Get 20M+ Full-Text Papers For Less Than $1.50/day. Start a 14-Day Trial for You or Your Team.

Learn More →

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016): late breaking abstracts

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC... Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 DOI 10.1186/s40425-016-0191-4 MEETING ABSTRACTS Open Access 31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016): late breaking abstracts National Harbor, MD, USA. 9-13 November 2016 Published: 8 December 2016 tumors [3]. By 29APR2016, 304 previously treated NSCLC pa- Biomarkers and Immune Monitoring tients, median 3 prior lines, received 10 mg/kg of durvalumab q2w ≤12 months. Baseline archived or fresh tumor biopsies O1 were analyzed for PD-L1 (Ventana/SP263) and CD8 (Ventana/ Combinatorial CD8+ and PD-L1+ cell densities correlate with SP239) by IHC. For the marker combination, slides were scored response and improved survival in non-small cell lung cancer using the product of PD-L1+ and CD8+ cell densities with (NSCLC) patients treated with durvalumab 1 2 2 1 Definiens’ Developer XD 2.1.4 software. For PD-L1 alone, ≥25% Sonja Althammer , Keith Steele , Marlon Rebelatto , Tze Heng Tan , 1 1 2 2 2 tumor cells stained for PD-L1 at any intensity were scored posi- Tobias Wiestler , Guenter Schmidt , Brandon Higgs , Xia Li , Li Shi , 2 2 2 2 tive. Clinical outcomes (ORR, PFS and OS) were analysed based Xiaoping Jin , Joyce Antal , Ashok Gupta , Koustubh Ranade , Gerd on CD8+ and PD-L1+ densities (n = 163 available) and PD-L1 Binning 1 2 alone in pre-treatment biopsies using a discovery (n = 84) and Definiens AG, Munich, Bayern, Germany; MedImmune, Gaithersburg, validation (n = 79) set. Datasets were matched on baseline PD- MD, USA L1 status, histology, ECOG, lines of therapy, and response. Correspondence: Brandon Higgs (higgsb@medimmune.com) Results Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O1 Patients with high pretreatment CD8+ and PD-L1+ densities (preva- lence = 36%) had better ORR, OS, and PFS compared to those with Background low CD8+ and PD-L1+ densities (Fig. 1), as well as high PD-L1 expres- Immunotherapies have improved patient responses and sur- sion alone. vival, though not all patients benefit. Effective biomarkers may Conclusions help to improve outcomes. Durvalumab is a human IgG1 mono- Automated image analysis of CD8+ and PD-L1+ cell densities in clonal antibody that inhibits PD-L1 binding to PD-1 and CD80, baseline tumor biopsies may identify patients with improved out- restoring antitumor immunity [1, 2]. PD-L1 expression on tumor comes to durvalumab. or tumor-infiltrating immune cells measured manually with dif- Trial Registration ferent immunohistochemistry (IHC) assays can enrich for pa- ClinicalTrials.gov identifier NCT01693562. tients responding to anti-PD-1/PD-L1 agents. Tumor-infiltrating cytotoxic CD8+ T cells may also have potential predictive utility for therapeutic response. We explored automated image ana- References lysis and pattern recognition of tumor biopsies to determine 1. MedImmune/AstraZeneca. Data on file. whether CD8+ and PD-L1+ cell densities could better identify 2. Ibrahim R, Stewart R, Shalabi A: PD-L1 blockade for cancer treatment: patients most likely to respond to durvalumab than PD-L1 IHC MEDI4736. Semin Oncol 2015, 42:474–483. alone. 3. Rizvi NA, Brahmer JR, Ou SHI, Segal NH, Khleif S, Hwu WJ, et al: Safety Methods and clinical activity of MEDI4736, an anti-programmed cell death- CP1108/NCT01693562 was a nonrandomized phase I/II trial ligand 1 (PD-L1 antibody, in patients with non-small lung cancer evaluating durvalumab in advanced NSCLC and other solid (NSCLC). J Clin Oncol 2015, 33(Suppl.):Abstract 8032. © The Author(s). 2016 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 224 of 242 Methods Eligible patients were enrolled regardless of PD-L1 expression and randomized 1:1 to pembrolizumab 200 mg Q3W for 24 months or investigator’schoice ofpaclitaxel175 mg/m Q3W, docetaxel 2 2 75 mg/m Q3W, or vinflunine 320 mg/m Q3W. Randomization was stratified by ECOG PS (0/1 vs 2), liver metastases (yes vs no), hemoglobin level (<10 vs ≥10 g/dL), and time from last chemo- therapy dose (<3 vs ≥3 months). The study had a group sequen- tial design to control for type I error. Primary endpoints were OS and PFS (RECIST v1.1 by blinded, independent central review). ORR was a key secondary endpoint. Differences in OS and PFS were assessed in the intention-to-treat population using the stratified log-rank test Fig. 1 (abstract O1). Clinical outcomes in CD8+/PD-L1+ or PD-L1 Results NSCLC patient subsets Between November 5, 2014 and November 13, 2015, 542 patients from 29 countries were enrolled: 270 in the pembrolizu- mab arm, 272 in the chemotherapy arm. As of September 7, 2016, median follow-up was 9.0 months; 49 (18.4%) patients remained on pembrolizumab and 3 (1.2%) patients remained on chemotherapy. Baseline characteristics were generally balanced between arms, with 87.3% with visceral disease, 34.3% with liver Clinical Trials: Cutting-Edge (Completed metastases, 1.1% with ECOG PS 2, 81.5% with hemoglobin ≥10 g/dL, and 38.2% with <3 months since most recent chemo- Trials) therapy. Pembrolizumab significantly improved OS over chemo- therapy (HR 0.73, P = 0.0022; median 10.3 vs 7.4 months) (Table O2 1). There was no difference in PFS (HR 0.98, P = 0.42) (Table 1). Keynote-045: open-label, phase III study of pembrolizumab versus ORR was significantly improved with pembrolizumab (21.1% vs investigator’s choice of paclitaxel, docetaxel, or vinflunine for 11.4%) (Table 1). Pembrolizumab was associated with fewer any- previously treated advanced urothelial cancer 1 2 3 4 grade (60.9% vs 90.2%) grade 3-5 treatment-related AEs (15.0% Joaquim Bellmunt, Ronald de Wit, David J Vaughn, Yves Fradet, Jae 5 6 7 8 vs 49.4%). 4 patients in each arm died due to treatment-related Lyun Lee, Lawrence Fong, Nicholas J Vogelzang, Miguel A Climent, 9 1 10 11 AEs. Daniel P Petrylak, Toni K Choueiri, Andrea Necchi, Winald Gerritsen, 12 13 14 Conclusions Howard Gurney, David I Quinn, Stéphane Culine, Cora N 15 16 16 16 Pembrolizumab demonstrated a statistically significant OS bene- Sternberg, Yabing Mai, Markus Puhlmann, Rodolfo F Perini, Dean fit over chemotherapy in the second-line advanced urothelial F Bajorin 1 2 cancer setting, making it the first therapy to demonstrate a Dana-Farber Cancer Institute, Boston, MA, USA; Erasmus MC survival benefit over an active comparator in this population. Cancer Institute, Rotterdam, Netherlands; Abramson Cancer Center The superior OS combined with the lower rate of any-grade of the University of Pennsylvania, Philadelphia, PA, USA; CHU de and high-grade treatment-related AEs support pembrolizumab Québec-Université Laval, Québec, QC, Canada; Asan Medical Center as a new standard of care for advanced urothelial cancer that and University of Ulsan College of Medicine, Seoul, South Korea; progressed on/after platinum-based chemotherapy. University of California, San Francisco, San Francisco, CA, USA; Trial registration Comprehensive Cancer Centers of Nevada, Las Vegas, NV, USA; ClinicalTrials.gov identifier: NCT02256436 Fundación Instituto Valenciano de Oncología, Valencia, Spain; Smilow Cancer Hospital at Yale University, New Haven, CT, USA; Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy; Radboud University Medical Center, Nijmegen, Netherlands; Westmead Hospital and Macquarie University, Sydney, NSW, Australia; Univeristy of Southern California Norris Comprehensive 14 Table 1 (abstract O2) Efficacy in KEYNOTE-045 Cancer Center and Hospital, Los Angeles, CA, USA; Hôpital Saint- Louis, Paris, France; San Camillo Forlanini Hospital, Rome, Italy; End point Pembrolizumab Chemotherapy 16 17 Merck & Co., Inc., Kenilworth, NJ, USA; Memorial Sloan Kettering N = 270 N = 272 Cancer Center, New York, NY, USA OS, no. of events 155 179 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O2 Median (95 % CI), months 10.3 (8.0-11.8) 7.4 (6.1-8.3) Background HR (95 % CI) 0.73 (0.59-0.91); P = 0.0022 There is no standard second-line therapy for advanced urothe- PFS, no. of events 218 219 lial cancer. Although paclitaxel, docetaxel, and vinflunine are commonly used, they provide limited clinical benefit. KEYNOTE- Median (95 % CI), months 2.1 (2.0-2.2) 3.3 (2.3-3.5) 045 compared the efficacy and safety of the anti–PD-1 antibody HR (95 % CI) 0.98 (0.81-1.19); P = 0.42 pembrolizumab versus investigator-choice chemotherapy as second-line therapy for advanced urothelial cancer that pro- ORR (95 % CI) 21.1 % (16.4-26.5) 11.4 % (7.9-15.8) gressed or recurred following first-line platinum-based Treatment difference, % (95 % CI) 9.6 (3.5-15.9); P = 0.0011 chemotherapy. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 225 of 242 O3 Conclusions Efficacy and safety of nivolumab plus ipilimumab in metastatic Second-line treatment with N1I3 may provide the most favorable urothelial carcinoma: first results from the phase I/II CheckMate benefit-risk ratio among the regimens studied. If these interim results 032 study are confirmed with longer follow-up, further development of the 1 2 3 Padmanee Sharma , Margaret K Callahan , Emiliano Calvo , Joseph W N1I3 combination in metastatic urothelial carcinoma is warranted. 4 5 6 7 8 Kim , Filipo de Braud , Patrick A Ott , Petri Bono , Rathi N Pillai , Michael Trial Registration 9 10 11 12 Morse , Dung T Le , Matthew Taylor , Pavlina Spilliopoulou , Johanna ClinicalTrials.gov identifier NCT01928394. 13 14 15 16 Bendell , Dirk Jaeger , Emily Chan , Scott J Antonia , Paolo A 17 18 18 18 Ascierto , Delphine Hennicken , Marina Tschaika , Alex Azrilevich , References Jonathan Rosenberg 1. Sharma P, Bono P, Kim JW, et al: Efficacy and safety of nivolumab University of Texas MD Anderson Cancer Center, Houston, TX, USA; monotherapy in metastatic urothelial cancer (mUC): Results from the 2 3 Memorial Sloan Kettering Cancer Center, New York, NY, USA; START phase I/II CheckMate 032 study. J Clin Oncol 2016, 34(15 suppl): Madrid, Centro Integral Oncológico Clara Campal, Madrid, Spain; Yale Abstract 4501. Cancer Center, New Haven, CT, USA; Istituto Nazionale dei Tumori- Università degli Studi di Milano, Milano, Lombardia, Italy; Dana-Farber Cancer Institute, Boston, MA, USA; Comprehensive Cancer Center, Coinhibition & Costimulation Helsinki University Hospital and University of Helsinki, Helsinki, Finland; 8 9 Emory Winship Cancer Institute, Atlanta, GA, USA; Duke University O4 Medical Center, Durham, NC, USA; Sidney Kimmel Comprehensive Computational identification, functional characterization, and Cancer Center at Johns Hopkins University, Baltimore, MD, USA; antibody blockade of a new immune checkpoint in the TIGIT 11 12 Oregon Health & Science University, Portland, OR, USA; Beatson West family of interacting molecules 1 2 1 2 of Scotland Cancer Centre, Glasgow, United Kingdom; Sarah Cannon Ofer Levy , Christopher Chan , Gady Cojocaru , Spencer Liang , Eran 1 3 1 2 1 Research Institute and Department of Medical Oncology, Tennessee Ophir , Sudipto Ganguly , Amir Toporik , Maya Kotturi , Tal Fridman Kfir , 3 2 1 2 Oncology, Nashville, TN, USA; Heidelberg University Hospital, Benjamin M. Murter , Kathryn Logronio , Liat Dassa , Ling Leung , Shirley 1 1 2 1 4 Heidelberg, Baden-Wurttemberg, Germany; Vanderbilt-Ingram Greenwald , Meir Azulay , Sandeep Kumar , Zoya Alteber , Xiaoyu Pan , 1 1 2 1 University Medical Center, Nashville, TN, USA; H. Lee Moffitt Cancer Arthur Machlenkin , Yair Benita , Andrew W. Drake , Ayelet Chajut , Ran 1 1 1 2 1 Center, Tampa, FL, USA; Istituto Nazionale Tumori Fondazione Pascale, Salomon , Ilan Vankin , Einav Safyon , John Hunter , Zurit Levine , Mark Naples, Italy, Napoli, Italy; Bristol-Myers Squibb, Princeton, NJ, USA White 1 2 Correspondence: Padmanee Sharma (padsharma@mdanderson.org) Compugen Ltd., Holon, Israel; Compugen Inc, USA, South San Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O3 Francisco, CA, USA; Johns Hopkins University, Baltimore, MD, USA; Bloomberg ~ Kimmel Institute for Cancer Immunotherapy, Johns Background Hopkins University, Baltimore, MD, USA Nivolumab is a programmed death-1 (PD-1) immune checkpoint in- Correspndence: John Hunter (johnh@cgen.com) hibitor associated with clinical benefit in previously treated patients Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O4 with metastatic urothelial carcinoma [1]. Preclinical and clinical data indicate that the combination of nivolumab plus ipilimumab, an anti- Background cytotoxic T-lymphocyte antigen-4 (CTLA-4) antibody, can improve an- While antibody blockade of the CTLA-4 and PD-1 pathways has titumor activity in other tumor types. Here, we report the first effi- emerged as an effective treatment modality for cancer, the majority cacy and safety results of combined nivolumab plus ipilimumab of patients do not derive long-term benefit, suggesting a need for given at two different dosing schedules in CheckMate 032 , an open- targeting of additional immune checkpoints. Employing our unique label, multicenter, phase I/II study of patients with metastatic urothe- computational algorithms to define new members of the B7/CD28 lial carcinoma who progressed after prior platinum-based therapy. family, we identified PVRIG, which is expressed by multiple subsets Methods of T and NK cells. We report here its expression pattern, functional Patients with locally advanced or metastatic urothelial carcinoma pre- characterization, and anti-tumor activity of blocking antibodies tar- viously treated with platinum-based therapy were included in the geting this molecule. study. Patients were treated with either of two combination sched- Methods ules, nivolumab 1 mg/kg + ipilimumab 3 mg/kg (N1I3) or nivolumab Utilizing Compugen’s Predictive Discovery platform we identified 3 mg/kg + ipilimumab 1 mg/kg (N3I1) every 3 weeks for four cycles, PVRIG as a potential novel immune checkpoint, after which a retro- followed by nivolumab 3 mg/kg every 2 weeks; or they were treated viral cell screening library was used to identify its cognate binding with nivolumab monotherapy 3 mg/kg (N3) every 2 weeks. All pa- counterpart. Target effects on T cell modulation were assessed with tients were treated until disease progression or unacceptable toxicity. primary and tumor-derived T cell assays, taking advantage of target The primary endpoint was investigator-assessed objective response overexpression, knockdown, and antagonist antibody approaches. rate (ORR) by RECIST v1.1. Secondary endpoints included safety and Antibodies against the human protein were screened for their ability duration of response (DoR). to enhance T cell activation in vitro, while antibodies targeting the Results mouse orthologue were assessed in vivo for effects on tumor growth Minimum follow-up was 3.9 months in the N1I3 (n = 26) group, inhibition in syngeneic models. 14.5 months in the N3I1 group (n = 104), and 13.8 months in N3 Results group (n = 78). ORR was 38.5% (95% confidence interval [CI], 20.2- A PVRIG-Fc-fusion protein was found to bind PVRL2, with binding 59.4), 26.0% (95% CI, 17.9-35.5), and 25.6% (95% CI, 16.4-36.8) in the specificity confirmed both by ELISA and flow cytometry analysis. N1I3, N3I1, and N3 groups, respectively. Median DoR has not been PVRIG demonstrated unique expression kinetics upon T cell activa- reached in any treatment group. The frequency of drug-related grade tion, with detection of the target on memory T cells, as well as on 3-4 adverse events was 30.8% (N1I3), 31.7% (N3I1), and 23.1% (N3). NK cells and γδ T cells. A panel of high affinity human antibodies Treatment-related adverse events led to discontinuation in 7.7% with the ability to block interaction of PVRIG with PVRL2 were gener- (N1I3), 13.5% (N3I1), and 3.8% (N3) of patients. One death was re- ated, which when tested in vitro were shown to enhance activation ported in the N3I1 group (pneumonitis) and two deaths were re- of both primary CD4+ and tumor-derived CD8+ T cells through a ported in the N3 group (pneumonitis and thrombocytopenia). PVRL2-dependent mechanism. The lead antibody, COM-701, is Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 226 of 242 currently in preclinical development. Since COM-701 is not mouse Results cross-reactive, in vivo studies were conducted with a surrogate block- As of the August 30, 2016 data cutoff, 159 patients were treated with ing anti-mouse PVRIG antibody. When combined with anti-PD-L1 the lirilumab plus nivolumab combination. Treatment-related adverse blockade, anti-mouse PVRIG inhibits growth of established tumors in events (TRAEs) and grade 3–4 TRAEs were reported in 72% (114/159) both the CT26 and MC38 colorectal cancer models. Combination and 15% (24/159) of patients, respectively. Discontinuations due to testing with additional immune checkpoint inhibitors, as well as in TRAEs occurred in 8% (12/159). Of the 41 patients with SCCHN PVRIG knockout mice, is ongoing. treated, 29 were evaluable for response. In this heavily pretreated, Conclusions checkpoint inhibitor–naïve group, ORR was 24% (7/29; confirmed We describe the identification of PVRIG as a novel immune check- and unconfirmed) and DCR was 52% (15/29). Maximum reduction in point on T cells, as well the development of a high affinity antagonis- target lesions are presented in Fig. 2 for 26 patients with available tic antibody, COM-701, that is currently in preclinical development. tumor assessments. Two patients classified as stable disease per COM-701 is able to enhance human T cell activation, and a surrogate RECIST v1.1 showed unconventional responses, with 100% and 37% antibody with similar characteristics shows synergy with PD-L1 reductions in target lesions. Among evaluable patients, five (17%) in vivo in multiple syngeneic models. Overall, our data demonstrate had reductions in tumor burden > 80%. Responses appear durable, the utility of targeting PVRIG in addition to other B7 family check- with the median DOR not reached (Fig. 3). Updated efficacy and pre- points for the treatment of cancer. liminary biomarker analyses (including PD-L1 and HPV status) will be presented. Conclusions Combinations: Immunotherapy/ Preliminary efficacy of lirilumab plus nivolumab in patients with ad- vanced platinum-refractory SCCHN demonstrates clinical benefit, with Immunotherapy encouraging response rates that were deep and durable responses O5 in some patients. This combination demonstrated a manageable Preliminary efficacy from a phase I/II study of the natural killer safety profile similar to that observed with nivolumab monotherapy. cell–targeted antibody lirilumab in combination with nivolumab in Further evaluation of this novel combination of an NK-cell inhibitor squamous cell carcinoma of the head and neck and an immune checkpoint inhibitor is ongoing. 1 2 3 4 Rom Leidner , Hyunseok Kang , Robert Haddad , Neil H Segal , Lori J Trial Registration 5 6 3 7 Wirth , Robert L Ferris , F Stephen Hodi , Rachel E Sanborn , Thomas F ClinicalTrials.gov identifier: NCT01714739 8 9 10 10 Gajewski , William Sharfman , Dan McDonald , Shivani Srivastava , 10 10 10 8 Xuemin Gu , Penny Phillips , Chaitali Passey , Tanguy Seiwert Earle A. Chiles Research Institute, Robert W. Franz Cancer Center, Providence Portland Medical Center, Portland, OR, USA; Johns Hopkins Sidney Kimmel Comprehensive Cancer Center, Baltimore, MD, USA; 3 4 Dana-Farber Cancer Institute, Boston, MA, USA; Memorial Sloan Kettering Cancer Center, New York, NY, USA; Massachusetts General Hospital, Boston, MA, USA; University of Pittsburg, Pittsburgh, PA, USA; Earle A. Chiles Research Institute, Providence Cancer Center, Portland, 8 9 OR, USA; University of Chicago Medical Center, Chicago, IL, USA; The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Lutherville, MD, USA; Bristol-Myers Squibb, Princeton, NJ, USA Correspondence: Rom Leidner (rom.leidner@providence.org) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O5 Fig. 2 (abstract O5). Maximum percent reduction in target lesions Background from baseline Natural killer (NK) cells and the innate immune system play a critical role in immunosurveillance, control of tumor growth, and metastasis. NK-cell activation is negatively regulated by inhibitory killer-cell immunoglobulin-like receptors (KIRs); therefore, blocking KIR function may potentiate an anti-tumor immune response and complement other immuno-oncology therapies that enhance T cell activity. We present preliminary efficacy results in patients with squamous cell carcinoma of the head and neck (SCCHN) from a phase I/II study of lirilumab, a fully human monoclonal antibody that blocks inhibitory KIRs on NK cells, in combination with nivolumab, a fully human IgG4 monoclonal antibody that targets the PD-1 receptor, in patients with solid tumors (NCT01714739). Methods During dose escalation, patients with advanced solid tumors who progressed after ≥ 1 prior therapy received lirilumab 0.1–3.0 mg/kg once every 4 weeks (Q4W) plus nivolumab 3.0 mg/kg Q2W. Cohort expansion was initiated at the maximum dose of lirilumab 3.0 mg/kg Q4W plus nivolumab 3.0 mg/kg Q2W in patients with advanced solid tumors. Key study endpoints include safety (primary), objective re- Fig. 3 (abstract O5). Percent change from baseline in target lesions sponse rate (ORR), disease control rate (DCR), duration of response over time (DOR), and biomarker assessments. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 227 of 242 Tumor Microenvironment References 1. Daud AI, Loo K, Pauli ML, Sanchez-Rodriguez R, Sandoval PM, Taravati K, et al: Tumor immune profiling predicts response to anti–PD-1 therapy Phase II study of intratumoral plasmid interleukin 12 (pIL-12) with in human melanoma. J Clin Invest 2016, 126(9):3447–3452. electroporation in combination with pembrolizumab in stage III/IV melanoma patients with low tumor infiltrating lymphocytes 1 1 1 1 Alain Algazi , Katy Tsai , Michael Rosenblum , Prachi Nandoskar , Robert Adoptive Cellular Therapy 2 1 3 3 HI Andtbacka , Amy Li , John Nonomura , Kathryn Takamura , Mary 3 3 3 3 3 Dwyer , Erica Browning , Reneta Talia , Chris Twitty , Sharron Gargosky , P1 3 4 4 Jean Campbell , Carmen Ballesteros-Merino , Carlo B. Bifulco , Bernard Chemo-immunotherapy with cyclophosphamide and tumor 4 5 3 1 Fox , Mai Le , Robert H Pierce , Adil Daud reactive CD4+ T cells lead to destruction of tumor vasculature and University of California, San Francisco, San Francisco, CA, USA; eventual tumor eradication Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, USA; Tsadik Habtetsion, Gang Zhou 3 4 OncoSec Medical Inc., San Diego, CA, USA; Robert W. Franz Cancer Augusta University, Augusta, GA, USA Research Center, Earle A. Chiles Research Institute, Providence Cancer Correspondence: Tsadik Habtetsion (tsadikg855@yahoo.com) Center, Portland, Oregon, USA; Doctor Hope, LLC, San Diego, CA, USA Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P1 Correspondence: Sharron Gargosky (sgargosky@oncosec.com) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):06 Background CD4 T cells are critical components of anti-tumor immunity and play Background a pivotal role in orchestrating anti-tumor immune responses. Mount- Low tumor infiltrating lymphocytes (TIL) are predictive for poor ing evidence from preclinical and clinical studies indicates that CD4 response to immunotherapy with anti-PD-1/PD-L1 agents. We T cells in combination with chemotherapy can control tumor growth have shown that melanoma patients with a low frequency of PD- and recurrence. CD4 T cells are suggested to mediate tumor rejec- 1hiCTLA-4 + TIL are unlikely to respond to pembrolizumab tion through mechanisms that include cytotoxic effects on tumor (Daud2016). Intratumoral electroporation of pIL-12 (IT-pIL12-EP) cells, inhibition of angiogenesis, and reprogramming of the tumor leads to an IFN-g signature suggestive of increased TIL as well as microenvironment. regression in both treated and untreated lesions. We hypothesize Methods that combination IT-pIL12-EP and pembrolizumab will improve In this project, we set out to study the cellular and molecular mecha- clinical outcomes in this low-response population. Preliminary re- nisms underlying the therapeutic effect of chemo-immunotherapy in sults from a multi-center, phase II, open-label trial testing this hy- the form of cyclophosphamide (CTX) and tumor specific CD4 T cells pothesis are presented. in a murine model of colorectal cancer. Mice were injected subcuta- Methods neously with colorectal cancer cells. When the tumor reached 140- Melanoma stage III/IV patients with accessible lesions were con- 160 mm in area, mice were injected with a low dose of cyclophos- sented and enrolled if they had a TIL status of hiCTLA-4+ in the phamide followed by adoptive transfer of tumor reactive CD4 T CD45 + CD8 + CD3+ gate by flow cytometry (FC). Patients were cells. treated with pembrolizumab (200 mg every 3 weeks) concurrently Results with IT-pIL12-EP on days 1, 5 and 8 every 6 weeks. Patients were In a murine model of colorectal cancer, we show that the combin- evaluated for overall response rate (ORR) every 12 weeks by ation therapy of CTX and tumor reactive CD4 T cells resulted in en- RECISTv1.1. Pre and post-treatment blood and tumor specimens were hanced necrosis of tumor cells in vivo, leading to eventual collected, and analyzed for immune phenotyping, gene expression, eradication of advanced tumors. By using immunofluorescence stain- TCR diversity, and changes in the tumor microenvironment with ing and blood perfusion imaging, we demonstrated that the combin- multispectral immunohistochemistry. ation therapy leads to destruction of the established tumor Results vasculature and reduced blood supply to tumor tissue. Furthermore, Interim ORR data is available on 15 patients. 13/15 patients had we assessed blood vessel permeability in the tumor tissue and found a frequency of PD-1hi CTLA-4+ TIL of < 22% (low TIL status), that the combination therapy increased extravasation of Evans blue phenotypes associated with a low probability of response to dye, suggesting an increase in vascular permeability. anti-PD-1 (Daud 2016). These 15 patients age 39-89 years, were Conclusions 53% male, 66% stage III and 34% stage IV. Treatment was well In summary, our findings suggest that the combination therapy of tolerated; 38% of adverse events (AE) were classified as treat- CTX + CD4 T cells leads to destruction of the tumor vasculature, ment site reactions (grade 1-2) that resolved. One SAE of cellu- resulting in extensive necrosis of tumor tissue and eventual tumor re- litis resolved with 5d antibiotics. One grade 3 AE of diarrhea gression. These findings may provide new insights into mechanisms resolved with corticosteroids. The ORR was 40% (4CR, 2PR) by of tumor rejection by CD4 T cells. RECISTv1.1. Analysis of tumor biopsies and blood demonstrated meaningful immunological changes including an increased num- ber and ratio of CD8+:PD-L1+ and CD8+:FoxP3 + TIL, tumoral P2 RNA signatures indicating an increase in CD8 and IFN-γ-related Preclinical development of tumor-infiltrating lymphocyte therapy gene expression and concordant immune phenotypes in the for pancreatic cancer periphery. Donastas Sakellariou-Thompson, Cara Haymaker, Caitlin Creasy, Mark Conclusions Hurd, Naohiro Uraoka, Jaime Rodriguez Canales, Scott Koptez, Patrick The combination IT-pIL12-EP with pembrolizumab in patients with Hwu, Anirban Maitra, Chantale Bernatchez an anti-PD-1 non-responsive phenotype engendered a 40% clinical University of Texas MD Anderson Cancer Center, Houston, TX, USA response with associated positive immune-based biomarker data Correspondence: Donastas Sakellariou-Thompson and an excellent safety profile. These data suggests that IT-pIL12-EP (dasakellariou@mdanderson.org) modulates the tumor microenvironment to enable an effective anti- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P2 PD-1 mAb response in patients otherwise unlikely to respond. Background Acknowledgements Immunotherapy has become an effective cancer therapy, particularly We thank Merck and Oncosec for supporting this trial with pembrolizumab in the case of checkpoint blockade and adoptive T cell therapy (ACT). and IT-pIL-12, respectfully. ACT exploits the presence of tumor-infiltrating lymphocytes (TIL) by Trial Registration exponentially expanding their numbers ex vivo and re-infusing them ClinicalTrials.gov identifier: NCT02493361 into the patient in an autologous setting. With the effectiveness of Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 228 of 242 TIL therapy already well established in multiple phase II studies in Results melanoma, there is a push to translate it to other cancers in dire Here we describe advances in our ON-switch CAR design that allow need of improved therapies. Pancreatic ductal adenocarcinoma for dose-dependent antigen-specific T cell activation in the presence (PDAC) is one such cancer for which the current therapy, surgery and of an FDA-approved non-immunosuppressive small molecule dimeri- chemotherapy, provides an overall 5-year survival rate of only 5%. zer. Furthermore, we demonstrate that our synNotch T cells are able The presence of TIL is correlated with increased survival in PDAC, to deliver therapeutic payloads capable of, but not limited to, modu- which suggests that TIL could effectively control the disease and pro- lating the tumor microenvironment and changing the cell-intrinsic vides a rationale to test TIL therapy in this setting. transcriptional properties of the synNotch T cells. Methods Conclusions To assess the feasibility, we characterized the immune component of We have successfully identified a heterodimerizing switch receptor PDAC, explored the ability to grow and expand TIL from tumor frag- that specifically activates T cells in the presence of an FDA-approved ments, and analyzed the clonality of these expanded TIL. non-immunosuppressive small molecule. The ability to control the Results potency of the CAR-mediated immune response in this way may re- Flow cytometry analysis detected low, CD4-rich T cell infiltration. These duce the toxicity associated with CAR-T therapy. Furthermore, we TIL were able to be expanded ex vivo and the addition of an agonistic have demonstrated that synNotch T cells are able to sculpt the anti- anti-41BB antibody to the cultures preferentially increased total TIL out- tumor immune response in both a T cell intrinsic (transcriptional pro- growth, particularly that of CD8 TIL. The success rate of TIL growth graming) and T cell extrinsic (therapeutic payload) manner, providing was increased from 23% to 50% for cultures grown without and with a customizable platform for altering T cell function and the tumor anti-41BB respectively. Sequencing of the T cell receptor CDR3-beta microenvironment. chain found specific T cell clones enriched at the tumor site in compari- son to the blood. IHC staining for MHC class I (MHCI) on PDAC tumor References samples showed that it is widely expressed but at low levels generally. 1. Wu C-Y, Roybal KT, Puchner EM, Onuffer J, Lim, WA: Remote control of Conclusions therapeutic T cells through a small molecule-gated chimeric receptor. In conclusion, it is possible to expand CD8 T cells from PDAC bear- Science 2015 350: aab4077. ing TCR sequences highly enriched in the tumor. Additionally, ex- 2. Roybal KT, Rupp LJ, Morsut L, Walker WJ, McNally KA, Park JS, Lim W: panded TIL would be able to target tumor cells as they are shown to Precision Tumor Recognition by T Cells With Combinatorial Antigen- express MHCI. Although there are barriers yet to overcome, the initial Sensing Circuits. Cell 164: 770–779. data suggest the feasibility of TIL therapy for PDAC. Biomarkers and Immune Monitoring P3 P4 Synthetic biology approaches to enhance adoptive cell therapy Evaluation of anticancer immunity in patients with thyroid cancer safety and precision with a focus towards developing effective combination 1 1 1 1 Scott M Coyle , Kole T Roybel , Levi J Rupp , Stephen P Santoro , immunotherapy 1 1 1 1 1 1 2 Stephanie Secrest , Michael Spelman , Hanson Ho , Tina Gomes , Tiffany Tarsem Moudgil , Carmen Ballesteros-Merino , Traci Hilton , Christopher 1 2 3 3 1 2 3 4 4 1 Tse , Chia Yung-Wu , Jack Taunton , Wendell Lim , Peter Emtage Paustian , Rom Leidner , David Page , Walter Urba , Bernard Fox , Bryan 1 2 3 1 Cell Design Labs, San Francisco, CA, USA; Amgen, San Francisco, CA, Bell , Ashish Patel 1 2 USA; University of California San Francisco, San Francisco, CA, USA Providence Portland Medical Center, Portland, OR, USA; UbiVac, Correspondence: Stephen P Santoro (steve@celldesignlabs.com) Portland, OR, USA; Earle A. Chiles Research Institute, Robert W. Franz Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P3 Cancer Center, Providence Portland Medical Center, Portland, OR, Portland, OR, USA; Earle A. Chiles Research Institute, Providence Cancer Background Center, Portland, OR, USA Chimeric antigen receptor T cells (CAR-T) have shown impressive effi- Correspondence: Bernard Fox (foxb@foxlab.com) cacy against numerous hematological malignancies, yet a high per- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P4 centage of individuals receiving these therapies experience toxicity in the form of cytokine release syndrome (CRS) and/or normal tissue Background destruction. Furthermore, solid tumors represent a substantive chal- Thyroid cancer is the most common endocrine-related cancer with lenge for CAR therapy due to a lack of tumor- specific antigens and 64,330 diagnoses expected this year. While the majority of these can- general inability of T cells to overcome immunosuppressive tumor cers are curable, almost 2% of these cancers are anaplastic thyroid microenvironments. We have sought to circumvent these obstacles cancers, which are highly aggressive and almost uniformly lethal. At by utilizing synthetic biology approaches to augment CAR-T function the same time, the thyroid is known for being inherently immuno- and specificity. genic. For these reasons and due to an active surgical practice pro- Methods viding regular resections of thyroid cancers, we undertook a study of We developed two platform technologies that aim to mitigate tox- thyroid cancer with the idea of developing an immunotherapy for icity associated with CAR-T therapy and endow T cells with environ- this disease. mental sensing capabilities that enhance tumor discrimination from Methods normal tissue and/or confer the ability to generate customizable re- We have developed a thyroid cancer tumor bank to complement our sponse outputs. Firstly, we engineered an “ON-switch” CAR that con- Oral, Head and Neck Cancer Program. This tumor bank cryopreserves sists of two protein modules that undergo heterodimerization and enzymatically isolated viable cells from resected tumors (n = 16).We become competent for signaling only in the presence of a small- are also attempting to develop primary cell lines and are isolating molecule dimerizing agent [1]. In addition, we created a “synthetic and assessing autologous tumor-specific functions of tumor- Notch” (synNotch) receptor, which we previously described in the infiltrating lymphocytes (TIL) (n = 7). context of combinatorial antigen sensing [2], that is capable of driv- Results ing expression of any number of downstream polypeptides in re- To date we have established 3 tumor cell lines from thyroid cancer sponse to antigen engagement. specimens and identified PD-L1 expression on 2 of 2 tested. While Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 229 of 242 numbers are small, preliminary analyses suggest that TIL cultures can (2.1 mSv/MBq) and LLI wall at 6.5 rem/mCi (1.8 mSv/MBq). A GLP be generated from 85% of thyroid cancer specimens and that autolo- toxicology study was conducted in cynomolgus monkeys that in- gous tumor-reactive TIL can be detected in 43% (n = 7) of thyroid cluded multiple dose cohorts of Zr-Df-IAB22M2C and a vehicle con- cancers. Since not every tumor appears to contain TIL capable of rec- trol. The results showed that doses up to 25 mg/kg of Zr-Df- ognizing autologous tumor, strategies to prime tumor-specific T cells IAB22M2C administered weekly to cynomolgus monkeys did not re- represents an area of interest. DPV-003 is a microvesicle vaccine, sult in any treatment-related findings in survival, clinical signs, body DRibble, that contains more than 80 proteins that are overexpressed weights, food consumption, ophthalmic examinations, electro- by thyroid cancer (TCGA provisional RNASeq n = 509 pts). The vac- cardiology, blood pressure, heart rate, clinical and anatomic path- cine also contains a number of DAMPs and agonist activity for mul- ology, peripheral blood lymphocyte population, and cytokine levels. tiple TLRs packed into stable double membrane microvesicles that Conclusions are targeted to CLEC9A+ antigen presenting cells. We are also devel- Zr-Df-IAB22M2C has the desired sensitivity and safety profile for im- oping a second thyroid-specific DRibble vaccine from a cell line de- aging CD8+ T cells and the first-in-human studies will commence in rived from an anaplastic thyroid cancer. the Q4 2016. Conclusions Almost half of thyroid cancers evaluated, including one anaplastic thyroid cancer, contain T cells capable of recognizing autologous P6 cancer cells and secreting IFN-g. However, the other 50% of thyroid High dose interleukin- 2 (HD IL-2) select trial in melanoma: a tissue cancers appear to lack tumor-reactive T cells and may benefit from and blood collection protocol to identify predictive biomarkers of combination immunotherapy strategies that include a vaccine. benefit to HD IL-2 in patients with advanced melanoma 1 2 3 4 Ryan J Sullivan , Yujin Hoshida , Theodore Logan , Nikhil Khushalani , 5 6 7 8 Acknowledgements Anita Giobbie-Hurder , Kim Margolin , Joanna Roder , Rupal Bhatt , 9 10 11 12 Support: Steve and Cindy Harder, Robert W. and Elsie Franz, Wes and Nancy Henry Koon , Thomas Olencki , Thomas Hutson , Brendan Curti , 13 8 14 7 Lematta, Lynn and Jack Loacker, and The Chiles foundation (BAF). Shauna Blackmon , James W Mier , Igor Puzanov , Heinrich Roder , 15 16 14 17 John Stewart , Asim Amin , Marc S Ernstoff , Joseph I Clark , Michael 18 19 20 8 B Atkins , Howard L Kaufman , Jeffrey Sosman , Sabina Signoretti , P5 David F McDermott Development and clinical translation of 89Zr-Df-IAB22M2C for Medical Oncology Department, Massachusetts General Hospital, Boston, detecting CD8+ T Cells for immunotherapy applications MA, USA; Hess Center for Science and Medicine; Tisch Cancer Institute, 1 2 1 1 Tove Olafsen , Daulet Satpayev , Michael Torgov , Filippo Marchioni , New York, NY, USA; Simon Cancer Center, Indiana University, 1 1 1 Jason Romero , Ziyue Karen Jiang , Charles Zamilpa , Jennifer S Indianapolis, IN, USA; H. Lee Moffitt Cancer Center, Tampa, FL, USA; 1 1 1 1 1 Keppler , Alessandro Mascioni , Fang Jia , Chen-Yu Lee , Jean Gudas Department of Biostatistics & Computational Biology, Boston, MA, USA; 1 2 ImaginAb Inc., Inglewood, CA, USA; AdicetBio, Inc., Menlo Park, CA, Department of Medical Oncology, City Of Hope, Duarte, CA, USA 7 8 USA Biodesix, Inc., Boulder, CO, USA; Beth Israel Deaconess Medical Center, Correspondence: Jean Gudas (jgudas@imaginab.com) Boston, MA, USA Case Western Reserve University, Cleveland, OH, USA; 10 11 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P5 The Ohio State University, Columbus, OH, USA; Texas Oncology- Baylor Charles A. Sammons Cancer Center, Dallas, TX, USA; Earle A. Background Chiles Research Institute, Providence Cancer Center, Portland, OR, USA; Immunotherapies are changing the landscape for cancer treatment; how- Massachusetts General Hospital Cancer Center, Boston, MA, USA; 14 15 ever, the field is hampered by the lack of biomarkers that can be used Roswell Park Cancer Institute, Buffalo, NY, USA; Wake Forest Baptist for patient selection and for monitoring treatment responses rapidly and Medical Center, Winston Salem, NC, USA; Levine Cancer Institute, non-invasively. To address this need, ImaginAb is developing Zr-Df- Carolinas HealthCare System, Charlotte, NC, USA; Loyola University IAB22M2C, an ~80 kDa minibody (Mb) with high affinity to the CD8 Medical Center, Maywood, IL, USA; Georgetown-Lombardi glycoprotein (binding EC = 0.4 nM) conjugated with desferrioxamine Comprehensive Cancer Center, Washington , DC, USA; Rutgers Cancer (Df) and radiolabeled with the positron emitting radionuclide Zirconium- Institute of New Jersey, New Brunswick, NJ, USA; Robert Lurie 89 ( Zr; T 78.4 hours) for imaging CD8+ T cells in humans. Comprehensive Cancer Center of Northwestern University, Chicago, IL, USA 1/2 Methods Correspondence: Ryan J Sullivan (rsullivan7@mgh.harvard.edu) A comprehensive preclinical program that included evaluation of the Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P6 in vitro and in vivo pharmacodynamics of IAB22M2C (unconjugated Mb), Df-IAB22M2C (conjugated Mb intermediate), Zr-Df-IAB22M2C (Zr Background chelated, conjugated non-radiolabeled form of final drug) and Zr- HD IL-2 provides objective responses in 15-20% and durable complete Df-IAB22M2C (radioactive final drug product) was conducted to dem- remission in 5-8% of patients with metastatic melanoma (MM). We pre- onstrate the safety and potential efficacy of the probe. viously identified a gene expression-based tumor subclass character- Results ized by immune related genes (Class 2; C2) associated with durable In vitro studies using human PBMCs from 10 individual human do- response to HD IL-2 compared to the remaining tumors that overex- nors showed no measurable or reproducible impact on proliferation, pressed lineage-associated genes (Class 1; C1). The primary objective of activation or depletion of CD8+ T cells and no consistent release of the HD IL-2 select trial in melanoma was to prospectively validate the cytokines when donor CD8+ T cells were exposed to soluble or favorable gene expression signature (C2). Secondary objectives were to immobilized Mb protein. Studies that evaluated the effect of saturat- seek serum and tissue biomarkers of durable response. ing concentrations of Zr-Df-IAB22M2C on proliferation and viability Methods of CD8+ T cells in vitro, also showed no impact on these parameters. 