title: Monitoring compound integrity with cytochrome P450 assays and qHTS.
abstract: The authors describe how room temperature storage of a 1120-member compound library prepared in either DMSO or in a hydrated-DMSO/water (67/33) mixture affects the reproducibility of potency values as monitored using cytochrome P450 1A2 and 2D6 isozyme assays. The bioluminescent assays showed Z' factors of 0.71 and 0.62, with 17% and 32% of the library found as active against the CYP 1A2 and 2D6 isozymes, respectively. The authors tested the library using quantitative high-throughput screening to generate potency values for every library member, which was measured at 7 time intervals spanning 37 weeks. They calculated the minimum significant ratio (MSR) from these potency values at each time interval and found that for the library stored in DMSO, the CYP 1A2 and 2D6 assay MSRs progressed from approximately 2.0 to 5.0. The hydrated conditions showed similar performance in both MSR progression and analytical quality control results. Based on this study, the authors recommend that DMSO samples be stored in 1536-well plates for <4 months at room temperature. Furthermore, the study illustrates the degree and time scale of apparent compound potency changes due to sample storage.
authors.affiliation: NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA.
authors.forename: Ryan
authors.initials: R
authors.lastname: MacArthur
authors.forename: William
authors.initials: W
authors.lastname: Leister
authors.forename: Henrike
authors.initials: H
authors.lastname: Veith
authors.forename: Paul
authors.initials: P
authors.lastname: Shinn
authors.forename: Noel
authors.initials: N
authors.lastname: Southall
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Austin
authors.forename: James
authors.initials: J
authors.lastname: Inglese
authors.forename: Douglas S
authors.initials: DS
authors.lastname: Auld
mesh_terms: D001681:Biological Assay / Q000295:instrumentation / Q000379:methods*; D002853:Chromatography, Liquid / Q000379:methods; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D055808:Drug Discovery / Q000295:instrumentation / Q000379:methods*; D004356:Drug Storage; D007527:Isoenzymes / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D004364:Pharmaceutical Preparations / Q000737:chemistry / Q000378:metabolism
chemical_list: D007527:Isoenzymes; D004364:Pharmaceutical Preparations; D003577:Cytochrome P-450 Enzyme System