2/21/2017. Tumor immunology. Catherine J. Wu, MD. Dana-Farber Cancer Institute, Boston, MA. DNA sequencing across cancers (n= >3000)

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1 Tumor immunology Catherine J. Wu, MD Dana-Farber Cancer Institute, Boston, MA DNA sequencing across cancers (n= >3000) Lawrence MS Nature

2 Evolving Understanding of CLL Heterogeneity Gruber & Wu Seminars in Hem

3 Fruits of NGS studies in CLL: systematic search for driving factors Diverse and heterogeneous somatic mutations in CLL (Wang NEJM 2011, Landau Tausch & Taylor Weiner Nature 2015; Puente Nature 2015) Somatic mutations suggest affected cellular pathways in CLL Intratumoral genetic heterogeneity is a common feature of CLL (Landau Cell 2013) Inference of earlier and later CLL drivers from aggregate frequencies possible (Landau Cell 2013) Preferred evolutionary trajectories (Wang Elife 2014; Landau Nature 2015) Intertumoral heterogeneity in CLL: independent evolutionary events Mutation profiles between individuals are vastly heterogeneous 538 cases, WES (278 GCLLSG-CLL DFCI-ICGC) Landau Tausch & Taylor Weiner, Nature

4 Lazarian, Guieze & Wu (submitted) Schreiber, Old & Smyth Science

5 The challenge Relationship between the immune system and human cancer is dynamic and complex Tumors: lots of somatic gene mutations and non-genetic dysregulated genes and proteins many potential foreign ANTIGENS But, the baseline relationship is one of TOLERANCE Tumors adapt and suppresses immunity 5

6 Natural immunity: How can we understand observed variations in the natural immune response? Therapeutic immunity: Why do certain patient tumors respond while others do not? The Cancer-Immunity Cycle Chen & Mellman Immunity

7 What can we learn from large-scale datasets about tumor immunity? TCGA, GTEX, CCLE datasets now available (1) What are potential drivers of immune cytolytic activity? (2) What genetic alterations are found in tumors under pressure from immune cytolytic activity? Can we generate a metric of CTL activity? Rooney Cell, 2015 Importance of TILs 1863: Virchow-immune infiltrates in tumors A plethora of data Spectrum of findings 7

8 Somatic mutations have the potential to generate neoantigens Classes of mutations that can generate potential tumor neoepitopes Missense Splice-site * LMPKHFIR (parental) LMPKLFIR (Mutated) Exon A * TGA Exon B Potential neoorfs Frame-shift * TGA Deletion or insertion Read-through * TGA INTRON TGA Gene fusion Gene A TGA Gene B 8

9 Classes of tumor antigens Neoantigens CT antigens Overexpressed Ag, shared Classes of tumor antigens Neoantigens CT antigens Overexpressed Ag, shared 9

10 Hitting the sweet spot Hacohen CIR 2013 Is CYT associated with count of mutations or neo epitopes per tumor? For each patient, impute HLA type For each patient, impute novel expressed peptides produced by each mutation Will the peptide bind any of the 6 MHC Class I alleles with IC50<500nM? Count of neo epitopes per patient 10

11 Multiple tumor types show positive association between cytolytic activity and the count of mutations Multiple tumor types show reduced rates of predicted HLA binding mutations vs. expected Antigen processing and presentation, q=1.2e 13 11

12 High mutational load related to neoantigen load and presence of T cell infiltrates Endometrial adenocarcinoma H & E CD3 CD8 MSS MSI POLE Howitt JAMA Onc 2015 A subset of driver genes are more likely to be non synonymously mutated in tumors with high cytolytic activity HLA A,B,C CASP8 B2M 12

13 Applied POLYSOLVER and HLA mutation detection pipeline to 3,708 samples across 12 tumor types Colon cancer identified as significant by MutSig Shukla Nat Biotech 2015 A more direct look. WES of 619 tumor/normal pairs of patients with CRC on FFPE specimens collected on 2 prospectively collected cohorts (NHS and HPFS) More than 20 year follow up Integration of genomic information, pathology (immune infiltration data) and clinical data Nosho et al. J Pathol 2010; 222: Immune cell subtype and infiltrate pattern previously examined Giannakis, Mu & Shukla L Garraway C Fuchs S Ogino 13

