Evolution of multi-drug resistant HCV clones from pre-existing resistant-associated variants during direct-acting antiviral therapy determined by third-generation sequencing.
Evolution of multi-drug resistant HCV clones from pre-existing resistant-associated variants during direct-acting antiviral therapy determined by third-generation sequencing.
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DOI：
10.1038/srep45605
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发表时间：
2017-03-31
影响因子：
4.6
通讯作者：
Marusawa H
中科院分区：
文献类型：
--
作者：
Takeda H;Ueda Y;Inuzuka T;Yamashita Y;Osaki Y;Nasu A;Umeda M;Takemura R;Seno H;Sekine A;Marusawa H
耐药相关变异(RAV)是直接作用抗病毒药物(DAA)治疗丙型肝炎病毒感染患者面临的最大临床挑战之一。我们使用第三代测序技术研究了接受DAA治疗的患者的病毒动力学。在283例1b型丙型肝炎患者中，32例(11.3%)未能达到持续病毒学应答( + )，其中32例(11.3%)未能达到持续病毒学应答。对104例患者(32例非SVR，72例SVR)进行常规超深测序，在所有非SVR患者中检测到具有代表性的RAV，其中28例(87.5%)检测到Y93H。通过第三代测序确定了来自6个有代表性的非SVR患者的12份血清中每个病毒克隆的NS3至NS5A区域的长连续序列，并显示在治疗失败时，在先前存在的RAV和优势分离株的一个亚群中，同时存在几个与耐药性相关的替换相关的同义突变。系统发育分析显示，在治疗失败时，先前存在的RAV和显性RAV之间的遗传距离很近。此外，在所有非SVR病例中，在DCV/ASV之后，先前存在的RAV上出现了多个耐药突变。总之，治疗失败时的多重耐药病毒克隆肯定起源于丙型肝炎病毒感染患者中先前存在的RAV的一个亚群。在DAA处理压力下，这些RAV被选中，并随着获得多个与抗性相关的替换而成为主导。
Resistance-associated variant (RAV) is one of the most significant clinical challenges in treating HCV-infected patients with direct-acting antivirals (DAAs). We investigated the viral dynamics in patients receiving DAAs using third-generation sequencing technology. Among 283 patients with genotype-1b HCV receiving daclatasvir + asunaprevir (DCV/ASV), 32 (11.3%) failed to achieve sustained virological response (SVR). Conventional ultra-deep sequencing of HCV genome was performed in 104 patients (32 non-SVR, 72 SVR), and detected representative RAVs in all non-SVR patients at baseline, including Y93H in 28 (87.5%). Long contiguous sequences spanning NS3 to NS5A regions of each viral clone in 12 sera from 6 representative non-SVR patients were determined by third-generation sequencing, and showed the concurrent presence of several synonymous mutations linked to resistance-associated substitutions in a subpopulation of pre-existing RAVs and dominant isolates at treatment failure. Phylogenetic analyses revealed close genetic distances between pre-existing RAVs and dominant RAVs at treatment failure. In addition, multiple drug-resistant mutations developed on pre-existing RAVs after DCV/ASV in all non-SVR cases. In conclusion, multi-drug resistant viral clones at treatment failure certainly originated from a subpopulation of pre-existing RAVs in HCV-infected patients. Those RAVs were selected for and became dominant with the acquisition of multiple resistance-associated substitutions under DAA treatment pressure.