline arm, reported dose modifications/delays will be displayed, since there is no detailed data collected on the administration. Note: Some of the tables may need to be split by regimen and/or by nodal status for the control arm. This will be decided on a case by case basis, in order to maximize usefulness for interpretation of the data. 6. Compliance The compliance to protocol treatment, as reviewed by the SC and/or the CRP, will be tabulated and listed, by treatment arm (for chemo randomized therapy). Other elements of compliance are being summarized as part of several tables planned in above sections, but can be summarized in the compliance section of the report as well. EORTC study 10041: analysis plan Version /21

310 EORTC Headquarters March 15, 2016 Compliance to the follow-up scheme (appropriate disease assessment over time) will be summarized by the various arms, in order to assess potential biases in the estimation of the endpoints For the R-T Randomization (AP) Table documenting general protocol compliance (section A4 on the medical review form). When documenting compliance to R-T (chemo vs no chemo), patients with changes in risk assessments will be documented separately since there is no unambigious definition of per protocol treatment for these patients For the R-C Randomization (ITT2) Table documenting whether the R-C randomization was followed by randomized treatment arm (section B2 on the medical review form) For the R-C Randomization (SP2) Table documenting the non-compliance with the regimen administration by treatment arm (section B3 on the medical review form). 7. Safety 7.1. Conventions for safety analyses No formal comparative analyses with p values will be carried out for safety, as the data collection was different between chemotherapy arms (more details were asked for the experimental arm). All grading of adverse events and laboratory values was and will be done according to Common Terminology Criteria for Adverse Events v.3.0 (CTCAE). Since no normal ranges were collected for the labvalues, only those labvalues with fixed ranges will be reported AE and laboratory abnormalities tables Randomized chemotherapy safety population (SP2) Adverse events and laboratory abnormalities during chemotherapy with variable cyclelb=1-8) will be tabulated (worst CTC grade per patient) by treatment arm. These are AEs recorded on a form8 (adverse events) with variable timeae=1-8 or lab values on a form5 (laboratory values). Separate tables and listings will be made for late toxicity as collected on the follow-up form (cardiac toxicity, secondary leukemia) Serious Adverse Events Serious Adverse Events will be listed as reported for all enrolled patients, stating the treatment they were undergoing at the time of the SAE Toxic deaths Narratives of all toxic deaths will be written Adverse events in safety population Adverse events will be tabulated (worst CTC grade per patient) in the APS population. EORTC study 10041: analysis plan Version /21

311 EORTC Headquarters March 15, Reasons off treatment Reasons off treatment will be prioritized as follows (in order of priority): Toxicity Patient refusal Major violation Intercurrent death Progressive disease Other (i.e. non-recodable other reasons) Loss to follow-up Normal completion 8.1. Reason off chemotherapy (AP dataset) The reason for going off chemotherapy treatment will be tabulated in the dataset of all patients assigned to chemotherapy, using as columns (R-C randomized to control arm, R-C randomized to experimental arm, non-randomized chemo). In this table, patients who never started chemotherapy will be identified as a special subcategory (with the reasons). Additional listings of specifications will be made. Enrolled in R-C randomization variable (ITT2) Randomized to Randomized to chemo received (outside of R-C) No chemo received TOTAL Arm 1 Arm 2 N (%) N (%) N (%) N (%) N (%) / EORTC study 10041: analysis plan Version /21

312 EORTC Headquarters March 15, Efficacy 9.1. Endpoints In this complex protocol, time to event endpoints (DMFS, OS, and DFS) will start at the following dates: For all overall analyses, and analyses related to R-T, time to event endpoints will start at the date the sample of the patient was received at Agendia. These are the analyses on the data sets AP, PT, PTS, PPT, PPS, ITT1, PPT1. For all analyses related to R-C, time to event endpoints will start at the date the date the sample of the patient was received at Agendia. These are the analyses on the data set ITT2. The last examination date is defined as the date on which at least one of the following is assessed on the follow-up form: box7 (presence of local recurrence since last follow-up), box 8 (presence of regional recurrence since last follow-up), box9 (presence of distant metastases since last follow-up) or the last baseline/treatment date pertaining to the patient Endpoint definitions Distant Metastasis Free Survival (DMFS) For the endpoint of DMFS only distant metastatic recurrences as defined in Section of the protocol and deaths (for any cause) will be taken as events. Contralateral breast cancer and secondary cancers will not be taken into account. If the patient is alive without distant metastatic recurrence, the censoring date will be the last examination date. DMFS is then defined as the time from the appropriate start date (as per section 9.1) to the date of death/distant metastasis/date of censoring. Patients who were found to be ineligible due to M1 status at baseline by medical review will be censored at time 0. Patients who died more than 2 years after their last examination date will be censored at the last examination date. A sensitivity analysis will be perfomed, which will include deaths occuring over two years after the last examination date as events Disease Free Survival (DFS) For the endpoint of DFS loco-regional and distant recurrences, ipsilateral/contralateral invasive or ductal in situ DCIS) breast cancer, invasive second primary cancer (non breast) and death all qualify as events. Lobular carcinoma in situ (LCIS) will not be considered an event for DFS. If the patient is alive without any observation of cancer, the censoring date will be the last examination date. DFS is then defined as the time from the appropriate start date (as per section 9.1) to the date of death/any cancer. Patients who were found to be ineligible due to M1 status at baseline by medical review will be censored at time 0. Patients who died more than 2 years after their last examination date will be censored at the last examination date. A sensitivity analysis will be perfomed, which will include deaths occuring over two years after the last examination date as events Overall Survival For overall survival, the patient s death (due to any cause) is considered an event. In case the patient is alive, the date the patient was last known to be alive will be used as censoring date. The overall duration of survival is then defined as the time from the appropriate start date (as per section 9.1) to the date of death/date of censoring General note to the above definitions Interruptions or delays in disease assessment will not lead to censoring, but will be summarized as per Section 6. EORTC study 10041: analysis plan Version /21

313 EORTC Headquarters March 15, Efficacy analyses for R-T Primary analysis for R-T DMFS at 5 years will be tested on the PT dataset (see also section ). As a sensitivity analysis for this primary analysis of R-T, the same test will be done on the PTE and PTS datasets Analyses on ITT1 dataset In the subgroup of clinical-pathological high risk and 70-gene signature low risk, DMFS, OS and DFS will be compared for randomization to chemo versus no chemo. As these comparisons are low-powered, primary attention should go to the confidence intervals. In the subgroup of clinical-pathological low risk and 70-gene signature high risk (at enrollment), DMFS, OS and DFS will be compared for randomization to chemo versus no chemo. Again, as these comparisons are low-powered, primary attention should go to the confidence intervals. The interaction between the effect of chemotherapy and the subgroup (clinical-pathological high risk and 70- gene signature low risk versus clinical-pathological low risk and 70-gene signature high risk, at enrollment) will also be tested. Just as for the subset tests, the interaction test will be low-powered, so it will be informative only if there is a large difference in the effect of chemotherapy between the subsets, a situation that is unlikely to occur. The above analyses will be repeated, as a sensitivity analysis, on the PP1 and PPS populations Analyses on AP dataset Kaplan-Meier curves for OS, DFS and DMFS will be calculated for the following six subgroups (for corrected risk): high risk for both risk assessments, low risk for both risk assessments, high risk for clinical-pathological criteria and low risk for 70-gene signature and randomized (R-T) to use clinical pathological assessment of risk high risk for clinical-pathological criteria and low risk for 70-gene signature and randomized (R-T) to use 70-gene signature assessment of risk low risk for clinical-pathological criteria and high risk for 70-gene signature and randomized (R-T) to use clinical pathological assessment of risk low risk for clinical-pathological criteria and high risk for 70-gene signature and randomized (R-T) to use 70-gene signature assessment of risk These six curves will be put on the same plot, and their qualitative interpretation will be of great importance for the evaluation of the two assessment methods. However no formal statistical testing will be performed on this plot other than those described for the ITT1 dataset Efficacy analyses for R-C The primary comparison of R-C will be for DFS, using the ITT2 dataset. Secondary comparisons are for DMFS and OS. For these three time-to-event endpoints, the analyses described in section will be performed. EORTC study 10041: analysis plan Version /21

314 EORTC Headquarters March 15, Pre-planned exploratory analyses per protocol or SAP Subgroup analyses for R-T The primary analysis for R-T (see section 9.2.1) and the analyses on the AP dataset for DMFS (see section 9.2.3) will be perfomed in a subgroup analysis by nodal status (N+ versus N-) and age (< 40 versus 40) and for the subgroup of ERpos-HER2neg patients. The analyses on the AP dataset for DMFS (see section 9.2.3) will be perfomed in a subgroup analysis by molecular subtype Subgroup analyses for R-C If a large effect of the type of chemotherapy (R-C) is observed, further subgroup analyses of the analyses on the ITT1 dataset may be performed by type of chemotherapy, to generate confidence intervals of effects per chemotherapy type Competing risk analyses Sensitivity analyses will be performed for overall survival analyses on the AP, PT, ITT1, ITT2 datasets, by taking only cancer related deaths into account. These analyses will use competing risk methods to describe cumulative incidence of cancer related deaths. In this analysis deaths due to treatment toxicity or other treatment related deaths will be interpreted as cancer related. Similarly, sensitivity analyses will be performed for DMFS analyses on the AP, PT, ITT1, ITT2 datasets, using time to distant metastasis, by taking only metastasis as an event, and not deaths. These analyses will also use competing risk methods to describe cumulative incidence of distant metastasis Assessment of the prognostic value of the 70-gene signature compared to existing clinical methods In this analysis the prognostic value of the signature will be assessed and compared to existing clinical methods and to estimate what it adds in terms of predictive accuracy. The model-based ROC methodology (Heagerty and Zheng) will be used. A separate statistical analysis plan will be written for these analyses. Heagerty PJ, Zheng Y: Survival model predictive accuracy and ROC curves. Biometrics Mar;61(1): Assessment of the clinical utility of the 70-gene signature We will develop a model that aims to predict the outcome of the 70-gene signature based on clinical factors (e.g. are there subgroups of patients for whom we can reliably predict the 70-gene outcome). Additionally we will combine clinical and genomic data to optimize the outcome prediction. A separate statistical analysis plan will be written for these analyses Assessment of the predictive effect of the 70-gene signature will be investigated We will make an estimation of the predictive effect of the 70-gene signature. Since patients with clgl and chgh risks were not randomized, we cannot make a direct assessment of the predictive effect. We will use additional estimations (e.g. adjuvant! Online) to make an estimation of the outcome in these strata. A separate statistical analysis plan will be written for these analyses Multivariate analyses Prognosis is a key element of the trial. However, evaluation of prognostic factors is confounded by chemotherapy administration (yes/no), because key prognostic factors were used to decide on chemotherapy. Therefore, chemotherapy will be included in all models below, in an attempt to adjust for the effect. If the chemotherapy comparison on ITT2 is significant, the variable will have three levels (no, chemo arm A, chemo arm B) otherwise only two (no chemo, chemo). Endocrine therapy will not start at the time of entry into the trial, and will not be taken into account as a covariate. The effect of factors defining EORTC study 10041: analysis plan Version /21

315 EORTC Headquarters March 15, 2016 endocrine therapy will thus include endocrine therapy use. On the AP dataset, the three time-to-event endpoints (DMFS, OS, DFS) will be subjected to a full multivariate analysis, using the following conventions: All variables listed below will be put into a multivariate Cox proportional hazards model, together with chemotherapy and chemotherapy-variable interactions, and the final model will be built using backward selection (until all p<0.05), while keeping chemotherapy and the factors to be retained (as listed below) in the model at all times. Variables will be included or excluded with their full categorization, so no regrouping or selection of categories is allowed. The focus of this analysis will be on determining the extent to which 70-gene signature risk assessment may or may not replace other risk factors. After determining a final model, it will be evaluated for calibration and discrimination ability, using bootstrapping. The stability of the selected covariates in the final model will also be evaluated by the bootstrap (Ref. 102), as an internal validation. All variables listed below will be put into a multivariate Cox proportional hazards model, together with chemotherapy. The coefficients of the variables in a univariate model will be added to the output table. Variables will be included or excluded with their full categorization, so no regrouping or selection of categories is allowed. The focus of this analysis will be on determining the extent to which 70-gene signature risk assessment may or may not replace other risk factors. The variables to be considered, and their categories, are: Age (<35, 35-49, 50-70) Baseline WHO performance status (WHO 0, WHO >0) Menopausal status Type of surgery (breast conserving surgery, radical mastectomy) Type of node evaluation (SNB, full axillary clearance) The below variables will be retained in all models: Tumor size (0-1 cm, >1-2 cm, >2-5 cm, >5 cm) Differentiation (grade I well differentiated, grade II moderately differentiated, grade III poorly differentiated) Nodal involvement (yes, no) HER-2 status (positive, negative) ER-PgR status (Positive (ER and/or PgR), Negative (both)) Clinical-pathological risk assessment (low, high) 70-gene signature risk assessment (low, high) Additional prognostic and predictive analyses are very likely, either to develop additional or improve existing prognostic gene signatures on the basis of microarray data for 6,000 patients, or to develop or validate predictive signatures for endocrine therapy or chemotherapy response. The strategy to perform these will be described in ad-hoc research protocols. These include, but are not restricted to, the analyses already mentioned in sections 9.4.4, and Note For the multivariate analysis, the central pathology results for ER, PgR, HER2 and grade will be used when available (local pathology will be used otherwise). EORTC study 10041: analysis plan Version /21

316 EORTC Headquarters March 15, 2016 Central pathology review will repeat immunohistochemistry for estrogen and progesterone receptors to give standardized levels of expression which will be expressed as a percentage of immunoreactive cells. Categories used will be: positive: ER and/or PgR 1% negative: both ER and PgR = 0% Central HER2 assessment: HER-2 expression was scored as 0, 1+, 2+, or 3+, according to the FDA scoring system. Tumors scored as 2+ were re-tested with FISH. Cases were considered HER-2-positive if scored 3+ by IHC and/or amplified by FISH (ratio 2). 10. Appendices Recommendations from the EORTC IDMC Recommendations of 15/12/ Concerning the timelines between surgery and enrolment: the IDMC recommends that the protocol be amended, taking the date of the second surgery (when present) as the date of the definitive surgery, but maintain the current limit of 56 days using the date of the second surgery (when present) as time zero. Recommendations of 31/03/ Regarding RNA extraction issue: A sensitivity analysis excluding the ±1300 patients enrolled during the period in question can be carried out after a further period of follow up Protocol texts Sample size In the set of patients who have a low risk gene prognosis signature using the 70-gene signature and high risk clinical-pathological criteria, and who were randomized (R-T) to use the 70-gene signature prognosis and thus received no chemotherapy (i.e. they truly did not receive chemotherapy), a null hypothesis of a 5-year distant metastasis free survival (DMFS) of 92% will be tested. With 6,600 patients accrued overall, this set has an expected size of 672 patients. The primary test will be a one-sided test at 97.5% confidence level. This primary test will be performed when two conditions are met: the standard error of DMFS at 5 years (obtained by estimating the standard error of log(-log(s(t)), and back-transformation) is 0.01 or less and at least one third of the patients in the above dataset have been followed for five years. To perform the test, a 95% two-sided confidence interval will be constructed for the DMFS at 5 years, using the log-log transform formula. The test will be significant if the confidence interval does not include the value of Using these conditions, this test has 80% power to reject the null hypothesis if the true 5-year DMFS is 95%. The primary test for R-C is to compare DFS between the two chemotherapy arms randomized. Due to insufficient numbers of patients randomized into this part of the study, there will be less than 80% power to detect the original alternative hypothesis of hazard ratio of 0.76 (for example a control 5-year DFS of 86% versus a 89.2% experimental 5-year DFS). The comparisons for R-C will be performed at the time of the primary test (as described above) at 5% alpha and an a posteriori power calculation will be performed. The primary test for R-E is to compare DFS between the two sequences of endocrine therapy randomized. Due to insufficient numbers of patients being randomized into this question, there will be less than 80% power to detect the original alternative hypothesis of hazard ratio of 0.75 (for example a DFS rate at 5 years of 86% on the control arm versus an experimental 5-year DFS of 89.3%). The primary analysis of the endocrine comparison will be performed when 379 events have been observed among the patients randomized for the endocrine question, yielding 80% power to detect a hazard ratio of This is projected to occur at a median follow up of 7.9 years. EORTC study 10041: analysis plan Version /21

317 EORTC Headquarters March 15, Randomization and stratifications For eligible patients with adequate material for microarray analysis, both the 70-gene signature and clinicalpathological risk will be assessed, after which they can be enrolled. Patients will be centrally enrolled and randomized as applicable for each of the three possible randomizations (R-T, R-C, R-E, for practical details, see chapter on enrollment / randomization procedure). Patients are evaluated as low clinical-pathological risk, if their 10-year disease specific survival (without chemotherapy or endocrine therapy) is estimated by Adjuvant! Online as greater than 88% for ER-positive patients, and greater than 92% for ER-negative patients. Patients who are at high risk according to both methods will receive chemotherapy, and patients who are at low risk according to both methods will not receive chemotherapy. As new versions of Adjuvant! Online (Standard version) become available, these will be implemented. The version at the time of the writing of this protocol is Standard Version 8.0. R-T: Patients for whom both methods are discordant will be randomized between chemotherapy-decisionmaking according to clinical criteria and chemotherapy-decision-making according to 70-gene signature prognosis. For R-T, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/low risk clinical vs. low risk genomic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), nodal involvement (yes, no), age (<50 vs. >=50), HER-2 (Positive vs. Negative vs. Unknown), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). In case PgR is unknown, the patient will be stratified to the HR negative group if ER is negative, and to the HR positive group if ER is positive. Patients who are candidates for chemotherapy (see also treatment section) will be proposed for R-C: randomize to anthracycline based chemotherapy or taxane/capecitabine chemotherapy. For R-C, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/low risk clinical vs. low risk genomic/high risk clinical vs. high risk genomic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), age (<50 vs. >=50), HER-2 (Positive vs. Negative vs. Unknown (at the time of R-C)), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). In case PgR is unknown, the patient will be stratified to the HR negative group if ER is negative, and to the HR positive group if ER is positive. All patients who are candidates for endocrine therapy will be offered randomization (R-E) to 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. For R-E, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/high risk clinical vs. high risk genomic/low risk clinical vs. low risk genomic/high risk clinical vs. low risk genomic/low risk clinical), chemotherapy (no chemotherapy, chemotherapy without R-C, R-C arm A, R-C arm B), type of endocrine sensitivity (both ER and PgR positive vs. either ER or PgR positive), age (<50 vs. >=50), HER-2/neu (Positive vs. Negative vs. Unknown (at the time of R)), method of axillary evaluation (sentinel only or dissection), type of surgery (mastectomy or quadrantectomy/tumorectomy). The three randomizations will be open-label Timing of analyses All analyses on the AP, PT, PTE, ITT1, PP1 datasets will be performed when the two conditions in section have been met. The final analysis on the ITT2, SP2 datasets will be performed at the time of the primary analysis on the PT dataset. The final analysis on the ITT3, SP3 datasets will be performed when 379 events have been observed on the ITT3 dataset. This is projected to occur at a median follow up of 7.9 years Table of acronyms used in this analysis plan Topic Acronym What it means Risk assessment cl Low clinical/pathological risk EORTC study 10041: analysis plan Version /21

318 EORTC Headquarters March 15, 2016 Randomizations Data sets used for analysis Efficacy endpoints ch gl gh R-T R-C R-E RP AP APS PTE PT PTS ITT1 PP1 ITT2 SP2 ITT3 SP3 DMFS High clinical/pathological risk Low 70-gene risk High 70-gene risk Randomization for patients with discordance between clinical/pathological and 70-gene risk, between chemotherapy and no chemotherapy Randomization for patients assigned to receive chemotherapy, between anthracycline-based regimens, and docetaxel/capecitabine Randomization for endocrinet therapy, between 2 years of Tamoxifen followed by 5 years of Letrozole, and 7 years of Letrozole. All registered patients: All patients who were entered into the preliminary registration for the study. These patients consented to sending their tumor sample to Agendia. However many of them never were enrolled into the study itself. All patients dataset: All patients who were enrolled into the study after receipt of the microarray analysis All patients safety dataset (APS): All patients who were enrolled into the study after receipt of the microarray analysis and who followed the allocated therapeutic strategy (chemo versus no chemo). Primary test dataset: The set of patients who have a low risk gene prognosis signature and high risk clinical-pathological criteria at the time of enrollment, and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy. This dataset is used for a sensitivity analysis to the primary test. Per protocol primary test dataset: The set of patients who were enrolled as having a low risk gene prognosis signature and high risk clinicalpathological criteria and thus receive no chemotherapy, and did not deviate from this: no change in risk status post enrollment and no chemotherapy received.. This dataset is used for the primary test. The subset of patients in the PPT dataset excluding those patients whose samples were received by Agendia between may and January (RNA extraction solution problem). This dataset is used for a second sensitivity analysis for the primary test. Intention-to-treat population for R-T: All patients randomized for R-T will be analyzed in the arm they were allocated by randomization. This is the dataset of all patients with discordant risk assessments at the time of enrollment, because these are the patients who were randomized by the platform at that time. Per protocol population for R-T: All patients who are eligible and randomized for R-T and have started their allocated treatment (at least one dose of chemotherapy if so randomized, respectively no chemotherapy). (Patients who were post enrollment found to have an error in either one or both of the risk assessements are excluded) Intention-to-treat population for R-C: All patients randomized for R-C will be analyzed in the arm they were allocated by randomization. Safety population for R-C: All patients who were randomized for R-C and who have started their allocated chemotherapy (at least one dose of the study drug(s)). Intention-to-treat population for R-E: All patients randomized for R-E will be analyzed in the arm they were allocated by randomization. Safety population for R-E: All patients who were randomized for R-E and who have started their allocated endocrine treatment (at least one dose of the study drug(s)). Distant Metastasis Free Survival EORTC study 10041: analysis plan Version /21

319 EORTC Headquarters March 15, 2016 Roles and responsibilities DFS OS SC CRP Disease Free Survival Overall Survival Study Coordinator Clinical Research Physician EORTC study 10041: analysis plan Version /21

