Therapeutic Poxviruses Induce Immunostimulating Extracellular Vesicles with Potent Antitumor Activity

Poxvirus-based vectors are a promising platform for cancer immunotherapy, designed to deliver immunostimulatory molecules and tumor-specific antigens. While viral infections are known to influence extracellular vesicle (EV) biogenesis, the role of therapeutic poxviral vectors in modulating EV secretion by immune cells and its impact on therapeutic efficacy remains largely unexplored. Understanding this interaction could unveil novel mechanisms of action for viral immunotherapies and potentially lead to the development of cell-free therapeutic strategies. This study addresses the gap in knowledge regarding how engineered poxviruses influence EV content and function in an antitumor context.

Researchers investigated how poxviruses, specifically Modified Vaccinia Ankara (MVA), influence small EV (sEV) secretion from peripheral blood mononuclear cells (PBMCs). They engineered an MVA vector to express therapeutic payloads: the model ovalbumin-derived peptide SIINFEKL (presented by MHC I), interleukin-12 (IL-12), and CD40 ligand (CD40L). sEVs were isolated from infected PBMCs using different methods. The study assessed in vitro SIINFEKL-specific CD8+ T cell stimulation. Crucially, they performed intravenous injections of these sEVs, and also the soluble secretome, into E.G7-OVA lymphoma-bearing mice to evaluate in vivo antitumor activity, comparing efficacy to direct virus administration.

Poxviruses, including MVA, were found to stimulate the secretion of small EVs (sEVs) from PBMCs, with these sEVs containing viral proteins and immune-related signatures. The engineered MVA vector successfully transferred virus-encoded therapeutic payloads, such as the OVA-derived peptide SIINFEKL (presented by MHC I), IL-12, and CD40L, into these sEVs. Depending on the isolation method, these sEVs stimulated SIINFEKL-specific CD8+ T cells with varying efficiencies in vitro.