Selection of systemic therapy in Advanced NSCLC based on biological factors. State of the Art

Selection of systemic therapy in Advanced NSCLC based on biological factors State of the Art Giorgio V. Scagliotti University of Torino Department of Clinical & Biological Sciences giorgio.scagliotti@unito.it UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Lung Cancer Four Main Histological Subtypes Squamous Cell Carcinoma Adenocarcinoma Large Cell Carcinoma Small Cell Carcinoma Travis WD et al. WHO Classification of Lung Tumors 2004

Lung Cancer Clinical Distinction for Therapy Non-Small Cell Lung Cancer Squamous Cell Carcinoma Adenocarcinoma Large Cell Carcinoma Small Cell Lung Cancer

Lung Cancer Clinical Distinction for Therapy Non-Small Cell Lung Cancer Similarities in Biological Behaviour & Clinical Outcomes Same Chemotherapy Options Squamous Cell Carcinoma Adenocarcinoma Large Cell Carcinoma Small Cell Lung Cancer

Prognostic Info ormation 180 160 140 120 100 80 60 40 20 0 Relative Contribution of Prognostic Factors in NSCLC 2 4 5 I=Institution; H=Histology; TS=Tumor size WL=Weight loss; ED=Extent of disease; PS=Performance Status 40 Prognostic Factors 70 I H TS WL ED PS WL+ED+PS 100 170 Stanley KE JNCI 1980; 65:25

Efficacy Plateau of Cytotoxic Chemotherapy in NSCLC Study Drugs # Pts Kelly,2001 SWOG 9503 Schiller,2002 ECOG 1594 Scagliotti,2002 ILCP Belani,2002 TAX 326 Vnr/Cis Tax225/Cb 202 208 %, St. IV 88 89 %, ORR 28 25 MST 8 8 %, 1-YS 33 36 Tax135/Cis 292 89 21.3 8.1 31 Gem/Cis 288 86 21 8.1 36 Txt/Cis 293 86 17.3 7.4 31 Tax225/Cb 290 86 15.3 8.3 35 Vnr/Cis Gem/Cis Tax225/Cb Vnr/Cis Txt/Cis TxT/Cb 201 205 201 404 408 402 81 81 82 67 67 67 30 30 32 25 32 24 9.5 9.8 9.9 10.1 11.3 9.4 37 37 43 41 46 38

Scagliotti G et al. J. Thorac. Oncol. 2009;4:1568

Retrospective Analysis of a 3-Arm Randomized Trial Pairwise Comparisons for Survival (P-values) Squamous Adenocarcinoma Large cell Other Squamous (N=187) - 0.0021 0.1607 0.9724 Adenocarcinoma (N=310) - 0.6953 0.0239 Large cell (N=45) - 0.2090 Other (N=65) - Squamous histology was associated with longer overall survival than adenocarcinoma To a lesser extent, other histology was associated with longer survival than adenocarcinoma Scagliotti G et al. J. Thorac. Oncol. 2009;4:1568

Retrospective Analysis by Histology in ECOG 1594 Regardless of histology types, overall survival and progression-free survival were similar in chemo-naive patients treated with standard platin-based doublets involving paclitaxel, docetaxel or gemcitabine Tien H, Dahlberg S, Schiller J, Johnson DJ., J. Thorac. Oncol. 2009; 4 (9 suppl.):s493

Prognostic & Predictive Role of Histology in Advanced NSCLC A literature search of the last 25 years of publications in NSCLC including phase II-III clinical trials, meta-analyses and retrospective reviews revealed : 11 reports found some degree of association between histology and prognosis In 7 reports histology predicted outcomes in patients treated with specific chemotherapeutic agents A prognostic/predictive role of histology was reported in 12 studies with EGFR TKis Hirsch F., Novello S. etal. JTO 2009

Selection Criteria for Treatment Clinical Factors Extent of disease, ECOG PS Age, Comorbidities Smoking Status, Ethnicity Histological Features Molecular Profile

