Tumor mutation burden as predictive biomarker from discovery to standardization

Transcription

1 ISTITUTO NAZIONALE PER LO STUDIO E LA CURA DEI TUMORI FONDAZIONE G. Pascale NAPOLI SC Biologia Cellulare e Bioterapie CENTRO RICERCHE ONCOLOGICHE MERCOGLIANO (AV) Laboratorio di Farmacogenomica Tumor mutation burden as predictive biomarker from discovery to standardization Nicola Normanno

2 Somatic mutation frequencies in cancer Lawrence Nature 2013

3 Tumor Mutation Burden in NSCLC Adeno (n=7,925) SCC (n=1,324) NSCLC NOS (n=1,773) SCLC (n=640) EGFR mutation (n=1,775) ALK or ROS1 fusion (n=489) METex14 alteration (n=286) BRAF mutation (n=493) KRAS mutation (n=3,155) Mean Mutations/ Mb TMB >10 (%) TMB >20 (%) Wilcoxon signed-rank test vs KRAS (30) 760 (10) 541 (41) 113 (9) 711 (40) 233 (13) 269 (42) 42 (7) 129 (7) 21 (1) 17 (3) 4 (1) 27 (9) 4 (1) 153 (31) 51 (10) p<0.001 p<0.001 p<0.001 p< ,238 (39) 298 (9) Spigel ASCO 2016

4 Estimate of the neoantigen repertoire in human cancer Schumacher Science 2015

5 Classes of human tumour antigens that are recognized by T lymphocytes Coulie Nat Rev Cancer 2014

6 Candidate neoantigens and response to pembrolizumab Rizvi Science 2015

7 Predicted Neoantigens Correlation Between TMB and Neoantigen Load 1000 R 2 =0.91 Spearman rho 0.92 Pan-Tumor Total Mutations Higher levels of TMB have been shown to correlate with higher neoantigen load 1-3 Melanoma, lung, bladder, liver, and stomach cancers are predicted to have the highest neoantigen loads 3 TMB=tumor mutation burden. 1. Hellmann M. Oral presentation at AACR Van Allen EM et al. Science. 2015;350(6257): Rooney MS et al. Cell. 2015;160:48-61.

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9 Correlation between TMB and Response Rate with Anti PD-1 or Anti PDL1 Therapy in 27 Tumor Types Yarchoan NEJM 2018

10 Whole Exome Sequencing Is a Technical and Complex Process NGS-based genomic profiling workflow 1 ACTGTGCGACGA CGCGTAGATCGC AACGTCGCGATA DNA Extraction Library Preparation Hybrid Capture DNA Sequencing Alignment Data Analysis Clinical Report Sample attrition may occur within the process, and technical complexity may require specialty labs 2 Use of consolidated gene panels may alleviate some of the challenges associated with wholeexome sequencing 2 1. Frampton GM et al. Nat Biotechnol. 2013;31(11): Ulahannan D. Br J Cancer. 2013; 109(4):

11 Assessment of TMB with targeted sequencing FoundationOne panel 315 genes Chalmers Genom Med 2017

12 TMB assessed by targeted NGS correlates with WES and durable clinical benefit (DCB) MSK-IMPACT panel 468 genes Rizvi JCO 2018

13 CheckMate 227: Nivo + Ipi in 1L NSCLC With High TMB ( 10 mut/mb) CheckMate 227 Part 1 Study Design a Patients for PD-L1 co-primary analysis N = 1189 Nivolumab 3 mg/kg Q2W Ipilimumab 1 mg/kg Q6W n = 396 Nivolumab + ipilimumab n = 396 1% PD-L1 expression R 1:1:1 Histology-based chemotherapy b n = 397 Chemotherapy b n = 397 Key Eligibility Criteria Stage IV or recurrent NSCLC No prior systemic therapy No known sensitizing EGFR/ALK alterations ECOG PS 0 1 Stratified by SQ vs NSQ Nivolumab 240 mg Q2W n = 396 Patients for TMB co-primary analysis c Nivolumab + ipilimumab n = 139 Chemotherapy b n = 160 N = 550 Nivolumab 3 mg/kg Q2W Ipilimumab 1 mg/kg Q6W n = 187 <1% PD-L1 expression R 1:1:1 Histology-based chemotherapy b n = 186 Nivolumab 360 mg Q3W + histology-based chemotherapy b n = 177 Co-primary endpoints: Nivolumab + ipilimumab vs chemotherapy OS in PD-L1 selected populations PFS in TMB-selected populations Database lock: January 24, 2018; minimum follow-up: 11.2 months a NCT b NSQ: pemetrexed + cisplatin or carboplatin, Q3W for 4 cycles, with optional pemetrexed maintenance following chemotherapy or nivolumab + pemetrexed maintenance following nivolumab + chemotherapy; SQ: gemcitabine + cisplatin, or gemcitabine + carboplatin, Q3W for 4 cycles; c The TMB co-primary analysis was conducted in the subset of patients randomized to nivolumab + ipilimumab or chemotherapy who had evaluable TMB 10 mut/mb 7

