Abstract Background Viral-based immunotherapy has the potential to overcome resistance to immune checkpoint blockade (ICB) and to fill the unmet needs of many cancer patients. Oncolytic viruses (OVs) are defined as engineered or naturally occurring viruses that selectively replicate in and kill cancer cells. OVs also induce antitumor immunity. The purpose of this study is to compare the antitumor effects of live OV-GM expressing murine granulocyte-macrophage colony-stimulating factor (mGM-CSF) versus inactivated OV-GM and elucidate the underlying immunological mechanisms. Methods In this study, we engineered a replication-competent, oncolytic vaccinia virus (OV-GM) by inserting a murine GM-CSF gene into the thymidine kinase (TK) locus of a mutant vaccinia E3LΔ83N, which lacks the Z-DNA-binding domain of vaccinia virulence factor E3. We compared the antitumor effects of intratumoral (IT) delivery of live OV-GM vs. heat-inactivated OV-GM (heat-iOV-GM) in a murine B16-F10 melanoma bilateral implantation model. Results Heat-iOV-GM infection of dendritic cells (DCs) and tumor cells in vitro induces type I IFN and pro inflammatory cytokines and chemokines, whereas live OV-GM does not. IT live OV-GM is less effective in generating systemic antitumor immunity compared with heat-iOV-GM. Similar to heat-iOV-GM, the antitumor effects of live OV-GM also require Batf3-dependent CD103+ dendritic cells. IT heat-iOV-GM induces higher numbers of infiltrating activated CD8+ and CD4+ T cells as well as higher levels of type I IFN, proinflammatory cytokines, and chemokines in the distant non-injected tumors, which is dependent on CD8+ T cells. When combined with systemic delivery of ICB, IT heat-iOV-GM is more effective in eradicating tumors compared with live OV-GM. Conclusions Tumor lysis induced by the replication of oncolytic DNA viruses has a limited effect on the generation of systemic antitumor immunity. The activation of Batf3-dependent CD103+ DCs is critical for antitumor effects induced by both live OV-GM and heat-iOV-GM. Heat-iOV-GM is more potent than live OV-GM in the induction of innate and adaptive immunity in both the injected and distant non-injected tumors. We propose that evaluations of both innate and adaptive immunity induced by IT oncolytic viral immunotherapy at injected and non-injected tumors should be included as potential biomarkers.

Elucidating Mechanisms of Antitumor Immunity Mediated by Live Oncolytic Vaccinia and Heat-Inactivated Vaccinia