Depletion of IL-10 in CAR-NK cells augments reprogramming of the tumor microenvironment and ameliorates therapeutic efficacy.

The ErbB2 (HER2)-specific CAR-engineered natural killer (NK) cell line NK-92/5.28.z is under investigation in a phase I clinical trial in glioblastoma patients. In preclinical studies, these cells demonstrated potent CAR-mediated cytotoxicity and stimulated endogenous antitumor immunity in immunocompetent animals. Pro-inflammatory cytokines can contribute to the CAR-NK cells' immunomodulatory activity, but this may be attenuated by immunosuppressive IL-10 that is also produced in substantial amounts by activated NK-92/5.28.z cells. To prevent IL-10 secretion, we modified the CAR-NK cells to express an intracellular anti-IL-10 antibody, which trapped IL-10 within the endoplasmic reticulum. This did not affect proliferation, phenotype, or cytotoxicity of the resulting NK-92/5.28.z/anti-IL10ER cells but strengthened their ability to mediate maturation of co-cultured dendritic cells and prevented M2-polarization of co-cultured macrophages induced by unmodified CAR-NK cells. In a syngeneic murine glioblastoma model, NK-92/5.28.z/anti-IL10ER cells exhibited enhanced antitumor activity and favored a pro-inflammatory tumor microenvironment characterized by reduced infiltration of IL-10-responsive immunosuppressive cell types. Our findings demonstrate that inhibiting IL-10 secretion improves the therapeutic potential of CAR-engineered NK-92 cells, suggesting this approach as a promising avenue for clinical translation.