TOPARP: From Genomics To Therapy

Prostate Cancer: From Genomics To TherapyJohann Sebastian de BonoDirector, Drug Development UnitProfessor of Experimental Cancer MedicineThe Institute of Cancer Research and Royal MarsdenLondon, United Kingdom

Qiagen Meeting January 2016

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Prostate CancerMost common cancer in men and second most common lethal cancer in men; metastatic disease invariably lethalMany new treatmentsAbiraterone, enzalutamide, cabazitaxel, radium 223Molecularly highly heterogeneousNo stratified treatment to datede Bono et al, NEJM 2011; de Bono et al, The Lancet 2010; Scher et al, NEJM 2012;Parker et al, NEJM 2014; Robinson et al, Cell 2015.

Seminar on prostate cancer published in The Lancet, 11th June 2015

Therapy: PARP Inhibitors in Advanced Prostate Cancer

HypothesisA molecular subclass of prostate cancers have DNA repair defects that render them vulnerable to synthetic lethal therapeutic strategies utilizing PARP inhibition.

Synthetic Lethality: Targeting the Achilles heelSynthetic lethal strategies pursue this Holy Grail: Killing tumor cells but sparing normal cells.

Synthetic lethality occurs when there a potent and lethal synergy between two otherwise non-lethal events: in this case a highly specific PARP inhibitor induces a DNA lesion and a tumor-restricted genetic loss of function for the DNA repair pathways required to repair it.Farmer et al, Nature 2005; Bryant et al, Nature 2005; Fong et al, NEJM 2009

Investigator-initiated multi-stage Phase II trialNCT-01682772; CR-UK/11/029Adaptive design focused on predictive biomarker identificationTest set (all comers; Part A) and validation set (patient selection; Part B)Possibility to proceed to a randomized Phase II of PARPi vs placeboOpen label, Olaparib (tabs) 400mg BIDStudy ObjectivesTo evaluate the antitumour activity of olaparib in mCRPCTo identify and clinically qualify a predictive biomarker suite for PARP inhibitor antitumour activity in CRPC

Trial design

Like the first abiraterone Phase I/II trial and the olaparib first in man Phase I trial, this trial was written at Flims.

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Primary endpoint: Response RateResponse as per RECIST 1.1PSA decline >50% (PCWG-2)CTC conversion (>5 to 4 weeks laterSecondary endpoints included: rPFS, OS, duration of responses, safety-tolerability.Exploratory endpoints:Study of diffusion-weighted MRI as response biomarker

Trial design

CTC Conversion for these 49 patients: Association with rPFS, OS

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po= RR 5%; p1= RR 20%=0.02; =0.10

Trial Design: Elucidating the Predictive Biomarker Suite

Investigator Initiated Trial Written at AACR/ESMO/ECCO Flims WorkshopTO-PARP-A; What I am presenting today.

Written with Shahneen Sandhu, now at Peter Mac, Melbourne

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po= RR 5%; p1= RR 20%=0.02; =0.10

Trial Design: Elucidating the Predictive Biomarker Suite

TOPARP-Part C

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Biomarker studies Mandated pre-and post-PARPi fresh tumour biopsies. Planned studies to analyse predictive biomarker suite:Whole exome and transcriptome analyses (HiSeq)Targeted NGS for validation (MiSeq; Qiagen M&Mv2)ctDNA: serial plasma samples (MiSeq; Qiagen M&Mv2)

PD studies in tumour tissuesGamma H2AX foci, RAD51 foci, 53BP1 foci

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15Statistical Analysis Plan Predictive Biomarker DefinitionDefinition of Biomarker positive (B+) patientPresence of a homozygous deletion AND/OR a putative deleterious mutation in a gene reported to be involved in DNA repair and/or sensitivity to PARP inhibition

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Trial populationMetastatic CRPC after 1-2 lines of taxane chemotherapy.Documented progressive disease by RECIST or PSA (PCWG2).ECOG Performance Status 0-2.Appropriate organ-function: Hb >10g/l, Neut>1.5x109/l, Plt>100x109/l, Bilir1yFocus on the 3 who stopped early: a false response and 2 patients with mielotoxCorrelation of PSA and CTC falls6 RECIST responses among patients with measurable disease.26

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PALB2: Partner and localizer of BRCA2. Pt 60: Biallelic HDAC2 loss and low HDAC1/2

Biomarker studies DNA repair defects associate with response

* Estimated from a logistic regression model

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Patients with and without DNA Repair Defects Time to progression and Overall Survival

Changes in PSA and CTC CountsMean counts during treatment by biomarker

Individual CTC Counts for TOPARP-A PatientsMolecular Characterization was prospectively planned but only available retrospectively

Biomarker studies BRCA2 loss sensitizes to olaparibIDGermline hit?Response Y/NRECIST responsePSA fall >50%CTC conversionTime on trial (weeks)14YESYESN/AYESYES3615YESYESPRYESYES3617NO, SOMATIC fs + het-delYESPRYESYES2420NO, SOMATIC fs + het-delYESPRYESN/E40+30YESYESN/AYESYES40+35NO, SOMATIC hom-delYESPRYESYES24+39NO, SOMATIC hom-delYESPRYESYES40+

All 7 achieved PSA falls over 50% and the 5 of them with measurable disease in CT scans achieved a RECIST response. Interestingly, only 3 had germline mutations detected, with the other 4 having somatic loss of BRCA2 (2 by hom-del and 2 by fs+del).REINFORCE idea germline mut were not known before trial.34

Somatic homozygous deletion of BRCA2

Another BRCA2 Responder

BASELINEWEEK 12Radiological PR lasting 9 months.