14 CheckMate 568: Identification of Optimal TMB Cutoff ( 10 mut/mb) for Nivo + Ipi in 1L NSCLC TMB Analysis With FoundationOne CDx Assay FoundationOne CDx TM (F1CDx) Tumor sample (FFPE) FoundationOne CDx TM FoundationOne CDx uses next-generation sequencing to detect substitutions, insertions and deletions, and copy number alterations in 324 genes and select gene rearrangements TMB: total number of synonymous and nonsynonymous variants ( 5% allele frequency) after filtering germline mutations Somatic mutations per megabase FoundationOne CDx TM TMB result FoundationOne CDx Technical Information. Cambridge, MA: Foundation Medicine Inc;

15 ORR (%) CheckMate 568: Identification of Optimal TMB Cutoff ( 10 mut/mb) for Nivo + Ipi in 1L NSCLC ORR by TMB a,b n/n 2/23 4/27 21/48 11/28 <5 c 5 to <10 d 10 e 15 f TMB (mut/mb) a Irrespective of PD-L1 expression; b 12% ORR for <10 mut/mb and 50% ORR for 10 to <15 mut/mb; c CR = 0; d CR = 4%; e CR = 8%; f CR = 7% 15

16 True-positive fraction True-positive fraction CheckMate 568: Identification of Optimal TMB Cutoff ( 10 mut/mb) for Nivo + Ipi in 1L NSCLC ROC Curves by Objective Response PD-L1 (n = 252) TMB (n = 98) mut/mb AUC = AUC = False-positive fraction False-positive fraction 16

17 PFS (%) CheckMate 227: Nivo + Ipi in 1L NSCLC With High TMB ( 10 mut/mb) Co-primary Endpoint: PFS With Nivolumab + Ipilimumab vs Chemotherapy in Patients With High TMB ( 10 mut/mb) a Nivo + ipi (n = 139) Chemo (n = 160) Median PFS, b mo HR c 97.5% CI , 0.81 P = y PFS = 43% Nivolumab + ipilimumab 20 1-y PFS = 13% 0 Chemotherapy No. at risk Months Nivo + ipi Chemo In patients with TMB <10 mut/mb treated with nivo + ipi vs chemo, the HR was 1.07 (95% CI: 0.84, 1.35) d a Per blinded independent central review (BICR); median (range) of follow-up in the co-primary analysis population was 13.6 mo (0.4, 25.1) for nivo + ipi and 13.2 mo (0.2, 26.0) for chemo; b 95% CI: nivo + ipi (5.5, 13.2 mo), chemo (4.4, 5.8 mo); c 95% CI: 0.43, 0.77 mo; d The P-value for the treatment interaction was

18 Blood-based TMB Gandara Nat Med 2018

19 TMB testing: Critical issues TMB is emerging as a relevant biomarker for response to immunotherapy. Standardization and harmonization of TMB testing are pivotal. Different open questions need to be answered: 1. Current methods for TMB measurement are not standardized and vary from WES to targeted sequencing panels (commercial kits, laboratory-developed assays ) 2. As a consequence, bioinformatic pipelines for TMB calculation differ among the different assays 3. No univocal cut-offs to identify low-medium-high TMB tumors exist (the only validated cut-off is 10mut/mb with FoundationOne CDX, Checkmate-227)

20 FoundationOne CDx 324 genes - Threshold 10 mut/mb FoundationOne 324 genes value 10 Thermo Fisher 409 genes value 12 Qiagen 400 genes value 12 WES value 200 Mutations/Mb Illumina 500 genes value 14

21 Aims of the study: a) to organize a pilot European EQA scheme for TMB; b) to harmonize the different TMB methods; c) to define recommendations for TMB assessment. Academia: ESMO, AIOM, Gen&Tiss, ESP, UKNEQAS, EMQN, ciqc, RCPA Quality Assurance Programs Sponsors: BMS, AstraZeneca, Roche/Foundation Medicine, Thermo Fisher Scientific, Illumina, Qiagen, Genentech

22 Tumour mutation burden: from recommendations for testing to external quality assessment schemes Project Kick-off Meeting Naples, 10 th May 2018 Participants: ESMO AIOM Gen&Tiss ESP EMQN ciqc BMS Thermo Fisher Illumina Qiagen Genentech Merck KGaA

23 Format of pilot EQA Selection of participants (survey) Release of EQA results and Scheme Report EQA material validation study EQA assessment EQA distribution Results submission - Reports - Data collection

