title: Mechanism of resistance of hepatitis C virus replicons to structurally distinct cyclophilin inhibitors.
abstract: The current standard of care for hepatitis C virus (HCV) infection, pegylated alpha interferon in combination with ribavirin, has a limited response rate and adverse side effects. Drugs targeting viral proteins are in clinical development, but they suffer from the development of high viral resistance. The inhibition of cellular proteins that are essential for viral amplification is thought to have a higher barrier to the emergence of resistance. Three cyclophilin inhibitors, the cyclosporine analogs DEBIO-025, SCY635, and NIM811, have shown promising results for the treatment of HCV infection in early clinical trials. In this study, we investigated the frequency and mechanism of resistance to cyclosporine (CsA), NIM811, and a structurally unrelated cyclophilin inhibitor, SFA-1, in replicon-containing Huh7 cells. Cross-resistance between all clones was observed. NIM811-resistant clones were selected only after obtaining initial resistance to either CsA or SFA-1. The time required to select resistance against cyclophilin inhibitors was significantly longer than that required for resistance selection against viral protein inhibitors, and the achievable resistance level was substantially lower. Resistance to cyclophilin inhibitors was mediated by amino acid substitutions in NS3, NS5A, and NS5B, with NS5A mutations conferring the majority of resistance. Mutation D320E in NS5A mediated most of the resistance conferred by NS5A. Taken together, the results indicate that there is a very low frequency and level of resistance to cyclophilin-binding drugs mediated by amino acid substitutions in three viral proteins. The interaction of cyclophilin with NS5A seems to be the most critical, since the NS5A mutations have the largest impact on resistance.
authors.affiliation: Novartis Institutes for Biomedical Research, Inc., Cambridge, MA 02139, USA.
authors.forename: Xiaoling
authors.initials: X
authors.lastname: Puyang
authors.forename: Danielle L
authors.initials: DL
authors.lastname: Poulin
authors.forename: Joanna E
authors.initials: JE
authors.lastname: Mathy
authors.forename: Leah J
authors.initials: LJ
authors.lastname: Anderson
authors.forename: Sue
authors.initials: S
authors.lastname: Ma
authors.forename: Zheng
authors.initials: Z
authors.lastname: Fang
authors.forename: Shejin
authors.initials: S
authors.lastname: Zhu
authors.forename: Kai
authors.initials: K
authors.lastname: Lin
authors.forename: Roger
authors.initials: R
authors.lastname: Fujimoto
authors.forename: Teresa
authors.initials: T
authors.lastname: Compton
authors.forename: Brigitte
authors.initials: B
authors.lastname: Wiedmann
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology; D002460:Cell Line; D021983:Cyclophilins / Q000037:antagonists & inhibitors*; D016572:Cyclosporine / Q000494:pharmacology*; D003524:Cyclosporins / Q000494:pharmacology; D024882:Drug Resistance, Viral / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016174:Hepacivirus / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D006801:Humans; D007783:Lactones / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D012367:RNA, Viral / Q000378:metabolism; D012093:Replicon / Q000235:genetics*; D013141:Spiro Compounds / Q000494:pharmacology; D014162:Transfection; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; D012324:RNA-Dependent RNA Polymerase
chemical_list: D000998:Antiviral Agents; D021983:Cyclophilins; D016572:Cyclosporine; D003524:Cyclosporins; D004791:Enzyme Inhibitors; D007783:Lactones; D012367:RNA, Viral; D013141:Spiro Compounds; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C121329:sanglifehrin A; C090438:(melle-4)cyclosporin; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase; C499715:alisporivir; C544142:SCY-635