title: Characterization of the metabolic activation of hepatitis C virus nucleoside inhibitor beta-D-2'-Deoxy-2'-fluoro-2'-C-methylcytidine (PSI-6130) and identification of a novel active 5'-triphosphate species.
abstract: beta-D-2'-Deoxy-2'-fluoro-2'-C-methylcytidine (PSI-6130) is a potent inhibitor of hepatitis C virus (HCV) replication in the subgenomic HCV replicon system, and its corresponding 5'-triphosphate is a potent inhibitor of the HCV RNA polymerase in vitro. In this study the formation of PSI-6130-triphosphate was characterized in primary human hepatocytes. PSI-6130 and its 5'-phosphorylated derivatives were identified, and the intracellular concentrations were determined. In addition, the deaminated derivative of PSI-6130, beta-d-2'-deoxy-2'-fluoro-2'-C-methyluridine (RO2433, PSI-6026) and its corresponding phosphorylated metabolites were identified in human hepatocytes after incubation with PSI-6130. The formation of the 5'-triphosphate (TP) of PSI-6130 (PSI-6130-TP) and RO2433 (RO2433-TP) increased with time and reached steady state levels at 48 h. The formation of both PSI-6130-TP and RO2433-TP demonstrated a linear relationship with the extracellular concentrations of PSI-6130 up to 100 mum, suggesting a high capacity of human hepatocytes to generate the two triphosphates. The mean half-lives of PSI-6130-TP and RO2433-TP were 4.7 and 38 h, respectively. RO2433-TP also inhibited RNA synthesis by the native HCV replicase isolated from HCV replicon cells and the recombinant HCV polymerase NS5B with potencies comparable with those of PSI-6130-TP. Incorporation of RO2433-5'-monophosphate (MP) into nascent RNA by NS5B led to chain termination similar to that of PSI-6130-MP. These results demonstrate that PSI-6130 is metabolized to two pharmacologically active species in primary human hepatocytes.
authors: Roche Palo Alto LLC, Palo Alto, California 94304, USA. han.ma@roche.com
authors: Han
authors: H
authors: Ma
authors: Wen-Rong
authors: WR
authors: Jiang
authors: Nicole
authors: N
authors: Robledo
authors: Vincent
authors: V
authors: Leveque
authors: Samir
authors: S
authors: Ali
authors: Teresa
authors: T
authors: Lara-Jaime
authors: Mohammad
authors: M
authors: Masjedizadeh
authors: David B
authors: DB
authors: Smith
authors: Nick
authors: N
authors: Cammack
authors: Klaus
authors: K
authors: Klumpp
authors: Julian
authors: J
authors: Symons
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology*; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D016174:Hepacivirus / Q000201:enzymology / Q000378:metabolism*; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D020128:Inhibitory Concentration 50; D008954:Models, Biological; D008956:Models, Chemical; D010710:Phosphates / Q000737:chemistry; D010766:Phosphorylation; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism; D014779:Virus Replication
chemical_list: D000998:Antiviral Agents; D010710:Phosphates; C504520:2'-deoxy-2'-fluoro-2'-C-methylcytidine; D003841:Deoxycytidine; D012324:RNA-Dependent RNA Polymerase; D012321:DNA-Directed RNA Polymerases