Kumar Sanjeev, Samuel Koppara, Subramanian Ramaswamy, Braun Matthew P, Stearns Ralph A, Chiu Shuet-Hing Lee, Evans David C, Baillie Thomas A
Department of Drug Metabolism, Merck Research Laboratories, Rahway, New Jersey 07065, USA.
J Pharmacol Exp Ther. 2002 Dec;303(3):969-78. doi: 10.1124/jpet.102.038992.
Diclofenac is eliminated predominantly (approximately 50%) as its 4'-hydroxylated metabolite in humans, whereas the acyl glucuronide (AG) pathway appears more important in rats (approximately 50%) and dogs (>80-90%). However, previous studies of diclofenac oxidative metabolism in human liver microsomes (HLMs) have yielded pronounced underprediction of human in vivo clearance. We determined the relative quantitative importance of 4'-hydroxy and AG pathways of diclofenac metabolism in rat, dog, and human liver microsomes. Microsomal intrinsic clearance values (CL(int) = V(max)/K(m)) were determined and used to extrapolate the in vivo blood clearance of diclofenac in these species. Clearance of diclofenac was accurately predicted from microsomal data only when both the AG and the 4'-hydroxy pathways were considered. However, the fact that the AG pathway in HLMs accounted for ~75% of the estimated hepatic CL(int) of diclofenac is apparently inconsistent with the 4'-hydroxy diclofenac excretion data in humans. Interestingly, upon incubation with HLMs, significant oxidative metabolism of diclofenac AG, directly to 4'-hydroxy diclofenac AG, was observed. The estimated hepatic CL(int) of this pathway suggested that a significant fraction of the intrahepatically formed diclofenac AG may be converted to its 4'-hydroxy derivative in vivo. Further experiments indicated that this novel oxidative reaction was catalyzed by CYP2C8, as opposed to CYP2C9-catalyzed 4'-hydroxylation of diclofenac. These findings may have general implications in the use of total (free + conjugated) oxidative metabolite excretion for determining primary routes of drug clearance and may question the utility of diclofenac as a probe for phenotyping human CYP2C9 activity in vivo via measurement of its pharmacokinetics and total 4'-hydroxy diclofenac urinary excretion.
双氯芬酸在人体内主要以其4'-羟基化代谢物的形式被清除（约50%），而酰基葡萄糖醛酸（AG）途径在大鼠（约50%）和狗（>80 - 90%）中似乎更为重要。然而，先前关于双氯芬酸在人肝微粒体（HLM）中氧化代谢的研究对人体体内清除率的预测明显偏低。我们确定了双氯芬酸在大鼠、狗和人肝微粒体中代谢的4'-羟基和AG途径的相对定量重要性。测定了微粒体固有清除率值（CL(int) = V(max)/K(m)），并用于推断这些物种中双氯芬酸的体内血药清除率。只有同时考虑AG和4'-羟基途径时，才能从微粒体数据准确预测双氯芬酸的清除率。然而，HLM中的AG途径占双氯芬酸估计肝CL(int)的约75%这一事实，显然与人类中4'-羟基双氯芬酸的排泄数据不一致。有趣的是，在与HLM孵育时，观察到双氯芬酸AG直接氧化代谢为4'-羟基双氯芬酸AG。该途径估计的肝CL(int)表明，肝内形成的双氯芬酸AG的很大一部分可能在体内转化为其4'-羟基衍生物。进一步的实验表明，这种新的氧化反应是由CYP2C8催化的，而不是由CYP2C9催化的双氯芬酸4'-羟基化反应。这些发现可能对使用总（游离+结合）氧化代谢物排泄来确定药物清除的主要途径具有普遍意义，并可能质疑双氯芬酸作为通过测量其药代动力学和总4'-羟基双氯芬酸尿排泄来体内表型化人CYP2C9活性的探针的效用。