24 Project Planning a) EQA Pilot scheme for TMB testing EQA sample validation FFPE cell lines with different mutational load (high, intermediate, low TMB) Different validating laboratories will be identified Perform sample validation (DNA extraction + TMB testing) Different TMB panels to test the samples (Thermo Fisher Qiagen Illumina - FoundationOne) EQA sample distribution Identification of 30 laboratories with a survey taking into account labs with molecular pathology expertise, already performing TMB testing, with different methods Analysis of EQA results/issue of results Specific scoring system TMB result and clinical interpretation

25 Project Planning b) Harmonization phase a) Creation of an harmonization network a) Identification of a panel of experts to provide FFPE material from NSCLC patients. b) Identify the appropriate number of centers to be involved b) Sample selection and analysis a) Collection of samples in order to obtain a significant number of analyses to allow the creation of a conversion table c) Comparative analysis a) Performance of comparative analysis of the scores resulting from different TMB tests to create a reference table that maps the score of one text to another, to enable harmonisation. d) Publication of the results/workshops

26 Harmonization phase (B) Validation phase (A) Project Timeline TMB project Timeline Jun/ Jul 2018 Aug/ Sept 2018 Oct/ Nov 2018 Dec 2018/ Jan 2019 Feb/ Mar 2019 Apr/ May 2019 Jun/ Jul 2019 Aug/ Sept 2019 Oct 2019 Tender for control material Identification of the participating centers (survey) Place order for EQA material Material manufacture Requirements for data collections EQA sample validation EQA sample distribution Analysis of EQA results/issue of results Creation of an harmonization network Sample selection and analysis Comparative analyses Publication of the results/workshops

27 Immuno-oncology Consortium with Leading EU Institutes Empower clinical trials with Ion Torrent immuno-oncology assays The immuno-oncology consortia members: Sabine Merkelbach-Bruse Institute of Pathology, University Hospital Cologne International Albrecht Stenzinger Institute of Pathology, Heidelberg University Hospital Michael Hummel Institute of Pathology, Charité - University Medical Center Berlin Wilko Weichert and Nicole Pfarr Institute of Pathology, Technical University of Munich Bea Bellosillo Hospital del Mar, Barcelona Nicola Normanno IRCCS National Cancer Institute "G. Pascale Foundation" Jose Carlos Machado i3s and Institute of Molecular Pathology and Immunology of the University of Porto (IPATIMUP), Portugal For Research Use Only. Not for use in diagnostic procedures. Maurice Jansen and Winand Dinjens Erasmus MC Cancer Institute, Rotterdam 27

28 Tumour Mutational Load assay project - Experimental plan Phase 1 - Controls 6 x cell line FFPE controls (AccuRef) predicted to be high and low TMB plus 2 x Acrometric Oncology Hotspot Control (AOHC) check reproducible results across 8 labs Phase 2 - Shared sample IPATIMUP providing 8 CRC samples for all labs which have MSI status, no exome or clinical response data 8 shared samples run as standard TML workflow 8plex chip without normal comparison TUM providing access to tumour-normal exome analysis via DKFZ Comparison to exome and interlay reproducibility Phase 3 - Own samples 16 samples from own biobank high and low mutational burden status Ideally with clinical response data and/or any supporting biomarker/exome data 28

29 TMB(Mut/Mb) Preliminary Phase I Results Cell-Ref FFPE Slides and Acrometrix Control A549 AOHC H2228 HCC2998 MCF7 SKMEL2 T47D Average TMB % CV 12% 9% 8% 6% 17% 9% 14% Range

30 Rep 2 (CROM) Preliminary data Phase II results (2 Laboratories) 70 Reproducibility of Shared Clinical Samples R² = 0, Rep 1 (IPATIMUP) 30

31 TMB testing in CRC cell lines (1) MSI Status Mutation Count (cbbioportal) TML (Oncomine TML Assay) RKO MSI-H ,62 LOVO MSI-H ,64 HCT116 MSI-H ,62 COLO320 MSS 23 15,89 HT29 MSS 64 15,89 SW1116 MSS 40 15,87 H1650 MSS 25 6,74 H1975 MSS 54 10,37

32 Mutation Count - CbioPortal TMB testing in CRC cell lines (2) R² = 0, TMB (Oncomine TML Assay)

33 Mutation Load/MB (Oncomine TML Assay) TMB in primary CRC FFPE samples MSI MSI-L/MSS

34 Genomics-Driven Oncology Garraway JCO 2013

35 Thermo Fisher Scientific and its affiliates are not endorsing, recommending, or promoting any use or application of Thermo Fisher Scientific products presented by third parties during this seminar. Information and materials presented or provided by third parties are provided as-is and without warranty of any kind, including regarding intellectual property rights and reported results. Parties presenting images, text and material represent they have the rights to do so.