The efficacy of adoptive T cell therapies for cancer treatment can be limited by suppressive signals from both extrinsic factors and intrinsic inhibitory checkpoints1,2. Targeted gene editing has the potential to overcome these limitations and enhance T cell therapeutic function3–10. Here we performed multiple genome-wide CRISPR knock-out screens under different immunosuppressive conditions to identify genes that can be targeted to prevent T cell dysfunction. These screens converged on RASA2, a RAS GTPase-activating protein (RasGAP) that we identify as a signalling checkpoint in human T cells, which is downregulated upon acute T cell receptor stimulation and can increase gradually with chronic antigen exposure. RASA2 ablation enhanced MAPK signalling and chimeric antigen receptor (CAR) T cell cytolytic activity in response to target antigen. Repeated tumour antigen stimulations in vitro revealed that RASA2-deficient T cells show increased activation, cytokine production and metabolic activity compared with control cells, and show a marked advantage in persistent cancer cell killing. RASA2-knockout CAR T cells had a competitive fitness advantage over control cells in the bone marrow in a mouse model of leukaemia. Ablation of RASA2 in multiple preclinical models of T cell receptor and CAR T cell therapies prolonged survival in mice xenografted with either liquid or solid tumours. Together, our findings highlight RASA2 as a promising target to enhance both persistence and effector function in T cell therapies for cancer treatment. Genome-wide CRISPR screens, biochemical studies and animal models show that RASA2 has a key role in regulating T cell function and has potential as a genetic target for enhancing anti-tumour immunity.
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T cells used in immunotherapy treatments can become exhausted by the task of fighting cancer cells or shut down when they enter tumours. However, a set of CRISPR screens allowed researchers to deactivate each gene in the genome, one at a time, in a pool of human T cells and the team found a handful of candidates that could make T cells resistant to key aspects of the immunosuppressive microenvironment often present in tumours. The researchers were particularly intrigued by a gene called RASA2, as it had never been associated with immune cell function before. The team created T cells with the RASA2 gene knocked out. They then subjected these T cells to various "stress tests" by repeatedly exposing them to cancer cells and models of the tumour microenvironment. They compared the performance of these cells to the original therapeutic T cells that still contained a functional RASA2 gene. Long after the original cells had lost their cancer-fighting abilities, the cells with knocked-out RASA2 remained remarkably tireless. The researchers thus made the therapeutic cells more resistant. This discovery could help overcome a major factor limiting the success of these promising therapies in the fight against solid and liquid tumours.