Drug Discov Ther. 2020;14(4):204-208. (DOI: 10.5582/ddt.2020.03060)

Phenotypic analysis of human CYP2C9 polymorphisms using fluorine-substituted tolbutamide

Kitamura Y, Saeki K


To investigate the effect of fluorine substitution on tolbutamide (TB) hydroxylation catalyzed by CYP2C9, the hydroxylation of TB and its fluorinated derivative 3'-fluoro-tolbutamide (3'-FTB) by recombinant human CYP2C9*1, CYP2C9*2, and CYP2C9*3 was analyzed. In general, fluorine substitution near the metabolic site may decrease enzymatic oxidation owing to its electron-withdrawing nature. Fluorine substitution reduced the Michaelis–Menten-derived Km of 4'-hydroxylation of TB catalyzed by CYP2C9*1 from 115 (TB) to 77 (3'-F-TB) μM. In the case of TB hydroxylation catalyzed by CYP2C9*2, the Km value of TB was also reduced by fluorine substitution from 129 to 88 μM. The greatest effect of fluorine substitution on the Km in TB hydroxylation was observed in the catalysis by CYP2C9*3, in which the Km value decreased from 287 to 117 μM. When a mixture containing TB and 3′-F-TB was hydroxylated by CYP2C9, the hydroxylated metabolite ratio in CYP2C9*3 was significantly increased compared with that in CYP2C9*1 and CYP2C9*2 (p < 0.01, Tukey–Kramer test). These results suggest that obtaining the metabolite profiles of fluorine-substituted analogs of the key substrate molecule may be useful as a new tool for phenotyping polymorphic CYP isoforms.

KEYWORDS: fluorine substitution, tolbutamide, CYP2C9 polymorphism, phenotyping

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