Contributions of UDP-Glucuronosyltransferases to Human Hepatic and Intestinal Metabolism of Ticagrelor and Inhibition of UGTs and Cytochrome P450 Enzymes by Ticagrelor and its Glucuronidated Metabolite

Ticagrelor is the first reversibly binding, direct-acting, oral P2Y12 receptor inhibitor. The contribution of UDP-glucuronosyltransferases (UGTs) enzymes to the metabolism of ticagrelor to its glucuronide conjugation, ticagrelor-O-glucuronide, in human liver microsomes (HLM) and human intestinal microsomes (HIM), was well characterized in the current study. The inhibition potential of human major UGTs by ticagrelor and ticagrelor-O-glucuronide was explored. The inhibitory effects of ticagrelor-O-glucuronide on cytochrome P450s (CYPs) enzymes were investigated as well. Ticagrelor glucuronidation exhibits substrate inhibition kinetics in both HLM and HIM with apparent Km values of 5.65 and 2.52 μM, Vmax values of 8.03 and 0.90 pmol min−1·mg protein−1, Ksi values of 1,343.0 and 292.9 respectively. The in vitro intrinsic clearances (Vmax/Km) for ticagrelor glucuronidation by HLM and HIM were 1.42 and 0.36 μl min−1·mg protein−1, respectively. Study with recombinant human UGTs suggested that multiple UGT isoforms including UGT1A9, UGT1A7, UGT1A3, UGT1A4, UGT1A1, UGT2B7 and UGT1A8 are involved in the conversion of ticagrelor to ticagrelor-O-glucuronide with UGT1A9 showing highest catalytic activity. The results were further supported by the inhibition studies on ticagrelor glucuronidation with typical UGT inhibitors in pooled HLM and HIM. Little or no inhibition of UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A9 and UGT2B7 by ticagrelor and ticagrelor-O-glucuronide was noted. Ticagrelor-O-glucuronide also exhibited limited inhibitory effects toward CYP2C8, CYP2D6 and CYP3A4. In contrast, ticagrelor-O-glucuronide weakly inhibited CYP2B6, CYP2C9 and CYP2C19 activity with apparent IC50 values of 45.0, 20.0 and 18.8 μM, respectively. The potential of ticagrelor-O-glucuronide to cause drug-drug interactions warrant further study.

Overexpression of P-glycoprotein, MRP2, and CYP3A4 impairs intestinal absorption of octreotide in rats with portal hypertension

Background Portal hypertension (PH) is the main cause of complications and death in liver cirrhosis. The effect of oral administration of octreotide (OCT), a drug that reduces PH by the constriction of mesenteric arteries, is limited by a remarkable intestinal first-pass elimination. Methods The bile duct ligation (BDL) was used in rats to induce liver cirrhosis with PH to examine the kinetics and molecular factors such as P-glycoprotein (P-gp), multidrug resistance-associated protein 2 (MRP2) and cytochrome P450 3A4 (CYP3A4) influencing the intestinal OCT absorption via in situ and in vitro experiments on jejunal segments, transportation experiments on Caco-2 cells and experiments using intestinal microsomes and recombinant human CYP3A4. Moreover, RT-PCR, western blot, and immunohistochemistry were performed. Results Both in situ and in vitro experiments in jejunal segments showed that intestinal OCT absorption in both control and PH rats was largely controlled by P-gp and, to a lesser extent, by MRP2. OCT transport mediated by P-gp and MRP2 was demonstrated on Caco-2 cells. The results of RT-PCR, western blot, and immunohistochemistry suggested that impaired OCT absorption in PH was in part due to the jejunal upregulation of these two transporters. The use of intestinal microsomes and recombinant human CYP3A4 revealed that CYP3A4 metabolized OCT, and its upregulation in PH likely contributed to impaired drug absorption. Conclusions Inhibition of P-gp, MRP2, and CYP3A4 might represent a valid option for decreasing intestinal first-pass effects on orally administered OCT, thereby increasing its bioavailability to alleviate PH in patients with cirrhosis.

