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Monitoring Compound Integrity With Cytochrome P450 Assays and qHTS

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA

William Leister

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA

Henrike Veith

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA

Paul Shinn

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA

Noel Southall

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA

Christopher P. Austin

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA

James Inglese

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA

Douglas S. Auld

NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892-3370, USA, dauld{at}mail.nih.gov

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. (Journal of Biomolecular Screening 2009:538-546)

Key Words: HTS • compound storage • DMSO • quantitative HTS

This version was published on June 1, 2009

Journal of Biomolecular Screening, Vol. 14, No. 5, 538-546 (2009)
DOI: 10.1177/1087057109336954


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