Identification and analysis of fitness of resistance mutations against the HCV protease inhibitor SCH 503034.
HCV NS3 protease variants resistant to the protease inhibitor SCH 503034 were selected. Three mutations, T54A, V170A and A156S mutations conferred low to moderate levels of resistance (<20-fold). Longer exposure (>10 passages) or selection with higher levels of compound led to the selection of a more resistant variant, A156T (>100-fold). [Lin, C., Lin, K., Luong, Y.P., Rao, B.G., Wei, Y.Y., Brennan, D.L., Fulghum, J.R., Hsiao, H.M., Ma, S., Maxwell, J.P., Cottrell, K.M., Perni, R.B., Gates, C.A., Kwong, A.D., 2004. In vitro resistance studies of hepatitis C virus serine protease inhibitors, VX-950 and BILN 2061: structural analysis indicates different resistance mechanisms. J. Biol. Chem. 279(17), 17508-17514; Lu, L., Pilot-Matias, T.J., Stewart, K.D., Randolph, J.T., Pithawalla, R., He, W., Huang, P.P., Klein, L.L., Mo, H., Molla, A., 2004. Mutations conferring resistance to a potent hepatitis C virus serine protease inhibitor in vitro. Antimicrob. Agents Chemother. 48(6), 2260-2266.] Combination with IFN-alpha drastically reduced the number of emergent colonies. Resistant colonies showed no change in sensitivity to IFN-alpha. Although the A156T mutation conferred the highest level of resistance to SCH 503034, it significantly reduced the colony formation efficiency (CFE) of the mutant replicon RNA, and rendered replicon cells less fit than those bearing wild-type replicons. Replicon cells bearing mutation A156S were less fit than wild-type in co-culture growth competition assays but showed no impact on CFE. The V170A mutation, on the other hand, did not affect replicon fitness in either assay, which was consistent with its emergence as the dominant mutant after 12 months of continuous selection. The reduced fitness of the most resistant variant suggests that it may be rare in naïve patients and that development of high-level resistance may be slow. Combination therapy with IFN-alpha should also greatly reduce the potential emergence of resistance.
Tong
Xiao
X
Department of Virology, Schering-Plough Research Institute, 2015 Galloping Hill Road, Kenilwoth, NJ 07033, USA. xiao.tong@spcorp.com
Chase
Robert
R
Skelton
Angela
A
Chen
Tong
T
Wright-Minogue
Jackie
J
Malcolm
Bruce A
BA
D015153:Blotting, Western; D045744:Cell Line, Tumor; D024882:Drug Resistance, Viral / Q000235:genetics; D004789:Enzyme Activation; D016174:Hepacivirus / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010641:Phenotype; D011392:Proline / Q000031:analogs & derivatives* / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology*; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D012093:Replicon / Q000187:drug effects / Q000235:genetics; D017421:Sequence Analysis; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics* / Q000378:metabolism; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D011392:Proline; D011480:Protease Inhibitors; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C512204:N-(3-amino-1-(cyclobutylmethyl)-2,3-dioxopropyl)-3-(2-((((1,1-dimethylethyl)amino)carbonyl)amino)-3,3-dimethyl-1-oxobutyl)-6,6-dimethyl-3-azabicyclo(3.1.0)hexan-2-carboxamide