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

• Authors: Jerome Deval, Alios BioPharma, Inc.; Jin Hong, Alios BioPharma, Inc.; Guangyi Wang, Alios BioPharma, Inc.; Josh Taylor, Alios BioPharma, Inc.; Lucas K. Smith, Alios BioPharma, Inc.; Amy Fung, Alios BioPharma, Inc.; Sarah K. Stevens, Alios BioPharma, Inc.; Hong Liu, Alios BioPharma, Inc.; Zhinan Jin, Alios BioPharma, Inc.; Natalia Dyatkina, Alios BioPharma, Inc.; Marija Prhavc, Alios BioPharma, Inc.; Antitsa D. Stoycheva, Alios BioPharma, Inc.; Vladimir Serebryany, Alios BioPharma, Inc.; Jyanwei Liu, Alios BioPharma, Inc.; David B. Smith, Alios BioPharma, Inc.; Yuen Tam, Alios BioPharma, Inc.; Qingling Zhang, Alios BioPharma, Inc.; Martin Moore, Emory University; Rachel Fearns, Boston University; Sushmita M. Chanda, Alios BioPharma, Inc.; Lawrence M. Blatt, Alios BioPharma, Inc.; Julian A. Symons, Alios BioPharma, Inc.; Leo Beigelman, Alios BioPharma, Inc.
• Journal Title: PLoS Pathogens
• Volume: Volume 11, Number 6
• Publisher: Public Library of Science | 2015-06, Pages e1004995-e1004995
• Type of Work: Article | Final Publisher PDF
• Publisher DOI: http://doi.org/10.1371/journal.ppat.1004995
• Subjects: Biology, Virology; Biology, Microbiology; Biology, Genetics
• 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

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.