EVIDENCE THAT CYP2C19 IS THE MAJOR (S)-MEPHENYTOIN 4'-HYDROXYLASE IN HUMANS

The present study assesses the role of members of the human CYP2C subf amily in the 4'-hydroxylation of(S)-mephenytoin, When recombinant CYP2 C proteins were expressed using a yeast cDNA expression system, 2C19 s tereospecifically 4'-hydroxylated (S)-mephenytoin with a turnover numb er at least 10 times higher than that of human liver microsomes. 2C9 ( both Ile(359) and Leu(359) alleles) and 2C18 (Thr(385) and Met(385) al leles) metabolized this substrate at a rate 100-fold lower than 2C19, and metabolism by these 2C proteins was not stereospecific for the S-e nantiomer. 2C8 exhibited very little mephenytoin 4'-hydroxylase activi ty. In contrast, the Ile(359) allele of 2C9 had a high turnover number for the hydroxylation of tolbutamide, while the Leu(359) allele was l ess active toward this substrate. Immunoblot analysis of 16 human live r donor samples indicated that (S)-mephenytoin 4'-hydroxylase activity correlated with the hepatic CYP2C19 content, but it did not correlate with the hepatic content of CYP2C9. Moreover, direct sequencing of th e polymerase chain reaction (PCR) products of 2C9 mRNA from six of the se human livers through areas of known allelic variations indicated th at the identity of the allele of 2C9 (Cys(144) vs Arg, Tyr(358) vs Cys , Ile(359) vs Leu, or Gly(417) VS Asp) did not appear to influence (S) -mephenytoin 4'-hydroxylase activity in these samples. These data indi cate that 2C19 is the principal determinant of (S)-mephenytoin 4'-hydr oxylase activity in human liver.