Integration of hepatic drug transporters and phase II metabolizing enzymes: Mechanisms of hepatic excretion of sulfate, glucuronide, and glutathione metabolites

Abstract

The liver is the primary site of drug metabolism in the body. Typically, metabolic conversion of a drug results in inactivation, detoxification, and enhanced likelihood for excretion in urine or feces. Sulfation, glucuronidation, and glutathione conjugation represent the three most prevalent classes of phase II metabolism, which may occur directly on the parent compounds that contain appropriate structural motifs, or, as is usually the case, on functional groups added or exposed by phase I oxidation. These three conjugation reactions increase the molecular weight and water solubility of the compound, in addition to adding a negative charge to the molecule. As a result of these changes in the physicochemical properties, phase II conjugates tend to have very poor membrane permeability, and necessitate carrier-mediated transport for biliary or hepatic basolateral excretion into sinusoidal blood for eventual excretion into urine. This review summarizes sulfation, glucuronidation, and glutathione conjugation reactions, as well as recent progress in elucidating the hepatic transport mechanisms responsible for the excretion of these conjugates from the liver. The discussion focuses on alterations of metabolism and transport by chemical modulators, and disease states, as well as pharmacodynamic and toxicological implications of hepatic metabolism and/or transport modulation for certain active phase II conjugates. A brief discussion of issues that must be considered in the design and interpretation of phase II metabolite transport studies follows.

Introduction

The liver has long been recognized as the primary site of drug metabolism in the body. Drug metabolism is classified into phase I and phase II reactions. Phase I metabolism usually does not result in a large change in molecular weight or water solubility of the substrate, but is of great importance because oxidative reactions add or expose sites where phase II metabolism can subsequently occur. In contrast, phase II conjugation typically results in an appreciable increase in molecular weight and water solubility. Phase I reactions are mediated primarily by the cytochrome P450 family of microsomal enzymes, but others (e.g. flavin monooxygenases, peroxidases, amine oxidases, dehydrogenases, xanthine oxidases, etc.) also catalyze oxidation of certain functional groups (Low, 1998). Inhibition, induction, and polymorphisms of cytochrome P450 enzymes are an intense area of research, because this class of metabolic enzymes is responsible for the biotransformation of the majority of drugs, thus their modulation has important therapeutic and toxic implications (Evans and Johnson, 2001, Rendic and Di Carlo, 1997). Most compounds undergo phase I oxidation prior to phase II conjugation [e.g. phenobarbital (Crayford and Hutson, 1980)], but molecules with sites amenable to conjugation may undergo phase II reactions directly [e.g. acetaminophen (Gram and Gillette, 1971)]. Molecules that undergo direct phase II conjugation may also undergo competing (e.g. acetaminophen) or additional [e.g. troglitazone (Kawai et al., 1997)] phase I oxidation.

Unbound compounds in sinusoidal blood are taken up into hepatocytes typically by a transporter-mediated mechanism or by diffusion across the basolateral membrane, depending on molecular lipophilicity, charge, size, and three-dimensional structure. Substrates of phase II metabolism are typically lipophilic (e.g. acetaminophen, 4-methylumbelliferone, dehydroepiandrosterone) and access the intracellular space by diffusion (Iida et al., 1989, Miyauchi et al., 1988, Miyazaki et al., 1983, Reuter and Mayer, 1995). In contrast, phase II conjugates formed inside the hepatocyte are typically too hydrophilic to passively diffuse across the canalicular membrane into bile or across the hepatic basolateral membrane into sinusoidal blood, and necessitate carrier-mediated transport to cross this diffusional barrier. Distinct transport proteins are present in the apical (canalicular) and basolateral (sinusoidal) domains of the hepatocyte's plasma membrane, where they efficiently pump substrates out of the cell.

This review provides a summary of the three most relevant phase II drug conjugation reactions, sulfation, glucuronidation, and glutathione conjugation, as well as documented interactions between these metabolites and hepatic efflux transporters (both canalicular and basolateral). Discussion of key issues in the study of hepatic transport of conjugates follows, with special emphasis on examples of co-modulation of phase II metabolism and transport, multiplicity of hepatic transport mechanisms, and choice of appropriate model systems for investigating the hepatic transport of conjugates.