Metabolism of the Anti-Hepatitis C Virus Nucleoside β- d - N ⁴ -Hydroxycytidine in Different Liver Cells

Beta-D-N4-hydroxycytidine (NHC) was found to have selective anti-hepatitis C virus (HCV) activity in the HCV replicon system (clone A). The intracellular metabolism of tritiated NHC was investigated in the HCV replicon system, Huh-7 cells, HepG2 cells, and primary human hepatocytes. Incubation of cells with 10 microM radiolabeled NHC demonstrated extensive and rapid phosphorylation in all liver cells. Besides the 5'-mono, -di-, and -triphosphate metabolites of NHC, other metabolites were characterized. These included cytidine and uridine mono-, di-, and triphosphates. UTP was the predominant early metabolite in Huh-7 cells and primary human hepatocytes, suggesting deamination of NHC as the primary catabolic pathway. The intracellular half-lives of radiolabeled NHC-triphosphate and of CTP and UTP derived from NHC incubation in Huh-7 cells were calculated to be 3.0 +/- 1.3, 10.4 +/- 3.3, and 13.2 +/- 3.5 h (means +/- standard deviations), respectively. Studies using monkey and human whole blood demonstrated more-rapid deamination and oxidation in monkey cells than in human cells, suggesting that NHC may not persist long enough in plasma to be delivered to liver cells.