title: Cell-based screen of HMG-CoA reductase inhibitors and expression regulators using LC-MS.
abstract: We present an integrated consolidation of previously reported methods for screening hydroxymethylglutaryl-coenzyme A reductase (HMGR) inhibitors in 96-well microtiter plates with rapid workup using established mammalian cell lines and liquid chromatography-mass spectrometry analysis. Inhibitors as well as expression regulators of HMGR (inducers or repressors) can be screened. To validate the method, three competitive inhibitors of HMGR (lovastatin, simvastatin, and atorvastatin), as well as a potent sterol repressor of HMGR synthesis (25-hydroxycholesterol), were assayed on two cell lines: HepG2, a human hepatic derived cell line, and L cells, a subline of NCTC clone 929 mouse fibroblasts. The direct inhibition of HMGR by statins, induction of HMGR synthesis by the same statins following incubation with the cells, and repression of HMGR synthesis by 25-hydroxycholesterol were confirmed.
authors.affiliation: Department of Chemical Engineering, 201 Gilman Hall, University of California, Berkeley, CA 94720-1462, USA.
authors.forename: Raphaele
authors.initials: R
authors.lastname: Gerber
authors.forename: Jessica D
authors.initials: JD
authors.lastname: Ryan
authors.forename: Douglas S
authors.initials: DS
authors.lastname: Clark
mesh_terms: D000069059:Atorvastatin; D001681:Biological Assay / Q000379:methods*; D002138:Calibration; D002460:Cell Line; D002853:Chromatography, Liquid; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic; D006538:Heptanoic Acids / Q000032:analysis / Q000494:pharmacology; D006801:Humans; D006888:Hydroxycholesterols / Q000032:analysis; D006903:Hydroxymethylglutaryl CoA Reductases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000032:analysis*; D008148:Lovastatin / Q000032:analysis / Q000494:pharmacology; D013058:Mass Spectrometry; D008798:Mevalonic Acid / Q000031:analogs & derivatives* / Q000032:analysis; D011758:Pyrroles / Q000032:analysis / Q000494:pharmacology; D019821:Simvastatin / Q000032:analysis / Q000494:pharmacology
chemical_list: D006538:Heptanoic Acids; D006888:Hydroxycholesterols; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D011758:Pyrroles; C015367:mevalonolactone; C007997:25-hydroxycholesterol; D008148:Lovastatin; D000069059:Atorvastatin; D019821:Simvastatin; D006903:Hydroxymethylglutaryl CoA Reductases; D008798:Mevalonic Acid