Novel nonnucleoside inhibitor of hepatitis C virus RNA-dependent RNA polymerase.

Howe, Anita Y M; Bloom, Johnathan; Baldick, Carl J; Benetatos, Christopher A; Cheng, Huiming; Christensen, Joel S; Chunduru, Srinivas K; Coburn, Glen A; Feld, Boris; Gopalsamy, Ariamala; Gorczyca, William P; Herrmann, Steve; Johann, Stephen; Jiang, Xiaoqun; Kimberland, Michelle L; Krisnamurthy, Girija; Olson, Matthew; Orlowski, Mark; Swanberg, Steve; Thompson, Ian; Thorn, Megan; Del Vecchio, Alfred; Young, Dorothy C; van Zeijl, Marja; Ellingboe, John W; Upeslacis, Janis; Collett, Marc; Mansour, Tarek S; O'Connell, John F

A novel nonnucleoside inhibitor of hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp), [(1R)-5-cyano-8-methyl-1-propyl-1,3,4,9-tetrahydropyano[3,4-b]indol-1-yl] acetic acid (HCV-371), was discovered through high-throughput screening followed by chemical optimization. HCV-371 displayed broad inhibitory activities against the NS5B RdRp enzyme, with 50% inhibitory concentrations ranging from 0.3 to 1.8 microM for 90% of the isolates derived from HCV genotypes 1a, 1b, and 3a. HCV-371 showed no inhibitory activity against a panel of human polymerases, including mitochondrial DNA polymerase gamma, and other unrelated viral polymerases, demonstrating its specificity for the HCV polymerase. A single administration of HCV-371 to cells containing the HCV subgenomic replicon for 3 days resulted in a dose-dependent reduction of the steady-state levels of viral RNA and protein. Multiple treatments with HCV-371 for 16 days led to a >3-log10 reduction in the HCV RNA level. In comparison, multiple treatments with a similar inhibitory dose of alpha interferon resulted in a 2-log10 reduction of the viral RNA level. In addition, treatment of cells with a combination of HCV-371 and pegylated alpha interferon resulted in an additive antiviral activity. Within the effective antiviral concentrations of HCV-371, there was no effect on cell viability and metabolism. The intracellular antiviral specificity of HCV-371 was demonstrated by its lack of activity in cells infected with several DNA or RNA viruses. Fluorescence binding studies show that HCV-371 binds the NS5B with an apparent dissociation constant of 150 nM, leading to high selectivity and lack of cytotoxicity in the antiviral assays.