Hamman M A, Thompson G A, Hall S D
Division of Clinical Pharmacology, Indiana University School of Medicine, Indianapolis 46202, U.S.A.
Biochem Pharmacol. 1997 Jul 1;54(1):33-41. doi: 10.1016/s0006-2952(97)00143-3.
The cytochrome P450s responsible for the regio- and stereoselectivity in the 2- and 3-hydroxylation of the chiral non-steroidal antiinflammatory drug ibuprofen were characterized in human liver microsomes. The rates of formation of both the 2- and 3-hydroxy metabolites exhibited monophasic (N = 2; N is the number of microsomal preparations) and biphasic (N = 2) substrate concentration dependence for both enantiomers of ibuprofen. The high affinity enzyme class parameters for S-ibuprofen (N = 4) were: 2-hydroxylation, Vmax = 566 +/- 213 pmol/min/mg, Km = 38 +/- 13 microM; 3-hydroxylation, Vmax = 892 +/- 630 pmol/min/mg, Km = 21 +/- 6 microM. For R-ibuprofen, the corresponding parameters were: 2-hydroxylation, Vmax = 510 +/- 117 pmol/min/mg, Km = 47 +/- 20 microM; 3-hydroxylation, Vmax = 593 +/- 113 pmol/min/mg, Km = 29 +/- 8 microM. cDNA-expressed CYP2C9 (Arg 144 and Cys 144) favored S-2- and S-3-hydroxyibuprofen formation, but CYP2C8 favored R-2-hydroxyibuprofen formation. Sulfaphenazole, retinol, and arachidonic acid competitively inhibited the rate of formation of all hydroxyibuprofens; Ki values (N = 3) for sulfaphenazole on the 2- and 3-hydroxylations of S-ibuprofen were 0.12 +/- 0.05 and 0.07 +/- 0.04 and of R-ibuprofen were 0.11 +/- 0.07 and 0.06 +/- 0.03 microM, respectively. Sulfaphenazole also competitively inhibited ibuprofen hydroxylation by cDNA-expressed CYP2C9 (Arg 144 and Cys 144) with Ki values in the range of 0.05 to 0.18 microM and CYP2C8 in the range of 0.36 to 0.55 microM. In a bank of 14 human liver microsome samples, significant correlations (r = 0.72 to 0.90; P < 0.01) were observed between the rates of formation of all four hydroxyibuprofens, and for each hydroxyibuprofen and prototypical CYP2C8/9 biotransformations. The regio- and stereoselectivities observed in vitro were consistent with those noted in vivo. The relative levels of both CYP2C8 and CYP2C9 and the expression of the corresponding variants may influence the disposition of ibuprofen in vivo.
在手性非甾体抗炎药布洛芬的2-和3-羟基化反应中负责区域和立体选择性的细胞色素P450酶在人肝微粒体中得到了表征。布洛芬两种对映体的2-和3-羟基代谢物的形成速率均呈现单相（N = 2；N为微粒体制剂的数量）和双相（N = 2）底物浓度依赖性。S-布洛芬（N = 4）的高亲和力酶类参数为：2-羟基化，Vmax = 566 ± 213 pmol/min/mg，Km = 38 ± 13 μM；3-羟基化，Vmax = 892 ± 630 pmol/min/mg，Km = 21 ± 6 μM。对于R-布洛芬，相应参数为：2-羟基化，Vmax = 510 ± 117 pmol/min/mg，Km = 47 ± 20 μM；3-羟基化，Vmax = 593 ± 113 pmol/min/mg，Km = 29 ± 8 μM。cDNA表达的CYP2C9（Arg 144和Cys 144）有利于S-2-和S-3-羟基布洛芬的形成，但CYP2C8有利于R-2-羟基布洛芬的形成。磺胺苯吡唑、视黄醇和花生四烯酸竞争性抑制所有羟基布洛芬的形成速率；磺胺苯吡唑对S-布洛芬2-和3-羟基化反应的Ki值（N = 3）分别为0.12 ± 0.05和0.07 ± 0.04，对R-布洛芬的Ki值分别为0.11 ± 0.07和0.06 ± 0.03 μM。磺胺苯吡唑还竞争性抑制cDNA表达的CYP2C9（Arg 144和Cys 144）介导的布洛芬羟基化反应，Ki值在0.05至0.18 μM范围内，对CYP2C8的抑制作用Ki值在0.36至0.55 μM范围内。在一组14个人肝微粒体样本中，观察到所有四种羟基布洛芬的形成速率之间以及每种羟基布洛芬与典型的CYP2C8/9生物转化之间存在显著相关性（r = 0.72至0.90；P < 0.01）。体外观察到的区域和立体选择性与体内观察到的一致。CYP2C8和CYP2C9的相对水平以及相应变体的表达可能会影响布洛芬在体内的处置。