320 Amendment to EORTC protocol (Microarray In Nodenegative Disease may Avoid ChemoTherapy) version 1.1 Amendment 1 PRC submission document Identification EORTC protocol number : Amendment number: 1 Date of submission to PRC: Classification Amendment to protocol document content: X Substantial, i.e. significantly impacting on: Inclusion criteria Blood sampling for future research Technical update for genomic prognosis Change to endocrine therapy Non-substantial Does this amendment concern urgent safety measures already implemented? Yes X No Amendment to trial arrangements (RAU to be consulted): X Substantial Non-substantial Internal PRC final decision on classification: Substantial Non-substantial Internal PRC not yet consulted FO5111 1/44 Template version: November 2005

321 EORTC protocol number, Amendment number We would like to submit a substantial amendment to the protocol, EORTC ( Microarray In Node-negative Disease may Avoid ChemoTherapy) There are 5 main changes: 1) To change the eligibility criteria from lymph node negative to also include 1-3 positive nodes following the validation of the 70-gene signature in this population. 2) As women with 1-3 positive lymph nodes will now be eligible for the study the chemotherapy randomization standard arm has been adapted to reflect this with FEC D as the standard arm chemotherapy regimen for lymph node positive patients. 3) To change the time point of the research blood samples from pre-treatment to pre-operative and add collection of whole blood. These blood samples would remain optional for patients. 4) To reduce the % of tumor cells required in the tumor biopsy sample to perform the genomic prognostic test following the validation with the lower % of cells. 5) To extend the obligation of ovarian function suppression from during the 5 or 7 years of Aromatase Inhibitor to also cover the initial 2 years of tamoxifen treatment. Validation of the genomic prognosis in women with 1-3 positive lymph nodes In the first retrospective validation study, by van de Vijver et al., the 70-gene profile was shown to be prognostic in lymph node-positive patients, with a hazard ratio for distant metastases of 5.5 (95% CI ; p<0.001) (Ref. 29). To further substantiate these results the TRANSBIG consortium conducted a validation study on 241 archival tumor samples from breast cancer patients with 1-3 positive lymph nodes. Patients had a unilateral T1, T2 or operable T3 tumor and were treated with breast conserving therapy or mastectomy with axillary lymph node dissection. One-hundred-seventyfour out of the 241 patients were diagnosed between 1996 and 2001, treated in the Netherlands Cancer Institute, Amsterdam and were up to 71 years of age at diagnosis. The remaining 67 patients were diagnosed between 1994 and 1998, treated at the European Institute of Oncology, Milan and were under the age of 61 years at diagnosis. Twenty patients (8.3%) received no adjuvant systemic therapy, 55 patients (22.8%) were treated with chemotherapy only, 81 patients (33.6%) were treated with endocrine therapy only and 73 patients (30.3%) received both chemo- and endocrine therapy. Frozen tumor samples were evaluated by gene expression profiling for the 70-gene profile using a custom designed microarray measuring the 70 genes in triplicate (MammaPrint). Patients were classified as genomic high risk or genomic low risk. Among the 241 patients, 99 (41%) were classified as genomic low risk and 142 (59%) as genomic high risk. At 5-years, the overall survival probability was 98% (SE 0.01) for the genomic low risk group and 88% (SE 0.03) for the genomic high risk group, respectively. Patients were also classified by Adjuvant! Online, using clinical-pathological criteria. The low clinical risk group was defined as a 10-year breast-cancer specific survival probability of more than 88% and more than 92% for ER positive and ER negative tumors, respectively (see paragraph 1.2.4) The estimated hazard ratio (HR) for the genomic risk was 5.4 (95%CI ; p<0.001) and 4.1 (95%CI ; p=0.002) for OS and distant metastases as first event, respectively, and 4.5 (95%CI ; p=0.002) and 3.4 (95%CI ; p=0.007) when adjusted for the clinical risk as defined by Adjuvant!. These results indicate that the 70-gene profile adds prognostic information to the risk assessment based on the clinical-pathological criteria. The 70-gene profile seemed to perform equally well in both the group of patients treated and those untreated with chemotherapy. In a multivariate analyses, the 70-gene profile, number of positive lymph nodes (3 versus 1) and adjuvant endocrine therapy were the only significant independent factors for prediction of OS with HRs of 5.3 (95%CI ; p=0.006), 3.6 (95%CI ; p=0.001) and 0.3 (95%CI ; p=0.01), respectively. In summary these results show that the 70-gene profile is a strong prognostic marker in breast cancer patients with 1-3 positive lymph nodes and adds prognostic information to the clinical-pathological criteria (paper in preparation). Moreover, the 70-gene profile was able to accurately identify a group of patients with 1-3 positive lymph nodes with an excellent survival who can be safely spared FO5111 2/44 Template version: November 2005

322 chemotherapy. These data provide sufficient evidence to enlarge the inclusion criteria of the to include patients with 1-3 positive lymph nodes as approved by the TRANSBIG and Steering Committees. With the change in the eligibility criteria to allow for 1-3 positive nodes, the standard - anthracycline arm of the chemotherapy question will be adapted for lymph node positive patients. The standard chemotherapy regimen for 1-3 positive node patients will be 3 cycles of FEC100 follow by 3 cycles of docetaxel (100 mg/m²). Standard arm Experimental arm N- One of the 5 possible anthracycline based Docetaxel - Capecitabine regimens (no changes) N+ FEC D Docetaxel - Capecitabine No adjustment of statistical assumptions at this time point The statistical assumptions have not been reviewed and adjusted at this time point as there are only 200 patients enrolled in the study and it was felt to be too small a sample to base any review on. There is a pre-planned review of the logistics and statistical assumptions at 800 patients and it was felt that this would be a more appropriate time to review the statistical assumptions of the trial. Also at this time we will have a better idea of the proportion of lymph node positive patients entered and this will enable a more accurate re-assessment of the expected global population. Blood sampling, change time-point and add whole blood sample Originally the blood sample for serum storage was planned after the primary breast cancer surgery and before the start of adjuvant treatment. A consensus from the Proteomics working group requested that the blood sample be taken before the primary surgery when the patient has the greatest tumor burden to facilitate the search for molecular markers linked to the presence of disease. In addition, whole blood will be taken for DNA extraction to enable future pharmacogenomic analysis (genetic predictors of response/side effects) to be performed. Reduce the % of tumor cells necessary to perform the genomic prognostic test For the development and validation of the genomic prognostic test (microarray) a cut-off of more than 50% tumor cells in the biopsy was used. Since the initial validation Agendia, the Biotechnology company that is performing the test, has validated the use of 30% tumor cells in the biopsy sample for Mammaprint their FDA cleared commercial molecular diagnostic test and for the test determined using whole genome arrays as used in. In their commercial activities they now routinely use samples with more than 30% tumor cells so we would like to reduce the percentage of biopsy tumor cellularity to the same as the current standard. Make ovarian function suppression mandatory during the whole duration of protocol endocrine therapy Currently, pre-menopausal women participating in the endocrine therapy randomization can chose in agreement with their physician whether to have ovarian function suppression during the initial 2 years of tamoxifen treatment. It was felt that this may induce a bias between the two arms so we would like to make ovarian function suppression mandatory also during the 2 years of tamoxifen treatment and hence the endocrine therapy randomization will only be proposed to menopausal patients either spontaneously or induced. - This amendment has been discussed and agreed by: - Study Chairpersons, TRANSBIG and Steering Committees - National Coordinating Groups/Centres - Companies: Novartis FO5111 3/44 Template version: November 2005

323 Hoffman la Roche Sanofi-Aventis FO5111 4/44 Template version: November 2005

324 1. Modifications related to the present amendment Current protocol 10041, 1.1 & Page 9, Objectives (Microarray In Node-negative Disease may Avoid ChemoTherapy) A prospective, randomized study comparing the Amsterdam 70-gene expression signature (70- gene signature) with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer. PRIMARY 1. The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to node-negative breast cancer patients. 2. The trial will also aim to compare a docetaxelcapecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens. Page 9 To compare the OS distributions associated with the 2 chemotherapy regimens To determine and compare the early and late toxicities associated with each regimen. Page 10 Long term research objectives: To set up several biological material bank resources (RNA, tumor tissue, serum) for future translational studies in both genomics and proteomics. Page 10, Diagnosis & main inclusion criteria women with histologically proven operable invasive breast cancer and a negative sentinel Amendment 2.0 (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy) A prospective, randomized study comparing the Amsterdam 70-gene expression signature (70- gene signature) with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0 to 3 positive nodes. PRIMARY 1. The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to breast cancer patients with 0 to 3 positive lymph nodes. 2. The trial will also aim to compare a docetaxelcapecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens (anthracyclines followed by docetaxel for node positive). To compare the OS distributions associated with the 2 chemotherapy arms To determine and compare the early and late toxicities associated with each arm. To set up several biological material bank resources (RNA, tumor tissue, blood & serum) for future translational studies in both genomics and proteomics. women with histologically proven operable invasive breast cancer, with 0 to 3 positive FO5111 5/44 Template version: November 2005

325 Current protocol 10041, 1.1 & node or negative axillary clearance (N0, M0). Page 10 the breast tumor must be unilateral, however DCIS or LCIS are allowed if invasive cancer is present. have adequate bone marrow reserves (neutrophil count >1.5 x109 /l and platelet count >100 x109/l), adequate renal function (creatinine clearance 50 ml/min (calculated according to Cockroft and Gault, see Appendix C), or serum creatinine 1.5 x upper limit of normal), hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline phosphatase 2.5 x upper limit of normal, total bilirubin 2.0 x upper limit of normal) and normal ECG compatible with chemotherapy administration. NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any cancer (other than breast cancer) in complete remission for 5 years. Page 11, Diagnosis & main inclusion criteria while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, injectables, combined oral contraceptives, some IUDs, sexual abstinence or vasectomised partner (Note 3 of the guidance on nonclinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). Page 12, Treatment The following women, if eligible, will be offered the chemotherapy randomization (R-C) comparing a commonly used anthracyclinebased regimen, selected from the list of acceptable regimens, and the new capecitabinedocetaxel combination: Page 12, Reference therapy, dose and mode of Amendment 2.0 lymph nodes and no distant metastases. the breast tumor must be unilateral. Multi focal tumors are allowed provided that they have identical histology. DCIS or LCIS are allowed if invasive cancer is present. have adequate bone marrow reserves (neutrophil count >1.5 x109 /l and platelet count >100 x109/l), adequate renal function (serum creatinine 1.5 x upper limit of normal), hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline phosphatase 2.5 x upper limit of normal and total bilirubin 2.0 x upper limit of normal). NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any invasive cancer (other than breast cancer) in complete remission for 5 years. while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, some IUDs, condoms, sexual abstinence or vasectomised partner (Note 3 of the guidance on nonclinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). The following women, if eligible, will be offered the chemotherapy randomization (R-C) comparing a commonly used anthracyclinebased regimen (anthracyclines followed by docetaxel for node positive), selected from the list of acceptable regimens, and the new capecitabine-docetaxel combination: FO5111 6/44 Template version: November 2005

326 Current protocol 10041, 1.1 & administration For the chemotherapy randomization (R-C) the standard arm is an anthracycline based chemotherapy regimen chosen from the list of allowed regimens. Page 12, Treatment, Test product, dose and mode of administration All postmenopausal endocrine-responsive (i.e. ER and/or PgR positive) patients on the study will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. It will be left to the decision of the investigator whether or not a premenopausal woman can be randomized into this question, with concomitant ovarian function suppression while taking letrozole. Page 13, Statistical methods Patients who are candidates for chemotherapy (see also treatment section) will be proposed for randomization R-C: between anthracycline based chemotherapy and docetaxel-capecitabine chemotherapy. Page 14, figure Delay between surgery and start of treatment ideally not more than 6 weeks, can not be > 8 weeks Visit Surgeon (T1, T2, or operable T3 & cn0 & M0) pn0 Page 14, figure R-C Chemotherapy Randomization of Anthracycline based CT vs Docetaxel/ Capecitabine Page 15, Trial Organization The investigational drugs (Docetaxel, Capecitabine and Letrozole) will be supplied by the industrial partners. Page 16, 1.1 General Introduction Current standard of care for those patients who Amendment 2.0 For the chemotherapy randomization (R-C) the standard arm is an anthracycline based chemotherapy regimen (anthracyclines followed by docetaxel for node positive) chosen from the list of allowed regimens. All postmenopausal endocrine-responsive (i.e. ER and/or PgR positive) patients on the study will be offered randomization (R-E) between 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. It will be left to the decision of the investigator whether or not a premenopausal woman can be randomized into this question, with concomitant ovarian function suppression (surgery, LHRH agonists or ovarian radiotherapy) during the complete duration of endocrine therapy. Patients who are candidates for chemotherapy (see also treatment section) will be proposed for randomization R-C: between anthracycline based chemotherapy (anthracyclines followed by docetaxel for node positive) and docetaxelcapecitabine chemotherapy. figure see attached Delay between surgery and start of treatment ideally not more than 6 weeks, can not be > 12 weeks Visit Surgeon (T1, T2, or operable T3 & M0) 0 to 3 positive nodes R-C Chemotherapy Randomization of Anthracycline based CT (FEC D for N+) vs Docetaxel/ Capecitabine The investigational drugs (Docetaxel*, Capecitabine and Letrozole) will be supplied by the industrial partners. *Footnote the docetaxel in the standard arm FEC D for lymph node positive patients will not be provided by the industrial partners Current standard of care for node negative FO5111 7/44 Template version: November 2005

327 Current protocol 10041, 1.1 & will receive chemotherapy is 6 cycles of anthracycline based chemotherapy with some experts favoring the incorporation of taxanes for node-positive disease. For patients with endocrine responsive disease, Page 16, 1.1 General Introduction In the last 20 years, little progress has been made with respect to assisting oncologists in determining which node-negative breast cancer patients require chemotherapy or other systemic therapy and which women can safely be treated with only loco-regional treatment. Great discomfort still persists in selecting those nodenegative women who need adjuvant chemotherapy (Ref. 6). Page 16, 1.2 Risk assessment in node-negative breast cancer Page 17, 1.2 Despite nodal status being one of the strongest prognostic factors used for predicting breast cancer clinical outcome, the Early Breast Cancer Trialist Collaborative Group (EBCTCG) overview recently reported Page 17, 1.2 There is however, much controversy related to the optimal definition of a low/minimal versus a moderate/high risk of relapse for these women with node-negative breast cancer. Page 17, 1.2 For women who have node-negative breast cancer, it is important to be able to accurately quantify the risk of recurrence and death so that the decisions about adjuvant chemotherapy can be taken appropriately. Page 17, Node-negative early breast cancer: who can be safely spared adjuvant chemotherapy? Page 17, Unfortunately little progress has been made with regards to new prognostic markers that can assist oncologists in treatment decision-making for node-negative early stage breast cancer. Amendment 2.0 patients for those patients who will receive chemotherapy is 6 cycles of anthracycline based chemotherapy with the incorporation of taxanes for node-positive disease. For patients with endocrine responsive disease, In the last 20 years, little progress has been made with respect to assisting oncologists in determining which early breast cancer patients require chemotherapy or other systemic therapy and which women can safely be treated with only loco-regional treatment. Great discomfort still persists in selecting those node-negative women and women with 1-3 positive nodes who need adjuvant chemotherapy (Ref. 6). 1.2 Risk assessment in early breast cancer Despite nodal status being historically one of the strongest prognostic factors used for predicting breast cancer clinical outcome, the Early Breast Cancer Trialist Collaborative Group (EBCTCG) overview recently reported There is however, much controversy related to the optimal definition of a low/minimal versus a moderate/high risk of relapse for these women with early breast cancer. For women who have early breast cancer, it is important to be able to accurately quantify the risk of recurrence and death so that the decisions about adjuvant chemotherapy can be taken appropriately Early breast cancer: who can be safely spared adjuvant chemotherapy? Unfortunately little progress has been made with regard to new prognostic markers that can assist oncologists in treatment decision-making for early stage breast cancer. FO5111 8/44 Template version: November 2005

328 Current protocol 10041, 1.1 & Amendment 2.0 Page Patients with 1 to 3 positive lymph nodes: Do all of them benefit from chemotherapy? The Oxford Overview published in 2005 suggested that the addition of chemotherapy to tamoxifen was beneficial for patients with hormone receptor positive cancer (1). But despite the clear difference it is evident that not all the patients obtain the same benefit. For example the benefit provided by chemotherapy on top of endocrine therapy varies according to patient s age, and there may be other situations where the addition of chemotherapy may not be translated in clinical benefit. An example of this can be found in an exploratory analysis of the SWOG 8814 trial performed by Albain et al (2). This retrospective analysis suggested that the subgroup of patients with ER positive, HER-2 negative and 1-3 positive lymph nodes did not obtain any benefit from the adition of chemotherapy to endocrine therapy. Other reports also suggest that not all patients with 1-3 positive lymph nodes may require chemotherapy (3) This data, in addition to the ongoing research on Luminal A type tumors, warrants further investigations aimed to the identification of a subgroup of patients with 1-3 positive lymph nodes who may safely be spared from chemotherapy. The St Gallen Consensus Guidelines consider 1-3 positive nodes as an intermediate risk group which, depending on other biological characteristics, may not necessarily need adjuvant chemotherapy (Ref 4) particularly in the case of endocrine responsive disease. Refs 1. Early Breast Cancer Trialists Collaborative Group. Effects of chemotherapy and hormonal FO5111 9/44 Template version: November 2005

329 Current protocol 10041, 1.1 & Amendment 2.0 therapy for early breast cancer on recurrence and 15-year survival: an overview of the randomized trials. Lancet. 2005;365: Albain K, Barlow W, O Malley F, et al. Concurrent (CAFT) versus sequential (CAF-T) chemohormonal therapy (cyclophosphamide, doxorubicin, 5-fluorouracil, tamoxifen) versus T alone for postmenopausal node-positive, estrogen (ER) and/or progesterone (PgR) receptor-positive breast cancer: mature outcomes and new biologic correlates on phase III intergroup trial 0100 (SWOG 8814). Breast Cancer Res Treat. 2005;90: Harbeck N, Kates RE, Look MP, Meijer-van Gelder ME, Klijn JGM, Jänicke F, Krüger A, Kiechle M, Schmitt M, Foekens JA. Enhanced benefit from adjuvant systemic chemotherapy in breast cancer patients classified high-risk according to upa and PAI-1 (n=3,424). Cancer Res 62 (16): , Page 20, Three years ago, in an attempt to better understand breast cancer biology and to find new clinically relevant molecular makers, the Netherlands Cancer Institute looked at Page 21 Page 21 The results of the validation study have been submitted for publication. Since the initial validation, the 70-gene signature has also been validated in a similar population of women aged from years. Page 21 signature has also been validated in a similar population of women aged from years. 4. A. Goldhirsch et al 2007, Progress and promise: highlights of the international expert consensus on the primary therapy of early breast cancer Annals of Oncology 18: , 2007 Six years ago, in an attempt to better understand breast cancer biology and to find new clinically relevant molecular makers, the Netherlands Cancer Institute looked at add sub-section title Initial validation in lymph node negative patients The results of the validation study have been published (Ref X). Since the initial validation, the 70-gene signature has also been validated in a similar population of women aged from years. Buyse et al, Journal of the National Cancer Institute, Vol. 98, No. 17, September 6, 2006 signature has also been validated in a similar population of women aged from years Subsequent validation in lymph node- FO /44 Template version: November 2005

330 Current protocol 10041, 1.1 & In summary, the TRANSBIG consortium has completed the first independent and external Amendment 2.0 positive patients In the first retrospective validation study, by van de Vijver et al., the 70-gene profile was shown to be prognostic in lymph nodepositive patients, with a hazard ratio for distant metastases of 5.5 (95% CI ; p<0.001) (Ref. 29). To further substantiate these results the TRANSBIG consortium conducted a validation study in breast cancer patients with 1-3 positive lymph nodes. This study was based on 241 archival tumor samples from breast cancer patients with 1-3 positive lymph nodes. Patients had a unilateral T1, T2 or operable T3 tumor and were treated with breast conserving therapy or mastectomy with axillary lymph node dissection. One-hundred-seventy-four out of the 241 patients were diagnosed between 1996 and 2001, treated in the Netherlands Cancer Institute, Amsterdam and were up to 71 years of age at diagnosis. The remaining 67 patients were diagnosed between 1994 and 1998, treated at the European Institute of Oncology, Milan and were under the age of 61 years at diagnosis. Twenty patients (8.3%) received no adjuvant systemic therapy, 55 patients (22.8%) were treated with chemotherapy only, 81 patients (33.6%) were treated with endocrine therapy only and 73 patients (30.3%) received both chemo- and endocrine therapy. Frozen tumor samples were evaluated by gene expression profiling for the 70-gene profile using a custom designed microarray measuring the 70 genes in triplicate (MammaPrint). Patients were classified as genomic high-risk or genomic low-risk. Among the 241 patients, 99 (41%) were classified as genomic low-risk and 142 (59%) as genomic high-risk. At 5-years, the overall survival probability was 98% (SE 0.01) for the genomic low-risk group and 88% (SE 0.03) for the genomic high-risk group, FO /44 Template version: November 2005

331 Current protocol 10041, 1.1 & Amendment 2.0 respectively. Patients were also classified by Adjuvant! Online, using clinical-pathological criteria. The low clinical risk group was defined as a 10-year breast-cancer specific survival probability of more than 88% and more than 92% for ER positive and ER negative tumors, respectively (see paragraph 1.2.4) The estimated hazard ratio (HR) for the genomic risk was 5.4 (95%CI ; p<0.001) and 4.1 (95%CI ; p=0.002) for OS and distant metastases as first event, respectively, and 4.5 (95%CI ; p=0.002) and 3.4 (95%CI ; p=0.007) when adjusted for the clinical risk as defined by Adjuvant!. These results indicate that the 70-gene profile adds prognostic information to the risk assessment based on the clinical-pathological criteria. The 70-gene profile seemed to perform equally well in both the group of patients treated and those untreated with chemotherapy. In a multivariate analyses, the 70-gene profile, number of positive lymph nodes (3 versus 1) and adjuvant endocrine therapy were the only significant independent factors for prediction of OS with HRs of 5.3 (95%CI ; p=0.006), 3.6 (95%CI ; p=0.001) and 0.3 (95%CI ; p=0.01), respectively. In summary these results show that the 70- gene profile is a strong prognostic marker in breast cancer patients with 1-3 positive lymph nodes and adds prognostic information to the clinical-pathological criteria (paper in preparation). Moreover, the 70-gene profile can accurately identify a group of patients with 1-3 positive lymph nodes with an excellent survival who can be safely spared chemotherapy. These data provide sufficient evidence to enlarge the inclusion criteria of the to include patients with 1-3 positive lymph nodes (ref X). FO /44 Template version: November 2005