170 patients with MM were enrolled at 15 Cytokine Working Group Preclinical imaging and biodistribution studies demonstrated favor- sites from 2010 to 2014. All patients had formalin-fixed paraffin- able pharmacokinetics and the ability of Zr-Df-IAB22M2C to detect embedded (FFPE) tumor tissues identified and blood drawn prior to infiltrating CD8+ T cells in a mouse hu-PBMC NSG™ GvHD model and HD IL-2. Tumor assessments used WHO criteria and investigator- TM in Matrigel plugs implanted with different numbers of human CD8 assessed outcomes. RNA extracted from FFPE tumor tissues was used TM + T cells. Radiation dosimetry studies conducted in hu-CD34 NSG for whole transcriptome profiling by RNA sequencing (114 samples mice and the results GLP dosimetry analysis showed that on average, yielded sufficient RNA, 101 passed default Quality Control (QC)). Pre- the organs receiving the largest dose equivalent were the kidneys at treatment serum from 114 patients served as the test set and was 8.0 rem/mCi (2.2 mSv/MBq) followed by the liver at 7.9 rem/mCi analyzed using matrix-assisted laser desorption/ionization (MALDI) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 230 of 242 and machine-based learning algorithms to identify a predictive pro- starting at week 6 for 4 infusions. Peripheral blood was obtained tein expression signature. (Paxgene RNA) at baseline and at weeks 4, 6, 9, and 15. Gene expres- Results sion (Agilent Microarray) was analyzed for changes in expression Thirty-one of 170 pts (18.2%) responded, and median overall survival level with treatment. Pharmacodynamic markers were identified with was 21.3 months, with a 40 month median follow-up. Analysis of a linear mixed effects model. False discovery was controlled with per- RNAseq from 101 patients whose specimens passed QC showed that mutation testing. a C2 signature was associated with response to HD IL-2 (normalized Results enrichment score 1.70, false discovery rate 0.004). Using MALDI, a Gene expression was measured in 16 patients in phase Ib. Most treat- protein expression signature enriched for acute phase proteins (in- ment effects on expression were seen after ipilimumab treatment, cluding CRP, IL-6, and SAA) was defined in the pre-treatment serum but there were a few effects in the initial T-VEC phase that passed and used to classify 39 patients into group A (non-acute phase pro- false discovery controls. These T-VEC effects included SELV, SYNPO, tein expression) and 75 patients in group B (acute phase protein ex- ZBTB32, IQCF2, CDC27, KLK1, PRR20B, CHST6, and IGH. ZBTB32 has pression). Complete response rate in group A was 21% and zero in been reported to control the proliferative burst of virus-specific nat- group B (p = 0.0001). Two-year PFS rate was 29% in group A com- ural killer cells responding to infection. The combination effects were pared to 4% in group B (p = 0.0005). enriched for genes involved in lymphoid tissue structure and devel- Conclusions opment and immune cell trafficking. 185 of these genes had signs of In this prospective biomarker validation study, HD IL-2 produced dur- a T-VEC effect in the monotherapy phase. These included increases able remissions and prolonged survival in patients with MM. A in GZMM, PDCD1, CD8B, CD8A, and CTLA4 and decreases in IL18, tumor-derived gene expression signature enriched for immune- IRAK3, and TXNRD1. related genes was associated with response. Additionally, preliminary Conclusions data with a serum protein signature appears to identify patients This hypothesis-generating microarray analysis identified genes up- most likely to have a complete response. regulated in circulating peripheral blood cells after T-VEC monother- Trial Registration apy and combination treatment. We plan to further evaluate these ClinicalTrials.gov identifier NCT01288963. genes and other potential pharmacodynamic markers in phase II. Trial Registration ClinicalTrials.gov identifier NCT01740297. P7 Pharmacodynamic gene expression changes from talimogene References laherparepvec (T-VEC) plus ipilimumab in a phase Ib study for 1. Puzanov I, Milhem MM, Minor D, Hamid O, Li A, Chen L, et al: metastatic melanoma Talimogene laherparepvec in combination with ipilimumab in 1* 2 3 Abraham A Anderson , Igor Puzanov , Mohammed M Milhem , Robert previously untreated, unresectable stage IIIB-IV melanoma. J Clin Oncol 4 5 1 1 HI Andtbacka , David Minor , Kevin S Gorski , Daniel M Baker , Omid 2016, 34:2619–2626. 6 7 Hamid , Howard L Kaufman 1 2 Amgen Inc., Thousand Oaks, CA, USA; Roswell Park Cancer Institute, Buffalo, NY, USA; University of Iowa Hospitals and Clinics, Iowa City, IA, Clinical Trials in Progress USA; University of Utah, Huntsman Cancer Institute, Salt Lake City, UT, 5 6 USA; California Pacific Melanoma Center, San Francisco, CA, USA; The P8 Angeles Clinic & Research Institute, Los Angeles, CA, USA; Rutgers Phase I study of alpha-tocopherlyoxyacetic acid in patients with Cancer Institute of New Jersey, New Brunswick, NJ, USA advanced cancer: immune response and pharmacokinetics results 1 2 2 Correspondence: Abraham A Anderson (andersoa@amgen.com) Emmanuel Akporiaye , Brendan Curti , Yoshinobu Koguchi , Rom 3 3 3 2 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P7 Leidner , Kim Sutcliffe , Kristie Conder , Walter Urba 1 2 Veana Therapeutics Inc, Portland, OR, USA; Earle A. Chiles Research Background Institute, Providence Cancer Center, Portland, OR, USA; Earle A. Chiles T-VEC is a herpes simplex virus type-1 based oncolytic immunother- Research Institute, Robert W. Franz Cancer Center, Providence Portland apy designed to selectively replicate in tumors, produce GM-CSF, and Medical Center, Portland, OR, USA stimulate antitumor immune responses. Ipilimumab is a checkpoint Correspondence: Emmanuel Akporiaye (etakporiaye@gmail.com) inhibitor that promotes T cell activation by blocking negative signal- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P8 ing through CTLA-4. Both agents have demonstrated activity as monotherapy in advanced melanoma. Based on the potential com- Background plementary MOA of the agents, tumor cell lysis and antigen presen- Alpha-tocopheryloxyacetic acid (α-TEA) targets tumor cell mitochon- tation (T-VEC) in combination with T cell checkpoint inhibition, we dria to release reactive oxygen species (ROS) that induce immuno- hypothesized that improved efficacy was possible when the agents genic cell death (ICD), antigen release, and enhanced antigen cross- are used in combination. Because the safety profiles are non- presentation in pre-clinical models. α-TEA is being evaluated for overlapping, the combination was not anticipated to have significant safety and tolerability in a first-in-human phase I trial in patients with increased toxicity. To address these hypotheses, a phase Ib/II study advanced cancers (NCT02192346). Tumor types in the ongoing trial evaluating the safety and efficacy of T-VEC plus ipilimumab for Stage include renal cancer, esophageal adenocarcinoma, thyroid cancer, III-IV metastatic melanoma was initiated. The phase Ib study was duodenal cancer, and squamous cell carcinoma of the head and completed (N = 19) with no DLTs (primary endpoint) or new safety neck. signals with combination treatment, and an ORR of 50% [1]. Phase Ib Methods also included biomarker analyses investigating potential pharmaco- α-TEA lysine salt is administered orally to patients and given daily in dynamic markers for T-VEC monotherapy and in combination with escalating doses for 28 days. Immune monitoring of peripheral whole ipilimumab. blood was conducted for all 12 patients at baseline, and at 1 week Methods and 4 weeks post-treatment. Plasma levels of α-TEA have been deter- 6 8 Nineteen patients received T-VEC at 10 PFU/mL at week 1, then 10 mined so far in patients receiving 2.4 mg/kg and 4.8 mg/kg α-TEA at PFU/mL Q2W from week 4. Ipilimumab was given at 3 mg/kg Q3W 1, 4, 8, and 24 hours after the first dose. Additional samples were Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 231 of 242 evaluated on days 8, 15, 22, and 29 before the planned α-TEA dose potential for cytolytic activity directed at normal tissues, and unin- on those days. tended toxicity in patients treated with MGD009. Results Methods Twelve patients have been treated so far at 2.4 mg/kg and 4.8 mg/ This multi-center, open-label trial is a phase I dose escalation/cohort kg dose levels. Eight patients have stable disease, lasting from 1 to expansion study. All patients must have advanced B7-H3-positive tu- 22+ months. One patient showed more than a 2-fold increase in the mors. Prior checkpoint inhibitor therapy will be allowed. Dose escal- number of activated (CD38+ HLA-DR+) effector CD8+ T cells 1 week ation uses a 3+3+3 design, with patients treated every 2 weeks with post-treatment. A second patient showed more than a 2-fold in- escalating doses of IV MGD009 (starting dose 0.3 ug/kg). The dose crease in the number of activated effector memory CD8+ T cells escalation phase enrolls patients with mesothelioma, bladder cancer, 4 weeks post-treatment. Both patients experienced stable disease melanoma, SCCHN, NSCLC, clear cell renal cell carcinoma, ovarian over 5 and 22 months, respectively. Evaluation of α-TEA levels at the cancer, thyroid cancer, triple-negative breast cancer, pancreatic can- 2.4 mg/kg and 4.8 mg/kg doses revealed a proportional increase in cer, colon cancer, soft tissue sarcoma, or prostate cancer. Cohort ex- α-TEA plasma levels over a 28-day interval without any indication pansions (n=16/cohort) treated at the maximum tolerated dose will that steady state plasma levels were reached. Of the 12 patients, 6 include patients with melanoma, SCCHN, mesothelioma, urothelial developed atrial fibrillation (AF) after starting α-TEA. The earliest cancer, and NSCLC. Pre- and on-study biopsies are required for mel- event occurred 7 days post-treatment, but AF was more common 29- anoma patients in the cohort expansion phase. Primary and second- 56 days post-treatment. Four of the 6 patients had a medical history ary study objectives include characterization of the safety, of AF. These were grade 2 events by CTCAE 4.0 criteria and managed pharmacokinetics, pharmacodynamics, and preliminary antitumor ac- with appropriate medication without further sequelae. tivity of MGD009. All tumor evaluations will be carried out by both Conclusions Response Evaluation Criteria in Solid Tumors (RECIST 1.1) and im- α-TEA treatment resulted in stable disease in 80% of patients lasting mune-related RECIST. between 1 and 22+ months. AF was observed commonly in patients Trial Registration with a medical history of AF, and was managed with appropriate ClinicalTrials.gov identifier NCT02628535. medication. No clinically meaningful grade 3 or 4 toxicities were ob- served. Plasma α-TEA levels increased proportionally without any in- dication that steady state levels were achieved. α-TEA may function P10 through enhancing pre-existing CD8+ T cell-mediated anti-tumor Intratumoral Flt3L and poly-ICLC combined with low dose activity. radiotherapy: a novel in situ vaccine for incurable indolent lymphomas 1 1 1 1 Thomas Marron , Nina Bhardwaj , Linda Hammerich , Fiby George , 1 2 2 2 P9 Seunghee Kim-Schulze , Tibor Keler , Tom Davis , Elizabeth Crowley , 3 4 Phase I, first-in-human, open label, dose escalation study of Andres Salazar , Joshua Brody 1 2 MGD009, a humanized B7-H3 x CD3 dual-affinity re-targeting Icahn School of Medicine at Mount Sinai, New York, NY, USA; Celldex (DART®) protein in patients with B7-H3-expressing neoplasms or Therapeutics, Hampton, NJ, USA Oncovir, Inc., Washington, DC, USA; B7-H3 expressing tumor vasculature Mount Sinai School of Medicine, New York, NY, USA 1 2 3 Anthony W Tolcher , Evan W Alley , Gurunadh Chichili , Jan E Correspondence: Thomas Marron (thomas.marron@mountsinai.org) 4 3 3 3 3 Baughman , Paul Moore , Ezio Bonvini , Syd Johnson , Sadhna Shankar , Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P10 3 3 5 James Vasselli , Jon Wigginton , John Powderly START - South Texas Accelerated Research Therapeutics, LLC, San Background 2 th Antonio, TX, USA; Penn Presbyterian Medical Center, University of Lymphomas are the 5 most common cancer in the United States, Pennsylvania, Philadelphia, PA, USA; MacroGenics, Inc., Rockville, MD, and unlike more aggressive lymphomas, indolent non-Hodgkin 4 5 USA; MacroGenics, Inc., South San Francisco, CA, USA; Carolina lymphoma (iNHL) and is incurable with standard therapy. A previous BioOncology Institute, PLLC, Huntersville, NC, USA trial of in situ vaccination in iNHL combined intratumoral injection of Correspondence: Jan E Baughman (baughmanj@macrogenics.com) a TLR9 ligand with radiation to induce a systemic immune response Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P9 against tumor. This approach induced tumor-specific CD8+ T cell re- sponses and durable clinical remissions of patients’ untreated sites of Background disease were seen in a minority of patients. One limitation in this MGD009 is a B7-H3 x CD3 dual affinity re-targeting (DART) protein. previous trial may have been the scarcity of intratumoral dendritic DART proteins are bispecific, antibody-based molecules that can bind cells (DCs); DCs are uniquely able to endocytose dying tumor cells two distinct antigens simultaneously. MGD009 is designed to redirect for cross-presentation to tumor antigen-specific CD8+ T cells. In our T cells to eliminate B7-H3-expressing target cells through co-engage- novel iteration of in situ vaccine, intratumoral FMS-like tyrosine kin- ment of B7-H3 on the target cell and CD3 on the T cell, each in a ase 3 ligand (Flt3L) is introduced as a priming step to increase the monovalent binding manner. MGD009 contains a mutated human presence of intratumoral DCs. Flt3L induced tumor leukocyte infiltra- IgG1 Fc domain to reduce/eliminate binding to Fc gamma receptors tion and regression in lymphoma tumors in pre-clinical trials, and (FcγRs) and complement, but retains binding to the neonatal Fc re- CDX-301- a formulation of Flt3L - was previously shown to mobilize ceptor (FcRn) enabling use of the IgG salvage pathway to prolong its BDCA-1 and BDCA-3 myeloid DC subsets in an early phase clinical tri- circulating half-life. B7-H3 protein expression is limited in normal hu- als. These DC subsets respond to several TLR agonists and cross- man tissues, but is overexpressed in a broad range of tumor types, present antigens more effectively than plasmacytoid DCs (pDCs). including melanoma, non-small cell lung cancer (NSCLC), squamous While pDCs are high expressers of TLR9, responsive to CpG, myeloid cell carcinoma of the head and neck (SCCHN), mesothelioma, and dendritic cells express a wider array of TLRs, including high levels of urothelial cancer. By binding to B7-H3 on tumor cells and CD3 on T TLR3. lymphocytes, MGD009 can recruit cytotoxic T cells, irrespective of Methods their ability to recognize cancer cells, and trigger T cell activation, ex- This phase I/II trial tests the hypothesis that this novel in situ vaccin- pansion, and T cell-mediated killing of B7-H3-expressing tumors. The ation will induce clinical remissions at distant (untreated) tumor sites limited expression of B7-H3 on normal cells should restrict the in two cohorts of patients with either previously untreated or Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 232 of 242 relapsed/refractory iNHL (n = 15 per group). Intratumoral CDX-301 experienced both grade 3 lymphopenia and grade 3 failure to thrive. 25ug/kg is injected into a palpable lymph node for 9 days, followed At twelve weeks post-treatment initiation, 83% of patients experi- by 2Gy local radiotherapy on day 9 and 10 to the target lymph node. enced response or disease stability. Three patients (50%) had absco- To activate local DCs, poly-ICLC 2 mg is injected on day 10, 14, 17, pal responses by RECIST criteria, two patients (33%) had stable and weekly thereafter for a total of 8 treatments. disease, and one patient (17%) had progressive disease. Results Conclusions Exploratory endpoints include measuring induction of systemic Pre-clinical data suggests that sequencing may be key to the success tumor-specific immune response in pre- and post-vaccine blood and of combinatorial strategies of PD-L1 blockade and RT. The safety run- tissue samples. Using flow cytometry and CyTOF, we have confirmed in for combining PD-L1 checkpoint inhibition with RT suggests that + + that CD1c + (BDCA1 ) and CD141+ (BDCA3 ) DCs home to treated tu- the combination is safe and tolerable in metastatic NSCLC. The trial mors following treatment with Flt3L and T cells attain a mature ef- will continue to accrual to evaluate different sequencing strategies fector phenotype. Tissue from initial bx is being sequenced, and for combining RT and PD-L1 checkpoint blockade. Further study is candidate neoantigens being determined in silico; these neoantigens needed to evaluate the efficacy and optimal sequencing of RT + PD- are then being synthesized and tested for potential to activate pa- L1 checkpoint blockade. tient pre- and post-vaccination T cells. Conclusions Combinations: Immunotherapy/ This trial is in process. Trial Registration Immunotherapy ClinicalTrials.gov identifier NCT01976585. P12 Tissue factor is a novel oncotarget in triple negative breast cancer P11 and BRAF- mutated melanoma for immunotherapy using a second Preliminary safety and efficacy data for radiotherapy and PD-L1 generation ICON (L-ICON) in monotherapy and combination checkpoint blockade in metastatic non-small cell lung cancer: is therapy 1 2 2 2 timing everything? Zhiwei Hu , Rulong Shen , Amanda Campbell , Elizabeth McMichael , 1 2 2 2 2 2 2 3 Arta Monjazeb , Megan E Daly , Jonathan Riess , Tianhong Li , William J Lianbo Yu , Bhuvaneswari Ramaswam , Cheryl A London , Tian Xu , 1 2 Murphy , Karen Kelly William Carson 1 2 University of California, Davis, Sacramento, CA, USA; University of The Ohio State University Wexner Medical Center and The OSU James California Davis Comprehensive Cancer Center, Sacramento, CA, USA Comprehensive Cancer Center, Columbus, OH, USA; The Ohio State Correspondence: Arta Monjazeb (ammonjazeb@ucdavis.edu) University, Columbus, OH, USA; Yale University, New Haven, CT, USA Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P11 Correspondence: Zhiwei Hu (zhiwei.hu@osumc.edu) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P12 Background Inhibition of the PD-1/PD-L1 checkpoint pathway can induce rapid Background and durable responses in patients with non-small cell lung cancer The objective of this study is to identify tissue factor (TF) as a novel (NSCLC). Unfortunately the majority of patients fail to respond and oncotarget for triple negative breast cancer (TNBC) and BRAF mu- there is interest in exploring combinatorial strategies to improve re- tated melanoma, both of which are very difficult to treat in clinic, sponse rates. One such strategy is combining checkpoint inhibition and to develop a novel TF-targeting agent for immunotherapy. To with radiotherapy (RT). We report here our pre-clinical data for com- achieve this goal, Hu developed a second generation TF-targeting bining radiotherapy with PD-L1 checkpoint blockade. These data ICON, named L-ICON, which consists of only the light chain (1-152 demonstrate a clear influence of the sequencing of combinatorial aa) of FVII fused to an IgG1Fc. The effects of L-ICON were evaluated therapy on its efficacy. Based on these data, we have initiated a clin- as monotherapy or combination therapy with interleukin 15 (IL-15) ical trial testing sequencing strategies of radiotherapy with PD-L1 in- for the malignancies. hibition in patients with metastatic NSCLC. Methods Methods TF expression was determined by immunohistochemistry or by flow Using syngeneic mouse tumor models, we tested the synergy of cytometry. L-ICON protein (GenBank accession no. KY760097) and combining RT with PD-L1 inhibition and the influence of the sequen- replication-deficient adenoviral vectors have been developed. Bind- cing of these therapies on efficacy. Based on this preclinical work, we ing activity of L-ICON was determined. Its ADCC effect was evaluated have initiated a phase II clinical trial testing this combinatorial strat- by an ADCC effector assay and coagulation activity by FVII chromo- egy with three cohorts. The three cohorts are concurrent therapy, genic activity assay. L-ICON efficacy in monotherapy and combin- radiotherapy followed by PD-L1 checkpoint blockade, and PD-L1 ation therapy with IL-15 was tested in mouse models of murine and blockade followed by radiotherapy. We report here the preliminary human breast cancer (4 T1 and TNBC MDA-MB-231) and melanoma safety, efficacy, and correlative science data from interim analysis of (B16F10 and BRAF mutated SK-Mel-28). the safety run-in for this trial. Results Results TF is over-expressed on TNBC cells and the tumor neovasculature In studies using syngeneic mouse tumor models, we find that PD-L1 in over 85% of TNBC patients (n = 14) when using standard inhibition provides no added benefit to radiotherapy alone when ad- paraffin-embedded tumor tissues or in nearly 60% of TNBC pa- ministered after radiotherapy. Conversely, priming of the immune tients (n = 157) when employing tissue microarray slides. Import- system with anti-PD-L1 prior to RT provides significant synergy of the antly, TF expression is not detected in normal breast tissues. combinatorial therapy. Our clinical trial has completed enrollment to L-ICON has several important improvements over its first gener- the safety-run in of 6 patients. In total, 2 patients experienced grade ation ICON, including (i) more than 50% reduction in molecular 3 dose limiting toxicities meeting the criteria for completion of the mass, (ii) complete elimination of coagulation activity, (iii) stron- safety-run in without the need for dose de-escalation. One patient ger binding activity to TNBC and (iv) more effective as monother- experienced asymptomatic grade 3 lymphopenia and one patient apy in vivo in orthotopic and subcutaneous mouse models of Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 233 of 242 human TNBC (MDA-MB-231) and murine cancer 4 T1 (an animal delay melanoma progression. Due to the availability of all three com- stage IV human breast cancer) and B16F10. L-ICON monotherapy ponents for use in humans, this therapeutic regimen can potentially and combination with IL-15 were effective for the treatment of be clinically translated to expand the population of metastatic mel- SK-Mel-28 in SCID mouse models. anoma patients who experience long term benefits from immune- Conclusions based therapies. TF is a novel biomarker and oncotarget in TNBC and BRAF- mutated melanoma. L-ICON, a novel TF-targeting ICON, was effective in mono- Acknowledgements therapy and combination therapy with IL-15 for the treatment of This work was supported by CURE Grant SAP #4100072562 (Pennsylvania murine and human TNBC and melanoma in vitro and in vivo in pre- Department of Health) and NIH/NCI 5 T32 CA060395 (KMK). IL-2 was clinical mouse models. generously provided by Prometheus Laboratories Inc. Acknowledgements This work was partly supported by a startup fund from OSUMC, a Seed Award P14 from the OSUCCC TT Program, a Phase 1 L-Pilot Award from OSU CCTS via Sequentially targeting upregulated TIM-3 and CTLA-4 does not NCATS Award Number UL1TR001070 and the Dr. Ralph and Marian Falk rescue the attenuated therapeutic efficacy of combination Medical Research Trust. IL-15 was obtained from the NCI Preclinical Repository. immunotherapy with OX40 costimulation and PD-1 blockade 1 2 2 Z.H. is the inventor of L-ICON and its uses (US Patent Application # 62/082,891). David J Messenheimer , Shawn Jensen , Bernard Fox 1 2 Earle A. Chiles Research Institute, Portland, OR, USA; Providence Cancer Center, Portland, OR, USA P13 Correspondence: David J Messenheimer (messenhe@ohsu.edu) Beta-adrenergic blockade improves the immunotherapeutic Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P14 response to melanoma 1 2 1 Kathleen M Kokolus , Elizabeth A Repasky , Todd D Schell , Joseph D Background Drabick Combination immunotherapy targeting checkpoint molecules has The Pennsylvania State University College of Medicine, Hershey, PA, shown substantial results against solid tumors. However as novel USA; Roswell Park Cancer Institute, Buffalo, NY, USA therapies targeting costimulatory molecules are introduced into the Correspondence: Kathleen M Kokolus (kokolusk@hmc.psu.edu) clinic, successful combination with checkpoint blockade remains un- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P13 certain. Some strongly immunogenic preclinical models have shown benefit when combining anti-OX40 and anti-PD-1 treatment. In con- Background trast, using a PD-1 refractory mammary tumor model we have dem- Recent developments in immunotherapy have made enormous onstrated that a significant anti-tumor effect generated with OX40 strides towards expanding the scope of cancer treatment by target- costimulation is significantly attenuated with the addition of PD-1 ing a patient’s own immune cells. Despite these advances, malignant blockade. We noted high levels of inflammatory cytokines in the melanoma remains a significant clinical issue with a high proportion serum of combination treated mice, and also saw a significant in- of patients remaining unresponsive to therapy and improved, but crease in inhibitory receptors TIM-3, LAG-3, and CTLA-4 on CD4 and still low, complete response rates. The body’s response to stress is CD8 T cells in the periphery of treated mice. We hypothesized that closely integrated with the immune response, yet few cancer treat- the upregulation of these other inhibitory receptors were limiting the ment strategies account for the relationship between these biological efficacy of anti-OX40 plus anti-PD-1 combination treatment, and systems. When the stress response is activated, neurotransmitters, tested whether sequentially blocking these receptors could augment including norepinephrine, which bind β-adrenergic receptors (βARs) therapeutic efficacy. located on the surface of immune cells, are released, leading to regu- Methods lation of various immune cell functions. βAR signaling can be pre- Established orthotopically transplanted MMTV-PyMT mammary tu- vented pharmaceutically with βAR antagonists (β-blockers) and mors in FVB/NJ mice were treated with three doses of anti-OX40 and considerable literature suggests that these drugs, which are com- anti-PD-1 every other day. Serum was taken at time points and monly prescribed for other indications including hypertension, are tested for cytokine concentration and spleens were taken two days associated with positive outcomes in cancer patients. We examined after treatment to measure surface expression of inhibitory and costi- the effects of βAR blockade on the efficacy of two immunotherapies mulatory receptors. Combination treated mice were then followed by approved to treat metastatic melanoma: IL-2, which promotes T cell three doses of anti-TIM-3 with or without anti-CTLA-4. proliferation and αPD-1, which impacts T cell activation. Results Methods The addition of anti-TIM-3 and CTLA-4 provided no additional impact C57BL/6 J mice with established B16.F10 melanomas were treated to tumor growth compared to that provided by anti-OX40 and anti- with β-blockers and immunotherapy (αPD-1, IL-2 or αPD-1/IL-2) and PD-1 (n = 7/group, two independent experiments). tumor growth was monitored throughout each treatment regimen. Conclusions The accumulation of immune cells within the tumors and lymphoid These data demonstrate that TIM-3 and CTLA-4 blockade was not tissues were evaluated by flow cytometry at multiple time points fol- sufficient to augment the inhibitory effects of the concurrent com- lowing treatment. bination of anti-OX40 and anti-PD-1, and suggest that targeting in- Results hibitory receptors upregulated after initial immunotherapy treatment Blockade of βAR signaling beginning after tumors were established will require more sophisticated biomarkers and immune monitoring. had no significant impact on tumor growth. In contrast, attenuation Alternatively, we noted significant increases in some costimulatory of tumor growth by each immune-based therapy was improved in receptors (ICOS, 4-1BB) and targeting these receptors may provide the presence of β-blockers. We observed significantly extended sur- an alternative to blocking inhibitory pathways. Additionally there vival in mice treated with αPD-1 or αPD-1/IL-2 combined with β- may be a tipping point where providing additional antibodies target- blockers compared to immunotherapy only mice. Most importantly, ing costimulatory or checkpoint molecules may be ineffective. T cells the combination of β-blockers, αPD-1 and IL-2 produced a highly sig- may become so heavily deregulated or over-stimulated that they be- nificant delay in tumor growth and prolonged survival compared to come permanently exhausted. In support of this we have previously αPD-1/IL-2 without β-blockers. shown that anti-OX40 combined sequentially with anti-PD-1 provides Conclusions superior therapeutic effect compared to concurrent combination, Blocking βAR signaling improved the efficacy of at least two types of with complete tumor regression in ~30% of animals treated. Sequen- immunotherapy, but was most effective when administered with tially treated animals generated T cells capable of significantly more dual-immunotherapy. We suggest that each therapeutic component long-term proliferation, suggesting that timing is critical when com- may improve a unique aspect of the immune response to maximally bining immunotherapies. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 234 of 242 and activation, and in a subcutaneous Lewis lung carcinoma model for their ability to mediate anti-tumor immunity, both alone and in combination with anti-PD-1 mAb. Results In normal mice, IL-2/mAb complexes potently expanded CD8+ T cells and NK cells with minimal expansion of Tregs. As single agent ther- apy, IL-2/mAb complexes or anti-PD-1 mAb reduced tumor growth, although most mice succumb to tumor growth eventually. Combin- ation of IL-2/mAb complexes with anti-PD-1 mAb therapy resulted in durable, complete responses in nearly half of the mice. Conclusions While immune based therapies such as anti-PD-1 mAb can be highly effective in select patients, even in those patients that obtain clinical benefit, disease may recur. Our results suggest that the addition of IL-2/mAb complexes to therapy with anti-PD-1 mAb could broadly in- crease the percentage of patients deriving benefit from immune- based therapy. References 1. Sato, et al: Biotherapy 1993, 6(3):225–231. 2. Boyman, et al: Science 2006, 311:1924–1927. Fig. 4 (abstract P14). The addition of anti-TIM + anti-CTLA-4 provides 3. Létourneau, et al: PNAS 2010, 107:2171–2176. no benefit to anti-OX40 + anti-PD-1 combination treatment P16 The combination of an IL-15/IL-15Ralpha complex (ALT-803) and anti-PD-1 mAb leads to superior anti-tumor immunity in a murine lung tumor model P15 1 1 1 John Wrangle , Kristina Andrijauskaite , Marzena Swiderska-syn , Peter Novel IL-2/mAb complexes mediate potent anti-tumor immunity 2 2 1 Rhode , Hing Wong , Mark Rubinstein which is augmented with anti-PD-1 mAb therapy 1 2 1 1 1 Medical University of South Carolina, Charleston, SC, USA; Altor Mark Rubinstein , Kristina Andrijauskaite , Marzena Swiderska-syn , Kristin 2 2 2 BioScience Corporation, Miramar, FL, USA Lind , Agnes Choppin , Marina K Roell 1 2 Correspondence: Mark Rubinstein (markrubinstein@musc.edu) Medical University of South Carolina, Charleston, SC, USA; XOMA Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P16 Corporation, Berkeley, CA, USA Correspondence: Mark Rubinstein (rubinsmp@musc.edu) Background Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P15 Administration of antibodies that block the PD-1/PD-L1 pathway has demonstrated unprecedented success in mediating clinical re- Background sponses in patients with advanced cancer. These antibodies are Recent success and FDA approval of immune checkpoint inhibitors thought to act by blocking the ability of PD-L1 to mediate an in- (CI) in a growing number of cancers are transforming cancer treat- hibitory signal to PD-1 expressing T cells during antigen- ment and revitalizing interest in immunotherapies. However, while recognition. These antibodies are now FDA-approved for multiple efficacy is observed in patients with advanced metastatic diseases cancers including non-small cell lung cancer (NSCLC) in patients treated with CI, not all patients respond and most responses are in- with disease that has progressed during or after platinum-based complete. Preclinical studies suggest that combinations of additional chemotherapy. In these patients, one in five patients can attain a modalities will provide opportunities to improve patient responses. clinical response while on checkpoint inhibitor therapy. While As both IL-2 and CI therapy can independently augment anti-tumor promising, this therapy fails to induce a durable clinical response immunity in patients, likely in mechanistically distinct ways, we hy- in most patients. To overcome this limitation, we hypothesized pothesized we could improve anti-tumor immunity by combining IL- that combinatorial therapy with anti-PD-1 mAb and a lymphocyte 2 and anti-PD-1 mAb therapy. growth factor would more effectively augment the expansion of Methods tumor-reactive lymphocytes. This would also provide a means to To improve IL-2 efficacy and therapeutic index, we generated novel not only remove inhibitory pathways but directly augment the anti-IL-2 mAbs which, when complexed with IL-2 (IL-2/mAb) offer ad- function of tumor-reactive lymphocytes. We chose to use an IL- vantages over standard IL-2 therapy [1-3]. First, binding to an anti-IL- 15/IL-15Ra complex (ALT-803) composed of an IL-15 mutant 2 mAb increases IL-2 half-life and biological activity. Second, depend- (N72D) that was pre-associated with the soluble IL-15Ra/Fc fusion ing on the epitope at which the mAb binds to IL-2, antibody binding protein. This superagonist complex has been shown to potently can modulate which IL-2 receptor subunits (alpha, beta, or gamma) expand and activate CD8 T cells and NK cells in various animal are engaged. Antibodies that interfere with binding of IL-2Rα can re- models. duce activation of high- IL-2Rα-expressing cell types, such as sup- pressive Tregs, and steer activity toward cell types expressing only IL- Methods 2Rβ and γ. In this way, these complexes may have more effective To assess the efficacy of combination therapy, we injected C57BL/6 anti-tumor activity [1-3]. We screened antibody phage libraries to mice subcutaneously with Lewis lung carcinoma. Mice with estab- identify antibodies that shift IL-2 receptor binding and activity differ- lished tumors were treated with anti-PD-1 mAb and/or IL-15/IL-15Ra entially on different cell types in vitro and in vivo. Complexes of these complex, and we monitored tumor progression and changes in im- antibodies were tested in vivo for their effects on T cell frequency mune cell populations in the periphery and tumor. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 235 of 242 Results findings provide a strategy for the selective targeting of Tregs in the The combination of anti-PD-1 mAb and the IL-15/IL-15Ra complex frame of cancer combination immunotherapy. was substantially more effective at inducing complete responses Conclusions compared with administration of either agent alone. Effective ther- These findings provide a new insight into CD4 T cell biology and apy was associated with the expansion of CD8 T cells and NK cells, offer a new strategy for selective targeting of Tregs in the frame of and the acquisition of the ability of CD8 T cells to produce IFNγ development of anti-cancer immunotherapies. after activation. Interestingly, in vitro, IFNγ led to upregulation of both MHC and PD-L1 on tumor cells, suggesting a mechanistic basis for the improved efficacy of the combination therapy. Combinations: Immunotherapy/ Conclusions Standard of Care Our results suggest that the efficacy of anti-PD-1 mAb therapy may be improved by co-administration of the IL-15/IL-15Ra complex. Our P18 results also suggest a mechanistic basis for why the combination Pembrolizumab in combination with chemoradiotherapy (CRT) for may be superior to single agent therapy. To determine if this com- locally-advanced squamous cell carcinoma of the head and neck bination would be of value in human patients, we have initiated a (SCCHN): Interim safety analysis (ISA) phase Ib/II clinical trial (NCT02523469) to assess the combination of 1 2 1 2 Steven Powell , Mark Gitau , Christopher Sumey , Andrew Terrell , anti-PD-1 mAb (nivoluamb) in combination with ALT-803 in patients 1 1 3 2 Michele Lohr , Ryan K Nowak , Steven McGraw , Ash Jensen , Miran with refractory advanced NSCLC. 2 4 4 1 Blanchard , Kathryn A Gold , Ezra EW Cohen , Christie Ellison , Lora 1 5 1 Black , John Lee , William Chad Spanos 1 2 Sanford Cancer Center, Sioux Falls, SD, USA; Sanford Roger Maris P17 Cancer Center, Fargo, ND, USA Sanford Health, Sioux Falls, SD, USA; Functional dichotomy of PI3K isoforms in CD4 T cells provides a Moores Cancer Center, University of California, San Diego, La Jolla, CA, strategy for selectively targeting regulatory T cells to enhance USA; NantKwest, Inc., Culver City, CA, USA anti-tumor immunotherapy 1 1 1 1 Correspondence: Steven Powell (steven.powell@sanfordhealth.org) Shamim Ahmad , Mason Webb , Rasha Abu-Eid , Rajeev Shrimali , Vivek 1 1 1 1 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P18 Verma , Atbin Doroodchi , Zuzana Berrong , David Yashar , Raed 2 1 1 Samara , Mikayel Mkrtichyan , Samir Khleif 1 2 Background Georgia Cancer Center, Augusta, GA, USA; Qiagen, Frederick, MD, USA Blockade of the programmed death receptor 1 (PD-1) interaction Correspondence: Samir Khleif (skhleif@augusta.edu) with its ligands (PD-L1, PD-L2) represents an active therapeutic Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P17 approach in recurrent and metastatic SCCHN [1]. This immune checkpoint may allow immune escape during standard treatment, Background including CRT [2]. Standard CRT in SCCHN has substantial toxicity The PI3K-Akt signaling pathway modulates diverse biological re- and the safety of adding PD-1 inhibition is unknown. We report sponses including signaling, proliferation and survival of T cells. The the first ISA of weekly cisplatin-based CRT in combination with identification of a signaling pathway, which differentially regulates pembrolizumab, a humanized IgG4 monoclonal antibody inhibitor regulatory T cells (Tregs) and conventional T cells (Tconvs), is crucial of the PD-1:PD-L1/2 interaction. for selectively modulating these two subsets. The differential role of Methods class IA PI3K isoform in regulating the survival and apoptosis of Tregs This is an open-label phase IB trial of pembrolizumab used in and Tconvs has not been elucidated yet. combination with cisplatin-based, definitive CRT in patents with Methods stage III-IVB SCCHN. The treatment schema is outlined in Fig. 5. For in vitro experiments sorted Tregs and Tconvs were labeled with TM Key inclusion and exclusion criteria are in Fig. 6. The primary CellTrace Violet Cell Proliferation stain (VCT) according to the man- endpoint of safety is assessed by dose-limiting grade ≥3adverse ufacturer’s protocol (Life Technologies, NY). Cells were stimulated events (AEs) and immune-related AEs (irAEs) per NCI-CTCAE v4.0 with and without inhibitors. For in vivo experiments C57BL/6 Mice criteria. Efficacy is measured by RECIST v1.1 complete response were injected subcutaneously (s.c.) with TC-1 tumor cells and moni- (CR) rate on 100-day post-CRT imaging and/or pathologic CR for tored for development of tumors. Vaccine was given weekly s.c. For those who undergo salvage surgery. Planned enrollment is 39 pa- therapeutic experiments vaccine was given weekly throughout the tients based on a hypothesized CR rate of >60% and a two-stage experiment. CAL-101 treatment was provided on the day when Simon minimax design (type I error rate of 0.05 and power of tumor size reached 3-4 mm 5-6 day before vaccination. 