14 Higher neoantigen load correlates with overall lymphocytic reaction, and CD45RO + (memory) immune-cell infiltrates Associated with a TIL pattern of reaction Prior antigen experience Giannakis, Shukla HLA mutations are enriched in CRC samples with high TILs All tumors MSS non-pole tumors Model: Giannakis, Mu & Shukla Cell Reports

15 Several loci undergo copy number alterations significantly associated with cytolytic activity CYT +CYT IDO1,2 PDL1,2 PDL1,2 Amp High CYT B2M Deletion High CYT ALOX Amp Low Cyt 15

16 Conclusions Predicted neoepitopes correlate with CYT, even in tumor types exhibiting viral infection High-CYT high-neopeptide count tumors are associated with improved survival, suggestive of protective immune response Mutations in HLA Class I and B2M in high CYT tumors are associated with high CYT with high significance, supporting immunoediting hypothesis for a subset of tumors (large datasets may support additional tumors). PDL1/2 and IDO1/2 are amplified, but PDL1/2 is positively associated with CYT while IDO1/2 is negatively associated (like ALOX genes). Supports immunoediting in tumors, and interaction of the immune response with tumor intrinsic features The genomic landscape of a tumor shapes and is shaped by anti-tumor immunity Natural immunity: How can we understand observed variations in the natural immune response? Therapeutic immunity: Why do certain patient tumors respond while others do not? 16

17 Principles underlying immunotherapeutics Taking advantage of existing spontaneous T cell responses Adoptive transfer of TILs Checkpoint inhibitors Oncolytic viruses Cancer vaccines Engineering new responses Antibody therapy CAR-Ts Oncolytic viruses Cancer vaccines JENNER 1798 Cowpox immunization prevents smallpox PASTEUR 1870s Inactivated pathogen 17

18 Whispers and murmurs: Coley s toxin-- the first adjuvant A patient with round cell sarcoma of the jaw and abdominal metastases seen by Coley in a Photograph after 63 injections with Coley's toxins; tumour had diminished to about half its original size. b Photograph after further treatment with Coley's toxins. In his 1910 lecture at the Royal Society of Medicine Coley reported that the patient was still alive and well. Images reproduced, with permission, from Ref. 17 (1910) Royal Society of Medicine. Effective established therapies 1. Allogeneic stem cell transplantation 2. Cytokine therapy 3. Monoclonal antibodies 18

19 1. Graft versus leukemia Curative The ultimate combination therapy Personal, multiple epitope, coordinated arms of immunity 2. Cytokines IL 2 therapy for renal cell cancer and melanoma First identified as a T cell growth factor in 1976 Proliferation signal for B and T cells, cytotoxic cells Antitumor effect: through proliferation of NK cells, lymphokine-activated killer cells and CTLs.Overall response rates of 15-20% Significant toxicities, reversible with therapy discontinuation 19

20 3. Monoclonal antibodies against tumor antigens can control tumor growth CD20 Her2/neu CD33 CD52 EGFR VEGF Topalian JCO 2011 Therapeutically Impacting the Cancer Immunity Cycle Cancer vaccines Adoptive Cellular Therapy Chen & Mellman Immunity 2013 Blockade of checkpoint molecules 20

21 Adoptive immunotherapy Adoptive T cell therapy Donor lymphocyte infusion unfractionated T cell subset selected antigen-stimulated (i.e. Anti viral cellular products) Adoptive-cell therapy directed at viral antigens Expansion of TILs CAR-T A ready army of T cells with the right specificity 21

22 Science

23 Science 2002 CD4+ T cells against NY-ESO1 raised by ex vivo stimulation against Cl II defined epitopes Prolonged persistance Epitope spreading 23