332 Current protocol 10041, 1.1 & Amendment 2.0 Mook S, Schmidt MK, Viale G, Pruneri G, Eekhout I, Piccart MJ, Cardoso F, Rutgers EJ and Van t Veer LJ, on behalf of the TRANSBIG consortium. Breast cancer patients with 1-3 positive lymph nodes and a low risk 70-gene profile have an excellent survival. Breast Cancer Research and Treatment 106 (supp 1): late breaking abstract #50 Page 22, section The success of this validation study allowed for the commencement of the large collaborative trial (Microarray In Node-negative Disease may Avoid ChemoTherapy), which will be the first prospective trial to validate a molecular-based signature for the adjuvant treatment of node negative early breast cancer patients. will enroll 6,000 patients to investigate the benefit/risk ratio of chemotherapy when the assessment of prognosis based on clinical-pathological features differs from that provided by the gene signature. It will use the 70-gene signature to classify node-negative early stage breast cancer patients into high and low risk of distant relapse and to compare this assessment to the Adjuvant! Online software risk assessment, which is widely used in clinical practice today and which takes into consideration traditional clinical-pathological factors. Page 22, For those patients receiving chemotherapy, an optional randomization will compare anthracyline-based regimens to a new docetaxelcapecitabine combination (see section 1.3). Page 23, section 1.3 In summary, the TRANSBIG consortium has completed the first independent and external The success of these validation studies allowed for the commencement of the large collaborative trial (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy), which will be the first prospective trial to validate a molecular-based signature for the adjuvant treatment of early breast cancer patients with 0 to 3 positive lymph nodes. will enroll 6,000 patients to investigate the benefit/risk ratio of chemotherapy when the assessment of prognosis based on clinical-pathological features differs from that provided by the gene signature. It will use the 70-gene signature to classify early stage breast cancer patients into high and low risk of distant relapse and to compare this assessment to the Adjuvant! Online software risk assessment, which is widely used in clinical practice today and which takes into consideration traditional clinical-pathological factors. For those patients receiving chemotherapy, an optional randomization will compare anthracyline-based regimens (anthracyclines followed by docetaxel for node positive patients) to a new docetaxel-capecitabine combination (see section 1.3). FO /44 Template version: November 2005

333 Current protocol 10041, 1.1 & Chemotherapy in the trial: can anthracycline-based chemotherapy be safely replaced by the combination of docetaxelcapecitabine as adjuvant treatment of nodenegative breast cancer? Page 23, Regimens such as FEC100, CAF/FAC and CEF, for 4 to 6 cycles, are considered the treatment of choice for node-positive and high risk nodenegative patients, based on the results of several randomized trials (Ref. 8;41-43) and the EBCTCG Overview. Page 25, Regimens comprising anthracyclines and taxanes, either concomitantly (e.g. TAC) or in sequence (e.g. EC/EC followed by taxane) Page 26, Chemotherapy in the trial The trial will ask an important question regarding chemotherapy optimization for nodenegative breast cancer: can a docetaxel-based regimen safely replace an anthracycline-based one? Page 26, In addition to their activity as single agents, promising preliminary results of their combined activity and the extent of experience with these two agents encourages their further investigation particularly in node-negative disease. Page 26, hopes to validate this new nonanthracycline-based chemotherapy regimen for use in high-risk women with node-negative breast cancer and to generate long-term safety data on this combination. Page 28, Due to the increasing use of screening mammography in women of 50 years of age in developed countries, the prevalence of nodenegative breast cancer has been continually increasing. Adjuvant therapy for women with node-negative disease is therefore of prime importance for the treating physician today. Amendment Chemotherapy in the trial: can anthracycline-based chemotherapy be safely replaced by the combination of docetaxelcapecitabine as adjuvant treatment of early breast cancer? Regimens such as FEC100, CAF/FAC and CEF, for 4 to 6 cycles, are considered the treatment of choice for high risk node-negative patients, based on the results of several randomized trials (Ref. 8;41-43) and the EBCTCG Overview. Regimens comprising anthracyclines and taxanes, either concomitantly (e.g. TAC) or in sequence (e.g. EC/FEC followed by taxane) The trial will ask an important question regarding chemotherapy optimization for early breast cancer: can a docetaxel-based regimen safely replace an anthracycline-based one? In addition to their activity as single agents, promising preliminary results of their combined activity and the extent of experience with these two agents encourages their further investigation. hopes to validate this new nonanthracycline-based chemotherapy regimen for use in high-risk women with early breast cancer and to generate long-term safety data on this combination. Due to the increasing use of screening mammography in women of 50 years of age in developed countries, the prevalence of nodenegative breast cancer or with few positive nodes has been continually increasing. Adjuvant therapy for these women is therefore of prime importance for the treating physician today. FO /44 Template version: November 2005

334 Current protocol 10041, 1.1 & Page 28, table 2 ²Relapse free survival Page 33, 2.1 The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to node-negative breast cancer patients. The trial will also aim to compare a docetaxelcapecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens. Page 34, 2.1 To compare the OS distributions associated with the 2 chemotherapy regimens. Page 35, 3.1 General eligibility criteria for inclusion in the trial women with histologically proven operable invasive breast cancer and a negative sentinel node or negative axillary clearance (N0, M0). Page 35, 3.1 the breast tumor must be unilateral, however DCIS or LCIS are allowed if invasive cancer is present. Page 35, 3.1 have adequate bone marrow reserves (neutrophil count >1.5 x109 /l and platelet count >100 x109), adequate renal function (creatinine clearance 50 ml/min (calculated according to Cockroft and Gault, see Appendix C), or serum creatinine 1.5 x upper limit of normal), hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline phosphatase 2.5 x upper limit of normal, total bilirubin 2.0 x upper limit of normal) and normal ECG compatible with chemotherapy administration. ²Disease free survival Amendment 2.0 The primary objective of the trial is to demonstrate the superiority of the molecular profiling approach over the usual clinical assessment in assigning adequate risk categories (and the need to receive adjuvant chemotherapy or not) to breast cancer patients with 0 to 3 positive nodes. The trial will also aim to compare a docetaxelcapecitabine regimen possibly associated with increased efficacy and reduced long-term toxicities to existing commonly used anthracycline-based chemotherapy regimens (anthracyclines followed by docetaxel for node positive). To compare the OS distributions associated with the 2 chemotherapy arms. women with histologically proven operable invasive breast cancer, 0 to 3 positive nodes and no distant metastases. the breast tumor must be unilateral. Multi focal tumors are allowed provided that they have identical histology. DCIS or LCIS are allowed if invasive cancer is present. have adequate bone marrow reserves (neutrophil count >1.5 x109 /l and platelet count >100 x109), adequate renal function (serum creatinine 1.5 x upper limit of normal), hepatic function (ALAT, ASAT 2.5 x upper limit of normal, alkaline phosphatase 2.5 x upper limit of normal and total bilirubin 2.0 x upper limit of normal). FO /44 Template version: November 2005

335 Current protocol 10041, 1.1 & Page 36, 3.1 NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any cancer (other than breast cancer) in complete remission for 5 years. Page 36, 3.1 while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, injectables, combined oral contraceptives, some IUDs, sexual abstinence or vasectomised partner (Note 3 of the guidance on non-clinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). Page 37, 3.3 WHO status 0 or 1 (see Appendix B) have cardiac function Page 37, 3.3 interval between definitive surgery and start of chemotherapy should ideally be no more than 6 weeks but can not exceed 8 weeks. Page 37, 3.4 premenopausal women must have ovarian function suppression during endocrine therapy with aromatase inhibitor (see 7.3 for details). Page 38, 4 It will also assess the efficacy of commonly used anthracycline-based chemotherapy regimens, selected from a list of acceptable regimens, to the new capecitabine-docetaxel combination. Page 38, 4 Node-negative women, will be offered those randomizations for which they are eligible, as shown below. Page 38, 4 Page 39, 4.1 Amendment 2.0 NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any invasive cancer (other than breast cancer) in complete remission for 5 years. while taking study medications patients should take adequate birth control measures which result in low failure rates (i.e. less then 1% per year) when used consistently and correctly such as implants, some IUDs, condoms, sexual abstinence or vasectomised partner (Note 3 of the guidance on non-clinical safety studies for the conduct of human clinical trials for pharmaceuticals CPMP/ICH/286/95). WHO status 0 or 1 (see Appendix B) normal ECG compatible with chemotherapy administration have cardiac function interval between definitive surgery and start of chemotherapy should ideally be no more than 6 weeks but can not exceed 12 weeks. premenopausal women must have ovarian function suppression during the whole of the duration of protocol endocrine therapy (see 7.3 for details). It will also assess the efficacy of commonly used anthracycline-based chemotherapy regimens (anthracyclines followed by docetaxel for node positive), selected from a list of acceptable regimens, to the new capecitabine-docetaxel combination. Women with 0 to 3 positive lymph nodes will be offered those randomizations for which they are eligible, as shown below. trial scheme figure see attached & as p14 FO /44 Template version: November 2005

336 Current protocol 10041, 1.1 & Prior to entering into the trial, at the time of surgery, when patient eligibility is not known, all patients who sign a screening PIS & IC to consent to the donation of a tumor sample, will be registered for screening. Page 39, 4.2 The date when the patient signs the PIS & IC 1 (after both the genomic and clinical risk assessments have been performed) is the date the patient enters the trial. This date will be referred to as the ENROLLMENT DATE. There are 3 possible randomizations in this trial and each of the 3 will only randomize a sub-set of the enrolled patients. Page 39, 4.4 The date when the patient signs the PIS & IC 1 (after both the genomic and clinical risk assessments have been performed) is the date the patient enters the trial. This date will be referred to as the ENROLLMENT DATE. Page 40, 4.4 Patients who are candidates for chemotherapy (also see treatment section) will be proposed randomization R-C between anthracycline based chemotherapy or docetaxel-capecitabine chemotherapy. Page 40, 4.4 The objective is to evaluate if the docetaxelcapecitabine regimen is a more effective and less toxic alternative to an anthracycline-based regimen, for high-risk node-negative patients. Page 41, 4.5 Pre-menopausal patients may be offered this randomization at the discretion of their treating physician. For these patients, the suppression of ovarian function (temporarily or definitively) is mandatory (until the patient is > 50 years for temporary suppression). Page 43, these hot nodes are not to be removed). Axillary Lymph Node Dissection (ALND) Traditionally, the indication for ALND Amendment 2.0 Prior to entering into the trial, at the time of surgery, when patient eligibility is not known, all patients who sign a screening PIS & IC to consent to the donation of a tumor and blood sample, will be registered for screening. The date when the patient signs the PIS & IC 1 (after both the genomic and clinical risk assessments have been performed) will be referred to as the ENROLLMENT DATE. There are 3 possible randomizations in this trial and each of the 3 will only randomize a sub-set of the enrolled patients. The date when the patient signs the PIS & IC 1 (after both the genomic and clinical risk assessments have been performed) will be referred to as the ENROLLMENT DATE. Patients who are candidates for chemotherapy (also see treatment section) will be proposed randomization R-C between anthracycline based chemotherapy (anthracyclines followed by docetaxel for node positive) or docetaxelcapecitabine chemotherapy. The objective is to evaluate if the docetaxelcapecitabine regimen is a more effective and less toxic alternative to an anthracycline-based regimen, for high-risk patients with 0 to 3 positive lymph nodes. Pre-menopausal patients may be offered this randomization at the discretion of their treating physician. For these patients, the suppression of ovarian function (temporarily or definitively) is mandatory during the whole duration of protocol endocrine therapy. these hot nodes are not to be removed). Local pathology categorization of lymph node status micromets (0.2mm - 2mm) pn1mi are FO /44 Template version: November 2005

337 Current protocol 10041, 1.1 & Amendment 2.0 considered as LN positive submicromets (<0.2mm) pn0(i+) are considered as LN negative Management of sentinel lymph node micrometastasis Patients with micrometastasis (0.2 to 2 mm) in a SLN should be considered as LN positive and undergo a complete axillary dissection unless they are randomized in a clinical trial evaluating treatment options for this patient population. In such case, patients can follow the allocated treatment. Page 43, 5.2 Studies have reported similar rates of locoregional recurrence in patients undergoing mastectomy without radiotherapy and patients undergoing complete tumor excision without mastectomy followed by radiotherapy to the breast. Radiotherapy to the breast is mandatory in all patients whose primary tumor surgery comprises a complete local excision without mastectomy. Radiotherapy is not mandatory for patients who have undergone a mastectomy. Tumor bed irradiation is mandatory for patients with non-resectable deep margin invasion. In order to minimize cardiac irradiation, all patients who receive internal mammary node (IMN) irradiation must have the radiotherapy planned using a technique that enables minimization of cardiac irradiation, preferably 3- D conformational CAT scan. This radiotherapy field is mandatory in patients with confirmed or suspicious IMN spread (by sentinel node technique) and optional in patients with tumors judged to be at high risk of IMN spread (such as central and medial hemisphere tumors). Page 44, 6.1 Treatment plan, anthracycline arm Axillary Lymph Node Dissection (ALND) Traditionally, the indication for ALND Radiotherapy to the breast is mandatory for all patients who have undergone breastconserving surgery. The use of a boost dose to the tumor bed should be considered, especially for patients 60 years of age or less, and is mandatory for patients with histologically incomplete tumor resection margins. Chest wall radiotherapy after mastectomy is optional, but mandatory for patients with incomplete tumor resection margins. Regional nodal irradiation (axillary, periclavicular and /or internal mammary chain) should be given according to local or national guidelines. For all patients, care should be taken to minimize exposure to heart and lung tissue. To achieve this, the use of a computer tomography (CT) based treatment simulation and planning is recommended, and is mandatory if the internal mammary lymph node chain is part of the target volume. 6.1 Treatment plan, anthracycline arm FO /44 Template version: November 2005

338 Current protocol 10041, 1.1 & The anthracycline arm will include Page 44, 6.1 policies to participate in the trial. Page 44, around 360 mg/m² for doxorubicin and 600 mg/m² for epirubicin. Page 44, The following are the accepted regimens and doses that can be used in the trial: FEC 100: Fluorouracil 500 mg/m² Page 45, mg/m², 50 mg/m², 600 mg/m², days 1) every 3 weeks for 4 cycles (Ref. 88) Amendment 2.0 The Anthracycline arm will be different for patients with lymph-node negative breast cancer and for patients with 1 to 3 positive lymph nodes, as the current standard therapies are different for each subgroup. Lymph node negative The anthracycline arm will include policies to participate in the trial. One to 3 positive lymph nodes The anthracycline arm will include one chemotherapy regimen, consisting of the sequence of 3 cycles of FEC 100 followed by 3 cycles of Docetaxel. Cycles should be given every 3 weeks. This regimen is currently perceived as standard treatment in several EU countriesfor this subpopulation, and therefore it will be the standard anthracycline-based treatment for patients with 1-3 positive nodes in the chemotherapy question. around 360 mg/m² for doxorubicin and 600 mg/m² for epirubicin. For toxicity information regarding Docetaxel, used in the FEC-D regimen for lymph node positive disease, please refer to section The following are the accepted regimens and doses that can be used in the trial: Options for node negative patients: FEC 100: Fluorouracil 500 mg/m² mg/m², 50 mg/m², 600 mg/m², days 1) every 3 weeks for 4 cycles (Ref. 88) Option for Node positive patients: FEC D: Fluorouracil 500 mg/m2, Epirubicin 100 mg/m2, and Cyclophosphamide 500 mg/m2 (FEC) intravenously on day 1 every 3 weeks for 3 FO /44 Template version: November 2005

339 Current protocol 10041, 1.1 & Page 45, The first cycle of chemotherapy should be administered within 4 weeks following the randomization R-C date and ideally no more than 6 weeks after surgery. If treatment has not started within 8 weeks of surgery the patient becomes ineligible. Page 45, If there is a 10% or greater decrease in body weight compared to baseline, the BSA will be recalculated. Page 45, Diagram anthracycline based regimens see attached Page 46, No primary prophylaxis with G-CSF/GM- CSF/PEG-filgrastim is allowed. Page 47, Table 3 Wait until ANC > 1.0x109/l (maximum time 3 weeks), then Pages 47 & 48 Tables 3 & 4 See attached add docetaxel dose reductions Page 51, Docetaxel 75 mg/m² should be administered as a 1-hour i.v. infusion, on day 1. Capecitabine 825 mg/m² will be given Page 51, For capecitabine, the daily dose will be derived by determining the body surface area (BSA). Use the tables of capecitabine doses in Table 5 to determine the appropriate dose for a given BSA. If body weight varies during the study, Amendment 2.0 cycles followed by Docetaxel 100 mg/m2 intravenously on day 1 every 3 weeks for 3 cycles (Ref. 55) The first cycle of chemotherapy should be administered within 4 weeks following the randomization R-C date and ideally no more than 8 weeks after surgery. If treatment has not started within 12 weeks of surgery the patient becomes ineligible. Page 45, If there is a 10% or greater change in body weight compared to baseline, the BSA will be recalculated. Add FEC D regimen No primary prophylaxis with G-CSF/GM- CSF/PEG-filgrastim is allowed. Pre-medication for docetaxel infusions is described in Wait until ANC > 1.0x109/l (maximum time up to 3 weeks), then Docetaxel 75 mg/m² should be administered as a 1-hour i.v. infusion, on day 1. If the calculated BSA of the patient is >2.2 m², the dose to be given to the patient will be calculated according to BSA = 2.2m². No ideal body weight should be used for the calculation of BSA. Capecitabine 825 mg/m² will be given For capecitabine, the daily dose will be derived by determining the body surface area (BSA). Use the tables of capecitabine doses in Table 5 to determine the appropriate dose for a given BSA. If the calculated BSA of the patient is >2.2 m², the dose to be given to the patient will be calculated according to BSA = 2.2m². If body weight varies during the study, FO /44 Template version: November 2005

340 Current protocol 10041, 1.1 & Page 51, Capecitabine (Xeloda ) will be provided as filmcoated 150 mg and 500 mg tablets, packed in polyethylene bottles containing 56 or 120 tablets. Tablets are not scored and should not be split. Page 52, table 5 see attached Page 54, Table 7 Wait until ANC > 1.0x109/l (maximum time 3 weeks), then Page 57, Table 9 see attached Page 60, 6.3 The patient will then be considered off-protocol chemotherapy treatment, but should be followed according to the protocol page 61, 7.3 All premenopausal women (until the age of 50 years) at diagnosis who decide to participate in the endocrine randomization in the trial must have adequate ovarian function suppression while taking letrozole. Each centre will decide, upfront, their policy regarding whether they will also provide ovarian function suppression for all of their premenopausal patients during the initial 2 years of tamoxifen treatment. Page 61, 7.3 Bilateral ovarian irradiation. Radiation should be given according to accepted guidelines. Biochemical verification of ovarian function suppression is required by 2 months using Page 62, 8.1 ECG (the pre-surgical ECG can be used provided it was normal) and within 21 days of the start of treatment for women receiving either chemotherapy or endocrine therapy: Page 63, To be performed within 21 days of randomization R-C and before the start of chemotherapy treatment. (see the summary table 11 below) Amendment 2.0 Capecitabine (Xeloda ) will be provided as filmcoated 150 mg and 500 mg tablets. Tablets are not scored and should not be split. Wait until ANC > 1.0x109/l (maximum time 3 up to weeks), then The patient will then be considered off-protocol chemotherapy treatment (however the patient remains eligible for R-E), but should be followed according to the protocol All premenopausal women (until the age of 50 years) at diagnosis who decide to participate in the endocrine randomization in the trial must have adequate ovarian function suppression or ablation for the whole duration of randomized endocrine therapy treatment in both arms. Bilateral ovarian irradiation. Radiation should be given according to accepted guidelines. Biochemical verification of ovarian function suppression is required after 2 months using and within a clinically acceptable period of the start of treatment for women receiving either chemotherapy or endocrine therapy: To be performed within a clinically acceptable period before randomization R-C. (see the summary table 11 below) FO /44 Template version: November 2005

341 Page 63, Cardiac function Current protocol 10041, 1.1 & Page 63, To be performed within 21 days of randomization R-E and before the start of endocrine therapy treatment. Page 66, Table 11 (see table attached) Page 69, The primary test for R-C is to compare DFS between the two chemotherapy regimens randomized. Page 70, R-T: Patients for whom both methods are stratifying for institution, risk group (high risk genetic/low risk clinical vs. low risk genetic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), age Page 70, For R-C, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genetic/low risk clinical vs. low risk genetic/high risk clinical vs. high risk genetic/high risk clinical), Page 70, For R-E, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genetic/high risk clinical vs. high risk genetic/low risk clinical vs. low risk genetic/high risk clinical vs. low risk genetic/low risk clinical), chemotherapy (no chemotherapy, chemotherapy without R-C, R-C arm A, R-C arm B), Page 72, Baseline characteristics will include age, menopausal status, ER/PgR status, HER-2 status, tumor size, differentiation, WHO performance status, Page 75, Differentiation (grade I well differentiated, grade II moderately differentiated, grade III Amendment 2.0 ECG (the pre-surgical ECG can be used provided it was normal) Cardiac function To be performed within a clinically acceptable period before randomization R-E and before the start of endocrine therapy treatment. The primary test for R-C is to compare DFS between the two chemotherapy arms randomized. R-T: Patients for whom both methods are stratifying for institution, risk group (high risk genomic/low risk clinical vs. low risk genomic/high risk clinical), HR status (Positive (ER and/or PgR) vs. Negative (both)), nodal involvement (yes, no), age For R-C, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/low risk clinical vs. low risk genomic /high risk clinical vs. high risk genomic/high risk clinical), For R-E, a minimization technique will be used for random treatment allocation stratifying for institution, risk group (high risk genomic/high risk clinical vs. high risk genomic/low risk clinical vs. low risk genomic/high risk clinical vs. low risk genomic /low risk clinical), chemotherapy (no chemotherapy, chemotherapy without R-C, R-C arm A, R-C arm B), Baseline characteristics will include age, menopausal status, ER/PgR status, HER-2 status, nodal involvement, tumor size, differentiation, WHO performance status, Differentiation (grade I well differentiated, grade II moderately differentiated, grade III poorly differentiated, unknown) FO /44 Template version: November 2005

