A robust comprehensive immunoinformatics approach for designing a potential multi-epitope based vaccine against a reiterated monkeypox virus

Mpox, a viral disease, caused by the monkeypox virus (MPXV) has been a public health emergency of international concern since 2024. The absence of any mpox-specific treatment or vaccine, along with the emergence of new variants like Clade Ib, underscores the urgent need for targeted vaccine development. To address the challenge, this study employed reverse vaccinology and immunoinformatics approaches to design a multi-epitope vaccine against MPXV. The vaccine construct includes four Linear B lymphocyte (LBL), nine Cytotoxic T lymphocyte (CTL), and seven Helper T lymphocyte (HTL) epitopes. LBL epitopes were selected from six membrane glycoproteins of the virus and the T-cell epitopes were selected from the experimentally validated conserved epitopes of the similar orthopoxviruses. These epitopes were combined with appropriate linkers and adjuvants to enhance structural flexibility, immunogenicity, and potency. The engineered vaccine underwent rigorous evaluation, considering physicochemical properties, structural integrity, population coverage, and immune system response through simulation. The 3D structure of the vaccine was predicted, optimized, and docking analysis revealed robust interactions with the human Toll-like receptor 2 and 4 (TLR-2 and TLR-4), supported by highly negative HADDOCK scores and low RMSD values. The stability of the vaccine construct and its stable interaction with TLR-2 and TLR-4 were confirmed by molecular dynamics (MD) simulation. Additionally, the immune simulation results showed that the vaccination significantly increased IgM levels during the primary response, while IgG subtypes as well as combined IgM and IgG levels nearly doubled in the secondary and tertiary responses. In silico expression in Escherichia coli (E. coli) further confirmed its potential for production. Overall, this study presents a highly immunogenic and promising vaccine candidate against MPXV that demands experimental validation for clinical application.