The development of a cocktail CYP2B6, CYP2C8, and CYP3A5 inhibition assay and a preliminary assessment of utility in a drug discovery setting.
Tools for studying the roles of CYP2B6, CYP2C8, and CYP3A5 in drug metabolism have recently become available. The level of interest in these enzymes has been elevated because investigations have revealed substrate promiscuity and/or polymorphic expression. In this study, we aimed to develop a single cocktail inhibition assay for the three enzymes and assess its utility in drug discovery. Bupropion hydroxylation, amodiaquine N-deethylation, and midazolam 1'-hydroxylation were chosen as probe reactions for CYP2B6, CYP2C8, and CYP3A5 and were analyzed using liquid chromatography-tandem mass spectrometry. Kinetic analyses were performed to establish suitable conditions for inhibition assays, which were subsequently automated. CYP2B6, CYP2C8, and CYP3A5 IC(50) values were determined for marketed drugs and almost 200 AstraZeneca discovery compounds from 16 separate discovery projects. For the marketed drugs, results obtained were comparable with literature values. Data were also compared with IC(50) values determined for CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4. In this dataset, the majority of compounds were more potent inhibitors of CYP2C9, CYP2C19, CYP2D6, and CYP3A4 than of CYP2B6, CYP2C8, or CYP3A5. The potential impact of these findings on a cytochrome P450 inhibition strategy is discussed.
O'Donnell
Charles J
CJ
Department of Physical and Metabolic Science, AstraZeneca R&D Charnwood, Bakewell Road, Loughborough, UK.
Grime
Ken
K
Courtney
Paul
P
Slee
Dean
D
Riley
Robert J
RJ
D001189:Aryl Hydrocarbon Hydroxylases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D001681:Biological Assay; D065702:Cytochrome P-450 CYP2B6; D065727:Cytochrome P-450 CYP2C8; D051544:Cytochrome P-450 CYP3A; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D015195:Drug Design; D004347:Drug Interactions; D004926:Escherichia coli / Q000235:genetics; D007654:Ketoconazole / Q000494:pharmacology; D007700:Kinetics; D010089:Oxidoreductases, N-Demethylating / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D004364:Pharmaceutical Preparations / Q000378:metabolism; D011794:Quercetin / Q000494:pharmacology; D013988:Ticlopidine / Q000494:pharmacology
D065607:Cytochrome P-450 Enzyme Inhibitors; D004364:Pharmaceutical Preparations; D003577:Cytochrome P-450 Enzyme System; D011794:Quercetin; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; C450259:CYP2C8 protein, human; C510164:CYP3A5 protein, human; D065702:Cytochrome P-450 CYP2B6; D065727:Cytochrome P-450 CYP2C8; D051544:Cytochrome P-450 CYP3A; D010089:Oxidoreductases, N-Demethylating; D013988:Ticlopidine; D007654:Ketoconazole