拉米夫定治疗慢性乙型肝炎过程中HBV发生YMDD变异的检测

Detection of YMDD mutation in HBV polymerase gene during the lamivudine treatment for the patients with hepatitis B virus infection

目的建立乙型肝炎病毒Ｐ基因区ＹＭＤＤ变异的检测方法，对用拉米夫定治疗５２周出现耐药的病人进行检测。方法从８８例治疗病人中选择１０例服用该药后ＨＢＶＤＮＡ转阴（＜１．６ｐｇ／ｍｌ），５２周时又反跳≥２０ｐｇ／ｍｌ的病人，用 ＰＣＲ后直接测序的方法，分别对 ０周和 ５２周标本进行 ＨＢＶ ＤＮＡ聚合酶活性区域的测序。结果 １０例服药５２周后ＨＢＶ ＤＮＡ反跳病人中７例测得ＨＢＶ ＹＭＤＤ变异。其变异分为两种类型：第一种类型为ＹＶＤＤ有５例，有核苷酸Ａ７４１→Ｇ，均伴有Ａ６６９→Ｃ的变异。第二种类型为ＹＩＤＤ有２例，有核苷酸Ｇ７４３→Ｔ，其中１例还有Ｔ７８１→Ｃ。结论少数病人在拉米夫定治疗过程中出现耐药，可能和ＨＢＶ ＹＭＤＤ变异有关。

Objective To detect YMDD mutation in HBV polymerase gene region of the patients with HBV infection during lamivudine therapy. Methods PCR and sequencing methods were used to determine this mutation in 10 of 88 treatment patients, which HBV DNA were elevated more than 20 pg/ml at 52 week from previous undetectable level(1 .6pg/ml). Results Analysis of gene sequence indicated that amino acid subshtutions in YNDD motif of HBV polymerase gene occurred in HBV DNA of 7 patients.The amino acid mutation divided two patterns:(1)YMDD → YVDD mutation were detected in 5 of specimens, which nucleotide changed from A^741 → G, all accompanied with A^669 → C, (2) YMDD → YIDD mutation were found in the remainder 2 patients, of which the nucleotide changed from G^743 → T,one of them accompanied by T^781 → C. Conclusion The result proved that the development of HBV resistance to lamivudine was related to YMDD mutation in HBV P gene region during lamivudine therapy.