Schinazi R F, Schlueter-Wirtz S, Stuyver L
Department of Veterans Affairs, Decatur, GA, USA.
Antivir Chem Chemother. 2001;12 Suppl 1:61-5.
A growing concern in the pursuit of new therapies for HIV-1 infection is the potential for the virus to develop drug resistance. With the advent of modern antiretroviral therapy and the common use of combined modalities, it is difficult to identify in the clinic the mutations associated with a specific drug. In general, drug selection of mutants using a relevant cell system, such as primary human lymphocytes, is a good prognosticator of what will happen in humans. In this study, HIV-infected human peripheral blood mononuclear cells were exposed, at a concentration of 1- to 10-fold the median effective antiviral concentration, to the nucleosides (-)-beta-2',3'-dideoxy-3'-thia-5-fluorocytidine [(-)-FTC] (-)-beta-2',3'-dideoxy-3'-thiacytidine (3TC), 3'-azido-2',3'-dideoxyuridine (CS-87, AZDU), 3'-azido-2',3'-dideoxy-5-methylcytidine (CS-92, AZMC), 2',3'-didehydro-3'-deoxythymidine (d4T), beta-L-2',3'-didehydro-2',3'-dideoxy-5-fluorocytidine (beta-L-D4FC), beta-L-2',3'-dideoxyadenine SATE[beta-L-ddAMP-bis(tbutylSATE)], beta-L-5-fluoro-2',3'-dideoxycytidine (L-FddC), and the protease inhibitors nelfinavir and amprenavir (VX-478). Virus from the culture supernatant was amplified by PCR and analysed by both HIV-1 reverse transcriptase and protease line probe assay. All the L-nucleoside analogues tested selected for the V184 mutation, including the L-pyrimidine nucleosides 3TC (-)-FTC, beta- L-FddC, beta-L-D4FC and the beta-L-purine nucleoside. beta-L-D4FC also selected for K/R65 in addition to V184, indicating that these two mutations are linked and compatible in vitro. No pattern of mutations leading to resistance or reduced susceptibility was discerned with d4T. Rapid genotyping analysis revealed the different kinetics and mutations obtained by in vitro selection in HIV-infected cells exposed to nucleoside analogues and protease inhibitors.
在寻求治疗HIV-1感染的新疗法过程中，一个日益受到关注的问题是病毒产生耐药性的可能性。随着现代抗逆转录病毒疗法的出现以及联合治疗方式的普遍使用，在临床上很难确定与特定药物相关的突变。一般来说，使用相关细胞系统（如原代人淋巴细胞）对突变体进行药物筛选，是预测人体中将会发生什么情况的良好指标。在本研究中，将感染HIV的人外周血单核细胞暴露于浓度为中位有效抗病毒浓度1至10倍的核苷（-）-β-2'，3'-二脱氧-3'-硫杂-5-氟胞苷[（-）-FTC]、（-）-β-2'，3'-二脱氧-3'-硫代胞苷（3TC）、3'-叠氮基-2'，3'-二脱氧尿苷（CS-87，AZDU）、3'-叠氮基-2'，3'-二脱氧-5-甲基胞苷（CS-92，AZMC）、2'，3'-二脱氢-3'-脱氧胸苷（d4T）、β-L-2'，3'-二脱氢-2'，3'-二脱氧-5-氟胞苷（β-L-D4FC）、β-L-2'，3'-二脱氧腺嘌呤SATE[β-L-ddAMP-双（叔丁基SATE）]、β-L-5-氟-2'，3'-二脱氧胞苷（L-FddC）以及蛋白酶抑制剂奈非那韦和安普那韦（VX-478）。通过PCR扩增培养上清液中的病毒，并通过HIV-1逆转录酶和蛋白酶线性探针检测进行分析。所有测试的L-核苷类似物都选择了V184突变，包括L-嘧啶核苷3TC、（-）-FTC、β-L-FddC、β-L-D4FC以及β-L-嘌呤核苷。除了V184之外，β-L-D4FC还选择了K/R65，这表明这两个突变在体外是相关且兼容的。未发现d4T导致耐药性或敏感性降低的突变模式。快速基因分型分析揭示了在暴露于核苷类似物和蛋白酶抑制剂的HIV感染细胞中通过体外选择获得的不同动力学和突变情况。
