Restricting efferocytosis pathway to CD91 via phosphatidylserine-targeting chimeric protein augments antitumor immune responses.

Phosphatidylserine is a representative eat-me signal on the outer membrane of apoptotic cells promoting phagocytosis/efferocytosis. It is recognized by various molecules, such as MFG-E8, which produce an immune-tolerizing environment. In order to develop immunological treatments targeting this mechanism, we designed a novel chimeric protein that consists of a phosphatidylserine-recognition domain from MFG-E8 and receptor-associated protein (RAP), which is a ligand of CD91 known as an immunostimulatory phagocytic receptor. This chimeric protein, PStRAP, can theoretically link phosphatidylserine-exposed apoptotic cells to phagocytes not through authentic immunosuppression but through an immunostimulative route via CD91. The inoculation of dying tumor cells expressing PStRAP, which served as an immunogen, induced potent adaptive immune responses against parental tumor cells in murine tumor models. The presence of PStRAP increased the phagocytosis of apoptotic cells in vitro and cross-priming to cytotoxic T-lymphocytes in vivo compared with the absence of it. Furthermore, PStRAP protein was supplied systemically by intravenous injection of PStRAP mRNA contained in lipid nanoparticles and along with cytotoxic drug to treat tumor-bearing mice. PStRAP significantly enhanced antitumor effects of cytotoxic drug alone. These results suggest that PStRAP can induce CD91-mediated phagocytosis of the apoptotic tumor cells and initiate meaningful adaptive immune responses against the tumor cells.