Journal Articles

Synthetic TRuC receptors engaging the complete T cell receptor for potent anti-tumor response

T cells expressing CD19-targeting chimeric antigen receptors (CARs) reveal high efficacy in the treatment of B cell malignancies. Here, we report that T cell receptor fusion constructs (TRuCs) comprising an antibody-based binding domain fused to T cell receptor (TCR) subunits can effectively reprogram an intact TCR complex to recognize tumor surface antigens. Unlike CARs, TRuCs become a functional component of the TCR complex. TRuC-T cells kill tumor cells as potently as second-generation CAR-T cells, but at significant lower cytokine release and despite the absence of an extra co-stimulatory domain. TRuC-T cells demonstrate potent anti-tumor activity in both liquid and solid tumor xenograft models. In several models, TRuC-T cells are more efficacious than respective CAR-T cells. TRuC-T cells are shown to engage the signaling capacity of the entire TCR complex in an HLA-independent manner.

Author Info: (1) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (2) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (3) TCR(2) Therapeutics, I

Author Info: (1) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (2) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (3) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (4) Department of Immunology, Faculty of Biology, BIOSS Center for Biological Signalling Studies, CIBSS-Centre for Integrative Biological Signalling Studies and Centre for Chronic Immunodeficiency CCI, University of Freiburg, Schanzlestrabetae 18, Freiburg, 79104, Germany. (5) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (6) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (7) Cellular Immunotherapy Program, Massachusetts General Hospital Cancer Center and Harvard Medical School, Bldg. 149 13th Street, Charlestown, MA, USA. (8) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (9) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (10) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (11) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (12) Department of Immunology, Faculty of Biology, BIOSS Center for Biological Signalling Studies, CIBSS-Centre for Integrative Biological Signalling Studies and Centre for Chronic Immunodeficiency CCI, University of Freiburg, Schanzlestrabetae 18, Freiburg, 79104, Germany. (13) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (14) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (15) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (16) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (17) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (18) Cellular Immunotherapy Program, Massachusetts General Hospital Cancer Center and Harvard Medical School, Bldg. 149 13th Street, Charlestown, MA, USA. (19) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. (20) Department of Immunology, Faculty of Biology, BIOSS Center for Biological Signalling Studies, CIBSS-Centre for Integrative Biological Signalling Studies and Centre for Chronic Immunodeficiency CCI, University of Freiburg, Schanzlestrabetae 18, Freiburg, 79104, Germany. (21) TCR(2) Therapeutics, Inc., 100 Binney Street, Cambridge, MA, 02142, USA. robert@tcr2.com.

Genetic diversity of tumors with mismatch repair deficiency influences anti-PD-1 immunotherapy response

Tumors with mismatch repair deficiency (MMR-d) are characterized by sequence alterations in microsatellites and can accumulate thousands of mutations. This high mutational burden renders tumors immunogenic and sensitive to programmed cell death-1 (PD-1) immune checkpoint inhibitors. Yet, despite their tumor immunogenicity, patients with MMR-deficient tumors experience highly variable responses, and roughly half are refractory to treatment. We present experimental and clinical evidence showing that the degree of microsatellite instability (MSI) and resultant mutational load, in part, underlies the variable response to PD-1 blockade immunotherapy in MMR-d human and mouse tumors. The extent of response is particularly associated with the accumulation of insertion-deletion (indel) mutational load. This study provides a rationale for the genome-wide characterization of MSI intensity and mutational load to better profile responses to anti-PD-1 immunotherapy across MMR-deficient human cancers.

