Identification of CYP1A2 as the main isoform for the phase I hydroxylated metabolism of genistein and a prodrug converting enzyme of methylated isoflavones.
This study determined the cytochrome P450 (P450) isoforms responsible for metabolism of isoflavones using human liver microsomes (HLM) and expressed P450s. The primary metabolite of genistein is 3'-OH-genistein, as identified with an authentic chemically synthesized standard. CYP1A2 was predominantly responsible for 3'-OH-genistein formation since its formation was inhibited (>50%, p < 0.05) by a monoclonal antibody specific for CYP1A2, was correlated with CYP1A2 activities of HLM, and was catalyzed by expressed CYP1A2. In addition to CYP1A2, CYP2E1 also catalyzed, although to a lesser extent, its formation. The contribution of these P450s to the formation of 3'-OH-genistein was also confirmed with a panel of expressed enzymes. Methylated isoflavones biochanin A, prunetin, and formononetin (10-100 microM) were rapidly converted by HLM and expressed CYP1A2 to more active genistein and daidzein. The conversion of biochanin A to genistein appears to be mainly mediated by CYP1A2 because of the strong correlation between the conversion rates and CYP1A2 activities in HLM. Thus, CYP1A2 is an effective prodrug-converting enzyme for less active methylated isoflavones. CYP1A2-catalyzed conversion of biochanin A to genistein (Km, 7.80 microM; Vmax, 903 pmol/min/mg of protein; Vmax/Km, 116 microl/min/mg of protein) was much faster than 3'-hydroxylation of genistein (Km, 12.7 microM and Vmax, 109 pmol/min/mg of protein; Vmax/Km, 8.6 microl/min/mg of protein). The interaction studies showed that genistein inhibited formation of acetaminophen from phenacetin with an IC50 value of 16 microM. Additional studies showed that phenacetin and genistein were mutually inhibitory. In conclusion, CYP1A2 and CYP2E1 metabolized genistein and CYP1A2 acted as prodrug-converting enzymes for other less active methylated isoflavones.
Hu
Ming
M
LAboratory of Molecular Carcinogenesis, National Cancer Institute, National Institutes of Healh, Bethesda, Maryland, USA. minghu@wsu.edu
Krausz
Kristopher
K
Chen
Jun
J
Ge
Xia
X
Li
Jianqi
J
Gelboin
Harry L
HL
Gonzalez
Frank J
FJ
D000098:Acetophenones / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000378:metabolism / Q000494:pharmacology; D001711:Biotransformation; D019388:Cytochrome P-450 CYP1A2 / Q000096:biosynthesis / Q000737:chemistry* / Q000493:pharmacokinetics; D004347:Drug Interactions; D004351:Drug Resistance / Q000502:physiology; D015870:Gene Expression; D019833:Genistein / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation; D007529:Isoflavones / Q000008:administration & dosage / Q000378:metabolism* / Q000493:pharmacokinetics; D008745:Methylation; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D015394:Molecular Structure; D010090:Oxidoreductases, O-Demethylating / Q000378:metabolism; D010615:Phenacetin / Q000378:metabolism; D011355:Prodrugs / Q000378:metabolism; D020033:Protein Isoforms
C482259:2'-hydroxy-genistein; C482258:3'-hydroxy-genistein; C482260:4,6-di(methoxymethoxy)-2-hydroxyacetophenone; D000098:Acetophenones; D000911:Antibodies, Monoclonal; D007529:Isoflavones; D011355:Prodrugs; D020033:Protein Isoforms; D019833:Genistein; D006899:Mixed Function Oxygenases; D010090:Oxidoreductases, O-Demethylating; D019388:Cytochrome P-450 CYP1A2; D010615:Phenacetin; C004541:biochanin A