Received for publication April 10, 2006.
Revised July 31, 2006.
Accepted for publication August 1, 2006.

Characterization of ebastine, hydroxyebastine, and carebastine metabolism by human liver microsomes and expressed cytochrome P450 enzymes: major roles for CYP2J2 and CYP3A

Abstract

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 (HLM) and expressed P450s to characterize the metabolism of ebastine and those of its metabolites, hydroxyebastine and carebastine. In HLM, ebastine was metabolized to desalkyl-, hydroxy-, and car-ebastine; hydroxyebastine to desalkyl- and car-ebastine; 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 (Cl_int) = 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), while 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 Cl_int 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 suggest that both CYP2J2 and CYP3A play important roles in ebastine sequential metabolism: dealkylation of ebastine and its metabolites is mainly catalyzed by CYP3A4 while 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.

Key words: cytochrome P450, drug disposition, enzyme kinetics, liver microsomes, mass spectrometry