Critical Issues with Lung Cancer Histology Relevanceofhistopathologicalsubtyping of lung cancer Subtyping problems in small samples (biopsies or FNA cytology) Role of immunohistochemical markers Tissueidentificationof prognosticand predictive factors(potentially useful for selecting therapy)

Histological Subtype of NSCLC and Adenocarcinoma Chemotherapy Selection Squamous cell carcinoma Pemetrexed (non-squamous) UFT (adenocarcinoma) EGFR TKIs (?papillary & BAC ) IGFR inhibitors Bevacizumab contraindicated

EGFR-TKIEfficacy in Selected Patient Populations Clinical factors: never smoker; adenocarcinoma; female sex Number of clinical factors ORR (%) MST (months) 3 56 14+ 2 30 12 1 9 5 0 3 3 Miller V, et al. J Clin Oncol 2004

Large Squam Small 2010: Lung Adenoca- Multiple Molecular Subsets Adeno ALK fusion ROS fusion BRAF PIK3CA PDGFR amp MEK1 HER2 KRAS EGFR Unknown

Large 2010: Never Smoker Lung Adenoca - Almost All Molecular Subsets Defined! Squam Small Adeno HER2 ALK fusion EGFR KRAS Pham et al 06; Stephens et al 04; Shaw et al 09 Riely et al 08 Unknown

20% NSCLC Oncogene Addicted Advanced NSCLC A New Treatment Paradigm NSCLC UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Impact of EGFR and k-rasmutations on Survival of Patients Treated with EGFR-TKis Jackman DM et al. Clin. Cancer Res. 2009; 15:5267

Impact of EGFR and k-ras Mutations on Survival of Patients Treated with EGFR-TKis Jackman DM et al. Clin. Cancer Res. 2009; 15:5267

EGFR-TKIs and EGFR Mutation -Directed Front - Line Studies Study Entry Criteria HR for PFS (EGFR mut+) IPASS Mok NEJM 2009 First SIGNAL Proc. IASLC 2009 WEJGSG002 Kobayashi ASCO 2009 Asiatic, never-& light smokers, adenocarcinoma (EGFR mut+ 59.7%) Adenocarcinoma, Neversmokers (EGFR mut+ 44%) 0.48 (0.36-0.66) 0.61 (0.30-1.22) EGFR Mutation + (all) 0.35 (0.25-0.50) HRforOS (EGFR mut+) 0.91* (0.76-1.10) *overall population 0.82 (0.35-1.92) EURTAC (EU) EGFR Mutation + (all)?? OPTIMAL (China) EGFR Mutation + (all) 0.16 (0.10-0.26) NA?

GefitinibvsCarbo/Pac in NEJSG Trial PFS and Overall Survival Maemondo M et al. N Engl J Med 2010;362:2380-2388 UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Maximum change in tumor size (%) Tumor Responses to Crizotinib for Patients with ALK-positive NSCLC 60 40 20 0 20 40 60 80 30% Progressive disease Stable disease Confirmed partial response Confirmed complete response 100 *Partial response patients with 100% change have non-target disease present * Bang Y et. Al. Proc. ASCO 2010

Lung Cancer Mutational Consortium 1000 patients Stage IV ECOG PS 0-2 Lung Adenocarcinoma Sufficient Tissue Informed Consent Mutational Analysis CLIA-Certified lab at LCMC site: KRAS, EGFR, EML4-ALK, BRAF, HER2, PIK3CA, NRAS, AKT1, MET amplification Central Confirmation of Adenocarcinoma Diagnosis (1 slide) Report to LCMC Virtual Database Report to Physician Use Data to Select Therapy (Erlotinib with EGFR mutation) Recommend Clinical Trial of Agent Specific for Target