Author Info: (1) Department of Otolaryngology-Head and Neck Surgery, Johns Hopkins University, Baltimore, MD 21287, USA. Bloomberg-Kimmel Institute for Cancer Immunotherapy at Johns Hopkins, Ba

Author Info: (1) Department of Otolaryngology-Head and Neck Surgery, Johns Hopkins University, Baltimore, MD 21287, USA. Bloomberg-Kimmel Institute for Cancer Immunotherapy at Johns Hopkins, Baltimore, MD 21287, USA. Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (2) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (3) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (4) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (5) Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA. (6) Computational and Systems Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (7) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (8) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (9) Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (10) Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (11) Department of Otolaryngology-Head and Neck Surgery, Weill Cornell New York Presbyterian Hospital, New York, NY 10065, USA. (12) Bloomberg-Kimmel Institute for Cancer Immunotherapy at Johns Hopkins, Baltimore, MD 21287, USA. Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA. Swim Across America Laboratory at Johns Hopkins, Baltimore, MD 21287, USA. (13) Swim Across America Laboratory at Johns Hopkins, Baltimore, MD 21287, USA. (14) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (15) Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (16) Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (17) Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (18) Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (19) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Head and Neck Service, Department of Surgery, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (20) Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (21) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (22) Bloomberg-Kimmel Institute for Cancer Immunotherapy at Johns Hopkins, Baltimore, MD 21287, USA. Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA. Swim Across America Laboratory at Johns Hopkins, Baltimore, MD 21287, USA. (23) Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Division of Solid Tumor Oncology, Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (24) Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. chant@mskcc.org. Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Immunogenomics and Precision Oncology Platform, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.

CD8(+) T cells regulate tumour ferroptosis during cancer immunotherapy

Cancer immunotherapy restores or enhances the effector function of CD8(+) T cells in the tumour microenvironment(1,2). CD8(+) T cells activated by cancer immunotherapy clear tumours mainly by inducing cell death through perforin-granzyme and Fas-Fas ligand pathways(3,4). Ferroptosis is a form of cell death that differs from apoptosis and results from iron-dependent accumulation of lipid peroxide(5,6). Although it has been investigated in vitro(7,8), there is emerging evidence that ferroptosis might be implicated in a variety of pathological scenarios(9,10). It is unclear whether, and how, ferroptosis is involved in T cell immunity and cancer immunotherapy. Here we show that immunotherapy-activated CD8(+) T cells enhance ferroptosis-specific lipid peroxidation in tumour cells, and that increased ferroptosis contributes to the anti-tumour efficacy of immunotherapy. Mechanistically, interferon gamma (IFNgamma) released from CD8(+) T cells downregulates the expression of SLC3A2 and SLC7A11, two subunits of the glutamate-cystine antiporter system xc(-), impairs the uptake of cystine by tumour cells, and as a consequence, promotes tumour cell lipid peroxidation and ferroptosis. In mouse models, depletion of cystine or cysteine by cyst(e)inase (an engineered enzyme that degrades both cystine and cysteine) in combination with checkpoint blockade synergistically enhanced T cell-mediated anti-tumour immunity and induced ferroptosis in tumour cells. Expression of system xc(-) was negatively associated, in cancer patients, with CD8(+) T cell signature, IFNgamma expression, and patient outcome. Analyses of human transcriptomes before and during nivolumab therapy revealed that clinical benefits correlate with reduced expression of SLC3A2 and increased IFNgamma and CD8. Thus, T cell-promoted tumour ferroptosis is an anti-tumour mechanism, and targeting this pathway in combination with checkpoint blockade is a potential therapeutic approach.