0.80). Secondary endpoints will include overall response, survival, Results and quality-of-life assessments. Here, we report that PI3Kd isoform is sufficient for TCR downstream Results signaling, proliferation, and survival for either Tconvs or Tregs. In At the time of ISA, 27 patients have been enrolled. Patient and Tregs, however, PI3Kd is a dominant isoform, where Tregs are fully disease characteristics are in Fig. 7. At data cut-off on 9/14/2016, dependent on PI3Kd to regulate these properties as PI3Kα and PI3Kb 22 patients have completed CRT. Of those patients, 3 required do not play any role in these biologic processes. On the other hand, cisplatin dose reductions and 6 required a dose discontinuation. in Tconvs, the two other isoforms, PI3Kα and PI3Kb combined, pro- Acute grade (G) ≥ 3 AEs for those completing CRT are in Fig. 8. vide redundant pathway to PI3Kd in the regulation of TCR signaling, Median cumulative cisplatindoseis225 mg/m2. Therewereno proliferation and survival. This redundant role provided by PI3Kα and radiation dose-limiting toxicities. Two patients (9%) discontinued PI3Kβ isoforms to PI3Kd in Tconvs offers a selective therapeutic ap- pembrolizumab due to irAEs (G2 peripheral motor neuropathy proach to inhibit Tregs, where by inhibiting PI3Kd, signaling, prolifer- and G3 transaminase elevation). No deaths have occurred. ation, and survival are inhibited in Tregs, while PI3Kα and PI3Kβ, will provide a path for Tconvs to proliferate and function. Conclusions Importantly, we demonstrate that our findings translate to thera- This represents one of the first studies to evaluate the safety of con- peutic efficacy in vivo, where the inhibition of PI3Kδ, enhanced anti- current CRT and PD-1 inhibition in SCCHN. Acute CRT-related toxic- tumor efficacy of antigen-specific vaccine by decreasing the suppres- ities are comparable to other SCCHN CRT studies. No new sive Tregs and increasing the number of vaccine-induced CD8 T cells, immunologic safety signals were seen. Further investigation of this thus showing synergistic therapeutic effect against tumors. Our approach is warranted. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 236 of 242 Acknowledgements Funding supported by the Merck Investigator Studies Program. Trial Registration ClinicalTrials.gov identifier: NCT02586207 References 1. Seiwert TY, et al: Safety and clinical activity of pembrolizumab for treatment of recurrent or metastatic squamous cell carcinoma of the head and neck (KEYNOTE-012): an open-label, multicentre, phase 1b trial. Lancet Oncol 2016, 17(7):956–965. 2. Parikh F, et al: Chemoradiotherapy-induced upregulation of PD-1 an- tagonizes immunity to HPV-related oropharyngeal cancer. Cancer Res 2014, 74(24):7205–7216. Fig. 8 (abstract P18). Acute Grade ≥3 Adverse Events with >1 Occurrence Diet, Exercise and/or Stress and Impact on the Immune System P19 Exercise training reduces splenic accumulation of MDSCs and delays tumor progression in a therapeutic breast cancer model 1 2 1 3 Erik Wennerberg , Emily Schwitzer , Claire Lhuillier , Graeme Koelwyn , 2 2 4 Rebecca Hiner , Lee Jones , Sandra Demaria 1 2 Weill Cornell Medicine, New York, NY, USA; Memorial Sloan Kettering Fig. 5 (abstract P18). Clinical Trial Schema Cancer Center, New York, NY, USA; New York University School of Medicine, New York, NY, USA; Department of Radiation Oncology, Weill Cornell Medicine, New York, NY, USA Correspondence: Erik Wennerberg (erw2010@med.cornell.edu) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P19 Background Epidemiological studies show a correlation between physical ac- tivity and cancer-related mortality [1]. However, the contribution of immune mediated anti-tumor immunity to the beneficial ef- fects of exercise has yet to be defined [2]. We sought to investi- gate if forced running would have a therapeutic benefit in mice bearing a poorly immunogenic breast cancer and investigate the Fig. 6 (abstract P18). Key Inclusion and Exclusion Criteria immunological changes occurring in response to exercise. Methods On day 0 Balb/c mice were inoculated with 4 T1 breast cancer cells subcutaneously in the right flank (n = 6/group). Starting on day 7, once tumors were palpable, mice were subjected to 30 minutes of forced treadmill running (18 cm/sec) five days per week. Control mice remained sedentary throughout the study. Analysis of immune cells in spleen and tumor was performed at day 17 and 32 and spon- taneous lung metastases were evaluated at day 32. Results We observed a significantly delayed primary tumor growth (tumor volume on day 31: 1167 ± 174 mm in sedentary versus 847 ± 124 mm in exercised mice, p < 0.01) and a tendency for re- duced metastatic burden in the lungs of exercised compared to sedentary mice. The progressive marked increase in myeloid- derived suppressor cells (MDSCs) and splenomegaly seen in sed- entary 4 T1 tumor-bearing mice was less pronounced in exercised mice. This difference was significant on day 17; with spleen weight (520 ± 110 mg in sedentary versus 330 ± 30 mg in exer- cised mice, p < 0.01) and MDSC frequency in spleen leukocytes (22.7 ± 2.6% in sedentary versus 14.3 ± 2.7% in exercised mice, p < 0.001) were significantly lower in exercised mice compared to sedentary mice. Furthermore, on day 32, the CD8+ T cell/Treg and CD8+ T cell/MDSC ratio showed a tendency to increase in tumors from exercised mice. Conclusions Our data demonstrate that exercise can slow tumor progression in a therapeutic setting. While the mechanisms of this effect re- Fig. 7 (abstract P18). Patient and Disease Characteristics quire further investigation, the observed decrease in the Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 237 of 242 proportion of immunosuppressive immune cells in spleen and Conclusions tumor of exercised mice is likely to play a role. Importantly, the These data support the possibility that NHS-muIL12 abrogates an ability of exercise to reduce immunosuppression locally and sys- immune-suppressive response within the TME, which might permit T temically supports testing exercise in combination with immuno- cells to execute their antitumor effects. NHS-huIL12 (MSB0010360N; therapy as a therapeutic modality that can increase responses M9241), is currently being evaluated against solid tumors in a phase without increasing toxicity. I clinical trial (NCT01417546). References Acknowledgements 1. Ballard-Barbash R, Friedenreich CM, Courneya KS, Siddiqi SM, McTiernan We acknowledge the kind contribution of NHS-muIL12 from EMD Serono, A, Alfano CM: Physical activity, biomarkers, and disease outcomes in Billerica, MA. cancer survivors: a systematic review. Natl Cancer Inst 2012, 104:815– 2. Koelwyn GJ, Wennerberg E, Demaria S, Jones LW: Exercise in Regulation Therapeutic Cancer Vaccines of Inflammation-Immune Axis Function in Cancer Initiation and Pro- gression. Oncology 2015, 29(12). P21 Intracellular trafficking of self-assembled immune signals Michelle Bookstaver, Christopher M Jewell Not Listed – Other Fischell Department of Bioengineering, University of Maryland - College P20 Park, College Park, MD, USA Systemic immunotherapeutic efficacy of an immunocytokine, NHS- Correspondence: Michelle Bookstaver (mlbooks@umd.edu) muIL12, in a superficial murine orthotopic bladder cancer model Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P21 Vandeveer Amanda, John W Greiner, Jeffrey Schlom Laboratory of Tumor Immunology and Biology, Center for Cancer Background Research, National Cancer Institute, Bethesda, MD, USA We recently exploited electrostatic interaction to design self- Correspondence: Vandeveer Amanda (amanda.vandeveer@nih.gov) assembling nanostructures comprised entirely from peptide anti- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P20 gens and toll-like receptor (TLR) agonists as adjuvants. These ma- terials simplify vaccine composition and exhibit unique properties Background such as direct control over the absolute and relative concentra- Interleukin-12 is one of the most powerful proinflammatory cytokines tions of each component and co-delivery of the signals to anti- capable of supporting T and NK cell function, inducing IFNγ while gen presenting cells. In pre-clinical models of melanoma, this driving a T 1 adaptive immune response. Its success as an antitumor approach leads to significantly enhanced anti-tumor immunity. agent in preclinical models has yet to be realized in a clinical setting Here, we study how the physicochemical features (e.g., peptide due to systemic toxicity. An IL-12 delivery system has been devel- charge) and relative concentration of each component impact the oped to maximize deposition of the cytokine directly in the tumor internalization, trafficking, and processing of the immune signals microenvironment (TME), while mitigating the dose-limiting systemic in antigen presenting cells. effects. Methods Methods FITC-labeled SIINFEKL peptide was modified with three or nine argi- NHS-IL12 is a novel immunocytokine, consisting of two molecules of nines, for use as a cationic anchor to support self-assembly with a human or murine IL-12 fused to a tumor necrosis-targeting human polyanionic nucleic acid-based TLR3 agonist, polyIC. Hollow capsules IgG1 (NHS76). NHS76 recognizes exposed chromatin-DNA found in built from these signals were synthesized by coating a sacrificial necrotic human/murine tumors. Previous studies have shown select- CaCO core with alternating layers of modified SIINFEKL and PolyIC. ive tumor uptake of NHS-IL12 in necrotic subcutaneous murine tu- After deposition, the core was removed using EDTA and capsules mors. Urothelial bladder cancer is known to respond favorably to were washed with buffer, resulting in stable capsules formed from immunotherapeutic agents due to many somatic mutations and TILs, immune signals. Capsule size was determined by image analysis and and response to Bacillus Calmette-Guerin (BCG). component loading levels were determined by fluorimetry using Results FITC-labeled peptide and Cy5-labeled TLRa. Stability studies were car- We evaluated the use of NHS-muIL12 in a murine orthotopic ried out by incubating capsules in media as a function of different Luc bladder cancer model. MB49 cells, instilled into the bladder pH and ionic strengths. For uptake and trafficking studies, murine form superficial, multifocal tumors which can be monitored with splenocytes were isolated and treated with different concentrations a luciferase-based intravital imaging system. NHS-muIL12 is a very of capsules. Cells were analyzed by flow cytometry and imaging in potent anti-tumor agent in both MB49 tumor models, reducing the presence or absence of inhibitors of endocytic processes and tumor volume in a dose-dependent manner. In the intravesical during staining with markers for surface proteins and intracellular bladder model, antitumor effects were seen at 2.5 mg/kg admin- organelles. istrated as three separate systemic injections. Mice were cured of Results tumor when treated at 20 mg/kgx3 NHS-muIL12 with durable Capsules loaded with FITC-SIINFEKL-R and PolyIC were 1-2 μmin tumor-free long-term survival. Immune analyses revealed TAA- diameter and exhibited similar size and shape for 2 weeks when in specific CTLs and IFN-γ responses, indicating the development of buffer. These materials exhibited tunable loading with a composition a specific anti-tumor immune response. An immune memory re- of 15.5% peptide and 84.5% TLRa used for trafficking studies. FITC- sponse protected mice following re-challenge with MB49 tumor SIINFEKL-R and PolyIC capsules were efficiently internalized through cells. Anti-tumor efficacy required CD4+ or CD8+ T cells as deple- energy dependent processes (i.e., endocytosis) when incubated with tion of either abrogated the anti-tumor effects. Evaluation of TILs primary dendritic cells within 1 hour of treatment. These effects were by FACS, revealed that NHS-muIL12 significantly reduced the also found to be dose-dependent and did not impact viability of number of immune suppressive cells such as MDSCs, 24 hours treated cells. post-treatment, which continued to the end of the study. Im- Conclusions munofluorescence showed correlative treatment-related modula- Initial studies reveal capsules comprised of FITC-SIIN-R and tion of CD4+ and CD8+ T cells as well as MDSCs and Tregs PolyIC are uptaken by primary immune cells quickly and effect- within the TME. Gene expression of RNA from bladder tumors, ively. Ongoing studies will assess the uptake of capsules by identified various immune components with immunosuppressive endocytosis in the presence of inhibitors to decipher the endocy- or immune potentiating roles, modulated by NHS-muIL12 tic pathway and trafficking of capsules through lysosomes and treatment. endosomes. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 238 of 242 Acknowledgements whether antibody and T cell responses were directed to mutations, This work was supported in part by NSF CAREER # 1351688 and Alliance for altered peptide ligands or overexpressed normal proteins. Cancer Gene Therapy # 15051543. Results Compared to vaccination alone or vaccination with GM-CSF, vaccin- ation with DPV-001 plus imiquimod significantly (p < 0.05) increased P22 the number of antibody responses that were four-fold higher at Analysis of B and T cell responses in non-small cell lung cancer week twelve. In the one patient where autologous tumor was avail- (NSCLC) patients enrolled in a phase II trial of cyclophosphamide able, vaccination increased the tumor-specific release of TNF-alpha with allogenic DRibble vaccine (DPV-001) by peripheral blood CD4 T cells. 1 1 2 3 Christopher Paustian , Andrew Gunderson , Brian Boulmay , Rui Li , Conclusions 4 4 5 6 Bradley Spieler , Kyle Happel , Tarsem Moudgil , Zipei Feng , Carmen Based on these studies, future trials will combine the adjuvant imi- 3 6 7 Ballesteros-Merino , Christopher Dubay , Brenda Fisher , Yoshinobu quimod with DRibble vaccine. Trial Registration 8 1 4 3 Koguchi , Sandra Aung , Eileen Mederos , Carlo B. Bifulco , Michael ClinicalTrials.gov identifier: NCT01909752 9 9 9 4 McNamara , Keith Bahjat , William Redmond , Augusto Ochoa , Hong- 10 11 11 6 Ming Hu , Adi Mehta , Fridtjof Lund-Johansen , Bernard Fox , Walter 8 8 1 Urba , Rachel E. Sanborn , Traci Hilton P23 1 2 UbiVac, Portland, OR, USA; Section of Hematology/Oncology, Louisiana An open-label phase I/IIa escalating dose study to evaluate safety State University, New Orleans, LA, USA; Robert W. Franz Cancer and T cell immunogenicity of PDS0101 in subjects with cervical Research Center, Earle A. Chiles Research Institute, Providence Cancer intraepithelial neoplasia (CIN) and high-risk HPV infection 4 1 1 1 1 Center, Portland, OR, USA; Louisiana State University Stanley S. Scott Frank Bedu-Addo , Greg Conn , Michael King , Panna Dutta , Robert 5 2 3 Cancer Center, New Orleans, LA, USA; PPMC, Portland, OR, USA; Shepard , Mark Einstein 6 7 1 2 Providence Cancer Center, Portland, OR, USA; Providence Medical PDS Biotech, New Brunswick, NJ, USA; PDS Biotech, Miami Beach, FL, 8 3 Center, Portland, OR, USA; Earle A. Chiles Research Institute, Providence USA; Rutgers, NJ Medical School, Newark, NJ, USA Cancer Center, Portland, OR, USA; Providence Medical Center, Portland, Correspondence: Robert Shepard (RShepard@Post.Harvard.edu) 10 11 OR, USA; UbiVac, Providence Medical Center, Portland, OR, USA; Oslo Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P23 University Hospital, Oslo, Norway Correspondence: Christopher Paustian Background (christopher.paustian@ubivac.com) Current HPV vaccines are effective at preventing infection. However, Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P22 there are no therapeutic vaccines to treat the infection or commonly associated diseases e.g. CIN, cervical, anal and oral cancers. A therapy Background that is simple, effective and safe enough to be administered to CIN DRibble vaccines are microvesicles derived from proteasome-blocked and early-stage cancer patients could be important in achieving the autophagosomes. The DPV-001 DRibble vaccine is derived from an goal of effective cancer prevention and treatment of pre-metastatic adenocarcinoma and a mixed histology cancer cell line. By mass cancer. We assessed whether PDS0101, a combination of modified spectroscopy they contain more than 130 potential NSCLC antigens, multi-epitope HPV16 peptides (HPVmix) and escalating doses of the many as prospective altered-peptide ligands, which could intensify synthetic Versamune® T cell activating platform could facilitate anti- their immunogenicity. In preclinical models, DRibble immunotherapy gen cross-presentation and safe immune activation leading to strong provided significant cross-protection against 8 of 9 tumors tested. HPV-specific CD8+ T cell induction in CIN. Additionally, Dribble vaccines are effective in treating established tu- Methods mors in preclinical combination immunotherapy models. We Safety and immunogenicity were assessed in an open label dose es- hypothesize that the efficacy of DRibbles’ vaccination can be attrib- calation study. Groups of 3-6 subjects received either low dose uted to their capacity to present tumor-derived short-lived proteins (1 mg), medium dose (3 mg) or high dose (10 mg) of Versamune® (SLiPs) and defective ribosomal products (DRiPs) that are typically cationic lipid with 2.4 mg of HPVmix. Each subject received one SC not processed and presented by professional antigen presenting dose every 3 weeks for a total of 3 doses. T cell response was evalu- cells. These SLiPs and DRiPs embody a prospective pool of tumor an- ated by IFN-γ and granzyme-b ELISpot using blood drawn from the tigens against which the host may be less tolerant. subjects pre-vaccination, 2 weeks after each vaccination and 90 days Methods after vaccination 3. The trial is registered at ClinicalTrials.gov (number Thirteen definitively-treated stage III NSCLC patients were vaccinated NCT02065973). at 3-week intervals. Patients were randomized such that some pa- Results tients’ intradermal vaccines were combined with administration of No serious adverse events were reported. No IND safety reports were imiquimod or GM-CSF as an adjuvant. PBMCs and serum were col- submitted. No subjects withdrew. Strong HPV-specific T cell re- lected at baseline and at each vaccination. For one patient, PBMCs sponses occurred at all 3 doses, even in those subjects with low pre- from the baseline visit and week 12 were tested against that patient’s vaccination T cell responses. PDS0101 vaccination led to strong T cell autologous tumor cell line to measure increased tumor specific T cell responses evaluated by both IFN-γ and granzyme-b ELISpot. Conclu- activation. Studies are currently underway to evaluate changes in sions: PDS0101 is safe and effectively performs antigen cross- TCR repertoires. CD4+ and CD8+ T cells from multiple time points presentation as demonstrated by HPV-specific T cell responses, in- were sorted and TCR sequencing is being performed to look at alter- cluding inducing active cytolytic T cells. Clinical benefit in CIN2/3 and ations in the T cell repertoire. The primary outcome measure of this cancer will be evaluated in larger phase II trials. clinical trial was to discover if vaccine alone, vaccine plus imiquimod, Conclusions or vaccine plus GM-CSF generated the greatest number of strong PDS0101 is safe and effectively performs antigen cross-presentation antibody response. as demonstrated by HPV-specific T cell responses, including inducing Serum from the baseline visit and week 12 was analyzed for in- active cytolytic T cells. Clinical benefit in CIN2/3 and cancer will be creased antibody response to >9000 human proteins using ProtoAr- evaluated in larger phase II trials. rays and Microsphere Affinity Proteomics. Where sufficient tumor Trial Registration was available, whole exome sequencing was done to evaluate ClinicalTrials.gov identifier: NCT02065973 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 239 of 242 Tumor Microenvironment Trial Registration ClinicalTrials.gov identifier: NCT01421017 P24 Effects of TLR7 agonist imiquimod combined with local References radiotherapy on the tumor microenvironment in women with 1. Panelli: Genome Biol 2008. metastatic breast cancer in a prospective trial 2. Dewan: CCR 2012. 1 2 3 1 1 Sylvia Adams , Ena Wang , Ping Jin , Yelena Novik , Debra Morrison , 1 2 4 1 Ruth Oratz , Franco M Marincola , David Stroncek , Judith Goldberg , 5 5 Sandra Demaria , Silvia C Formenti P25 Perlmutter Cancer Center, New York University School of Medicine, Immunoscore as a prognostic marker in stage I-III colon cancer: NYC, NY, USA; Sidra Medical and Research Center, Doha, Qatar; results of a SITC-led global validation study 3 1 1 1 2 National Institutes of Health Clinical Center Department of Transfusion Jérôme Galon , Bernhard Mlecnik , Florence Marliot , Fang-Shu Ou , 4 3 4 4 4 Medicine, Bethesda, MD, USA; National Institutes of Health Clinical Carlo B Bifulco , Alessandro Lugli , Inti Zlobec , Tilman T Rau , Iris D 5 5 5 6 6 Center, Bethesda, MD, USA; Weill Cornell Medicine, Department of Nagtegaal , Elisa Vink-Borger , Arndt Hartmann , Carol Geppert , Michael 7 7 7 7 Radiation Oncology, New York, NY, USA H. Roehrl , Prashant Bavi , Pamela S Ohashi , Julia Y Wang , Linh T 7 7 7 7 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P24 Nguyen , SeongJun Han , Heather L MacGregor , Sara Hafezi-Bakhtiari , 7 8 8 8 Bradley G Wouters , Yutaka Kawakami , Boryana Papivanova , Mingli Xu , 8 9 9 9 Correspondence: Sylvia Adams (sylvia.adams@nyumc.org) Tomonobu Fujita , Shoichi Hazama , Nobuaki Suzuki , Hiroaki Nagano , 10 11 12 12 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P24 Kiyotaka Okuno , Kyogo Itoh , Eva Zavadova , Michal Vocka , Jan 12 12 12 12 Spacek , Lubos Petruzelka , Bohuslav Konopasek , Pavel Dundr , 12 13 13 Background Helena Skalova , Toshihiko Torigoe , Noriyuki Sato , Tomohisa 13 13 14 Application of TLR7 activator imiquimod (IMQ) onto BCC of the skin Furuhata , Ichiro Takemasa , Marc Van den Eynde , Anne Jouret- 14 14 1 1 leads to an early tumoral transcriptional profile of immunological re- Mourin , Jean-Pascal Machiels , Tessa Fredriksen , Lucie Lafontaine , 1 1 1 1 jection (ICR) preceding complete remission as shown in a random- Bénédicte Buttard , Sarah Church , Pauline Maby , Helen Angell , 1 1 1 1 ized trial [1]. Here we employed the same methodology evaluating Mihaela Angelova , Angela Vasaturo , Gabriela Bindea , Anne Berger , 1 15 15 15 serial FNA tumor biopsies from breast cancer patients treated on a Christine Lagorce , Prabhu S Patel , Hemangini H Vora , Birva Shah , 15 15 15 clinical trial of the combination of IMQ and radiotherapy (RT), to de- Jayendrakumar B Patel , Kruti N Rajvik , Shashank J Pandya , Shilin N 15 16 16 17 lineate dynamic changes associated with ICR in breast cancer and to Shukla , Yili Wang , Guanjun Zhang , Giuseppe V Masucci , Emilia K 17 18 18 19 understand the contribution of each treatment modality to antitumor Andersson , Fabio Grizzi , Luigi Laghi Gerardo Botti , Fabiana 19 19 19 19 immunity in vivo. We previously demonstrated synergy of IMQ/RT in Tatangelo , Paolo Delrio , Gennaro Cilberto , Paolo A Ascierto , 20 2 3 1 the poorly immunogenic TSA model with enhanced T cell-mediated Franco Marincola , Daniel J Sargent , Bernard A Fox , Franck Pages inhibition of treated and untreated tumors [2]. INSERM, Université Pierre et Marie Curie, Université Paris Descartes, 2 3 Methods Paris, France; Mayo Clinic, Rochester, MN, USA; Earle A. Chiles Research Clinical trial (NCT01421017): for patients with metastatic breast can- Institute, Providence Cancer Center, Portland, Oregon, USA; Institute of cer to the skin. Treatment: topical IMQ to one metastasis, IMQ and Pathology, University of Bern, Bern, Switzerland; Radboud University RT to another metastasis. IMQ self-applied 5xX/per week x 8 weeks, Nijmegen Medical Center, Nijmegen, Netherlands; University Erlangen- RT started with first IMQ (6Gyx5 over 10 days). Cyclophosphamide Nürnberg, Erlangen, Germany; Princess Margaret Cancer Centre, (200 mg/m2 IV) was administered a week before in a subset of pa- University Health Network, Toronto, ON, Canada; Division of Cellular tients. An untreated, measurable lesion (skin or distant metastases) Signaling, Institute for Advanced Medical Research, Keio University outside the IMQ and radiation fields was observed as systemic re- School of Medicine, Tokyo, Japan; Department of Gastroenterological, sponse read-out per RECIST1.1. Local response defined as PR or CR in Breast and Endocrine Surgery, Yamaguchi University Graduate School of treated lesions. FNA of IMQ and IMQ/RT treated metastases: at base- Medicine, Ube, Japan; Department of Surgery, Kinki University Faculty line, 2 and 3 weeks, RNA isolation/amplification performed per SOPs. of Medicine, Osaka-Sayama, Japan; Division of Clinical Research, Gene expression: Affymetrix Human GeneArray 1.0 ST/Partek Genom- Research Center for Innovative Cancer Therapy, Kurume University ics suite software with special emphasis on expected immune School of Medicine, Kurume, Japan; First Faculty of Medicine, Charles signature. University and General University Hospital, Prague, Czech Republic; 13 14 Results Sapporo Medical University, Sapporo, Japan; Institut Roi Albert II, Serial FNA samples are available from 18 patients. Gene expression Cliniques universitaires St-Luc, Université Catholique de Louvain, profiles of baseline biopsy treated with IMQ/RT identified 2309 differ- Brussels, Belgium; The Gujarat Cancer & Research Institute, entially expressed genes (p < 0.005) between CR and PR. Among Ahmedabad, India; Institute for Cancer Research, Xi’an Jiaotong them, ICR genes such as GZMB, GZMH, PRF1, GNLY, CD8A and TBX21 University, Xi’an, China; Karolinska Institutet, Karolinska University, are over expressed in CR. Significant altered gene expression was ob- Stockholm, Sweden; Humanitas Clinical and Research Center, Rozzano, 19 20 served in progressing lesions (week 3 vs baseline, 1854 genes) in Italy; Istituto Nazionale Tumori Fondazione Pascale, Naples, Italy; Sidra contrast to responding metastases (PR: 53 genes, CR: 23 genes) post Medical and Research Center, Doha, Qatar IMQ/RT suggesting active wound healing and tumor progression sig- Correspondence: Bernard A Fox (bernard.fox@providence.org) nature. For the IMQ alone treated metastases, differential gene ex- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P25 pression was observed at baseline distinguishing subsequent PR and PD (n = 189, p < 0.005). Systemic response was observed as a marked Background baseline gene expression difference (n = 1177, p < 0.005) predicting Increasing evidence has illustrated that enhanced lymphocytic infil- abscopal phenomena (CR, PR, SD and PD). tration is a powerful prognostic marker in colon cancer (CC). The Conclusions Immunoscore (IM) methodology was developed as a standardized The ICR signature in tumors before IMQ-RT treatment is positively assay to quantify the in situ immune cell infiltrate. correlated with complete local response, which validates the ICR hy- Methods pothesis in metastatic breast cancer. Systemic response consistent The Society for Immunotherapy of Cancer (SITC) led an international with induction and/or boosting of adaptive immunity is predicted by consortium, initiated with 23 expert centers from 17 countries, to evalu- significant enrichment of immune signature. ate the Immunoscore in routine clinical settings. CC patients (pts) stages I/II/III with no prior neo-adjuvant treatment were included in this Acknowledgements study. Overall, 3855 pts split into a training set (TS), internal validation 1RO1CA161891 set (IVS), and external validation set (EVS) were quantified for IM using Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 240 of 242 immunohistochemistry with CD3/CD8 antibodies and digital pathology documented clinical follow up. We identified a preliminary population quantification of whole slide sections. All statistical analyses were pre- of 31 stage II-III melanoma patients diagnosed between 2000 and defined and performed by external statisticians. The primary endpoint 2012, with characteristics shown in Fig. 9 for whom pathology from was time-to-recurrence (TTR); multivariate analyses were performed the primary biopsy was shown. Clinical follow up was available on 18 using Cox models adjusted for IM, age, gender, T-stage, N-stage, and patients of whom 9 patients were alive with no evidence of recur- stratified by participating center. rence, 1 had died of another malignancy, and 7 had died of melan- Results oma. 15 patients had more than 24 months of survival information Across centers, the median recurrent follow-up was 126.6 months. Pt available but no detailed clinical information. 5 μm slides from either characteristics: 51.5% male, median age 69 years, and 17%/54%/29% the primary biopsy or subsequent wide local excision procedure were stage I/II/III, respectively. Among pts with stages I-III CC in the TS, TTR stained using Opal multiplex IHC for DAPI, CD3 (LN10, Leica), CD8 was shorter among 152 pts (22%) with Low-IM CC vs. 548 pts with (4B11, Leica), CD68 (KP1, Biogenex), SOX10 (BC34, Biocare), HLA-DR High-IM CC (HR [95% CI], 0.41 [0.28-0.61]; P < 0.0001). In the IVS, TTR (LN-3, Abcam) and Ki67 (MIB1, Abcam). Cell phenotypes within repre- was also shorter among 155 pts with Low-IM CC vs. 481 pts with sentative fields pre-selected by a trained dermato-pathologist and High-IM CC (0.41 [0.27-0.65]; P < 0.0001). In the EVS, TTR was also were visualized using the Mantra quantitative pathology workstation shorter among 225 pts with Low-IM CC vs. 744 pts with High-IM CC (Perkin Elmer), and analysis of spatial distribution of CD3 + CD8+ cells (0.51 [0.38-0.68]; P < 0.0001). These results were independent of age, analyzed as shown in Figs. 10 and 11 using inForm® image analysis sex, tumor stage, and sidedness. Among secondary objectives, Immu- software (Perkin Elmer), and Spotfire software (TIBCO). noscore groups (High, Int, Low) predicted time to recurrence in the Results TS (0.19 [0.10-0.37]; P < 0.0001), IVS (0.27 [0.14-0.53]; P < 0.0001), and CD3 + CD8+ cells are closer to both tumor (SOX10+) and CD68+ cells EVS (0.33 [0.22-0.49]; P < 0.0001). In stage II CC pts (1433), the differ- when they express HLA-DR (p < 0.001). Conversely, CD3 + CD8+ cells ence in TTR was significant between the Low and High-Immunoscore are significantly farther from Sox10+ cells when they express Ki-67. groups (0.36 [0.23-0.56]; P < 0.0001). In multivariate models, Immuno- Among patients with clinical follow up, CD3 + CD8+ cells in non- score grouping (2, 3, or 5) was significant (C-index : 0.73 [0.66-0.80], recurrent patients were closer to SOX10+HLA-DR+ cells than they all P < 0.0001). Multivariate models including MSI and sidedness were were in recurrent patients(p < 0.001). performed and will also be presented. Reproducibility of the IM assay Conclusions was validated across centers. If proximity is a surrogate for interaction, these data may indicate Conclusions that HLA-DR expression enhances interaction with T cells for both The primary and secondary endpoints of the global Immunoscore CD68+ infiltrating cells and Sox10+ tumor cells. In addition, study were reached. Overall, TTR was significantly longer in pts with CD3+CD8+ cells were closer to SOX10+HLA-DR+ cells in patients who stages I-III CC defined as High-IM. Moreover, a subgroup of patients did not recur, which is interesting in light of recent data showing that with high-risk stage II CC was also identified by Low-IM. expression of HLA-DR by tumor cells increases likelihood of response to anti-PD1. Further staining and analysis of annotated tumor samples Acknowledgements from the complete HICCC cohort 2000-2012 is ongoing and results will This initiative was supported by a variety of sources, including funding from be updated at time of presentation. Definiens, Prometheus, and a grant from the Czech Ministry of Health, 15- 28188A and League against cancer. P26 Defining critical features of the immune microenvironment in melanoma 1 1 2 1 3 Robyn Gartrell , Douglas Marks , Edward Stack , Yan Lu , Daisuke Izaki , 4 4 4 4 Kristen Beck , Dan Tong Jia , Paul Armenta , Ashley White-Stern , Yichun 4 1 5 1 1 Fu , Zoe Blake , Howard L Kaufman , Bret Taback , Basil Horst , Yvonne M Saenger 1 2 Columbia University Medical Center, New York, NY, USA; Perkin Elmer, Hopkinton, MA, USA; Columbia University, New York, NY, USA; Columbia University College of Physicians and Surgeons, New York, NY, USA; Rutgers Cancer Institute of New Jersey, New Brunswick, NJ, USA; New York Presbyterian/Columbia University Medical Center, New York, NY, USA Correspondence: Robyn Gartrell (rdg2129@columbia.edu) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P26 Background Precise biomarkers are urgently needed to characterize the tumor immune micro-environment, both for prognostication and to predict the benefit of immuno-therapeutic intervention. HLA-DR on tumor cells and Ki67 on cytotoxic (CD8+) T cells have been proposed as biomarkers of anti-PD1 activity. Multiplex immunohistochemistry (mIHC) allows for automated quantitation of phenotypes and spatial distributions of immune cell populations within formalin fixed paraf- fin embedded (FFPE) tissues. Methods In order to test whether mIHC can better characterize the tumor im- mune microenvironment, we screened databases at the Herbert Irving Cancer Center (HICC) at Columbia University for early stage melanoma Fig. 9 (abstract P26). Demographic Characteristics of Melanoma patients with available FFPE primary melanoma tissue and Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 241 of 242 microenvironment (flipping immunosuppressive M2 macrophages to an anti-tumor M1 state), and T cell expansion and activation via den- dritic cell maturation and antigen presentation. Clinically, Imprime has demonstrated promising efficacy in clinical trials when combined with tumor-targeting or anti-angiogenic antibodies. Phase II studies with pembrolizumab are starting in both metastatic triple negative breast cancer and metastatic melanoma. Herein, we have employed multi- plexed immunofluorescence to profile the immune microenvironment in preclinical tumor tissues. Methods The B16F10 experimental metastasis model was used to interro- gate Imprime’s anti-tumor activity in vivo. B16F10 melanoma cells were injected into the tail vein of syngeneic C57BL/6 mice, seed- ing the lungs with B16 foci. Outgrowth of these metastatic foci was assessed after treatment with the tumor-targeting anti-tryp1 Fig. 10 (abstract P26). Distance between CD3 + CD8+ T cells and antibody TA99, Imprime, or the combination. At various times nearest neighbor, Left: Mean distance from CD3 + CD8+ to CD68 + post tumor injection, lungs were examined via multiplexed im- HLA-DR- (light blue) or CD68 + HLA-DR+ (dark blue), Center: Mean munofluorescence (IFC) for markers of immune infiltration and distance from CD3 + CD8+ to SOX10 + HLA-DR- (pink) or SOX10 + activation. IFC was performed using 7-color staining (Opal tech- HLA-DR+ red), Right: Mean distance from CD3 + CD8+ to SOX10 + nology, PerkinElmer) combined with in situ hybridization (RNA- Ki67- (pink) or SOX10 + Ki67+ red) Scope, ACD). Images were acquired with the Vectra3 multispectral imaging system and cells segmented using Inform (PerkinElmer). Imaging data were transformed into “.fcs” files and analyzed using Flowjo flow cytometry software (Treestar)). Relational pa- rameters such as immune cell clustering and tumor infiltration were performed via custom algorithms in R. Results TA99 alone suppressed the outgrowth of B16 lung metastases by 54% when compared to vehicle treatment. The combination of Imprime with TA99 reduced the number of metastases even more profoundly (96% vs vehicle). IFC analyses showed that Imprime specifically accumulates in the tumor stroma, binds to macro- phages and elicits increased iNOS production, indicating the re- polarization of these macrophages to a more M1-like, inflamma- tory state. Imprime-treatment also triggered the formation of large immune cell clusters, possibly representing resolved tumor nests or the establishment of tertiary lymphoid tissues, both of which have been identified as predictors of successful anti-tumor im- mune responses. Finally, Imprime treatment and localization at the tumor site corresponds with substantial upregulation of the gene Mx1- a type 1 interferon-responsive gene. Conclusions Imprime is a potent immunomodulator that induces a coordi- nated immune attack in vivo demonstrated by immune cell binding, M1 re-polarization and a type-1 interferon signature Fig. 11 (abstract P26). Distance between CD3 + CD8+ T cells and that coincides with reduced outgrowth of established ling nearest neighbor be recurrence status, Top Left: Mean distance from metastases. CD3 + CD8+ to CD68 + HLA-DR- by recurrence status, Lower Left: Mean distance from CD3 + CD8+ to CD68 + HLA-DR+ by recurrence status. Top Right: Mean distance from CD3 + CD8+ to SOX10 + HLA- P28 DR- by recurrence status, Bottom Right: Mean distance from CD3 + Local convection-enhanced delivery of PD-1 blockade antibody in CD8+ to SOX10 + HLA-DR+ by recurrence status de novo murine model of glioblastoma Jennifer S Sims, Liang Lei, Takashi Tsujiuchi, Jeffrey N Bruce, Peter Canoll Columbia University Medical Center, New York, NY, USA P27 Correspondence: Jennifer S Sims (jennifer.s.sims@gmail.com) Immune profiling via multiplexed immunofluorescence shows that Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P28 Imprime based anti-cancer efficacy involves profound changes in macrophage polarization, type 1 IFN signaling and the formation Background of immune cell clusters Systemic delivery of anti-PD1 antibody therapy has proven rela- Steven Leonardo, Keith Gorden, Ross B Fulton, Kathryn Fraser, Takashi O tively safe in glioma patients, but therapeutic response remains Kangas, Richard Walsh, Kathleen Ertelt, Jeremy Graff, Mark Uhlik unpredictable and persistently low. Checkpoint blockade anti- Biothera Pharmaceuticals Inc., Eagan, MN, USA bodies face numerous potential confounders in these tumors, Correspondence: Steven Leonardo (sleonardo@biothera.com) such as the blood-brain barrier, poor local or lymph node presen- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P27 tation of tumor antigens, and unknown dependency on PD-1/PD- L1 activity during tumor progression. Here, we conducted a pilot Background study using intracranial convection-enhanced delivery (CED) of Imprime PGG (Imprime) is a soluble, intravenously (iv) administered anti-PD1 (mDX400) into a de novo murine glioma model to the yeast 1,3/1,6 β-glucan PAMP (pathogen-associated molecular pattern). dissect tumor and immune perturbations following local treat- As a PAMP, Imprime triggers innate immune function, including direct ment, and to compare the efficacy of treating during early or late tumor killing, repolarization of the immunosuppressive tumor tumor progression. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 242 of 242 Methods produced several “long-term survivors”, who lived up to 158 days Transgenic C57BL/6-PTEN(fl/fl) mice were injected with a retro- with stable tumor burden. While substantial T cell infiltration was virus expressing PDGFb and cre recombinase, inducing tumori- detected in the end-stage tumors of both mDX400- and isotype- genesis as previously described. In this model, with a median treated mice by immunohistochemistry (CD3e), expression of im- survival of 80 days post-tumor induction (D80), convergence to a mune signaling pathways (e.g., Fc receptor and Toll-like receptor stereotyped subset of genomic rearrangements occurs by ap- families, phagosome/lysosome components), was significantly higher proximately D35. Intracranial osmotic pumps filled with mDX400 among three long-surviving mDX400-treated mice than in three or isotype control antibody solution were implanted at the tumor isotype-treated mice. site for 14-day windows spanning (D28-D42) or following (D42- Conclusions D56) this developmental transition, then removed. Tumor burden Our pilot study of mDX400 administration by CED identified an impact was monitored by bioluminescence (luciferase reporter), and mice on tumor burden during and following therapy, but a lack of survival were sacrificed upon presentation of tumor-related morbidity. Tis- benefit for D28-D42 treatment. While additional experiments are needed sue was formalin-fixed for histopathology and cryopreserved for to statistically evaluate survival benefit for the later treatment window, gene expression analysis. differentially high intratumoral expression of genes reflecting immune ac- Results tivation among mDX400-treated, long-surviving mice demonstrates that During both treatment windows, tumor burden decreased differen- molecular study in this model may elucidate intratumoral conditions as- tially in mDX400-treated mice. While survival time between mDX400- sociated with response to anti-PD1 blockade in glioma. and isotype-treated mice was nearly identical for D28-D42 (both median D70), for mice treated between D42-D56, median survival Acknowledgements differed (D88 vs. D68), but without statistical significance between This pre-clinical study is supported by Merck & Co. Investigator-Initiated the groups (p = 0.25). Interestingly, the D42-D56 mDX400 group Sponsored Projects grant LKR146174. Submit your next manuscript to BioMed Central and we will help you at every step: • We accept pre-submission inquiries � Our selector tool helps you to find the most relevant journal � We provide round the clock customer support � Convenient online submission � Thorough peer review � Inclusion in PubMed and all major indexing services � Maximum visibility for your research Submit your manuscript at www.biomedcentral.com/submit http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Journal for ImmunoTherapy of Cancer Springer Journals