24 A single clone of neoantigen-specific T cells mediates rejection of human metastatic disease Tumor Biopsy Characterize neoantigen profile Screen TILs for NeoAg reactivity TILs Identify dominant reactive T cell clone for adoptive therapy. Tran et al. Science ENGINEERED T CELLS (CAR Ts) Engineered to express the antigenbinding part of a monoclonal antibody (single chain Fv) on its surface Can recognize antigen on the cell surface in a non-mhc dependent manner, Has the affinity and specificity of monoclonal antibodies Curran et al, J. of Gene Med

25 Breakthrough in CAR mediated therapy-cd19 CAR T cells in CLL Chimeric Antigen Receptor Modified T Cells in Chronic Lymphoid Leukemia David L. Porter, M.D., Bruce L. Levine, Ph.D., Michael Kalos, Ph.D., Adam Bagg, M.D., and Carl H. June, M.D. N Engl J Med 2011; 365:

26 No detectable CLL after CD19- CAR T infusion Porter et al, N Eng J Med 2011 Fig. 5 Evaluation of clinical responses after infusion of CART19 cells. M. Kalos et al., Sci Transl Med 2011;3:95ra73 Published by AAAS 26

27 CART for multiple myeloma BEFORE AFTER 2017 FDA approval is very likely for DLBCL, ALL and other diseases Garfall, Maus NEJM 2015 ENGINEERED T CELLS tories/detail/a new kind of clinical trial 27

28 Recent adoptive cell therapy trials using CAR engineered T cells CD19 (>30) for B cell malignancies, ALL, CLL, MCL, HD BCMA (1) MM CD20 (1) NHL CD22 (1) B cell malignancies CD30 (3) HD, NHL, CD30+ lymphoma CD33 (1) AML CD139 (1) myeloma CD171 (1) Neuroblastoma CEA (1) EGFR (2) GBM and solid tumors EGFRviii (1) GBM ErbB (1) H and Neck Ca FAP (1) mesothelioma GD2 (3) neuroblastoma and sarcoma Glypican 3 (1) HCC Her2 (4) sarcoma, GBM, lung cancer, solid tumors Gill Si et al, Blood Rev

29 Are we ready to cure all cancers with CAR T cells?, Neurologic toxicity Molecular Therapy 2010: 18, Checkpoint blockade inhibitors 29

30 Check point blockade (CPB) therapies Mellman 2011 Releasing the brakes 30

31 Anti-CTLA4 treatment (post vaccine) Ovarian cancer Hodi...Dranoff 2008 Hodi...Dranoff

32 Dramatic clinical responses after checkpoint blockade antibodies for solid tumors Anti-CTLA4 for metastatic melanoma Study Ipi 3mg/kg 100 patients with ipi alone 100 ipi/gp gp100 Results - Median survival 6 10 months Conclusions - Anti-CTLA4 induces clinically effective tumor immunity Hodi FS, et al. NEJM 2010 Caveats - Autoimmune effects - Helps only a minority of patients 32

33 Clinical activity of α-pd1 Ab Topalian SL, et al. NEJM 2012 Clinical activity of α-pdl1 Ab Brahmer JR., et al. NEJM

34 Who responds? PD-1 blockade in R/R Hodgkin s lymphoma Elevated PD-L1/2 expression 9p24.1 amplification JAK2 amplification EBV infection Universal 9p24 abnormalities Green Blood 2010; Green CCR 2012 Roemer JCO 2016 Chen CCR 2013; Roemer JCO

35 Change From Baseline, % Cohort 1: Progressed after ASCT and subsequent BV therapy 63 (95%) patients had a reduction in tumor size Change From Baseline, % -80 Very frequent -100 PD 100 L1/L2 expression Change From Baseline, % Cohort 3: Failed ASCT and not treated with BV 56 (93%) patients had a reduction in tumor size Cohort 2: Failed salvage chemotherapy, ineligible for ASCT, and failed BV therapy 73 (92%) patients had a reduction in tumor size Armand P; Ansell S 2016 KEYNOTE-024 trial; Front-line advanced NSCLC; Need to have PDL1 levels of 50% or more 35

