title: The novel nucleoside analog R1479 (4'-azidocytidine) is a potent inhibitor of NS5B-dependent RNA synthesis and hepatitis C virus replication in cell culture.
abstract: Hepatitis C virus (HCV) polymerase activity is essential for HCV replication. Targeted screening of nucleoside analogs identified R1479 (4'-azidocytidine) as a specific inhibitor of HCV replication in the HCV subgenomic replicon system (IC(50) = 1.28 microM) with similar potency compared with 2'-C-methylcytidine (IC(50) = 1.13 microM). R1479 showed no effect on cell viability or proliferation of HCV replicon or Huh-7 cells at concentrations up to 2 mM. HCV replicon RNA could be fully cleared from replicon cells after prolonged incubation with R1479. The corresponding 5'-triphosphate derivative (R1479-TP) is a potent inhibitor of native HCV replicase isolated from replicon cells and of recombinant HCV polymerase (NS5B)-mediated RNA synthesis activity. R1479-TP inhibited RNA synthesis as a CTP-competitive inhibitor with a K(i) of 40 nM. On an HCV RNA-derived template substrate (complementary internal ribosome entry site), R1479-TP showed similar potency of NS5B inhibition compared with 3'-dCTP. R1479-TP was incorporated into nascent RNA by HCV polymerase and reduced further elongation with similar efficiency compared with 3'-dCTP under the reaction conditions. The S282T point mutation in the coding sequence of NS5B confers resistance to inhibition by 2'-C-MeATP and other 2'-methyl-nucleotides. In contrast, the S282T mutation did not confer cross-resistance to R1479.
authors.affiliation: Roche Palo Alto LLC, CA 94304, USA. klaus.klumpp@roche.com
authors.forename: Klaus
authors.initials: K
authors.lastname: Klumpp
authors.forename: Vincent
authors.initials: V
authors.lastname: Lévêque
authors.forename: Sophie
authors.initials: S
authors.lastname: Le Pogam
authors.forename: Han
authors.initials: H
authors.lastname: Ma
authors.forename: Wen-Rong
authors.initials: WR
authors.lastname: Jiang
authors.forename: Hyunsoon
authors.initials: H
authors.lastname: Kang
authors.forename: Caroline
authors.initials: C
authors.lastname: Granycome
authors.forename: Margaret
authors.initials: M
authors.lastname: Singer
authors.forename: Carl
authors.initials: C
authors.lastname: Laxton
authors.forename: Julie Qi
authors.initials: JQ
authors.lastname: Hang
authors.forename: Keshab
authors.initials: K
authors.lastname: Sarma
authors.forename: David B
authors.initials: DB
authors.lastname: Smith
authors.forename: Dieter
authors.initials: D
authors.lastname: Heindl
authors.forename: Chris J
authors.initials: CJ
authors.lastname: Hobbs
authors.forename: John H
authors.initials: JH
authors.lastname: Merrett
authors.forename: Julian
authors.initials: J
authors.lastname: Symons
authors.forename: Nick
authors.initials: N
authors.lastname: Cammack
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Martin
authors.forename: Rene
authors.initials: R
authors.lastname: Devos
authors.forename: Isabel
authors.initials: I
authors.lastname: Nájera
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D003562:Cytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D016174:Hepacivirus / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D012367:RNA, Viral / Q000096:biosynthesis*; D012324:RNA-Dependent RNA Polymerase / Q000037:antagonists & inhibitors*; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors*; D014779:Virus Replication / Q000187:drug effects*
chemical_list: D000998:Antiviral Agents; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins; D003562:Cytidine; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase; C508950:4'-azidocytidine