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016): late breaking abstracts

Althammer, Sonja; Steele, Keith; Rebelatto, Marlon; Tan, Tze; Wiestler, Tobias; Schmidt, Guenter; Higgs, Brandon; Li, Xia; Shi, Li; Jin, Xiaoping; Antal, Joyce; Gupta, Ashok; Ranade, Koustubh; Binning, Gerd; Bellmunt, Joaquim; de Wit, Ronald; Vaughn, David; Fradet, Yves; Lee, Jae; Fong, Lawrence; Vogelzang, Nicholas; Climent, Miguel; Petrylak, Daniel; Choueiri, Toni; Necchi, Andrea; Gerritsen, Winald; Gurney, Howard; Quinn, David; Culine, Stéphane; Sternberg, Cora; Mai, Yabing; Puhlmann, Markus; Perini, Rodolfo; Bajorin, Dean; Sharma, Padmanee; Callahan, Margaret; Calvo, Emiliano; Kim, Joseph; de Braud, Filipo; Ott, Patrick; Bono, Petri; Pillai, Rathi; Morse, Michael; Le, Dung; Taylor, Matthew; Spilliopoulou, Pavlina; Bendell, Johanna; Jaeger, Dirk; Chan, Emily; Antonia, Scott; Ascierto, Paolo; Hennicken, Delphine; Tschaika, Marina; Azrilevich, Alex; Rosenberg, Jonathan; Levy, Ofer; Chan, Christopher; Cojocaru, Gady; Liang, Spencer; Ophir, Eran; Ganguly, Sudipto; Toporik, Amir; Kotturi, Maya; Kfir, Tal; Murter, Benjamin; Logronio, Kathryn; Dassa, Liat; Leung, Ling; Greenwald, Shirley; Azulay, Meir; Kumar, Sandeep; Alteber, Zoya; Pan, Xiaoyu; Machlenkin, Arthur; Benita, Yair; Drake, Andrew; Chajut, Ayelet; Salomon, Ran; Vankin, Ilan; Safyon, Einav; Hunter, John; Levine, Zurit; White, Mark; Leidner, Rom; Kang, Hyunseok; Haddad, Robert; Segal, Neil; Wirth, Lori; Ferris, Robert; Hodi, F.; Sanborn, Rachel; Gajewski, Thomas; Sharfman, William; McDonald, Dan; Srivastava, Shivani; Gu, Xuemin; Phillips, Penny; Passey, Chaitali; Seiwert, Tanguy; Habtetsion, Tsadik; Zhou, Gang; Sakellariou-Thompson, Donastas; Haymaker, Cara; Creasy, Caitlin; Hurd, Mark; Uraoka, Naohiro; Canales, Jaime; Koptez, Scott; Hwu, Patrick; Maitra, Anirban; Bernatchez, Chantale; Coyle, Scott; Roybel, Kole; Rupp, Levi; Santoro, Stephen; Secrest, Stephanie; Spelman, Michael; Ho, Hanson; Gomes, Tina; Tse, Tiffany; Yung-Wu, Chia; Taunton, Jack; Lim, Wendell; Emtage, Peter; Moudgil, Tarsem; Ballesteros-Merino, Carmen; Hilton, Traci; Paustian, Christopher; Leidner, Rom; Page, David; Urba, Walter; Fox, Bernard; Bell, Bryan; Patel, Ashish; Olafsen, Tove; Satpayev, Daulet; Torgov, Michael; Marchioni, Filippo; Romero, Jason; Jiang, Ziyue; Zamilpa, Charles; Keppler, Jennifer; Mascioni, Alessandro; Jia, Fang; Lee, Chen-Yu; Gudas, Jean; Sullivan, Ryan; Hoshida, Yujin; Logan, Theodore; Khushalani, Nikhil; Giobbie-Hurder, Anita; Margolin, Kim; Roder, Joanna; Bhatt, Rupal; Koon, Henry; Olencki, Thomas; Hutson, Thomas; Curti, Brendan; Blackmon, Shauna; Mier, James; Puzanov, Igor; Roder, Heinrich; Stewart, John; Amin, Asim; Ernstoff, Marc; Clark, Joseph; Atkins, Michael; Kaufman, Howard; Sosman, Jeffrey; Signoretti, Sabina; McDermott, David; Anderson, Abraham; Puzanov, Igor; Milhem, Mohammed; Andtbacka, Robert; Minor, David; Gorski, Kevin; Baker, Daniel; Hamid, Omid; Kaufman, Howard; Akporiaye, Emmanuel; Curti, Brendan; Koguchi, Yoshinobu; Leidner, Rom; Sutcliffe, Kim; Conder, Kristie; Urba, Walter; Marron, Thomas; Bhardwaj, Nina; Hammerich, Linda; George, Fiby; Kim-Schulze, Seunghee; Keler, Tibor; Davis, Tom; Crowley, Elizabeth; Salazar, Andres; Brody, Joshua; Monjazeb, Arta; Daly, Megan; Riess, Jonathan; Li, Tianhong; Murphy, William; Kelly, Karen; Hu, Zhiwei; Shen, Rulong; Campbell, Amanda; McMichael, Elizabeth; Yu, Lianbo; Ramaswam, Bhuvaneswari; London, Cheryl; Xu, Tian; Carson, William; Kokolus, Kathleen; Repasky, Elizabeth; Schell, Todd; Drabick, Joseph; Messenheimer, David; Jensen, Shawn; Fox, Bernard; Rubinstein, Mark; Andrijauskaite, Kristina; Swiderska-syn, Marzena; Lind, Kristin; Choppin, Agnes; Roell, Marina; Wrangle, John; Andrijauskaite, Kristina; Swiderska-syn, Marzena; Rhode, Peter; Wong, Hing; Rubinstein, Mark; Ahmad, Shamim; Webb, Mason; Abu-Eid, Rasha; Shrimali, Rajeev; Verma, Vivek; Doroodchi, Atbin; Berrong, Zuzana; Yashar, David; Samara, Raed; Mkrtichyan, Mikayel; Khleif, Samir; Powell, Steven; Gitau, Mark; Sumey, Christopher; Terrell, Andrew; Lohr, Michele; Nowak, Ryan; McGraw, Steven; Jensen, Ash; Blanchard, Miran; Gold, Kathryn; Cohen, Ezra; Ellison, Christie; Black, Lora; Lee, John; Spanos, William; Wennerberg, Erik; Schwitzer, Emily; Lhuillier, Claire; Koelwyn, Graeme; Hiner, Rebecca; Jones, Lee; Demaria, Sandra; Amanda, Vandeveer; Greiner, John; Schlom, Jeffrey; Bookstaver, Michelle; Jewell, Christopher; Paustian, Christopher; Gunderson, Andrew; Boulmay, Brian; Li, Rui; Spieler, Bradley; Happel, Kyle; Moudgil, Tarsem; Feng, Zipei; Ballesteros-Merino, Carmen; Dubay, Christopher; Fisher, Brenda; Koguchi, Yoshinobu; Aung, Sandra; Mederos, Eileen; Bifulco, Carlo; McNamara, Michael; Bahjat, Keith; Redmond, William; Ochoa, Augusto; Hu, Hong-Ming; Mehta, Adi; Lund-Johansen, Fridtjof; Fox, Bernard; Urba, Walter; Sanborn, Rachel; Hilton, Traci; Bedu-Addo, Frank; Conn, Greg; King, Michael; Dutta, Panna; Shepard, Robert; Einstein, Mark; Adams, Sylvia; Wang, Ena; Jin, Ping; Novik, Yelena; Morrison, Debra; Oratz, Ruth; Marincola, Franco; Stroncek, David; Goldberg, Judith; Demaria, Sandra; Formenti, Silvia; Galon, Jérôme; Mlecnik, Bernhard; Marliot, Florence; Ou, Fang-Shu; Bifulco, Carlo; Lugli, Alessandro; Zlobec, Inti; Rau, Tilman; Nagtegaal, Iris; Vink-Borger, Elisa; Hartmann, Arndt; Geppert, Carol; Roehrl, Michael; Bavi, Prashant; Ohashi, Pamela; Wang, Julia; Nguyen, Linh; Han, SeongJun; MacGregor, Heather; Hafezi-Bakhtiari, Sara; Wouters, Bradley; Kawakami, Yutaka; Papivanova, Boryana; Xu, Mingli; Fujita, Tomonobu; Hazama, Shoichi; Suzuki, Nobuaki; Nagano, Hiroaki; Okuno, Kiyotaka; Itoh, Kyogo; Zavadova, Eva; Vocka, Michal; Spacek, Jan; Petruzelka, Lubos; Konopasek, Bohuslav; Dundr, Pavel; Skalova, Helena; Torigoe, Toshihiko; Sato, Noriyuki; Furuhata, Tomohisa; Takemasa, Ichiro; Van den Eynde, Marc; Jouret-Mourin, Anne; Machiels, Jean-Pascal; Fredriksen, Tessa; Lafontaine, Lucie; Buttard, Bénédicte; Church, Sarah; Maby, Pauline; Angell, Helen; Angelova, Mihaela; Vasaturo, Angela; Bindea, Gabriela; Berger, Anne; Lagorce, Christine; Patel, Prabhu; Vora, Hemangini; Shah, Birva; Patel, Jayendrakumar; Rajvik, Kruti; Pandya, Shashank; Shukla, Shilin; Wang, Yili; Zhang, Guanjun; Masucci, Giuseppe; Andersson, Emilia; Grizzi, Fabio; Laghi, Luigi; Botti, Gerardo; Tatangelo, Fabiana; Delrio, Paolo; Cilberto, Gennaro; Ascierto, Paolo; Marincola, Franco; Sargent, Daniel; Fox, Bernard; Algazi, Alain; Tsai, Katy; Rosenblum, Michael; Nandoskar, Prachi; Andtbacka, Robert; Li, Amy; Nonomura, John; Takamura, Kathryn; Dwyer, Mary; Browning, Erica; Talia, Reneta; Twitty, Chris; Gargosky, Sharron; Campbell, Jean; Ballesteros-Merino, Carmen; Bifulco, Carlo; Fox, Bernard; Le, Mai; Pierce, Robert; Daud, Adil; Gartrell, Robyn; Marks, Douglas; Stack, Edward; Lu, Yan; Izaki, Daisuke; Beck, Kristen; Jia, Dan; Armenta, Paul; White-Stern, Ashley; Fu, Yichun; Blake, Zoe; Kaufman, Howard; Taback, Bret; Horst, Basil; Saenger, Yvonne; Leonardo, Steven; Gorden, Keith; Fulton, Ross; Fraser, Kathryn; Kangas, Takashi; Walsh, Richard; Ertelt, Kathleen; Graff, Jeremy; Uhlik, Mark; Sims, Jennifer; Lei, Liang; Tsujiuchi, Takashi; Bruce, Jeffrey; Canoll, Peter; Tolcher, Anthony; Alley, Evan; Chichili, Gurunadh; Canoll, Jan; Moore, Paul; Bonvini, Ezio; Johnson, Syd; Shankar, Sadhna; Vasselli, James; Wigginton, Jon; Powderly, John
Journal for ImmunoTherapy of Cancer , Volume 4 (2) – Dec 8, 2016

