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Author Notes:

Correspondence: jdeval@aliosbiopharma.com (JD); lbeigelman@aliosbiopharma.com (LB)

Conceived and designed the experiments: JD JH GW ZJ ADS JL DBS QZ SKS LB SMC.

Performed the experiments: JD JT LKS AF SKS HL ZJ ND MP ADS VS YT.

Analyzed the data: JD JH GW ZJ ADS JL DBS QZ.

Contributed reagents/materials/analysis tools: MLMRF.

Wrote the paper: JD MLM RF ADS MP DBS JAS LMB LB SMC.

We thank Mark Peeples from The Ohio State University, USA, for his help in developing the RSV replicon assay licensed from Apath LLC, USA.

We also thank April Kinkade, Humaira Sarker, and Andreas Jekle from Alios BioPharma for their help with the influenza and rhinovirus assays, as well as Kenneth Shaw and Hua Tan for their input in the HCV and VSV cellbased inhibition assays.

We also wish to thank the staff at BIOQUAL Inc., Rockville, MD for their expertise in the conduct of the African Green monkey efficacy study, especially Hanne Andersen Elyard for study direction, Brad Finneyfrock, Steve and Jeff Harbaugh for animal care and procedures; Jack Greenhouse for viral RNA assays, and Liz Peters for sample processing and study coordination.

Finally, we thank Peggy Korn for her editorial review of the manuscript, and Bruno Canard from CNRS, France, for helpful discussions.

The authors of this manuscript have the following competing interests: JD, JH, GW, JT, LKS, AF, SKS, HL, ZJ, ND, MP, ADS, VS, JL, DBS, YT, QZ, SMC, LMB, JAS, and LB are current or former employees of Alios BioPharma, a Janssen Pharmaceutical Company of Johnson & Johnson.

This does not alter our adherence to all PLOS policies on sharing data and materials.

Subjects:

Research Funding:

The authors received no specific funding for this work.

Keywords:

  • Science & Technology
  • Life Sciences & Biomedicine
  • Microbiology
  • Parasitology
  • Virology
  • T-705 FAVIPIRAVIR
  • STRUCTURAL BASIS
  • CELL-CULTURE
  • ACTIVE-SITE
  • INFECTION
  • REPLICATION
  • SEQUENCE
  • TRANSCRIPTASE
  • INFLUENZA
  • DOMAINS
  • Polymerase
  • RNA polymerase
  • Ribonucleoproteins
  • Hepatitis C virus
  • Nucleosides
  • RNA virus
  • RNA synthesis
  • Viral replication

Molecular Basis for the Selective Inhibition of Respiratory Syncytial Virus RNA Polymerase by 2 '-Fluoro-4 '-Chloromethyl-Cytidine Triphosphate

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Journal Title:

PLoS Pathogens

Volume:

Volume 11, Number 6

Publisher:

, Pages e1004995-e1004995

Type of Work:

Article | Final Publisher PDF

Abstract:

Respiratory syncytial virus (RSV) causes severe lower respiratory tract infections, yet no vaccines or effective therapeutics are available. ALS-8176 is a first-in-class nucleoside analog prodrug effective in RSV-infected adult volunteers, and currently under evaluation in hospitalized infants. Here, we report the mechanism of inhibition and selectivity of ALS-8176 and its parent ALS-8112. ALS-8176 inhibited RSV replication in non-human primates, while ALS-8112 inhibited all strains of RSV in vitro and was specific for paramyxoviruses and rhabdoviruses. The antiviral effect of ALS-8112 was mediated by the intracellular formation of its 5'-triphosphate metabolite (ALS-8112-TP) inhibiting the viral RNA polymerase. ALS-8112 selected for resistance-associated mutations within the region of the L gene of RSV encoding the RNA polymerase. In biochemical assays, ALS-8112-TP was efficiently recognized by the recombinant RSV polymerase complex, causing chain termination of RNA synthesis. ALS-8112-TP did not inhibit polymerases from host or viruses unrelated to RSV such as hepatitis C virus (HCV), whereas structurally related molecules displayed dual RSV/HCV inhibition. The combination of molecular modeling and enzymatic analysis showed that both the 2'F and the 4'ClCH<inf>2</inf> groups contributed to the selectivity of ALS-8112-TP. The lack of antiviral effect of ALS-8112-TP against HCV polymerase was caused by Asn291 that is well-conserved within positive-strand RNA viruses. This represents the first comparative study employing recombinant RSV and HCV polymerases to define the selectivity of clinically relevant nucleotide analogs. Understanding nucleotide selectivity towards distant viral RNA polymerases could not only be used to repurpose existing drugs against new viral infections, but also to design novel molecules.

Copyright information:

© 2015 Deval et al.

This is an Open Access work distributed under the terms of the Creative Commons Attribution 4.0 International License (https://creativecommons.org/licenses/by/4.0/).
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