342 Current protocol 10041, 1.1 & poorly differentiated, unknown) HER-2 status (positive, negative, unknown) Page 77, The patient population recruited upfront for the trial is not biased : clinicians are entering in the trial a reasonable spectrum of node-negative disease and not just Page 79, optional blood sample (for serum storage) for proteomics analysis and biological materials bank storage to be taken between enrollment and the start of treatment Page 80, 12.3 The initial proteomics studies will be performed at the proteomics facility in Swansea, UK. Therefore, at the end of accrual serum samples will be shipped from each participating centre (on dry ice) to the TRANSBIG biological materials bank in Brussels; from there one of the 0.5 ml serum aliquots will be dispatched to Southwest Wales Cancer Institute, Swansea UK for analysis. Page 81, 12.4 Briefly, one section will be stained with HE to determine the percentage tumor cells in the frozen biopsy sample. When the tumor cell percentage is sufficient (> 50%), RNA will be isolated from the sample. Page 81, 12.5 To establish an early breast cancer biological materials bank with fresh frozen tumor tissue samples, RNA, serum and tissue microarrays. Page 82, 12.5 The serum samples will be shipped to the TRANSBIG biological materials bank at the end of accrual and then an aliquot transferred to Swansea for proteomics analysis. Page 82, 13 All abstracts or manuscripts as well as detailed descriptions of any planned oral presentations must be submitted to the EORTC and to the TRANSBIG Steering Committee for approval Amendment 2.0 nodal involvement (yes, no) HER-2 status (positive, negative, unknown) The patient population recruited upfront for the trial is not biased : clinicians are entering in the trial a reasonable spectrum of node-negative and 1-3 positive node disease and not just 1 optional blood sample (for whole blood and serum storage) for genetic and proteomics analysis and biological materials bank storage for future research to be taken before surgery. The initial proteomics studies will be performed as agreed by the TRANSBIG and Steering Committees. Therefore, at the end of accrual (or sooner if required) serum samples will be shipped from each participating centre (on dry ice) to the TRANSBIG biological materials bank in Brussels. From there 0.5 ml aliquots will be transferred to the research laboratories for analysis. Briefly, one section will be stained with HE to determine the percentage tumor cells in the frozen biopsy sample. When the tumor cell percentage is sufficient (> 30%), RNA will be isolated from the sample. To establish an early breast cancer biological materials bank with fresh frozen tumor tissue samples, RNA, whole blood, serum and tissue microarrays. The whole blood and serum samples will be shipped to the TRANSBIG biological materials bank at the end of accrual, or sooner if required. All abstracts, manuscripts as well as detailed descriptions of any planned oral presentations must be submitted to the EORTC and to the and TRANSBIG Steering Committees for approval (via FO /44 Template version: November 2005

343 Current protocol 10041, 1.1 & and at the earliest practicable time, but at least within four weeks prior to any proposed submission for publication of any manuscript or any presentation or other public disclosure. Abstracts must be sent to the EORTC and TRANSBIG Steering Committee at the earliest practicable time, but at least within ten days before submission. Page 83, 14 A signed initial Material Transfer Agreement (MTA) to be sent to TRANSBIG. (Confirm MTA online at the end of accrual listing all sample SeqIDs) Page 84, 15 register patient for screening on web page and receive patient identification number (SeqID) breast cancer surgery, freeze tumor biopsy & Page 84, All trial authorized investigators can enroll/randomize patients directly on the website, 24 hours a day, 7 days a week ( Page 84, 15.2 a paper enrollment checklist (detailing the eligibility criteria) will be provided and should be filled in before connecting to the website to enroll a patient Page 85, 15.2 (results of clinical and genomic prognostic tests) Inform patient of proposed treatment Page 85, 15.2 Amendment and transbig@bordet.be) at the earliest practicable time. For publication of any manuscript or any presentation or other public disclosure must be at least within four weeks prior to any proposed submission. Abstracts must be sent at least within ten days before submission. Research projects must follow the application policy approved by the and TRANSBIG Steering Committees, information available via the same addresses. A signed initial Acknowledgement of Transfer of biological Materials (ATM) to be sent to TRANSBIG. (Confirm ATM online at the end of accrual listing all sample SeqIDs) register patient for screening on web page and receive patient identification number (SeqID) take blood sample for future research (optional for patient) breast cancer surgery, freeze tumor biopsy & All trial authorized investigators can enroll/randomize patients directly on the website, 24 hours a day, 7 days a week ( a paper enrollment checklist (detailing the eligibility criteria) will be provided and can be filled in before connecting to the website to enroll a patient (results of clinical and genomic prognostic tests) either print the completed enrollment checklist screen print from the web based platform or complete a paper checklist, attach the sample sticker and sign it. Send it to your NCC/EORTC within 3 weeks of enrolling the patient. Inform patient of proposed treatment Sign and date subsequent checklist(s)/ FO /44 Template version: November 2005

344 Current protocol 10041, 1.1 & Sign and date the enrollment checklist and send to NCC/G Page 85, hospital patient's chart number (if available) Note this item is optional. Page 85, The patient s age, ER status, tumor grade and tumor size will be passed on to Adjuvant! Online by the web based platform. Comorbidity will be defaulted to Minor problems and positive nodes to 0. If Adjuvant! Online is upgraded during the trial the new version will be used. If a new variable is added (for example HER-2 status) the new variable will be incorporated into the clinical prognosis. Page 86, The risk assessment results and allocated prognostic tool, where applicable, must be recorded on the enrollment checklist which will be signed and dated by the Principal Investigator and sent to the NCC/G. Page 86, anthracycline based chemotherapy 2. docetaxel-capecitabine chemotherapy This completes the chemotherapy randomization. The allocated treatment must be recorded on the randomization checklist which will be signed and dated by the Principal Investigator and sent to the NCC/G. Page 86, If a patient who should be proposed chemotherapy does not wish to participate in the chemotherapy randomization (R-C), she should receive the hospital s standard chemotherapy. Page 87, The allocated treatment must be recorded on the randomization checklist which will be signed Amendment 2.0 screenprint(s) and send to your NCC/G or EORTC The patient s age, ER status, tumor grade, tumor size and nodal status will be passed on to Adjuvant! Online by the web based platform. Comorbidity will be defaulted to Minor problems. If Adjuvant! Online is upgraded during the trial the new version will be used. If a new variable is added the new variable will be incorporated into the clinical prognosis. The enrollment checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks of enrollment. 1. anthracycline based chemotherapy (FEC D for N+) 2. docetaxel-capecitabine chemotherapy This completes the chemotherapy randomization. The randomization checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks. If a patient who should be proposed chemotherapy does not wish to participate in the chemotherapy randomization (R-C), she should receive the hospital s standard chemotherapy. You will still need to complete the first 2 questions of the chemotherapy checklist and then say which non-randomized chemotherapy the patient will receive. The randomization checklist/screenprint will be signed and dated by the Principal Investigator FO /44 Template version: November 2005

345 Current protocol 10041, 1.1 & and dated by the Principal Investigator and sent to the NCC/G. Page 87, If a patient with an endocrine responsive tumor does not wish to participate in the endocrine therapy randomization (R-E), she should receive the hospital s standard endocrine therapy. Page 88, 16.1 A a signed and dated hardcopy/paper copy of the enrollment check list, must be sent either to your NCC/G or directly to the EORTC Data Center (if the EORTC is your NCC/G). Page 88, 1.6.1A Further randomization checklist(s) (chemotherapy R-C & endocrine therapy R-E as applicable), must also be sent either Page 90, All SAEs that are simply signs and symptoms of the disease being studied do NOT need to be collected! Page 91, The product reference documents are For marketed products: Summary Of Product Characteristics which can be found on m# For non-marketed products: Current version of the Investigators Brochure For the causality assessment, the following definitions Amendment 2.0 and sent to the NCC/G or EORTC within 3 weeks. If a patient with an endocrine responsive tumor does not wish to participate in the endocrine therapy randomization (R-E), she should receive the hospital s standard endocrine therapy. You will still need to complete the first 2 questions of the endocrine therapy checklist and then say which non-randomized endocrine therapy the patient will receive. a signed and dated screenprint/paper copy of the enrollment check list, must be sent either to your NCC/G or directly to the EORTC Data Center (if the EORTC is your NCC/G) within 3 weeks of enrollment. Further randomization checklist(s)/ screenprint(s) (chemotherapy R-C & endocrine therapy R-E as applicable), must also be sent either All SAEs that are simply signs and symptoms of the disease being studied do NOT need to be collected. All SAEs that are related to standard primary breast cancer surgery and adjuvant radiotherapy do NOT need to be reported. The product reference documents are For the experimental arm drugs (Docetaxel, Capecitabine and Letrozole) the Investigator Brochures will be the reference documents. For the standard arm drugs the SmPCs listed below will be the reference documents. 5-FU: SmPC of Fluorouracil Injection, 50 mg/ml (Medac) cyclophosphamide : SmPC of Endoxan 500mg Injection (Baxter Healthcare Limited) doxorubicin: SmPC of Doxorubicin Rapid Dissolution_(Pharmacia Limited) epirubicin: SmPC of Pharmorubicin (Pharmacia Limited) methotrexate: SmPC of Methotrexate 2.5mg/ml Injection (Mayne_Pharma) tamoxifen : SmPC of Nolvadex (AstraZeneca) For the causality assessment, the following definitions FO /44 Template version: November 2005

346 Current protocol 10041, 1.1 & Page 91, It should be recognized that Serious Adverse Reactions (SAR) which have not been previously documented in the Investigators Brochure, or which occur in a more severe form than anticipated (i.e. they are unexpected by nature or severity), are subject to rapid reporting to the Regulatory Authorities. Page 92, 18.2 add subsection 18.2 Audits Amendment 2.0 It should be recognized that Serious Adverse Reactions (SAR) which have not been previously documented in the applicable Reference Document, or which occur in a more severe form than anticipated (i.e. they are unexpected by nature or severity), are subject to rapid reporting to the Regulatory Authorities Quality Assurance and Quality Control Program An integrated Quality Assurance and Quality Control (QA & QC) Program that encompasses the usual EORTC extensive central monitoring checks, data quality checks and data timeliness and a quality check visit for selected sites will be carried out in collaboration between the EORTC and the NCC/Gs. This will insure that we can demonstrate a high level of quality throughout the trial. The QA & QC Program also details the audit plan for the study. Details of the Program and responsibilities can be found in the QA & QC Program and Intergroup agreements. Practically it will mean one 1 day visit for selected centers, with a potential additional visit for high recruiting centers, performed either by the EORTC and/or the NCC/G. The aim of these visits is to identify any areas where sites may require additional help or explanation and to demonstrate compliance to the protocol. Page 92, 18.2 This procedure does not apply to non-eortc investigators who should contact their NCC/G for the information on eventual audits performed by their group. Page 92, 19.1 The responsible investigator will ensure that this 18.3 Audits Whether other groups center's will participate in the audit part of the QA & QC Program or in their own Group's audit programs will be agreed between the Collaborative Group and the EORTC. For details please contact your Group The responsible investigator will ensure that this study is conducted in agreement with either the FO /44 Template version: November 2005

347 Current protocol 10041, 1.1 & study is conducted in agreement with either the Declaration of Helsinki (Tokyo, Page 93, 19.2 In order to avoid identification errors, patient s code (maximum of 4 letters), date of birth and local chart number (if available) will also be reported on the case report forms. Page 94, 20.1 The centre of the tumor is often necrotic, while the edge of the tumor often contains many stromal cells, which in both cases will lead to percentage of tumor cells less than 50% and therefore not enough to isolate tumor RNA from. Page 94, 20.1 Tumor biopsy cellularity will be determined by Agendia/NKI along with other quality checks. Page 94, 20.1 The frozen biopsy will be used for microarray analysis only when a patient is confirmed to be lymph node-negative. Page 94, 20.2 Giuseppe.Viale@ieo.it Page 94, 20.3 Serum sample schedule For those patients who consent to the optional blood sampling for serum storage for proteomics analysis, blood should be taken after the patient has signed the PIS & IC 1 and before the start of any protocol treatment. Serum should be separated and frozen according to SOP 15 and stored at - 80 C until the end of accrual when it should be sent to the TRANSBIG biological materials bank in Belgium. Amendment 2.0 Declaration of Helsinki, see appendix C (Tokyo, In order to avoid identification errors, patient s code (maximum of 4 letters or numbers) and date of birth will also be reported on the case report forms. The centre of the tumor is often necrotic, while the edge of the tumor often contains many stromal cells, which in both cases will lead to percentage of tumor cells less than 30% and therefore not enough to isolate tumor RNA from. Tumor biopsy cellularity will be determined by Agendia along with other quality checks. The frozen biopsy will be used for microarray analysis only once local lymph node status is confirmed and the patient is perceived likely to 20.3 Blood and Serum sample schedule For those patients who consent to the optional blood sampling for whole blood and serum storage for proteomics analysis, blood should be taken after the patient has signed the screening PIS & IC and before surgery, preferably during a routine blood sampling. For whole blood, the sample should be taken into an EDTA blood tube. The whole blood should be aliquoted in 0.5 ml aliquots into cryovials prelabeled with coded stickers according to the instructions provided in SOP MA 15. For serum, the sample should be taken into a serum blood tube with a gel separation system. Serum should be separated and frozen according to SOP 15 and stored at - 80 C until the end of accrual when it should be sent to the TRANSBIG biological materials bank in Belgium. FO /44 Template version: November 2005

348 Current protocol 10041, 1.1 & Page 95, Figure (see attached) Page 110, Appendix C cut appendix calculation of the glomerular filtration rate Page 112, Appendix E (spis & IC) gives consent for the donation of a piece of tumor tissue and for its being sent to a specialized cancer center and for the microarray analysis to be performed. their center s standard PIS & IC if they are confident that it covers the sending of tissue and the microarray analysis. and does not state that an EORTC clinical trials insurance is in place for the screening period Page 112, Appendix E There will be a maximum of 3 PIS & ICs for this study. The first one, the PIS & IC 1, will be signed by all eligible patients Page 113, Appendix E Patients with aggressive tumors need more than local treatment and are given chemotherapy or hormonal therapy Page 113, spis & IC The purpose of the research study this hospital is involved in is to find out if this new test is really better than traditional pathological examinations of tumor tissue for those patients who do not have cancer cells in the lymph nodes (called lymph node-negative breast cancer). Page 113, spis & IC To have the gene expression test done, we would need to send a piece of tumor tissue to the Netherlands Page 114, spis & IC 1. If there are no tumor cells found in your axillary lymph nodes you will be offered the Amendment 2.0 Appendix C Declaration of Helsinki (spis & IC) gives consent for the donation of a piece of tumor tissue and for its being sent to a specialized cancer center and for the microarray analysis to be performed. Also for the donation of a blood sample. their center s standard PIS & IC if they are confident that it covers the sending of tissue, the microarray analysis and the donation of a blood sample. donation of a blood sample There will be a maximum of 4 PIS & ICs for this study. All screened patients will sign the spis & IC. The PIS & IC 1 will be signed by all eligible patients Patients with aggressive tumors need more than local treatment and are given chemotherapy and/or hormonal therapy The purpose of the research study this hospital is involved in is to find out if this new test is really better than traditional pathological examinations of tumor tissue. The new test has been developed for those patients who do not have cancer cells in the lymph nodes (called lymph node-negative breast cancer) and later for those patients with tumor cells found in 1 to 3 lymph nodes. To have the gene expression test done, we would need to send a piece of your left over tumor tissue to the Netherlands 1. If there are no tumor cells found in your axillary lymph nodes or if there are tumor cells in only 1 to 3 lymph nodes, you will be offered FO /44 Template version: November 2005

349 Current protocol 10041, 1.1 & opportunity to participate in the research study mentioned above. Page 114, spis & IC b) you may decide not to participate in this research study in which case you may ask for the piece of your tumor tissue that was sent to the Netherlands to be destroyed. Page 114, spis & IC 2. If tumor cells have been identified in one or more of your axillary lymph nodes the research study is not appropriate to your situation. Page 114, spis & IC a) you may prefer that the piece of your tumor tissue that was sent to the Netherlands is destroyed. Page 114, spis & IC add section before What are my rights? Amendment 2.0 the opportunity to participate in the research study mentioned above. b) you may decide not to participate in this research study in which case you may ask for the piece of your tumor tissue that was sent to the Netherlands to be destroyed or returned. 2. If tumor cells have been identified in more than 3 of your axillary lymph nodes the research study is not appropriate to your situation. a) you may prefer that the piece of your tumor tissue that was sent to the Netherlands is destroyed or returned. Donation of a blood sample for future research As well as deciding whether you would like to donate any remaining tumor tissue after the performance of the prognostic test for future research, we would like to ask you to donate a 20 ml (2 table spoons) blood sample. This blood sample would be taken before the surgery to remove your breast cancer. It will be stored and used for future research projects to increase our understanding of cancer with a view to improving patient care. The blood sample will be used for both serum storage for proteomics analysis (looking at proteins) and whole blood storage for potential genetic research (looking at how your genes may have information about cancer risk). In the future, we hope that proteomics will help us predict if or when tumors will come back. Because proteomics is still a new type of research, your blood sample will not immediately be useful for seeing when and if your cancer may come back. However, it will help us to further develop this new technology, which will help other breast cancer patients in the future. Genetic research is also developing rapidly so FO /44 Template version: November 2005

350 Current protocol 10041, 1.1 & Page 114, spis & IC What are my rights? Your privacy will be protected at all times and your tumor tissue will not be used to test whether your breast cancer is hereditary (that is, breast cancer runs in your family). Your tumor tissue will not be used for any other purpose unless you are contacted again with more information and a new request for your consent Page 114, spis & IC for analysis to the Netherlands. Regardless of whether you decide to have a piece of your tumor sent to the Netherlands or not, not, your relationship with your physician will in no way be affected. Amendment 2.0 we don t yet know the kind of research projects that may be possible. Your blood sample will be stored without personal information so that researchers will not know who you are and will be analyzed with many other samples to increase our understanding of cancer. The biological samples (blood and tumor), will be stored in a biological materials bank. The biological materials bank will be under the guardianship of an international, academic, non-profit organization. Please be informed that all future research projects must be approved by the relevant research and ethics committees before the samples can be used. If you consent to have a piece of your tumour tissue sent to the Netherlands, your privacy will be protected at all times. In agreeing to the genomic prognostic test your tumor tissue will not be used to test whether your breast cancer is hereditary (that is, breast cancer runs in your family). If you agree to an additional blood sample for research or to donate the remainder of your tumor tissue for future research, this may involve genetic research.. If you agree to donate a blood sample collected for future research, this will probably be taken at the same time as a routine blood samples, in this case you will not need an additional needle prick/puncture. Regardless of whether you decide to have a piece of your left over tumor sent to the Netherlands or not, not, your relationship with your physician will in no way be affected. Consent is voluntary and can be withdrawn at anytime without you having to give a reason and without having any affect on your relationship with your treating physician and clinical team. If new information becomes available about the new test, you will be FO /44 Template version: November 2005

351 Current protocol 10041, 1.1 & Amendment 2.0 Page 116, spis & IC I accept that a part of my tumor tissue will be sent to the Netherlands, where it will be analyzed using the new genomic prognostic test. If tumor cells are found in my axillary lymph nodes and the new genomic test is not appropriate for me, I would like to donate the piece of my tumor tissue to research studies not defined at the present time. Page 116, spis & IC Page 118, PIS & IC 1 Now we would like to invite you to participate in the clinical trial (a research study) called (Microarray In Node-negative Disease may Avoid ChemoTherapy). This study Page 118, informed of this by your treating physician. I accept that a part of my left over tumor tissue will be sent to the Netherlands, where it will be analyzed using the new genomic prognostic test. I would like to donate any remaining tumor tissue after the performance of the new test for future research. If the new genomic test is not appropriate for me or I decide not to participate in the proposed study, I would still like to donate the piece of my tumor tissue to research studies not defined at the present time. I would like to donate a blood sample for future research. If the new genomic test is not appropriate for me or I decide not to participate in the proposed study, I prefer that my tumor tissue that has been send to the Netherlands will be destroyed or returned Now we would like to invite you to participate in the clinical trial (a research study) called (Microarray In node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy). This study (* details are provided in section 15) (* details are provided in section 14) Page 120, PIS & IC 1 Diagram 1 Node-negative breast cancer Node-negative breast cancer & 1-3 positive lymph nodes breast cancer Page 122, 9. Research on biological material - objectives and description You have already donated a piece of your tumor and consented for the new genomic prognostic test to be performed. In addition to this tumor donation, if you participate in this study, you will be asked to donate an optional blood sample before the start of treatment and perhaps another optional blood sample at the end of chemotherapy. These samples will be stored in what we call a biological materials bank. These samples will be used for measuring the types and amounts of proteins in your blood. FO /44 Template version: November 2005

352 Current protocol 10041, 1.1 & Amendment 2.0 This type of research is called proteomics. In the future, we hope that proteomics will help us predict if or when tumors will come back. Because proteomics is still a new type of research, your blood sample will not immediately be useful for seeing when and if your cancer may come back. However, it will help us to further develop this new technology, which will help other breast cancer patients in the future. The blood sample donation is optional; if you decide to donate 1 or maybe 2, 10 ml (1 tablespoon) blood samples, they will be stored anonymously in a biological materials bank. The biological materials bank will be under the guardianship of the international, academic, nonprofit organization the Breast International Group (BIG*). (*see section 15 for details) Blood samples from the biological materials bank will be made available to scientific researchers in the future who want to improve the understanding and treatment of breast cancer. However, any researchers who want to use the blood samples for research will first have to have their project approved by a committee of academic researchers representing the groups participating in. All research projects that use samples from the biological materials bank will also have to be approved by local and national ethics committees. Your blood sample will not be used for any genetic tests (for example, hereditary tests to see if cancer runs in your family), nor to do research on any disease other than breast cancer. Page 122, 10. Voluntary participation and renumbering of other section titles Page 123, Only authorized persons (EORTC research staff, national and/or foreign health authority representatives) may have Page 124, 14 Responsabilities coordinated and analyzed by the European 9. Voluntary participation Only authorized persons (EORTC research staff and their representatives, national and/or foreign health authority representatives) may have coordinated and analyzed by the European FO /44 Template version: November 2005