Loading next page...
 
/lp/springer-journals/31st-annual-meeting-and-associated-programs-of-the-society-for-6K1dBHSaYG

References (15)

Publisher
Springer Journals
Copyright
Copyright © 2016 by The Author(s).
Subject
Medicine & Public Health; Oncology; Immunology
eISSN
2051-1426
DOI
10.1186/s40425-016-0191-4
Publisher site
See Article on Publisher Site

Abstract

Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 DOI 10.1186/s40425-016-0191-4 MEETING ABSTRACTS Open Access 31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016): late breaking abstracts National Harbor, MD, USA. 9-13 November 2016 Published: 8 December 2016 tumors [3]. By 29APR2016, 304 previously treated NSCLC pa- Biomarkers and Immune Monitoring tients, median 3 prior lines, received 10 mg/kg of durvalumab q2w ≤12 months. Baseline archived or fresh tumor biopsies O1 were analyzed for PD-L1 (Ventana/SP263) and CD8 (Ventana/ Combinatorial CD8+ and PD-L1+ cell densities correlate with SP239) by IHC. For the marker combination, slides were scored response and improved survival in non-small cell lung cancer using the product of PD-L1+ and CD8+ cell densities with (NSCLC) patients treated with durvalumab 1 2 2 1 Definiens’ Developer XD 2.1.4 software. For PD-L1 alone, ≥25% Sonja Althammer , Keith Steele , Marlon Rebelatto , Tze Heng Tan , 1 1 2 2 2 tumor cells stained for PD-L1 at any intensity were scored posi- Tobias Wiestler , Guenter Schmidt , Brandon Higgs , Xia Li , Li Shi , 2 2 2 2 tive. Clinical outcomes (ORR, PFS and OS) were analysed based Xiaoping Jin , Joyce Antal , Ashok Gupta , Koustubh Ranade , Gerd on CD8+ and PD-L1+ densities (n = 163 available) and PD-L1 Binning 1 2 alone in pre-treatment biopsies using a discovery (n = 84) and Definiens AG, Munich, Bayern, Germany; MedImmune, Gaithersburg, validation (n = 79) set. Datasets were matched on baseline PD- MD, USA L1 status, histology, ECOG, lines of therapy, and response. Correspondence: Brandon Higgs (higgsb@medimmune.com) Results Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O1 Patients with high pretreatment CD8+ and PD-L1+ densities (preva- lence = 36%) had better ORR, OS, and PFS compared to those with Background low CD8+ and PD-L1+ densities (Fig. 1), as well as high PD-L1 expres- Immunotherapies have improved patient responses and sur- sion alone. vival, though not all patients benefit. Effective biomarkers may Conclusions help to improve outcomes. Durvalumab is a human IgG1 mono- Automated image analysis of CD8+ and PD-L1+ cell densities in clonal antibody that inhibits PD-L1 binding to PD-1 and CD80, baseline tumor biopsies may identify patients with improved out- restoring antitumor immunity [1, 2]. PD-L1 expression on tumor comes to durvalumab. or tumor-infiltrating immune cells measured manually with dif- Trial Registration ferent immunohistochemistry (IHC) assays can enrich for pa- ClinicalTrials.gov identifier NCT01693562. tients responding to anti-PD-1/PD-L1 agents. Tumor-infiltrating cytotoxic CD8+ T cells may also have potential predictive utility for therapeutic response. We explored automated image ana- References lysis and pattern recognition of tumor biopsies to determine 1. MedImmune/AstraZeneca. Data on file. whether CD8+ and PD-L1+ cell densities could better identify 2. Ibrahim R, Stewart R, Shalabi A: PD-L1 blockade for cancer treatment: patients most likely to respond to durvalumab than PD-L1 IHC MEDI4736. Semin Oncol 2015, 42:474–483. alone. 3. Rizvi NA, Brahmer JR, Ou SHI, Segal NH, Khleif S, Hwu WJ, et al: Safety Methods and clinical activity of MEDI4736, an anti-programmed cell death- CP1108/NCT01693562 was a nonrandomized phase I/II trial ligand 1 (PD-L1 antibody, in patients with non-small lung cancer evaluating durvalumab in advanced NSCLC and other solid (NSCLC). J Clin Oncol 2015, 33(Suppl.):Abstract 8032. © The Author(s). 2016 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 224 of 242 Methods Eligible patients were enrolled regardless of PD-L1 expression and randomized 1:1 to pembrolizumab 200 mg Q3W for 24 months or investigator’schoice ofpaclitaxel175 mg/m Q3W, docetaxel 2 2 75 mg/m Q3W, or vinflunine 320 mg/m Q3W. Randomization was stratified by ECOG PS (0/1 vs 2), liver metastases (yes vs no), hemoglobin level (<10 vs ≥10 g/dL), and time from last chemo- therapy dose (<3 vs ≥3 months). The study had a group sequen- tial design to control for type I error. Primary endpoints were OS and PFS (RECIST v1.1 by blinded, independent central review). ORR was a key secondary endpoint. Differences in OS and PFS were assessed in the intention-to-treat population using the stratified log-rank test Fig. 1 (abstract O1). Clinical outcomes in CD8+/PD-L1+ or PD-L1 Results NSCLC patient subsets Between November 5, 2014 and November 13, 2015, 542 patients from 29 countries were enrolled: 270 in the pembrolizu- mab arm, 272 in the chemotherapy arm. As of September 7, 2016, median follow-up was 9.0 months; 49 (18.4%) patients remained on pembrolizumab and 3 (1.2%) patients remained on chemotherapy. Baseline characteristics were generally balanced between arms, with 87.3% with visceral disease, 34.3% with liver Clinical Trials: Cutting-Edge (Completed metastases, 1.1% with ECOG PS 2, 81.5% with hemoglobin ≥10 g/dL, and 38.2% with <3 months since most recent chemo- Trials) therapy. Pembrolizumab significantly improved OS over chemo- therapy (HR 0.73, P = 0.0022; median 10.3 vs 7.4 months) (Table O2 1). There was no difference in PFS (HR 0.98, P = 0.42) (Table 1). Keynote-045: open-label, phase III study of pembrolizumab versus ORR was significantly improved with pembrolizumab (21.1% vs investigator’s choice of paclitaxel, docetaxel, or vinflunine for 11.4%) (Table 1). Pembrolizumab was associated with fewer any- previously treated advanced urothelial cancer 1 2 3 4 grade (60.9% vs 90.2%) grade 3-5 treatment-related AEs (15.0% Joaquim Bellmunt, Ronald de Wit, David J Vaughn, Yves Fradet, Jae 5 6 7 8 vs 49.4%). 4 patients in each arm died due to treatment-related Lyun Lee, Lawrence Fong, Nicholas J Vogelzang, Miguel A Climent, 9 1 10 11 AEs. Daniel P Petrylak, Toni K Choueiri, Andrea Necchi, Winald Gerritsen, 12 13 14 Conclusions Howard Gurney, David I Quinn, Stéphane Culine, Cora N 15 16 16 16 Pembrolizumab demonstrated a statistically significant OS bene- Sternberg, Yabing Mai, Markus Puhlmann, Rodolfo F Perini, Dean fit over chemotherapy in the second-line advanced urothelial F Bajorin 1 2 cancer setting, making it the first therapy to demonstrate a Dana-Farber Cancer Institute, Boston, MA, USA; Erasmus MC survival benefit over an active comparator in this population. Cancer Institute, Rotterdam, Netherlands; Abramson Cancer Center The superior OS combined with the lower rate of any-grade of the University of Pennsylvania, Philadelphia, PA, USA; CHU de and high-grade treatment-related AEs support pembrolizumab Québec-Université Laval, Québec, QC, Canada; Asan Medical Center as a new standard of care for advanced urothelial cancer that and University of Ulsan College of Medicine, Seoul, South Korea; progressed on/after platinum-based chemotherapy. University of California, San Francisco, San Francisco, CA, USA; Trial registration Comprehensive Cancer Centers of Nevada, Las Vegas, NV, USA; ClinicalTrials.gov identifier: NCT02256436 Fundación Instituto Valenciano de Oncología, Valencia, Spain; Smilow Cancer Hospital at Yale University, New Haven, CT, USA; Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy; Radboud University Medical Center, Nijmegen, Netherlands; Westmead Hospital and Macquarie University, Sydney, NSW, Australia; Univeristy of Southern California Norris Comprehensive 14 Table 1 (abstract O2) Efficacy in KEYNOTE-045 Cancer Center and Hospital, Los Angeles, CA, USA; Hôpital Saint- Louis, Paris, France; San Camillo Forlanini Hospital, Rome, Italy; End point Pembrolizumab Chemotherapy 16 17 Merck & Co., Inc., Kenilworth, NJ, USA; Memorial Sloan Kettering N = 270 N = 272 Cancer Center, New York, NY, USA OS, no. of events 155 179 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O2 Median (95 % CI), months 10.3 (8.0-11.8) 7.4 (6.1-8.3) Background HR (95 % CI) 0.73 (0.59-0.91); P = 0.0022 There is no standard second-line therapy for advanced urothe- PFS, no. of events 218 219 lial cancer. Although paclitaxel, docetaxel, and vinflunine are commonly used, they provide limited clinical benefit. KEYNOTE- Median (95 % CI), months 2.1 (2.0-2.2) 3.3 (2.3-3.5) 045 compared the efficacy and safety of the anti–PD-1 antibody HR (95 % CI) 0.98 (0.81-1.19); P = 0.42 pembrolizumab versus investigator-choice chemotherapy as second-line therapy for advanced urothelial cancer that pro- ORR (95 % CI) 21.1 % (16.4-26.5) 11.4 % (7.9-15.8) gressed or recurred following first-line platinum-based Treatment difference, % (95 % CI) 9.6 (3.5-15.9); P = 0.0011 chemotherapy. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 225 of 242 O3 Conclusions Efficacy and safety of nivolumab plus ipilimumab in metastatic Second-line treatment with N1I3 may provide the most favorable urothelial carcinoma: first results from the phase I/II CheckMate benefit-risk ratio among the regimens studied. If these interim results 032 study are confirmed with longer follow-up, further development of the 1 2 3 Padmanee Sharma , Margaret K Callahan , Emiliano Calvo , Joseph W N1I3 combination in metastatic urothelial carcinoma is warranted. 4 5 6 7 8 Kim , Filipo de Braud , Patrick A Ott , Petri Bono , Rathi N Pillai , Michael Trial Registration 9 10 11 12 Morse , Dung T Le , Matthew Taylor , Pavlina Spilliopoulou , Johanna ClinicalTrials.gov identifier NCT01928394. 13 14 15 16 Bendell , Dirk Jaeger , Emily Chan , Scott J Antonia , Paolo A 17 18 18 18 Ascierto , Delphine Hennicken , Marina Tschaika , Alex Azrilevich , References Jonathan Rosenberg 1. Sharma P, Bono P, Kim JW, et al: Efficacy and safety of nivolumab University of Texas MD Anderson Cancer Center, Houston, TX, USA; monotherapy in metastatic urothelial cancer (mUC): Results from the 2 3 Memorial Sloan Kettering Cancer Center, New York, NY, USA; START phase I/II CheckMate 032 study. J Clin Oncol 2016, 34(15 suppl): Madrid, Centro Integral Oncológico Clara Campal, Madrid, Spain; Yale Abstract 4501. Cancer Center, New Haven, CT, USA; Istituto Nazionale dei Tumori- Università degli Studi di Milano, Milano, Lombardia, Italy; Dana-Farber Cancer Institute, Boston, MA, USA; Comprehensive Cancer Center, Coinhibition & Costimulation Helsinki University Hospital and University of Helsinki, Helsinki, Finland; 8 9 Emory Winship Cancer Institute, Atlanta, GA, USA; Duke University O4 Medical Center, Durham, NC, USA; Sidney Kimmel Comprehensive Computational identification, functional characterization, and Cancer Center at Johns Hopkins University, Baltimore, MD, USA; antibody blockade of a new immune checkpoint in the TIGIT 11 12 Oregon Health & Science University, Portland, OR, USA; Beatson West family of interacting molecules 1 2 1 2 of Scotland Cancer Centre, Glasgow, United Kingdom; Sarah Cannon Ofer Levy , Christopher Chan , Gady Cojocaru , Spencer Liang , Eran 1 3 1 2 1 Research Institute and Department of Medical Oncology, Tennessee Ophir , Sudipto Ganguly , Amir Toporik , Maya Kotturi , Tal Fridman Kfir , 3 2 1 2 Oncology, Nashville, TN, USA; Heidelberg University Hospital, Benjamin M. Murter , Kathryn Logronio , Liat Dassa , Ling Leung , Shirley 1 1 2 1 4 Heidelberg, Baden-Wurttemberg, Germany; Vanderbilt-Ingram Greenwald , Meir Azulay , Sandeep Kumar , Zoya Alteber , Xiaoyu Pan , 1 1 2 1 University Medical Center, Nashville, TN, USA; H. Lee Moffitt Cancer Arthur Machlenkin , Yair Benita , Andrew W. Drake , Ayelet Chajut , Ran 1 1 1 2 1 Center, Tampa, FL, USA; Istituto Nazionale Tumori Fondazione Pascale, Salomon , Ilan Vankin , Einav Safyon , John Hunter , Zurit Levine , Mark Naples, Italy, Napoli, Italy; Bristol-Myers Squibb, Princeton, NJ, USA White 1 2 Correspondence: Padmanee Sharma (padsharma@mdanderson.org) Compugen Ltd., Holon, Israel; Compugen Inc, USA, South San Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O3 Francisco, CA, USA; Johns Hopkins University, Baltimore, MD, USA; Bloomberg ~ Kimmel Institute for Cancer Immunotherapy, Johns Background Hopkins University, Baltimore, MD, USA Nivolumab is a programmed death-1 (PD-1) immune checkpoint in- Correspndence: John Hunter (johnh@cgen.com) hibitor associated with clinical benefit in previously treated patients Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O4 with metastatic urothelial carcinoma [1]. Preclinical and clinical data indicate that the combination of nivolumab plus ipilimumab, an anti- Background cytotoxic T-lymphocyte antigen-4 (CTLA-4) antibody, can improve an- While antibody blockade of the CTLA-4 and PD-1 pathways has titumor activity in other tumor types. Here, we report the first effi- emerged as an effective treatment modality for cancer, the majority cacy and safety results of combined nivolumab plus ipilimumab of patients do not derive long-term benefit, suggesting a need for given at two different dosing schedules in CheckMate 032 , an open- targeting of additional immune checkpoints. Employing our unique label, multicenter, phase I/II study of patients with metastatic urothe- computational algorithms to define new members of the B7/CD28 lial carcinoma who progressed after prior platinum-based therapy. family, we identified PVRIG, which is expressed by multiple subsets Methods of T and NK cells. We report here its expression pattern, functional Patients with locally advanced or metastatic urothelial carcinoma pre- characterization, and anti-tumor activity of blocking antibodies tar- viously treated with platinum-based therapy were included in the geting this molecule. study. Patients were treated with either of two combination sched- Methods ules, nivolumab 1 mg/kg + ipilimumab 3 mg/kg (N1I3) or nivolumab Utilizing Compugen’s Predictive Discovery platform we identified 3 mg/kg + ipilimumab 1 mg/kg (N3I1) every 3 weeks for four cycles, PVRIG as a potential novel immune checkpoint, after which a retro- followed by nivolumab 3 mg/kg every 2 weeks; or they were treated viral cell screening library was used to identify its cognate binding with nivolumab monotherapy 3 mg/kg (N3) every 2 weeks. All pa- counterpart. Target effects on T cell modulation were assessed with tients were treated until disease progression or unacceptable toxicity. primary and tumor-derived T cell assays, taking advantage of target The primary endpoint was investigator-assessed objective response overexpression, knockdown, and antagonist antibody approaches. rate (ORR) by RECIST v1.1. Secondary endpoints included safety and Antibodies against the human protein were screened for their ability duration of response (DoR). to enhance T cell activation in vitro, while antibodies targeting the Results mouse orthologue were assessed in vivo for effects on tumor growth Minimum follow-up was 3.9 months in the N1I3 (n = 26) group, inhibition in syngeneic models. 14.5 months in the N3I1 group (n = 104), and 13.8 months in N3 Results group (n = 78). ORR was 38.5% (95% confidence interval [CI], 20.2- A PVRIG-Fc-fusion protein was found to bind PVRL2, with binding 59.4), 26.0% (95% CI, 17.9-35.5), and 25.6% (95% CI, 16.4-36.8) in the specificity confirmed both by ELISA and flow cytometry analysis. N1I3, N3I1, and N3 groups, respectively. Median DoR has not been PVRIG demonstrated unique expression kinetics upon T cell activa- reached in any treatment group. The frequency of drug-related grade tion, with detection of the target on memory T cells, as well as on 3-4 adverse events was 30.8% (N1I3), 31.7% (N3I1), and 23.1% (N3). NK cells and γδ T cells. A panel of high affinity human antibodies Treatment-related adverse events led to discontinuation in 7.7% with the ability to block interaction of PVRIG with PVRL2 were gener- (N1I3), 13.5% (N3I1), and 3.8% (N3) of patients. One death was re- ated, which when tested in vitro were shown to enhance activation ported in the N3I1 group (pneumonitis) and two deaths were re- of both primary CD4+ and tumor-derived CD8+ T cells through a ported in the N3 group (pneumonitis and thrombocytopenia). PVRL2-dependent mechanism. The lead antibody, COM-701, is Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 226 of 242 currently in preclinical development. Since COM-701 is not mouse Results cross-reactive, in vivo studies were conducted with a surrogate block- As of the August 30, 2016 data cutoff, 159 patients were treated with ing anti-mouse PVRIG antibody. When combined with anti-PD-L1 the lirilumab plus nivolumab combination. Treatment-related adverse blockade, anti-mouse PVRIG inhibits growth of established tumors in events (TRAEs) and grade 3–4 TRAEs were reported in 72% (114/159) both the CT26 and MC38 colorectal cancer models. Combination and 15% (24/159) of patients, respectively. Discontinuations due to testing with additional immune checkpoint inhibitors, as well as in TRAEs occurred in 8% (12/159). Of the 41 patients with SCCHN PVRIG knockout mice, is ongoing. treated, 29 were evaluable for response. In this heavily pretreated, Conclusions checkpoint inhibitor–naïve group, ORR was 24% (7/29; confirmed We describe the identification of PVRIG as a novel immune check- and unconfirmed) and DCR was 52% (15/29). Maximum reduction in point on T cells, as well the development of a high affinity antagonis- target lesions are presented in Fig. 2 for 26 patients with available tic antibody, COM-701, that is currently in preclinical development. tumor assessments. Two patients classified as stable disease per COM-701 is able to enhance human T cell activation, and a surrogate RECIST v1.1 showed unconventional responses, with 100% and 37% antibody with similar characteristics shows synergy with PD-L1 reductions in target lesions. Among evaluable patients, five (17%) in vivo in multiple syngeneic models. Overall, our data demonstrate had reductions in tumor burden > 80%. Responses appear durable, the utility of targeting PVRIG in addition to other B7 family check- with the median DOR not reached (Fig. 3). Updated efficacy and pre- points for the treatment of cancer. liminary biomarker analyses (including PD-L1 and HPV status) will be presented. Conclusions Combinations: Immunotherapy/ Preliminary efficacy of lirilumab plus nivolumab in patients with ad- vanced platinum-refractory SCCHN demonstrates clinical benefit, with Immunotherapy encouraging response rates that were deep and durable responses O5 in some patients. This combination demonstrated a manageable Preliminary efficacy from a phase I/II study of the natural killer safety profile similar to that observed with nivolumab monotherapy. cell–targeted antibody lirilumab in combination with nivolumab in Further evaluation of this novel combination of an NK-cell inhibitor squamous cell carcinoma of the head and neck and an immune checkpoint inhibitor is ongoing. 1 2 3 4 Rom Leidner , Hyunseok Kang , Robert Haddad , Neil H Segal , Lori J Trial Registration 5 6 3 7 Wirth , Robert L Ferris , F Stephen Hodi , Rachel E Sanborn , Thomas F ClinicalTrials.gov identifier: NCT01714739 8 9 10 10 Gajewski , William Sharfman , Dan McDonald , Shivani Srivastava , 10 10 10 8 Xuemin Gu , Penny Phillips , Chaitali Passey , Tanguy Seiwert Earle A. Chiles Research Institute, Robert W. Franz Cancer Center, Providence Portland Medical Center, Portland, OR, USA; Johns Hopkins Sidney Kimmel Comprehensive Cancer Center, Baltimore, MD, USA; 3 4 Dana-Farber Cancer Institute, Boston, MA, USA; Memorial Sloan Kettering Cancer Center, New York, NY, USA; Massachusetts General Hospital, Boston, MA, USA; University of Pittsburg, Pittsburgh, PA, USA; Earle A. Chiles Research Institute, Providence Cancer Center, Portland, 8 9 OR, USA; University of Chicago Medical Center, Chicago, IL, USA; The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Lutherville, MD, USA; Bristol-Myers Squibb, Princeton, NJ, USA Correspondence: Rom Leidner (rom.leidner@providence.org) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):O5 Fig. 2 (abstract O5). Maximum percent reduction in target lesions Background from baseline Natural killer (NK) cells and the innate immune system play a critical role in immunosurveillance, control of tumor growth, and metastasis. NK-cell activation is negatively regulated by inhibitory killer-cell immunoglobulin-like receptors (KIRs); therefore, blocking KIR function may potentiate an anti-tumor immune response and complement other immuno-oncology therapies that enhance T cell activity. We present preliminary efficacy results in patients with squamous cell carcinoma of the head and neck (SCCHN) from a phase I/II study of lirilumab, a fully human monoclonal antibody that blocks inhibitory KIRs on NK cells, in combination with nivolumab, a fully human IgG4 monoclonal antibody that targets the PD-1 receptor, in patients with solid tumors (NCT01714739). Methods During dose escalation, patients with advanced solid tumors who progressed after ≥ 1 prior therapy received lirilumab 0.1–3.0 mg/kg once every 4 weeks (Q4W) plus nivolumab 3.0 mg/kg Q2W. Cohort expansion was initiated at the maximum dose of lirilumab 3.0 mg/kg Q4W plus nivolumab 3.0 mg/kg Q2W in patients with advanced solid tumors. Key study endpoints include safety (primary), objective re- Fig. 3 (abstract O5). Percent change from baseline in target lesions sponse rate (ORR), disease control rate (DCR), duration of response over time (DOR), and biomarker assessments. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 227 of 242 Tumor Microenvironment References 1. Daud AI, Loo K, Pauli ML, Sanchez-Rodriguez R, Sandoval PM, Taravati K, et al: Tumor immune profiling predicts response to anti–PD-1 therapy Phase II study of intratumoral plasmid interleukin 12 (pIL-12) with in human melanoma. J Clin Invest 2016, 126(9):3447–3452. electroporation in combination with pembrolizumab in stage III/IV melanoma patients with low tumor infiltrating lymphocytes 1 1 1 1 Alain Algazi , Katy Tsai , Michael Rosenblum , Prachi Nandoskar , Robert Adoptive Cellular Therapy 2 1 3 3 HI Andtbacka , Amy Li , John Nonomura , Kathryn Takamura , Mary 3 3 3 3 3 Dwyer , Erica Browning , Reneta Talia , Chris Twitty , Sharron Gargosky , P1 3 4 4 Jean Campbell , Carmen Ballesteros-Merino , Carlo B. Bifulco , Bernard Chemo-immunotherapy with cyclophosphamide and tumor 4 5 3 1 Fox , Mai Le , Robert H Pierce , Adil Daud reactive CD4+ T cells lead to destruction of tumor vasculature and University of California, San Francisco, San Francisco, CA, USA; eventual tumor eradication Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, USA; Tsadik Habtetsion, Gang Zhou 3 4 OncoSec Medical Inc., San Diego, CA, USA; Robert W. Franz Cancer Augusta University, Augusta, GA, USA Research Center, Earle A. Chiles Research Institute, Providence Cancer Correspondence: Tsadik Habtetsion (tsadikg855@yahoo.com) Center, Portland, Oregon, USA; Doctor Hope, LLC, San Diego, CA, USA Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P1 Correspondence: Sharron Gargosky (sgargosky@oncosec.com) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):06 Background CD4 T cells are critical components of anti-tumor immunity and play Background a pivotal role in orchestrating anti-tumor immune responses. Mount- Low tumor infiltrating lymphocytes (TIL) are predictive for poor ing evidence from preclinical and clinical studies indicates that CD4 response to immunotherapy with anti-PD-1/PD-L1 agents. We T cells in combination with chemotherapy can control tumor growth have shown that melanoma patients with a low frequency of PD- and recurrence. CD4 T cells are suggested to mediate tumor rejec- 1hiCTLA-4 + TIL are unlikely to respond to pembrolizumab tion through mechanisms that include cytotoxic effects on tumor (Daud2016). Intratumoral electroporation of pIL-12 (IT-pIL12-EP) cells, inhibition of angiogenesis, and reprogramming of the tumor leads to an IFN-g signature suggestive of increased TIL as well as microenvironment. regression in both treated and untreated lesions. We hypothesize Methods that combination IT-pIL12-EP and pembrolizumab will improve In this project, we set out to study the cellular and molecular mecha- clinical outcomes in this low-response population. Preliminary re- nisms underlying the therapeutic effect of chemo-immunotherapy in sults from a multi-center, phase II, open-label trial testing this hy- the form of cyclophosphamide (CTX) and tumor specific CD4 T cells pothesis are presented. in a murine model of colorectal cancer. Mice were injected subcuta- Methods neously with colorectal cancer cells. When the tumor reached 140- Melanoma stage III/IV patients with accessible lesions were con- 160 mm in area, mice were injected with a low dose of cyclophos- sented and enrolled if they had a TIL status of hiCTLA-4+ in the phamide followed by adoptive transfer of tumor reactive CD4 T CD45 + CD8 + CD3+ gate by flow cytometry (FC). Patients were cells. treated with pembrolizumab (200 mg every 3 weeks) concurrently Results with IT-pIL12-EP on days 1, 5 and 8 every 6 weeks. Patients were In a murine model of colorectal cancer, we show that the combin- evaluated for overall response rate (ORR) every 12 weeks by ation therapy of CTX and tumor reactive CD4 T cells resulted in en- RECISTv1.1. Pre and post-treatment blood and tumor specimens were hanced necrosis of tumor cells in vivo, leading to eventual collected, and analyzed for immune phenotyping, gene expression, eradication of advanced tumors. By using immunofluorescence stain- TCR diversity, and changes in the tumor microenvironment with ing and blood perfusion imaging, we demonstrated that the combin- multispectral immunohistochemistry. ation therapy leads to destruction of the established tumor Results vasculature and reduced blood supply to tumor tissue. Furthermore, Interim ORR data is available on 15 patients. 13/15 patients had we assessed blood vessel permeability in the tumor tissue and found a frequency of PD-1hi CTLA-4+ TIL of < 22% (low TIL status), that the combination therapy increased extravasation of Evans blue phenotypes associated with a low probability of response to dye, suggesting an increase in vascular permeability. anti-PD-1 (Daud 2016). These 15 patients age 39-89 years, were Conclusions 53% male, 66% stage III and 34% stage IV. Treatment was well In summary, our findings suggest that the combination therapy of tolerated; 38% of adverse events (AE) were classified as treat- CTX + CD4 T cells leads to destruction of the tumor vasculature, ment site reactions (grade 1-2) that resolved. One SAE of cellu- resulting in extensive necrosis of tumor tissue and eventual tumor re- litis resolved with 5d antibiotics. One grade 3 AE of diarrhea gression. These findings may provide new insights into mechanisms resolved with corticosteroids. The ORR was 40% (4CR, 2PR) by of tumor rejection by CD4 T cells. RECISTv1.1. Analysis of tumor biopsies and blood demonstrated meaningful immunological changes including an increased num- ber and ratio of CD8+:PD-L1+ and CD8+:FoxP3 + TIL, tumoral P2 RNA signatures indicating an increase in CD8 and IFN-γ-related Preclinical development of tumor-infiltrating lymphocyte therapy gene expression and concordant immune phenotypes in the for pancreatic cancer periphery. Donastas Sakellariou-Thompson, Cara Haymaker, Caitlin Creasy, Mark Conclusions Hurd, Naohiro Uraoka, Jaime Rodriguez Canales, Scott Koptez, Patrick The combination IT-pIL12-EP with pembrolizumab in patients with Hwu, Anirban Maitra, Chantale Bernatchez an anti-PD-1 non-responsive phenotype engendered a 40% clinical University of Texas MD Anderson Cancer Center, Houston, TX, USA response with associated positive immune-based biomarker data Correspondence: Donastas Sakellariou-Thompson and an excellent safety profile. These data suggests that IT-pIL12-EP (dasakellariou@mdanderson.org) modulates the tumor microenvironment to enable an effective anti- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P2 PD-1 mAb response in patients otherwise unlikely to respond. Background Acknowledgements Immunotherapy has become an effective cancer therapy, particularly We thank Merck and Oncosec for supporting this trial with pembrolizumab in the case of checkpoint blockade and adoptive T cell therapy (ACT). and IT-pIL-12, respectfully. ACT exploits the presence of tumor-infiltrating lymphocytes (TIL) by Trial Registration exponentially expanding their numbers ex vivo and re-infusing them ClinicalTrials.gov identifier: NCT02493361 into the patient in an autologous setting. With the effectiveness of Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 228 of 242 TIL therapy already well established in multiple phase II studies in Results melanoma, there is a push to translate it to other cancers in dire Here we describe advances in our ON-switch CAR design that allow need of improved therapies. Pancreatic ductal adenocarcinoma for dose-dependent antigen-specific T cell activation in the presence (PDAC) is one such cancer for which the current therapy, surgery and of an FDA-approved non-immunosuppressive small molecule dimeri- chemotherapy, provides an overall 5-year survival rate of only 5%. zer. Furthermore, we demonstrate that our synNotch T cells are able The presence of TIL is correlated with increased survival in PDAC, to deliver therapeutic payloads capable of, but not limited to, modu- which suggests that TIL could effectively control the disease and pro- lating the tumor microenvironment and changing the cell-intrinsic vides a rationale to test TIL therapy in this setting. transcriptional properties of the synNotch T cells. Methods Conclusions To assess the feasibility, we characterized the immune component of We have successfully identified a heterodimerizing switch receptor PDAC, explored the ability to grow and expand TIL from tumor frag- that specifically activates T cells in the presence of an FDA-approved ments, and analyzed the clonality of these expanded TIL. non-immunosuppressive small molecule. The ability to control the Results potency of the CAR-mediated immune response in this way may re- Flow cytometry analysis detected low, CD4-rich T cell infiltration. These duce the toxicity associated with CAR-T therapy. Furthermore, we TIL were able to be expanded ex vivo and the addition of an agonistic have demonstrated that synNotch T cells are able to sculpt the anti- anti-41BB antibody to the cultures preferentially increased total TIL out- tumor immune response in both a T cell intrinsic (transcriptional pro- growth, particularly that of CD8 TIL. The success rate of TIL growth graming) and T cell extrinsic (therapeutic payload) manner, providing was increased from 23% to 50% for cultures grown without and with a customizable platform for altering T cell function and the tumor anti-41BB respectively. Sequencing of the T cell receptor CDR3-beta microenvironment. chain found specific T cell clones enriched at the tumor site in compari- son to the blood. IHC staining for MHC class I (MHCI) on PDAC tumor References samples showed that it is widely expressed but at low levels generally. 