title: Discovery of 5-benzyl-3-phenyl-4,5-dihydroisoxazoles and 5-benzyl-3-phenyl-1,4,2-dioxazoles as potent firefly luciferase inhibitors.
abstract: Luciferase reporter assays are commonly used in high-throughput screening methods. Here, we report new firefly luciferase (FLuc) inhibitors based on 5-benzyl-3-phenyl-4,5-dihydroisoxazoles and 5-benzyl-3-phenyl-1,4,2-dioxazoles, which showed up as "false positives" in a luciferase reporter gene-based assay for nuclear receptor antagonists. The inhibition was shown to be noncompetitive for both natural enzyme substrates (d-luciferin and ATP) and selective to FLuc and proven to arise from a direct interaction between the enzyme and the inhibitor. Of the 63 evaluated compounds, 28 showed significantly better inhibition potency than the well-known inhibitor resveratrol (IC(50) = 59 nM), with five compounds having distinctly subnanomolar IC(50) values. The most efficient compounds inhibited the luminescence at concentrations lower than (1)/(100) in comparison to resveratrol (lowest IC(50) = 0.26 nM) and can thus be considered to belong to the most potent FLuc inhibitors reported thus far. Overall, the novel inhibitors form a unique molecular library for structure-activity relationship (SAR) analyses.
authors.affiliation: School of Pharmacy, University of Eastern Finland , P.O. Box 1627, FI-70211 Kuopio, Finland.
authors.forename: Pekka K
authors.initials: PK
authors.lastname: Poutiainen
authors.forename: Jorma J
authors.initials: JJ
authors.lastname: Palvimo
authors.forename: Ari E
authors.initials: AE
authors.lastname: Hinkkanen
authors.forename: Arto
authors.initials: A
authors.lastname: Valkonen
authors.forename: Topi K
authors.initials: TK
authors.lastname: Väisänen
authors.forename: Reino
authors.initials: R
authors.lastname: Laatikainen
authors.forename: Juha T
authors.initials: JT
authors.lastname: Pulkkinen
mesh_terms: D000818:Animals; D001393:Azoles / Q000737:chemistry* / Q000494:pharmacology*; D002460:Cell Line; D055808:Drug Discovery; D004353:Drug Evaluation, Preclinical; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology*; D048888:Fireflies / Q000201:enzymology; D020128:Inhibitory Concentration 50; D008156:Luciferases / Q000037:antagonists & inhibitors*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular
chemical_list: D001393:Azoles; D004791:Enzyme Inhibitors; D008156:Luciferases