353 Current protocol 10041, 1.1 & Organization for Research and Treatment of Cancer (EORTC) Page 125, PIS & IC 1 I agree to participate in the clinical trial called ( Microarray in up Nodenegative Disease may Avoid Chemo Therapy. A prospective, randomized study comparing the 70-gene signature with common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer), EORTC study number and BIG study number BIG Page 125, PIS & IC 1 I agree to donate 1 or possibly 2, 10 ml sample(s) of my blood (1 tablespoon) for research purposes only. Page 125, PIS & IC 1 In case of the EORTC or the BIG / TRANSBIG research consortium agreeing to a collaboration with a third party (not already foreseen in the protocol), I agree that my biological material can be used by: Another academic institution/organization. A commercial company (i.e. one of the pharmaceutical companies supporting ) Page 128, PIS & IC 2 As part of the - Microarray In Nodenegative Disease may Avoid ChemoTherapy trial (A prospective, randomised study comparing the 70-gene signature with common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer) Can anthracycline-based chemotherapy be safely replaced by the combination of Docetaxel- Capecitabine as adjuvant treatment of nodenegative breast cancer? Amendment 2.0 Organisation for Research and Treatment of Cancer (EORTC) I agree to participate in the clinical trial called ( Microarray In Nodenegative and 1 to 3 positive lymph node Disease may Avoid Chemo Therapy. A prospective, randomized study comparing the 70-gene signature with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0 to 3 positive nodes), EORTC study number and BIG study number BIG I agree that my biological material can be used in the case of the EORTC or the BIG / TRANSBIG research consortium agreeing to a collaboration with a third party (not already foreseen in the protocol). As part of the - Microarray In Nodenegative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy trial (A prospective, randomized study comparing the 70-gene signature with common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0-3 positive nodes) Can anthracycline-based chemotherapy be safely replaced by the combination of Docetaxel- Capecitabine as adjuvant treatment of early breast cancer? FO /44 Template version: November 2005

354 Current protocol 10041, 1.1 & Page 129, PIS & IC 2 Treatment Selection Your doctor will call a central statistical office, which will assign one of the two treatments to you. Page 129, Chemotherapy B The four drugs that can be used in different Page 129 is 3 or 4 weeks) of this chemotherapy regimen. If you participate in this study Page 129, Results of the study will not be available until after the study is finished (recruitment of patient is expected to finish in 2009 and the study to have initial results 2012), but Page 131 Chemotherapy B General Side-effects with the drugs combination Page 132, doctor following the standard of care applicable in the hospital. Amendment 2.0 Your doctor will contact a central statistical office, which will assign one of the two treatments to you. Chemotherapy B Depending whether your breast cancer was lymph node-negative or -positive, the chemotherapy options are different. As it is considered that patients needing chemotherapy may be at a higher risk when they are node-positive, the chemotherapy for these patients is more aggressive. The chemotherapy options in this arm are well know and are the standard treatment for patients in your situation. Chemotherapy B for node-negative patients The four drugs that can be used in different is 3 or 4 weeks) of this chemotherapy regimen. Chemotherapy B for node-positive patients. The option here is a chemotherapy (called FEC D) consisting of 3 cycles of a combination of fluorouracil, cyclophosphamide and epirubicin, followed by 3 cycles of docetaxel. This chemotherapy is given on one day every 3 weeks, up to a total of 6 cycles. If you participate in this study Results of the study will not be available until after the study is finished (recruitment of patient is expected to finish in 2010 and the study to have initial results 2013), but Chemotherapy B Chemotherapy B for node-negative patients General Side-effects with the drugs combination doctor following the standard of care applicable in the hospital. Chemotherapy B for node-positive patients. FO /44 Template version: November 2005

355 Current protocol 10041, 1.1 & Page 132, 8 Data Protection Duly authorized persons (EORTC research staff, national and/or foreign health authority representatives) may have access Page 134, PIS & IC 2 I agree to participate in the clinical research study entitled - Microarray In Node-negative Disease may Avoid ChemoTherapy in the chemotherapy therapy question titled Can anthracyclinebased chemotherapy be safely replaced by the combination of Docetaxel- Capecitabine as adjuvant treatment of node-negative breast cancer? Page 136, PIS & IC 3 As part of the - Microarray In Nodenegative Disease may Avoid ChemoTherapy trial (A prospective, randomised study comparing the 70-gene signature with common clinical pathological criteria in selecting patients for adjuvant chemotherapy in node-negative breast cancer). Page 137, PIS & IC 3 Treatment Selection Your doctor will call a central statistical office, which will assign one of the two treatments to you. Page 138, 5. Description of foreseeable risks and discomforts Less frequent side effects affecting less than 1% of patients are: tumor pain, leucopenia, hypercholestrolaemia (high blood cholesterol), Page 140, 13. Contact person If you consent to join this trial, you will be given a telephone number of the hospital that you can contact at any time if you feel unwell or have further questions. With your agreement, your family doctor (if applicable) will also be Amendment 2.0 For the first 3 cycles of FEC, the side effects are described above. For the following 3 cycles of docetaxel, please refer to the section describing Docetaxel toxicity in Chemotherapy A. Duly authorized persons (EORTC research staff and their representatives, national and/or foreign health authority representatives) may have access I agree to participate in the clinical research study entitled Microarray In Node-negative and 1-3 positive lymph node Disease may Avoid ChemoTherapy in the chemotherapy therapy question titled Can anthracycline-based chemotherapy be safely replaced by the combination of Docetaxel- Capecitabine as adjuvant treatment of breast cancer with 0-3 positive nodes? As part of the - Microarray In Nodenegative and 1-3 positive lymph node Disease may Avoid ChemoTherapy trial (A prospective, randomized study comparing the 70-gene signature with common clinical pathological criteria in selecting patients for adjuvant chemotherapy in breast cancer) with 0-3 positive nodes. Your doctor will contact a central statistical office, which will assign one of the two treatments to you. Less frequent side effects affecting less than 1% of patients are: leucopenia, hypercholestrolaemia (high blood cholesterol), With your agreement, your family doctor (if applicable) will also be informed about your taking part in this trial and what is involved. FO /44 Template version: November 2005

356 Current protocol 10041, 1.1 & informed about your taking part in this trial and what is involved, if you agree. Page 141, PIS & IC 3 I agree to participate in the clinical research study entitled Microarray In Node-negative Disease may Avoid ChemoTherapy in the endocrine therapy question titled: Amendment 2.0 I agree to participate in the clinical research study entitled Microarray In Node-negative and 1-3 positive lymph node Disease may Avoid ChemoTherapy in the endocrine therapy question titled: provide the original text in the left column; provide modified text (in bold) in the right column 2. Other non-substantial modifications to the protocol (i.e. administrative ) Current protocol 10041, 1.1 & Page 1 In Title (Microarray In Node-negative Disease may Avoid chemotherapy): A prospective, randomized study comparing the 70-gene signature with the common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in node negative breast cancer Collaborative Groups: remove ICORG Amendment 2.0 (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid chemotherapy): A prospective, randomized study comparing the 70-gene signature with the common clinical-pathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0 to 3 positive nodes add CEEOG & NCRI Breast Group The EORTC logo will be updated for all instances for which it is used. Page 2, Writing Committee: J. Bogaerts, EORTC Data Center, Brussels, Belgium J. Harrison, EORTC Data Center, Brussels, Belgium P. Therasse, EORTC Data Center, Brussels, Belgium Writing Committee: J. Bogaerts, EORTC Headquarters, Brussels, Belgium J. Harrison, EORTC Headquarters, Brussels, Belgium P. Therasse, EORTC Headquarters, Brussels, Belgium FO /44 Template version: November 2005

357 Current protocol 10041, 1.1 & Clinical fellows: G. Demonty, EORTC Data Center, Brussels, Belgium Page 2 Contacts add & remove collaborative groups Remove ICORG Irish Clinical Oncology Research Group Page 8 appendix C cut Calculation of Glomerular filtration rate and replace it with the Declaration of Helsinki Page 15, Trial Organization Participation in this trial is only possible through a national structure (coordinating group/center, NCC/G or EORTC Data Center) Please contact the EORTC Data Center directly at Page 58, Table 10 Moderate symptoms: any symptom not listed above (mildsymptoms) or Page 68, Investigators who do not have access to Internet can contact the EORTC Data Center to receive a copy by post. Page 82, 13 The final publication of the trial results will be written by the Study Coordinators from the EORTC Breast Cancer Group on the basis of the final analysis performed at the EORTC Data Center and then approved by the EORTC Data Center. After revision by the EORTC Data Center and other co-authors (and the industrial partners) the manuscript will be sent to a major scientific journal. The EORTC Group Chairman, Study Coordinators, EORTC Data Center Team, the Amendment 2.0 Clinical fellows: G. Demonty, EORTC Headquarters, Brussels, Belgium Add NCRI Breast Group Add CEEOG Central and Eastern European Oncology Group Table of Appendices & as appendix (pages X) Add Declaration of Helsinki Participation in this trial is only possible through a national structure (coordinating group/center, NCC/G or EORTC Headquarters) Please contact the EORTC Headquarters directly at Moderate symptoms: any symptom not listed above (mild symptoms) or Investigators who do not have access to Internet can contact the EORTC Headquarters to receive a copy by post. The final publication of the trial results will be written by the Study Coordinators from the EORTC Breast Cancer Group on the basis of the final analysis performed at the EORTC Headquarters and then approved by the EORTC Headquarters. After revision by the EORTC Headquarters and other co-authors (and the industrial partners) the manuscript will be sent to a major scientific journal. The EORTC Group Chairman, Study Coordinators, EORTC Headquarters Team, the FO /44 Template version: November 2005

358 Current protocol 10041, 1.1 & TRANSBIG Steering Committee or any Page 84, 14 All the above mentioned documents are available at their NCC/G or EORTC Data Center Page 86, The Principal Investigator, NCC/G where applicable and the EORTC Data Center will receive an Page 86, The Principal Investigator, NCC/G if applicable and the EORTC Data Center will receive Page 87, The Principal Investigator, NCC/G if applicable and the EORTC Data Center will receive Page 87, 16 All RDC forms will be specifically designed by the EORTC Data Center for this study. Page 87, 16.1 and then signed and returned to their NCC/G or directly to the EORTC data center in countries where the EORTC is the NCC/G. Forms to be sent to the EORTC Data Center directly should be sent to Breast cancer group Data Manager EORTC Data Center Page 88, 16.2 The list of staff members authorized to enter forms must be identified on the signature log and sent to the Data Center by the responsible Page 89, 16.2 On the data received, the EORTC Data Center will perform extensive the original must be sent by regular mail to your NCC/G or to the EORTC Data Center. needs to modify a CRF after the form has been sent to the EORTC Data Center, he/she should Amendment 2.0 TRANSBIG Steering Committee or any All the above mentioned documents are available at their NCC/G or EORTC Headquarters The Principal Investigator, NCC/G where applicable and the EORTC Headquarters will receive an The Principal Investigator, NCC/G if applicable and the EORTC Headquarters will receive The Principal Investigator, NCC/G if applicable and the EORTC Headquarters will receive All RDC forms will be specifically designed by the EORTC Headquarters for this study. and then signed and returned to their NCC/G or directly to the EORTC Headquarters in countries where the EORTC is the NCC/G. Forms to be sent to the EORTC Headquarters directly should be sent to Breast cancer group Data Manager EORTC Headquarters The list of staff members authorized to enter forms must be identified on the signature log and sent to the EORTC Headquarters by the responsible On the data received, the EORTC Headquarters will perform extensive the original must be sent by regular mail to your NCC/G or to the EORTC Headquarters. needs to modify a CRF after the form has been sent to the EORTC Headquarters, he/she FO /44 Template version: November 2005

359 Current protocol 10041, 1.1 & use a Data Correction Sheet and sent it to their NCC/G or the EORTC Data Center. Page 92, 18.1 Data forms will be entered in the database of the EORTC Data Center using the RDC system. Page 93, 19.2 The name of the patient will not be asked for nor recorded at the Data Center. Page 93, 19.3 Documented informed consent must be obtained for all patients included in the study before they are enrolled or randomized at the EORTC Data Center. Page 94, 20.1 Immediately after taken the punch biopsy sample place it in the labeled tube provided, firmly close the tube and submerge the tube in liquid nitrogen. Page 95, 21.1 The Study Coordinator (in cooperation with the Data Center) will be responsible Page 96, 21.2 The EORTC Data Center The EORTC Data Center will be responsible for reviewing All methodological questions should be addressed to the EORTC Data Center. EORTC DATA CENTER 83, avenue Emmanuel Mounier, Bte 11 Page 96, 21.2 Registration of patients: Page 97, 21.2 Data Manager: Inge Delmotte Phone: Fax: inge.delmotte@eortc.be Page 97, 21.2 Project Managers: Frédéric Hénot Amendment 2.0 should use a Data Correction Sheet and sent it to their NCC/G or the EORTC Headquarters. Data forms will be entered in the database of the EORTC Headquarters using the RDC system. The name of the patient will not be asked for nor recorded at the EORTC Headquarters. Documented informed consent must be obtained for all patients included in the study before they are enrolled or randomized at the EORTC Headquarters. Immediately after taking the punch biopsy sample place it in the labeled tube provided, firmly close the tube and submerge the tube in liquid nitrogen. The Study Coordinator (in cooperation with the EORTC Headquarters) will be responsible 21.2 The EORTC Headquarters The EORTC Headquarters will be responsible for reviewing All methodological questions should be addressed to the EORTC Headquarters. EORTC HEADQUARTERS 83, avenue Emmanuel Mounier, Bte 11 Registration of patients: Data Manager: Kaat Vansevenant Phone: Fax: kaat.vansevenant@eortc.be Project Manager: Jillian Harrison FO /44 Template version: November 2005

360 Current protocol 10041, 1.1 & Phone: Fax: frederic.henot@eortc.be Jillian Harrison Phone: Fax: jillian.harrison@eortc.be Coordinating Physician: Dr. Patrick Therasse Phone: Fax: patrick.therasse@eortc.be Page 97, 21.2 F. Hoffmann La Roche Cristella LaRosa Clinical Trial Manager-Consultant Pharma Business, Hoffman-LaRoche Inc. 340 Kingsland Street Nutley, NJ Tel: Fax: Page 98, 21.4 EORTC Breast Cancer Group Chairman: Dr. Emiel Rutgers Phone: Fax: E.Rutgers@nki.nl Secretary: Dr. Hervé Bonnefoi Phone: Fax: // herve.bonnefoi@hcuge.ch Page 98, 21.5 Belgium EORTC Data Center 83, avenue Emmanuel Mounier, Bte 11, B-1200 Brussels, Belgium Tel: /1655 Fax: Page 99, 21.5 Amendment 2.0 Phone: Fax: jillian.harrison@eortc.be Coordinating Physician: Dr. Gaston Demonty Phone: Fax: gaston.demonty@eortc.be Séverine Wollenschneider Ph.D. Clinical Trial Manager XELODA F. Hoffmann-La Roche Ltd Grenzacherstrasse 124 Pharmaceuticals Division Pharma Business Medical Bldg. 074/4W.219 CH-4070 Basel Phone: Fax: severine.wollenschneider@roche.com EORTC Breast Cancer Group Chairman: Dr. Hervé Bonnefoi Tel.: Fax: Bonnefoi@bergonie.org Secretary: Dr David Cameron Tel.: Fax: d.cameron@ncrn.org.uk Belgium EORTC Headquarters 83, avenue Emmanuel Mounier, Bte 11, B-1200 Brussels, Belgium Tel: /1061 Fax: Pharmacovigilance and local trial management FO /44 Template version: November 2005

361 Current protocol 10041, 1.1 & Pharmacovigilance & mailbox functions Fédération Nationale des Centres de Lutte Contre le Cancer (FNCLCC) 101, rue de Tolbiac, PARIS cedex 13, FRANCE Tel : Fax : al-martin@fnclcc.fr Page 99, 21.5 West German Study Group (WSG) gynaecological hospital - university clinics Duesseldorf, Mooresnstraße 5, Duesseldorf , Germany Tel / Fax / nitzu@uni-duesseldorf.de Page 99, 21.5 Ireland Irish Clinical Oncology Research Group (ICORG) 120 Pembroke Road, Ballsbridge, Dublin 4, Ireland Tel: Fax: brian.moulton@icorg.ie Page 99, 21.5 Serbia Montenegro EORTC Data Center Page 100, 21.5 SOLTI SANT LLORENÇ, 23-1º, 08202, Spain Tel: ; Fax: josep.vazquez@gruposolti.org Page 98, 21.5 The Netherlands Cancer Institute-Antoni van Leeuwenhoekziekenhuis Plesmanlann 121, NL 1066 CX Amsterdam, The Netherlands. Tel Fax E.Rutgers@nki.nl Pharmacovigilance & mailbox functions Amendment 2.0 Fédération Nationale des Centres de Lutte Contre le Cancer (FNCLCC) 101, rue de Tolbiac, PARIS cedex 13, FRANCE Tel : Fax : al-martin@fnclcc.fr /@fnclcc.fr West German Study Group (WSG) ggmbh Ludwig-Weber-Strasse Mönchengladbach Tel / Fax / corinna.buehne@wsg-online.com Serbia Montenegro EORTC Headquarters SOLTI SANT LLORENÇ, 23 - bajos, Barcelona 08202, Spain Tel: ; Fax: josep.vazquez@gruposolti.org The Netherlands Cancer Institute-Antoni van Leeuwenhoekziekenhuis Plesmanlann 121, NL 1066 CX Amsterdam, The Netherlands. Tel Fax E.Rutgers@nki.nl FO /44 Template version: November 2005

362 Current protocol 10041, 1.1 & University Hospital Maastricht (UM) Internal medicine, PO Box 5800, Maastricht, 6202 AZ, The Netherlands Tel Fax Page 98, 21.5 UK South West Wales Cancer Institute Singleton Hospital Sketty Lane Swansea SA2 8QA Tel: Fax: Page 100, 22 The Director General of the EORTC is: Professor Françoise Meunier EORTC Central Office Page 101, 23 Clinical trial insurance is only valid if the treatment is given in a center authorized by the EORTC Data Center and which has obtained Therefore all centers will have to declare to their NCC/G, who will inform the EORTC Data Center, of details Page 105, References Ref 55. Roché H, Fumoleau P Spielmann M et al. Five years analysis of the PACS 01 trial: 6 cycles of FEC100 vs 3 cycles of FEC100 followed by 3 cycles of docetaxel (D) for the adjuvant treatment of node positive breast cancer. Breast Cancer Res Treat 88, S Page 111, Appendix D The EORTC Data Center web site Amendment 2.0 ISD Cancer Clinical Trials Team (Partner in CaCTUS - Cancer Clinical Trials Unit Scotland) Open Plan Area 159C, 1st Floor Gyle Square 1 South Gyle Crescent Edinburgh, EH12 9EB Scotland Tel: Fax: kirsten.murray@isd.csa.scot.nhs.uk University of Dundee Ninewells Hospital and Medical School Prof A Thompson Tel Fax a.m.thompson@dundee.ac.uk The Director General of the EORTC is: Professor Françoise Meunier EORTC Clinical trial insurance is only valid if the treatment is given in a center authorized by the EORTC Headquarters and which has obtained Therefore all centers will have to declare to their NCC/G, who will inform the EORTC Headquarters, of details Ref 55. Roché H, Fumoleau P, Spielmann M et al. Sequential adjuvant epirubicin-based and docetaxel chemotherapy for node-positive breast cancer patients: the FNCLCC PACS 01 Trial. J Clin Oncol Dec 20;24(36): The EORTC Headquarters web site FO /44 Template version: November 2005

363 Current protocol 10041, 1.1 & provides Page 113, Appendix E Consequently, many scientists now think that by looking at the genes of tumor cells we will get Page 117, Appendix F send a copy of the approved translated document to the EORTC Data Center who will then Page 127, Appendix G send a copy of the approved translated document to the EORTC Data Center who will then Page 135, Appendix H send a copy of the approved translated document to the EORTC Data Center who will then Page 129, Chemotherapy B The drugs will be given intravenously in the day care unit usually once every 3 or 4 weeksweeks (it will take about 2 hours in the day care unit). Page 130 Especially anticoagulants, antacids, folic acid, leucovorin, phenytoin (Dilantin), asprin and vitamins. Page 143, Appendix J: Abbreviations Page 143, Appendix J: Abbreviations Microarray In Node-negative Disease may Avoid ChemoTherapy provides Amendment 2.0 Consequently, many scientists now think that by looking at the genes of tumor cells, we will get send a copy of the approved translated document to the EORTC Headquarters who will then send a copy of the approved translated document to the EORTC Headquarters who will then send a copy of the approved translated document to the EORTC Headquarters who will then The drugs will be given intravenously in the day care unit usually once every 3 or 4 weeks (it will take about 2 hours in the day care unit). Especially anticoagulants, antacids, folic acid, leucovorin, phenytoin (Dilantin), aspirin and vitamins. IRC Independent Review Committee Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy provide the original text in the left column; provide modified text (in bold) in the right column FP amend 1 - addendum.doc Table doc Table 11.doc Table doc timelines dia protocol P.ppt Chemo regimens dia doc Capecitabine pill combinations.doc FO /44 Template version: November 2005

364 EORTC protocol (Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy): A prospective, randomized study comparing the 70-gene signature with the common clinicalpathological criteria in selecting patients for adjuvant chemotherapy in breast cancer with 0 to 3 positive nodes. EORTC BIG 3-04 Intergroup Study (EudraCT number ) Amendment 2 Identification: Classification - Amendment to protocol document content: EORTC protocol number: Amendment number: Protocol document(s) affected: Full protocol PIS IC / Group Specific Appendix (GSA) Coordinating Full protocol Would you like to recompile the affected protocol document(s)? Yes No Scientific, i.e. significantly impacting on: the safety, physical or mental integrity of the patients; and/or the scientific value of the trial; and/or risk benefit (for individual patient or population at risk) and or may affect the patient s decision to participate (for PIS & IC) Administrative Does this amendment concern urgent safety measures already implemented? Yes No To be filled in by Protocol Development Unit (PDU): Date of submission to PRC: 27/05/2011 Recompiled (Y/N) Y New protocol version number* 3.0 * non recompiled amendments will have a hypothetical numbering but no recompiled document will be made e.g. for the first non recompiled administrative amendment 1.1 Internal PRC final decision on classification: Scientific Administrative Presented at the IPRM: 24/05/2011 EORTC approved by: PRC Chair Date: 31/05/2011 PD-004-AF-01 Page 1 of 46 Version: OCT 2009

