A Novel Engineered AAV-based Neoantigen Vaccine in Combination with Radiotherapy Eradicates Tumors.

The potency of tumor-specific antigen (TSA) vaccines, such as neoantigen-based cancer vaccines, can be compromised by host immune checkpoint inhibitory mechanisms, such as programmed cell death protein 1 (PD-1)/programmed death ligand 1 (PD-L1), that attenuate neoantigen presentation on dendritic cells (DCs) and hinder T cell-mediated cytotoxicity. To overcome PD-1/PD-L1 inhibition in DCs, we developed a novel adeno-associated virus (meAAV) neoantigen vaccine, modified with TLR9 inhibitory fragments, PD-1 trap, and PD-L1 miRNA, which extend the persistence of meAAV and activate neoantigen-specific T-cell responses in immune-competent colorectal and breast cancer murine models. Moreover, we found that in combination with radiotherapy, the meAAV-based neoantigen cancer vaccine not only elicited higher antigen presentation ability, but also maintained neoantigen-specific cytotoxic T lymphocyte (CTL) responses. These functional PD-1 traps and PD-L1 miRNAs overcome host PD1/PD-L1 inhibitory mechanisms and boost the therapeutic efficacy of radiotherapy. More importantly, combined radiotherapy and meAAV neoantigen cancer vaccines significantly enhanced neoantigen-specific CTL responses, increased CTL infiltration in tumor microenvironment, and decreased tumor-associated immunosuppression. This process led to the complete elimination of colorectal cancer and delayed tumor growth of breast cancer in tumor-bearing mice. Taken together, our results demonstrated a novel strategy that combines neoantigen cancer vaccine and radiotherapy to increase the therapeutic efficacy against colorectal and breast cancers.