The decreased in vivo clearance of CYP2D6 substrates by CYP2D6*10 might be caused not only by the low-expression but also by low affinity of CYP2D6

CYP2D6 exhibits genetic polymorphism with interindividual differences in metabolic activity, We have found a significant influence on the pharmacokinetics of venlafaxine by the CYP2D6*10 allele in a Japanese population. CYP2D6.10, which is translated from CYP2D6*10, has two amino acid substitutions: Pro34 --> Ser and Ser486 --> Thr, In this study, CYP2D6.10 was expressed in Saccharomyces cerevisiae and its catalytic activity for CYP2D6 substrates was investigated. The CYP2D6*10B- and *10C-associated cDNA were isolated from human lymphocyte genotyped as CYP2D6*10, In addition, three forms of CYP2D6, Pro34/Thr486 (PT), Ser34/Ser486 (SS), and Pro34/Ser486 (wild type, CYP2D6.1), were constructed by PCR-site mutagenesis to clarify the effects of the two amino-acid substitutions. The expression of CYP2D6 protein was confirmed by immunoblotting using CYP2D antibody. The absorbance at 450 nm was measured by CO-reduced difference spectra from five ail microsome preparations. The CYP2D6 forms with Pro34 --> Ser amino acid substitution were at a lower expression than CYP2D6.1 from the findings of immunoblotting and spectral analysis, The apparent K-m values of CYP2D6.1, CYP2D6.10A, and CYP2D6.10C were 1.7, 8.5, and 49.7 mu M, respectively, for bufuralol 1'-hydroxylation, and 9.0, 51.9, and 117.4 mu M, respectively, for venlafaxine O-demethylation, respectively, The V-max values were not significantly different among the three variants. These findings suggest that the decreased in vivo clearance by CYP2D6*10 was caused not only by low expression of but also the increased K-m value of CYP2D6. (C) 2000 Academic Press.