1. Wu C-Y, Roybal KT, Puchner EM, Onuffer J, Lim, WA: Remote control of Conclusions therapeutic T cells through a small molecule-gated chimeric receptor. In conclusion, it is possible to expand CD8 T cells from PDAC bear- Science 2015 350: aab4077. ing TCR sequences highly enriched in the tumor. Additionally, ex- 2. Roybal KT, Rupp LJ, Morsut L, Walker WJ, McNally KA, Park JS, Lim W: panded TIL would be able to target tumor cells as they are shown to Precision Tumor Recognition by T Cells With Combinatorial Antigen- express MHCI. Although there are barriers yet to overcome, the initial Sensing Circuits. Cell 164: 770–779. data suggest the feasibility of TIL therapy for PDAC. Biomarkers and Immune Monitoring P3 P4 Synthetic biology approaches to enhance adoptive cell therapy Evaluation of anticancer immunity in patients with thyroid cancer safety and precision with a focus towards developing effective combination 1 1 1 1 Scott M Coyle , Kole T Roybel , Levi J Rupp , Stephen P Santoro , immunotherapy 1 1 1 1 1 1 2 Stephanie Secrest , Michael Spelman , Hanson Ho , Tina Gomes , Tiffany Tarsem Moudgil , Carmen Ballesteros-Merino , Traci Hilton , Christopher 1 2 3 3 1 2 3 4 4 1 Tse , Chia Yung-Wu , Jack Taunton , Wendell Lim , Peter Emtage Paustian , Rom Leidner , David Page , Walter Urba , Bernard Fox , Bryan 1 2 3 1 Cell Design Labs, San Francisco, CA, USA; Amgen, San Francisco, CA, Bell , Ashish Patel 1 2 USA; University of California San Francisco, San Francisco, CA, USA Providence Portland Medical Center, Portland, OR, USA; UbiVac, Correspondence: Stephen P Santoro (steve@celldesignlabs.com) Portland, OR, USA; Earle A. Chiles Research Institute, Robert W. Franz Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P3 Cancer Center, Providence Portland Medical Center, Portland, OR, Portland, OR, USA; Earle A. Chiles Research Institute, Providence Cancer Background Center, Portland, OR, USA Chimeric antigen receptor T cells (CAR-T) have shown impressive effi- Correspondence: Bernard Fox (foxb@foxlab.com) cacy against numerous hematological malignancies, yet a high per- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P4 centage of individuals receiving these therapies experience toxicity in the form of cytokine release syndrome (CRS) and/or normal tissue Background destruction. Furthermore, solid tumors represent a substantive chal- Thyroid cancer is the most common endocrine-related cancer with lenge for CAR therapy due to a lack of tumor- specific antigens and 64,330 diagnoses expected this year. While the majority of these can- general inability of T cells to overcome immunosuppressive tumor cers are curable, almost 2% of these cancers are anaplastic thyroid microenvironments. We have sought to circumvent these obstacles cancers, which are highly aggressive and almost uniformly lethal. At by utilizing synthetic biology approaches to augment CAR-T function the same time, the thyroid is known for being inherently immuno- and specificity. genic. For these reasons and due to an active surgical practice pro- Methods viding regular resections of thyroid cancers, we undertook a study of We developed two platform technologies that aim to mitigate tox- thyroid cancer with the idea of developing an immunotherapy for icity associated with CAR-T therapy and endow T cells with environ- this disease. mental sensing capabilities that enhance tumor discrimination from Methods normal tissue and/or confer the ability to generate customizable re- We have developed a thyroid cancer tumor bank to complement our sponse outputs. Firstly, we engineered an “ON-switch” CAR that con- Oral, Head and Neck Cancer Program. This tumor bank cryopreserves sists of two protein modules that undergo heterodimerization and enzymatically isolated viable cells from resected tumors (n = 16).We become competent for signaling only in the presence of a small- are also attempting to develop primary cell lines and are isolating molecule dimerizing agent [1]. In addition, we created a “synthetic and assessing autologous tumor-specific functions of tumor- Notch” (synNotch) receptor, which we previously described in the infiltrating lymphocytes (TIL) (n = 7). context of combinatorial antigen sensing [2], that is capable of driv- Results ing expression of any number of downstream polypeptides in re- To date we have established 3 tumor cell lines from thyroid cancer sponse to antigen engagement. specimens and identified PD-L1 expression on 2 of 2 tested. While Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 229 of 242 numbers are small, preliminary analyses suggest that TIL cultures can (2.1 mSv/MBq) and LLI wall at 6.5 rem/mCi (1.8 mSv/MBq). A GLP be generated from 85% of thyroid cancer specimens and that autolo- toxicology study was conducted in cynomolgus monkeys that in- gous tumor-reactive TIL can be detected in 43% (n = 7) of thyroid cluded multiple dose cohorts of Zr-Df-IAB22M2C and a vehicle con- cancers. Since not every tumor appears to contain TIL capable of rec- trol. The results showed that doses up to 25 mg/kg of Zr-Df- ognizing autologous tumor, strategies to prime tumor-specific T cells IAB22M2C administered weekly to cynomolgus monkeys did not re- represents an area of interest. DPV-003 is a microvesicle vaccine, sult in any treatment-related findings in survival, clinical signs, body DRibble, that contains more than 80 proteins that are overexpressed weights, food consumption, ophthalmic examinations, electro- by thyroid cancer (TCGA provisional RNASeq n = 509 pts). The vac- cardiology, blood pressure, heart rate, clinical and anatomic path- cine also contains a number of DAMPs and agonist activity for mul- ology, peripheral blood lymphocyte population, and cytokine levels. tiple TLRs packed into stable double membrane microvesicles that Conclusions are targeted to CLEC9A+ antigen presenting cells. We are also devel- Zr-Df-IAB22M2C has the desired sensitivity and safety profile for im- oping a second thyroid-specific DRibble vaccine from a cell line de- aging CD8+ T cells and the first-in-human studies will commence in rived from an anaplastic thyroid cancer. the Q4 2016. Conclusions Almost half of thyroid cancers evaluated, including one anaplastic thyroid cancer, contain T cells capable of recognizing autologous P6 cancer cells and secreting IFN-g. However, the other 50% of thyroid High dose interleukin- 2 (HD IL-2) select trial in melanoma: a tissue cancers appear to lack tumor-reactive T cells and may benefit from and blood collection protocol to identify predictive biomarkers of combination immunotherapy strategies that include a vaccine. benefit to HD IL-2 in patients with advanced melanoma 1 2 3 4 Ryan J Sullivan , Yujin Hoshida , Theodore Logan , Nikhil Khushalani , 5 6 7 8 Acknowledgements Anita Giobbie-Hurder , Kim Margolin , Joanna Roder , Rupal Bhatt , 9 10 11 12 Support: Steve and Cindy Harder, Robert W. and Elsie Franz, Wes and Nancy Henry Koon , Thomas Olencki , Thomas Hutson , Brendan Curti , 13 8 14 7 Lematta, Lynn and Jack Loacker, and The Chiles foundation (BAF). Shauna Blackmon , James W Mier , Igor Puzanov , Heinrich Roder , 15 16 14 17 John Stewart , Asim Amin , Marc S Ernstoff , Joseph I Clark , Michael 18 19 20 8 B Atkins , Howard L Kaufman , Jeffrey Sosman , Sabina Signoretti , P5 David F McDermott Development and clinical translation of 89Zr-Df-IAB22M2C for Medical Oncology Department, Massachusetts General Hospital, Boston, detecting CD8+ T Cells for immunotherapy applications MA, USA; Hess Center for Science and Medicine; Tisch Cancer Institute, 1 2 1 1 Tove Olafsen , Daulet Satpayev , Michael Torgov , Filippo Marchioni , New York, NY, USA; Simon Cancer Center, Indiana University, 1 1 1 Jason Romero , Ziyue Karen Jiang , Charles Zamilpa , Jennifer S Indianapolis, IN, USA; H. Lee Moffitt Cancer Center, Tampa, FL, USA; 1 1 1 1 1 Keppler , Alessandro Mascioni , Fang Jia , Chen-Yu Lee , Jean Gudas Department of Biostatistics & Computational Biology, Boston, MA, USA; 1 2 ImaginAb Inc., Inglewood, CA, USA; AdicetBio, Inc., Menlo Park, CA, Department of Medical Oncology, City Of Hope, Duarte, CA, USA 7 8 USA Biodesix, Inc., Boulder, CO, USA; Beth Israel Deaconess Medical Center, Correspondence: Jean Gudas (jgudas@imaginab.com) Boston, MA, USA Case Western Reserve University, Cleveland, OH, USA; 10 11 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P5 The Ohio State University, Columbus, OH, USA; Texas Oncology- Baylor Charles A. Sammons Cancer Center, Dallas, TX, USA; Earle A. Background Chiles Research Institute, Providence Cancer Center, Portland, OR, USA; Immunotherapies are changing the landscape for cancer treatment; how- Massachusetts General Hospital Cancer Center, Boston, MA, USA; 14 15 ever, the field is hampered by the lack of biomarkers that can be used Roswell Park Cancer Institute, Buffalo, NY, USA; Wake Forest Baptist for patient selection and for monitoring treatment responses rapidly and Medical Center, Winston Salem, NC, USA; Levine Cancer Institute, non-invasively. To address this need, ImaginAb is developing Zr-Df- Carolinas HealthCare System, Charlotte, NC, USA; Loyola University IAB22M2C, an ~80 kDa minibody (Mb) with high affinity to the CD8 Medical Center, Maywood, IL, USA; Georgetown-Lombardi glycoprotein (binding EC = 0.4 nM) conjugated with desferrioxamine Comprehensive Cancer Center, Washington , DC, USA; Rutgers Cancer (Df) and radiolabeled with the positron emitting radionuclide Zirconium- Institute of New Jersey, New Brunswick, NJ, USA; Robert Lurie 89 ( Zr; T 78.4 hours) for imaging CD8+ T cells in humans. Comprehensive Cancer Center of Northwestern University, Chicago, IL, USA 1/2 Methods Correspondence: Ryan J Sullivan (rsullivan7@mgh.harvard.edu) A comprehensive preclinical program that included evaluation of the Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P6 in vitro and in vivo pharmacodynamics of IAB22M2C (unconjugated Mb), Df-IAB22M2C (conjugated Mb intermediate), Zr-Df-IAB22M2C (Zr Background chelated, conjugated non-radiolabeled form of final drug) and Zr- HD IL-2 provides objective responses in 15-20% and durable complete Df-IAB22M2C (radioactive final drug product) was conducted to dem- remission in 5-8% of patients with metastatic melanoma (MM). We pre- onstrate the safety and potential efficacy of the probe. viously identified a gene expression-based tumor subclass character- Results ized by immune related genes (Class 2; C2) associated with durable In vitro studies using human PBMCs from 10 individual human do- response to HD IL-2 compared to the remaining tumors that overex- nors showed no measurable or reproducible impact on proliferation, pressed lineage-associated genes (Class 1; C1). The primary objective of activation or depletion of CD8+ T cells and no consistent release of the HD IL-2 select trial in melanoma was to prospectively validate the cytokines when donor CD8+ T cells were exposed to soluble or favorable gene expression signature (C2). Secondary objectives were to immobilized Mb protein. Studies that evaluated the effect of saturat- seek serum and tissue biomarkers of durable response. ing concentrations of Zr-Df-IAB22M2C on proliferation and viability Methods of CD8+ T cells in vitro, also showed no impact on these parameters. 170 patients with MM were enrolled at 15 Cytokine Working Group Preclinical imaging and biodistribution studies demonstrated favor- sites from 2010 to 2014. All patients had formalin-fixed paraffin- able pharmacokinetics and the ability of Zr-Df-IAB22M2C to detect embedded (FFPE) tumor tissues identified and blood drawn prior to infiltrating CD8+ T cells in a mouse hu-PBMC NSG™ GvHD model and HD IL-2. Tumor assessments used WHO criteria and investigator- TM in Matrigel plugs implanted with different numbers of human CD8 assessed outcomes. RNA extracted from FFPE tumor tissues was used TM + T cells. Radiation dosimetry studies conducted in hu-CD34 NSG for whole transcriptome profiling by RNA sequencing (114 samples mice and the results GLP dosimetry analysis showed that on average, yielded sufficient RNA, 101 passed default Quality Control (QC)). Pre- the organs receiving the largest dose equivalent were the kidneys at treatment serum from 114 patients served as the test set and was 8.0 rem/mCi (2.2 mSv/MBq) followed by the liver at 7.9 rem/mCi analyzed using matrix-assisted laser desorption/ionization (MALDI) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 230 of 242 and machine-based learning algorithms to identify a predictive pro- starting at week 6 for 4 infusions. Peripheral blood was obtained tein expression signature. (Paxgene RNA) at baseline and at weeks 4, 6, 9, and 15. Gene expres- Results sion (Agilent Microarray) was analyzed for changes in expression Thirty-one of 170 pts (18.2%) responded, and median overall survival level with treatment. Pharmacodynamic markers were identified with was 21.3 months, with a 40 month median follow-up. Analysis of a linear mixed effects model. False discovery was controlled with per- RNAseq from 101 patients whose specimens passed QC showed that mutation testing. a C2 signature was associated with response to HD IL-2 (normalized Results enrichment score 1.70, false discovery rate 0.004). Using MALDI, a Gene expression was measured in 16 patients in phase Ib. Most treat- protein expression signature enriched for acute phase proteins (in- ment effects on expression were seen after ipilimumab treatment, cluding CRP, IL-6, and SAA) was defined in the pre-treatment serum but there were a few effects in the initial T-VEC phase that passed and used to classify 39 patients into group A (non-acute phase pro- false discovery controls. These T-VEC effects included SELV, SYNPO, tein expression) and 75 patients in group B (acute phase protein ex- ZBTB32, IQCF2, CDC27, KLK1, PRR20B, CHST6, and IGH. ZBTB32 has pression). Complete response rate in group A was 21% and zero in been reported to control the proliferative burst of virus-specific nat- group B (p = 0.0001). Two-year PFS rate was 29% in group A com- ural killer cells responding to infection. The combination effects were pared to 4% in group B (p = 0.0005). enriched for genes involved in lymphoid tissue structure and devel- Conclusions opment and immune cell trafficking. 185 of these genes had signs of In this prospective biomarker validation study, HD IL-2 produced dur- a T-VEC effect in the monotherapy phase. These included increases able remissions and prolonged survival in patients with MM. A in GZMM, PDCD1, CD8B, CD8A, and CTLA4 and decreases in IL18, tumor-derived gene expression signature enriched for immune- IRAK3, and TXNRD1. related genes was associated with response. Additionally, preliminary Conclusions data with a serum protein signature appears to identify patients This hypothesis-generating microarray analysis identified genes up- most likely to have a complete response. regulated in circulating peripheral blood cells after T-VEC monother- Trial Registration apy and combination treatment. We plan to further evaluate these ClinicalTrials.gov identifier NCT01288963. genes and other potential pharmacodynamic markers in phase II. Trial Registration ClinicalTrials.gov identifier NCT01740297. P7 Pharmacodynamic gene expression changes from talimogene References laherparepvec (T-VEC) plus ipilimumab in a phase Ib study for 1. Puzanov I, Milhem MM, Minor D, Hamid O, Li A, Chen L, et al: metastatic melanoma Talimogene laherparepvec in combination with ipilimumab in 1* 2 3 Abraham A Anderson , Igor Puzanov , Mohammed M Milhem , Robert previously untreated, unresectable stage IIIB-IV melanoma. J Clin Oncol 4 5 1 1 HI Andtbacka , David Minor , Kevin S Gorski , Daniel M Baker , Omid 2016, 34:2619–2626. 6 7 Hamid , Howard L Kaufman 1 2 Amgen Inc., Thousand Oaks, CA, USA; Roswell Park Cancer Institute, Buffalo, NY, USA; University of Iowa Hospitals and Clinics, Iowa City, IA, Clinical Trials in Progress USA; University of Utah, Huntsman Cancer Institute, Salt Lake City, UT, 5 6 USA; California Pacific Melanoma Center, San Francisco, CA, USA; The P8 Angeles Clinic & Research Institute, Los Angeles, CA, USA; Rutgers Phase I study of alpha-tocopherlyoxyacetic acid in patients with Cancer Institute of New Jersey, New Brunswick, NJ, USA advanced cancer: immune response and pharmacokinetics results 1 2 2 Correspondence: Abraham A Anderson (andersoa@amgen.com) Emmanuel Akporiaye , Brendan Curti , Yoshinobu Koguchi , Rom 3 3 3 2 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P7 Leidner , Kim Sutcliffe , Kristie Conder , Walter Urba 1 2 Veana Therapeutics Inc, Portland, OR, USA; Earle A. Chiles Research Background Institute, Providence Cancer Center, Portland, OR, USA; Earle A. Chiles T-VEC is a herpes simplex virus type-1 based oncolytic immunother- Research Institute, Robert W. Franz Cancer Center, Providence Portland apy designed to selectively replicate in tumors, produce GM-CSF, and Medical Center, Portland, OR, USA stimulate antitumor immune responses. Ipilimumab is a checkpoint Correspondence: Emmanuel Akporiaye (etakporiaye@gmail.com) inhibitor that promotes T cell activation by blocking negative signal- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P8 ing through CTLA-4. Both agents have demonstrated activity as monotherapy in advanced melanoma. Based on the potential com- Background plementary MOA of the agents, tumor cell lysis and antigen presen- Alpha-tocopheryloxyacetic acid (α-TEA) targets tumor cell mitochon- tation (T-VEC) in combination with T cell checkpoint inhibition, we dria to release reactive oxygen species (ROS) that induce immuno- hypothesized that improved efficacy was possible when the agents genic cell death (ICD), antigen release, and enhanced antigen cross- are used in combination. Because the safety profiles are non- presentation in pre-clinical models. α-TEA is being evaluated for overlapping, the combination was not anticipated to have significant safety and tolerability in a first-in-human phase I trial in patients with increased toxicity. To address these hypotheses, a phase Ib/II study advanced cancers (NCT02192346). Tumor types in the ongoing trial evaluating the safety and efficacy of T-VEC plus ipilimumab for Stage include renal cancer, esophageal adenocarcinoma, thyroid cancer, III-IV metastatic melanoma was initiated. The phase Ib study was duodenal cancer, and squamous cell carcinoma of the head and completed (N = 19) with no DLTs (primary endpoint) or new safety neck. signals with combination treatment, and an ORR of 50% [1]. Phase Ib Methods also included biomarker analyses investigating potential pharmaco- α-TEA lysine salt is administered orally to patients and given daily in dynamic markers for T-VEC monotherapy and in combination with escalating doses for 28 days. Immune monitoring of peripheral whole ipilimumab. blood was conducted for all 12 patients at baseline, and at 1 week Methods and 4 weeks post-treatment. Plasma levels of α-TEA have been deter- 6 8 Nineteen patients received T-VEC at 10 PFU/mL at week 1, then 10 mined so far in patients receiving 2.4 mg/kg and 4.8 mg/kg α-TEA at PFU/mL Q2W from week 4. Ipilimumab was given at 3 mg/kg Q3W 1, 4, 8, and 24 hours after the first dose. Additional samples were Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 231 of 242 evaluated on days 8, 15, 22, and 29 before the planned α-TEA dose potential for cytolytic activity directed at normal tissues, and unin- on those days. tended toxicity in patients treated with MGD009. Results Methods Twelve patients have been treated so far at 2.4 mg/kg and 4.8 mg/ This multi-center, open-label trial is a phase I dose escalation/cohort kg dose levels. Eight patients have stable disease, lasting from 1 to expansion study. All patients must have advanced B7-H3-positive tu- 22+ months. One patient showed more than a 2-fold increase in the mors. Prior checkpoint inhibitor therapy will be allowed. Dose escal- number of activated (CD38+ HLA-DR+) effector CD8+ T cells 1 week ation uses a 3+3+3 design, with patients treated every 2 weeks with post-treatment. A second patient showed more than a 2-fold in- escalating doses of IV MGD009 (starting dose 0.3 ug/kg). The dose crease in the number of activated effector memory CD8+ T cells escalation phase enrolls patients with mesothelioma, bladder cancer, 4 weeks post-treatment. Both patients experienced stable disease melanoma, SCCHN, NSCLC, clear cell renal cell carcinoma, ovarian over 5 and 22 months, respectively. Evaluation of α-TEA levels at the cancer, thyroid cancer, triple-negative breast cancer, pancreatic can- 2.4 mg/kg and 4.8 mg/kg doses revealed a proportional increase in cer, colon cancer, soft tissue sarcoma, or prostate cancer. Cohort ex- α-TEA plasma levels over a 28-day interval without any indication pansions (n=16/cohort) treated at the maximum tolerated dose will that steady state plasma levels were reached. Of the 12 patients, 6 include patients with melanoma, SCCHN, mesothelioma, urothelial developed atrial fibrillation (AF) after starting α-TEA. The earliest cancer, and NSCLC. Pre- and on-study biopsies are required for mel- event occurred 7 days post-treatment, but AF was more common 29- anoma patients in the cohort expansion phase. Primary and second- 56 days post-treatment. Four of the 6 patients had a medical history ary study objectives include characterization of the safety, of AF. These were grade 2 events by CTCAE 4.0 criteria and managed pharmacokinetics, pharmacodynamics, and preliminary antitumor ac- with appropriate medication without further sequelae. tivity of MGD009. All tumor evaluations will be carried out by both Conclusions Response Evaluation Criteria in Solid Tumors (RECIST 1.1) and im- α-TEA treatment resulted in stable disease in 80% of patients lasting mune-related RECIST. between 1 and 22+ months. AF was observed commonly in patients Trial Registration with a medical history of AF, and was managed with appropriate ClinicalTrials.gov identifier NCT02628535. medication. No clinically meaningful grade 3 or 4 toxicities were ob- served. Plasma α-TEA levels increased proportionally without any in- dication that steady state levels were achieved. α-TEA may function P10 through enhancing pre-existing CD8+ T cell-mediated anti-tumor Intratumoral Flt3L and poly-ICLC combined with low dose activity. radiotherapy: a novel in situ vaccine for incurable indolent lymphomas 1 1 1 1 Thomas Marron , Nina Bhardwaj , Linda Hammerich , Fiby George , 1 2 2 2 P9 Seunghee Kim-Schulze , Tibor Keler , Tom Davis , Elizabeth Crowley , 3 4 Phase I, first-in-human, open label, dose escalation study of Andres Salazar , Joshua Brody 1 2 MGD009, a humanized B7-H3 x CD3 dual-affinity re-targeting Icahn School of Medicine at Mount Sinai, New York, NY, USA; Celldex (DART®) protein in patients with B7-H3-expressing neoplasms or Therapeutics, Hampton, NJ, USA Oncovir, Inc., Washington, DC, USA; B7-H3 expressing tumor vasculature Mount Sinai School of Medicine, New York, NY, USA 1 2 3 Anthony W Tolcher , Evan W Alley , Gurunadh Chichili , Jan E Correspondence: Thomas Marron (thomas.marron@mountsinai.org) 4 3 3 3 3 Baughman , Paul Moore , Ezio Bonvini , Syd Johnson , Sadhna Shankar , Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P10 3 3 5 James Vasselli , Jon Wigginton , John Powderly START - South Texas Accelerated Research Therapeutics, LLC, San Background 2 th Antonio, TX, USA; Penn Presbyterian Medical Center, University of Lymphomas are the 5 most common cancer in the United States, Pennsylvania, Philadelphia, PA, USA; MacroGenics, Inc., Rockville, MD, and unlike more aggressive lymphomas, indolent non-Hodgkin 4 5 USA; MacroGenics, Inc., South San Francisco, CA, USA; Carolina lymphoma (iNHL) and is incurable with standard therapy. A previous BioOncology Institute, PLLC, Huntersville, NC, USA trial of in situ vaccination in iNHL combined intratumoral injection of Correspondence: Jan E Baughman (baughmanj@macrogenics.com) a TLR9 ligand with radiation to induce a systemic immune response Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P9 against tumor. This approach induced tumor-specific CD8+ T cell re- sponses and durable clinical remissions of patients’ untreated sites of Background disease were seen in a minority of patients. One limitation in this MGD009 is a B7-H3 x CD3 dual affinity re-targeting (DART) protein. previous trial may have been the scarcity of intratumoral dendritic DART proteins are bispecific, antibody-based molecules that can bind cells (DCs); DCs are uniquely able to endocytose dying tumor cells two distinct antigens simultaneously. MGD009 is designed to redirect for cross-presentation to tumor antigen-specific CD8+ T cells. In our T cells to eliminate B7-H3-expressing target cells through co-engage- novel iteration of in situ vaccine, intratumoral FMS-like tyrosine kin- ment of B7-H3 on the target cell and CD3 on the T cell, each in a ase 3 ligand (Flt3L) is introduced as a priming step to increase the monovalent binding manner. MGD009 contains a mutated human presence of intratumoral DCs. Flt3L induced tumor leukocyte infiltra- IgG1 Fc domain to reduce/eliminate binding to Fc gamma receptors tion and regression in lymphoma tumors in pre-clinical trials, and (FcγRs) and complement, but retains binding to the neonatal Fc re- CDX-301- a formulation of Flt3L - was previously shown to mobilize ceptor (FcRn) enabling use of the IgG salvage pathway to prolong its BDCA-1 and BDCA-3 myeloid DC subsets in an early phase clinical tri- circulating half-life. B7-H3 protein expression is limited in normal hu- als. These DC subsets respond to several TLR agonists and cross- man tissues, but is overexpressed in a broad range of tumor types, present antigens more effectively than plasmacytoid DCs (pDCs). including melanoma, non-small cell lung cancer (NSCLC), squamous While pDCs are high expressers of TLR9, responsive to CpG, myeloid cell carcinoma of the head and neck (SCCHN), mesothelioma, and dendritic cells express a wider array of TLRs, including high levels of urothelial cancer. By binding to B7-H3 on tumor cells and CD3 on T TLR3. lymphocytes, MGD009 can recruit cytotoxic T cells, irrespective of Methods their ability to recognize cancer cells, and trigger T cell activation, ex- This phase I/II trial tests the hypothesis that this novel in situ vaccin- pansion, and T cell-mediated killing of B7-H3-expressing tumors. The ation will induce clinical remissions at distant (untreated) tumor sites limited expression of B7-H3 on normal cells should restrict the in two cohorts of patients with either previously untreated or Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 232 of 242 relapsed/refractory iNHL (n = 15 per group). Intratumoral CDX-301 experienced both grade 3 lymphopenia and grade 3 failure to thrive. 