Engineering with EP2/EP4 knockout and IL-15 transpresentation renders stem cell-derived NK cells self-persistent and...

Adoptive Natural Killer (NK) cell transfer is a promising therapy for the treatment of cancer. We developed a GMP-compliant protocol to generate NK cells from hematopoietic stem and progenitor cells (HSPC-NK). However, the immunosuppressive tumor microenvironment - especially prostaglandin E2 (PGE2) signaling via E-prostanoid receptors EP2 and EP4- and lack of persistence of allogeneic NK cells hinders HSPC-NK therapeutic efficacy. Here, we enhanced the therapeutic efficacy of HSPC-NK cells by interfering with both EP2 and EP4 with antagonists or CRISPR-KO, and through engineering with IL-15 tethered to membrane-bound IL-15Rα (tIL15). We found that CRISPR interference of EP2 and EP4 fully rescued proliferation and potency of HSPC-NK cells under PGE2 rich conditions. In addition, combined EP2/EP4 KO and tIL15 engineering renders HSPC-NK cells resistant to PGE2, while improving their survival and functionality against different tumor targets in cytokine-deprived environments. Altogether, our EP2/EP4-KO tIL15-HSPC-NK cells pose a promising therapy for cancer.