拉米夫定耐药慢性乙型肝炎患者的HBV逆转酶区突变模式

目的研究慢性乙型肝炎患者在拉米夫定（LAM）治疗过程中出现耐药后，HBV逆转录酶区基因的突变模式及临床特征。方法采用巢式PCR方法对2007年4月至2010年12月于沈阳市第六人民医院肝病门诊或住院诊治的共260例慢性乙型肝炎LAM耐药患者的HBV聚合酶基因逆转录酶区进行扩增，对PCR产物进行直接测序，回顾性分析LAM耐药时HBV聚合酶基因的不同突变模式及患者的临床特征。结果260例患者诊断为LAM耐药，215例患者检测到LAM相关的HBV聚合酶基因突变。患者血清HBVDNA为（5．41±1．29）log10拷贝／ml。98．1％（211／215）患者存在YMDD基序突变。其中3种主要突变类型分别为：单位点rtM204I突变占40．0％（86／215）；rtL180M＋rtM204V占37．2％（80／215）；rtL180M＋rtM204I占13．0％（28／215）。与rtM2041相比，rtM204V多以联合rtL180M突变的形式存在（P〈0．05）。LAM耐药时，单位点突变与联合突变类型患者的血清HBVDNA、ALT、年龄、HBeAgFH性与阴性，肝硬化与慢性乙型肝炎差异均无统计学意义（P〉0．05）。测出基因突变患者与伴生化学突破患者HBVDNA载量较高（P〈0．05）。结论YMDD基序突变是LAM耐药后HBV聚合酶基因突变的主要模式，LAM耐药后患者临床病情轻重可能与HBV聚合酶基因突变类型无关。

The mutation patterns of HBV polymerase gene in chronic hepatitis B patients with lamivudine resistant

Objective To investigate the clinical features and mutation patterns of HBV polymerase gene in patients with chronic hepatitis B （CHB） after the emergence of drug-resisance during lamivudine （LAM） therapy. Methods The HBVP gene RT region from 260 serum samples was amplified by nest PCR, and PCR products were directly sequenced. The outpatients and hospitalized CHB patients with LAM- resistant in the No. 6 People＇s Hospital of Shenyang from April 2007 to December 2009 were collected. Clinical features after the emergence of LAM-resistant mutations were analyzed, retrospectively, Results Total of 260 patients with CHB were diagnosed as LAM-resistant. Among them 215 patients were found to had LAM-resistant-associated mutations in the polymerase gene. The median of serum HBV DNA was （5.41 ± 1.29） log10 copies/ml. There were 98.1% （211/215） patients had YMDD mutations. Three major mutation patterns of LAM-resistant HBV were identified as rtM204I with 40.0% （86/215）, rtL180M ＋ rtM204V with 37.2% （80/215） and rtL180M ＋ rtM204I with 13.0% （28/215）. The rtM204V mutation was accompanied more frequently by the rtL180M mutation compared with the rtM204I mutation （P 〈 0.05）. There were no significant difference in ALT or HBV DNA levels and age, gender, HBeAg status, cirrhosis or not were found among patients with one point mutation and joint mutation patterns （P 〈 0.05）. HBV viral load was higher in patients who were detected gene mutation and accompanied with biochemical breakthrough. Conclusions YMDD is the major mutation pattern of HBV polymerase gene after emergence of LAM-resistant. The mutation patterns of HBV polymerase gene are possibly not related to the clinical severity of CHB patients during LAM therapy.