Effect of UGT2B7*2 and CYP2C8*4 polymorphisms on diclofenac metabolism

Abstract

The use of diclofenac is associated with rare but severe drug-induced liver injury (DILI) in a very small number of patients. The factors which predispose susceptible patients to hepatotoxicity of diclofenac are still incompletely understood. Formation of protein-reactive metabolites by UDP-glucuronosyl transferases and cytochromes P450 is commonly considered to play an important role, as indicated by the detection of covalent protein adducts and antibodies in the serum of patients suffering from diclofenac-induced liver injury. Since no associations have been found with HLA-alleles, polymorphisms of genes encoding for proteins involved in the disposition of diclofenac may be important. Previous association studies showed that possession of the UGT2B7*2 and CYP2C8*4 alleles is more common in cases of diclofenac-induced DILI. In the present study, the metabolism of diclofenac by UGT2B7*2 and CYP2C8*4 was compared with their corresponding wild-type enzymes. Enzyme kinetic analysis revealed that recombinant UGT2B7*2 showed an almost 6-fold lower intrinsic clearance of diclofenac glucuronidation compared to UGT2B7*1. The mutant CYP2C8*4 showed approximately 35% reduced activity in the 4'-hydroxylation of diclofenac acyl glucuronide. Therefore, a decreased hepatic exposure to diclofenac acyl glucuronide is expected in patients with the UGT2B7*2 genotype. The increased risk for hepatotoxicity, therefore, might be the result from a shift to oxidative bioactivation to cytotoxic quinoneimines.

Biochemical alterations in diclofenac-treated rats: Effect of selenium on oxidative stress, inflammation, and hematological changes

“…DCF is safe at therapeutic doses and can be toxic in humans and animals when administered in large doses. Toxcities associated with DCF are as a result of its reactive metabolites, 4 0 -OH-DCF and 5 0 -OH-DCF 19,22,43 and the highly reactive benzoquinone imines, 19,20 conjugation with reduced GSH and inactivated by GSTs, 23,24 enhances DCF-metabolite excretion 44 with subsequent reduction in DCF toxicity. Induction of biodisposition enzyme containing Sel and enhancement of cellular antioxidant status will therefore reduce DCF-associated toxicities.…”

“…11,16 In the liver, DCF is metabolized to 4-hydroxydiclofenac (4 0 -OH-DCF) by CYP2C9 and a minor metabolite, 5-hydroxydiclofenac (5 0 -OH-DCF), by CYP 3A4 17,18 and other hydroxylated forms, which further undergo phase II reaction such as glucuronidation and sulfation prior to excretion in the urine (65%) and bile (35%). 10 4 0 -OH-DCF and 5 0 -OH-DCF can further be metabolized into highly reactive benzoquinone imines 19,20 by cytochrome CYP450 2C9. This reactive intermediate selectively reacts with protein cysteine 19 causing damage to rats' livers.…”

“…21 Increase in DCF concentration leads to increase in the activities of hepatic transaminases, lactate dehydrogenase (LDH), and DCF reactive metabolites. 17 Inhibition of CYPs is reported to confer protection on rat hepatocytes 20 ; however, DCF quinoneimines have been shown to be inactivated by glutathione-S-transferases (GSTs), 22 NAD(P)H:quinone oxidoreductase1 by conjugation with glutathione (GSH), and reduction reaction. 20,23,24 Earlier studies attribute the mechanisms of DCF toxicity to induction of oxidative stress, mitochondrial damage, and the interaction of its reactive metabolite with cellular macromolecules, leading to alteration of proteins integrity.…”

“…17 Inhibition of CYPs is reported to confer protection on rat hepatocytes 20 ; however, DCF quinoneimines have been shown to be inactivated by glutathione-S-transferases (GSTs), 22 NAD(P)H:quinone oxidoreductase1 by conjugation with glutathione (GSH), and reduction reaction. 20,23,24 Earlier studies attribute the mechanisms of DCF toxicity to induction of oxidative stress, mitochondrial damage, and the interaction of its reactive metabolite with cellular macromolecules, leading to alteration of proteins integrity. 16,25 Low intracellular level of nicotinamide adenine dinucleotide (NADH), GSH, nicotinamide adenine dinucleotide phosphate (NADPH), and other reducing agents potentiate the binding of DCF to macromolecules and important hepatic proteins.…”

Prediction of Human Hepatic Clearance for Cytochrome P450 Substrates <i>via</i> a New Culture Method Using the Collagen Vitrigel Membrane Chamber and Fresh Hepatocytes Isolated from Liver Humanized Mice

“…4) Additionally, there is a possibility of the involvement of other CYP and/or non-CYP enzymes, even when the test substrates are selective for one CYP. For example, diclofenac is metabolized by CYP2C9 and uridine 5′-diphospho-glucuronosyltransferase (UGT), 20) resulting in a large in vivo and in vitro variation. Regarding alprazolam, a CYP3A substrate, CL was overestimated 7.2 times in the vitrigel culture method.…”