Y. Yoon

Inhibitory effects of fruit juices on CYP3A activity

There have been very limited reports on the effects of commercial fruit juices on human CYP3A activity. Therefore, the inhibitory effects of readily available commercial fruit juices on midazolam 1′-hydroxylase activity, a marker of CYP3A, were evaluated in pooled human liver microsomes. The fruit juices investigated were black raspberry, black mulberry, plum, and wild grape. White grapefruit, pomegranate, and orange juice were used as positive and negative controls. The black mulberry juice showed the most potent inhibition of CYP3A except for grapefruit juice. The inhibition depended on the amount of a fruit juice added to the incubation mixture. The inhibitory potential of human CYP3A was in the order: grapefruit > black mulberry > wild grape > pomegranate > black raspberry. The IC50 values of all fruit juices tested were reduced after preincubation with microsomes in the presence of the NADPH-generating system, suggesting that a mechanism-based inhibitory component was present in these fruit juices, as in the case of grapefruit. The results suggest that, like grapefruit juice, commercial fruit juices also have the potential to inhibit CYP3A-catalzyed midazolam 1′-hydroxylation. Therefore, in vivo studies investigating the interactions between fruit juices such as black mulberry and wild grape and CYP3A substrates are necessary to determine whether inhibition of CYP3A activity by fruit juices is clinically relevant.

Characterization of Ebastine, Hydroxyebastine, and Carebastine Metabolism by Human Liver Microsomes and Expressed Cytochrome P450 Enzymes: Major Roles for CYP2J2 and CYP3A

Ebastine undergoes extensive metabolism to form desalkylebastine and hydroxyebastine. Hydroxyebastine is subsequently metabolized to carebastine. Although CYP3A4 and CYP2J2 have been implicated in ebastine N-dealkylation and hydroxylation, the enzyme catalyzing the subsequent metabolic steps (conversion of hydroxyebastine to desalkylebastine and carebastine) have not been identified. Therefore, we used human liver microsomes (HLMs) and expressed cytochromes P450 (P450s) to characterize the metabolism of ebastine and that of its metabolites, hydroxyebastine and carebastine. In HLMs, ebastine was metabolized to desalkyl-, hydroxy-, and carebastine; hydroxyebastine to desalkyl- and carebastine; and carebastine to desalkylebastine. Of the 11 cDNA-expressed P450s, CYP3A4 was the main enzyme catalyzing the N-dealkylation of ebastine, hydroxyebastine, and carebastine to desalkylebastine [intrinsic clearance (CLint) = 0.44, 1.05, and 0.16 μl/min/pmol P450, respectively]. Ebastine and hydroxyebastine were also dealkylated to desalkylebastine to some extent by CYP3A5. Ebastine hydroxylation to hydroxyebastine is mainly mediated by CYP2J2 (0.45 μl/min/pmol P450; 22.5- and 7.5-fold higher than that for CYP3A4 and CYP3A5, respectively), whereas CYP2J2 and CYP3A4 contributed to the formation of carebastine from hydroxyebastine. These findings were supported by chemical inhibition and kinetic analysis studies in human liver microsomes. The CLint of hydroxyebastine was much higher than that of ebastine and carebastine, and carebastine was metabolically more stable than ebastine and hydroxyebastine. In conclusion, our data for the first time, to our knowledge, suggest that both CYP2J2 and CYP3A play important roles in ebastine sequential metabolism: dealkylation of ebastine and its metabolites is mainly catalyzed by CYP3A4, whereas the hydroxylation reactions are preferentially catalyzed by CYP2J2. The present data will be very useful to understand the pharmacokinetics and drug interaction of ebastine in vivo.

