Naturally occurring single-nucleotide variants (SNVs) in T cells can affect their functionality, to the point of clinical pathology, and so Walsh and Shah et al. hypothesized that precise base editing of essential T cell genes could uncover variants with superior T cell activity. High-throughput adenine base editor screens in human T cells generated a library of SNVs targeting relevant loci in 102 genes critical to T cell function. Several variants (particularly in PIK3CD) improved T cell proliferation, activation, and cytokine production. Editing these SNVs into TCR T cells or CD19 CAR T cells boosted antigen-specific cytokine production and tumor cell killing.
Contributed by Alex Najibi
ABSTRACT: Single-nucleotide variants (SNVs) in key T cell genes can drive clinical pathologies and could be repurposed to improve cellular cancer immunotherapies. Here, we perform massively parallel base-editing screens to generate thousands of variants at gene loci annotated with known or potential clinical relevance. We discover a broad landscape of putative gain-of-function (GOF) and loss-of-function (LOF) mutations, including in PIK3CD and the gene encoding its regulatory subunit, PIK3R1, LCK, SOS1, AKT1 and RHOA. Base editing of PIK3CD and PIK3R1 variants in T cells with an engineered T cell receptor specific to a melanoma epitope or in different generations of CD19 chimeric antigen receptor (CAR) T cells demonstrates that discovered GOF variants, but not LOF or silent mutation controls, enhanced signaling, cytokine production and lysis of cognate melanoma and leukemia cell models, respectively. Additionally, we show that generations of CD19 CAR T cells engineered with PIK3CD GOF mutations demonstrate enhanced antigen-specific signaling, cytokine production and leukemia cell killing, including when benchmarked against other recent strategies.
Author Info: (1) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medic
Author Info: (1) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (2) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (3) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. (4) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. (5) Department of Systems Biology, Columbia University Irving Medical Center, New York, NY, USA. (6) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (7) Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. Department of Genetics and Development, Columbia University Medical Center, New York, NY, USA. (8) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (9) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (10) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (11) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (12) Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (13) Department of Systems Biology, Columbia University Irving Medical Center, New York, NY, USA. (14) Department of Pediatrics, Columbia University Irving Medical Center, New York, NY, USA. (15) Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. Department of Genetics and Development, Columbia University Medical Center, New York, NY, USA. (16) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. Columbia Center for Translational Immunology, New York, NY, USA. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. (17) Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA. bi2175@cumc.columbia.edu. Department of Medicine, Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, NY, USA. bi2175@cumc.columbia.edu. Columbia Center for Translational Immunology, New York, NY, USA. bi2175@cumc.columbia.edu. Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY, USA. bi2175@cumc.columbia.edu. Department of Systems Biology, Columbia University Irving Medical Center, New York, NY, USA. bi2175@cumc.columbia.edu.