36 Checkpoint blockade therapy BEFORE AFTER METASTATIC MELANOMA COLORECTAL CANCER Neoantigen load associated with NSCLC response to Pembro Discovery set: 16 pts Validation set: 18 pts Rivzi., Science,

37 Ipilimumab and melanoma cohort study: clinical correlates WES on FFPE pretreatment samples from 110 pts with metastatic melanoma Matched RNA seq on 40 Van Allen, Miao, Schilling et al, Science 2015 Checkpoint blockade inhibition Gubin, Nature Van Rooij. JCO

38 High mutation load Low mutation load 25 patients, 56% with response (4 CR, 10 PR) in both virus + and virus negative tumors α CTLA4 + α PD1 therapy Wolchok JD, et al. NEJM

39 Most patients do not have complete responses anti PD1 alone 9% complete response Larkin V, et al. NEJM 2015 Toxic autoimmunity is common anti PD1 alone 16% grade 3 or 4 adverse event 7.7% discontinued treatment Larkin V, et al. NEJM

40 Oncolytic viruses FDA approved % responses with TVEC, vs 2% GM CSF Dual mechanism direct tumor killing induction of systemic anti tumor immunity Challenges difficult delivery, may be eliminated early Cancer vaccines 40

41 Expand and broaden the T cell repertoire by inducing neoantigenspecific T cells Generate highly specific anti-tumor immunity with fewer side effects on vital tissues Challenges and potential solutions 41

42 Challenges and potential solutions 2a. Common adjuvants Clinicaltrials.gov, Sept

43 2b. GM-CSF 2c. Local production of GM-CSF improves tumor antigen presentation Intense localized skin reactions at the injection sites CD4+ CD20+ CD8+ IgG,K Associated with new influx of immune mediators Associated with subsequent immune mediated tumor rejection at distant metastatic sites Soiffer, JCO

44 2d. Common adjuvants act through TLR signaling 2e. Poly ICLC is a highly effective vaccine adjuvant - Nucleic acid ligands of TLR/RLRs are effective adjuvants - CpG DNA is difficult to obtain for trials - dsrna stimulates several key pathogen sensors - Stabilization of pic in a complex with carboxymethylcellulose, poly-lysine and pic Caskey M J Exp Med Caskey M J Exp Med

45 3. Diverse formats of immunogen Clinicaltrials.gov, Sept

46 Selection and delivery of antigen Antigen-specific vaccination: the opportunity to focus the response and broaden the T cell repertoire Less impressive. Single immunogen vaccine studies Glimmers of success! Whole tumor cell vaccines Improved delivery Long peptides (HPV) Dendritic cell-based vaccines (DC fusion, Provenge) Whole tumor cell vaccines Complex vaccines-many antigens available to stimulate B and T cell responses Potential to be a personal vaccine Potentially poorer expression of any one ag +GM-CSF Low-ish activity 46

47 Sipuleucel: DC-based vaccine 47

48 Study 9 E6 & 4 E7 peptides (30 35mer) Montanide (DMSO/PBS) 0.3mg peptide/patient Results 47% CR with: - 100% CD4+ T responses - 83% CD8+ T (76% E6, 11% E7) - CD8: mean of ~3 epitopes Conclusions First demonstration that long peptide vaccines generate clinically significant tumor immunity Caveats CD8 responses cover few epitopes per patient 48

49 A long peptide neoantigen vaccine can mediate tumor control + prophylactic therapeutic Gubin Nature 2014 Yadav Nature

50 Carreno Science 2015 (submitted) Final points Immunological approaches have been attempted for more than 100 years but only past decade do w see real promise Important strides in understanding of underlying biology The (past) and future lies in combination therapy how to integration of conventional approaches (surg/chemo to reduce tumor load) or novel targeted therapy with immunotherapy Precision immuno oncology how can we match patients to the right therapeutic modality? 50