Metabolism of Ticlopidine in Rats: Identification of the Main Biliary Metabolite as a Glutathione Conjugate of TiclopidineS-Oxide

Abstract

ABSTRACT:We have identified several novel metabolites of ticlopidine, a well known antiplatelet agent and have revealed its metabolic route in rats. The main biliary metabolite of ticlopidine was characterized as a glutathione (GSH) conjugate of ticlopidine S-oxide, in which conjugation had occurred at carbon 7a in the thienopyridine moiety. Quantitative analysis revealed that 29% of the dose was subjected to the formation of reactive intermediates followed by conjugation with GSH after oral administration of ticlopidine (22 mg/ kg) to rats. In vitro incubation of ticlopidine with rat liver 9000g supernatant fraction (S9) fractions led to the formation of multiple metabolites, including 2-oxo-ticlopidine, the precursor for the pharmacologically active ticlopidine metabolite, [1-(2-chlorobenzyl)-4-mercaptopiperidin-(3Z)-ylidene] acetic acid. A novel thiophene ring-opened metabolite with a thioketone group and a carboxylic acid moiety has also been detected after incubation of 2-oxo-ticlopidine with rat liver microsomes or upon incubation of ticlopidine with rat liver S9 fractions.Ticlopidine is a well known antiplatelet agent (Quinn and Fitzgerald, 1999) and has been widely used for the secondary prevention of atherothrombosis (Jacobson, 2004). The metabolism of ticlopidine is complex because of extensive oxidation in the liver (Saltiel and Ward, 1987). For example, less than 1% of the parent compound was detected in urine, whereas approximately 60 and 25% of radioactivity was recovered in urine after a single oral administration of [ 14 C]ticlopidine to humans (Noble and Goa, 1996) and rats (Tuong et al., 1981), respectively. Furthermore, a large difference was observed between the total radioactivity and unchanged ticlopidine level in the plasma (Panak et al., 1983). In the 1980s, metabolic studies suggested that N-dealkylation, N-oxidation, and oxidation of the thiophene ring followed by ring opening appeared to be the main routes, but numerous highly polar urinary and biliary metabolites in both humans and animals remained unidentified (Tuong et al., 1981;Panak et al., 1983). More recently, novel ticlopidine metabolites such as the S-oxide form (Ha-Duong et al., 2001) and the pharmacologically active metabolite (Yoneda et al., 2004) have been detected. However, the whole metabolism of ticlopidine has remained unclarified.In the present study, we have identified several novel metabolites of ticlopidine and revealed the metabolic pathways of ticlopidine quantitatively. Moreover, an in vitro experiment was conducted to elucidate the whole metabolic route of ticlopidine including the biotransformation to the pharmacologically active metabolite.

Ticlopidine, a Cholestatic Liver Injury-Inducible Drug, Causes Dysfunction of Bile Formation via Diminished Biliary Secretion of Phospholipids: Involvement of Biliary-Excreted Glutathione-Conjugated Ticlopidine Metabolites

“…The biliary output of TIC-SGs in rats treated with SKF-525A and TIC was much less than that in rats treated with TIC alone (Fig. 2B), consistent with the previous report that TIC-SGs were formed in the liver after P450s-mediated metabolism (Shimizu et al, 2009).…”

“…Because TIC reportedly undergoes extensive metabolism in the liver and ,1% of the parent compound is detected in urine (Shimizu et al, 2009), we examined the involvement of TIC metabolites in the altered bile composition. We pretreated SD rats with SKF-525A, a nonspecific inhibitor of P450s (Yang et al, 2009), prior to the administration of TIC.…”

“…Because two types of GSH-conjugated TIC metabolites were previously detected in the bile of TIC-administered rats (Shimizu et al, 2009), we isolated these metabolites (TIC-SG1 and TIC-SG2) from rat bile and constructed quantification methods by LC-MS/MS (Supplemental Fig. S1).…”

“…Two types of GSHconjugated TIC metabolites (TIC-SGs), referred to as TIC-SG1 and TIC-SG2 (Supplemental Fig. S1), were isolated from the rat bile according to a previously reported process (Shimizu et al, 2009). Briefly, bile specimens collected from SD rats that were continuously infused with TIC at 13 mg/kg/h were applied to solid-phase extraction columns (Mega Bond Elut C 18 ; Varian Inc., Palo Alto, CA).…”

Pharmacokinetics, Metabolism, and Excretion of [14C]Axitinib, a Vascular Endothelial Growth Factor Receptor Tyrosine Kinase Inhibitor, in Humans

“…The fragment ion at m/z 372 indicated a loss of 31 amu, which was consistent with the loss observed in the fragmentation of axitinib. The fragment ion at m/z 355 resulted from a loss of 48 amu and was characteristic of a loss of a sulfoxide moiety from the molecule (Bu et al, 2007;Shimizu et al, 2009). The ions at m/z 344 and 327 indicated a loss of 28 amu from m/z 372 and 355, respectively, and suggested a loss of either a carbonyl group from the acylium intermediate that was formed from cleavage of the N-methylamide or loss of nitrogen from the indazole ring.…”