Ortqvist Pernilla, Vema Aparna, Ehrenberg Angelica E, Dahl Göran, Rönn Robert, Akerblom Eva, Karlén Anders, Danielson U Helena, Sandström Anja
Department of Medicinal Chemistry, Uppsala University, Sweden.
Antivir Ther. 2010;15(6):841-52. doi: 10.3851/IMP1655.
HCV infections are a serious threat to public health. An important drug target is the NS3 protease, for which several inhibitors are in clinical trials. Because of the high mutation rate of the virus, resistance against any HCV-specific drug is likely to become a substantial problem. Structure-activity data for the major resistant variants are therefore needed to guide future designs of protease inhibitors.
The inhibitory potency of tripeptide NS3 protease inhibitors, with either a P2 proline or phenylglycine, in combination with different P3 and P1-P1' groups, was assessed in enzyme activity assays using the full-length NS3 protein with known resistance-conferring substitutions A156T or D168V. The results obtained from these variants were compared with the inhibition of the wild-type enzyme. Molecular modelling was used to rationalize the biochemical results.
Inhibitors combining the P2 proline and P1 (1R,2S)-1-amino-2-vinylcyclopropyl-carboxylic acid (vinylACCA) lost much of their potency on the resistant variants. Exchange of the P2 proline for phenylglycine yielded inhibitors that were equipotent on the wild-type and on the A156T and D168V variants. The same result was obtained from the combination of either the P2 residue with a norvaline or an aromatic scaffold in the P1 position.
The combination of a substituted P2 proline and P1 vinylACCA appears to be the main problem behind the observed resistance. Molecular modelling suggests an enforced change in binding conformation for the P2 proline-based inhibitors, whereas the phenylglycine-based inhibitors retained their wild-type binding conformation in the substituted forms of the enzyme.
丙型肝炎病毒（HCV）感染对公众健康构成严重威胁。一个重要的药物靶点是NS3蛋白酶，目前有几种抑制剂正处于临床试验阶段。由于该病毒的高突变率，对任何HCV特异性药物产生耐药性可能会成为一个重大问题。因此，需要主要耐药变体的构效关系数据来指导未来蛋白酶抑制剂的设计。
使用具有已知耐药性替代A156T或D168V的全长NS3蛋白，通过酶活性测定评估含有P2脯氨酸或苯甘氨酸的三肽NS3蛋白酶抑制剂与不同P3和P1-P1'基团组合的抑制效力。将这些变体获得的结果与野生型酶的抑制情况进行比较。利用分子建模对生化结果进行合理化分析。
结合P2脯氨酸和P1（1R,2S）-1-氨基-2-乙烯基环丙基羧酸（乙烯基ACCA）的抑制剂在耐药变体上失去了大部分效力。将P2脯氨酸替换为苯甘氨酸得到的抑制剂对野生型以及A156T和D168V变体具有同等效力。P2残基与正缬氨酸或P1位置的芳香族支架组合也得到了相同的结果。
取代的P2脯氨酸和P1乙烯基ACCA的组合似乎是观察到的耐药性背后的主要问题。分子建模表明，基于P2脯氨酸的抑制剂在结合构象上发生了强制变化，而基于苯甘氨酸的抑制剂在酶的取代形式中保留了其野生型结合构象。
