Department of Immunology & Microbial Science, The Scripps Research Institute, La Jolla, California, USA.
Antimicrob Agents Chemother. 2012 Oct;56(10):5113-21. doi: 10.1128/AAC.00919-12. Epub 2012 Jul 16.
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.
阿利西尤单抗是目前处于 III 期研究阶段的最先进的靶向宿主抗病毒环孢素（Cyp）抑制剂，在降低感染患者的丙型肝炎病毒（HCV）病毒血症方面显示出巨大的潜力。为了进一步阐明阿利西尤单抗的作用机制，筛选出对该药物具有抗性的 HCV 复制子。有趣的是，NS5A 结构域 II 中的突变不断出现。为了证明这些突变是导致耐药的原因，将它们重新引入亲本 HCV 基因组中，并在存在阿利西尤单抗或不存在阿利西尤单抗靶标 CypA 的情况下，测试由此产生的突变病毒的复制情况。我们还检查了突变对 NS5A 自身结合（寡聚化）、CypA、RNA 和 NS5B 的影响。重要的是，这些突变不影响任何这些相互作用。此外，这些突变并不能使 NS5A-CypA 相互作用免受阿利西尤单抗的破坏。NS5A 突变本身使 HCV 对阿利西尤单抗的耐药性仅略有增加。相比之下，当多个 NS5A 突变结合时，观察到明显的耐药性。在 NS5A 中引入多个突变显著恢复了 CypA 敲低细胞中的病毒复制。有趣的是，NS5A 突变的组合使 HCV 对所有 Cyp 抑制剂均具有耐药性。这项研究表明，需要在 NS5A 的 II 结构域中结合多个突变，而不是单个突变，才能使 HCV 对 Cyp 抑制剂具有显著且普遍的耐药性。这与体内数据一致，表明阿利西尤单抗与病毒耐药性发展的低潜力相关。
