MMP3 overexpression enhances CAR-T cell infiltration and antitumor activity in a CAF-enriched solid tumor model

BACKGROUND: Chimeric antigen receptor (CAR) T cell therapy has shown remarkable success in hematologic malignancies but faces substantial challenges in solid tumors. One of the main obstacles is the extracellular matrix (ECM), which serves as the physical barrier that hinders T cell infiltration into tumor tissues. METHODS: We engineered CAR-T cells targeting mesothelin or B7H3 to co-express matrix metalloproteinase-3 (MMP3). We evaluated the effects of MMP3 overexpression on CAR-T cell proliferation, activation, cytotoxicity, and tumor infiltration using both in vitro Matrigel-based assays and in vivo xenograft and syngeneic models enriched with cancer-associated fibroblasts (CAFs). RESULTS: MMP3 overexpression did not impair CAR-T cell proliferation, activation, or cytotoxicity. However, it significantly enhanced their capacity to invade through ECM and improved tumor cell killing in vitro. In CAF-enriched xenograft models, MMP3-engineered CAR-T cells demonstrated superior tumor infiltration, expansion, and antitumor activity. Notably, MMP3 overexpression rescued the function of B7H3 CAR-T cells in the stringent CAF-enriched tumor microenvironment, while conventional CAR-T cells showed limited activity. Importantly, MMP3 overexpression also conferred potent antitumor activity in an immunocompetent mouse model, underscoring its therapeutic benefit in a more physiologically and clinically related setting. CONCLUSIONS: These findings suggest that MMP3 engineering is a simple yet effective strategy to overcome stromal barriers and enhance the efficacy of CAR-T cell therapy in solid tumors.