VandenBranden M, Ring B J, Binkley S N, Wrighton S A
Department of Drug Metabolism and Disposition, Lilly Research Laboratories, Eli Lilly and Co., Lilly Corporate Center, Indianapolis, IN 46285, USA.
Pharmacogenetics. 1996 Feb;6(1):81-91. doi: 10.1097/00008571-199602000-00007.
The interactions in vitro of LY307640 with the cytochromes P450 (P450s) were studied using human liver microsomes, specific inhibitors of the P450s, and cDNA expressed enzymes. The kinetics of formation of the two major oxidative metabolites, desmethyl LY307640 and LY307640 sulfone, were determined using two human liver microsomal samples. The kinetic data indicated that high and low affinity sites were present for the production of both metabolites of LY307640. The Km(apparent) and Vmax(apparent) for desmethyl LY307640 formation by microsomes from human liver E (HL-E) for the high affinity site were 18.8 +/- 4.4 microM and 402 +/- 52 pmol product/min/mg protein. The high affinity site Km(apparent) and Vmax(apparent) for LY307640 sulfone formation by microsomes from HL-E were 4.4 +/- 2.1 microM and 81.8 +/- 18 pmol product/min/mg protein. The rates of desmethyl LY307640 and LY307640 sulfone formation by the high affinity site were determined using 14 human liver microsomal samples characterized for P450 marker catalytic activities and immunoquantified levels of the P450s. Rates of formation of desmethyl LY307640 significantly correlated with the immunoquantified levels of CYP 2C19 and the ability of the microsomes to 4'-hydroxylate S-mephenytoin. LY307640 sulfone formation significantly correlated with the immunoquantified levels of CYP 3A and the ability of the microsomes to 1'-hydroxylate midazolam. Inhibition studies and use of expressed cytochrome P450 systems confirmed the correlation data demonstrating that CYP 2C19 catalyzed the formation of desmethyl LY307640 and CYP 3A and catalyzed LY307640 sulfone formation. Further, LY307640 competitively inhibited S-mephenytoin 4'-hydroxylation and midazolam 1'-hydroxylation as did the structurally related compound, omeprazole. For the inhibition of S-mephenytoin 4'-hydroxylation and midazolam 1'-hydroxylation, LY307640 had higher Ki(apparent) values than that of omeprazole. These studies demonstrate that the high affinity enzymes which catalyze the formation of the desmethyl and sulfone metabolites of LY307640 are, respectively, CYP 2C19 and CYP 3A. In addition, the inhibition data suggest that LY307640 has less potential to inhibit the metabolism of CYP 2C19 substrates compared to omeprazole, and that LY307640 and omeprazole have a similarly low potential to inhibit the metabolism of CYP 3A substrates.
利用人肝微粒体、细胞色素P450（P450s）的特异性抑制剂和cDNA表达酶，研究了LY307640在体外与细胞色素P450的相互作用。使用两份人肝微粒体样品测定了两种主要氧化代谢产物去甲基LY307640和LY307640砜形成的动力学。动力学数据表明，LY307640的两种代谢产物生成均存在高亲和力和低亲和力位点。人肝E（HL-E）微粒体形成去甲基LY307640的高亲和力位点的表观米氏常数（Km(apparent)）和表观最大反应速率（Vmax(apparent)）分别为18.8±4.4微摩尔和402±52皮摩尔产物/分钟/毫克蛋白。HL-E微粒体形成LY307640砜的高亲和力位点的表观米氏常数（Km(apparent)）和表观最大反应速率（Vmax(apparent)）分别为4.4±2.1微摩尔和81.8±18皮摩尔产物/分钟/毫克蛋白。使用14份经P450标记催化活性和P450免疫定量水平表征的人肝微粒体样品，测定了高亲和力位点形成去甲基LY307640和LY307640砜的速率。去甲基LY307640的形成速率与CYP 2C19的免疫定量水平以及微粒体4'-羟基化S-美芬妥因的能力显著相关。LY307640砜的形成与CYP 3A的免疫定量水平以及微粒体1'-羟基化咪达唑仑的能力显著相关。抑制研究和表达的细胞色素P450系统的使用证实了相关数据，表明CYP 2C19催化去甲基LY307640的形成，CYP 3A催化LY307640砜的形成。此外，LY307640竞争性抑制S-美芬妥因4'-羟基化和咪达唑仑1'-羟基化，结构相关化合物奥美拉唑也有此作用。对于S-美芬妥因4'-羟基化和咪达唑仑1'-羟基化的抑制，LY307640的表观抑制常数（Ki(apparent)）值高于奥美拉唑。这些研究表明，催化LY307640去甲基和砜代谢产物形成的高亲和力酶分别是CYP 2C19和CYP 3A。此外，抑制数据表明，与奥美拉唑相比，LY307640抑制CYP 2C19底物代谢的潜力较小，并且LY307640和奥美拉唑抑制CYP 3A底物代谢的潜力同样较低。
