By screening ~12,000 human ORFs overexpressed in primary human T cells, and using a novel method of single-cell sequencing coupled with ORF capture, Legut et al. identified genes spanning diverse functions that increased CD4+/CD8+ T cell activation via CD3/CD28. T cell expression of the top-ranked ORF, Lymphotoxin-β receptor, typically expressed by stromal and myeloid cells, induced NF-κB-dependent transcriptional and epigenomic changes to boost exhaustion/apoptosis resistance and proinflammatory cytokine secretion. Expression of top-ranked ORFs in CD19-CAR T cells and tumor-reactive γδ T cells improved their antigen-specific responses.
Contributed by Paula Hochman
ABSTRACT: The engineering of autologous patient T cells for adoptive cell therapies has revolutionized the treatment of several types of cancer1. However, further improvements are needed to increase response and cure rates. CRISPR-based loss-of-function screens have been limited to negative regulators of T cell functions2-4 and raise safety concerns owing to the permanent modification of the genome. Here we identify positive regulators of T cell functions through overexpression of around 12,000 barcoded human open reading frames (ORFs). The top-ranked genes increased the proliferation and activation of primary human CD4+ and CD8+ T cells and their secretion of key cytokines such as interleukin-2 and interferon-γ. In addition, we developed the single-cell genomics method OverCITE-seq for high-throughput quantification of the transcriptome and surface antigens in ORF-engineered T cells. The top-ranked ORF-lymphotoxin-β receptor (LTBR)-is typically expressed in myeloid cells but absent in lymphocytes. When overexpressed in T cells, LTBR induced profound transcriptional and epigenomic remodelling, leading to increased T cell effector functions and resistance to exhaustion in chronic stimulation settings through constitutive activation of the canonical NF-κB pathway. LTBR and other highly ranked genes improved the antigen-specific responses of chimeric antigen receptor T cells and γδ T cells, highlighting their potential for future cancer-agnostic therapies5. Our results provide several strategies for improving next-generation T cell therapies by the induction of synthetic cell programmes.
Author Info: (1) New York Genome Center, New York, NY, USA. mateusz.legut@gmail.com. Department of Biology, New York University, New York, NY, USA. mateusz.legut@gmail.com. Department of Neuros
Author Info: (1) New York Genome Center, New York, NY, USA. mateusz.legut@gmail.com. Department of Biology, New York University, New York, NY, USA. mateusz.legut@gmail.com. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. mateusz.legut@gmail.com. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. mateusz.legut@gmail.com. (2) New York Genome Center, New York, NY, USA. Department of Biology, New York University, New York, NY, USA. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. (3) New York Genome Center, New York, NY, USA. Department of Biology, New York University, New York, NY, USA. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. (4) New York Genome Center, New York, NY, USA. Department of Biology, New York University, New York, NY, USA. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. Beam Tx, Cambridge, MA, USA. (5) New York Genome Center, New York, NY, USA. Department of Biology, New York University, New York, NY, USA. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. (6) New York Genome Center, New York, NY, USA. Department of Biology, New York University, New York, NY, USA. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. (7) New York Genome Center, New York, NY, USA. Department of Biology, New York University, New York, NY, USA. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. (8) New York Genome Center, New York, NY, USA. Department of Biology, New York University, New York, NY, USA. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. (9) Technology Innovation Lab, New York Genome Center, New York, NY, USA. Immunai, New York, NY, USA. (10) Technology Innovation Lab, New York Genome Center, New York, NY, USA. (11) Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, USA. (12) Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. (13) Technology Innovation Lab, New York Genome Center, New York, NY, USA. Immunai, New York, NY, USA. (14) New York Genome Center, New York, NY, USA. neville@sanjanalab.org. Department of Biology, New York University, New York, NY, USA. neville@sanjanalab.org. Department of Neuroscience and Physiology, New York University School of Medicine, New York, NY, USA. neville@sanjanalab.org. Perlmutter Cancer Center, New York University School of Medicine, New York, NY, USA. neville@sanjanalab.org.
