Case studies of the development of targeted therapies for cancer: 1. BRAF inhibitors in melanoma 2. Autophagy Inhibitors

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1 Case studies of the development of targeted therapies for cancer: 1. BRAF inhibitors in melanoma 2. Autophagy Inhibitors Ravi Amaravadi, MD Assistant Professor of Medicine Division of Hematology-Oncology Department of Medicine Abramson Cancer Center Perelman School of Medicine University of Pennsylvania Disclosures Laboratory Grant Funding NCI Abramson Cancer Center Penn-Pfizer Alliance Millenium Pharmaceuticals Tetralogics Pharmaceuticals Industry Support for clinical trials Advisory Board/Speakers Fee Pfizer Tetralogics Millenium Novartis Roche Roche Merck Bayer Millenium (Takeda) Genentech Patent Pending US 61/177,697 Combination antineoplastic therapy with mtor and autophagy inhibitors US 61/486,641 Novel autophagy inhibitors for cancer therapy 1

2 Case 1: BRAF inhibitors for metastatic melanoma Treatment of Metastatic Melanoma: No standard 1 st Line therapy identified Interferon Interleukin-2 Cisplatin, Vincristine, Dacarbazine Temozolomide High dose temozolomide Tamoxifen v. Dacarbazine Median Survival: 8 months Median Survival Brain Metastases: 3 months 2

3 2010 NCCN guidelines for first line treatment of Stage IV melanoma Clinical trial preferred Or Interleukin 2 Or Dacarbazine Or Temozolomide ± cisplatin and vinblastine 3

4 Ibrahim and Haluska Ann rev. pathology mech. Disease 2009 Suppression of BRAF induces cell death in BRAF mutant cell lines Karasarides et al. Oncogene (2004) 23,

5 Sorafenib Cl CF 3 N H O N H O N O N H Kinase assays IC 50 C-Raf 2 nm mvegfr2, VEGFR3 wt B-Raf, V599E B- Raf 6-10 nm nm p38, PDGFrβ nm FLT-3, c-kit EGFR, PKC, MEK, ERK nm Inactive at 10 mm Wan PT et al. Cell Mar 19;116(6): Sorafenib in Melanoma 2002: phase II trial of sorafenib in melanoma: 2% response rate. 2002: phase I/II trial of sorafenib + carboplatin and paclitaxel 105 patients: 27% response rate/8 month PFS 2005: Sorafenib approved for renal cell ca 2007 Sorafenib approved for HCC : 2009: 2 large randomized phase III trials (n=>1000 patients) of sorafenib + carbo/taxol versus carbo/taxol found absolutely no difference in response rate, or survival with the addition of sorafenib to chemotherapy 5

6 Could targeting BRAF be too simplistic or even misguided? 33,345 mutations were found in the melanoma genome compared to the normal genome from the same patient. 187 mutations were predicted to alter the known function of specific proteins. Nature 463, (14 January 2010) doi: /nature08658; Received 30 July 2009; Accepted 4 November 2009; Published online 16 December 2009; Corrected 14 January 2010 A comprehensive catalogue of somatic mutations from a human cancer genome Erin D. Pleasance 1,8 et al. 6

7 Pharmacodynamics: Does the drug hit its target in patient tumors? Pre-treatment Post-treatment Sorafenib: No PLX4032: Yes 7

8 Phase I Results of PLX4032 in patients with BRAF mutant melanoma Dose and schedule 960 mg bid maximal tolerated dose and phase II dose Safety Common side effects include a rash, squamous cell carcinoma, and fatigue. Response Definition No % Complete response No evidence of disease 1 4 Partial Response Stable Disease Progressive Disease >30% tumor shrinkage from baseline <30% shrinkage and <20% growth from baseline >20% growth from baseline or new tumors (all had shrinkage) Total Day 0 V600E+ melanoma patient PET scan at baseline and day +15 after PLX4032 treatment at 720 mg BID Day 15 Flaherty, ASCO

9 1st interim analysis found that Vemurafenib reduces the risk of progression by 74% and the risk of dying by 63% compared to chemotherapy Chapman et al NEJM 2011 The rapid success and approval of vemurafenib in BRAF mutant melanoma depended on the following: 1) The molecular target was identified and validated in preclinical models 2) The target is present in a large fraction of patients 3) Lessons learned from the failure of the first generation Raf inhibitor sorafenib were used by persistent investigators 4) A more potent and specific second generation inhibitor was found 5) Phase I testing with PK and PD analysis was performed with rigor 6) The phase I dose was increased to the maximal tolerated dose 7) The drug was tested in a biomarker-restricted population (BRAF mutant melanoma) 8) An OPEN collaboration between multiple academic investigators, Pharma teams, and the FDA 9) LUCK 9