Author Info: (1) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel

Author Info: (1) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (2) Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. Department of Radiation Oncology, University of Michigan School of Medicine, Ann Arbor, MI, USA. (3) Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. Department of Pathology, University of Michigan School of Medicine, Ann Arbor, MI, USA. Michigan Center for Translational Pathology, University of Michigan School of Medicine, Ann Arbor, MI, USA. (4) Cayman Chemical Company, Ann Arbor, MI, USA. (5) Cayman Chemical Company, Ann Arbor, MI, USA. (6) Cayman Chemical Company, Ann Arbor, MI, USA. (7) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (8) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. Department of Radiation Oncology, University of Michigan School of Medicine, Ann Arbor, MI, USA. (9) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (10) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (11) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (12) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (13) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (14) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (15) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (16) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (17) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (18) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (19) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. (20) Institute for Cancer Genetics, Department of Pathology and Cell Biology, and Herbert Irving Comprehensive Cancer Center, College of Physicians and Surgeons, Columbia University, New York, NY, USA. (21) Department of Obstetrics and Gynecology, University of Michigan School of Medicine, Ann Arbor, MI, USA. (22) Department of Radiation Oncology, University of Michigan School of Medicine, Ann Arbor, MI, USA. (23) Department of Chemical Engineering, University of Texas at Austin, Austin, TX, USA. Department of Molecular Biosciences, University of Texas at Austin, Austin, TX, USA. (24) Department of Chemical Engineering, University of Texas at Austin, Austin, TX, USA. Department of Molecular Biosciences, University of Texas at Austin, Austin, TX, USA. (25) Department of Pathology, University of Michigan School of Medicine, Ann Arbor, MI, USA. Department of Computational Medicine & Bioinformatics, University of Michigan School of Medicine, Ann Arbor, MI, USA. (26) Department of Chemical Engineering, University of Texas at Austin, Austin, TX, USA. Department of Molecular Biosciences, University of Texas at Austin, Austin, TX, USA. (27) Department of Chemical Engineering, University of Texas at Austin, Austin, TX, USA. Department of Molecular Biosciences, University of Texas at Austin, Austin, TX, USA. (28) Immunogenomics and Precision Oncology Platform, Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY, USA. (29) Department of Pathology, University of Michigan School of Medicine, Ann Arbor, MI, USA. Michigan Center for Translational Pathology, University of Michigan School of Medicine, Ann Arbor, MI, USA. Howard Hughes Medical Institute, University of Michigan School of Medicine, Ann Arbor, MI, USA. (30) Department of Surgery, University of Michigan School of Medicine, Ann Arbor, MI, USA. wzou@med.umich.edu. Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA. wzou@med.umich.edu. Department of Pathology, University of Michigan School of Medicine, Ann Arbor, MI, USA. wzou@med.umich.edu. Graduate Program in Immunology, University of Michigan School of Medicine, Ann Arbor, MI, USA. wzou@med.umich.edu. Graduate Program in Cancer Biology, University of Michigan School of Medicine, Ann Arbor, MI, USA. wzou@med.umich.edu.

Prophylactic TNF blockade uncouples efficacy and toxicity in dual CTLA-4 and PD-1 immunotherapy

Combined PD-1 and CTLA-4-targeted immunotherapy with nivolumab and ipilimumab is effective against melanoma, renal cell carcinoma and non-small-cell lung cancer(1-3). However, this comes at the cost of frequent, serious immune-related adverse events, necessitating a reduction in the recommended dose of ipilimumab that is given to patients(4). In mice, co-treatment with surrogate anti-PD-1 and anti-CTLA-4 monoclonal antibodies is effective in transplantable cancer models, but also exacerbates autoimmune colitis. Here we show that treating mice with clinically available TNF inhibitors concomitantly with combined CTLA-4 and PD-1 immunotherapy ameliorates colitis and, in addition, improves anti-tumour efficacy. Notably, TNF is upregulated in the intestine of patients suffering from colitis after dual ipilimumab and nivolumab treatment. We created a model in which Rag2(-/-)Il2rg(-/-) mice were adoptively transferred with human peripheral blood mononuclear cells, causing graft-versus-host disease that was further exacerbated by ipilimumab and nivolumab treatment. When human colon cancer cells were xenografted into these mice, prophylactic blockade of human TNF improved colitis and hepatitis in xenografted mice, and moreover, immunotherapeutic control of xenografted tumours was retained. Our results provide clinically feasible strategies to dissociate efficacy and toxicity in the use of combined immune checkpoint blockade for cancer immunotherapy.

