Department of Medicine and Molecular Science, Division of Frontier Medical Science, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima, Japan.
Hepatology. 2011 Sep 2;54(3):781-8. doi: 10.1002/hep.24460. Epub 2011 Aug 2.
Telaprevir is a potent inhibitor of hepatitis C virus (HCV) NS3-4A protease. However, the emergence of drug-resistant strains during therapy is a serious problem, and the susceptibility of resistant strains to interferon (IFN), as well as the details of the emergence of mutant strains in vivo, is not known. We previously established an infectious model of HCV using human hepatocyte chimeric mice. Using this system we investigated the biological properties and mode of emergence of mutants by ultra-deep sequencing technology. Chimeric mice were injected with serum samples obtained from a patient who had developed viral breakthrough during telaprevir monotherapy with strong selection for resistance mutations (A156F [92.6%]). Mice infected with the resistant strain (A156F [99.9%]) developed only low-level viremia and the virus was successfully eliminated with interferon therapy. As observed in patients, telaprevir monotherapy in viremic mice resulted in breakthrough, with selection for mutations that confer resistance to telaprevir (e.g., a high frequency of V36A [52.2%]). Mice were injected intrahepatically with HCV genotype 1b clone KT-9 with or without an introduced resistance mutation, A156S, in the NS3 region, and treated with telaprevir. Mice infected with the A156S strain developed lower-level viremia compared to the wildtype strain but showed strong resistance to telaprevir treatment. Although mice injected with wildtype HCV showed a rapid decline in viremia at the beginning of therapy, a high frequency (11%) of telaprevir-resistant NS3 V36A variants emerged 2 weeks after the start of treatment.
Using deep sequencing technology and a genetically engineered HCV infection system, we showed that the rapid emergence of telaprevir-resistant HCV was induced by mutation from the wildtype strain of HCV in vivo.
特拉匹韦是一种有效的丙型肝炎病毒（HCV）NS3-4A 蛋白酶抑制剂。然而，在治疗过程中出现耐药株是一个严重的问题，耐药株对干扰素（IFN）的敏感性以及体内突变株出现的细节尚不清楚。我们之前使用人源肝细胞嵌合小鼠建立了 HCV 感染模型。利用该系统，我们通过超深度测序技术研究了突变株的生物学特性和出现方式。嵌合小鼠被注射来自于一名患者的血清样本，该患者在接受特拉匹韦单药治疗时出现病毒突破，对耐药突变有强烈选择（A156F[92.6%]）。感染耐药株（A156F[99.9%]）的小鼠仅出现低水平病毒血症，用干扰素治疗可成功清除病毒。与患者观察结果一致，在有病毒血症的小鼠中单用特拉匹韦治疗导致突破，出现对特拉匹韦耐药的突变（例如，V36A 高频出现[52.2%]）。将 HCV 基因型 1b 克隆 KT-9 或在 NS3 区引入耐药突变 A156S 注射到小鼠肝内，并给予特拉匹韦治疗。与野生型株相比，感染 A156S 株的小鼠病毒血症水平较低，但对特拉匹韦治疗有很强的耐药性。虽然感染野生型 HCV 的小鼠在治疗开始时病毒血症迅速下降，但在治疗开始 2 周后出现了高频（11%）的对特拉匹韦耐药的 NS3 V36A 变异体。
利用深度测序技术和基因工程 HCV 感染系统，我们表明体内野生型 HCV 突变快速诱导了对特拉匹韦耐药的 HCV 的出现。
