title: Establishment of a new cell line inducibly expressing HIV-1 protease for performing safe and highly sensitive screening of HIV protease inhibitors.
abstract: 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.
authors.affiliation: Laboratory of Molecular Biology of Infectious Agents, Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-machi, Nagasaki, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Fuse
authors.forename: Ken
authors.initials: K
authors.lastname: Watanabe
authors.forename: Kaio
authors.initials: K
authors.lastname: Kitazato
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Kobayashi
mesh_terms: D002460:Cell Line; D004318:Doxycycline / Q000494:pharmacology; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D016333:HIV Protease / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D017320:HIV Protease Inhibitors / Q000494:pharmacology*; D006801:Humans; D019888:Nelfinavir / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D019258:Saquinavir / Q000494:pharmacology; D012680:Sensitivity and Specificity
chemical_list: D017320:HIV Protease Inhibitors; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D016333:HIV Protease; D019888:Nelfinavir; D019258:Saquinavir; D004318:Doxycycline