Document 0715
 DOCN  M9650715
 TI    Anti-Tat MTT assay: a novel anti-HIV drug screening system using the
       viral regulatory network of replication.
 DT    9605
 AU    Kira T; Merin JP; Baba M; Shigeta S; Okamoto T; Department of
       Microbiology, Fukushima Medical College, Japan.
 SO    AIDS Res Hum Retroviruses. 1995 Nov;11(11):1359-66. Unique Identifier :
       AIDSLINE MED/96159133
 AB    Since the recognition of its pivotal role in viral replication, Tat
       activity has become an interesting target for chemotherapeutic
       intervention of HIV infection. Here, we report a sensitive and simple
       colorimetric assay for the screening of Tat inhibitors. We have
       constructed a plasmid that contains the hygromycin B phosphotransferase
       gene under the control of the HIV-1 long terminal repeat (LTR) and HIV-1
       tat gene constitutively expressed from the cytomegalovirus promoter.
       This plasmid has been stably transfected to the CD4+ T cell line CEM,
       which is rendered resistant to hygromycin B through the action of Tat.
       The inhibitory activity of the anti-Tat drugs was assessed by the extent
       of cytotoxicity in the presence of hygromycin B as a consequence of the
       suppressed expression of the hygromycin B phosphotransferase gene.
       Spectrophotometric quantitation of cell viability was done utilizing
       3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) dye
       as the indicator. Using this assay system, we have confirmed that known
       anti-Tat compound Ro5-3335 and its derivative Ro24-7429 could inhibit
       Tat-mediated gene expression although their selectivities (anti-Tat
       activity versus nonselective cytotoxicity) were narrow. Since this
       method offers the advantage of not handling infectious particles or
       radioactive materials, it can offer wide applicability as a screening
       system for anti-Tat compounds.
 DE    Antiviral Agents/*PHARMACOLOGY  Base Sequence  Cell Line  Colorimetry
       Drug Screening/*METHODS  Dyes  DNA Primers  Gene Products,
       tat/*ANTAGONISTS & INHIB  Human  Hygromycin B/ANALOGS &
       DERIVATIVES/PHARMACOLOGY  HIV-1/*DRUG EFFECTS/PHYSIOLOGY  Molecular
       Sequence Data  Support, Non-U.S. Gov't  Tetrazolium Salts  Thiazoles
       Virus Replication/DRUG EFFECTS  JOURNAL ARTICLE

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