Chimeric antigen receptor (CAR)-natural killer (NK) cell therapies hold promise for solid tumours but remain limited because of poor tumour infiltration, persistence and resistance in the tumour microenvironment1–4. Here, to identify gain-of-function targets that enhance CAR-NK cell efficacy, we performed an unbiased in vivo CRISPR activation screen followed by a barcoded targeted in vivo open reading frame screen in primary human CAR-NK cells. We identified and comprehensively validated OR7A10, a G protein-coupled receptor (GPCR), as the top candidate. Engineering CAR-NK cells with OR7A10 cDNA (a CRISPR-independent method with a simple manufacturing strategy) enhanced their proliferation, activation, degranulation, cytokine production, death ligand expression, chemokine receptor expression, cytotoxicity, persistence, metabolic fitness and tumour microenvironment resistance. Moreover, exhaustion in primary human NK cells derived from multiple peripheral blood and cord blood donors was reduced. OR7A10 gain-of-function CAR-NK cells displayed strong in vivo efficacy across multiple solid tumour models. For example, 100% complete response with long-term tumour control and survival benefit in an orthotopic breast cancer mouse model were achieved. These findings establish OR7A10-engineered CAR-NK cells as a highly potent and scalable off-the-shelf therapeutic for solid tumours.