20% NSCLC Oncogene Addicted Advanced NSCLC A New Treatment Paradigm JMDB HR 0.81 cisplatin- pemetrexed NSCLC non-squamous subset beva-eligible ECOG HR 0.80 AVAiL HR 0.98 doublet + bevacizumab *Beva-eligible: non-squamous, no gr 2 haemoptysis, no invasion of major vessels, no cavitation, no uncontrolled hypertension, no recent history of thrombosis, no hemorrhagic disorders, no recent anticoagulation squamous platinum doublet

ECOG 4599: Survival by Histology Subtype Baseline Characteristics Total N PC (n = 444) Median N Months PCB (n = 434) Median N Months Hazard Ratio 95% CI All patients 878 444 10.3 434 12.3 0.80 0.69-0.93 Histologic type - Adenocarcinoma 602 302 10.3 300 14.2 0.69 0.58-0.83 - Large Cell 48 30 8.7 18 10.0 1.15 0.60-2.24 - Squamous 3 2 12.3 1 22.4 0.00 0.00- - BAC 23 11 17.7 12 10.0 1.48 0.57-3.69 - NSCLC,NOS 165 86 10.0 79 9.5 1.16 0.84-1.61 - Other 34 11 12.6 23 8.4 0.92 0.43-1.98 Sandler A. et al. J Thorac Oncol. 2008;3(11 Suppl 4): Abstract 133.

Cis/Pem vs. Cis/Gem in Advanced NSCLC Scagliotti GV et al. J. Clin. Oncol 2008; 26:3543 UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Survival without grade 3-4 drug related toxicity for patients with non squamous histology Scagliotti G. et al. Eur. J. Cancer 2009; 45: 2298

Efficacy by Histology in Pemetrexed Studies NSCLC Histologic Group Second-line Pem vs. Docetaxel First-line Pem/Cis vs. Gem/Cis Maintenance Pem vs. Placebo Pem Doc Cis/Pem Cis/Gem Pem Placebo Non-squamous n=205 n=194 n=618 n=634 n=325 n=156 Median OS, months 9.3 8.0 11.0 10.1 15.5 10.3 Adjusted HR (95% CI) P value 0.78 (0.61 1.00) 0.048 0.84 (0.74 0.96) 0.011 0.70 (0.56 0.88) 0.002 Squamous n=78 n=94 n=244 n=229 n=116 n=66 Median OS, months 6.2 7.4 9.4 10.8 9.9 10.8 Adjusted HR (95% CI) P value 1.56 (1.08 2.26) 0.018 1.23 (1.00 1.51) 0.050 Non-squamous = adenocarcinoma, large cell carcinoma, and other/indeterminate NSCLC histology 1.07 (0.77 1.50) 0.678 Scagliotti GV et al. Oncologist 2009

CLINICAL & BIOLOGICAL SCIENCES Thymidilate Synthase Expression in Normal Lung Tissue & Lung Cancer Snap Frozen Tissues FFPE Tissues levels UNIVERSTY OF TORINO DEPT. OF Significantly Higher in Lung Cancer than in normal lung tissue TS mrna Significantly Higher in Squamous Cell Carcinoma of the Lung Ceppi P. et al. Cancer 2006

Cis/Pem vs. Cis/Pem : Subgroup Analyses Scagliotti GV et al. JCO 2008; 26:3543 UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Meta-Analysis of Postoperative Adjuvant CT with Tegafur-Uracil in NSCLC Hamada C. et al. JCO 2005; 23:4999 UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Biopsy Techniques in Lung Cancer Diagnosis Sputum cytology Bronchial brushings and washings Fluids FNA cytology primary or mets Transbronchial biopsy Bronchial biopsy Core biopsy primary or mets Liver biopsy Mediastinoscopy Lymph node excision VATS biopsy / resection Thoracotomy & tumour excision Increase in Cell number and Tissue architecture Kerr K., 2008

The norm Operates at all levels Morphology Protein expression mrna Gene copy number Mutations Sample Heterogeneity Differences between primary tumour and metastases? Small biopsy sample vs the tumour overall? Makes scoring and interpretation difficult Sampling becomes important