10 As Presented at the NCCS Cancer Policy Roundtable Case 2: Autophagy a new therapeutic target in cancer Autophagy, the cell s garbage disposal and built in recycling plant gets rid of waste and feeds the cancer cell when it is faced with stresses. Damaged cell component Autophagic Vesicle Recycled building blocks Lysosome 10

11 Chloroquine derivatives impair the lysosome, blocking the last step of autophagy Autophagy Inhibition augments cancer cell death in the context of Nutrient Limitation Lum et al. Cell 2005 Growth Factor Limitation Lum et al. Cell 2005 P53 activation or Alkylating chemotherapy Amaravadi et al. JCI 2007 Tumorigenesis McLean et al. JCI 2008 HDAC inhibitor Carew et al. Blood 2007 Dormant tumor cells Luo et al. JCI 2008 Akt inhibitor Degtyarev M (Genentech) J Cell Bio 2008 Imatinib Bellodi JCI 2009 Velcade Ding et al. MBC

12 Could chloroquine (CQ) or hydroxychloroquine (HCQ) be used to treat cancer? Chloroquine derivatives used in humans since 1940 s Used for decades for malaria, rheumatoid arthritis Extensive preclinical testing, hundreds of thousands of patients worth of safety data Hydroxychloroquine is a safer drug than chloroquine at high doses Generic drug with no pharmaceutical company interested in marketing for cancer indications Excellent brain penetration Drug Monthly Cost Erbitux $ 9600 Avastin $ Sorafenib $ 5614 Gleevec $ 3816 Herceptin $ 3195 Tarceva $ 2697 Hydroxychloroquine $

13 Other investigators have already tested chloroquine for cancer Randomized phase III trial of RT/carmustine +/- low dose CQ in newly diagnosed Glioma (Brain Tumors) Sotelo, et al. Ann Int. Med Kaplan Meier Analysis of Overall Survival in GBM patients treated on the phase III trial of CQ/carmustine/RT v. placebo/carmustine/rt ( p>0.05) HCQ Trial #1: ABTC 0603: A phase I/II trial of hydroxychloroquine with radiation therapy and concomitant and adjuvant temozolomide in patients with newly diagnosed GBM WK RT TMZ Initiation Cycle Radiation 59.4 Gy/33 fractions Temozolomide 75 mg/m2 po qd PI: Myrna Rosenfeld, MD, PhD Sample Size: Phase I: 6-18 Phase II: 76 80% power to detect a 40% v. 26% 2-year overall survival rate. HCQ Hydroxychloroquine 200 mg po qd MTD Maintenance Cycle: Setting ABTC (15 centers) DAY TMZ HCQ TMZ 150 mg/m 2 po d1-5 HCQ 200 mg po qd MTD 13

14 Condition Intervention Phase Sponsors Collaborators Multiple myeloma HCQ + Bortezomib I/II UPenn, Millenium Brain, central nervous system tumors Adult solid tumors HCQ+ Temozolomide/RT HCQ+ temozolomide I/II I UPenn,,CTEP, NCI UPenn, Schering Adult solid tumors HCQ+ temsirolimus I UPenn, Wyeth Adult Solid Tumors HCQ + Vorinostat Prostate cancer Docetaxel hydroxychloroquine I San Antonio, NCI, Merck ClinicalTrials.go v Identifier NCT NCT NCT NCT NCT Title A Phase I/II Trial of HCQAdded to Bortezomib for Relapsed/Refractory Myeloma A Phase I/II Trial of HCQ in Conjunction with Radiation Therapy and Concurrent and Adjuvant Temozolomide in Patients With Newly Diagnosed Glioblastoma Multiforme A Phase I Study of HCQ in Combination with temozolomide in Patients with Advanced Solid Tumors A Phase I Study of HCQ in Combination with temsirolimus in Patients with Advanced Solid Tumors A phase I pharmacokinetic and Pharmacodynamic Study of Hydroxychloroquine in combination with vorinostat for the treatment of patients with advanced solid tumors II CINJ, NCI NCT A Phase II Study of Docetaxel and Modulation of Autophagy with HCQ for Metastatic Hormone Refractory Prostate Cancer Prostate cancer HCQ II CINJ, NCI NCT Autophagic Cell Death in Patients with Hormone-Dependent Prostate-Specific Antigen Progression after Local Therapy for Prostate Cancer Breast cancer HCQ+ ixabepilone I/II CINJ, NCI NCT Phase I/II Study of Ixabepilone in Combination with the Autophagy Inhibitor HCQ for the Treatment of Patients with Metastatic Breast Cancer Lung cancer HCQ+Bevacizumab carboplatin paclitaxel Advanced cancer HCQ + sunitinib Non-small cell lung cancer HCQ + Gefitinib I/II Singapore, MGH B-cell chronic lymphocytic leukemia Pancreas Cancer HCQ II North Shore Long Island Jewish HCQ+ Gemcitabine I/II CINJ, NCI NCT Modulation of Autophagy with HCQ in Combination with Carboplatin, Paclitaxel and Bevacizumab in Patients with Advanced/Recurrent Non-Small Cell Lung Cancer -A Phase I/II Study I CINJ, NCI NCT Anti-Angiogenic Therapy in Patients with Advanced Malignancies: A Phase I Trial of Sunitinib and HCQ NCT NCT A Phase II with a Lead in Phase I Study to Examine the Tolerability, Safety Profile and Efficacy of HCQ and Gefitinib in Advanced Non- Small Cell Lung Cancer Autophagic Modulation with Phase II Study to Evaluate the Tolerability and Efficacy of Treatment of Previously Untreated B- Cell Chronic Lymphocytic Leukemia (B-CLL) Patients with HCQ I U. Pittsburgh Pending Phase I/II study of Preoperative Gemcitabine in Combination with Oral HCQ in Subjects With Resectable Stage IIb Or III Pancreatic Adenocarcinoma Renal Cancer HCQ IA U. Pittsburgh Pending Neoadjuvant Study of Preoperative HCQ in Patients with Resectable Renal Cell Carcinoma A PBMC samples Baseline 10 Week 3 9 Week * * * * Patient HCQ (Mg/Day) C Pre-treatment 3 wks 800 mg HCQ /TMZ/RT Autophagic vesicles/cell B * Actin P 3W LC3 I LC3 II 2 µm 14

