[Mpox virus (MPXV) DNA replication machinery: Structural mechanisms and antiviral targets]: A review

Current control of mpox virus (MPXV) still relies mainly on smallpox-derived vaccines and antiviral agents, but their clinical use remains limited by uncertain efficacy, potential toxicity, and increasing resistance. Meanwhile, the key mechanisms underlying MPXV replication and its potential druggability remain insufficiently understood. Because viral DNA replication is directly linked to viral proliferation, the core proteins of the replication machinery are not only essential for maintaining the viral life cycle but may also represent more selective therapeutic targets. Therefore, systematic analysis of the composition, structure, and function of the MPXV replication machinery, together with related antiviral strategies, is of important theoretical and translational value. This review summarizes the core replication machinery of MPXV, including the F8-A22-E4 polymerase holoenzyme, the E5 helicase-primase, and the I3 single-stranded DNA-binding protein. Recent cryo-EM studies have shown that F8 and E4 form a unique closed ring-like channel around template single-stranded DNA. This "forward-sliding" mechanism differs from the canonical PCNA-dependent organization of B-family polymerases and provides a potential structural basis for selective drug design. Meanwhile, E5 participates in DNA unwinding and RNA primer synthesis, whereas I3 stabilizes single-stranded DNA and helps organize replication-associated factors to support viral DNA replication. Finally, this review summarizes advances in antiviral development targeting the polymerase complex and E5. Although cidofovir and brincidofovir remain limited by safety concerns, most next-generation inhibitors are still at the preclinical stage. Future efforts should prioritize target validation, combination therapy optimization, and resistance surveillance to support safer and more effective anti-MPXV therapeutics.