365 PRC submission document Amendment to EORTC protocol EORTC Rationale for the amendment and it s classification EORTC protocol 10041, Amendment This is a scientific amendment which mainly adjusts timelines from patient screening to randomization and randomization to start of treatment, clarifies rules of dose modifications in the docetaxel-capecitabine arm as well, and finally increases the sample size by 600 patients to compensate the non compliance rate of treatment assigned in the patient group of clinical high and genomic low risk. The PIS/IC will be impacted. This amendment has been discussed and agreed by: the EORTC Headquarters, the study coordinators and steering committee. The trial sponsor companies (Agendia, Roche, Novartis and Sanofi-aventis) have been fully informed. The companies, IDMC and collaborative groups are informed of the scope of the amendment and will receive the final version. PD-004-AF-01 Page 2 of 46 Version AUG 2010

366 PRC submission document Amendment to EORTC protocol EORTC Scientific modifications Page 10-11, Protocol summary Current protocol 10041, Version 2.0 / 25 April, 2008 Number of patients Enrollment of 6,000 patients is planned with those having discordant risk assessments (an estimated 1920 patients) being randomized for treatment decision making. 4,000 patients are planned for the chemotherapy randomization and 3,500 patients planned for the endocrine therapy randomization. Diagnosis and main criteria for inclusion patients with unresectable positive deep margins who receive adjuvant radiotherapy are eligible provided that all other margins are negative. patients should be aged between 18 and 70 years at randomization. (Elderly patients for whom adjuvant treatment is considered could be offered participation in one of the BIG Elderly trials.) NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any invasive cancer (other than breast cancer) in complete remission for 5 years. NOT have received previous chemotherapy, hormonal therapy or radiotherapy. Second Amendment Version 3.0 Enrollment of 6,000 6,600 patients is planned with those having discordant risk assessments (an estimated 1920 patients) being randomized for treatment decision making. Up to 4,000 patients are planned for the chemotherapy randomization and 3,500 patients are planned for the endocrine therapy randomization. patients with unresectable positive deep margins who will receive adjuvant radiotherapy are eligible, provided that all other margins are negative. Patients should be aged between 18 and 70 years at randomization. (Elderly patients for whom adjuvant treatment is considered could be offered participation in one of the BIG Elderly trials.) NOT have previous or concurrent cancer, possible exceptions are:; adequately treated carcinoma in situ of the cervix, nonmelanoma skin cancer, lobular or ductal carcinoma in situ of the breast and any invasive cancer (other than breast cancer) in complete remission for 5 years. NOT have received (neo) adjuvant previous chemotherapy, hormonal (neo) adjuvant endocrine therapy or radiotherapy PD-004-AF-01 Page 3 of 46 Version AUG 2010

367 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 for the primary breast cancer considered for this trial. Page 12, Protocol summary: Treatment Test product, dose and mode of administration setting during the trial. Treatment with trastuzumab is recommended to start after the end of chemotherapy. Page 13, Protocol summary Statistical methods With 6,000 patients accrued overall, this set has an expected size of 672 patients. With an accrual of 3 years, and a total duration of 6 years (so 3 to 6 years follow up for each patient), a one-sided test at 97.5% confidence level has 80% power to reject this hypothesis if the true 5-year DMFS is 95%. Test of hypothesis for R-C: Assuming a control 5-year DFS of 86%, with an expected 4,000 patients in this section of the study, there is 80% power to detect a hazard ratio of 0.76 (or 89% experimental 5-year DFS) at the time of the primary analysis. Test of hypothesis for R-E: With an expected 3,500 patients being randomized into this question, setting during the trial. Treatment with trastuzumab can be proposed either sequential or concomitant with chemotherapy (including patients in capectibaine-docetaxel arm). is recommended to start after the end of chemotherapy. Statistical methods With 6,000 6,600 patients accrued overall, this set has an expected size of 672 patients. With an accrual of 3 years, and a total duration of 6 years (so 3 to 6 years follow up for each patient), The primary test will be a one-sided test at 97.5% confidence level. has 80% power to reject this hypothesis if the true 5-year DMFS is 95%. Test of hypothesis for R-C: Assuming a control 5-year DFS of 86%, due to an insufficient number of patients randomized into this part of the study with an expected 4,000 patients in this section of the study, there will be less than is 80% power to detect the original alternative hypothesis of a hazard ratio of 0.76 (or 89% experimental 5-year DFS) at the time of the primary analysis. Test of hypothesis for R-E: Due to insufficient number of patients With an expected 3,500 PD-004-AF-01 Page 4 of 46 Version AUG 2010

368 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 and assuming a DFS rate at 5 years of 86% on the control arm, there is 80% power to detect a hazard ratio of 0.75 at the time of the primary analysis (i.e. a 5-year DFS of 86% vs. 89.3%). patients being randomized into this question, and assuming a DFS rate at 5 years of 86% on the control arm, there will be less than is 80% power to detect the original alternative hypothesis of a hazard ratio of 0.75 at the time of the primary analysis (i.e. a 5-year DFS of 86% vs. 89.3%). The final analysis of the endocrine comparison will be performed when 379 events have been observed among the patients randomized for the endocrine question, yielding 80% power to detect a hazard ratio of This is projected to occur at a median follow up of 7.9 years. Page 14, Trial Design See new version of the figure (attachment 6) Page 23, The trial: The first trial to evaluate the clinical value of a genomic tool will enroll 6,000 patients to investigate the benefit/risk ratio of chemotherapy when the assessment... All 6,000 patients enrolled in the study will have their risk of recurrence assessed by both clinical-pathological and 70-gene signature methods. Page 37 and 38, 3.1 General eligibility criteria for inclusion in the trial patients with unresectable positive deep margins who receive adjuvant radiotherapy are eligible provided that all other margins are negative. patients should: be aged between 18 and 70 years at randomization. (Elderly patients for whom adjuvant treatment is will enroll 6,000 6,600 patients to investigate the benefit/risk ratio of chemotherapy when the assessment All 6,000 6,600 patients enrolled in the study will have their risk of recurrence assessed by both clinical-pathological and 70-gene signature methods. patients with unresectable positive deep margins who will receive adjuvant radiotherapy are eligible provided that all other margins are negative. Patients should: be aged between 18 and 70 years at randomization. (Elderly patients for whom adjuvant treatment is PD-004-AF-01 Page 5 of 46 Version AUG 2010

369 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 considered could be offered participation in one of the BIG Elderly trials.) NOT have previous or concurrent cancer, possible exceptions are; adequately treated carcinoma in situ of the cervix, non-melanoma skin cancer, and any invasive cancer (other than breast cancer) in complete remission for 5 years. NOT have received previous chemotherapy, hormonal therapy or radiotherapy. Second Amendment Version 3.0 considered could be offered participation in one of the BIG Elderly trials.) NOT have previous or concurrent cancer, possible exceptions are; : adequately treated carcinoma in situ of the cervix, nonmelanoma skin cancer, lobular or ductal carcinoma in situ of the breast and any invasive cancer (other than breast cancer) in complete remission for 5 years. NOT have received (neo) adjuvant previous chemotherapy, hormonal (neo) adjuvant endocrine therapy or radiotherapy for the primary breast cancer considered for this trial. Page 39, 3.3 Selection criteria for chemotherapy randomization (R-C) Interval between definitive surgery and start of chemotherapy should ideally be no more than 6 weeks but can not exceed 12 weeks Page 39, 3.4 Selection criteria for endocrine therapy randomization (R-E) women with a history of breast cancer chemoprevention with anti-estrogens can be included if at least 18 months has elapsed between discontinuation of chemoprevention and clinical or radiological diagnosis of breast cancer. Interval between definitive surgery (second surgery, if applicable) and start of chemotherapy should ideally be not more than 6 8 weeks but cannot exceed 12 weeks 120 days women with a history of breast cancer chemoprevention or osteoporosis treatment with anti-estrogens (e.g. tamoxifen, raloxifen) can be included if at least 18 months has elapsed between anti-estrogens discontinuation of chemoprevention and clinical or radiological diagnosis of breast cancer. PD-004-AF-01 Page 6 of 46 Version AUG 2010

370 PRC submission document Amendment to EORTC protocol EORTC Page 40, 4 Trial Design Figure trial design Page 41, 4.1 Screening Current protocol 10041, Version 2.0 / 25 April, 2008 all patients who sign a screening PIS & IC to consent to the donation of a tumor and blood sample, will be registered for screening. Second Amendment Version 3.0 See new version figure trial design (attachment 6) Change in the limit date from surgery to start of randomized CT to 120 days. all patients who sign a screening PIS & IC to consent to the donation of a tumor and (optional) blood sample, will be registered for screening. Page Treatment decision randomization for chemotherapy allocation and for the other half chemotherapy will be assigned according to the 70-gene signature risk. 4.3 Treatment decision randomization for chemotherapy allocation and for the other half chemotherapy will be assigned according to the 70-gene signature risk. Patients with HER-2 positive tumors which have both methods discordant and were randomized to no chemotherapy, can receive adjuvant trastuzumab alone or with adjuvant endocrine therapy (if hormonal receptor positive), if decided by the treating physician. Patients with HER-2 positive tumors that are classified low risk by both methods can receive adjuvant trastuzumab alone or with adjuvant endocrine therapy (if hormonal receptor positive), if decided by the treating physician and if no issues for trastuzumab reimbursement exist in the investigator s country. PD-004-AF-01 Page 7 of 46 Version AUG 2010

371 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, Chemotherapy randomization trial design setting during the trial. Treatment with trastuzumab is recommended to start after the end of chemotherapy. Second Amendment Version 3.0 setting during the trial. Treatment with trastuzumab can be proposed either sequential or concomitant with chemotherapy (including patients in capectibaine-docetaxel arm). is recommended to start after the end of chemotherapy. Page 43, 4.4 Endocrine therapy randomization trial design Hormone therapy treatment should be started for consenting patients. Hormone Endocrine therapy treatment should be started for consenting patients. Page 46, 6 Chemotherapy: therapeutic regimens, expected toxicity, dose modifications 6.1 Treatment plan, anthracycline arm The first cycle of chemotherapy will be administered within 4 weeks following the randomization R-C date and ideally not more than 8 weeks after the definitive surgery. If chemotherapy has not started within 120 days of the definitive surgery the patient becomes ineligible for R-C. 6.1 Treatment plan, anthracycline arm Page 47, Drug administration, anthracycline arm The first cycle of chemotherapy should be administered within 4 weeks following the randomization R-C date and ideally no more than 8 weeks after surgery. If treatment has not started within 12 weeks of surgery the patient becomes ineligible. The first cycle of chemotherapy should be administered within 4 weeks following the randomization R-C date and ideally no more than 8 weeks after surgery. If treatment has not started within 12 weeks of surgery the patient becomes ineligible. Page 48, Drug administration, anthracycline arm PD-004-AF-01 Page 8 of 46 Version AUG 2010

372 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 If there is will be recalculated. No ideal body Second Amendment Version 3.0 If there is will be recalculated. If the calculated BSA of the patient is >2.2 m 2, it is recommended to calculate the dose to be given using BSA=2.2m 2. No ideal body Page 49, Dose adjustments anthracycline arm Full blood... observed during the study). Doses will be reduced Full blood... observed during the study). Rounding of dosage is allowed if the rounding is within 5% of the calculated dose. Doses will be reduced Page 51, Table 4 Dose modification of Anthracycline-Regimens for Non-Hematological Toxicities Table 4 See new version table 4 (attachment 2) Page 55, Drug administration docetaxel-capecitabine arm If body weight varies during the study, it is assumed that the body surface area will remain approximately constant (i.e. no dose adjustments for changes in body weight will be done). If body weight varies during the study, it is assumed that the body surface area will remain approximately constant (i.e. no dose adjustments for changes in body weight will be done). If there is a 10% or more change in body weight between cycles, the BSA will be recalculated and capecitabine dose should be adjusted, according to table 5. Page 57, Dose adjustments docetaxel-capecitabine arm Full blood count will be performed on day 1 of the cycle and adjustments will be based on these results (and also based on the Full blood count will be performed on day 1 of the cycle and adjustments will be based on these results (and also based on the PD-004-AF-01 Page 9 of 46 Version AUG 2010

373 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 guidelines in tables 7 and 8 below). No changes should be done toxicities. Docetaxel therapy should be discontinued if toxicities do not resolve to grade 0 or I within 3 weeks; in that case the patient should be withdrawn from the chemotherapy part of the study and be treated at the discretion of the attending physician. Capecitabine should be resumed at 75% of the starting dose on resolution of the toxicity to grade 0 or I. Capecitabine following the recommendations in table 9. If the calculated creatinine a dose reduction. Second Amendment Version 3.0 guidelines in tables 7 and 8 below). The full blood counts are to be performed on day 1 of each cycle and/or if symptoms are present. In case there is an isolated grade 3 or 4 neutropenia/granulocytopenia without fever during the cycle, capecitabine will not be interrupted. No changes should be done toxicitites. If Docetaxel and/or Capecitabine therapy should be discontinued, due to if toxicities and/or interruption lasts for more than 3 weeks, do not resolve to grade 0 or I within 3 weeks; in that case the patient should be withdrawn from the chemotherapy part of the study and be treated at the discretion of the attending physician. Capecitabine should be resumed at 75% of the starting dose on resolution of the toxicity to grade 0 or I. However, if the patient needs discontinuation of one of the drugs due to toxicity, it is at the investigators discretion to continue with the remaining drug or to stop overall. Capecitabine following the recommendations in table 9. In case blood count test is performed during the 14 days of Capecitabine administration, treatment should only be interrupted if a non-hematological adverse event (grade 2) coincides with a grade 3 or 4 neutropenia. In that case, one should wait until resolution of both adverse events (including the neutropenia) to a grade 0 or I. It is recommended to use G- CSF or PEG-filgrastim in subsequent cycles. The doses for toxicity grade II and III occurrence should be adjusted as mentioned above. If the calculated creatinine a dose reduction. PD-004-AF-01 Page 10 of 46 Version AUG 2010

374 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Page 58, Table 7 Dose modification for Hematological Toxicities - Docetaxel-Capecitabine combination Table 7 Page 59, Table 8 Dose modification for non-hematological Toxicities - Docetaxel-Capecitabine combination Table 8 Second Amendment Version 3.0 See new version table 7 (attachment 3) Clarified Neutropenia grade 4 See new version table 8 (attachment 4) Modified Neurologic AE, HFS grade 4 deleted, Mucositis grade I deleted Page 64, 6.3 Treatment withdrawal criteria The patient will then be considered off-protocol chemotherapy treatment (however the patient remains eligible for R-E), but should be followed according to the protocol. The patient will then be considered off-protocol randomized chemotherapy treatment, (however the patient remains eligible for R-E) and, but should be followed according to the protocol. Page 64, 7 Endocrine therapy Page 65, 7.2 Side effects of drugs No standard dosage modifications are prescribed. (new paragraph to be added before section 7.1) The first dose of endocrine therapy should be administered within 4 weeks following the randomization (R-E) date. If treatment has not started within 300 days after definitive surgery, the patient becomes ineligible for randomized endocrine therapy. No standard dosage modifications are prescribed. PD-004-AF-01 Page 11 of 46 Version AUG 2010

375 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 Patients in the Tamoxifen-Letrozole arm, who are under tamoxifen treatment and need to discontinue tamoxifen due to toxicity, can switch early to letrozole and complete the 7 years of treatment per protocol. Patients who discontinue the treatment are considered off randomized endocrine therapy and should be treated at the discretion of treating physician. All patients should be followed up according to study protocol. Missing doses due to treatment interruption should not be replaced. Treatment cannot be interrupted for more than 90 consecutive days (see treatment withdrawal criteria). Page 65, 7.3 Ovarian function suppression Bilateral surgical oopherectomy. Page 66, 7 Endocrine therapy Considering the and weight bearing exercise (see ASCO guidelines on bone care (Ref. 98)). Bilateral surgical oopheorectomy. Considering the and weight bearing exercise (see ASCO guidelines on bone care (Ref.98)). 7.5 Treatment withdrawal criteria The treatment should be withdrawn if: The patient, at any time, withdraws consent to participate The Investigator judges that the decision is in the best interest of the patient The treatment must be interrupted for more than 90 consecutive days PD-004-AF-01 Page 12 of 46 Version AUG 2010

376 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 There is evidence of disease recurrence/relapse The patient becomes pregnant The patient is lost to follow-up The patient will then be followed-up according to the protocol. Page 66, 8.1 Baseline evaluations Chest x-ray Chest x-ray or Thoracic CT scan Page 70, Table 11 3 If standard of care in the institute 3 If standard of care in the institute (thoracic CT allowed) Page 71, Radiological assessments A Chest X-ray should be performed according to standard of care in the institute and preferably yearly. Posterior-anterior (PA) and lateral views are preferred to local practice. A Chest X-ray or Thoracic CT scan should be performed prior to enrollment (baseline evaluation) according to standard of care in the instituteion and preferably yearly if clinically indicated during follow-up. For chest X-ray Pposterior-anterior (PA) and lateral views are preferred to local practice. Page 72, Overall Survival (OS) Patients still known to be alive. Patients still known to be alive. Early death will be defined as death occurring within 30 days of last protocol treatment or within 90 days from enrollment/randomization. PD-004-AF-01 Page 13 of 46 Version AUG 2010

377 PRC submission document Amendment to EORTC protocol EORTC Page 73, Sample size Current protocol 10041, Version 2.0 / 25 April, 2008 to use the 70-gene signature prognosis and thus receive no chemotherapy, a null hypothesis of a 5-year distant metastasis free survival (DMFS) of 92% will be tested. With 6,000 patients accrued overall, this set has an expected size of 672 patients. With an accrual of 3 years, and a total duration of 6 years (so 3 to 6 years follow up for each patient), a one-sided test at 97.5% confidence level has 80% power to reject this hypothesis if the true 5-year DMFS is 95%. The primary test will be performed when three conditions are met: and back-transformation) is 0.01or less, at least one third of the patients in the above dataset have been followed for five years, and the number of events for the primary test for R-C has been reached. two chemotherapy arms randomized. With an expected 4,000 patients in this section of the study, there is 80% power to detect a hazard ratio of 0.76 (for example a control 5-year DFS of 86% versus a 89.2% experimental 5-year DFS) with at least 422 events (deaths or recurrences among the patients randomized for R-C). The Second Amendment Version 3.0 to use the 70-gene signature prognosis and thus received no chemotherapy (i.e. they truly did not receive chemotherapy), a null hypothesis of a 5-year distant metastasis free survival (DMFS) of 92% will be tested. With 6,600 6,000 patients accrued overall, this set has an expected size of 672 patients. With an accrual of 3 years, and a total duration of 6 years (so 3 to 6 years follow up for each patient), The primary test will be a one-sided test at 97.5% confidence level. has 80% power to reject this hypothesis if the true 5-year DMFS is 95%. This The primary test will be performed when two three conditions are met: and back-transformation) is 0.01 or less, and at least one third of the patients in the above dataset have been followed for five years., and the number of events for the primary test for R-C has been reached. Using these conditions, this test has 80% power to reject the null hypothesis if the true 5-year DMFS is 95%. two chemotherapy arms randomized. Due to insufficient numbers of patients randomized into this part of the study, there will be less than 80% power With an expected 4,000 patients in this section of the study, there is 80% power to detect the original alternative hypothesis of a hazard ratio of 0.76 (for example a control 5-year DFS of 86% versus a 89.2% experimental PD-004-AF-01 Page 14 of 46 Version AUG 2010

378 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 comparisons for R-C will be performed after at least 422 events have been observed for this comparison. After 211 events have been observed, an interim test will be performed for this comparison, using a rho spending function (Ref. 100) with rho=2.5, which is somewhat less conservative than an O Brien-Fleming spending function. If the number of events at the time of the interim test is exactly 211, the tests at interim and at final analysis will have nominal alpha values of and , respectively. If the true hazard ratio is 0.6, this interim test has 85% power of rejecting the null hypothesis. two sequences of endocrine therapy randomized. With an expected 3,500 patients being randomized into this question, there is 80% power to detect a hazard ratio of 0.75 (for example a DFS rate at 5 years of 86% on the control arm versus an experimental 5-year DFS of 89.3%) with at least 382 events. After 191 events have been observed, an interim test will be performed for this comparison, also using a rho spending function with rho=2.5. If the number of events at the time of the interim test is exactly 191, the tests at interim and at final analysis will have nominal alpha values of and , respectively. If the true hazard ratio is 0.6, this interim test has 81% power. Second Amendment Version year DFS) with at least 422 events (deaths or recurrences among the patients randomized for R-C). The comparisons for R-C will be performed at the time of the primary test (as described above) at 5% alpha and an a posteriori power calculation will be performed. after at least 422 events have been observed for this comparison. After 211 events have been observed, an interim test will be performed for this comparison, using a rho spending function (Ref.100) with rho=2.5, which is somewhat less conservative than an O Brien-Fleming spending function. If the number of events at the time of the interim test is exactly 211, the tests at interim and at final analysis will have nominal alpha values of and , respectively. If the true hazard ratio is 0.6, this interim test has 85% power of rejecting the null hypothesis. two sequences of endocrine therapy randomized. Due to insufficient numbers of patients With an expected 3,500 patients being randomized into this question, there will be less than is 80% power to detect the original alternative hypothesis of a hazard ratio of 0.75 (for example a DFS rate at 5 years of 86% on the control arm versus an experimental 5-year DFS of 89.3%). with at least 382 events. After 191 events have been observed, an interim test will be performed for this comparison, also using a rho spending function with rho=2.5. If the number of events at the time of the interim test is exactly 191, the tests at interim and at final analysis will have nominal alpha values of and , respectively. If the true hazard ratio is 0.6, this interim test has 81% power. The primary analysis of the endocrine comparison will be performed when 379 events have been observed among the PD-004-AF-01 Page 15 of 46 Version AUG 2010