15 A B HCQ concentration (ng/ml) C Estimated peak HCQ concentration (ng/ml) HCQ concentration (ng/ml) HCQ sulfate daily dose (mg) Significant AV accumulation Time on study (hrs) Figure 5. Whole blood hydroxychloroquine (HCQ) concentrations and pharmacodynamic response. (A) Range of HCQ concentrations demonstrates a proportional dose-concentration relationship. (B) HCQ concentrations are maintained over the course of months. (C) Patients with significant AV accumulation had significantly higher estimated peak HCQ concentrations; *p= (A-C) Patients/samples available: 14/45. Population pharmacokinetic modeling indicated data are best described using a onecompartment model with first-order elimination. Encouraging activity of autophagy inhibition combined with mtor inhibition Phase I trial of temsirolimus + HCQ completed dose escalation 27 pts Dose expansion in melanoma patients Safe and tolerable regimen 65% & 73% stable disease rate in solid tumors and melanoma Temsirolimus as a single agent has a 0% stable disease rate in melanoma MVP mean: MVP mean: MVP mean: Baseline Temsirolimus Tem + HCQ Presented at AACR 2011 and SMR

16 Designing a more potent autophagy inhibitor Lys01/Lys05 (water soluble) are novel potent autophagy inhibitors designed by employing first principles of chemistry. Patent was filed April 2011 A few pharmaceutical companies have expressed initial interest Jeffrey Winkler, PhD Merriam Professor of Organic Chemistry Department of Chemistry School of the Arts and Sciences University of Pennsylvania Within 4 hours of treatment Lys05 treatment results in an increased size and number of accumulated autophagic vesicles compared to HCQ PBS HCQ 10 µm Lys05 10 µm 16

17 In vivo autophagy inhibition and antitumor efficacy of Lys05 in 2 different melanoma xenograft mouse models Identifying the molecular target of lysosomal Autophagy inhibitors Step 1:Subtractive chemical proteomics for target identification Active autophagy inhibitor 1 Lys Inactive autophagy inhibitor 1 Lys

18 A biomarker of sensitivity to autophagy inhibition has been identified Autophagy inhibitor Biomarker + - Figure 1. An autophagy inhibitor biomarker Predicts sensitivity to HCQ Figure 2. IHC and serum DNA methylation Assays can detect the candidate biomarker In patient samples Can autophagy inhibitors help cancer patients? 1) Molecular target identified and validated in preclinical models? Not fully- we need to get more specific 2) Is autophagy and a biomarker of sensitivity to autophagy inhibitors present in a large fraction of patients Yes 3) Are first generation autophagy inhibitor trials up and running? Yes 4) Can we find a more potent and specific drug? Yes-early stages 5) Does phase I testing include PK and PD analysis? Yes 6) Are we pushing the dose to the maximal tolerated dose? Not fully done 7) Are we testing the drug in a biomarker-restricted population? Not yet- early stages of biomarker development and validation 8) Is there an OPEN collaboration between multiple academic investigators, Pharma teams, and the FDA? Emerging interest in pharmaceutical companies, but may need to be championed by other causes. 9) LUCK 18

19 Take Home Messages The successful development of vemurafenib for BRAF mutant melanoma can serve as a model For development of other targeted therapies for cancer. Autophagy is an emerging target in cancer that if properly studied could yield multiple drugs that benefit a large subfraction of cancer patients Thank you to all of the patients and families who have participated in clinical research 19