Substrate specificity for rat cytochrome P450 (CYP) isoforms: screening with cDNA-expressed systems of the rat.
In this study, we performed a screening of the specificities of rat cytochrome P450 (CYP) isoforms for metabolic reactions known as the specific probes of human CYP isoforms, using 13 rat CYP isoforms expressed in baculovirus-infected insect cells or B-lymphoblastoid cells. Among the metabolic reactions studied, diclofenac 4-hydroxylation (DFH), dextromethorphan O-demethylation (DMOD) and midazolam 4-hydroxylation were specifically catalyzed by CYP2C6, CYP2D2 and CYP3A1/3A2, respectively. These results suggest that diclofenac 4-hydroxylation, dextromethorphan O-demethylation and midazolam 4-hydroxylation are useful as catalytic markers of CYP2C6, CYP2D2 and CYP3A1/3A2, respectively. On the other hand, phenacetin O-deethylation and 7-ethoxyresorufin O-deethylation were catalyzed both by CYP1A2 and by CYP2C6. Benzyloxyresorufin O-dealkylation and pentoxyresorufin O-dealkylation were also catalyzed by CYP1A2 in addition to CYP2B1. Bufuralol 1'-hydroxylation was extensively catalyzed by CYP2D2 but also by CYP2C6 and CYP2C11. p-Nitrophenol 2-hydroxylation and chlorzoxazone 6-hydroxylation were extensively catalyzed by CYP2E1 but also by CYP1A2 and CYP3A1. Therefore, it is necessary to conduct further study to clarify whether these activities in rat liver microsomes are useful as probes of rat CYP isoforms. In contrast, coumarin 7-hydroxylation and S- and R-mephenytoin 4'-hydroxylation did not show selectivity toward any isoforms of rat CYP studied. Therefore, activities of coumarin 7-hydroxylation and S- and R-mephenytoin 4'-hydroxylation are not able to be used as catalytic probes of CYP isoforms in rat liver microsomes. These results may provide useful information regarding catalytic probes of rat CYPs for studies using rat liver microsomal samples.
Kobayashi
Kaoru
K
Laboratory of Biochemical Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, Chiba University, Chiba, Japan. kaoruk@p.chiba-u.ac.jp
Urashima
Kikuko
K
Shimada
Noriaki
N
Chiba
Kan
K
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D016367:Baculoviridae / Q000235:genetics; D002478:Cells, Cultured; D002753:Chlorzoxazone / Q000378:metabolism; D003374:Coumarins / Q000378:metabolism; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D003915:Dextromethorphan / Q000378:metabolism; D004008:Diclofenac / Q000378:metabolism; D004983:Ethanolamines / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D007313:Insecta / Q000166:cytology; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008617:Mephenytoin / Q000378:metabolism; D008874:Midazolam / Q000378:metabolism; D009596:Nitrophenols / Q000378:metabolism; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D010615:Phenacetin / Q000378:metabolism; D051381:Rats; D013379:Substrate Specificity; D013739:Testosterone / Q000378:metabolism; D014407:Tumor Cells, Cultured
D003374:Coumarins; D018076:DNA, Complementary; D004983:Ethanolamines; D007527:Isoenzymes; D009596:Nitrophenols; D004364:Pharmaceutical Preparations; D004008:Diclofenac; D013739:Testosterone; D003915:Dextromethorphan; C010831:bufuralol; D003577:Cytochrome P-450 Enzyme System; C030123:coumarin; D001189:Aryl Hydrocarbon Hydroxylases; C465138:Cyp3a23-3a1 protein, rat; D051544:Cytochrome P-450 CYP3A; D010615:Phenacetin; D002753:Chlorzoxazone; D008617:Mephenytoin; D008874:Midazolam; C024836:4-nitrophenol