A macrophage-targeted bivalent saRNA vaccine protects against ocular and cutaneous HSV-1 infection in mice

Herpes simplex virus type 1 (HSV-1) infects over 1.5 million individuals annually, causing herpes simplex keratitis (HSK) with irreversible vision loss, as well as skin and mucosal infections. The virus establishes lifelong latency in the nervous system, rendering current therapies ineffective for eliminating latent reservoirs or preventing recurrence. To address this challenge, we developed a novel bivalent self-amplifying RNA (saRNA) vaccine, termed MTL-bivalent-IL12, which incorporates saRNAs encoding HSV-1 glycoproteins gD and gB, formulated within ganglioside GM3-engineered lipid nanoparticles (LNPs) for specific targeting of CD169⁺ macrophage, and is co-delivered with IL12 mRNA as an integrated adjuvant. In murine models of ocular and cutaneous HSV-1 challenge, prime-boost immunization with MTL-bivalent-IL12 elicited robust neutralizing antibodies and antigen-specific T-cell responses, resulting in a significant reduction in viral loads in trigeminal ganglia (TG) and dorsal root ganglia (DRG). Notably, subconjunctival administration preserved corneal integrity and effectively blocked neuroinvasion, whereas intramuscular immunization prevented epithelial shedding and skin lesion development, with both routes achieving 100% survival under high-dose challenge conditions. Critically, the developed macrophage-targeted LNP vaccine exhibited a favorable safety profile with no evidence of systemic toxicity. This integrated strategy confers robust cross-tissue protection against HSV-1 by enabling coordinated CD169⁺ macrophage-specific delivery, bivalent immunogen expression, and localized immune activation, positioning it as a promising translational platform for preemptive prophylaxis.