Author Info: (1) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. Department of Oncology,

Author Info: (1) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. Department of Oncology, Clinica Universidad de Navarra, Pamplona, Spain. Department of Oncology, Hospital Costa del Sol, Marbella, Spain. Instituto de Investigacion Biomedica de Malaga (IBIMA), Hospitales Universitarios Regional y Virgen de la Victoria, Malaga, Spain. (2) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. (3) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. (4) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. (5) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain. (6) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. (7) Navarra Institute for Health Research (IDISNA), Pamplona, Spain. Department of Pathology, Clinica Universidad de Navarra, Pamplona, Spain. (8) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Department of Oncology, Clinica Universidad de Navarra, Pamplona, Spain. (9) Navarra Institute for Health Research (IDISNA), Pamplona, Spain. Department of Oncology, Clinica Universidad de Navarra, Pamplona, Spain. Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain. (10) Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain. Department of Oncology, Hospital General Universitario Gregorio Maranon, Madrid, Spain. (11) Department of Oncology, Hospital Universitario Virgen de la Victoria, Malaga, Spain. (12) Instituto de Investigacion Biomedica de Malaga (IBIMA), Hospitales Universitarios Regional y Virgen de la Victoria, Malaga, Spain. Laboratorio de Biologia Molecular del Cancer, Centro de Investigaciones Medico-Sanitarias (CIMES), Universidad de Malaga, Malaga, Spain. Department of Pathology, Faculty of Medicine, Universidad de Malaga, Malaga, Spain. (13) Instituto de Investigacion Biomedica de Malaga (IBIMA), Hospitales Universitarios Regional y Virgen de la Victoria, Malaga, Spain. Laboratorio de Biologia Molecular del Cancer, Centro de Investigaciones Medico-Sanitarias (CIMES), Universidad de Malaga, Malaga, Spain. (14) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. (15) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain. (16) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. pberraondol@unav.es. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. pberraondol@unav.es. Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain. pberraondol@unav.es. (17) Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain. imelero@unav.es. Navarra Institute for Health Research (IDISNA), Pamplona, Spain. imelero@unav.es. Department of Oncology, Clinica Universidad de Navarra, Pamplona, Spain. imelero@unav.es. Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain. imelero@unav.es. Department of Immunology and Immunotherapy, Clinica Universidad de Navarra, Pamplona, Spain. imelero@unav.es.

Endogenous CD4+ T cells recognize neoantigens in lung cancer patients, including recurrent oncogenic KRAS and ERBB2 (Her2) driver mutations

T cells specific for neoantigens encoded by mutated genes in cancers are increasingly recognized as mediators of tumor destruction after immune checkpoint inhibitor therapy or adoptive cell transfer. Much of the focus has been on identifying epitopes presented to CD8+ T cells by class I MHC. However, CD4+ class II MHC-restricted T cells have been shown to have an important role in antitumor immunity. Unfortunately, the vast majority of neoantigens recognized by CD8+ or CD4+ T cells in cancer patients result from random mutations and are patient-specific. Here, we screened the blood of 5 NSCLC patients for T-cell responses to candidate mutation-encoded neoepitopes. T-cell responses were detected to 8.8% of screened antigens, with 1-7 antigens identified per patient. A majority of responses were to random, patient-specific mutations. However, CD4+ T cells that recognized the recurrent KRASG12V and the ERBB2 (Her2) internal tandem duplication (ITD) oncogenic driver mutations, but not the corresponding wildtype sequences, were identified in two patients. Two different T-cell receptors (TCRs) specific for KRASG12V and one T-cell receptor specific for Her2-ITD were isolated and conferred antigen specificity when transfected into T cells. Deep sequencing identified the Her2-ITD-specific TCR in the tumor but not non-adjacent lung. Our results showed that CD4+ T-cell responses to neoantigens, including recurrent driver mutations, can be derived from the blood of NSCLC patients. These data support the use of adoptive transfer or vaccination to augment CD4+ neoantigen-specific T cells and elucidate their role in human antitumor immunity.

Author Info: (1) Clinical Research Division, Fred Hutchinson Cancer Research Center jveatch@fhcrc.org. (2) Immunology, Fred Hutchinson Cancer Research Center. (3) Otto Loewi Research Center, Di

Author Info: (1) Clinical Research Division, Fred Hutchinson Cancer Research Center jveatch@fhcrc.org. (2) Immunology, Fred Hutchinson Cancer Research Center. (3) Otto Loewi Research Center, Division of Pharmacology, Medical University of Graz. (4) Bioinformatics Shared Resource, Fred Hutchinson Cancer Research Center. (5) University of Washington. (6) Department of Medicine, University of Washington. (7) Department of Medicine, Division of Medical Oncology, University of Washington. (8) Clinical Research, Fred Hutchinson Cancer Research Center. (9) Fred Hutchinson Cancer Research Center.

