拉米夫定与恩替卡韦序贯治疗中的乙型肝炎病毒准种动力学

目的 研究HBV准种群体在拉米夫定(LAM)与恩替卡韦(ETV)序贯治疗过程中的动态演变及其临床意义. 方法 对2例采用LAM-ETV序贯治疗出现不同临床结果的患者进行了近4年的随访,用多聚酶链反应-克隆-测序的方法研究HBV准种组成的长程动态演变,用最大似然法建立遗传进化树分析代表序列的遗传进化关系,结合血清学和病毒学指标分析HBV准种演变与临床过程的关系. 结果 2例患者均出现了LAM耐药病毒学突破,采用ETV治疗后,1例患者获得了持续病毒学应答,另一例患者在ETV治疗72周时又出现病毒学突破.病毒学突破均发生在原有对药物敏感的优势准种被耐药株替代时,ETV耐药与rtL180M+S202G+M204V三联突变株成为优势准种密切相关,准种动力学和遗传进化分析结果提示LAM治疗筛选出的rtL180M合并M204V联合突变株在HBV种群中的累积与此三联突变株的出现有直接关系.结论 HBV准种组成与核苷类药物敏感性密切相关,LAM耐药株可进一步演化为ETV耐药株,对使用核苷类药物治疗的患者进行HBV准种监测具有重要临床意义.

Evolution of hepatitis B virus quasispecies during lamivudine-entecavir sequential therapy

Objective To study the evolution of HBV quasispecies under the pressures of lamivudine (LAM) - entecavir (ETV) sequential therapy and its clinical significance. Methods Consecutive serum samples from 2 patients underwent LAM-ETV sequential therapy were extensively studied for HBV quasispecies composition and evolution, using PCR-cloning-sequencing method. Maximum likelihood trees were built to analyze the genetic relationship between representative sequences. Correlation between HBV quasispecies evolution and serological/virological data was analyzed to determined the clinical significance of the evolution of HBV quasispecies during prolonged nucleotide analog therapy. Results Virological breakthrough was observed in both patients. Patient I acquired sustained virological response after switching to ETV rescue therapy, whereas Patient II suffered from virological breakthrough after 72 weeks of ETV therapy. Each virological breakthrough was accompanied with the replacement of previous drug susceptible dominant quasispecies with a drug resistant variant, indicating a close correlation between quasispecies composition and drug susceptibility. The rtL180M+S202G+M204V triple mutant, which was most likely a descendant of the LAM resistant rtL180M+M204V variant, was closely correlated with ETV resistant in Patient II. Conclusions Quasispecies composition of HBV is closely correlated with nucleotide analog susceptibility. ETV resistant variant can emerge from a LAM resistant viral population. Dynamic monitoring of HBV quasispecies composition is of great importance during nucleotide analog therapy.