A Single-Chain Mpox mRNA Vaccine Elicits Protective Immune Response in Mice.

The re-emerging mpox virus (MPXV) has spread to numerous countries and raised global concern. There is an urgent need for a safe and effective mRNA vaccine candidate against MPXV infection. Previously, we developed a penta-component mRNA vaccine that contained five distinct antigen-encoded mRNAs encapsulated within lipid nanoparticles (LNPs). Here, we sought to develop a single-chain mRNA vaccine that encodes antigens derived from both intracellular mature virion (IMV) and extracellular enveloped virion (EEV). A single-chain mRNA vaccine encoding a fusion protein comprising the ectodomains of M1R (eM1R) and A35R (eA35R) (MPXV) was developed and characterized, while an admixed formulation of two individual mRNA-LNPs encoding separate antigens was developed as the control (MPXV). Meanwhile, based on the same strategy, we designed a single-chain mRNA vaccine encoding dimeric antigens (MPXV). Mice were immunized with two doses of the candidate vaccines, and both humoral and cellular immune responses were evaluated. The protective efficacy of the candidate vaccines was evaluated based on body weight monitoring and tissue viral load measurement after challenge with vaccinia virus (VACV). Immunization with two doses of MPXV elicited robust levels of neutralizing antibodies and antigen-specific cellular immune response. Importantly, MPXV demonstrated protective efficacy in a VACV challenge mouse model and showed superior capacity in preventing weight loss post-challenge compared to MPXV. Similarly, MPXV exhibited comparable or superior immunogenicity and protective efficacy compared to the admixed formulations. The single-chain mRNA vaccine elicited a protective immune response in mice, offering significant advantages in terms of manufacturing processes and quality control. Our single-chain mRNA vaccine platform presents a promising strategy for the next generation design of mpox vaccines and contributes to the mitigation of MPXV endemic worldwide.