Establishment of a new cell line inducibly expressing HIV-1 protease for performing safe and highly sensitive screening of HIV protease inhibitors
Establishment of a new cell line inducibly expressing HIV-1 protease for performing safe and highly sensitive screening of HIV protease inhibitors
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DOI：
10.1016/j.micinf.2006.02.016
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发表时间：
2006-06-01
影响因子：
5.8
通讯作者：
Kobayashi, Nobuyuki
中科院分区：
文献类型：
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作者：
Fuse, Takayuki;Watanabe, Ken;Kobayashi, Nobuyuki
人类免疫缺陷病毒 1 型 (HIV-1) 蛋白酶 (PR) 在将病毒多蛋白加工成成熟蛋白方面发挥着重要作用。因此，它是开发抗艾滋病药物的主要目标。然而，由于HIV耐药性的迅速出现，迫切需要开发新型HIV PR抑制剂。我们最近建立了一个新的细胞系E-PR293，它可以作为一种安全、方便、高效的检测系统来筛选HIV-1 PR抑制剂。在细胞中，HIV-1 PR 以与绿色荧光蛋白 (GFP) 的嵌合蛋白的形式表达。该测定测量 PR 活性作为 GFP 荧光或细胞中表达的 HIV-1 PR 细胞毒性活性的函数。 E-PR293 细胞在多西环素存在下维持，多西环素抑制 HIV-1 PR 的表达。强力霉素的去除会诱导HIV-1 PR的表达，用于筛选HIV-1 PR抑制剂。在 E-PR293 细胞中，奈非那韦和沙奎那韦的细胞毒性作用的 50% 抑制浓度低至纳摩尔水平，几乎等于 HIV 感染测定中发现的浓度。 (c) 2006 年爱思唯尔 SAS。版权所有。
The human immunodeficiency virus type 1 (HIV-1) protease (PR) plays an essential role in processing viral polyproteins into mature proteins. As a result, it is a major target for the development of drugs against AIDS. However, due to the rapid emergence of drug-resistant HIV, the development of novel HIV PR inhibitors is urgently needed. We recently established a new cell line E-PR293 which can be used as a safe, convenient and highly efficient assay system to screen HIV-1 PR inhibitors. In the cells, the HIV-1 PR is expressed in a chimeric protein with the green fluorescence protein (GFP). This assay measures the PR activity as a function of either the fluorescence of GFP or the cytotoxic activity of HIV-1 PR which is expressed in the cell. E-PR293 cells were maintained in the presence of doxycycline, which suppresses the expression of HIV-1 PR. The removal of doxycycline induces the expression of HIV-1 PR, which is used to screen HIV-1 PR inhibitors. In E-PR293 cells, the 50% inhibitory concentration of the cytotoxic effects by nelfinavir and saquinavir were as low as nanomolar levels, almost equal to those found in the HIV-infection assay. (c) 2006 Elsevier SAS. All rights reserved.