Multi-omics discovery of exome-derived neoantigens in hepatocellular carcinoma

BACKGROUND: Although mutated HLA ligands are considered ideal cancer-specific immunotherapy targets, evidence for their presentation is lacking in hepatocellular carcinomas (HCCs). Employing a unique multi-omics approach comprising a neoepitope identification pipeline, we assessed exome-derived mutations naturally presented as HLA class I ligands in HCCs. METHODS: In-depth multi-omics analyses included whole exome and transcriptome sequencing to define individual patient-specific search spaces of neoepitope candidates. Evidence for the natural presentation of mutated HLA ligands was investigated through an in silico pipeline integrating proteome and HLA ligandome profiling data. RESULTS: The approach was successfully validated in a state-of-the-art dataset from malignant melanoma, and despite multi-omics evidence for somatic mutations, mutated naturally presented HLA ligands remained elusive in HCCs. An analysis of extensive cancer datasets confirmed fundamental differences of tumor mutational burden in HCC and malignant melanoma, challenging the notion that exome-derived mutations contribute relevantly to the expectable neoepitope pool in malignancies with only few mutations. CONCLUSIONS: This study suggests that exome-derived mutated HLA ligands appear to be rarely presented in HCCs, inter alia resulting from a low mutational burden as compared to other malignancies such as malignant melanoma. Our results therefore demand widening the target scope for personalized immunotherapy beyond this limited range of mutated neoepitopes, particularly for malignancies with similar or lower mutational burden.

Author Info: (1) Department of General, Visceral and Transplant Surgery, University Hospital Tubingen, Hoppe-Seyler-Str. 3, D-72076, Tubingen, Germany. Markus.Loeffler@uni-tuebingen.de. Interfa