Characterization of Benidipine and Its Enantiomers' Metabolism by Human Liver Cytochrome P450 Enzymes

Benidipine is a dihydropyridine calcium antagonist that has been used clinically as an antihypertensive and antianginal agent. It is used clinically as a racemate, containing the (-)-α and (+)-α isomers of benidipine. This study was performed to elucidate the metabolism of benidipine and its enantiomers in human liver microsomes (HLMs) and to characterize the cytochrome P450 (P450) enzymes that are involved in the metabolism of benidipine. Human liver microsomal incubation of benidipine in the presence of NADPH resulted in the formation of two metabolites, N-desbenzylbenidipine and dehydrobenidipine. The intrinsic clearance (CLint) of the formation of N-desbenzylbenidipine and dehydrobenidipine metabolites from (-)-α isomer was similar to those from the (+)-α isomer (1.9 ± 0.1 versus 2.3 ± 2.3 μl/min/pmol P450 and 0.5 ± 0.2 versus 0.6 ± 0.6 μl/min/pmol P450, respectively). Correlation analysis between the known P450 enzyme activities and the rate of the formation of benidipine metabolites in the 15 HLMs showed that benidipine metabolism is correlated with CYP3A activity. The P450 isoform-selective inhibition study in liver microsomes and the incubation study of cDNA-expressed enzymes also showed that theN-debenzylation and dehydrogenation of benidipine are mainly mediated by CYP3A4 and CYP3A5. The total CLint values of CYP3A4-mediated metabolite formation from (-)-α isomer were similar to those from (+)-α isomer (17.7 versus 14.4 μl/min/pmol P450, respectively). The total CLint values of CYP3A5-mediated metabolite formation from (-)-α isomer were also similar to those from (+)-α isomer (8.3 versus 11.0 μl/min/pmol P450, respectively). These findings suggest that CYP3A4 and CYP3A5 isoforms are major enzymes contributing to the disposition of benidipine, but stereoselective disposition of benidipine in vivo may be influenced not by stereoselective metabolism but by other factors.

Potential of pranlukast and zafirlukast in the inhibition of human liver cytochrome P450 enzymes.

1. The potential of zafirlukast to inhibit several human cytochrome P450 enzymes is well known. However, pranlukast, a structural analogue of zafirlukast, has not been studied. Accordingly, the inhibitory potential of pranlukast was evaluated and compared with that of zafirlukast, a known CYP2C9 inhibitor, in in vitro microsomal incubation studies. 2. Both pranlukast and zafirlukast showed moderate inhibition of CYP2C9-catalysed tolbutamide 4-methylhydroxylation, competitively inhibiting tolbutamide 4-methylhydroxylation with estimated mean Ki values of 3.82 ± 0.50 and 5.86 ± 0.08 μM, respectively. 3. Pranlukast had no effect on CYP2C19-catalysed S-mephenytoin 4′-hydroxylation or CYP3A4-catalysed midazolam 1-hydroxylation. However, zafirlukast showed minor inhibition of these reactions. Neither pranlukast nor zafirlukast inhibited CYP1A2-catalysed phenacetin O-deethylation, CYP2D6-catalysed dextromethorphan O-demethylation or CYP2E1-catalysed chlorzoxazone 6-hydroxylation. 4. The results suggest that like zafirlukast, pranlukast also has the potential moderately to inhibit CYP2C9-catalysed tolbutamide 4-methylhydroxylation. Therefore, the inhibitory potential of pranlukast should be considered when it is co-administered with CYP2C9 substrates with narrow therapeutic ranges (e.g. S-warfarin, phenytoin).

Potential of hydroxyebastine and terfenadine alcohol to inhibit the human cytochrome P450 2J2 isoform

Although selective inhibitors of cytochrome P450 enzymes can be used to determine relative contributions of the enzymes to xenobiotic metabolism, characterization of CYP2J2 in drug metabolism is more challenging due to lack of selective, well-characterized inhibitors. Thus, selectivity of hydroxyebastine, which has high affinity for recombinant CYP2J2, was studied. The IC50 value of hydroxyebastine in CYP2J2-mediated astemizole O-demethylation activity was lower than that of its structural analog, terfenadine alcohol. Terfenadine alcohol inhibited several other P450 activities, such as CYP2D6, more potently than CYP2J2, and is thus not suitable as a CYP2J2-selective inhibitor. Inhibitory potential values of hydroxyebastine in CYP2J2-catalyzed astemizole O-demethylation, tolbutamide hydroxylation (CYP2C9), S-mephenytoin hydroxylation (CYP2C19), and dextromethorphan O-demethylation (CYP2D6) were 0.45, 2.74, 10.22, and 3.83 μM, respectively. The inhibitory potential of other P450 enzymes, such as CYP1A2, CYP2B6, CYP2E1, and CYP3A, was negligible. Although hydroxyebastine was a relatively potent inhibitor of CYP2J2, it provided a selectivity of only > 6-fold (CYP2J2 vs. other P450s). However, hydroxyebastine can serve as a relatively selective inhibitor of CYP2J2 and can be used to characterize the contribution of CYP2J2 to xenobiotic metabolism due to the lack of a more specific inhibitor.

