Multiple mutations in hepatitis C virus NS5A domain II are required to confer a significant level of resistance to alisporivir.
Alisporivir is the most advanced host-targeting antiviral cyclophilin (Cyp) inhibitor in phase III studies and has demonstrated a great deal of promise in decreasing hepatitis C virus (HCV) viremia in infected patients. In an attempt to further elucidate the mechanism of action of alisporivir, HCV replicons resistant to the drug were selected. Interestingly, mutations constantly arose in domain II of NS5A. To demonstrate that these mutations are responsible for drug resistance, they were reintroduced into the parental HCV genome, and the resulting mutant viruses were tested for replication in the presence of alisporivir or in the absence of the alisporivir target, CypA. We also examined the effect of the mutations on NS5A binding to itself (oligomerization), CypA, RNA, and NS5B. Importantly, the mutations did not affect any of these interactions. Moreover, the mutations did not preserve NS5A-CypA interactions from alisporivir rupture. NS5A mutations alone render HCV only slightly resistant to alisporivir. In sharp contrast, when multiple NS5A mutations are combined, significant resistance was observed. The introduction of multiple mutations in NS5A significantly restored viral replication in CypA knockdown cells. Interestingly, the combination of NS5A mutations renders HCV resistant to all classes of Cyp inhibitors. This study suggests that a combination of multiple mutations in domain II of NS5A rather than a single mutation is required to render HCV significantly and universally resistant to Cyp inhibitors. This in accordance with in vivo data that suggest that alisporivir is associated with a low potential for development of viral resistance.
Department of Immunology & Microbial Science, The Scripps Research Institute, La Jolla, California, USA.
Jose A
JA
Garcia-Rivera
Michael
M
Bobardt
Udayan
U
Chatterji
Sam
S
Hopkins
Matthew A
MA
Gregory
Barrie
B
Wilkinson
Kai
K
Lin
Philippe A
PA
Gallay
D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D016572:Cyclosporine / Q000494:pharmacology*; D024882:Drug Resistance, Viral / Q000235:genetics; D005838:Genotype; D016174:Hepacivirus / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D017361:Viral Nonstructural Proteins / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase
D000998:Antiviral Agents; D016572:Cyclosporine; D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase; C499715:alisporivir