Genome editing or small molecule inhibition of KMT5A in CAR-T cells enhances antitumor immunity

Background Adoptive T-cell therapy has emerged as a promising therapeutic strategy for cancer treatment. However, clinical challenges persist, including the limited ability of CD8+T cells to infiltrate solid tumors and efficiently eliminate tumor cells. Given the critical role of epigenetic mechanisms in antitumor immunity, targeting epigenetic regulators represents a critical step toward optimizing adoptive T-cell therapies for solid tumors. Methods To investigate the role of KMT5A in CD8+Tcell function, we employed CRISPR screening to identify KMT5A as a negative regulator. We then genetically deleted KMT5A in human CD8+T cells and systematically evaluated its impact on the antitumor efficacy of chimeric antigen receptor (CAR)-T cells using xenograft models. Furthermore, we used the small-molecule inhibitor UNC0379 to pharmacologically inhibit KMT5A, meticulously assessing the consequent effects on CAR-T cell activation, cytotoxicity, and antitumor activity. Results We report that lysine methyltransferase KMT5A acts as a negative regulator of CD8+Tcell function, identified via CRISPR screening. KMT5A deletion in human CD8+T cells significantly enhances the antitumor efficacy of CAR CD8+T cells in xenograft models. In vitro immunophenotyping reveals that KMT5A deletion improves effector functions, cytokine secretion, and early activation of CD8+T cells. Mechanistically, KMT5A depletion increases the expression and chromatin accessibility of multiple effector-related genes in CD8+T cells. KMT5A-mediated histone H4 modifications and chromatin remodeling suppress CD8+Tcell effector functions, partially via inhibition of the transcription factor SP1. Notably, pharmacological inhibition of KMT5A using the small-molecule inhibitor UNC0379 enhanced activation, cytotoxicity, and antitumor activity in human CD8+T cells—a novel finding in this study. Conclusions Our findings establish KMT5A as an epigenetic regulator that impairs CD8+Tcell function. These findings demonstrate that genetic or pharmacological (eg, UNC0379) targeting of KMT5A in CD8+T cells represents a viable therapeutic strategy to augment effector functions and improve adoptive T-cell therapies, particularly CAR-T cells, for solid tumors.