Author Info: (1) Department of General, Visceral and Transplant Surgery, University Hospital Tubingen, Hoppe-Seyler-Str. 3, D-72076, Tubingen, Germany. Markus.Loeffler@uni-tuebingen.de. Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. Markus.Loeffler@uni-tuebingen.de. German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ) Partner Site Tubingen, Tubingen, Germany. Markus.Loeffler@uni-tuebingen.de. Department of Clinical Pharmacology, University Hospital Tubingen, Auf der Morgenstelle 8, D-72076, Tubingen, Germany. Markus.Loeffler@uni-tuebingen.de. (2) Institute for Translational Bioinformatics, University Hospital Tubingen, Tubingen, Germany. Quantitative Biology Center (QBiC), University of Tubingen, Auf der Morgenstelle 10, D-72076, Tubingen, Germany. (3) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. Center for Bioinformatics, University of Tubingen, Sand 14, D-72076, Tubingen, Germany. Department of Computer Science, Applied Bioinformatics, Sand 14, D-72076, Tubingen, Germany. (4) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ) Partner Site Tubingen, Tubingen, Germany. (5) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. Center for Bioinformatics, University of Tubingen, Sand 14, D-72076, Tubingen, Germany. Department of Computer Science, Applied Bioinformatics, Sand 14, D-72076, Tubingen, Germany. Present address: European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), Wellcome Trust Genome Campus, Hinxton, Cambridgeshire, CB10 1SD,, United Kingdom. (6) Institute of Medical Genetics and Applied Genomics, University Hospital Tubingen, Calwerstr. 7, D-72076, Tubingen, Germany. (7) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. (8) Institute of Medical Genetics and Applied Genomics, University Hospital Tubingen, Calwerstr. 7, D-72076, Tubingen, Germany. (9) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. (10) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. (11) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. Present address: Immatics Biotechnologies GmbH, Paul-Ehrlich-Str. 15, D-72076, Tubingen, Germany. (12) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. Present address: Immatics Biotechnologies GmbH, Paul-Ehrlich-Str. 15, D-72076, Tubingen, Germany. (13) Institute of Medical Genetics and Applied Genomics, University Hospital Tubingen, Calwerstr. 7, D-72076, Tubingen, Germany. (14) Institute of Medical Genetics and Applied Genomics, University Hospital Tubingen, Calwerstr. 7, D-72076, Tubingen, Germany. (15) Institute of Medical Genetics and Applied Genomics, University Hospital Tubingen, Calwerstr. 7, D-72076, Tubingen, Germany. NGS Competence Center Tubingen (NCCT), University of Tubingen, Tubingen, Germany. (16) Quantitative Biology Center (QBiC), University of Tubingen, Auf der Morgenstelle 10, D-72076, Tubingen, Germany. (17) Quantitative Biology Center (QBiC), University of Tubingen, Auf der Morgenstelle 10, D-72076, Tubingen, Germany. (18) Department of General, Visceral and Transplant Surgery, University Hospital Tubingen, Hoppe-Seyler-Str. 3, D-72076, Tubingen, Germany. (19) Department of General, Visceral and Transplant Surgery, University Hospital Tubingen, Hoppe-Seyler-Str. 3, D-72076, Tubingen, Germany. (20) Department of General, Visceral and Transplant Surgery, University Hospital Tubingen, Hoppe-Seyler-Str. 3, D-72076, Tubingen, Germany. (21) Department of General, Visceral and Transplant Surgery, University Hospital Tubingen, Hoppe-Seyler-Str. 3, D-72076, Tubingen, Germany. Present address: Department of General and Visceral Surgery, Schwarzwald-Baar Hospital, Klinikstr. 11, D-78052, Villingen-Schwenningen, Germany. (22) Institute of Pathology and Neuropathology, University Hospital Tubingen, Liebermeisterstr. 8, D-72076, Tubingen, Germany. (23) Institute of Pathology and Neuropathology, University Hospital Tubingen, Liebermeisterstr. 8, D-72076, Tubingen, Germany. (24) Interfaculty Institute for Cell Biology, Proteome Center Tubingen (PCT), University of Tubingen, Auf der Morgenstelle 15, 72076, Tubingen, Germany. (25) Interfaculty Institute for Cell Biology, Proteome Center Tubingen (PCT), University of Tubingen, Auf der Morgenstelle 15, 72076, Tubingen, Germany. (26) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ) Partner Site Tubingen, Tubingen, Germany. Internal Medicine, Department for Oncology, Hematology, Immunology, Rheumatology and Pulmonology, University of Tubingen, Otfried-Muller-Str. 10, D-72076, Tubingen, Germany. (27) Cancer Immunoregulation Unit, Istituto Nazionale per lo Studio e la Cura dei Tumori, "Fondazione Pascale" - IRCCS, 80131, Naples, Italy. (28) German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ) Partner Site Tubingen, Tubingen, Germany. Institute for Translational Bioinformatics, University Hospital Tubingen, Tubingen, Germany. Quantitative Biology Center (QBiC), University of Tubingen, Auf der Morgenstelle 10, D-72076, Tubingen, Germany. Center for Bioinformatics, University of Tubingen, Sand 14, D-72076, Tubingen, Germany. Department of Computer Science, Applied Bioinformatics, Sand 14, D-72076, Tubingen, Germany. NGS Competence Center Tubingen (NCCT), University of Tubingen, Tubingen, Germany. Max Planck Institute for Developmental Biology, Biomolecular Interactions, Spemannstr. 35, D-72076, Tubingen, Germany. (29) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ) Partner Site Tubingen, Tubingen, Germany. (30) Department of General, Visceral and Transplant Surgery, University Hospital Tubingen, Hoppe-Seyler-Str. 3, D-72076, Tubingen, Germany. German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ) Partner Site Tubingen, Tubingen, Germany. (31) Interfaculty Institute for Cell Biology, Department of Immunology, University of Tubingen, Auf der Morgenstelle 15, D-72076, Tubingen, Germany. German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ) Partner Site Tubingen, Tubingen, Germany.

