Single-chain A35R-M1R-B6R trivalent mRNA vaccines protect mice against both mpox virus and vaccinia virus.

Background: Mpox has spread to many countries around the world. While the existing live attenuated mpox vaccines are effective, advances in 21st century technologies now enable the development of vaccines with more specific antigens, clearer mechanisms, and more controllable side effects.

Methods: We systematically evaluated the immunogenicity and protective efficacy of the A35R, M1R and B6R antigens of the mpox virus (MPXV). With these findings, we designed three single-chain trivalent mRNA vaccines (AMAB-wt, AMAB-C140S and AMB-C140S) by integrating the soluble regions of these antigens into single mRNA-encoded polypeptides based on their protein structures. Then, the immunogenicity and protective efficacy of these single-chain mRNA vaccines were evaluated in mice models against both VACV and MPXV.

Findings: The three single-chain vaccines elicited neutralising antibodies that effectively neutralised both VACV and MPXV. The single-chain vaccines or cocktail vaccine containing mRNAs encoding soluble antigen (sA35R + sM1R + sB6R) exhibited 100% or 80% protection against a lethal dose of VACV challenge, while the cocktail of full-length antigens (A35 + M1 + B6) and the live attenuated vaccine, VACV Tian Tan (VACV-VTT), completely failed to protect mice. Moreover, the single-chain vaccines significantly reduced viral load in the lungs and ovaries of MPXV-challenged mice.

Interpretation: Compared with the cocktail vaccines, our single-chain designs demonstrated similar or superior immunogenicity and protective efficacy. Importantly, the simplicity of the single-chain vaccines enhances both the controllability and accessibility of mpox vaccines. We believe these single-chain vaccines qualify as the next-generation mpox vaccines.

Funding: National Natural Science Foundation of China and Youth Innovation Promotion Association of the CAS.