Immunohistochemistry of Lung Cancer IHC is the most rapid (1 day) and less expensive method (2-8 euro x reaction) to display cell differentiation when morphology cannot distinguish clear-cut criteria Poorly-differentiated NSCLC??? IHC TTF-1 + Poorly-differentiated adenocarcinoma

Can Immunohistochemistry Help to Shed TTF-1 Light on NSCLC -NOS? Surfactant Apoproteins (A & B) NapsinA CK 7 ADENOCARCINOMA p63 HMWCK (CK5-6, 34βE12) Desmocollin 3 SQUAMOUS CELL CARCINOMA

NSCLC on H&E SQC at IHC NSCLC on H&E ADC at IHC H&E TTF-1 p63 UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Diagnostic Algorithm for Advanced Lung Cancer Metastatic Identify Malignancy Mucin stains P63 & CK5/6 TTF1 SCLC Primary Subtype prediction 83% accurate Overall only 7% cases not typed Not SCLC Squamous Adenocarcinoma NSCLC-NOS Rarer Subtype? 25-40% of cases Probably Squamous Probably Adenoca IHC not predictive

Is There Enough Material on Which To Perform These Studies? Morphological diagnosis Diagnostic IHC Predictive marker IHC mrna Mutational analysis Tissue biopsy Aspiration cytology Fluid cytology? % tumour in bronchial biopsy samples In malignant bronchial biopsy samples 33-50% of fragments do not contain tumour Coghlin CL et al, JTO 2010, 5:448-452

Predictive Molecular Markers for Response to Chemotherapy in NSCLC Gene Abnormality Drug Response p53 Mutation Multiple KRAS Mutation Platinum β-tubulin Increased isotype 3 Taxanes RRM1 Increased expression Gemcitabine ERCC1 Increased expression Platinum BRCA1 Increased expression Platinum Thymidylate synthase Increased expression Antifolates EGFR mutation Present Platinum

Cumulative survival Cumulative survival ERCC1 and RRM1 Gene Expression in Advanced NSCLC Treated with Cisplatin and Gemcitabine 1,0 0,8 0,6 0,4 0,2 0 1,0 0,8 0,6 0,4 0,2 ERCC1 < 4 4 p=0.0032 0 10 20 30 40 50 60 70 Time (months) Low expression in both genes Others p=0.0023 0 0 10 20 30 40 50 60 70 Time (months) Concomitant Low Expression of ERCC1 and RRM1 Cumulative survival 1,0 0,8 0,6 0,4 0,2 0 1,0 0,8 0,6 0,4 0,2 RRM1 < 7.5 7.5 p=0.0390 0 10 20 30 40 50 60 70 Time (months) At least one gene with low expression Others p=0.0216 0 0 10 20 30 40 50 60 70 Time (months) Low Expression in At Least One Gene Ceppi P et al. Ann. Oncol 2006;17:1818 25

Gene Expression Modulation in 64 Cases of ΔΔCt 7.0 6.6 6.2 5.8 5.4 5.0 4.6 4.2 3.8 3.4 3.0 2.6 2.2 1.8 1.4 1.0 0.6 0.2 0.2 0.6 1.0 1.4 1.8 2.2 2.6 3.0 Resected NSCLC: DNA Repair Genes DSB-repair Direct BER NER-repair PR-repair DNA-replication Telomere repair repair maintenance Saviozzi S et al. Cancer Res 2009;69:3390 6

Molecular Predictive and Prognostic Markers in NSCLC Good prognosis High ERCC1 (qrt-pcr) High RRM1 (Aqua-IHC) KRASwild type EGFR mutation Poor prognosis High BRCA1 (qrt-pcr) High p53 (IHC) High beta-tubulin (IHC) TP53 mutation KRAS mutation? High EGFRcopy number Coate LE et al. Lancet Oncology 2009; 10: 1001-1010