25ug/kg is injected into a palpable lymph node for 9 days, followed At twelve weeks post-treatment initiation, 83% of patients experi- by 2Gy local radiotherapy on day 9 and 10 to the target lymph node. enced response or disease stability. Three patients (50%) had absco- To activate local DCs, poly-ICLC 2 mg is injected on day 10, 14, 17, pal responses by RECIST criteria, two patients (33%) had stable and weekly thereafter for a total of 8 treatments. disease, and one patient (17%) had progressive disease. Results Conclusions Exploratory endpoints include measuring induction of systemic Pre-clinical data suggests that sequencing may be key to the success tumor-specific immune response in pre- and post-vaccine blood and of combinatorial strategies of PD-L1 blockade and RT. The safety run- tissue samples. Using flow cytometry and CyTOF, we have confirmed in for combining PD-L1 checkpoint inhibition with RT suggests that + + that CD1c + (BDCA1 ) and CD141+ (BDCA3 ) DCs home to treated tu- the combination is safe and tolerable in metastatic NSCLC. The trial mors following treatment with Flt3L and T cells attain a mature ef- will continue to accrual to evaluate different sequencing strategies fector phenotype. Tissue from initial bx is being sequenced, and for combining RT and PD-L1 checkpoint blockade. Further study is candidate neoantigens being determined in silico; these neoantigens needed to evaluate the efficacy and optimal sequencing of RT + PD- are then being synthesized and tested for potential to activate pa- L1 checkpoint blockade. tient pre- and post-vaccination T cells. Conclusions Combinations: Immunotherapy/ This trial is in process. Trial Registration Immunotherapy ClinicalTrials.gov identifier NCT01976585. P12 Tissue factor is a novel oncotarget in triple negative breast cancer P11 and BRAF- mutated melanoma for immunotherapy using a second Preliminary safety and efficacy data for radiotherapy and PD-L1 generation ICON (L-ICON) in monotherapy and combination checkpoint blockade in metastatic non-small cell lung cancer: is therapy 1 2 2 2 timing everything? Zhiwei Hu , Rulong Shen , Amanda Campbell , Elizabeth McMichael , 1 2 2 2 2 2 2 3 Arta Monjazeb , Megan E Daly , Jonathan Riess , Tianhong Li , William J Lianbo Yu , Bhuvaneswari Ramaswam , Cheryl A London , Tian Xu , 1 2 Murphy , Karen Kelly William Carson 1 2 University of California, Davis, Sacramento, CA, USA; University of The Ohio State University Wexner Medical Center and The OSU James California Davis Comprehensive Cancer Center, Sacramento, CA, USA Comprehensive Cancer Center, Columbus, OH, USA; The Ohio State Correspondence: Arta Monjazeb (ammonjazeb@ucdavis.edu) University, Columbus, OH, USA; Yale University, New Haven, CT, USA Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P11 Correspondence: Zhiwei Hu (zhiwei.hu@osumc.edu) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P12 Background Inhibition of the PD-1/PD-L1 checkpoint pathway can induce rapid Background and durable responses in patients with non-small cell lung cancer The objective of this study is to identify tissue factor (TF) as a novel (NSCLC). Unfortunately the majority of patients fail to respond and oncotarget for triple negative breast cancer (TNBC) and BRAF mu- there is interest in exploring combinatorial strategies to improve re- tated melanoma, both of which are very difficult to treat in clinic, sponse rates. One such strategy is combining checkpoint inhibition and to develop a novel TF-targeting agent for immunotherapy. To with radiotherapy (RT). We report here our pre-clinical data for com- achieve this goal, Hu developed a second generation TF-targeting bining radiotherapy with PD-L1 checkpoint blockade. These data ICON, named L-ICON, which consists of only the light chain (1-152 demonstrate a clear influence of the sequencing of combinatorial aa) of FVII fused to an IgG1Fc. The effects of L-ICON were evaluated therapy on its efficacy. Based on these data, we have initiated a clin- as monotherapy or combination therapy with interleukin 15 (IL-15) ical trial testing sequencing strategies of radiotherapy with PD-L1 in- for the malignancies. hibition in patients with metastatic NSCLC. Methods Methods TF expression was determined by immunohistochemistry or by flow Using syngeneic mouse tumor models, we tested the synergy of cytometry. L-ICON protein (GenBank accession no. KY760097) and combining RT with PD-L1 inhibition and the influence of the sequen- replication-deficient adenoviral vectors have been developed. Bind- cing of these therapies on efficacy. Based on this preclinical work, we ing activity of L-ICON was determined. Its ADCC effect was evaluated have initiated a phase II clinical trial testing this combinatorial strat- by an ADCC effector assay and coagulation activity by FVII chromo- egy with three cohorts. The three cohorts are concurrent therapy, genic activity assay. L-ICON efficacy in monotherapy and combin- radiotherapy followed by PD-L1 checkpoint blockade, and PD-L1 ation therapy with IL-15 was tested in mouse models of murine and blockade followed by radiotherapy. We report here the preliminary human breast cancer (4 T1 and TNBC MDA-MB-231) and melanoma safety, efficacy, and correlative science data from interim analysis of (B16F10 and BRAF mutated SK-Mel-28). the safety run-in for this trial. Results Results TF is over-expressed on TNBC cells and the tumor neovasculature In studies using syngeneic mouse tumor models, we find that PD-L1 in over 85% of TNBC patients (n = 14) when using standard inhibition provides no added benefit to radiotherapy alone when ad- paraffin-embedded tumor tissues or in nearly 60% of TNBC pa- ministered after radiotherapy. Conversely, priming of the immune tients (n = 157) when employing tissue microarray slides. Import- system with anti-PD-L1 prior to RT provides significant synergy of the antly, TF expression is not detected in normal breast tissues. combinatorial therapy. Our clinical trial has completed enrollment to L-ICON has several important improvements over its first gener- the safety-run in of 6 patients. In total, 2 patients experienced grade ation ICON, including (i) more than 50% reduction in molecular 3 dose limiting toxicities meeting the criteria for completion of the mass, (ii) complete elimination of coagulation activity, (iii) stron- safety-run in without the need for dose de-escalation. One patient ger binding activity to TNBC and (iv) more effective as monother- experienced asymptomatic grade 3 lymphopenia and one patient apy in vivo in orthotopic and subcutaneous mouse models of Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 233 of 242 human TNBC (MDA-MB-231) and murine cancer 4 T1 (an animal delay melanoma progression. Due to the availability of all three com- stage IV human breast cancer) and B16F10. L-ICON monotherapy ponents for use in humans, this therapeutic regimen can potentially and combination with IL-15 were effective for the treatment of be clinically translated to expand the population of metastatic mel- SK-Mel-28 in SCID mouse models. anoma patients who experience long term benefits from immune- Conclusions based therapies. TF is a novel biomarker and oncotarget in TNBC and BRAF- mutated melanoma. L-ICON, a novel TF-targeting ICON, was effective in mono- Acknowledgements therapy and combination therapy with IL-15 for the treatment of This work was supported by CURE Grant SAP #4100072562 (Pennsylvania murine and human TNBC and melanoma in vitro and in vivo in pre- Department of Health) and NIH/NCI 5 T32 CA060395 (KMK). IL-2 was clinical mouse models. generously provided by Prometheus Laboratories Inc. Acknowledgements This work was partly supported by a startup fund from OSUMC, a Seed Award P14 from the OSUCCC TT Program, a Phase 1 L-Pilot Award from OSU CCTS via Sequentially targeting upregulated TIM-3 and CTLA-4 does not NCATS Award Number UL1TR001070 and the Dr. Ralph and Marian Falk rescue the attenuated therapeutic efficacy of combination Medical Research Trust. IL-15 was obtained from the NCI Preclinical Repository. immunotherapy with OX40 costimulation and PD-1 blockade 1 2 2 Z.H. is the inventor of L-ICON and its uses (US Patent Application # 62/082,891). David J Messenheimer , Shawn Jensen , Bernard Fox 1 2 Earle A. Chiles Research Institute, Portland, OR, USA; Providence Cancer Center, Portland, OR, USA P13 Correspondence: David J Messenheimer (messenhe@ohsu.edu) Beta-adrenergic blockade improves the immunotherapeutic Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P14 response to melanoma 1 2 1 Kathleen M Kokolus , Elizabeth A Repasky , Todd D Schell , Joseph D Background Drabick Combination immunotherapy targeting checkpoint molecules has The Pennsylvania State University College of Medicine, Hershey, PA, shown substantial results against solid tumors. However as novel USA; Roswell Park Cancer Institute, Buffalo, NY, USA therapies targeting costimulatory molecules are introduced into the Correspondence: Kathleen M Kokolus (kokolusk@hmc.psu.edu) clinic, successful combination with checkpoint blockade remains un- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P13 certain. Some strongly immunogenic preclinical models have shown benefit when combining anti-OX40 and anti-PD-1 treatment. In con- Background trast, using a PD-1 refractory mammary tumor model we have dem- Recent developments in immunotherapy have made enormous onstrated that a significant anti-tumor effect generated with OX40 strides towards expanding the scope of cancer treatment by target- costimulation is significantly attenuated with the addition of PD-1 ing a patient’s own immune cells. Despite these advances, malignant blockade. We noted high levels of inflammatory cytokines in the melanoma remains a significant clinical issue with a high proportion serum of combination treated mice, and also saw a significant in- of patients remaining unresponsive to therapy and improved, but crease in inhibitory receptors TIM-3, LAG-3, and CTLA-4 on CD4 and still low, complete response rates. The body’s response to stress is CD8 T cells in the periphery of treated mice. We hypothesized that closely integrated with the immune response, yet few cancer treat- the upregulation of these other inhibitory receptors were limiting the ment strategies account for the relationship between these biological efficacy of anti-OX40 plus anti-PD-1 combination treatment, and systems. When the stress response is activated, neurotransmitters, tested whether sequentially blocking these receptors could augment including norepinephrine, which bind β-adrenergic receptors (βARs) therapeutic efficacy. located on the surface of immune cells, are released, leading to regu- Methods lation of various immune cell functions. βAR signaling can be pre- Established orthotopically transplanted MMTV-PyMT mammary tu- vented pharmaceutically with βAR antagonists (β-blockers) and mors in FVB/NJ mice were treated with three doses of anti-OX40 and considerable literature suggests that these drugs, which are com- anti-PD-1 every other day. Serum was taken at time points and monly prescribed for other indications including hypertension, are tested for cytokine concentration and spleens were taken two days associated with positive outcomes in cancer patients. We examined after treatment to measure surface expression of inhibitory and costi- the effects of βAR blockade on the efficacy of two immunotherapies mulatory receptors. Combination treated mice were then followed by approved to treat metastatic melanoma: IL-2, which promotes T cell three doses of anti-TIM-3 with or without anti-CTLA-4. proliferation and αPD-1, which impacts T cell activation. Results Methods The addition of anti-TIM-3 and CTLA-4 provided no additional impact C57BL/6 J mice with established B16.F10 melanomas were treated to tumor growth compared to that provided by anti-OX40 and anti- with β-blockers and immunotherapy (αPD-1, IL-2 or αPD-1/IL-2) and PD-1 (n = 7/group, two independent experiments). tumor growth was monitored throughout each treatment regimen. Conclusions The accumulation of immune cells within the tumors and lymphoid These data demonstrate that TIM-3 and CTLA-4 blockade was not tissues were evaluated by flow cytometry at multiple time points fol- sufficient to augment the inhibitory effects of the concurrent com- lowing treatment. bination of anti-OX40 and anti-PD-1, and suggest that targeting in- Results hibitory receptors upregulated after initial immunotherapy treatment Blockade of βAR signaling beginning after tumors were established will require more sophisticated biomarkers and immune monitoring. had no significant impact on tumor growth. In contrast, attenuation Alternatively, we noted significant increases in some costimulatory of tumor growth by each immune-based therapy was improved in receptors (ICOS, 4-1BB) and targeting these receptors may provide the presence of β-blockers. We observed significantly extended sur- an alternative to blocking inhibitory pathways. Additionally there vival in mice treated with αPD-1 or αPD-1/IL-2 combined with β- may be a tipping point where providing additional antibodies target- blockers compared to immunotherapy only mice. Most importantly, ing costimulatory or checkpoint molecules may be ineffective. T cells the combination of β-blockers, αPD-1 and IL-2 produced a highly sig- may become so heavily deregulated or over-stimulated that they be- nificant delay in tumor growth and prolonged survival compared to come permanently exhausted. In support of this we have previously αPD-1/IL-2 without β-blockers. shown that anti-OX40 combined sequentially with anti-PD-1 provides Conclusions superior therapeutic effect compared to concurrent combination, Blocking βAR signaling improved the efficacy of at least two types of with complete tumor regression in ~30% of animals treated. Sequen- immunotherapy, but was most effective when administered with tially treated animals generated T cells capable of significantly more dual-immunotherapy. We suggest that each therapeutic component long-term proliferation, suggesting that timing is critical when com- may improve a unique aspect of the immune response to maximally bining immunotherapies. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 234 of 242 and activation, and in a subcutaneous Lewis lung carcinoma model for their ability to mediate anti-tumor immunity, both alone and in combination with anti-PD-1 mAb. Results In normal mice, IL-2/mAb complexes potently expanded CD8+ T cells and NK cells with minimal expansion of Tregs. As single agent ther- apy, IL-2/mAb complexes or anti-PD-1 mAb reduced tumor growth, although most mice succumb to tumor growth eventually. Combin- ation of IL-2/mAb complexes with anti-PD-1 mAb therapy resulted in durable, complete responses in nearly half of the mice. Conclusions While immune based therapies such as anti-PD-1 mAb can be highly effective in select patients, even in those patients that obtain clinical benefit, disease may recur. Our results suggest that the addition of IL-2/mAb complexes to therapy with anti-PD-1 mAb could broadly in- crease the percentage of patients deriving benefit from immune- based therapy. References 1. Sato, et al: Biotherapy 1993, 6(3):225–231. 2. Boyman, et al: Science 2006, 311:1924–1927. Fig. 4 (abstract P14). The addition of anti-TIM + anti-CTLA-4 provides 3. Létourneau, et al: PNAS 2010, 107:2171–2176. no benefit to anti-OX40 + anti-PD-1 combination treatment P16 The combination of an IL-15/IL-15Ralpha complex (ALT-803) and anti-PD-1 mAb leads to superior anti-tumor immunity in a murine lung tumor model P15 1 1 1 John Wrangle , Kristina Andrijauskaite , Marzena Swiderska-syn , Peter Novel IL-2/mAb complexes mediate potent anti-tumor immunity 2 2 1 Rhode , Hing Wong , Mark Rubinstein which is augmented with anti-PD-1 mAb therapy 1 2 1 1 1 Medical University of South Carolina, Charleston, SC, USA; Altor Mark Rubinstein , Kristina Andrijauskaite , Marzena Swiderska-syn , Kristin 2 2 2 BioScience Corporation, Miramar, FL, USA Lind , Agnes Choppin , Marina K Roell 1 2 Correspondence: Mark Rubinstein (markrubinstein@musc.edu) Medical University of South Carolina, Charleston, SC, USA; XOMA Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P16 Corporation, Berkeley, CA, USA Correspondence: Mark Rubinstein (rubinsmp@musc.edu) Background Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P15 Administration of antibodies that block the PD-1/PD-L1 pathway has demonstrated unprecedented success in mediating clinical re- Background sponses in patients with advanced cancer. These antibodies are Recent success and FDA approval of immune checkpoint inhibitors thought to act by blocking the ability of PD-L1 to mediate an in- (CI) in a growing number of cancers are transforming cancer treat- hibitory signal to PD-1 expressing T cells during antigen- ment and revitalizing interest in immunotherapies. However, while recognition. These antibodies are now FDA-approved for multiple efficacy is observed in patients with advanced metastatic diseases cancers including non-small cell lung cancer (NSCLC) in patients treated with CI, not all patients respond and most responses are in- with disease that has progressed during or after platinum-based complete. Preclinical studies suggest that combinations of additional chemotherapy. In these patients, one in five patients can attain a modalities will provide opportunities to improve patient responses. clinical response while on checkpoint inhibitor therapy. While As both IL-2 and CI therapy can independently augment anti-tumor promising, this therapy fails to induce a durable clinical response immunity in patients, likely in mechanistically distinct ways, we hy- in most patients. To overcome this limitation, we hypothesized pothesized we could improve anti-tumor immunity by combining IL- that combinatorial therapy with anti-PD-1 mAb and a lymphocyte 2 and anti-PD-1 mAb therapy. growth factor would more effectively augment the expansion of Methods tumor-reactive lymphocytes. This would also provide a means to To improve IL-2 efficacy and therapeutic index, we generated novel not only remove inhibitory pathways but directly augment the anti-IL-2 mAbs which, when complexed with IL-2 (IL-2/mAb) offer ad- function of tumor-reactive lymphocytes. We chose to use an IL- vantages over standard IL-2 therapy [1-3]. First, binding to an anti-IL- 15/IL-15Ra complex (ALT-803) composed of an IL-15 mutant 2 mAb increases IL-2 half-life and biological activity. Second, depend- (N72D) that was pre-associated with the soluble IL-15Ra/Fc fusion ing on the epitope at which the mAb binds to IL-2, antibody binding protein. This superagonist complex has been shown to potently can modulate which IL-2 receptor subunits (alpha, beta, or gamma) expand and activate CD8 T cells and NK cells in various animal are engaged. Antibodies that interfere with binding of IL-2Rα can re- models. duce activation of high- IL-2Rα-expressing cell types, such as sup- pressive Tregs, and steer activity toward cell types expressing only IL- Methods 2Rβ and γ. In this way, these complexes may have more effective To assess the efficacy of combination therapy, we injected C57BL/6 anti-tumor activity [1-3]. We screened antibody phage libraries to mice subcutaneously with Lewis lung carcinoma. Mice with estab- identify antibodies that shift IL-2 receptor binding and activity differ- lished tumors were treated with anti-PD-1 mAb and/or IL-15/IL-15Ra entially on different cell types in vitro and in vivo. Complexes of these complex, and we monitored tumor progression and changes in im- antibodies were tested in vivo for their effects on T cell frequency mune cell populations in the periphery and tumor. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 235 of 242 Results findings provide a strategy for the selective targeting of Tregs in the The combination of anti-PD-1 mAb and the IL-15/IL-15Ra complex frame of cancer combination immunotherapy. was substantially more effective at inducing complete responses Conclusions compared with administration of either agent alone. Effective ther- These findings provide a new insight into CD4 T cell biology and apy was associated with the expansion of CD8 T cells and NK cells, offer a new strategy for selective targeting of Tregs in the frame of and the acquisition of the ability of CD8 T cells to produce IFNγ development of anti-cancer immunotherapies. after activation. Interestingly, in vitro, IFNγ led to upregulation of both MHC and PD-L1 on tumor cells, suggesting a mechanistic basis for the improved efficacy of the combination therapy. Combinations: Immunotherapy/ Conclusions Standard of Care Our results suggest that the efficacy of anti-PD-1 mAb therapy may be improved by co-administration of the IL-15/IL-15Ra complex. Our P18 results also suggest a mechanistic basis for why the combination Pembrolizumab in combination with chemoradiotherapy (CRT) for may be superior to single agent therapy. To determine if this com- locally-advanced squamous cell carcinoma of the head and neck bination would be of value in human patients, we have initiated a (SCCHN): Interim safety analysis (ISA) phase Ib/II clinical trial (NCT02523469) to assess the combination of 1 2 1 2 Steven Powell , Mark Gitau , Christopher Sumey , Andrew Terrell , anti-PD-1 mAb (nivoluamb) in combination with ALT-803 in patients 1 1 3 2 Michele Lohr , Ryan K Nowak , Steven McGraw , Ash Jensen , Miran with refractory advanced NSCLC. 2 4 4 1 Blanchard , Kathryn A Gold , Ezra EW Cohen , Christie Ellison , Lora 1 5 1 Black , John Lee , William Chad Spanos 1 2 Sanford Cancer Center, Sioux Falls, SD, USA; Sanford Roger Maris P17 Cancer Center, Fargo, ND, USA Sanford Health, Sioux Falls, SD, USA; Functional dichotomy of PI3K isoforms in CD4 T cells provides a Moores Cancer Center, University of California, San Diego, La Jolla, CA, strategy for selectively targeting regulatory T cells to enhance USA; NantKwest, Inc., Culver City, CA, USA anti-tumor immunotherapy 1 1 1 1 Correspondence: Steven Powell (steven.powell@sanfordhealth.org) Shamim Ahmad , Mason Webb , Rasha Abu-Eid , Rajeev Shrimali , Vivek 1 1 1 1 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P18 Verma , Atbin Doroodchi , Zuzana Berrong , David Yashar , Raed 2 1 1 Samara , Mikayel Mkrtichyan , Samir Khleif 1 2 Background Georgia Cancer Center, Augusta, GA, USA; Qiagen, Frederick, MD, USA Blockade of the programmed death receptor 1 (PD-1) interaction Correspondence: Samir Khleif (skhleif@augusta.edu) with its ligands (PD-L1, PD-L2) represents an active therapeutic Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P17 approach in recurrent and metastatic SCCHN [1]. This immune checkpoint may allow immune escape during standard treatment, Background including CRT [2]. Standard CRT in SCCHN has substantial toxicity The PI3K-Akt signaling pathway modulates diverse biological re- and the safety of adding PD-1 inhibition is unknown. We report sponses including signaling, proliferation and survival of T cells. The the first ISA of weekly cisplatin-based CRT in combination with identification of a signaling pathway, which differentially regulates pembrolizumab, a humanized IgG4 monoclonal antibody inhibitor regulatory T cells (Tregs) and conventional T cells (Tconvs), is crucial of the PD-1:PD-L1/2 interaction. for selectively modulating these two subsets. The differential role of Methods class IA PI3K isoform in regulating the survival and apoptosis of Tregs This is an open-label phase IB trial of pembrolizumab used in and Tconvs has not been elucidated yet. combination with cisplatin-based, definitive CRT in patents with Methods stage III-IVB SCCHN. The treatment schema is outlined in Fig. 5. For in vitro experiments sorted Tregs and Tconvs were labeled with TM Key inclusion and exclusion criteria are in Fig. 6. The primary CellTrace Violet Cell Proliferation stain (VCT) according to the man- endpoint of safety is assessed by dose-limiting grade ≥3adverse ufacturer’s protocol (Life Technologies, NY). Cells were stimulated events (AEs) and immune-related AEs (irAEs) per NCI-CTCAE v4.0 with and without inhibitors. For in vivo experiments C57BL/6 Mice criteria. Efficacy is measured by RECIST v1.1 complete response were injected subcutaneously (s.c.) with TC-1 tumor cells and moni- (CR) rate on 100-day post-CRT imaging and/or pathologic CR for tored for development of tumors. Vaccine was given weekly s.c. For those who undergo salvage surgery. Planned enrollment is 39 pa- therapeutic experiments vaccine was given weekly throughout the tients based on a hypothesized CR rate of >60% and a two-stage experiment. CAL-101 treatment was provided on the day when Simon minimax design (type I error rate of 0.05 and power of tumor size reached 3-4 mm 5-6 day before vaccination. 0.80). Secondary endpoints will include overall response, survival, Results and quality-of-life assessments. Here, we report that PI3Kd isoform is sufficient for TCR downstream Results signaling, proliferation, and survival for either Tconvs or Tregs. In At the time of ISA, 27 patients have been enrolled. Patient and Tregs, however, PI3Kd is a dominant isoform, where Tregs are fully disease characteristics are in Fig. 7. At data cut-off on 9/14/2016, dependent on PI3Kd to regulate these properties as PI3Kα and PI3Kb 22 patients have completed CRT. Of those patients, 3 required do not play any role in these biologic processes. On the other hand, cisplatin dose reductions and 6 required a dose discontinuation. in Tconvs, the two other isoforms, PI3Kα and PI3Kb combined, pro- Acute grade (G) ≥ 3 AEs for those completing CRT are in Fig. 8. vide redundant pathway to PI3Kd in the regulation of TCR signaling, Median cumulative cisplatindoseis225 mg/m2. Therewereno proliferation and survival. This redundant role provided by PI3Kα and radiation dose-limiting toxicities. Two patients (9%) discontinued PI3Kβ isoforms to PI3Kd in Tconvs offers a selective therapeutic ap- pembrolizumab due to irAEs (G2 peripheral motor neuropathy proach to inhibit Tregs, where by inhibiting PI3Kd, signaling, prolifer- and G3 transaminase elevation). No deaths have occurred. ation, and survival are inhibited in Tregs, while PI3Kα and PI3Kβ, will provide a path for Tconvs to proliferate and function. Conclusions Importantly, we demonstrate that our findings translate to thera- This represents one of the first studies to evaluate the safety of con- peutic efficacy in vivo, where the inhibition of PI3Kδ, enhanced anti- current CRT and PD-1 inhibition in SCCHN. Acute CRT-related toxic- tumor efficacy of antigen-specific vaccine by decreasing the suppres- ities are comparable to other SCCHN CRT studies. No new sive Tregs and increasing the number of vaccine-induced CD8 T cells, immunologic safety signals were seen. Further investigation of this thus showing synergistic therapeutic effect against tumors. Our approach is warranted. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 236 of 242 Acknowledgements Funding supported by the Merck Investigator Studies Program. Trial Registration ClinicalTrials.gov identifier: NCT02586207 References 1. Seiwert TY, et al: Safety and clinical activity of pembrolizumab for treatment of recurrent or metastatic squamous cell carcinoma of the head and neck (KEYNOTE-012): an open-label, multicentre, phase 1b trial. Lancet Oncol 2016, 17(7):956–965. 2. Parikh F, et al: Chemoradiotherapy-induced upregulation of PD-1 an- tagonizes immunity to HPV-related oropharyngeal cancer. Cancer Res 2014, 74(24):7205–7216. Fig. 8 (abstract P18). Acute Grade ≥3 Adverse Events with >1 Occurrence Diet, Exercise and/or Stress and Impact on the Immune System P19 Exercise training reduces splenic accumulation of MDSCs and delays tumor progression in a therapeutic breast cancer model 1 2 1 3 Erik Wennerberg , Emily Schwitzer , Claire Lhuillier , Graeme Koelwyn , 2 2 4 Rebecca Hiner , Lee Jones , Sandra Demaria 1 2 Weill Cornell Medicine, New York, NY, USA; Memorial Sloan Kettering Fig. 5 (abstract P18). Clinical Trial Schema Cancer Center, New York, NY, USA; New York University School of Medicine, New York, NY, USA; Department of Radiation Oncology, Weill Cornell Medicine, New York, NY, USA Correspondence: Erik Wennerberg (erw2010@med.cornell.edu) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P19 Background Epidemiological studies show a correlation between physical ac- tivity and cancer-related mortality [1]. However, the contribution of immune mediated anti-tumor immunity to the beneficial ef- fects of exercise has yet to be defined [2]. We sought to investi- gate if forced running would have a therapeutic benefit in mice bearing a poorly immunogenic breast cancer and investigate the Fig. 6 (abstract P18). Key Inclusion and Exclusion Criteria immunological changes occurring in response to exercise. Methods On day 0 Balb/c mice were inoculated with 4 T1 breast cancer cells subcutaneously in the right flank (n = 6/group). Starting on day 7, once tumors were palpable, mice were subjected to 30 minutes of forced treadmill running (18 cm/sec) five days per week. Control mice remained sedentary throughout the study. Analysis of immune cells in spleen and tumor was performed at day 17 and 32 and spon- taneous lung metastases were evaluated at day 32. Results We observed a significantly delayed primary tumor growth (tumor volume on day 31: 1167 ± 174 mm in sedentary versus 847 ± 124 mm in exercised mice, p < 0.01) and a tendency for re- duced metastatic burden in the lungs of exercised compared to sedentary mice. The progressive marked increase in myeloid- derived suppressor cells (MDSCs) and splenomegaly seen in sed- entary 4 T1 tumor-bearing mice was less pronounced in exercised mice. This difference was significant on day 17; with spleen weight (520 ± 110 mg in sedentary versus 330 ± 30 mg in exer- cised mice, p < 0.01) and MDSC frequency in spleen leukocytes (22.7 ± 2.6% in sedentary versus 14.3 ± 2.7% in exercised mice, p < 0.001) were significantly lower in exercised mice compared to sedentary mice. Furthermore, on day 32, the CD8+ T cell/Treg and CD8+ T cell/MDSC ratio showed a tendency to increase in tumors from exercised mice. Conclusions Our data demonstrate that exercise can slow tumor progression in a therapeutic setting. While the mechanisms of this effect re- Fig. 7 (abstract P18). Patient and Disease Characteristics quire further investigation, the observed decrease in the Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 237 of 242 proportion of immunosuppressive immune cells in spleen and Conclusions tumor of exercised mice is likely to play a role. Importantly, the These data support the possibility that NHS-muIL12 abrogates an ability of exercise to reduce immunosuppression locally and sys- immune-suppressive response within the TME, which might permit T temically supports testing exercise in combination with immuno- cells to execute their antitumor effects. NHS-huIL12 (MSB0010360N; therapy as a therapeutic modality that can increase responses M9241), is currently being evaluated against solid tumors in a phase without increasing toxicity. I clinical trial (NCT01417546). References Acknowledgements 1. Ballard-Barbash R, Friedenreich CM, Courneya KS, Siddiqi SM, McTiernan We acknowledge the kind contribution of NHS-muIL12 from EMD Serono, A, Alfano CM: Physical activity, biomarkers, and disease outcomes in Billerica, MA. cancer survivors: a systematic review. Natl Cancer Inst 2012, 104:815– 2. Koelwyn GJ, Wennerberg E, Demaria S, Jones LW: Exercise in Regulation Therapeutic Cancer Vaccines of Inflammation-Immune Axis Function in Cancer Initiation and Pro- gression. Oncology 2015, 29(12). P21 Intracellular trafficking of self-assembled immune signals Michelle Bookstaver, Christopher M Jewell Not Listed – Other Fischell Department of Bioengineering, University of Maryland - College P20 Park, College Park, MD, USA Systemic immunotherapeutic efficacy of an immunocytokine, NHS- Correspondence: Michelle Bookstaver (mlbooks@umd.edu) muIL12, in a superficial murine orthotopic bladder cancer model Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P21 Vandeveer Amanda, John W Greiner, Jeffrey Schlom Laboratory of Tumor Immunology and Biology, Center for Cancer Background Research, National Cancer Institute, Bethesda, MD, USA We recently exploited electrostatic interaction to design self- Correspondence: Vandeveer Amanda (amanda.vandeveer@nih.gov) assembling nanostructures comprised entirely from peptide anti- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P20 gens and toll-like receptor (TLR) agonists as adjuvants. These ma- terials simplify vaccine composition and exhibit unique properties Background such as direct control over the absolute and relative concentra- Interleukin-12 is one of the most powerful proinflammatory cytokines tions of each component and co-delivery of the signals to anti- capable of supporting T and NK cell function, inducing IFNγ while gen presenting cells. In pre-clinical models of melanoma, this driving a T 1 adaptive immune response. Its success as an antitumor approach leads to significantly enhanced anti-tumor immunity. agent in preclinical models has yet to be realized in a clinical setting Here, we study how the physicochemical features (e.g., peptide due to systemic toxicity. An IL-12 delivery system has been devel- charge) and relative concentration of each component impact the oped to maximize deposition of the cytokine directly in the tumor internalization, trafficking, and processing of the immune signals microenvironment (TME), while mitigating the dose-limiting systemic in antigen presenting cells. effects. Methods Methods FITC-labeled SIINFEKL peptide was modified with three or nine argi- NHS-IL12 is a novel immunocytokine, consisting of two molecules of nines, for use as a cationic anchor to support self-assembly with a human or murine IL-12 fused to a tumor necrosis-targeting human polyanionic nucleic acid-based TLR3 agonist, polyIC. Hollow capsules IgG1 (NHS76). NHS76 recognizes exposed chromatin-DNA found in built from these signals were synthesized by coating a sacrificial necrotic human/murine tumors. Previous studies have shown select- CaCO core with alternating layers of modified SIINFEKL and PolyIC. ive tumor uptake of NHS-IL12 in necrotic subcutaneous murine tu- After deposition, the core was removed using EDTA and capsules mors. Urothelial bladder cancer is known to respond favorably to were washed with buffer, resulting in stable capsules formed from immunotherapeutic agents due to many somatic mutations and TILs, immune signals. Capsule size was determined by image analysis and and response to Bacillus Calmette-Guerin (BCG). component loading levels were determined by fluorimetry using Results FITC-labeled peptide and Cy5-labeled TLRa. Stability studies were car- We evaluated the use of NHS-muIL12 in a murine orthotopic ried out by incubating capsules in media as a function of different Luc bladder cancer model. MB49 cells, instilled into the bladder pH and ionic strengths. For uptake and trafficking studies, murine form superficial, multifocal tumors which can be monitored with splenocytes were isolated and treated with different concentrations a luciferase-based intravital imaging system. NHS-muIL12 is a very of capsules. Cells were analyzed by flow cytometry and imaging in potent anti-tumor agent in both MB49 tumor models, reducing the presence or absence of inhibitors of endocytic processes and tumor volume in a dose-dependent manner. In the intravesical during staining with markers for surface proteins and intracellular bladder model, antitumor effects were seen at 2.5 mg/kg admin- organelles. istrated as three separate systemic injections. Mice were cured of Results tumor when treated at 20 mg/kgx3 NHS-muIL12 with durable Capsules loaded with FITC-SIINFEKL-R and PolyIC were 1-2 μmin tumor-free long-term survival. Immune analyses revealed TAA- diameter and exhibited similar size and shape for 2 weeks when in specific CTLs and IFN-γ responses, indicating the development of buffer. These materials exhibited tunable loading with a composition a specific anti-tumor immune response. An immune memory re- of 15.5% peptide and 84.5% TLRa used for trafficking studies. FITC- sponse protected mice following re-challenge with MB49 tumor SIINFEKL-R and PolyIC capsules were efficiently internalized through cells. Anti-tumor efficacy required CD4+ or CD8+ T cells as deple- energy dependent processes (i.e., endocytosis) when incubated with tion of either abrogated the anti-tumor effects. Evaluation of TILs primary dendritic cells within 1 hour of treatment. These effects were by FACS, revealed that NHS-muIL12 significantly reduced the also found to be dose-dependent and did not impact viability of number of immune suppressive cells such as MDSCs, 24 hours treated cells. post-treatment, which continued to the end of the study. Im- Conclusions munofluorescence showed correlative treatment-related modula- Initial studies reveal capsules comprised of FITC-SIIN-R and tion of CD4+ and CD8+ T cells as well as MDSCs and Tregs PolyIC are uptaken by primary immune cells quickly and effect- within the TME. Gene expression of RNA from bladder tumors, ively. Ongoing studies will assess the uptake of capsules by identified various immune components with immunosuppressive endocytosis in the presence of inhibitors to decipher the endocy- or immune potentiating roles, modulated by NHS-muIL12 tic pathway and trafficking of capsules through lysosomes and treatment. endosomes. