Bräunlein et al. identified three neoantigens from a patient with metastatic melanoma using immunopeptidomics (SYTL4S363F and NCAPG2P333L) or in silico prediction (KIF2CP13L), and characterized the immune response against the selected neoantigens. Seven T cell receptors (TCRs) that recognized these antigens were isolated following in vitro stimulation and expansion. TCR-transgenic CD8+ T cells showed high antigen specificity, low cross-reactivity, and in vivo antitumor potential. TCRs with lower functional avidity occurred at high frequencies; persisted long-term within tumor tissues, lymph nodes, and blood samples; and showed at least equal antitumor control in vivo.
Contributed by Shishir Pant
BACKGROUND: Neoantigens derived from somatic mutations correlate with therapeutic responses mediated by treatment with immune checkpoint inhibitors. Neoantigens are therefore highly attractive targets for the development of therapeutic approaches in personalized medicine, although many aspects of their quality and associated immune responses are not yet well understood. In a case study of metastatic malignant melanoma, we aimed to perform an in-depth characterization of neoantigens and respective T-cell responses in the context of immune checkpoint modulation. METHODS: Three neoantigens, which we identified either by immunopeptidomics or in silico prediction, were investigated using binding affinity analyses and structural simulations. We isolated seven T-cell receptors (TCRs) from the patient's immune repertoire recognizing these antigens. TCRs were compared in vitro by multiparametric analyses including functional avidity, multicytokine secretion, and cross-reactivity screenings. A xenograft mouse model served to study in vivo functionality of selected TCRs. We investigated the patient's TCR repertoire in blood and different tumor-related tissues over 3_years using TCR beta deep sequencing. RESULTS: Selected mutated peptide ligands with proven immunogenicity showed similar binding affinities to the human leukocyte antigen complex and comparable disparity to their wild-type counterparts in molecular dynamic simulations. Nevertheless, isolated TCRs recognizing these antigens demonstrated distinct patterns in functionality and frequency. TCRs with lower functional avidity showed at least equal antitumor immune responses in vivo. Moreover, they occurred at high frequencies and particularly demonstrated long-term persistence within tumor tissues, lymph nodes and various blood samples associated with a reduced activation pattern on primary in vitro stimulation. CONCLUSIONS: We performed a so far unique fine characterization of neoantigen-specific T-cell responses revealing defined reactivity patterns of neoantigen-specific TCRs. Our data highlight qualitative differences of these TCRs associated with function and longevity of respective T cells. Such features need to be considered for further optimization of neoantigen targeting including adoptive T-cell therapies using TCR-transgenic T cells.
Author Info: (1) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (2) Klinik und Poliklinik fr Innere Medizin III, Klini
Author Info: (1) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (2) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (3) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (4) Laboratory of Protein Biochemistry, Institute for Chemistry and Biochemistry, Freie Universitt Berlin, Berlin, Germany. (5) Institute of Molecular Oncology and Functional Genomics, TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Center for Translational Cancer Research (TranslaTUM), TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (6) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (7) TUM School of Life Sciences and Center for Integrated Protein Science Munich, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (8) Institute of Molecular Oncology and Functional Genomics, TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Center for Translational Cancer Research (TranslaTUM), TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Department of Medicine II, Klinikum rechts der Isar, TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (9) Institute of Molecular Oncology and Functional Genomics, TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Center for Translational Cancer Research (TranslaTUM), TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (10) Laboratory of Protein Biochemistry, Institute for Chemistry and Biochemistry, Freie Universitt Berlin, Berlin, Germany. (11) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (12) Institute for Medical Microbiology Immunology and Hygiene, Technische Universitt Mnchen, Mnchen, Germany. (13) Institute of Pathology, School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. MRI-TUM-Biobank at the Institute of Pathology, School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (14) Institute of Pathology, School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Core Facility Experimental Pathology, School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. German Cancer Consortium (DKTK), partner-site Munich, and German Cancer Research Center (DKFZ) Heidelberg, Heidelberg, Germany. (15) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (16) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (17) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. German Cancer Consortium (DKTK), partner-site Munich, and German Cancer Research Center (DKFZ) Heidelberg, Heidelberg, Germany. (18) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Center for Translational Cancer Research (TranslaTUM), TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. German Cancer Consortium (DKTK), partner-site Munich, and German Cancer Research Center (DKFZ) Heidelberg, Heidelberg, Germany. (19) Institute of Pathology, School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. MRI-TUM-Biobank at the Institute of Pathology, School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Core Facility Experimental Pathology, School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. German Cancer Consortium (DKTK), partner-site Munich, and German Cancer Research Center (DKFZ) Heidelberg, Heidelberg, Germany. (20) Institute for Medical Microbiology Immunology and Hygiene, Technische Universitt Mnchen, Mnchen, Germany. (21) Institute of Molecular Oncology and Functional Genomics, TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Center for Translational Cancer Research (TranslaTUM), TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. Department of Medicine II, Klinikum rechts der Isar, TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. German Cancer Consortium (DKTK), partner-site Munich, and German Cancer Research Center (DKFZ) Heidelberg, Heidelberg, Germany. (22) Laboratory of Protein Biochemistry, Institute for Chemistry and Biochemistry, Freie Universitt Berlin, Berlin, Germany. (23) TUM School of Life Sciences and Center for Integrated Protein Science Munich, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. (24) Klinik und Poliklinik fr Innere Medizin III, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany angela.krackhardt@tum.de. Center for Translational Cancer Research (TranslaTUM), TUM School of Medicine, Klinikum rechts der Isar der Technischen Universitt Mnchen, Mnchen, Germany. German Cancer Consortium (DKTK), partner-site Munich, and German Cancer Research Center (DKFZ) Heidelberg, Heidelberg, Germany.
