Identification of a novel resistance mutation for benzimidazole inhibitors of the HCV RNA-dependent RNA polymerase.
Non-nucleoside inhibitors of the RNA-dependent RNA polymerase of the hepatitis C virus that are based on a benzimidazole or indole scaffold have been reported to interact with thumb domain 1 of the enzyme. Escape mutants that confer in vitro resistance to these inhibitors map to amino acids P495, P496 or V499. We here report a novel resistance mutation (T389S/A) that was identified following resistance selection with the benzimidazole non-nucleoside polymerase inhibitor JT-16 in HCV Con-1 subgenomic replicon (genotype 1b). This JT-16 resistant replicon retained wild-type susceptibility to protease inhibitors and nucleoside polymerase inhibitors. Replicons that carry mutations T389A and T389S have moderate levels of resistance to JT-16 (7- and 13-fold, respectively). Mutation P495A is associated with high-level (44-fold) resistance. Surprisingly, this previously reported 'key' mutation for benzimidazole resistance, P495A, was detected in only 15% of the resistant population. Furthermore, the replication fitness of the T389S mutant was significantly higher than that of the P495A mutant. By means of molecular modeling a structural hypothesis was formulated to explain the emergence of the T389S/A mutation in the JT-16 resistant replicon. Our data demonstrate that low-level resistant, but fit, variants can develop during in vitro resistance selection with the benzimidazole inhibitor JT-16. Moreover, different substitutions to the benzimidazole scaffold can affect the (pattern of) resistance mutations that emerge during resistance selection.
Department of Microbiology and Immunology, Rega Institute for Medical Research, K.U.Leuven, Belgium.
Leen
L
Delang
Mathy
M
Froeyen
Piet
P
Herdewijn
Johan
J
Neyts
D000081:Acetamides / Q000737:chemistry / Q000494:pharmacology*; D000998:Antiviral Agents / Q000737:chemistry / Q000494:pharmacology*; D001562:Benzimidazoles / Q000737:chemistry / Q000494:pharmacology*; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D024882:Drug Resistance, Viral / Q000235:genetics; D005838:Genotype; D016174:Hepacivirus / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D012324:RNA-Dependent RNA Polymerase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics
C571516:2-(4-((4-(acetylamino)-4'-chloro-(1,1'-biphenyl)-2-yl)methoxy)phenyl)-1-cyclohexyl-1H-benzimidazole-5-carboxylic acid; D000081:Acetamides; D000998:Antiviral Agents; D001562:Benzimidazoles; D012324:RNA-Dependent RNA Polymerase