Discovery of a Series of Potent and Selective Nucleotide Prodrug Inhibitors of Respiratory Syncytial Virus (RSV) Replication

Abstract

BackgroundRSV can cause severe respiratory tract infections in infants and the elderly. Current experimental therapies include polymerase and fusion inhibitors, but their clinical use may be limited by toxicity or rapid emergence of viral resistance. Here we report new nucleotide prodrugs that are selective for and highly active against RSV replication in vitro.MethodsNovel nucleotide prodrugs were synthesized and tested for their ability to inhibit RSV replication in 3-dimensional preparations of differentiated normal human bronchial epithelial (dNHBE) cells. Drug selectivity was assessed in the anti-RSV assays at concentrations up to 100 µg/mL, and in 14-day exposures with human bone marrow stem cells and 3-day exposures with human induced pluripotent (iPS) cardiomyocytes at concentrations up to 100 µM. The formation and half-lives (t½) of analog triphosphates (TPs) of selected prodrugs were measured in phytohaemagglutinin-stimulated human peripheral blood mononuclear cells (PBMCs) incubated with 100 µM prodrug. After 8 hours, medium was replaced with fresh medium without drug and cell extracts were prepared at various time points and analyzed for intracellular levels of TPs. After single oral dosing of Golden Syrian hamsters with selected prodrugs (~60 mg/kg), plasma pharmacokinetics and lung levels of TPs were determined at 4 and 24 hours or at 24 and 72 hours post dose.ResultsThe most potent nucleotide prodrugs inhibited RSV replication by 90% at concentrations (EC90) as low as 0.021 µM. None of the prodrugs tested showed significant cytotoxicity with dNHBE cells, bone marrow stem cells or cardiomyocytes. The t½ of the TPs formed in human PBMCs ranged from 1.3 to >5 days. In hamsters, plasma parent drug levels were ≤1 ng/mL, yet significant levels of the corresponding TPs were detected in lung tissue. Furthermore, the highest TP concentrations (up to 1344 ng/g) were observed at the latest sampling time point (up to 72 hours).ConclusionThe data indicate that these potent new nucleotide prodrugs are metabolized to TPs that prevent RSV replication likely by inhibition of the viral RNA polymerase. Additionally, the long t½ observed for many of the TPs suggest that it might be possible to cure RSV infections with a single dose. IND enabling studies are ongoing, targeting clinical evaluation in early 2018.Disclosures S. Good, Atea Pharmaceuticals, Inc.: Employee and Shareholder, Salary; A. Moussa, Atea Pharmaceuticals, Inc.: Employee and Shareholder, Salary; J. C. Meillon, Oxeltis: Employee and Shareholder, Salary; X. J. Zhou, Atea Pharmaceuticals, Inc.: Employee and Shareholder, Salary; K. Pietropaolo, Atea Pharmaceuticals, Inc.: Employee and Shareholder, Salary; J. P. Sommadossi, Atea Pharmaceuticals, Inc.: Board Member, Employee and Shareholder, Salary