title: The identification and biochemical characterization of drug-like compounds that inhibit botulinum neurotoxin serotype A endopeptidase activity.
abstract: A robust, high-throughput, two-tiered assay for screening small molecule inhibitors against botulinum neurotoxin serotype A was developed and employed to screen 16,544 compounds. Thirty-four compounds were identified as potent hits employing the first-tier assay. Subsequently, nine were confirmed as actives by our second-tier confirmatory assay. Of these, one displayed potent inhibitory efficacy, possessing an IC(50)=16 microM (+/-1.6 microM) in our in vitro assay. This inhibitor (0831-1035) is highly water-soluble, and possesses an IC(50)=47 microM (+/-7.0 microM) in our primary cell culture assay (with virtually no cytotoxicity up to 500 microM), suggesting that this inhibitor is a good candidate for further development as a therapeutic countermeasure to treat botulism resulting from botulinum neurotoxin serotype A intoxication. An enzyme kinetics study indicated that this inhibitor exhibits mixed non-competitive inhibition, with a K(I)=9 microM.
authors.affiliation: Botulinum Research Center, and Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, 285 Old Westport Road, North Dartmouth, MA 02747, USA.
authors.forename: Shuowei
authors.initials: S
authors.lastname: Cai
authors.forename: Paul
authors.initials: P
authors.lastname: Lindo
authors.forename: Jong-Beak
authors.initials: JB
authors.lastname: Park
authors.forename: Kruti
authors.initials: K
authors.lastname: Vasa
authors.forename: Bal Ram
authors.initials: BR
authors.lastname: Singh
mesh_terms: D000818:Animals; D015153:Blotting, Western; D019274:Botulinum Toxins, Type A / Q000037:antagonists & inhibitors*; D010450:Endopeptidases / Q000187:drug effects / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D031541:Fluorescence Resonance Energy Transfer; D007700:Kinetics; D011480:Protease Inhibitors / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D011480:Protease Inhibitors; D010450:Endopeptidases; D019274:Botulinum Toxins, Type A