title: Fluorometric assay for phenotypic differentiation of drug-resistant HIV mutants.
abstract: Convenient drug-resistance testing of viral mutants is indispensable to effective treatment of viral infection. We developed a novel fluorometric assay for phenotypic differentiation of drug-resistant mutants of human immunodeficiency virus-I protease (HIV-PR) which uses enzymatic and peptide-specific fluorescence (FL) reactions and high-performance liquid chromatography (HPLC) of three HIV-PR substrates. This assay protocol enables use of non-purified enzyme sources and multiple substrates for the enzymatic reaction. In this study, susceptibility of HIV mutations to drugs was evaluated by selective formation of three FL products after the enzymatic HIV-PR reaction. This proof-of-concept study indicates that the present HPLC-FL method could be an alternative to current phenotypic assays for the evaluation of HIV drug resistance.
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Qinchang
authors.initials: Q
authors.lastname: Zhu
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Zhiqiang
authors.initials: Z
authors.lastname: Yu
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Kabashima
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Sheng
authors.initials: S
authors.lastname: Yin
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Shpend
authors.initials: S
authors.lastname: Dragusha
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Ahmed F M
authors.initials: AF
authors.lastname: El-Mahdy
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Valon
authors.initials: V
authors.lastname: Ejupi
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Shibata
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14, Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Masaaki
authors.initials: M
authors.lastname: Kai
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D024882:Drug Resistance, Viral / Q000235:genetics*; D005470:Fluorometry; D015658:HIV Infections / Q000188:drug therapy; D016333:HIV Protease / Q000235:genetics*; D017320:HIV Protease Inhibitors / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D019469:Indinavir / Q000494:pharmacology; D061466:Lopinavir / Q000494:pharmacology; D008826:Microbial Sensitivity Tests / Q000379:methods*; D019438:Ritonavir / Q000494:pharmacology; D019258:Saquinavir / Q000494:pharmacology
chemical_list: D019380:Anti-HIV Agents; D017320:HIV Protease Inhibitors; D061466:Lopinavir; D019469:Indinavir; D016333:HIV Protease; D019258:Saquinavir; D019438:Ritonavir