Identification of the human cytochromes P450 catalysing the rate‐limiting pathways of gliclazide elimination

Abstract

What is already known about this subject • Gliclazide is a widely used oral hypoglycaemic agent. • The major metabolites of gliclazide formed in vivo have been identified. • However, the cytochrome P450 enzymes catalysing the rate‐limiting pathways of gliclazide elimination are unknown. What this study adds • CYP2C9 is the major enzyme involved in the various hydroxylation pathways of gliclazide, although a contribution of CYP2C19 to tolymethylhydroxylation, the major metabolic route, cannot be discounted. • Factors known to influence CYP2C9 activity will provide the main source of variability in gliclazide pharmacokinetics. Aims To identify the human cytochrome P450 (CYP) enzymes responsible for the formation of the 6β‐hydroxy (6β‐OHGz), 7β‐hydroxy (7β‐OHGz) and hydroxymethyl (MeOH‐Gz) metabolites of gliclizide (Gz). Methods 6β‐OHGz, 7β‐OHGz and MeOH‐Gz formation by human liver microsomes and a panel of recombinant human P450s was measured using a high‐performance liquid chromatography procedure, and the kinetics of metabolite formation was determined for each pathway. Effects of prototypic CYP enzyme selective inhibitors were characterized for each of the microsomal metabolic pathways. Results Microsomes from six human livers converted Gz to its 6β‐OHGz, 7β‐OHGz, and MeOH‐Gz metabolites, with respective mean (± SD) Km values of 461 ± 139, 404 ± 143 and 334 ± 75 µm and mean Vmax values of 130 ± 55, 82 ± 31 and 268 ± 115 pmol min−1 mg−1, respectively. Vmax/Km ratios for the microsomal reactions parallelled relative metabolite formation in vivo. Sulfaphenazole inhibited microsomal 6β‐OHGz, 7β‐OHGz and MeOH‐Gz formation by 87, 83 and 64%, respectively, whereas S‐mephenytoin caused significant inhibition (48%) of only MeOH‐Gz formation. Recombinant CYP2C9, CYP2C18 and CYP2C19 catalysed all hydroxylation pathways, whereas CYP2C8 formed only 6β‐OHGz and 7β‐OHGz. Conclusion Taken together, the results indicate that CYP2C9 is the major contributor to Gz metabolic clearance, although CYP2C19 may also be involved in MeOH‐Gz formation (the major metabolic pathway). Factors known to influence CYP2C9 activity will provide the main source of variability in Gz pharmacokinetics.

Applications of CYP450 Testing in the Clinical Setting

“…They are mainly metabolised by CYP2C9 [138], with a contribution of CYP2C19 for several of these sulfonylureas [139, 140]. The PK properties of sulfonylureas have been evaluated in few studies.…”

Role of Cytochrome P450 2C8 in Drug Metabolism and Interactions

“…Additionally, CYP2C8 is involved, to a minor extent, in the metabolism of the sulfonylureas gliclazide, glyburide, and tolbutamide, the dipeptidyl peptidase 4 inhibitor sitagliptin, and the PPARa agonist sipoglitazar (Table 1; Relling et al, 1990;Srivastava et al, 1991;Veronese et al, 1993;Elliot et al, 2007;Vincent et al, 2007;Zharikova et al, 2009;Nishihara et al, 2012). Tolbutamide p-methyl hydroxylation has been used as a marker reaction for CYP2C8 activity in several in vitro studies.…”

Pharmacogenetics in diabetes

“…This variation in hydroxylation of tolbutamide was subsequently shown to be due to variation in CYP2C9 [2]. CYP2C9 has also been shown to be a rate-limiting enzyme in the metabolism of other sulfonylureas, including glibenclamide [3], gliclazide [4], glipizide [5], and glimepiride [6]. Two variants in CYP2C9 affect the catalytic function of the enzyme: Arg144Cys (2C9*2; allele frequency 11%) and Ile359Leu (2C9*3; allele frequency 7%).…”