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 238 of 242 Acknowledgements whether antibody and T cell responses were directed to mutations, This work was supported in part by NSF CAREER # 1351688 and Alliance for altered peptide ligands or overexpressed normal proteins. Cancer Gene Therapy # 15051543. Results Compared to vaccination alone or vaccination with GM-CSF, vaccin- ation with DPV-001 plus imiquimod significantly (p < 0.05) increased P22 the number of antibody responses that were four-fold higher at Analysis of B and T cell responses in non-small cell lung cancer week twelve. In the one patient where autologous tumor was avail- (NSCLC) patients enrolled in a phase II trial of cyclophosphamide able, vaccination increased the tumor-specific release of TNF-alpha with allogenic DRibble vaccine (DPV-001) by peripheral blood CD4 T cells. 1 1 2 3 Christopher Paustian , Andrew Gunderson , Brian Boulmay , Rui Li , Conclusions 4 4 5 6 Bradley Spieler , Kyle Happel , Tarsem Moudgil , Zipei Feng , Carmen Based on these studies, future trials will combine the adjuvant imi- 3 6 7 Ballesteros-Merino , Christopher Dubay , Brenda Fisher , Yoshinobu quimod with DRibble vaccine. Trial Registration 8 1 4 3 Koguchi , Sandra Aung , Eileen Mederos , Carlo B. Bifulco , Michael ClinicalTrials.gov identifier: NCT01909752 9 9 9 4 McNamara , Keith Bahjat , William Redmond , Augusto Ochoa , Hong- 10 11 11 6 Ming Hu , Adi Mehta , Fridtjof Lund-Johansen , Bernard Fox , Walter 8 8 1 Urba , Rachel E. Sanborn , Traci Hilton P23 1 2 UbiVac, Portland, OR, USA; Section of Hematology/Oncology, Louisiana An open-label phase I/IIa escalating dose study to evaluate safety State University, New Orleans, LA, USA; Robert W. Franz Cancer and T cell immunogenicity of PDS0101 in subjects with cervical Research Center, Earle A. Chiles Research Institute, Providence Cancer intraepithelial neoplasia (CIN) and high-risk HPV infection 4 1 1 1 1 Center, Portland, OR, USA; Louisiana State University Stanley S. Scott Frank Bedu-Addo , Greg Conn , Michael King , Panna Dutta , Robert 5 2 3 Cancer Center, New Orleans, LA, USA; PPMC, Portland, OR, USA; Shepard , Mark Einstein 6 7 1 2 Providence Cancer Center, Portland, OR, USA; Providence Medical PDS Biotech, New Brunswick, NJ, USA; PDS Biotech, Miami Beach, FL, 8 3 Center, Portland, OR, USA; Earle A. Chiles Research Institute, Providence USA; Rutgers, NJ Medical School, Newark, NJ, USA Cancer Center, Portland, OR, USA; Providence Medical Center, Portland, Correspondence: Robert Shepard (RShepard@Post.Harvard.edu) 10 11 OR, USA; UbiVac, Providence Medical Center, Portland, OR, USA; Oslo Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P23 University Hospital, Oslo, Norway Correspondence: Christopher Paustian Background (christopher.paustian@ubivac.com) Current HPV vaccines are effective at preventing infection. However, Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P22 there are no therapeutic vaccines to treat the infection or commonly associated diseases e.g. CIN, cervical, anal and oral cancers. A therapy Background that is simple, effective and safe enough to be administered to CIN DRibble vaccines are microvesicles derived from proteasome-blocked and early-stage cancer patients could be important in achieving the autophagosomes. The DPV-001 DRibble vaccine is derived from an goal of effective cancer prevention and treatment of pre-metastatic adenocarcinoma and a mixed histology cancer cell line. By mass cancer. We assessed whether PDS0101, a combination of modified spectroscopy they contain more than 130 potential NSCLC antigens, multi-epitope HPV16 peptides (HPVmix) and escalating doses of the many as prospective altered-peptide ligands, which could intensify synthetic Versamune® T cell activating platform could facilitate anti- their immunogenicity. In preclinical models, DRibble immunotherapy gen cross-presentation and safe immune activation leading to strong provided significant cross-protection against 8 of 9 tumors tested. HPV-specific CD8+ T cell induction in CIN. Additionally, Dribble vaccines are effective in treating established tu- Methods mors in preclinical combination immunotherapy models. We Safety and immunogenicity were assessed in an open label dose es- hypothesize that the efficacy of DRibbles’ vaccination can be attrib- calation study. Groups of 3-6 subjects received either low dose uted to their capacity to present tumor-derived short-lived proteins (1 mg), medium dose (3 mg) or high dose (10 mg) of Versamune® (SLiPs) and defective ribosomal products (DRiPs) that are typically cationic lipid with 2.4 mg of HPVmix. Each subject received one SC not processed and presented by professional antigen presenting dose every 3 weeks for a total of 3 doses. T cell response was evalu- cells. These SLiPs and DRiPs embody a prospective pool of tumor an- ated by IFN-γ and granzyme-b ELISpot using blood drawn from the tigens against which the host may be less tolerant. subjects pre-vaccination, 2 weeks after each vaccination and 90 days Methods after vaccination 3. The trial is registered at ClinicalTrials.gov (number Thirteen definitively-treated stage III NSCLC patients were vaccinated NCT02065973). at 3-week intervals. Patients were randomized such that some pa- Results tients’ intradermal vaccines were combined with administration of No serious adverse events were reported. No IND safety reports were imiquimod or GM-CSF as an adjuvant. PBMCs and serum were col- submitted. No subjects withdrew. Strong HPV-specific T cell re- lected at baseline and at each vaccination. For one patient, PBMCs sponses occurred at all 3 doses, even in those subjects with low pre- from the baseline visit and week 12 were tested against that patient’s vaccination T cell responses. PDS0101 vaccination led to strong T cell autologous tumor cell line to measure increased tumor specific T cell responses evaluated by both IFN-γ and granzyme-b ELISpot. Conclu- activation. Studies are currently underway to evaluate changes in sions: PDS0101 is safe and effectively performs antigen cross- TCR repertoires. CD4+ and CD8+ T cells from multiple time points presentation as demonstrated by HPV-specific T cell responses, in- were sorted and TCR sequencing is being performed to look at alter- cluding inducing active cytolytic T cells. Clinical benefit in CIN2/3 and ations in the T cell repertoire. The primary outcome measure of this cancer will be evaluated in larger phase II trials. clinical trial was to discover if vaccine alone, vaccine plus imiquimod, Conclusions or vaccine plus GM-CSF generated the greatest number of strong PDS0101 is safe and effectively performs antigen cross-presentation antibody response. as demonstrated by HPV-specific T cell responses, including inducing Serum from the baseline visit and week 12 was analyzed for in- active cytolytic T cells. Clinical benefit in CIN2/3 and cancer will be creased antibody response to >9000 human proteins using ProtoAr- evaluated in larger phase II trials. rays and Microsphere Affinity Proteomics. Where sufficient tumor Trial Registration was available, whole exome sequencing was done to evaluate ClinicalTrials.gov identifier: NCT02065973 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 239 of 242 Tumor Microenvironment Trial Registration ClinicalTrials.gov identifier: NCT01421017 P24 Effects of TLR7 agonist imiquimod combined with local References radiotherapy on the tumor microenvironment in women with 1. Panelli: Genome Biol 2008. metastatic breast cancer in a prospective trial 2. Dewan: CCR 2012. 1 2 3 1 1 Sylvia Adams , Ena Wang , Ping Jin , Yelena Novik , Debra Morrison , 1 2 4 1 Ruth Oratz , Franco M Marincola , David Stroncek , Judith Goldberg , 5 5 Sandra Demaria , Silvia C Formenti P25 Perlmutter Cancer Center, New York University School of Medicine, Immunoscore as a prognostic marker in stage I-III colon cancer: NYC, NY, USA; Sidra Medical and Research Center, Doha, Qatar; results of a SITC-led global validation study 3 1 1 1 2 National Institutes of Health Clinical Center Department of Transfusion Jérôme Galon , Bernhard Mlecnik , Florence Marliot , Fang-Shu Ou , 4 3 4 4 4 Medicine, Bethesda, MD, USA; National Institutes of Health Clinical Carlo B Bifulco , Alessandro Lugli , Inti Zlobec , Tilman T Rau , Iris D 5 5 5 6 6 Center, Bethesda, MD, USA; Weill Cornell Medicine, Department of Nagtegaal , Elisa Vink-Borger , Arndt Hartmann , Carol Geppert , Michael 7 7 7 7 Radiation Oncology, New York, NY, USA H. Roehrl , Prashant Bavi , Pamela S Ohashi , Julia Y Wang , Linh T 7 7 7 7 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P24 Nguyen , SeongJun Han , Heather L MacGregor , Sara Hafezi-Bakhtiari , 7 8 8 8 Bradley G Wouters , Yutaka Kawakami , Boryana Papivanova , Mingli Xu , 8 9 9 9 Correspondence: Sylvia Adams (sylvia.adams@nyumc.org) Tomonobu Fujita , Shoichi Hazama , Nobuaki Suzuki , Hiroaki Nagano , 10 11 12 12 Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P24 Kiyotaka Okuno , Kyogo Itoh , Eva Zavadova , Michal Vocka , Jan 12 12 12 12 Spacek , Lubos Petruzelka , Bohuslav Konopasek , Pavel Dundr , 12 13 13 Background Helena Skalova , Toshihiko Torigoe , Noriyuki Sato , Tomohisa 13 13 14 Application of TLR7 activator imiquimod (IMQ) onto BCC of the skin Furuhata , Ichiro Takemasa , Marc Van den Eynde , Anne Jouret- 14 14 1 1 leads to an early tumoral transcriptional profile of immunological re- Mourin , Jean-Pascal Machiels , Tessa Fredriksen , Lucie Lafontaine , 1 1 1 1 jection (ICR) preceding complete remission as shown in a random- Bénédicte Buttard , Sarah Church , Pauline Maby , Helen Angell , 1 1 1 1 ized trial [1]. Here we employed the same methodology evaluating Mihaela Angelova , Angela Vasaturo , Gabriela Bindea , Anne Berger , 1 15 15 15 serial FNA tumor biopsies from breast cancer patients treated on a Christine Lagorce , Prabhu S Patel , Hemangini H Vora , Birva Shah , 15 15 15 clinical trial of the combination of IMQ and radiotherapy (RT), to de- Jayendrakumar B Patel , Kruti N Rajvik , Shashank J Pandya , Shilin N 15 16 16 17 lineate dynamic changes associated with ICR in breast cancer and to Shukla , Yili Wang , Guanjun Zhang , Giuseppe V Masucci , Emilia K 17 18 18 19 understand the contribution of each treatment modality to antitumor Andersson , Fabio Grizzi , Luigi Laghi Gerardo Botti , Fabiana 19 19 19 19 immunity in vivo. We previously demonstrated synergy of IMQ/RT in Tatangelo , Paolo Delrio , Gennaro Cilberto , Paolo A Ascierto , 20 2 3 1 the poorly immunogenic TSA model with enhanced T cell-mediated Franco Marincola , Daniel J Sargent , Bernard A Fox , Franck Pages inhibition of treated and untreated tumors [2]. INSERM, Université Pierre et Marie Curie, Université Paris Descartes, 2 3 Methods Paris, France; Mayo Clinic, Rochester, MN, USA; Earle A. Chiles Research Clinical trial (NCT01421017): for patients with metastatic breast can- Institute, Providence Cancer Center, Portland, Oregon, USA; Institute of cer to the skin. Treatment: topical IMQ to one metastasis, IMQ and Pathology, University of Bern, Bern, Switzerland; Radboud University RT to another metastasis. IMQ self-applied 5xX/per week x 8 weeks, Nijmegen Medical Center, Nijmegen, Netherlands; University Erlangen- RT started with first IMQ (6Gyx5 over 10 days). Cyclophosphamide Nürnberg, Erlangen, Germany; Princess Margaret Cancer Centre, (200 mg/m2 IV) was administered a week before in a subset of pa- University Health Network, Toronto, ON, Canada; Division of Cellular tients. An untreated, measurable lesion (skin or distant metastases) Signaling, Institute for Advanced Medical Research, Keio University outside the IMQ and radiation fields was observed as systemic re- School of Medicine, Tokyo, Japan; Department of Gastroenterological, sponse read-out per RECIST1.1. Local response defined as PR or CR in Breast and Endocrine Surgery, Yamaguchi University Graduate School of treated lesions. FNA of IMQ and IMQ/RT treated metastases: at base- Medicine, Ube, Japan; Department of Surgery, Kinki University Faculty line, 2 and 3 weeks, RNA isolation/amplification performed per SOPs. of Medicine, Osaka-Sayama, Japan; Division of Clinical Research, Gene expression: Affymetrix Human GeneArray 1.0 ST/Partek Genom- Research Center for Innovative Cancer Therapy, Kurume University ics suite software with special emphasis on expected immune School of Medicine, Kurume, Japan; First Faculty of Medicine, Charles signature. University and General University Hospital, Prague, Czech Republic; 13 14 Results Sapporo Medical University, Sapporo, Japan; Institut Roi Albert II, Serial FNA samples are available from 18 patients. Gene expression Cliniques universitaires St-Luc, Université Catholique de Louvain, profiles of baseline biopsy treated with IMQ/RT identified 2309 differ- Brussels, Belgium; The Gujarat Cancer & Research Institute, entially expressed genes (p < 0.005) between CR and PR. Among Ahmedabad, India; Institute for Cancer Research, Xi’an Jiaotong them, ICR genes such as GZMB, GZMH, PRF1, GNLY, CD8A and TBX21 University, Xi’an, China; Karolinska Institutet, Karolinska University, are over expressed in CR. Significant altered gene expression was ob- Stockholm, Sweden; Humanitas Clinical and Research Center, Rozzano, 19 20 served in progressing lesions (week 3 vs baseline, 1854 genes) in Italy; Istituto Nazionale Tumori Fondazione Pascale, Naples, Italy; Sidra contrast to responding metastases (PR: 53 genes, CR: 23 genes) post Medical and Research Center, Doha, Qatar IMQ/RT suggesting active wound healing and tumor progression sig- Correspondence: Bernard A Fox (bernard.fox@providence.org) nature. For the IMQ alone treated metastases, differential gene ex- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P25 pression was observed at baseline distinguishing subsequent PR and PD (n = 189, p < 0.005). Systemic response was observed as a marked Background baseline gene expression difference (n = 1177, p < 0.005) predicting Increasing evidence has illustrated that enhanced lymphocytic infil- abscopal phenomena (CR, PR, SD and PD). tration is a powerful prognostic marker in colon cancer (CC). The Conclusions Immunoscore (IM) methodology was developed as a standardized The ICR signature in tumors before IMQ-RT treatment is positively assay to quantify the in situ immune cell infiltrate. correlated with complete local response, which validates the ICR hy- Methods pothesis in metastatic breast cancer. Systemic response consistent The Society for Immunotherapy of Cancer (SITC) led an international with induction and/or boosting of adaptive immunity is predicted by consortium, initiated with 23 expert centers from 17 countries, to evalu- significant enrichment of immune signature. ate the Immunoscore in routine clinical settings. CC patients (pts) stages I/II/III with no prior neo-adjuvant treatment were included in this Acknowledgements study. Overall, 3855 pts split into a training set (TS), internal validation 1RO1CA161891 set (IVS), and external validation set (EVS) were quantified for IM using Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 240 of 242 immunohistochemistry with CD3/CD8 antibodies and digital pathology documented clinical follow up. We identified a preliminary population quantification of whole slide sections. All statistical analyses were pre- of 31 stage II-III melanoma patients diagnosed between 2000 and defined and performed by external statisticians. The primary endpoint 2012, with characteristics shown in Fig. 9 for whom pathology from was time-to-recurrence (TTR); multivariate analyses were performed the primary biopsy was shown. Clinical follow up was available on 18 using Cox models adjusted for IM, age, gender, T-stage, N-stage, and patients of whom 9 patients were alive with no evidence of recur- stratified by participating center. rence, 1 had died of another malignancy, and 7 had died of melan- Results oma. 15 patients had more than 24 months of survival information Across centers, the median recurrent follow-up was 126.6 months. Pt available but no detailed clinical information. 5 μm slides from either characteristics: 51.5% male, median age 69 years, and 17%/54%/29% the primary biopsy or subsequent wide local excision procedure were stage I/II/III, respectively. Among pts with stages I-III CC in the TS, TTR stained using Opal multiplex IHC for DAPI, CD3 (LN10, Leica), CD8 was shorter among 152 pts (22%) with Low-IM CC vs. 548 pts with (4B11, Leica), CD68 (KP1, Biogenex), SOX10 (BC34, Biocare), HLA-DR High-IM CC (HR [95% CI], 0.41 [0.28-0.61]; P < 0.0001). In the IVS, TTR (LN-3, Abcam) and Ki67 (MIB1, Abcam). Cell phenotypes within repre- was also shorter among 155 pts with Low-IM CC vs. 481 pts with sentative fields pre-selected by a trained dermato-pathologist and High-IM CC (0.41 [0.27-0.65]; P < 0.0001). In the EVS, TTR was also were visualized using the Mantra quantitative pathology workstation shorter among 225 pts with Low-IM CC vs. 744 pts with High-IM CC (Perkin Elmer), and analysis of spatial distribution of CD3 + CD8+ cells (0.51 [0.38-0.68]; P < 0.0001). These results were independent of age, analyzed as shown in Figs. 10 and 11 using inForm® image analysis sex, tumor stage, and sidedness. Among secondary objectives, Immu- software (Perkin Elmer), and Spotfire software (TIBCO). noscore groups (High, Int, Low) predicted time to recurrence in the Results TS (0.19 [0.10-0.37]; P < 0.0001), IVS (0.27 [0.14-0.53]; P < 0.0001), and CD3 + CD8+ cells are closer to both tumor (SOX10+) and CD68+ cells EVS (0.33 [0.22-0.49]; P < 0.0001). In stage II CC pts (1433), the differ- when they express HLA-DR (p < 0.001). Conversely, CD3 + CD8+ cells ence in TTR was significant between the Low and High-Immunoscore are significantly farther from Sox10+ cells when they express Ki-67. groups (0.36 [0.23-0.56]; P < 0.0001). In multivariate models, Immuno- Among patients with clinical follow up, CD3 + CD8+ cells in non- score grouping (2, 3, or 5) was significant (C-index : 0.73 [0.66-0.80], recurrent patients were closer to SOX10+HLA-DR+ cells than they all P < 0.0001). Multivariate models including MSI and sidedness were were in recurrent patients(p < 0.001). performed and will also be presented. Reproducibility of the IM assay Conclusions was validated across centers. If proximity is a surrogate for interaction, these data may indicate Conclusions that HLA-DR expression enhances interaction with T cells for both The primary and secondary endpoints of the global Immunoscore CD68+ infiltrating cells and Sox10+ tumor cells. In addition, study were reached. Overall, TTR was significantly longer in pts with CD3+CD8+ cells were closer to SOX10+HLA-DR+ cells in patients who stages I-III CC defined as High-IM. Moreover, a subgroup of patients did not recur, which is interesting in light of recent data showing that with high-risk stage II CC was also identified by Low-IM. expression of HLA-DR by tumor cells increases likelihood of response to anti-PD1. Further staining and analysis of annotated tumor samples Acknowledgements from the complete HICCC cohort 2000-2012 is ongoing and results will This initiative was supported by a variety of sources, including funding from be updated at time of presentation. Definiens, Prometheus, and a grant from the Czech Ministry of Health, 15- 28188A and League against cancer. P26 Defining critical features of the immune microenvironment in melanoma 1 1 2 1 3 Robyn Gartrell , Douglas Marks , Edward Stack , Yan Lu , Daisuke Izaki , 4 4 4 4 Kristen Beck , Dan Tong Jia , Paul Armenta , Ashley White-Stern , Yichun 4 1 5 1 1 Fu , Zoe Blake , Howard L Kaufman , Bret Taback , Basil Horst , Yvonne M Saenger 1 2 Columbia University Medical Center, New York, NY, USA; Perkin Elmer, Hopkinton, MA, USA; Columbia University, New York, NY, USA; Columbia University College of Physicians and Surgeons, New York, NY, USA; Rutgers Cancer Institute of New Jersey, New Brunswick, NJ, USA; New York Presbyterian/Columbia University Medical Center, New York, NY, USA Correspondence: Robyn Gartrell (rdg2129@columbia.edu) Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P26 Background Precise biomarkers are urgently needed to characterize the tumor immune micro-environment, both for prognostication and to predict the benefit of immuno-therapeutic intervention. HLA-DR on tumor cells and Ki67 on cytotoxic (CD8+) T cells have been proposed as biomarkers of anti-PD1 activity. Multiplex immunohistochemistry (mIHC) allows for automated quantitation of phenotypes and spatial distributions of immune cell populations within formalin fixed paraf- fin embedded (FFPE) tissues. Methods In order to test whether mIHC can better characterize the tumor im- mune microenvironment, we screened databases at the Herbert Irving Cancer Center (HICC) at Columbia University for early stage melanoma Fig. 9 (abstract P26). Demographic Characteristics of Melanoma patients with available FFPE primary melanoma tissue and Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 241 of 242 microenvironment (flipping immunosuppressive M2 macrophages to an anti-tumor M1 state), and T cell expansion and activation via den- dritic cell maturation and antigen presentation. Clinically, Imprime has demonstrated promising efficacy in clinical trials when combined with tumor-targeting or anti-angiogenic antibodies. Phase II studies with pembrolizumab are starting in both metastatic triple negative breast cancer and metastatic melanoma. Herein, we have employed multi- plexed immunofluorescence to profile the immune microenvironment in preclinical tumor tissues. Methods The B16F10 experimental metastasis model was used to interro- gate Imprime’s anti-tumor activity in vivo. B16F10 melanoma cells were injected into the tail vein of syngeneic C57BL/6 mice, seed- ing the lungs with B16 foci. Outgrowth of these metastatic foci was assessed after treatment with the tumor-targeting anti-tryp1 Fig. 10 (abstract P26). Distance between CD3 + CD8+ T cells and antibody TA99, Imprime, or the combination. At various times nearest neighbor, Left: Mean distance from CD3 + CD8+ to CD68 + post tumor injection, lungs were examined via multiplexed im- HLA-DR- (light blue) or CD68 + HLA-DR+ (dark blue), Center: Mean munofluorescence (IFC) for markers of immune infiltration and distance from CD3 + CD8+ to SOX10 + HLA-DR- (pink) or SOX10 + activation. IFC was performed using 7-color staining (Opal tech- HLA-DR+ red), Right: Mean distance from CD3 + CD8+ to SOX10 + nology, PerkinElmer) combined with in situ hybridization (RNA- Ki67- (pink) or SOX10 + Ki67+ red) Scope, ACD). Images were acquired with the Vectra3 multispectral imaging system and cells segmented using Inform (PerkinElmer). Imaging data were transformed into “.fcs” files and analyzed using Flowjo flow cytometry software (Treestar)). Relational pa- rameters such as immune cell clustering and tumor infiltration were performed via custom algorithms in R. Results TA99 alone suppressed the outgrowth of B16 lung metastases by 54% when compared to vehicle treatment. The combination of Imprime with TA99 reduced the number of metastases even more profoundly (96% vs vehicle). IFC analyses showed that Imprime specifically accumulates in the tumor stroma, binds to macro- phages and elicits increased iNOS production, indicating the re- polarization of these macrophages to a more M1-like, inflamma- tory state. Imprime-treatment also triggered the formation of large immune cell clusters, possibly representing resolved tumor nests or the establishment of tertiary lymphoid tissues, both of which have been identified as predictors of successful anti-tumor im- mune responses. Finally, Imprime treatment and localization at the tumor site corresponds with substantial upregulation of the gene Mx1- a type 1 interferon-responsive gene. Conclusions Imprime is a potent immunomodulator that induces a coordi- nated immune attack in vivo demonstrated by immune cell binding, M1 re-polarization and a type-1 interferon signature Fig. 11 (abstract P26). Distance between CD3 + CD8+ T cells and that coincides with reduced outgrowth of established ling nearest neighbor be recurrence status, Top Left: Mean distance from metastases. CD3 + CD8+ to CD68 + HLA-DR- by recurrence status, Lower Left: Mean distance from CD3 + CD8+ to CD68 + HLA-DR+ by recurrence status. Top Right: Mean distance from CD3 + CD8+ to SOX10 + HLA- P28 DR- by recurrence status, Bottom Right: Mean distance from CD3 + Local convection-enhanced delivery of PD-1 blockade antibody in CD8+ to SOX10 + HLA-DR+ by recurrence status de novo murine model of glioblastoma Jennifer S Sims, Liang Lei, Takashi Tsujiuchi, Jeffrey N Bruce, Peter Canoll Columbia University Medical Center, New York, NY, USA P27 Correspondence: Jennifer S Sims (jennifer.s.sims@gmail.com) Immune profiling via multiplexed immunofluorescence shows that Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P28 Imprime based anti-cancer efficacy involves profound changes in macrophage polarization, type 1 IFN signaling and the formation Background of immune cell clusters Systemic delivery of anti-PD1 antibody therapy has proven rela- Steven Leonardo, Keith Gorden, Ross B Fulton, Kathryn Fraser, Takashi O tively safe in glioma patients, but therapeutic response remains Kangas, Richard Walsh, Kathleen Ertelt, Jeremy Graff, Mark Uhlik unpredictable and persistently low. Checkpoint blockade anti- Biothera Pharmaceuticals Inc., Eagan, MN, USA bodies face numerous potential confounders in these tumors, Correspondence: Steven Leonardo (sleonardo@biothera.com) such as the blood-brain barrier, poor local or lymph node presen- Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):P27 tation of tumor antigens, and unknown dependency on PD-1/PD- L1 activity during tumor progression. Here, we conducted a pilot Background study using intracranial convection-enhanced delivery (CED) of Imprime PGG (Imprime) is a soluble, intravenously (iv) administered anti-PD1 (mDX400) into a de novo murine glioma model to the yeast 1,3/1,6 β-glucan PAMP (pathogen-associated molecular pattern). dissect tumor and immune perturbations following local treat- As a PAMP, Imprime triggers innate immune function, including direct ment, and to compare the efficacy of treating during early or late tumor killing, repolarization of the immunosuppressive tumor tumor progression. Journal for ImmunoTherapy of Cancer 2016, 4(Suppl 2):91 Page 242 of 242 Methods produced several “long-term survivors”, who lived up to 158 days Transgenic C57BL/6-PTEN(fl/fl) mice were injected with a retro- with stable tumor burden. While substantial T cell infiltration was virus expressing PDGFb and cre recombinase, inducing tumori- detected in the end-stage tumors of both mDX400- and isotype- genesis as previously described. In this model, with a median treated mice by immunohistochemistry (CD3e), expression of im- survival of 80 days post-tumor induction (D80), convergence to a mune signaling pathways (e.g., Fc receptor and Toll-like receptor stereotyped subset of genomic rearrangements occurs by ap- families, phagosome/lysosome components), was significantly higher proximately D35. Intracranial osmotic pumps filled with mDX400 among three long-surviving mDX400-treated mice than in three or isotype control antibody solution were implanted at the tumor isotype-treated mice. site for 14-day windows spanning (D28-D42) or following (D42- Conclusions D56) this developmental transition, then removed. Tumor burden Our pilot study of mDX400 administration by CED identified an impact was monitored by bioluminescence (luciferase reporter), and mice on tumor burden during and following therapy, but a lack of survival were sacrificed upon presentation of tumor-related morbidity. Tis- benefit for D28-D42 treatment. While additional experiments are needed sue was formalin-fixed for histopathology and cryopreserved for to statistically evaluate survival benefit for the later treatment window, gene expression analysis. differentially high intratumoral expression of genes reflecting immune ac- Results tivation among mDX400-treated, long-surviving mice demonstrates that During both treatment windows, tumor burden decreased differen- molecular study in this model may elucidate intratumoral conditions as- tially in mDX400-treated mice. While survival time between mDX400- sociated with response to anti-PD1 blockade in glioma. and isotype-treated mice was nearly identical for D28-D42 (both median D70), for mice treated between D42-D56, median survival Acknowledgements differed (D88 vs. D68), but without statistical significance between This pre-clinical study is supported by Merck & Co. Investigator-Initiated the groups (p = 0.25). Interestingly, the D42-D56 mDX400 group Sponsored Projects grant LKR146174. Submit your next manuscript to BioMed Central and we will help you at every step: • We accept pre-submission inquiries � Our selector tool helps you to find the most relevant journal � We provide round the clock customer support � Convenient online submission � Thorough peer review � Inclusion in PubMed and all major indexing services � Maximum visibility for your research Submit your manuscript at www.biomedcentral.com/submit

Journal

Journal for ImmunoTherapy of CancerSpringer Journals

Published: Dec 8, 2016

There are no references for this article.