(-)- N -3-Benzylphenobarbital Is Superior to Omeprazole and (+)- N -3-Benzylnirvanol as a CYP2C19 Inhibitor in Suspended Human Hepatocytes.

Early assessment of metabolism pathways of new chemical entities guides the understanding of drug-drug interactions. Selective enzyme inhibitors are indispensable in CYP reaction phenotyping. The most commonly applied CYP2C19 inhibitor, omeprazole, lacks selectivity. Two promising alternatives, (+)- N -3-benzylnirvanol and (-)- N -3-benzylphenobarbital, are already used as CYP2C19 inhibitors in some in vitro studies with suspended human hepatocytes. However, a full validation proving their suitability in terms of CYP and non-CYP selectivity has not been presented in literature. The present study provides a thorough comparison between omeprazole, (+)- N -3-benzylnirvanol, and (-)- N -3-benzylphenobarbital in terms of potency and selectivity and shows the superiority of (-)- N -3-benzylphenobarbital as a CYP2C19 inhibitor in suspended human hepatocytes. Furthermore, we evaluated the application of (-)- N -3-benzylphenobarbital to predict the in vivo contribution of CYP2C19 to drug metabolism [fraction metabolized (fm) of CYP2C19, fm _CYP2C19 ]. A set of 10 clinically used CYP2C19 substrates with reported in vivo fm _CYP2C19 data was evaluated. fm _CYP2C19 , which was predicted using data from suspended human hepatocyte incubations, underestimated the in vivo fm _CYP2C19 The use of a different hepatocyte batch with a different CYP3A4/CYP2C19 activity ratio showed the impact of intrinsic CYP activities on the determination of fm _CYP2C19 Overall, this study confirms the selective CYP2C19 inhibition by (-)- N -3-benzylphenobarbital over other CYP isoforms (CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2D6, and CYP3A4) and clinically relevant non-CYP enzymes [aldehyde oxidase, flavin-containing monooxygenase 3, N-acetyltransferase 2, uridine diphosphate glucuronosyltransferase (UGT) 1A1, UGT1A4, UGT2B7, UGT2B15] in suspended human hepatocytes. (-)- N -3-benzylphenobarbital is therefore the preferred CYP2C19 inhibitor to assess fm _CYP2C19 in suspended human hepatocytes in comparison with omeprazole and (+)- N -3-benzylnirvanol. SIGNIFICANCE STATEMENT: (-)- N -3-Benzylphenobarbital is a more potent and selective inhibitor of CYP2C19 in suspended human hepatocytes than omeprazole and (+)- N -3-benzylnirvanol. (-)- N -3-Benzylphenobarbital can be used to predict the fraction metabolized by CYP2C19 in suspended human hepatocytes.
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