834 Rational design of recombinant modified vaccinia virus Ankara for cancer immunotherapy

Abstract

Poxviruses are large cytoplasmic DNA viruses and modified vaccinia virus Ankara (MVA) is a highly attenuated vaccinia strain. Intratumoral (IT) delivery of inactivated MVA (iMVA) eradicates injected tumors and generates systemic antitumor immunity, which synergizes with immune chekpoint blockade antibodies (Dai et al. Science Immunology, 2017). The antitumor effects induced by iMVA requires CD8+ T cells and CD103+ dendritic cells (DCs), and is dependent on cGAS/STING. We hypothesize that MVA encodes viral inhibitor(s) of the cGAS/STING pathway and therefore deleting such inhibitor(s) from MVA would improve the antitumor effects of the virus. We conducted a systematic screening of vaccinia early genes for potential inhibitors of the cGAS/STING pathway and identified several candidates. Among them, the vaccinia E5 protein encoded by the E5R gene has a dominant inhibitory function. Infection of bone marrow-derived DCs (BMDCs) with MVAΔE5R induces 80-fold higher IFN-β secretion compared with MVA and 10-fold higher IFN-β secertion compared with iMVA. IT MVAΔE5R results in strong antitumor effects that is dependent on the cGAS/STING/IFNAR pathway. MVA also expresses another virulence factor C7, which is encoded by the C7L gene. C7 binds to the transcription factors IRF3 and Stat2 and thereby inhibits type I IFN production and IFNAR signaling. IT recombinant MVA with deletion of both C7L and E5R, and with expression of two transgenes human Flt3L, a growth factor for DCs, and murine OX40L, a costimulatory factor for T cells, results in enhanced antitumor immunity compared with iMVA in murine B16-F10 melanoma model and other tumor models. Our results demonstrate the feasibility of dramatically improving MVA-based cancer immunotherapeutics through rational design.