Immune modulation of innate and adaptive responses restores immune surveillance and establishes anti-tumor immunological memory.

Current immunotherapies have proven effective in strengthening anti-tumor immune responses, but constant opposing signals from tumor cells and the surrounding microenvironment eventually lead to immune escape. We hypothesized that in situ release of antigens and regulation of both the innate and adaptive arms of the immune system would provide a robust and long-term anti-tumor effect by creating immunological memory against tumors. To achieve this, we developed CARG-2020, a genetically modified virus-like vesicle (VLV) that is a self-amplifying RNA with oncolytic capacity and encodes immune regulatory genes. CARG-2020 carries three immune modulators: 1) the pleiotropic antitumor cytokine, IL12, in which the subunits (p35 and p40) are tethered together; 2) the extracellular domain (ECD) of the pro-tumor IL17RA, which serves as a dominant-negative antagonist; and 3) a shRNA targeting PD-L1. Using a mouse model of ovarian cancer, we demonstrated the oncolytic effect and immune-modulatory capacities of CARG-2020. By enhancing IL12 and blocking IL17 and PD-L1, CARG-2020 successfully reactivated immune surveillance by promoting M1, instead of M2, macrophage differentiation, inhibiting MDSC expansion and establishing a potent CD8+ T cell-mediated anti-tumoral response. Furthermore, we demonstrated that this therapeutic approach provided tumor-specific and long-term protection against the establishment of new tumors. Our results provide rationale for the further development of this platform as a therapeutic modality for ovarian cancer patients to enhance anti-tumor responses and prevent recurrence.