Identification of human cytochrome P450 isoforms that contribute to all-trans-retinoic acid 4-hydroxylation.
The 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 cells and correlation analysis using different microsome preparations were also performed. With expressed isoforms, evidence for a role for CYP2C8, CYP3A4, CYP2C9, and CYP1A1 in 4-hydroxylation was obtained, with the highest catalytic efficiency being observed for CYP2C8. Using inhibition studies and correlation analysis, we also concluded that CYP2C8 was the major all-trans-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-trans-retinoic acid. Culture in the presence of all-trans-retinoic acid decreased the half-life twofold and resulted in an increased sensitivity of retinoic acid degredation to ketoconazole. Since no induction of either CYP1A1, CYP2C8, CYP2C9, or CYP3A4 was detected using immunoblotting and as mRNA encoding another cytochrome P450 enzyme, CYP26, has been previously demonstrated to be induced by retinoic acid treatment of HepG2 cells and to be highly sensitive to ketoconazole, this enzyme in addition to CYP2C8, CYP2C9 and CYP3A4 likely plays a role in all-trans-retinoic acid oxidation in the liver at high retinoic acid levels.
McSorley
L C
LC
Department of Pharmacological Sciences, University of Newcastle upon Tyne, Medical School, Newcastle upon Tyne, UK.
Daly
A K
AK
D000906:Antibodies / Q000494:pharmacology; D001189:Aryl Hydrocarbon Hydroxylases; D002478:Cells, Cultured; D065727:Cytochrome P-450 CYP2C8; D065729:Cytochrome P-450 CYP2C9; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006900:Hydroxylation; D007527:Isoenzymes / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000032:analysis; D039181:Steroid 16-alpha-Hydroxylase; D013250:Steroid Hydroxylases / Q000032:analysis; D014212:Tretinoin / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D000906:Antibodies; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D014212:Tretinoin; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D013250:Steroid Hydroxylases; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C450259:CYP2C8 protein, human; C104464:CYP3A protein, human; D065727:Cytochrome P-450 CYP2C8; D051544:Cytochrome P-450 CYP3A; D039181:Steroid 16-alpha-Hydroxylase; C510163:CYP3A4 protein, human