Department of Therapeutic Research and Medicines Evaluation, Istituto Superiore di Sanità, Rome, Italy.
J Virol Methods. 2010 Sep;168(1-2):272-6. doi: 10.1016/j.jviromet.2010.06.004. Epub 2010 Jun 15.
Macrophages represent an important site for productive infection of HIV-1 and the evaluation of integrase (IN) inhibitors on this cell subset is of fundamental importance. In this report, preclinical evaluation of IN inhibitors on primary human macrophages was attempted successfully using a 96-well microtiter phenotypic assay developed recently for the evaluation of IN inhibitors in a cell-based system by taking advantage of HIV-derived lentiviral vectors expressing luciferase. IN inhibitors were also tested using a lentiviral vector containing an IN with introduced T66I/S153Y mutations, known to affect the activity of azido-group-containing diketo acid (DKA) IN inhibitors. Utilizing different classes of HIV integrase inhibitors against the wild-type IN and the mutant mentioned above, some of the IN inhibitors used were also active on this particular mutant, suggesting that should HIV-1 develop additional or different mutations to become resistant to such anti-IN drugs, new drugs can be developed with a better resistance profile. This assay provides a standardized method for the preclinical evaluation of the efficacy of IN inhibitors on wild-type and mutated IN that can be adapted easily for the evaluation of anti-IN activity on IN sequences derived from patients.
巨噬细胞是 HIV-1 有效感染的重要部位，评估整合酶（IN）抑制剂对这个细胞亚群具有重要意义。本报告中，我们成功地利用最近开发的基于细胞的 IN 抑制剂检测方法，通过利用表达荧光素酶的 HIV 衍生慢病毒载体，在 96 孔微量滴定板表型测定中，对原发性人巨噬细胞中的 IN 抑制剂进行了临床前评估。我们还使用含有 IN 引入 T66I/S153Y 突变的慢病毒载体对 IN 抑制剂进行了测试，这些突变已知会影响含有叠氮基团的二酮酸（DKA）IN 抑制剂的活性。利用针对野生型 IN 和上述突变体的不同类别的 HIV 整合酶抑制剂，一些 IN 抑制剂对该特定突变体也具有活性，这表明如果 HIV-1 发生更多或不同的突变以对这类抗 IN 药物产生耐药性，那么可以开发具有更好耐药谱的新药。该测定法为评估 IN 抑制剂对野生型和突变型 IN 的疗效提供了一种标准化方法，该方法可以很容易地适应于评估来自患者的 IN 序列的抗 IN 活性。