Classification of Human Lung Carcinomas by mrna Expression Profiling Reveals Distinct Adenocarcinoma Subclasses SCLC Very High TS ; Squamous Intermediate TS, Adenocarcinoma Low TS Bhattacharjee A et al. PNAS 2001; 98:13790

TS mrna Expression in Lung Cancer (n=146) Monica V. et al. Clin. Cancer Res. 2009; 15:7547 UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

Phase III Study ED-SCLC of Pemetrexed/ Carboplatin vsetoposide/ Carboplatin Socinski MA. JCO 2009;27:4787 UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES

TS Immunoreactivity in Cytological NSCLC-NOS Cytological sample Histological sample UNIVERSTY OF TORINO DEPT. OF CLINICAL & BIOLOGICAL SCIENCES TS negative TS positive

CLINICAL & BIOLOGICAL SCIENCES UNIVERSTY OF TORINO DEPT. OF Phase II Study of Preoperative Cis/Pem in Stage IIIa N2 non-squamous NSCLC N=33 Stage IIIa N2 Cis75 mg/ m2 & Restaging by by Med (+CT Pem500 mg/m2 CT scan and Scan + PET) x 3 q3wks PET Biomarker Assessment TS, TTF-1, E2F1, ERCC1, BRCA1 (Functional Imaging Changes) Primary end point : ORR Secondary end points :pcr,pfs,os, tumor downstaging, toxicity Surgery

Gene Expression According to Histology Squamous Cell Carcinoma Median(range) Adenocarcinoma Median(range) P value ERCC1 1.41 (0.45-7.34) 0.72 (0.23-2.45) 0.0001 MZF1 0.62 (0.06-6.72) 0.25(0.03-1.49) 0.0001 Twist 10.37 (0.30-76.01) 2.50 (0.14-19.16) 0.0001 RRM1 2 (0.6-6.9) 1.2 (0.4-2.9) 0.0001 TRX 2.13 (0.40-11.88) 0.91 (0.31-7.94) 0.0001 Tpd1 1.7 (0.6-7.3) 1.3 (0.1-2.6) 0.02 NFAT 0.4 (0.1-2.3) 0.5 (0.1-1.8) 0.65 BRCA1 4.26 (0.55-18.48) 1.50 (0.09-8.08) 0.0001 BubR1 16.3 (1.4-90) 7 (0.8-25) 0.0001 Rosell R. et al. PLoS ONE 2:e1129

Are Other Genomic Markers Differentially Expressed in NSCLC? Author Squamous Adenocarcinoma P Value Olauseen, NEJM 2006 ERRC1 Positive * 70% 45% ERCC1 Negative * 21% 40% Zheng, NEJM 2007 Median ERCC1** 56.8 (1.9-178.7) Median RRM1 ** 62.3 (13.2-96.2) * H Score (semiquantitative IHC) ; ** AQUA Scores 68.0 (6.6-153.1) 40.5 (8.3-95.1) 0.001

Five Gene Signature and Outcome in NSCLC High-risk Low-risk P value Original Cohort (n=101) Adenocarcinoma 61% 36% 0.03 Squamous Cell Ca. 32% 47% Others 7% 17% Validation Cohort (n=60) Adenocarcinoma 32% 50% 0.19 Squamous Cell Ca. 59% 42% Others 9% 8% Chen HY et al. NEJM 2007; 356:11

Conclusion WHO classification should remain the common language for exchanging information and treatment decision. There is no apparent prognostic role of histology with modern cytotoxic chemotherapy. Preliminary evidence indicate a prognostic role for some molecular markers. Histologic subtyping should be taken into account in making treatment decisions for toxicity and efficacy issues. IHC may help in separating squamous from non squamous histology also in limited cytological samples. Mandatory need of histology (pharmacogenomic) -driven prospective trials in any stage of NSCLC.