Antigen-dependent interplay of formulation, systemic innate responses, and antibody responses to multi-component replicon RNA vaccination.

Replicon RNA (repRNA) vaccines are a transformative tool in combating infectious diseases, eliciting robust immune responses at lower doses. However, systemic inflammatory responses to lipid nanoparticle (LNP)-formulated RNA vaccines may compromise safety and efficacy. repRNA vaccines delivered with cationic nanocarriers like LION offer a promising alternative by localizing innate immune responses required to induce a robust adaptive response at the site of injection. Here, we show, in pigtail macaques, that a multi-component repRNA vaccine against two viral pathogens formulated with LION induces antigen-specific antibody (Ab) responses against enterovirus D68 (EV-D68) while reducing systemic inflammatory responses compared to the same vaccine formulated with LNP. Notably, early systemic interferon (IFN) levels inversely correlate with EV-D68 binding and neutralizing Ab titers, indicating that excessive systemic innate immune responses can impair RNA vaccine immunogenicity. These findings suggest that LION-mediated delivery offers a safer and more effective platform for RNA vaccines than conventional LNP formulations. By mitigating systemic cytokine induction, LION enhances vaccine immunogenicity and safety-key considerations for optimizing RNA vaccine design.