M. J. Pishvaian* , R. Slack, A. Witkiewicz + , A. R. He,

A Phase II study of ABT-888 + temozolomidein patients with heavily pretreated, metastatic colorectal cancer M. J. Pishvaian*, R. Slack*, A. Witkiewicz+, A. R. He*, J. J. Hwang*, A. Hankin*, K. Dorsch-Vogel*, D. Kuda*, T. McAndrew*, L. M. Weiner*, J. Marshall*, J. R. Brody+ *Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC +Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA

Financial Disclosures • This clinical trial is funded by the • Otto J. Ruesch Center for the Cure of GI Cancers, Lombardi Comprehensive Cancer Center. • Abbott Inc. has provided research funding for a portion of the correlative science • I have no personal financial disclosures Georgetown Lombardi

Mechanisms of DNA Repair: PARP (Poly(ADP-ribose) polymerase) Environmental factors (UV, radiation, chemicals) Chemotherapy (e.g. alkylating agents) Normal physiology (DNA replication, ROS) Radiotherapy PNK 1 XRCC1 pol β LigIII PARP DNA DAMAGE • PARP • Critical DNA repair enzyme (SSB, BER) • Often overexpressed in cancer cells • Confers resistance to chemotherapy and radiation • Inhibition of PARP • Prevents recruitment of DNA repair enzymes • Leads to failure of single strand break repair • Unrepaired break site  replication fork arrest • Leads to degeneration into double-strand breaks • Ultimately  chromosomal catastrophe cell death Cell Death Georgetown Lombardi Tutt, A, et al, JCO 27:18s, 2009 (suppl; abstr CRA501) Helleday T, et al. Nat Rev Cancer. 2008;8:193-204

Background • ABT-888 • Oral PARP-1, 2 inhibitor • Proven PARP inhibition in vitro, and in vivo • Potentiates activity of multiple chemotherapies in • pre-clinical models including temozolomide • Temozolomide (TMZ) • Oral potent atypical alkylating agent • Ongoing trials in combination with ABT-888 Donawho, CK, et al, Clin Cancer Res 2007;13(9) May1, 2007 Palma, JP, et al, Clin Cancer Res 2009;15(23):7277–90 Kummar, S, et al, JCO. 2009 Jun 1;27(16):2705-11 Delaney, CA, et al, Clin Cancer Res, 6: 2860-2867, 2000 Raymond, E, et al, Clin Cancer Res, 3: 1769-1774, 1997. Georgetown Lombardi

PARP is a Promising Target in CRC DMSO TMZ ABT-888 TMZ + ABT-888 TMZ + ABT-888 • Overexpression of PARP in CRC • leads to chemotherapy resistance • ABT-888 + temozolomide  cell death • Many CRCs may be exquisitely sensitive to PARP inhibitor-based therapy • 5-7% with mismatch repair gene defects (dMMR) • Up to 40% PTEN deficient • (defect in homologous recombination) 100000 *** 75000 50000 Cell Number *** 25000 0 SW480 HCT116 Loupakis F, et al, JCO 27:2622-9, 2009 McCabe N, et al, Can Res 66:8109-15, 2006 Liu, X., et al, Mol Cancer Res, 7: 1686-1692, 2009 Horton, TM, et al, Mol Cancer Ther, 8: 2232-2242, 2009 Georgetown Lombardi

Metastatic CRC Measurable or evaluable disease Adequate hepatic, bone marrow, and renal function Age ≥ 18 years ECOG performance status 0-2 Progression on or ineligible for all standard therapies: Fluoropyrimidine Oxaliplatin Irinotecan Bevacizumab Cetuximab/Panitumumab Inclusion/Exclusion Criteria • Inclusion Criteria • Exclusion Criteria • Untreated CNS metastases • Active severe infection • Active cardiovascular disease • Women who were pregnant or breastfeeding • Anticipated patient survival under 3 months Georgetown Lombardi

Trial Design- Single Arm, Phase II 8 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 III III III III III II II TMZ - 150mg/m2 QD ABT-888 - 40 mg BID 4 week cycles 4 week cycles ABT-888 ABT-888 A T A T TMZ TMZ 16 12 Weeks on study Restaging studies every 8 weeks Days of the week Georgetown Lombardi

Trial Design • Primary endpoint - Disease control rate (DCR) • Complete response, partial response, or stable disease (after two cycles) • Secondary endpoints • Objective response rate, progression free survival, overall survival • Correlation between DCR and MMR and PTEN expression • Simon’s two stage optimal design • P0 = 10%, P1 = 25%, a=10%, b=10% (Power = 90%) • Stage I = 21 patients (3/21 = 14% respond, then proceed to Stage II) • Stage II = 29 additional patients (50 total) • If DCR ≥ 8/50 = 16%  Further study justified Georgetown Lombardi

