Identification of human cytochrome P450 isoforms that contribute to all-trans-Retinoic Acid 4-hydroxylation

Thr role of specific human cytochrome P450 (CYP) isoforms in the oxidative metabolism of all-trans-retinoic acid was investigated by studies in human liver microsomes using isoform-specific chemical inhibitors and inhibitory antibodies. Studies using individual isoforms expressed in lymphoblastoid c ells and correlation analysis using different microsome preparations were a lso performed. With expressed isoforms, evidence for a role for CYP2C8, CYP 3A4, CYP2C9, and CYP1A1 in 4-hydroxylation was obtained, with the highest c atalytic efficiency being observed for CYP2C8. Using inhibition studies and correlation analysis, we also concluded that CYP2C8 was the major all-tran s-retinoic acid 4-hydroxylating cytochrome P450 in human liver microsomes, though CYP3A4 and, to a lesser extent CYP2C9, also made a contribution. In addition, we compared the rate of retinoic acid degredation in HepG2 cells when cultured in the absence and presence of 3-methylcholanthrene or all-tr ans-retinoic acid. Culture in the presence of all-trans-retinoic acid decre ased the half-life twofold and resulted in an increased sensitivity of reti noic acid degredation to ketoconazole. Since no induction of either CYP1A1, CYP2C8, CYP2C9, or CYP3A4 was detected using immunoblotting and as mRNA en coding another cytochrome P450 enzyme, CYP26, has been previously demonstra ted to be induced by retinoic acid treatment of HepG2 cells and to be highl y sensitive to ketoconazole, this enzyme in addition to CYP2C8, CYP2C9 and CYP3A4 likely plays a role in all-trans-retinoic acid oxidation in the live r at high retinoic acid levels. BIOCHEM PHARMACOL 60;4:517-526, 2000. (C) 2 000 Elsevier Science Inc.