High-Throughput Cytochrome P450 Inhibition Assays Using Laser Diode Thermal Desorption-Atmospheric Pressure Chemical Ionization-Tandem Mass Spectrometry

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• 作者：Jin Wu ; Christopher S. Hughes ; Pierre Picard ; Sylvain Letarte ; Mireille Gaudreault ; Jean-Franç ; ois Lé ; vesque ; Deborah A. Nicoll-Griffith ; Kevin P. Bateman
• 刊名：Analytical Chemistry
• 出版年：2007
• 出版时间：June 15, 2007
• 年：2007
• 卷：79
• 期：12
• 页码：4657 - 4665
• 全文大小：162K
• 年卷期：v.79,no.12(June 15, 2007)
• ISSN：1520-6882

文摘

This paper describes the development of a high-throughput method for the analysis of cytochrome P450 (CYP)inhibition assay incubation samples using laser diodethermal desorption interfaced with atmospheric pressurechemical ionization mass spectrometry (LDTD-APCI-MS).Data for the CYP isoforms 3A4, 2D6, 2C9, and 1A2 fromcompetitive inhibition assays are shown. The potential forinhibition of the CYP isoforms was measured by monitoring the level of the metabolites 6-hydroxytestosterone(3A4), dextrorphan (2D6), 4'-hydroxydiclofenac (2C9),and acetaminophen (1A2) formed in the presence of drugcandidates using an eight-point titration. The analyticalmethod involves plating of the inhibition samples onspecially designed 96-well plates with stainless steelbottoms, followed by direct analysis using the LDTDsource. Validation of the LDTD-MS method was performed by testing for interferences, reproducibility, dynamic range, ion suppression, and the ability of the sourceto produce comparable results to previously validatedLC-MS methods. IC₅₀ values for each CYP isoform using33 different test compounds showed excellent agreementbetween LDTD-APCI-MS and LC-MS methods and literature values where available. Assay analysis time usingthe LDTD-APCI source is reduced to less than 30 minfor a single 96-well plate compared to greater than 10 husing the LC-MS method. The LDTD-APCI-MS and LC-MS methods and results are compared and limitationsand future potential for LDTD-APCI-MS are discussed.