Identification of the Human Liver Cytochrome P450 Isoenzyme Responsible for the 6-Methylhydroxylation of the Novel Anticancer Drug 5,6-Dimethylxanthenone-4-Acetic Acid
In vitro studies were conducted to identify the hepatic cytochrome P450 (CYP) isoenzyme involved in the 6-methylhydroxylation of 5,6-dimethylxanthenone-4-acetic acid (DMXAA) by using a human liver library (n = 14). The metabolite 6-hydroxymethyl-5-methylxanthenone-4-acetic acid (6-OH-MXAA) was determined by HPLC with fluorescence detection. The metabolite formed in human liver microsomes and by cDNA-expressed CYP isoform was identified by liquid chromatography mass spectrometry as 6-OH-MXAA. In human liver microsomes (n = 14), 6-methylhydroxylation of DMXAA followed monophasic Michaelis-Menten kinetics, with a mean apparent Km of 21 ± 5 μM and Vmax of 0.043 ± 0.019 nmol/min/mg. An approximate 10-fold interindividual variation in the intrinsic clearance (Vmax/Km) of DMXAA 6-methylhydroxylation in human liver microsomes was observed. The involvement of CYP1A2 in DMXAA metabolism by human livers was demonstrated by the following: 1) the potent inhibition of DMXAA metabolism by furafylline (kinact = 0.23 ± 0.04 min−1, K′app= 15.6 ± 6.7 μM) and α-naphthoflavone (Ki = 0.036 μM), but not by cimetidine, ketoconazole, tolbutamide, quinidine, chlorzoxazone, diethyldithiocarbamate, troleandomycin, and sulfaphenazole; 2) when incubated with human lymphoblastoid cell microsomes containing cDNA-expressed CYP isoenzymes, DMXAA was metabolized only by CYP1A2, with an apparent Km of 6.2 ± 1.5 μM and Vmax of 0.014 ± 0.001 nmol/min/mg, but not by CYP2A6, CYP2B6, CYP2C9 (Arg144), CYP2C19, CYP2D6 (Val374), CYP2E1, and CYP3A4; 3) a significant correlation (r = 0.90; P < .001) between 6-methylhydroxylation of DMXAA and 7-ethoxyresorufinO-deethylation; and 4) a significant correlation (r = 0.75; P < .01) between the CYP1A protein level determined by Western blots and DMXAA 6-methylhydroxylation.
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Citation / Publisher Attribution
Drug Metabolism and Disposition, v. 28, issue 12, p. 1449-1456
Scholar Commons Citation
Zhou, Shufeng; Paxton, James W.; Tingle, Malcolm D.; and Kestell, Philip, "Identification of the Human Liver Cytochrome P450 Isoenzyme Responsible for the 6-Methylhydroxylation of the Novel Anticancer Drug 5,6-Dimethylxanthenone-4-Acetic Acid" (2000). Pharmacy Faculty Publications. 51.