Identification of Neoantigen-Reactive Tumor-Infiltrating Lymphocytes in Primary Bladder Cancer

Immune checkpoint inhibitors are effective in treating a variety of malignancies, including metastatic bladder cancer. A generally accepted hypothesis suggests that immune checkpoint inhibitors induce tumor regressions by reactivating a population of endogenous tumor-infiltrating lymphocytes (TILs) that recognize cancer neoantigens. Although previous studies have identified neoantigen-reactive TILs from several types of cancer, no study to date has shown whether neoantigen-reactive TILs can be found in bladder tumors. To address this, we generated TIL cultures from patients with primary bladder cancer and tested their ability to recognize tumor-specific mutations. We found that CD4(+) TILs from one patient recognized mutated C-terminal binding protein 1 in an MHC class II-restricted manner. This finding suggests that neoantigen-reactive TILs reside in bladder cancer, which may help explain the effectiveness of immune checkpoint blockade in this disease and also provides a rationale for the future use of adoptive T cell therapy targeting neoantigens in bladder cancer.

Author Info: (1) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. Hematology Branch, National Heart, Lung and Blood Institute, National Institutes o

Author Info: (1) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20982. (2) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. (3) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. (4) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. (5) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. (6) Genitourinary Malignancies Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892; and. (7) Urologic Oncology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. (8) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. (9) Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892; Yong-Chen.Lu@nih.gov.

Efficacy and Safety Analysis of Nelipepimut-S Vaccine to Prevent Breast Cancer Recurrence: A Randomized, Multicenter, Phase III Clinical Trial

PURPOSE: In phase I/II studies, nelipepimut-S (NP-S) plus granulocyte-macrophage colony-stimulating factor (GM-CSF) vaccine was well tolerated and effectively raised HER2-specific immunity in breast cancer patients. Results from a prespecified interim analysis of a phase III trial assessing NP-S+GM-CSF are reported. EXPERIMENTAL DESIGN: This multicenter, randomized, double-blind phase III study enrolled females >/=18 years with T1-T3, HER2 low-expressing (immunohistochemistry 1+/2+), node-positive breast cancer in the adjuvant setting. Patients received 1000 mug NP-S+250 microg GM-CSF or placebo+GM-CSF monthly for 6 months, then every 6 months through 36 months. The primary objective was disease-free survival (DFS). Protocol-specified imaging occurred annually. New abnormalities were categorized as recurrence events; biopsy confirmation was not mandated. The interim analysis was conducted as specified in the protocol after 73 DFS events. RESULTS: Seven hundred fifty-eight patients (mean age 51.8 years) were randomized. Adverse events were similar between groups; most common were injection-associated: erythema (84.3%), induration (55.8%), pruritus (54.9%). There was no significant between-arms difference in DFS events at interim analysis at median follow-up (16.8 months). In the NP-S arm, imaging detected 54.1% of recurrence events in asymptomatic patients versus 29.2% in the placebo arm (P=0.069). CONCLUSIONS: NP-S was well tolerated. There was no significant difference in DFS events between NP-S and placebo. Use of mandated annual scans and image-detected recurrence events hastened the interim analysis contributing to early trial termination. Trial registration ID: NCT01479244.

Author Info: (1) Department of Surgery, Brigham and Women's Hospital emittendorf@bwh.harvard.edu. (2) Independent Statistical Contractor. (3) Hematology/Oncology, University of California, San

Author Info: (1) Department of Surgery, Brigham and Women's Hospital emittendorf@bwh.harvard.edu. (2) Independent Statistical Contractor. (3) Hematology/Oncology, University of California, San Francisco. (4) Division of Medical Oncology, University of Alberta, Cross Cancer Institute. (5) Oncology and Medical Radiology Department, Dnipropetrovsk Multifield Clinical Hospital. (6) Cancer Centre, University Hosptials of North Midlands and Keele University. (7) mamology depatment, Kyiv Regional Oncologic Dispensary. (8) Clinical and Expeimental Pathology, Masaryk Memorial Cancer Institute. (9) Metis Foundation, Cancer Vaccine Development Program.