The effect of CYP2C19 genotype on the time course of platelet aggregation inhibition after clopidogrel administration

We evaluated the effect of CYP2C19 genotype over time on the antiplatelet response of clopidogrel in healthy subjects. Seventy subjects enrolled for a pharmacodynamic study and 22 subjects for a pharmacokinetic and pharmacodynamic study took 300 mg clopidogrel on the first day and 75 mg once daily for six consecutive days. The subjects with CYP2C19 poor metabolizers (PM, N = 22) and intermediate metabolizers (IM, N = 37) had significantly delayed time to inhibition of platelet aggregation (IPA) compared with CYP2C19 extensive metabolizers (EM, N = 33) (12 vs. 9 vs. 2 hours as median Tmax, P < .05) after a 300 mg of clopidogrel. During maintenance doses of clopidogrel, IPA values of only CYP2C19 PM subjects were gradually decreased from 30.0 ± 21.9% on day 2 to 23.7 ± 16.6% on day 8 (P > .05 for time effect; P < .05 for time and genotype interaction effect). CYP2C19 PM had decreased Cmax and AUC of thiol metabolite compared with CYP2C19 EM (0.42‐ and 0.37‐fold on day 1, P < .01; 0.39‐ and 0.34‐fold on day 7, P < .01, respectively). Delayed time to reach maximal IPA as well as decreased IPA may influence the increased risk of the acute cardiac events in CYP2C19 PM and IM.

PI-63The effects of pomegranate juice on pharmacokinetics of simvastatin in healthy Korean subjects

Previous in vitro and animal experiments reported that pomegranate juice increased markedly the bioavailability of carbamazepine due to a potent inhibition of CYP3A4. This study was aimed to evaluate the effect of pomegranate on the disposition of simvastatin and its active metabolite, simvastatin acid, compared with grapefruit juice in 12 healthy volunteers.

Effect of itraconazole on the pharmacokinetics and pharmacodynamics of fexofenadine in subjects with known genotype of MDR1 3435C>T allele

Clinical Pharmacology & Therapeutics (2003) 73, P58–P58; doi:

Herba Epimedii Extract Increases The Digoxin Transport in Transwell Culture of CaCO‐2 Cells in Vitro

Clinical Pharmacology & Therapeutics (2003) 73, P41–P41; doi:

Extensive evaluation of genetic polymorphism of CYP2C19 in a Korean population: interethnic comparison among Asian populations

We evaluated the frequency distribution of the cytochrome P450 2C19 (CYP2C19) allelic variants in a Korean population and also measured the metabolic ratio of omeprazole, a probe drug for CYP2C19 according to the genotypes. The results were compared with those of other ethnics including Vietnamese data which we had studied. Genotyping of CYP2C19 (*2 and *3) was carried out in 282 Korean subjects using PCR‐RFLP and ninety‐four subjects were participated in phenotyping study using omeprazole. Plasma concentrations of omeprazole and its metabolite, hydroxy‐omeprazole were measured at 2hr after 20mg omeprazole intake. Forty subjects were identified to have CYP2C19 genotype of PM with the frequency of 8.9% CYP2C19*2/*2, 3.9% CYP2C192/*3, and 1.4% CYP2C19*3/*3, while 242 Korean subjects carried extensive metabolizer (EM) genotypes which consists of CYP2C19*1/*1 (41.8%), CYP2C19*1/*2 (35.5%), and CYP2C19*1/*3 (8.5%). The frequency distribution of metabolic ratio was distinctly bimodal. Ten Korean subjects were identified to be PMs with an antimode of 1.2. The frequency (12.6 %) of PM in Korean population is significantly higher than those in Vietnamese (7.1 %) and Thai (6.5), but lower than those in Chinese (19.8 %), Filipinos (22.6 %) and Japanese (18.8%).