CARM1-mediated hypoxanthine-enriched exosome biogenesis rewires inosine metabolism of CD8+ T cells and impairs their antitumor ability

Cancer cells engage in complex interactions with tumor infiltrating immune cells to overcome barriers to tumor progression. Targeting biogenesis of tumor-derived exosomes is a promising strategy for disrupting intracellular communication. Herein, we identify co-activator-associated arginine methyltransferase 1 (CARM1) as a key regulator of tumor-derived exosome biogenesis and metabolite sorting, thereby inhibiting CD8+ T cell-mediated antitumor immunity. Genetic ablation of CARM1 in breast cancer cells impairs the secretion of immunosuppressive exosomes, leading to enhanced CD8+ T cell infiltration, proliferation, and effector function in tumors. Mechanistically, CARM1 methylates apoptosis-linked gene-2 interacting protein X (ALIX) at Arg757, facilitating its interaction with endosomal sorting complex required transport (ESCRT) components, and promoting the biogenesis of tetraspanin-enriched exosomes. CARM1-dependent ALIX methylation enables selective packaging hypoxanthine into exosomes through direct binding to ALIX at the F676 pocket. Excess hypoxanthine disrupts inosine metabolism in the activated CD8+ T cells, inhibiting glycolysis, nucleotide synthesis, and effector cytokine production. Co-administration of CARM1 inhibitor with inosine significantly increases tumor infiltrating CD8+ T cell cytotoxicity, reduces exhaustion markers programmed cell death 1 and T cell immunoglobulin mucin 3 (PD-1+TIM-3+) expression and suppresses tumor growth. Collectively, the CARM1-ALIX-hypoxanthine axis functions as immunosuppressive mechanism in tumor growth, and targeting CARM1 combined with inosine supplement may represent a novel therapeutic regimen for breast cancer therapy.