title: High-throughput identification of promiscuous inhibitors from screening libraries with the use of a thiol-containing fluorescent probe.
abstract: Testing small molecules for their ability to modify cysteine residues of proteins in the early stages of drug discovery is expected to accelerate our ability to develop more selective drugs with lesser side effects. In addition, this approach also enables the rapid evaluation of the mode of binding of new drug candidates with respect to thiol reactivity and metabolism by glutathione. Herein, we describe the development of a fluorescence-based high-throughput assay that allows the identification of thiol-reactive compounds. A thiol-containing fluorescent probe, MSTI, was synthesized and used to evaluate small molecules from the Library of Pharmacologically Active Compounds (LOPAC) collection of bioactive molecules. LOPAC compounds that are known to react with sulfur nucleophiles were identified with this assay, for example, irreversible protease inhibitors, nitric oxide-releasing compounds, and proton-pump inhibitors. The results confirm that both electrophilic and redox reactive compounds can be quickly identified in a high-throughput manner, enabling the assessment of screening libraries with respect to thiol-reactive compounds.
authors: University of Wisconsin, Milwaukee, WI, USA.
authors: Megan M
authors: MM
authors: McCallum
authors: Premchendar
authors: P
authors: Nandhikonda
authors: Jonathan J
authors: JJ
authors: Temmer
authors: Charles
authors: C
authors: Eyermann
authors: Anton
authors: A
authors: Simeonov
authors: Ajit
authors: A
authors: Jadhav
authors: Adam
authors: A
authors: Yasgar
authors: David
authors: D
authors: Maloney
authors: Alexander Leggy
authors: AL
authors: Arnold
mesh_terms: D055808:Drug Discovery / Q000379:methods; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry*; D005978:Glutathione / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000379:methods*; D009569:Nitric Oxide / Q000378:metabolism; D010084:Oxidation-Reduction / Q000187:drug effects; D011480:Protease Inhibitors / Q000494:pharmacology; D054328:Proton Pump Inhibitors / Q000494:pharmacology; D054852:Small Molecule Libraries / Q000494:pharmacology*; D013438:Sulfhydryl Compounds / Q000737:chemistry*
chemical_list: D005456:Fluorescent Dyes; D011480:Protease Inhibitors; D054328:Proton Pump Inhibitors; D054852:Small Molecule Libraries; D013438:Sulfhydryl Compounds; D009569:Nitric Oxide; D005978:Glutathione
keywords: fluorescence; glutathione; high-throughput screening; promiscuous inhibitors; thiol-reactive or electrophilic compound