Interplay of Phase II Enzymes and Transporters in Futile Cycling: Influence of Multidrug Resistance-Associated Protein 2-Mediated Excretion of Estradiol 17β-d-Glucuronide and Its 3-Sulfate Metabolite on Net Sulfation in Perfused TR⁻and Wistar Rat Liver Preparations

Abstract

ABSTRACT:The hepatic disposition of estradiol 17␤-D-glucuronide (E 2 17G), a substrate of the organic anion-transporting polypeptides Oatp1a1, Oatp1a4, and Oatp1b2, was investigated in Wistar and E 2 3S17G (11؋) in liver and reduced net sulfation (40 ؎ 6 from 77 ؎ 6% dose, P < 0.05) were observed at 2 h for the TR ؊ versus the Wistar rats. With use of a physiologically based pharmacokinetic model, analytical solutions for the areas under the curve for the precursor and metabolite were obtained to reveal how enzymeand transporter-mediated processes affected the hepatic disposition of the precursor and metabolite in futile cycling. The analytical solutions were useful to explain transporter-enzyme interplay in futile cycling and predicted that a shutdown of Mrp2 function led to decreased net sulfation of E 2 17G by raising the intracellular concentration of the metabolite, E 2 3S17G, which readily refurnished E 2 17G via desulfation.The liver is the most important drug eliminatory organ and involves enzymes and basolateral influx/efflux and canalicular transporters for entry and elimination. The basolateral influx transporters [organic anion-transporting polypeptides (Oatps), sodium-dependent taurocholate-cotransporting polypeptide, organic anion transporter 2, and organic cation transporter 1] regulate the entry of substrate into the cell to undergo metabolism by phase I and/or phase II enzymes. Then both the parent drug and phase II metabolites are subject to canalicular transport via P-glycoprotein, multidrug resistance-associated protein (MRP) 2, the bile salt export pump, and breast cancer resistance protein, whereas basolateral efflux occurs via MRP3 and MRP4 into sinusoidal blood. Although metabolism is normally deemed to be irreversible, the metabolite, on occasion, may re-form the parent drug. This phenomenon, known as "reversible metabolism" or "futile cycling," describes the interconversion between the parent drug and its metabolite and can occur between a precursor and its phase I (Meffin et al., 1983;Baillie et al., 2001) or phase II (Hansel and Morris, 1996;Grubb et al., 1999) metabolites. For example, the sulfated metabolite formed via sulfotransferases (SULTs) in the cytosol may access the arylsulfatases in the endoplasmic reticulum to become desulfated to re-form the parent drug (Ratna et al., 1993;Kauffman et al., 1998). Acinar heterogeneity of the SULTs further complicates the scenario (Xu et al., 1993;Tan et al., 2001).A well-stirred liver model with membrane barriers has been shown to be useful to relate the physiological and biochemical factors to hepatic drug and metabolite processing (Sirianni and Pang, 1997;Liu and Pang, 2005;Sun and Pang, 2010). In brief, transporter or enzyme activity is denoted by the intrinsic clearance or the ratio of V max , the maximum rate of the enzyme or transporter-mediated process, and K m , the Michaelis-Menten constant, under linear conditions. CL in and Article, publication date, and citation information can be found at

Impact of Intestinal Glucuronidation on the Pharmacokinetics of Raloxifene

“…Therefore, prediction of Fg for such compounds, which are often eliminated through phase II metabolism and subsequent excretion, is becoming essential. For anionic compounds, the interactions of conjugating enzymes with transporters have been increasingly recognized as an important process of elimination (Nies et al, 2008;Pang et al, 2009;Sun et al, 2010). Therefore, comprehensive studies including metabolism and transport are required.…”

Hepatocellular Exposure of Troglitazone Metabolites in Rat Sandwich-Cultured Hepatocytes Lacking Bcrp and Mrp2: Interplay between Formation and Excretion

“…Overall, decreased biliary excretion combined with increased glucuronidation led to significantly increased cellular accumulation of TG in TR -compared with WT rat SCH. Medium concentrations of TG also were increased in TR -rat SCH, consistent with increased glucuronidation and increased expression of the basolateral efflux transporter Mrp3 (Johnson et al, 2006;Sun et al, 2010).…”

“…In rats, TS formation is four times faster in male rats compared with female rats, which is consistent with greater expression of Sult1a1 and Sult1e1 in male rats (Demyan et al, 1992;Liu and Klaassen, 1996). Interestingly, protein levels of hepatic Sult1a1 and Sult1e1 were comparable between WT and TR -rats (Johnson et al, 2006;Sun et al, 2010), and our in vitro metabolism study using S9 fractions from livers of WT and TR -rats revealed that TS formation rates were comparable. A decrease in net sulfation of estradiol-17 b-glucuronide (E 2 17G) reported in TR -rats was attributed to increased desulfation of estradiol-3-sulfate-17b-glucuronide (Sun et al, 2010).…”

“…Interestingly, protein levels of hepatic Sult1a1 and Sult1e1 were comparable between WT and TR -rats (Johnson et al, 2006;Sun et al, 2010), and our in vitro metabolism study using S9 fractions from livers of WT and TR -rats revealed that TS formation rates were comparable. A decrease in net sulfation of estradiol-17 b-glucuronide (E 2 17G) reported in TR -rats was attributed to increased desulfation of estradiol-3-sulfate-17b-glucuronide (Sun et al, 2010). However, desulfation of TS to TGZ was not observed in S9 fraction from livers of WT or TR -rats.…”

“…TR -rat SCH also may have higher cellular concentrations of endogenous substrates (e.g., bile acids) that deplete PAPS, leading to decreased recovery of TS. Compared with WT rats, hepatic sulfation activities in TR -rats have been reported to be modestly increased (i.e., acetaminophen, 4-methylumbelliferone), similar (i.e., harmol, E 2 17G), or decreased (i.e., resveratrol) in isolated perfused liver studies (Xiong et al, 2000;Zamek-Gliszczynski et al, 2005;Zamek-Gliszczynski et al, 2006;Maier-Salamon et al, 2008;Zamek-Gliszczynski et al, 2008;Sun et al, 2010). However, with the exception of the harmol and E 2 17G studies, recovery was measured only in perfusate and bile, and mass balance did not account for recovery in the liver tissue.…”