Characterization of the inhibition of hepatitis C virus RNA replication by nonnucleosides.
The RNA-dependent RNA polymerase of hepatitis C virus (HCV) is necessary for the replication of viral RNA and thus represents an attractive target for drug development. Several structural classes of nonnucleoside inhibitors (NNIs) of HCV RNA polymerase have been described, including a promising series of benzothiadiazine compounds that efficiently block replication of HCV subgenomic replicons in tissue culture. In this work we report the selection of replicons resistant to inhibition by the benzothiadiazine class of NNIs. Four different single mutations were identified in separate clones, and all four map to the RNA polymerase gene, validating the polymerase as the antiviral target of inhibition. The mutations (M414T, C451R, G558R, and H95R) render the HCV replicons resistant to inhibition by benzothiadiazines, though the mutant replicons remain sensitive to inhibition by other nucleoside and NNIs of the HCV RNA polymerase. Additionally, cross-resistance studies and synergistic inhibition of the enzyme by combinations of a benzimidazole and a benzothiadiazine indicate the existence of nonoverlapping binding sites for these two structural classes of inhibitors.
Tomei
Licia
L
Department of Biochemistry, Istituto di Ricerche di Biologia Molecolare P. Angeletti (IRBM), Pomezia, Italy.
Altamura
Sergio
S
Bartholomew
Linda
L
Bisbocci
Monica
M
Bailey
Carolyn
C
Bosserman
Michele
M
Cellucci
Antonella
A
Forte
Eleonora
E
Incitti
Ilario
I
Orsatti
Laura
L
Koch
Uwe
U
De Francesco
Raffaele
R
Olsen
David B
DB
Carroll
Steven S
SS
Migliaccio
Giovanni
G
D000998:Antiviral Agents / Q000494:pharmacology*; D001562:Benzimidazoles / Q000494:pharmacology*; D001581:Benzothiadiazines / Q000494:pharmacology*; D001665:Binding Sites; D002460:Cell Line; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016174:Hepacivirus / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D012367:RNA, Viral / Q000096:biosynthesis; D012324:RNA-Dependent RNA Polymerase / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication / Q000187:drug effects*
D000998:Antiviral Agents; D001562:Benzimidazoles; D001581:Benzothiadiazines; D004791:Enzyme Inhibitors; D012367:RNA, Viral; C031000:benzimidazole; D012324:RNA-Dependent RNA Polymerase