Heat-inactivated modified vaccinia virus Ankara boosts Th1-biased cellular and humoral immune responses as a vaccine adjuvant by activating the STING-mediated cytosolic DNA-sensing pathway

Protein or peptide-based subunit vaccines are promising platforms for combating human cancers and infectious diseases. However, one primary concern regarding subunit vaccines is the relatively weak immune responses induced by proteins or peptides. Therefore, developing novel and effective vaccine adjuvants is critical for the success of subunit vaccines. Modified vaccinia virus (MVA) is a safe and effective vaccine against smallpox and monkeypox. In this study, we explored the potential of heat-inactivated MVA (heat-iMVA; by incubating MVA at 37C for one hour) as a novel vaccine adjuvant. We previously reported that heat-iMVA enters dendritic cells to activate the cytosolic DNA-sensing pathway, leading to the production of type I interferon and proinflammatory cytokines and chemokines. Heat-iMVA infection does not express viral proteins, which include inhibitors of innate immune sensing pathways. Here, we show that co-administration of protein- or peptide-based immunogens with heat-iMVA dramatically enhances Th1-biased cellular and humoral immune responses. This adjuvant effect of heat-iMVA is dependent on the Stimulator of Interferon Genes (STING), and the antigen-specific CD8+ T cell response requires Batf3-dependent CD103+/CD8a+ dendritic cells (DCs). Heat-iMVA infection of bone marrow-derived DCs (BMDCs) promoted antigen cross-presentation, whereas live MVA infection did not. RNA-seq analyses revealed that heat-iMVA is a more potent activator of the STING pathway than live MVA, leading to more robust induction of type I interferon (IFN) pathway and IFN-stimulated genes in BMDCs. Additionally, combining tumor neoantigen peptides with heat-iMVA delayed tumor growth and extended the median survival and in a therapeutic vaccination model. Furthermore, co-administration of SARS-CoV-2 spike protein and heat-iMVA induced higher titers of spike-specific antibodies with potent neutralizing activities than spike protein immunization alone. Taken together, we provide evidence to support that heat-iMVA is a safe and potent vaccine adjuvant for cancer and infectious disease applications.