HCV-796 (Nesbuvir)

Nesbuvir is a nonnucleoside inhibitor of the hepatitis C virus (HCV) nonstructural protein 5B (NS5B) polymerase.

Signaling Pathways >> Anti-infection >> HCV

Research Areas >> Infection

EC50: 9 nM (NS3V170A), 13 nM (NS3V170A), 15 nM (NS3K583T), 13 nM (NS5BI424V)[1]

Replicon cells are treated with 1 mg/mL G418 and combinations of the two compounds. Nesbuvir (HCV-796) is added to 40 or 80 nM (approximately 10 and 20 times the EC50 in a 3-day replicon inhibition assay, respectively) and Boceprevir is added to 400 or 800 nM (approximately 2 and 4 times the EC50, respectively). The EC50s for Nesbuvir and Boceprevir for the parental replicon in the transient expression assay are comparable to those obtained in the 3-day inhibition assay with the stable replicon cells; the EC50 for Nesbuvir in the transient expression assay is 14 nM, whereas it is 5 nM for the stable replicon; and the EC50 for Boceprevir in the transient expression assay is 608 nM, whereas it is 201 nM for the stable replicon[1].

Among a huge variety of yet characterized nucleoside and non-nucleoside inhibitors (NNI), the benzofurane derivative NNI Nesbuvir (HCV-796) is demonstrated to yield significant antiviral effects in mice with chimeric human livers and in patients infected with HCV. HCV-796 binds to a hydrophobic binding pocket at the “palm” domain of NS5B; however, its mode of inhibition remains to be defined[2].

Huh7-BB7 cells are seeded at a density of 20,000 cells per 100 mm dish in DMEM supplemented with 2% FBS, 1 mg/mL G418, and various concentrations of Nesbuvir and/or Boceprevir with DMSO at a final concentration of 0.5% (vol/vol). The medium is removed and is replaced with fresh medium with the appropriate compound concentrations every 3 or 4 days. After 7 days, the cells are split 1 to 10, placed into fresh 100 mm dishes, and incubated with medium with the appropriate compound concentrations. After 20 days, the medium is removed and the cells are fixed with 7% (wt/vol) formaldehyde and stained with 1% (wt/vol) crystal violet in 50% (vol/vol) ethanol[1].

[1]. Flint M, et al. Selection and characterization of hepatitis C virus replicons dually resistant to the polymerase and protease inhibitors HCV-796 and boceprevir (SCH 503034). Antimicrob Agents Chemother. 2009 Feb;53(2):401-11.

[2]. Reich S, et al. Mechanisms of activity and inhibition of the hepatitis C virus RNA-dependent RNA polymerase. J Biol Chem. 2010 Apr 30;285(18):13685-93.

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