1. Academic Validation
  2. Molnupiravir promotes SARS-CoV-2 mutagenesis via the RNA template

Molnupiravir promotes SARS-CoV-2 mutagenesis via the RNA template

  • J Biol Chem. 2021 Jul;297(1):100770. doi: 10.1016/j.jbc.2021.100770.
Calvin J Gordon 1 Egor P Tchesnokov 1 Raymond F Schinazi 2 Matthias Götte 3
Affiliations

Affiliations

  • 1 Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada.
  • 2 Laboratory of Biochemical Pharmacology, Department of Pediatrics, Center for AIDS Research, Emory University School of Medicine, and Children's Healthcare of Atlanta, Atlanta, Georgia, USA.
  • 3 Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada; Li Ka Shing Institute of Virology at University of Alberta, Edmonton, Alberta, Canada. Electronic address: gotte@ualberta.ca.
Abstract

The RNA-dependent RNA polymerase of the severe acute respiratory syndrome coronavirus 2 is an important target in current drug development efforts for the treatment of coronavirus disease 2019. Molnupiravir is a broad-spectrum Antiviral that is an orally bioavailable prodrug of the nucleoside analogue β-D-N4-hydroxycytidine (NHC). Molnupiravir or NHC can increase G to A and C to U transition mutations in replicating coronaviruses. These increases in mutation frequencies can be linked to increases in Antiviral effects; however, biochemical data of molnupiravir-induced mutagenesis have not been reported. Here we studied the effects of the active compound NHC 5'-triphosphate (NHC-TP) against the purified severe acute respiratory syndrome coronavirus 2 RNA-dependent RNA polymerase complex. The efficiency of incorporation of natural nucleotides over the efficiency of incorporation of NHC-TP into model RNA substrates followed the order GTP (12,841) > ATP (424) > UTP (171) > CTP (30), indicating that NHC-TP competes predominantly with CTP for incorporation. No significant inhibition of RNA synthesis was noted as a result of the incorporated monophosphate in the RNA primer strand. When embedded in the template strand, NHC-monophosphate supported the formation of both NHC:G and NHC:A base pairs with similar efficiencies. The extension of the NHC:G product was modestly inhibited, but higher nucleotide concentrations could overcome this blockage. In contrast, the NHC:A base pair led to the observed G to A (G:NHC:A) or C to U (C:G:NHC:A:U) mutations. Together, these biochemical data support a mechanism of action of molnupiravir that is primarily based on RNA mutagenesis mediated via the template strand.

Keywords

Covid-19; RNA-dependent RNA polymerase; SARS-CoV-2; antiviral agent; coronavirus; drug development; mutagen; nucleoside analogue.

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