Previously unknown germline BRCA2 mutation + loss of 2nd copy in tumor (no family history of cancer)

CTC COUNT

PSA

Is BRCA carrier CRPC underdiagnosed? Probably.

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Biomarker studies ATM defects and response to olaparibIDalteration detectedResponse Y/NRECIST responsePSA fall >50%CTC conversionTime on trial (weeks)1SOMATIC p.N2875S (kinase domain)YESN/EYESYES735GERMLINE stop gain p.W2638* (kinase domain)YESN/EYESYES56+6SOMATIC fs deletion p.-2288fs (kinase domain)YESSDNO (30%)YES17*12GERMLINE fs del p.-1970 + SOMATIC p.I2752F (kinase domain)NOPDNONO1136GERMLINE fs del p.-1267fs + SOMATIC LOHYESSDYESYES57

*Patient 6 came off study after repeated dose reductions for anemia

Some germline events too37

Prolonged response in a patient post docetaxel, cabazitaxel, abiraterone and enzalutamide.

The patient is still responding after 10+ months. Has stopped opioids and his mobility has improved.Germline FS del ATM + LOH in tumor.

Transcriptome analysis: very low ATM expression in tumor.

CTC COUNT

PSA

TO-PARP Responder: Biallelic ATM LossPatient on trial > 1 year

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TO-PARP Responder:ATM Genomic characterization by LOH analysis.ChromosomeStartEndMean Zygosity DeviationLog2 Coverage RatioCopy Number RatioProbabilityClassificationHeterozygous SNPsTargeted Exons111066471351087231020.414-0.6070.65656111 Copy41208ATM, GUCY1A2

ATMATM is on a region of one copy loss in the tumor. The reference allele is lost in the tumor and the frameshifted variant allele is retained.

ATM expression across a cancer compendiumRed arrow is this patients ATM expressionTO-PARP Responder Case 3:Low ATM mRNA Expression

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T001014 Pre Castration Resistant Prostate Cancer (Poorly diff adenocarcinoma)IHC: Nuclear H score: 0 Cytoplasm H score: 0

IHC: Nuclear H score: 0 Cytoplasm H score: 0

Responding patient

Biomarker studies ATM Truncating Mutation and response to olaparib

Baseline+3 months

PSACTC countNGS of the baseline bone marrow biopsy revealed a somatic missense mutation within the ATM PI3K catalytic-domain (p.N2875S) without evidence of genomic loss of the second allele and maintained ATM expression by IHC. ? Dominant negative (Bakkenist & Kastan, Nature 2003); Haploinsufficiency?

Somatic missense mutation disrupting the kinase domain of ATM, Was on trial for 14 months. 42

Serial Plasma DNA NGS: TOPARP

Patient T001-15-V4081-DB

Patient T001-14-V4038-TS

Patient T001-09-V5116-BC

TO-PARP Trial ConclusionsTherapeutic trials in CRPC mandating fresh biopsies feasible and can inform on disease biologyA proportion of mCRPC have DNA repair defects that can be targeted through a synthetic lethal strategy by PARPi.Approximately 20-30% of mCRPC patients warrant molecular stratification by analyses of DNA repair gene defects It is envisioned that targeted NGS will impact the treatment of CRPC patients not only in the metastatic setting but also earlier in the management

Acknowledgements

Experimental Cancer Medicine Centre To all patients and their families for their participation and support.To the AACR-ESMO-ECCO Flims workshop, where this protocol was developed.Supported by Cancer Research UK (CRUK/11/029; C12540/A12829; C1491/A15955; C12540/A13230) through Collaboration between AstraZeneca and National Cancer Research Network.SU2C-PCF Prostate Cancer Dream TeamPCF Challenge Award (Knudsen, Feng, Rubin & de Bono).All the Cancer Biomarkers group at the ICRTOPARP investigators and all the staff at the participating sites, Trial Management Group, Trial Steering Committee and IDMC. Staff at Clinical Trials and Statistics Units at the ICR.

Prostate Cancer UK is a registered charity in England andWales (1005541) and in Scotland (SC039332). Registered company 2653887

in partnership with

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Title, Location, Date51

Sheet1Landmark analysis: what is the prognostic values of CTC response at 4,8 and 12 weeks to predict future events?4 weeks8 weeks12 weeksNo. HR (95% CI)p-valueNo. HR (95% CI)p-valueNo. HR (95% CI)p-valuerPFSCTC conversion460.48 (0.23, 0.996)0.049400.37 (0.18, 0.77)0.0072.10 (0.43, 10.39)0.36OSCTC conversion490.32 (0.13, 0.77)0.011470.31 (0.14, 0.68)0.004430.36 (0.16, 0.81)0.01CTC conversionPatients who are alive and progression-free and have responded by CTC by 4 weeks have lower risk of progressionPatients who are alive and have responded by CTC at 4 weeks have lower risk of dyingIdem at 8 weeksLandmark at 12 weeks for progression is not informative, as many events happen by 12 weeks, few progressions occur afterwards. Though HR is >1, please note the wide CI and small sample size!!!!For Os, patients who are alive and have responded by 12 weeks have a lower risk of dying Fold change in CTC: fold change in CTC, HR1: higher risk if CTC increases

CTC conversion as a time dependent covariate: HR (95% CI)p-valuerPFS0.30 (0.13, 0.66)0.003OS0.58 (0.26, 1.30)0.19Patients that respond by CTc seem to have lower risk of progressing and dying ( HR