Breast Cancer Resistance Protein and Multidrug Resistance Protein 2 Mediate the Disposition of Leonurine-10-O-β-glucuronide

Background: Leonurine (Leo), a promising antilipemic agent that has been approved for clinical trials, is extensively metabolized into bioactive Leonurine-10-O-β-glucuronide (L-10-G) vivo. Objective: To explore the effects of breast cancer resistance protein (Bcrp) and multidrug resistance protein 2 (Mrp2) on the disposition of L-10-G. Methods: The pharmacokinetics, tissue distribution and intestinal perfusion of Leo were studied by using efflux transporter gene knockout mouse models. The enzyme kinetics via liver and intestinal microsomes were also examined. Results: After intravenous injection with Leo, the AUC0-∞ values of L-10-G in Bcrp1-/- and Mrp2-/- mice were 1.55-fold and 16.80-fold higher, respectively, than those in wild-type FVB mice (P < 0.05). After oral administration, the AUC0-∞ value of L-10-G showed a 2.82-fold increase in Mrp2-/- mice compared with wild-type FVB mice (P < 0.05). After gavage with Leo for 10 and 25 min, the bile accumulation of L-10-G in Mrp2-/- mice was 3-fold and 22-fold lower, respectively, than that in wild-type FVB mice (P < 0.05). Besides, the intestinal excreted amount of L-10-G showed 2.22-fold and 2.68-fold decrease in Bcrp1-/- and Mrp2-/- mice, respectively, compared with that in wild-type FVB mice (P < 0.05). The clearance of L-10-G decreased in liver microsomes and increased in intestinal microsomes of Bcrp1-/- and Mrp2-/- mice compared to the wild-type FVB mice (P < 0.05). Conclusion: Both Bcrp and Mrp2 are involved in the disposition of L-10-G, and Mrp2 exhibits a superior influence.

Dronedarone: the effect of diet-induced obesity on its metabolism and experimental hyperlipidemia on its metabolism and tissue distribution in the rat

Dronedarone biodistribution in hyperlipidemia and dronedarone metabolism in hyperlipidemia or obesity were assessed. Male Sprague–Dawley rats were given either normal standard chow with water or various high-fat or high-carbohydrate diets for 14 weeks. There was also a nonobese hyperlipidemic group given poloxamer 407 intraperitoneally. Liver and intestinal microsomes were prepared and the metabolic conversion of dronedarone to desbutyldronedarone was followed. A biodistribution study of dronedarone given orally was conducted in hyperlipidemic and control normolipidemic rats. The metabolism of dronedarone to desbutyldronedarone in control rats was consistent with substrate inhibition. However in the treatment groups, the formation of desbutyldronedarone did not follow substrate inhibition; hyperlipidemia and high-calorie diets created remarkable changes in dronedarone metabolic profiles and reduction in formation velocities. Tissue concentrations of dronedarone were much higher than in plasma. Furthermore, in hyperlipidemia, plasma and lung dronedarone concentrations were significantly higher compared to normolipidemia.

Survey of Drug Oxidation Activities in Hepatic and Intestinal Microsomes of Individual Common Marmosets, a New Nonhuman Primate Animal Model

Background: Common marmosets (Callithrix jacchus) are potentially useful nonhuman primate models for preclinical studies. Information for major drug-metabolizing cytochrome P450 (P450) enzymes is now available that supports the use of this primate species as an animal model for drug development. Here, we collect and provide an overview of information on the activities of common marmoset hepatic and intestinal microsomes with respect to 28 typical human P450 probe oxidations. Results: Marmoset P450 2D6/8-dependent R-metoprolol O-demethylation activities in hepatic microsomes were significantly correlated with those of midazolam 1′- and 4-hydroxylations, testosterone 6β-hydroxylation, and progesterone 6β-hydroxylation, which are probe reactions for marmoset P450 3A4/5/90. In marmosets, the oxidation activities of hepatic microsomes and intestinal microsomes were roughly comparable for midazolam and terfenadine. Overall, multiple forms of marmoset P450 enzymes in livers and intestines had generally similar substrate recognition functionalities to those of human and/or cynomolgus monkey P450 enzymes. Conclusion: The marmoset could be a model animal for humans with respect to the first-pass extraction of terfenadine and related substrates. These findings provide a foundation for understanding individual pharmacokinetic and toxicological results in nonhuman primates as preclinical models and will help to further support understanding of the molecular mechanisms of human P450 function.