379 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 patients randomized for the endocrine question, yielding 80% power to detect a hazard ratio of This is projected to occur at a median follow up of 7.9 years. Page 74, Randomization and stratifications R-T: Patients for...hr status (Positive (ER and/or PgR) vs. Negative (both)), nodal (mastectomy or quadrantectomy/tumorectomy). Patients who are...hr status (Positive (ER and/or PgR) vs. Negative (both)), age (mastectomy or quadrantectomy/tumorectomy). R-T: Patients for...hr status (Positive (ER and/or PgR) vs. Negative (both)), nodal (mastectomy or quadrantectomy/tumorectomy). In case PgR is unknown, the patient will be stratified to the HR negative group if ER is negative, and to the HR positive group if ER is positive. Patients who are...hr status (Positive (ER and/or PgR) vs. Negative (both)), age (mastectomy or qudrantectomy/tumorectomy). In case PgR is unknown, the patient will be stratified to the HR negative group if ER is negative, and to the HR positive group if ER is positive. Page 75, 10.2 Statistical analysis plan Analysis populations and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy. This dataset is used for the primary test. and who were randomized (R-T) to use the 70-gene signature risk and thus receive no chemotherapy. and did not deviate from this: no change in risk status post enrollment and no chemotherapy received. This dataset is used for the primary test. Primary test dataset as enrolled (PTE): The set of patients who have a low risk gene prognosis signature and high risk PD-004-AF-01 Page 16 of 46 Version AUG 2010

380 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 clinical-pathological criteria, and who were randomized (R- T) to use the 70-gene signature risk and thus receive no chemotherapy, regardless of compliance to the no chemotherapy assignment. A patient will be considered to be eligible if she did not have any major deviations from the patient entry criteria listed in chapter 3. Eligibility will be assessed by the Study Coordinator based on the review of each patient file. A patient will be considered to be eligible if she did not have any major deviations from the patient entry criteria listed in chapter 3 of the protocol. Eligibility will be assessed by the Study Coordinator based on the review of each patient file. Potential eligibility problems will be assessed by the Clinical Research Physician (CRP) at time of medical review. If needed the CRP can ask the help of the Study Coordinators Timing of analyses Interim analysis on the ITT2, SP2 datasets will be performed at the time at least 211 events for DFS have been observed on the ITT2 dataset. If that analysis is significant as described above, this will be the final analysis for the chemotherapy comparison. If not, the final analysis on the ITT2, SP2 datasets will be performed at the time at least 422 events for DFS have been observed on the ITT2 dataset Timing of analyses Interim analysis on the ITT2, SP2 datasets will be performed at the time at least 211 events for DFS have been observed on the ITT2 dataset. If that analysis is significant as described above, this will be the final analysis for the chemotherapy comparison. If not, the The final analysis on the ITT2, SP2 datasets will be performed at the time at least 422 events for DFS have been observed on the ITT2 dataset of the primary analysis on the PT dataset. All analyses on the AP, PT, ITT1, PP1 datasets will be performed when the three conditions in section have been met. All analyses on the AP, PT, PTE, ITT1, PP1 datasets will be performed when the two three conditions in section have been met. PD-004-AF-01 Page 17 of 46 Version AUG 2010

381 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Interim analysis on the ITT3, SP3 datasets will be performed at the time at least 191 events for DFS have been observed on the ITT3 dataset. If that analysis is significant as described above, this will be the final analysis for the endocrine comparison. If not, the final analysis on the ITT3, SP3 datasets will be performed at the time at least 382 events for DFS have been observed on the ITT3 dataset. All analyses are driven on the number of events observed, and so the timing depends on accrual, relative size of the appropriate analysis set, event rates, and loss to follow-up. Under the assumption of a homogeneous 3 year accrual, and under the alternative hypotheses stated above for the respective tests, the expected timing of the analyses is as follows: - interim test for chemotherapy (ITT2): 4 years after start of accrual - interim test for endocrine therapy (ITT3): 4.5 years after start of accrual - primary test on AP, analyses for ITT1, final test for chemotherapy (ITT2): 6.3 years after start of accrual - primary test for endocrine therapy (ITT3): 7 years after start of accrual Second Amendment Version 3.0 Interim analysis on the ITT3, SP3 datasets will be performed at the time at least 191 events for DFS have been observed on the ITT3 dataset. If that analysis is significant as described above, this will be the final analysis for the endocrine comparison. If not, the The final analysis on the ITT3, SP3 datasets will be performed at the time at least 382 events for DFS have been observed when 379 events have been observed on the ITT3 dataset. This is projected to occur at a median follow up of 7.9 years. All analyses are driven on the number of events observed, and so the timing depends on accrual, relative size of the appropriate analysis set, event rates, and loss to follow-up. Under the assumption of a homogeneous 3 year accrual, and under the alternative hypotheses stated above for the respective tests, the expected timing of the analyses is as follows: - interim test for chemotherapy (ITT2): 4 years after start of accrual - interim test for endocrine therapy (ITT3): 4.5 years after start of accrual - primary test on AP, analyses for ITT1, final test for chemotherapy (ITT2): 6.3 years after start of accrual - primary test for endocrine therapy (ITT3): 7 years after start of accrual Page 76, Statistical methods Full analysis plan The outline of analysis given in this section of the protocol will be further detailed in a separate analysis plan that will be PD-004-AF-01 Page 18 of 46 Version AUG 2010

382 PRC submission document Amendment to EORTC protocol EORTC General All statistical tests Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 finalized prior to any analyses being performed. General All statistical tests Page 79, Prognostic factor analyses on the basis of microarray data for 6,000 patients, or to develop or validate on the basis of microarray data for 6,600 6,000 patients, or to develop or validate Page 81, 10.3 End of study 3. The database has been fully cleaned and frozen for this analysis Pilot study of the trial phase III randomized trial that will include approximately 6,000 patients, will Page 82, Interim analyses for the chemotherapy and the endocrine questions Interim analyses for the chemotherapy and the endocrine questions An interim analysis of the primary comparison for R-C will be performed as described in section This interim analysis will be reviewed by the EORTC-IDMC. It will be restricted to the ITT2 dataset, and only consider the comparison of the two randomized regimens of chemotherapy. A significant outcome may lead to early publication of results, restricted to this particular comparison. An interim analysis of the primary comparison for R-E will be 3. The database has been fully cleaned and frozen for this analysis these analyses phase III randomized trial that will include approximately 6,600 6,000 patients, will Interim analyses for the chemotherapy and the endocrine questions An interim analysis of the primary comparison for R-C will be performed as described in section This interim analysis will be reviewed by the EORTC-IDMC. It will be restricted to the ITT2 dataset, and only consider the comparison of the two randomized regimens of chemotherapy. A significant outcome may lead to early publication of results, restricted to this particular comparison. An interim analysis of the primary comparison for R-E will be PD-004-AF-01 Page 19 of 46 Version AUG 2010

383 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 performed as described in section This interim analysis will also be reviewed by the EORTC-IDMC. It will be restricted to the ITT3 dataset, and only consider the comparison of the two randomized sequences of endocrine therapy. A significant outcome may lead to early publication of results, restricted to this particular comparison. The following table gives an overview of the analyses planned for each of the three primary endpoints. Table 12: Overview of interim and final analyses of primary endpoints Primary endpoint for Endpoint Analysis population Alpha spending function Analysis R-T DMFS PT NA Final R-C DFS ITT2 R-E DFS ITT3 Rho = 2.5 Rho = 2.5 Interim Final Interim Final Timing See events 422 events 191 events 382 events Nominal 2-sided alpha level * * * * * These are the nominal levels if the interim analysis is at exactly half the planned number of events. If the number of events at interim is slightly more, the correct values according to the spending function will be used. These are very close to the tabulated values. Second Amendment Version 3.0 performed as described in section This interim analysis will also be reviewed by the EORTC-IDMC. It will be restricted to the ITT3 dataset, and only consider the comparison of the two randomized sequences of endocrine therapy. A significant outcome may lead to early publication of results, restricted to this particular comparison. The following table gives an overview of the analyses planned for each of the three primary endpoints. Table 12: Overview of interim and final analyses of primary endpoints Primary endpoint for Endpoint Analysis population Alpha spending function Analysis R-T DMFS PT NA Final R-C DFS ITT2 R-E DFS ITT3 Rho = 2.5 Rho = 2.5 Interim Final Interim Final Timing See events 422 events 191 events 382 events Nominal 2-sided alpha level * * * * * These are the nominal levels if the interim analysis is at exactly half the planned number of events. If the number of events at interim is slightly more, the correct values according to the spending function will be used. These are very close to the tabulated values. PD-004-AF-01 Page 20 of 46 Version AUG 2010

384 PRC submission document Amendment to EORTC protocol EORTC Page 83, 12.1 Sample collection Current protocol 10041, Version 2.0 / 25 April, optional blood sample (for whole blood and serum storage) for genetics and proteomics analysis and biological materials bank storage for future research to be taken before surgery. Second Amendment Version optional blood sample (for whole blood and serum storage) for genetics and proteomics analysies and biological materials bank storage for future research to be taken during the study (preferably before surgery). Page 88, Access to the interactive randomization program All trial authorized investigators can enroll/randomize patients directly on the website, 24 hours a day, 7 days a week ( To access the website containing the interactive randomization program, investigators need a username and a password. All authorized investigators will be provided with a username and password prior to site initiation. EORTC participants can use their ORTA username and password to access the website and if they have forgotten their ORTA password, they can interactively request a password reminder from the EORTC ORTA website ( Page 88, 15.2 Overview of patient screening and enrollment Access to the interactive randomization program Access to RDC for registration/enrollment/randomization Patients are registered, enrolled and randomized directly on RDC (RDC= Remote Data Capture), All trial authorized investigators can enroll/randomize patients directly on the website, accessible 24 hours a day, 7 days a week ( Guidelines on how to register, enroll and randomize patients in RDC are provided to all centers by the EORTC. To access the website containing the interactive randomization program RDC, investigators need a username and a password. All authorized investigators will be provided with a username and password prior to site initiation. EORTC participants can use their ORTA username and password to access the website RDC. and if If an ORTA password has been they have forgotten their ORTA password, it they can interactively be requested by a password reminder through from the EORTC ORTA website ( Steps to be taken for patient registration for screening (also see Steps to be taken for patient registration for screening (also see PD-004-AF-01 Page 21 of 46 Version AUG 2010

385 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 diagram chapter 20) take blood sample for future research (optional for patient) breast cancer surgery, freeze tumor biopsy and register tumor sample as available for collection on website as soon as lymph node status is confirmed by local pathology this information must be recorded on the website (then the genomic test will be performed for eligible patients) Second Amendment Version 3.0 diagram chapter 20) take blood sample for future research (optional for patient). Collected during the study, preferably before surgery. date of breast cancer surgery, freeze tumor biopsy and register tumor sample as available for collection on website RDC and Samples website as soon as lymph node status is confirmed by local pathology this information must be recorded on the log lymph node form (RG3) on the website RDC (then the genomic test will be performed for eligible patients). Note that, in case of reoperation, to achieve axillary clearance (e.g. axillary lymph node dissection due to positive sentinel node biopsy) the lymph node status must be confirmed only after the definitive surgery. Page 89, 15.2 Overview of patient screening and enrollment a paper enrollment checklist (detailing the eligibility criteria) will be provided and can be filled in before connecting to the website to enroll a patient connect to website, input SeqID & confirm date of birth then input clinical data and date of signing of PIS & IC 1 to enroll the patient. Comparison of clinical and genomic risk assessments and receive the treatment allocation (results of clinical and genomic prognostic tests) a paper enrollment checklist (detailing the eligibility criteria) will be provided and can be filled in before connecting to the website to enroll a patient connect to website RDC, select the correct input SeqID & confirm date of birth, create the enrollment checklist (RG5) then and input clinical data and date of signing of PIS & IC 1 to enroll the patient. Comparison of clinical and genomic risk assessments and receive the treatment allocation (results of PD-004-AF-01 Page 22 of 46 Version AUG 2010

386 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 clinical and genomic prognostic tests). After the previous forms and the enrollment checklist (RG5) have been submitted, the results form (RG5trt) will be available on RDC. After completing Rg5trt, clinical and genomic risk result and treatment decision outcome will be available on this results form (RG5trt). either print the completed enrollment checklist screen print from the web based platform or complete a paper checklist, attach the sample sticker and sign it. Send it to your NCC/EORTC within 3 weeks of enrolling the patient. Sign and date subsequent checklist(s)/screenprints and send to your NCC/G or EORTC Page 89, Patient registration for screening Patients will be registered for screening before or at the time of surgery and tumor sample availability. When a patient is registered for screening some clinical information will be requested and the web based platform will generate a unique sequential identification number (SeqID) for each patient which will also serve to identify the patient during the trial if the patient is eligible and enrolled. This SeqID must be quoted at all subsequent sessions on the website. The SeqID number attributed to the patient at the end of the registration for screening procedure must be recorded on the enrollment checklist, and must be reported on all case report forms. either print the completed enrollment checklist screen print from the web based platform or complete a paper checklist, attach the sample sticker and sign it. Send it to your NCC/EORTC within 3 weeks of enrolling the patient. Sign and date subsequent checklist(s)/screenprints and send to your NCC/G or EORTC Page 89, Patient registration for screening Patients will be registered for screening before or at the time of surgery and tumor sample availability. When a patient is registered for screening some clinical information will be requested and the web based platform will generate a unique sequential identification number (SeqID) for each patient which will also serve to identify the patient during the trial if the patient is eligible and enrolled. This SeqID must be quoted at all subsequent sessions on the website. The SeqID number attributed to the patient at the end of the registration for screening procedure must be recorded on the enrollment checklist, and must be reported on all case report forms. PD-004-AF-01 Page 23 of 46 Version AUG 2010

387 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 PD-004-AF-01 Page 24 of 46 Version AUG 2010

388 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Page 89, Patient enrollment and randomization for treatment decision making tool (R-T) A patient can be enrolled and randomized only after verification of eligibility. This must be done before the start of the protocol treatment. An exhaustive list of questions to be answered during the enrollment/randomization procedure is included in the enrollment checklist, which is part of the case report forms. This checklist should be completed by the responsible investigator before the patient is enrolled. In order to enroll a patient in the trial, the interactive randomization program needs to be accessed via the website and the following questions answered: provide SeqID confirm Date Of Birth (DOB) (day/month/year) patient's code (maximum 4 letters or digits chosen at the discretion of the investigator) date of written informed consent Eligibility will be confirmed by collection of all the data requested on the enrollment checklist. The risk according to clinical-pathological criteria will be calculated by the Adjuvant! Online software incorporated into the web based platform. The patient s age, ER status, tumor grade, tumor size and Second Amendment Version 3.0 Page 89, Patient enrollment and randomization for treatment decision making tool (R-T) A patient can be enrolled and randomized only after verification of eligibility. This must be done before the start of the protocol treatment. An exhaustive list of questions to be answered during the enrollment/randomization procedure is included in the enrollment checklist (RG5), which is part of the case report forms. This checklist should be completed on RDC by the responsible investigator before the patient is enrolled. In order to enroll a patient in the trial, the interactive randomization program needs to be accessed via the website and the following questions answered: provide SeqID confirm Date Of Birth (DOB) (day/month/year) patient's code (maximum 4 letters or digits chosen at the discretion of the investigator) date of written informed consent Eligibility will be confirmed by collection of all the data requested on the enrollment checklist. The risk according to clinical-pathological criteria will be calculated by the Adjuvant! Online software incorporated into the web based platform RDC. The patient s age, ER status, HER2 status, tumor PD-004-AF-01 Page 25 of 46 Version AUG 2010

389 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 nodal status will be passed on to Adjuvant! Online by the web based platform. Co-morbidity will be defaulted to Minor problems This completes enrollment. The enrollment checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks of enrollment. The Principal Investigator, NCC/G where applicable and the EORTC Headquarters will receive an confirming patient enrollment with details of both the genomic and the clinical risk assessment results and the result of randomization for all discordant patients Page 90-91, Randomization of chemotherapy (R-C) Once a patient has been assigned to receive chemotherapy, she can be offered the randomization to anthracycline based chemotherapy or docetaxel-capecitabine chemotherapy. This randomization can be done immediately after the assignment to chemotherapy, or later. If the chemotherapy randomization (R-C) is done later, the interactive randomization program needs to be accessed and the sequential identification number attributed to the patient provided. The computer will display all known information entered for this patient, including whether this patient has been allocated to receive chemotherapy. Fill in the chemotherapy randomization checklist and then log-on to Second Amendment Version 3.0 grade, tumor size and nodal status will be passed on to Adjuvant! Online by the web based platform RDC. Co-morbidity will be defaulted to Minor problems This completes enrollment. All resulting outcomes (clinical risk, genomic risk and treatment decision outcome) are visible on the results form (RG5trt) which the Principal Investigator has to create after the enrollment checklist (RG5) is sent. The enrollment checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks of enrollment. The Principal Investigator, NCC/G where applicable and the EORTC Headquarters will receive an confirming patient enrollment with details of both the genomic and the clinical risk assessment results and the result of randomization for all discordant patients Once a patient has been assigned to receive chemotherapy, she can be offered the randomization to anthracycline based chemotherapy or docetaxel-capecitabine chemotherapy. This randomization can be done immediately after the assignment to chemotherapy, or later. If the chemotherapy randomization (R-C) is done later, the interactive randomization program needs to be accessed and the sequential identification number attributed to the patient provided. In case the treatment decision outcome is chemotherapy, the chemotherapy checklist (RG6) has to be completed on RDC. The computer will display all known information entered for this patient, including whether this patient has been allocated to receive PD-004-AF-01 Page 26 of 46 Version AUG 2010

390 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 the website to randomize the patient. If the patient agrees to the chemotherapy randomization, fill in all the information from the chemotherapy randomization checklist and the computer will allocate her to This completes the chemotherapy randomization. The randomization checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks. The Principal Investigator, NCC/G if applicable and the EORTC Headquarters will receive an confirming the chemotherapy treatment allocation. If a patient who should be proposed chemotherapy does not wish to participate in the chemotherapy randomization (R-C), she should receive the hospital s standard chemotherapy. You will still need to complete the first 2 questions of the chemotherapy checklist and then say which non-randomized chemotherapy the patient will receive. The patient remains on protocol and a summary of the chemotherapy treatment given should be recorded on the non-randomized chemotherapy CRF. Second Amendment Version 3.0 chemotherapy. Fill in the chemotherapy randomization checklist and then log-on to the website to randomize the patient If the patient agrees to the chemotherapy randomization, complete the section about chemotherapy randomization fill in all the information from on the chemotherapy randomization checklist (RG6) and the computer randomization algorithm will allocate her to After sending the chemotherapy checklist (RG6), the chemotherapy treatment allocation will be visible on the chemotherapy checklist (RG6) on RDC. This completes the chemotherapy randomization. The randomization checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks. The Principal Investigator, NCC/G if applicable and the EORTC Headquarters will receive an confirming the chemotherapy treatment allocation. If a patient who should be proposed chemotherapy does not wish to participate in the chemotherapy randomization (R-C), she should receive the hospital s standard chemotherapy. You The Principal Investigator will still need to complete the first 2 questions and section 1 of the chemotherapy checklist on RDC. and then say which non-randomized chemotherapy the patient will receive. The patient remains on protocol follow-up and a summary of the chemotherapy treatment given should be recorded on the nonrandomized chemotherapy CRF and the patient will still be a PD-004-AF-01 Page 27 of 46 Version AUG 2010

391 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 candidate to participate to the endocrine therapy randomization. Page 91, Randomization of endocrine therapy (R-E) All hormone receptor positive patients (i.e. ER &/or PgR positive) can be offered the endocrine therapy randomization to 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. In order to randomize a patient, complete the endocrine therapy randomization checklist, then connect to the website. Provide the SeqID attributed to the patient and confirm her date of birth then the computer will display all known information entered for this patient, including whether this patient is eligible to receive endocrine therapy. If the patient agrees to the endocrine therapy randomization, fill in all the information from the endocrine therapy randomization checklist and the computer will allocate her to 1. 2 years of tamoxifen followed by 5 years of letrozole 2. 7 years of letrozole All hormone receptor positive patients (i.e. ER &/or PgR positive) can be offered the endocrine therapy randomization to 2 years of tamoxifen followed by 5 years of letrozole or 7 years of letrozole. In order to randomize a patient, complete the endocrine therapy randomization checklist (RG8) on RDC. then connect to the website. Provide the SeqID attributed to the patient and confirm her date of birth then the computer will display all known information entered for this patient, including whether this patient is eligible to receive endocrine therapy. If the patient agrees to the endocrine therapy randomization, complete the section about endocrine therapy randomization fill in all the information from on the endocrine therapy randomization checklist (RG8) and the computer randomization algorithm will allocate her to 1. 2 years of tamoxifen followed by 5 years of letrozole 2. 7 years of letrozole After sending the endocrine therapy checklist (RG8), the endocrine treatment allocation will be visible on the endocrine therapy checklist (RG8) on RDC. PD-004-AF-01 Page 28 of 46 Version AUG 2010

392 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 This completes the endocrine therapy randomization. The randomization checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks. The Principal Investigator, NCC/G if applicable and the EORTC Headquarters will receive an confirming the endocrine therapy treatment allocation. If a patient with an endocrine responsive tumor does not wish to participate in the endocrine therapy randomization (R-E), she should receive the hospital s standard endocrine therapy. You will still need to complete the first 2 questions of the endocrine therapy checklist and then say which nonrandomized endocrine therapy the patient will receive. The patient remains on protocol and the endocrine therapy treatment given should be recorded on the adjuvant endocrine therapy CRF. Page 91, 16.1 Electronic case report forms and schedule for completion All further data will be reported on electronic forms which will be completed and submitted using Remote Data capture (RDC). Guidelines on how to use RDC will be provided to all centers by the EORTC. All RDC forms will be specifically designed by the EORTC Headquarters for this study. These electronic forms will be used by all participants. Queries will be generated by the EORTC and sent (as.pdf) either to the National Coordinating Centers/Groups (NCC/G) to distribute or to the centers directly to centers in countries where the EORTC is the NCC/G. The query.pdf forms will be printed and filled in on paper by the PI (and anyone authorized to sign forms by the PI and included on the signature log) and then signed and Second Amendment Version 3.0 This completes the endocrine therapy randomization. The randomization checklist/screenprint will be signed and dated by the Principal Investigator and sent to the NCC/G or EORTC within 3 weeks. The Principal Investigator, NCC/G if applicable and the EORTC Headquarters will receive an confirming the endocrine therapy treatment allocation. If a patient with an endocrine responsive tumor does not wish to participate in the endocrine therapy randomization (R-E), she should receive the hospital s standard endocrine therapy. You The Principal Investigator will still need to complete the first 2 questions and section 1 of the endocrine therapy checklist on RDC. and then say which non-randomized endocrine therapy the patient will receive. The patient remains on protocol and the endocrine therapy treatment given should be recorded on the adjuvant endocrine therapy CRF. All further data will be reported on electronic forms which will also be completed and submitted using Remote Data capture (RDC). Once the results form (RG5trt) is completed and the patient is enrolled, all case report forms will be visible and available for completion on RDC. Completion gguidelines on how to use RDC for all forms will be are provided to all centers by the EORTC. All RDC forms will be are specifically designed by the EORTC Headquarters for this study. These electronic forms will be used by all participants. Electronic queries Queries will be generated by the EORTC. These queries will be displayed in RDC and should be answered in RDC. and sent (as.pdf) either to the National PD-004-AF-01 Page 29 of 46 Version AUG 2010