A library of Neo Open Reading Frame peptides (NOPs) as a sustainable resource of common neoantigens in up to 50% of cancer patients

Somatic mutations in cancer can result in neoantigens against which patients can be vaccinated. The quest for tumor specific neoantigens has yielded no targets that are common to all tumors, yet foreign to healthy cells. Single base pair substitutions (SNVs) at best can alter 1 amino acid which can result in a neoantigen; with the exception of rare site-specific oncogenic driver mutations (such as RAS) such mutations are private. Here, we describe a source of common neoantigens induced by frame shift mutations, based on analysis of 10,186 TCGA tumor samples. We find that these frame shift mutations can produce long neoantigens. These are completely new to the body, and indeed recent evidence suggests that frame shifts can be highly immunogenic. We report that many different frame shift mutations converge to the same small set of 3' neo open reading frame peptides (NOPs), all encoded by the Neo-ORFeome. We find that a fixed set of only 1,244 neo-peptides in as much as 30% of all TCGA cancer patients. For some tumor classes this is higher; e.g. for colon and cervical cancer, peptides derived from only ten genes (saturated at 90 peptides) can be applied to 39% of all patients. 50% of all TCGA patients can be achieved at saturation (using all those peptides in the library found more than once). A pre-fabricated library of vaccines (peptide, RNA or DNA) based on this set can provide off the shelf, quality certified, 'personalized' vaccines within hours, saving months of vaccine preparation. This is crucial for critically ill cancer patients with short average survival expectancy after diagnosis.

Author Info: (1) Amsterdam UMC, University of Amsterdam, Department of Oncogenomics, Meibergdreef 9, Amsterdam, The Netherlands. jankoster@amc.uva.nl. (2) myTomorrows, Antoni Fokkerweg 61, Amst

Author Info: (1) Amsterdam UMC, University of Amsterdam, Department of Oncogenomics, Meibergdreef 9, Amsterdam, The Netherlands. jankoster@amc.uva.nl. (2) myTomorrows, Antoni Fokkerweg 61, Amsterdam, The Netherlands. ronald.plasterk@frametherapeutics.com. Founder/CEO, Frame Cancer Therapeutics, Science Park 106, Amsterdam, 1098 XG, The Netherlands. ronald.plasterk@frametherapeutics.com. Amsterdam UMC, Department of Clinical Epidemiology, Biostatistics and Bioinformatics, Amsterdam, The Netherlands. ronald.plasterk@frametherapeutics.com.

Adaptive Immune Resistance Emerges from Tumor-Initiating Stem Cells

Our bodies are equipped with powerful immune surveillance to clear cancerous cells as they emerge. How tumor-initiating stem cells (tSCs) that form and propagate cancers equip themselves to overcome this barrier remains poorly understood. To tackle this problem, we designed a skin cancer model for squamous cell carcinoma (SCC) that can be effectively challenged by adoptive cytotoxic T cell transfer (ACT)-based immunotherapy. Using single-cell RNA sequencing (RNA-seq) and lineage tracing, we found that transforming growth factor beta (TGF-beta)-responding tSCs are superior at resisting ACT and form the root of tumor relapse. Probing mechanism, we discovered that during malignancy, tSCs selectively acquire CD80, a surface ligand previously identified on immune cells. Moreover, upon engaging cytotoxic T lymphocyte antigen-4 (CTLA4), CD80-expressing tSCs directly dampen cytotoxic T cell activity. Conversely, upon CTLA4- or TGF-beta-blocking immunotherapies or Cd80 ablation, tSCs become vulnerable, diminishing tumor relapse after ACT treatment. Our findings place tSCs at the crux of how immune checkpoint pathways are activated.

Author Info: (1) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. (2) Robin Che

Author Info: (1) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. (2) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. (3) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. (4) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. (5) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. (6) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. (7) Department of Surgery, Laboratory of Epithelial Cancer Biology and Molecular Cytology Core Facility, Memorial Sloan Kettering Cancer Center, New York, New York, NY 10065, USA. (8) Department of Dermatology, University of California, San Francisco, San Francisco, CA 94143, USA. (9) Robin Chemers Neustein Laboratory of Mammalian Cell Biology and Development, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA. Electronic address: fuchslb@rockefeller.edu.