Hearps Anna C, Greengrass Vicki, Hoy Jennifer, Crowe Suzanne M
Clinical Research Laboratory, Centre for Virology, Burnet Institute for Medical Research and Public Health, Melbourne, Vic. 3004, Australia.
Sex Health. 2009 Dec;6(4):305-9. doi: 10.1071/SH09041.
The integrase inhibitors (e.g. Raltegravir) are a new class of antiretroviral drugs that have recently become available for the treatment of patients with multi-drug resistant HIV-1 within Australia. The emergence of mutations that confer resistance to the integrase inhibitors has been observed in vivo; however, no commercial genotyping assay is currently available to screen for resistance to these drugs.
The HIV-1 integrase gene was amplified from plasma-derived HIV-1 viral RNA via reverse transcription-polymerase chain reaction and genotype determined via population DNA sequencing. Drug resistance mutations and polymorphisms were detected using the Stanford University online HIV database. Assay sensitivity and reproducibility were determined using clinical and laboratory-derived samples.
Our in-house assay was capable of genotyping the integrase gene from all samples tested (n = 30) of HIV-1 subtypes B, C, D, F, CFR01_AE and CRF02_AG and can amplify the integrase region from plasma samples containing as few as 50 HIV RNA copies/mL. The assay is highly reproducible (average nucleotide concordance = 99.6%, n = 4) and is capable of detecting resistance-associated mutations.
This assay is suitable for routine drug resistance screening of plasma samples from HIV-infected patients receiving integrase inhibitor antiretroviral drugs and also serves as a useful research tool.
整合酶抑制剂（如雷特格韦）是一类新型抗逆转录病毒药物，最近在澳大利亚已可用于治疗多重耐药HIV-1患者。体内已观察到对整合酶抑制剂产生耐药性的突变的出现；然而，目前尚无商业基因分型检测方法可用于筛查对这些药物的耐药性。
通过逆转录-聚合酶链反应从血浆来源的HIV-1病毒RNA中扩增HIV-1整合酶基因，并通过群体DNA测序确定基因型。使用斯坦福大学在线HIV数据库检测耐药性突变和多态性。使用临床和实验室来源的样本确定检测灵敏度和可重复性。
我们的内部检测方法能够对所有检测样本（n = 30）中的HIV-1 B、C、D、F、CFR01_AE和CRF02_AG亚型的整合酶基因进行基因分型，并且能够从每毫升仅含50个HIV RNA拷贝的血浆样本中扩增整合酶区域。该检测方法具有高度可重复性（平均核苷酸一致性 = 99.6%，n = 4），并且能够检测与耐药性相关的突变。
该检测方法适用于对接受整合酶抑制剂抗逆转录病毒药物治疗的HIV感染患者的血浆样本进行常规耐药性筛查，也是一种有用的研究工具。