393 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 returned to their NCC/G or directly to the EORTC Headquarters in countries where the EORTC is the NCC/G. Forms to be sent to the EORTC Headquarters directly should be sent to: Breast Cancer Group Data Manager EORTC Headquarters Avenue Emmanuel Mounier, 83, bte 11 B-1200 Brussels, Belgium A. Before the adjuvant treatment starts: the patient must be enrolled/randomized through the web based platform a signed and dated screenprint/paper copy of the enrollment check list, must be sent either to your NCC/G or directly to the EORTC Headquarters (if the EORTC is your NCC/G) within 3 weeks of enrollment. The optimal way to work is to complete the enrollment checklist first and to enroll/randomize the patient as soon as it is completed. The date of registration and patient sequential identification number are then completed on the checklist, and this form can be sent to the Headquarters. Further randomization checklist(s)/screenprint(s) (chemotherapy R-C & endocrine therapy R-E as applicable), must also be sent either to your NCC/G or directly to the EORTC Headquarters (if the EORTC Second Amendment Version 3.0 Coordinating Centers/Groups (NCC/G) to distribute or to the centers directly to centers in countries where the EORTC is the NCC/G. The query.pdf forms will be printed and filled in on paper by the PI (and anyone authorized to sign forms by the PI and included on the signature log) and then signed and returned to their NCC/G or directly to the EORTC Headquarters in countries where the EORTC is the NCC/G. Forms to be sent to the EORTC Headquarters directly should be sent to: Breast Cancer Group Data Manager EORTC Headquarters Avenue Emmanuel Mounier, 83, bte 11 B-1200 Brussels, Belgium A. Before the adjuvant treatment starts: the patient must be enrolled/randomized through the web based platform a signed and dated screenprint/paper copy of the enrollment check list, must be sent either to your NCC/G or directly to the EORTC Headquarters (if the EORTC is your NCC/G) within 3 weeks of enrollment. The optimal way to work is to complete the enrollment checklist first and to enroll/randomize the patient as soon as it is completed. The date of registration and patient sequential identification number are then completed on the checklist, and this form can be sent to the Headquarters. Further randomization checklist(s)/screenprint(s) (chemotherapy R- C & endocrine therapy R-E as applicable), must also be sent either to your NCC/G or directly to the EORTC Headquarters (if the PD-004-AF-01 Page 30 of 46 Version AUG 2010

394 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 is your NCC/G). EORTC is your NCC/G). Page 93, 16.2 Data Flow (Remote Data Capture) All forms must be electronically completed according to the schedule defined in the CRF guidelines through the EORTC web based Remote Data Capture (RDC) system. The list of staff members authorized to enter forms must be identified on the signature log and sent to the Headquarters by the responsible investigator before the start of the study. In all cases, it remains the responsibility of the investigator to check that On the data received, the EORTC Headquarters will perform extensive consistency checks and issue queries in case of inconsistent data. Queries will be sent by (PDF), and must be filled out on the printed paper copy. A copy should be kept on site, the original must be sent by regular mail to your NCC/G or to the EORTC Headquarters. The EORTC data manager will subsequently apply the corrections into the database. If an investigator (or an authorized staff member) needs to modify a CRF after the form has been sent to the EORTC Headquarters, he/she should use a Data Correction Sheet and sent it to their NCC/G or the EORTC Headquarters. Page 96, 18.1 Control of data consistency Data forms will be entered in the database of the EORTC All The forms must be completed electronically completed according to the schedule defined in the CRF guidelines through the EORTC web based Remote Data Capture (RDC) system. The list of site staff members, authorized to enter forms data, must be identified on the signature log and sent to the Headquarters by the responsible investigator before the start of the study. In all cases, it remains the responsibility of the principal investigator to check that On the data received, the The EORTC Headquarters will perform extensive consistency checks on the received data and will issue queries in case of inconsistent data. Queries will be sent by (PDF), and must be filled out on the printed paper copy. A copy should be kept on site, the original must be sent by regular mail to your NCC/G or to the EORTC Headquarters. These queries will appear in the RDC system and must be answered there directly. The EORTC data manager will subsequently apply the corrections into the database. If an investigator (or an authorized staff member) needs to modify a CRF after the form has been electronically sent to the EORTC Headquarters, he/she should create a request for data correction in the RDC system. use a Data Correction Sheet and sent it to their NCC/G or the EORTC Headquarters. Data forms will be entered in electronically sent to the database of PD-004-AF-01 Page 31 of 46 Version AUG 2010

395 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Headquarters using the RDC system. Computerized and manual consistency checks will be performed on newly entered forms; queries will be issued in case of inconsistencies. Consistent forms will be validated by the Data Manager to be entered on the master database. Inconsistent forms will be kept "pending" until resolution of the inconsistencies. Second Amendment Version 3.0 the EORTC Headquarters by the RDC (Remote Data Capture) system.using the RDC system. Computerized and manual consistency checks will be performed on newly entered received forms; queries will be issued in case of inconsistencies. Consistent forms will be validated by the Ddata M manager(s) to be entered ion the master database. Inconsistent forms will be kept "pending" until resolution of the inconsistencies. Page 97, 19.1 Patient protection The responsible investigator will ensure that this study is conducted in agreement with either the Declaration of Helsinki, see Appendix D (Tokyo, Venice, Hong Kong, Somerset West and Edinburgh amendments) or the laws and regulations of the country, whichever provides the greatest protection of the patient. The protocol has been written, and the study will be conducted according to the ICH Harmonized Tripartite Guideline for Good Clinical Practice (ref: The protocol will be approved by the Local, Regional or National Ethics Committees. Page 99, 20.1 Frozen tumor sample schedule One, or if feasible 2, biopsies of 3 mm are acceptable. The responsible investigator will ensure that this study is conducted in agreement with either the Declaration of Helsinki (available on the World Medical Association website ( see Appendix D (Tokyo, Venice, Hong Kong, Somerset West and Edinburgh amendments) and/or the laws and regulations of the country, whichever provides the greatest protection of the patient. The protocol has been written, and the study will be conducted according to the ICH Harmonized Tripartite Guideline for Good Clinical Practice (ICH-GCP, available online at (ref: The protocol must will be approved by the competent ethics committee(s) as required by the applicable national legislation. the Local, Regional or National Ethics Committees. One, or if feasible 2 biopsies of 3 mm are acceptable. In case of multifocal tumors; if the histopathological profile of all foci are PD-004-AF-01 Page 32 of 46 Version AUG 2010

396 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Page 99, 20.3 Blood and serum sample schedule For those patients for proteomics analysis, blood should be taken after the patient has signed the screening PIS & IC and before surgery, preferably during a routine blood sampling. Page 100, Figure study timeline guidelines from diagnosis to start of treatment 21.1 The study coordinator The Study Coordinator (in cooperation with the EORTC Headquarters) will be responsible for writing the protocol, reviewing all clinical data entered onto electronic case report forms and documenting his/her review on evaluation forms, discussing the contents of the reports with the Data Manager and the Statistician, and for publishing the study results. He will also generally be responsible for answering all clinical questions concerning eligibility, treatment, and the evaluation of the patients The EORTC Headquarters Second Amendment Version 3.0 similar (histopathological type, tumor grade, HR, HER2 status), the biopsy for the genomic assessment should be taken from the largest focus. For those patients for proteomics analysis, blood should be taken after the patient has signed the screening PIS & IC but can be collected and before surgery, preferably during the study a routine blood sampling (preferably before surgery) Trial logistics study timelines guidelines, from diagnosis to start of treatment (see new version of the figure attachment 5) 21.1 The study coordinator The Study Coordinator(s) (in cooperation with the EORTC Headquarters) will be responsible for writing the protocol, reviewing all clinical data entered onto electronic case report forms and documenting his/her review on evaluation forms contributing to the medical review, discussing the contents of the reports with the ddata mmanager(s) and the sstatistician, and for publishing the study results. He/she will assist the Clinical Research Physician also generally be responsible for answering some all clinical questions concerning eligibility, treatment, and the medical review evaluation of the patients The EORTC Headquarters PD-004-AF-01 Page 33 of 46 Version AUG 2010

397 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 The EORTC Headquarters will be responsible for reviewing the protocol, collecting electronic case report forms, controlling the quality of the reported data, and generating reports and analyses in cooperation with the Study Coordinator. Second Amendment Version 3.0 The EORTC Headquarters will be responsible for writing the protocol and PIS/IC, reviewing the protocol, setting up the trial, collecting electronic case report forms, controlling the quality of the reported data, organizing the medical review and generating reports and analyses in cooperation with the Study Coordinator. Page 119, Appendix E: Screening patient information sheet and informed consent document. This blood sample would be taken before the surgery to remove your breast cancer. Page 126, Appendix F Patient Information Sheet - Comparison of risk assessment methods in the trial The trial will include 6,000 women with similar disease to yours over a period of about 3 years or slightly more. have received it according to the clinical-pathological test (an estimated 672 women of the total 6,000 participants) will be very closely monitored. Page 128, Appendix F Patient Information Sheet - Comparison of risk assessment methods in the trial Tumor and blood samples (biological material) In, this group of experts will review a small fragment of some of the tumors of the 6,000 women participating in this trial. Administrative modifications This blood sample would be taken during the study (preferably before surgery). before the surgery to remove your breast cancer. The trial will include 6,000 6,600 women with a similar disease to yours over a period of about 3 5 years or slightly more. have received it according to the clinical-pathological test (an estimated 672 women of the total 6,000 6,600 participants) will be very closely monitored. In, this group of experts will review a small fragment of some of the tumors of the 6,000 6,600 women participating in this trial. PD-004-AF-01 Page 34 of 46 Version AUG 2010

398 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 To include - Signature Page See Signature page (Attachment 1) Page 1 Study coordinators Study Coordinator: Emiel Rutgers Vice-coordinators: Fatima Cardoso and Martine Piccart Page 2, Contacts Writing Committee: F. Cardoso, Institut Jules Bordet, Brussels, Belgium Clinical fellows: Page , 21.1 Study co-coordinator: Dr Fatima Cardoso Address: Institut Jules Bordet 121 Blvd de Waterloo B-1000 Brussels, Belgium Phone: Fax: fatima.cardoso@bordet.be Study Coordinators: Emiel Rutgers ViceCo-coordinators: Fatima Cardoso and Martine Piccart Writing Committee: F. Cardoso, Champalimaud Cancer Center, Lisbon, Portugal Institut Jules Bordet, Brussels, Belgium Clinical fellows: (to be added) G. Werutsky, EORTC Headquarters, Brussels, Belgium C. Moulin, EORTC Headquarters, Brussels, Belgium Study co-coordinator: Dr Fatima Cardoso Address: Institut Jules Bordet 121 Blvd de Waterloo B-1000 Brussels, Belgium Phone: Fax: fatima.cardoso@bordet.be Address: Champalimaud Cancer Center Av. De Brasília - Doca de Pedrouços Lisbon, Portugal Phone: +351 (210) PD-004-AF-01 Page 35 of 46 Version AUG 2010

399 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 Fax: +351 (213) fatimacardoso@fundacaochampalimaud.pt Page , 21.2 The EORTC Headquarters Data Manager: Kaat Vansevenant Phone: Fax: kaat.vansevenant@eortc.be Data Managers: Kaat Vansevenant Virginie Soete Phone: Fax: kaat.vansevenant@eortc.be virginie.soete@eortc.be Roel Goossens Phone: Fax: roel.goossens@eortc.be Project Manager: Jillian Harrison Phone: Fax: jillian.harrison@eortc.be Coordinating Physician: Dr Gaston Demonty Phone: Fax: gaston.demonty@eortc.be Project Manager: Jillian Harrison Kristel Engelen Phone: Fax: jillian.harrison@eortc.be kristel.engelen@eortc.be Coordinating Clinical Research Physician: Dr Gaston Demonty Gustavo Werutsky Phone: Fax: gaston.demonty@eortc.be gustavo.werutsky@eortc.be PD-004-AF-01 Page 36 of 46 Version AUG 2010

400 PRC submission document Amendment to EORTC protocol EORTC Current protocol 10041, Version 2.0 / 25 April, 2008 Second Amendment Version 3.0 Senior Clinical Research Physician Dr. Sandrine Marreaud Phone: Fax: sandrine.marreaud@eortc.be PD-004-AF-01 Page 37 of 46 Version AUG 2010

401 Attachment 1 Chapter: Signature page Chapter Signature page-eortc (update: optional) Protocol Sponsor: Denis Lacombe (Date) Director, EORTC Headquarters EORTC European Organisation for Research and Treatment of Cancer AISBL-IVZW Avenue E. Mounierlaan 83/11 Brussel 1200 Bruxelles België - Belgique Phone: +32 (0) Fax: +32 (0) denis.lacombe@eortc.be Website: Study Coordinator: Emiel Rutgers (Date) The Netherlands Cancer Institute-Antoni Van Leeuwenhoekziekenhuis Plesmanlaan 121 NL 1066 CX Amsterdam The Netherlands Phone: Fax: E.Rutgers@nki.nl PD-004-AF-01 Page 38 of 46 Version: OCT 2009

402 PRC submission document Amendment to EORTC protocol EORTC Investigator: (if applicable) Investigator s name (printed clearly) (Date) PD-004-AF-01 Page 39 of 46 Version AUG 2010

403 PRC submission document Amendment to EORTC protocol EORTC Attachment 2 Table 4 Dose modification of Anthracycline-Regimens for Non-Hematological Toxicities Toxicity Grade Treatment of toxicity Change to chemotherapy Nausea Any Local guidelines antiemetics and/or corticoid Vomiting Any Local guidelines antiemetics and/or corticoid Diarrhea III-IV Stop treatment until grade I, then change dose Mucositis III-IV Stop treatment until grade I, then change dose Dysphagia III-IV Stop treatment until grade I, then change dose Cystitis (If using Cyclophosphamide) I-II III-IV Careful follow-up plus orientation Stop treatment until grade I, then change dose No modification No modification 25% dose reduction (Anthracycline) 1st occurrence- 25% dose reduction (anthracycline or docetaxel for N+ subgroup if applicable) 2nd occurrence discontinue chemotherapy treatment 25% dose reduction (Anthracycline or docetaxel for N+ subgroup if applicable) No change 1st occurrence- 25% dose reduction (cyclophosphamide) 2nd occurrence discontinue chemotherapy treatment PD-004-AF-01 Page 40 of 46 Version AUG 2010

404 PRC submission document Amendment to EORTC protocol EORTC Bilirubin I Wait until within normal limits, then change dose Reduce docetaxel by 25% Alkaline Phosphatase III-IV Wait until Grade II, then change dose ASAT or ALAT III-IV Wait until Grade II, then change dose Alkaline Phosphatase and ASAT or ALAT Reduce docetaxel by 25% Reduce docetaxel by 25% II Change dose Reduce docetaxel by 25% Edema Any No dose change; further treatment at physician s discretion Neurologic II Wait until subsequent cycle; if I, no change If there is no resolution Reduce docetaxel by 25% If persisting after dose reduction, discontinue docetaxel III Discontinue docetaxel Arthralgia/Myalgia Any Symptomatic treatment Skin rash Any Symptomatic treatment Asthenia III Symptomatic treatment 1 st occurrence- reduce docetaxel by 25% 2 nd occurrence reduce docetaxel by 25% Other III Stop treatment until grade I, then change dose Reduce 25% (anthracycline and accompanying drugs or docetaxel for N+ subgroup if applicable) IV Stop treatment until grade I, then change dose 1st occurrence- 25% dose reduction (anthracycline and accompanying drugs or docetaxel for N+ subgroup if applicable) 2nd occurrence discontinue chemotherapy treatment PD-004-AF-01 Page 41 of 46 Version AUG 2010

405 PRC submission document Amendment to EORTC protocol EORTC Attachment 3 Table 7 Dose modification for Hematological Toxicities - Docetaxel-Capecitabine combination Toxicity Treatment of Modification of chemotherapy toxicity ANC < 1.0x10 9 /l on day of infusion (day 1 of cycle) None Wait until ANC > 1.0x10 9 /l (maximum time up to 3 weeks), then use full dose. G-CSF or PEG-Filgrastim may be used in subsequent cycles. Febrile Neutropenia or Neutrophil Counts 0.5x 10 9 /L (grade 4) for more than 7 days. According to standard care in institute Wait until ANC > 1.0x10 9 /l (maximum time up to 3 weeks), then use full dose. G- CSF or PEG-Filgrastim may be used in subsequent cycles. 2 nd episode of Febrile Neutropenia or Neutrophil Counts 0.5x 10 9 /L (grade 4) for more than 7 days. 3 rd episode of Febrile Neutropenia or Neutrophil Counts 0.5x 10 9 /L (grade 4) for more than 7 days. Platelets < 75x10 9 /l on the day of treatment Platelets < 50x10 9 /l on the day of treatment Platelet transfusion According to standard care in institute According to standard care in institute None None Reduce dose of docetaxel and capecitabine by 25% Discontinue chemotherapy treatment Wait until platelet count > 75x10 9 /l and reduce dose of docetaxel and capecitabine by 25% Wait until platelet count > 75x10 9 /l and reduce dose of docetaxel and capecitabine by 25% Wait until platelet count > 75x10 9 /l and reduce dose of docetaxel and capecitabine by 25% PD-004-AF-01 Page 42 of 46 Version AUG 2010

406 PRC submission document Amendment to EORTC protocol EORTC Attachment 4: Table 8 Dose modification of docetaxel-capecitabine for non-hematological toxicities Toxicity Grade Treatment of toxicity Change to chemotherapy Bilirubin I Wait until within normal limits, then change dose Reduce docetaxel by 25% Alkaline Phosphatase III-IV Wait until Grade II, then change dose Reduce docetaxel by 25% ASAT or ALAT III-IV Wait until Grade II, then change dose Reduce docetaxel by 25% Alkaline Phosphatase and ASAT or ALAT II Change dose Reduce docetaxel by 25% Edema Any No dose change; further treatment at physician discretion Neurologic II Wait until subsequent cycle; if I, no change If there is no resolution Reduce docetaxel by 25% If persisting after dose reduction, discontinue docetaxel III Discontinue chemotherapy treatment Arthralgia/Myalgia Any Symptomatic treatment Skin rash Any Symptomatic treatment Asthenia III Symptomatic treatment 1 st occurrence- reduce docetaxel by 25% 2 nd occurrence reduce capecitabine by 25% Nausea Any Local guidelines antiemetics and/or corticoid No modification Vomiting Any Local guidelines antiemetics and/or corticoid No modification Hand-foot syndrome II-III Stop capecitabine until grade I, then change dose 1 st occurrence reduce capecitabine by 25% 2 nd occurrence reduce capecitabine by a further 25% (to 50% of initial dose) 3 rd occurrence - stop docetaxel and capecitabine PD-004-AF-01 Page 43 of 46 Version AUG 2010

407 PRC submission document Amendment to EORTC protocol EORTC Table 8 - Dose modification of docetaxel-capecitabine for Non-Hematological Toxicities Continued Toxicity Grade Treatment Change Diarrhea I None II-III Stop capecitabine until grade I, then change dose and treat diarrhea IV Stop capecitabine until grade I, then change dose and treat diarrhea Mucositis II Stop capecitabine until grade I, then change dose III-IV Stop capecitabine until grade 1, then change dose Dysphagia III Stop capecitabine until grade 1, then change dose 1 st occurrence reduce capecitabine by 25% 2 nd occurrence reduce capecitabine by a further 25% (i.e. to 50% of initial dose) 3 rd occurrence - stop chemotherapy treatment 1 st occurrence reduce capecitabine and docetaxel by 25% 2 nd occurrence stop chemotherapy treatment 1 st occurrence reduce capecitabine by 25% 2 nd occurrence reduce capecitabine by a further 25% (i.e. to 50% of initial dose) 3 rd occurrence - stop chemotherapy treatment 1 st occurrence reduce capecitabine and docetaxel by 25% 2 nd occurrence stop chemotherapy treatment Reduce capecitabine by 25% IV Stop capecitabine until grade 1, then change dose Reduce docetaxel and capecitabine by 25% Creatinine II No treatment; perform a sensitive test to ascertain the real value Stop capecitabine if Cr clearance 30 ml/min; if it increases to 30 ml/min, then resume it Other III Wait until I, then change dose Reduce capecitabine and docetaxel by 25% IV Stop treatment until grade I, then change dose 1 st occurrence- 25% dose reduction 2 nd occurrence stop chemotherapy treatment PD-004-AF-01 Page 44 of 46 Version AUG 2010

408 Attachment 5 study timeline guidelines from diagnosis to start of treatment max 84 days max 56 days max 120 days Signing of spis & IC Patient receives general information Surgeon / research nurse A. B. R I. T1 T2 Surgery* Hospitalization T3a T3b T4 Diagnosis Surgical Oncologist post-op Start post-op op visit visit Treatment C. Cancer Institute Key T= Hospital visit/time point R= randomization PIS-IC = Patient information & Informed Consent D. G. F. E. Discussion Lymph node RNA QC good + status known Proposal for YES/NO Randomizations Feedback 0-3 +ve LN according to risk from patient microarray Chemotherapy and Endocrine therapy H. Signing of PIS & IC 2 or 3 Bring to Center F. during LVEF visit or post to Center Information & signing of PIS & IC 1 (Discordant patients are randomized) A. Tumor diagnosis B. Patient signs screening PIS & IC C. Tumor sample sent to Cancer Institute D. Local pathology, Lymph node status determination E. RNA quantity & quality good, for 0-3 +ve LN PI told patient eligible & microarray analysis performed F. Genomic prognostic test performed, result communicated to PI after signing of PIS & IC 1 G. Patient returns home with R-C / R-E information to decide on participation H. Patient signs PIS & IC 2 or 3 and brings/posts it to Center/signs at the start of Treatment I. Patient inclusion in and randomization for discordant patients * For patients who underwent more than 1 surgery, the timelines start from date of last (definitive) surgery. V 5p PD-004-AF-01 Page 45 of 46