Results - Patients • 09/09 to 06/11, 49 patients enrolled • 51% KRAS mutant • Median Age – 55 years • Range 36 to 72 • 29 Male, 21 Female • Median ECOG PS – 1 • 0 n=15 • 1 n=32 • 2 n=2 • Median number of prior • chemotherapy regimens - 3.5 • Range 2 to 7 Prior Therapies 100 90 80 70 60 Percent of Patients 50 40 30 20 10 0 Other Irinotecan Oxaliplatin Bevacizumab Fluoropyrimidine Cetux/Panitumumab Georgetown Lombardi

Adverse Events • Only 1 patient withdrew due to toxicity - pancytopenia Georgetown Lombardi

Evidence of Anti-Cancer Activity 100 80 60 Percent Surviving 40 20 0 0 5 10 15 20 Months • As of 06-01-2011 - ITT analysis of 49 patients • Disease Control • 11 (2 confirmed PRs) • DCR = 22% • Median Duration of Disease Control • 22 weeks • Range 15 – 40 weeks • Median overall survival • 6.3 months Georgetown Lombardi

Evidence of Anti-Cancer Activity 40 Duration of Disease Control 30 = Disease Control (22%) = Partial Response (4%) = Progression (78%) Weeks 20 10 0 Individual Patients Georgetown Lombardi

Patient 26 – 69 year old female April, 2010 34% reduction 34 wks on study August, 2010 Georgetown Lombardi

Correlative Studies • Good Tissue Acquisition • 45/49 archived surgical specimens evaluable • 7/9 fresh serial tumor evaluable • Pre-treatment and Day 8 • Fixed AND frozen samples • Mismatch repair protein expression • Increased DCR in dMMR tumors • PTEN expression • Increased DCR in PTEN-deficient tumors Georgetown Lombardi

Correlative Studies - MMR • Assessed MMR status by IHC on paraffin samples • MLH-1, MSH-2, PMS-2, and MSH-6 • 35 of 49 samples assessed to date • No MMR defects detected in any of the patients • Can not assess for any association between MMR status and DCR Georgetown Lombardi Lindor, et al, JCO, 20: 1043-1048, 2002. Wright, CL, et al, Am J Surg Pathol 2003;27:1393–1406

Cancer Normal Correlative Studies - PTEN • Assessed PTEN protein expression by IHC • To date, no clear relationship between expression and DCR PTEN Positive CRC N=13 N=5 Cancer PTENNegative CRC H-score = (%1+cellsX1) + (%2+cellsX2) + (%3+cellsX3) Georgetown Lombardi Loupakis, F, et al, JCO 2009, 27:2622-2629

Correlative Studies PTEN MMR MGMT Methylation Investigational Markers DNA Repair SNPs Serum microRNA DNA Repair Expression Panel ? Pre-clinically Supported Markers No Apparent Correlation ? ? dMMR Patients Needed Results Pending Patient Tumor Samples Georgetown Lombardi

Correlative Studies - DNA Repair Expression Panel • RNA from tumor samples • Mini-DNA array • 90 DNA repair genes • Pre-treatment vs. Day 8 • Expand analysis • Predictive markers • Responsive signature • Resistance markers Georgetown Lombardi

Conclusions • ABT-888 plus TMZ is a well tolerated oral combination • There is evidence of anti-cancer activity in refractory colon cancer patients: • Partial response • Prolonged stable disease • Anti-cancer activity even in MMR-proficient patients • Correlative studies for predictive subgroups are pending • Good tissue acquisition rate Georgetown Lombardi

Study Expansion 20 Patient – dMMR Cohort Pre-clinically selected Predictive marker Primary Cohort (50 Patients) Enrollment completed 20 Patient – High dose TMZ Cohort Increased TMZ dose Greater DNA damage Mandatory “Fresh” pre-tx biopsies • Evaluate efficacy in MMR-deficient subgroup • Identify predictive markers of response • Aim to initiate phase II trial in selected subgroups Georgetown Lombardi

Biostatistician Rebecca Slack, MS Clinical Care and CRMO John Marshall, MD Louis M. Weiner, MD Jimmy Hwang, MD A. Ruth He, MD, PhD Amy Hankin, PA Karen Vogel, RN Divyesh Kukadiya, BS Marion Hartley, PhD Thomas Jefferson Jonathan Brody, PhD Agnieszka Witkiewicz, MD Indivumed Nina Gabelia, MD, MPH Lombardi Research Anton Wellstein, MD, PhD Narayan Shivapurkar, PhD Histopathology and Tissue Shared Resource Abbott Meeta Jaiswal, PhD Acknowledgments • Otto J. Ruesch Center for the Cure of GI Cancer • The patients and their families Georgetown Lombardi

M. J. Pishvaian* , R. Slack, A. Witkiewicz + , A. R. He,