Efficient incorporation and template-dependent polymerase inhibition are major determinants for the broad-spectrum antiviral activity of remdesivir

Götte, M.; Gordon, C. J.; Lee, H. W.; Tchesnokov, E. P.; Perry, J. K.; Feng, J. Y.; Bilello, J. P.; Porter, D. P.

doi:10.1101/2021.10.14.464416

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bioRxiv · 10.1101/2021.10.14.464416

Efficient incorporation and template-dependent polymerase inhibition are major determinants for the broad-spectrum antiviral activity of remdesivir

Götte, M.·Gordon, C. J.·Lee, H. W.·Tchesnokov, E. P.·Perry, J. K.·Feng, J. Y.·Bilello, J. P.·Porter, D. P.

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Abstract

Remdesivir (RDV) is a direct antiviral agent that is approved in several countries for the treatment of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). RDV exhibits broad-spectrum antiviral activity against positive-sense RNA viruses, e.g., SARS-CoV-2 and hepatitis C virus (HCV) and non-segmented negative-sense RNA viruses, e.g., Nipah virus (NiV), while several segmented negative-sense RNA viruses such as influenza (Flu) virus or Crimean-Congo hemorrhagic fever virus (CCHFV) are not sensitive to the drug. The reasons for this apparent pattern are unknown. Here, we expressed and purified representative RNA-dependent RNA polymerases (RdRp) and studied three biochemical parameters that have been associated with the inhibitory effects of RDV-triphosphate (TP): (i) selective incorporation of the nucleotide substrate RDV-TP, (ii) the effect of the incorporated RDV-monophosphate (MP) on primer extension, and (iii) the effect of RDV-MP in the template during incorporation of the complementary UTP. The results of this study revealed a strong correlation between antiviral effects and efficient incorporation of RDV-TP. Delayed chain-termination is heterogeneous and usually inefficient at higher NTP concentrations. In contrast, template-dependent inhibition of UTP incorporation opposite the embedded RDV-MP is seen with all polymerases. Molecular modeling suggests a steric conflict between the 1-cyano group of RDV-MP and conserved residues of RdRp motif F. We conclude that future efforts in the development of nucleotide analogues with a broader spectrum of antiviral activities should focus on improving rates of incorporation while capitalizing on the inhibitory effects of a bulky 1-modification.

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Götte, M., Gordon, C. J., Lee, H. W., Tchesnokov, E. P., Perry, J. K., Feng, J. Y., Bilello, J. P., Porter, D. P.. 2021-10-15. Efficient incorporation and template-dependent polymerase inhibition are major determinants for the broad-spectrum antiviral activity of remdesivir. https://doi.org/10.1101/2021.10.14.464416

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