Perspectives in Pharmacology Pharmacological and Physiological Functions of the Polyspecific Organic Cation Transporters: OCT1, 2, and 3 (SLC22A1-3)

For the elimination of environmental toxins and metabolic waste products, the body is equipped with a range of broadspecificity transporters that are generally present in the liver, kidney, and intestine. The polyspecific organic cation transporters OCT1, 2, and 3 (SLC22A1-3) mediate the facilitated transport of a variety of structurally diverse organic cations, including many drugs, toxins, and endogenous compounds. OCT1 and OCT2 are found in the basolateral membrane of hepatocytes, enterocytes, and renal proximal tubular cells. OCT3 has a more widespread tissue distribution and is considered to be the major component of the extraneuronal monoamine transport system (or uptake-2), which is responsible for the peripheral elimination of monoamine neurotransmitters. Studies with knockout mouse models have directly demonstrated that these transporters can have a major impact on the pharmacological behavior of various substrate organic cations. The recent identification of polymorphic genetic variants of human OCT1 and OCT2 that severely affect transport activity thus suggests that some of the interpatient differences in response and sensitivity to cationic drugs may be caused by variable activity of these transporters. The body is continuously exposed to a variety of environmental toxins and metabolic waste products. To rid itself of these compounds, it is equipped with various detoxification mechanisms such as metabolizing enzymes and transport proteins mediating their inactivation and excretion. For excretion, a plethora of transmembrane transport proteins is present in the major excretory organs: liver, kidney, and intestine. The solute carrier (SLC) superfamily is by far the largest superfamily of transporters, consisting of about 225 members in humans (see Human Genome Organization at http://www.gene.ucl.ac.uk/nomenclature). Whereas most of these transporters are highly specialized, mediating facilitated transport of essential nutrients (e.g., glucose, amino acids, nucleosides, and fatty acids), some members are more generalized. Due to their broad substrate specificity, the latter are also termed polyspecific transporters. They play a major role in the elimination of, and protection against, noxious compounds. Among the SLC superfamily, two families (SLC21 and 22) with polyspecific members have been identified, together mediating the transport of a variety of structurally diverse organic anions, cations, and uncharged compounds. The SLC21 family of organic anion transporting polypeptides currently consists of nine members in humans, transporting a range of relatively large (usually 450 Da), mostly anionic amphipathic compounds, including bile salts, eicosanoids, steroid hormones, and their conjugates (for review, see Hagenbuch and Meier, 2003). The SLC22 family currently consists of 12 members in humans and rats, encompassing organic cation transporters (OCTs), the carnitine transporter (OCTN2/SLC22A5) (Wu et al., 1998b), the urate anion-exchanger (URAT1/SLC22A12) (Enomoto et al., 2002), and several organic anion transporters (for review, see Sweet et al., 2001). In this review, we focus on the polyspecific organic Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. DOI: 10.1124/jpet.103.053298. ABBREVIATIONS: SLC, solute carrier superfamily; OCT, organic cation transporter in human and rat; TMD, transmembrane domain; EMT, extraneuronal monoamine transporter; TEA, tetraethylammonium; MPP , 1-methyl-4-phenylpyridinium; RT-PCR, reverse transcription-polymerase chain reaction; Oct, organic cation transporter in mice. 0022-3565/04/3081-2–9$20.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 308, No. 1 Copyright © 2004 by The American Society for Pharmacology and Experimental Therapeutics 53298/1123140 JPET 308:2–9, 2004 Printed in U.S.A. 2 at A PE T Jornals on July 9, 2017 jpet.asjournals.org D ow nladed from cation transporters (OCT1, 2, and 3) that belong to the family SLC22 (Koepsell and Endou, 2003; Koepsell et al., 2003). Cloning and Functional Characteristics The first identified member of the organic cation transporter family, OCT1 (SLC22A1), was isolated by expression cloning from rat kidney (Gründemann et al., 1994). In this initial study, it was shown that rOCT1 had functional characteristics similar to the previously described organic cation transport process in the basolateral membrane of renal proximal tubules and hepatocytes. rOCT1 encodes a 556-amino acid protein and has a proposed secondary structure displaying 12 transmembrane domains (TMD). It contains a large extracellular loop, located between the first and second TMD, with three predicted N-linked glycosylation sites (MeyerWentrup et al., 1998) that may be involved in protein stability, intracellular routing, or protection from extracellular proteases (Fig. 1). Currently, mammalian orthologs of OCT1 have been cloned from mouse (Schweifer and Barlow, 1996), human (Gorboulev et al., 1997; Zhang et al., 1997b), and rabbit (Terashita et al., 1998). By homology screening, a second member of the organic cation transporter family, designated OCT2 (SLC22A2), was isolated from rat kidney (Okuda et al., 1996) and later also cloned from human (Gorboulev et al., 1997), pig (Gründemann et al., 1997), mouse (Mooslehner and Allen, 1999), and rabbit (Zhang et al., 2002). rOCT2 encodes a 593-amino acid protein with a calculated molecular mass of 66 kDa and 67% identity with rOCT1. The third member of the organic cation transporter family, designated OCT3 (SLC22A3), was independently cloned and identified as the extraneuronal monoamine transporter (EMT, see section catecholamine transport) by two different groups (Gründemann et al., 1998b; Kekuda et al., 1998; Wu et al., 1998a) and later also cloned from mouse (Verhaagh et al., 1999). rOCT3 encodes a 551amino acid protein with a predicted molecular mass of 61 kDa and 48% identity with rOCT1 (Kekuda et al., 1998). The functional characteristics of these transporters have been extensively investigated using cRNA injected Xenopus laevis oocytes and transfected mammalian cell lines. OCT1, 2, and 3 all mediate the facilitated transport of a broad range of structurally diverse organic cations, and they have extensively overlapping substrate specificities. In general, the OCTs mediate the (bidirectional) transport of small hydrophilic compounds, ranging in size from about 60 to 350 Da, with at least one positively charged amine moiety at physiological pH. Although many compounds have been shown to inhibit or modulate transport activity of the OCTs, not all of them are transported substrates. Substrates for which transport has been directly demonstrated include the model substrate tetraethylammonium (TEA), the parkinsonian neurotoxin 1-methyl-4-phenylpyridinium (MPP ), clinically used drugs such as antiparkinsonians (amantadine and memantine), antidiabetics (biguanides) and the H2 receptor agonist cimetidine, biogenic amines (dopamine, norepinephrine), and several other endogenous compounds (choline and creatinine). In addition to organic cations, it has also been demonstrated that hOCT1 and hOCT2 mediate the transport of some anionic prostaglandins (Kimura et al., 2002), indicating that a positive charge is no absolute prerequisite for OCT substrates (Table 1). Expression and Subcellular Localization Northern analysis and RNA in situ hybridization demonstrated that in rats, rOCT1 mRNA is expressed in liver, kidney, and intestine (Gründemann et al., 1994). In humans, hOCT1 is primarily expressed in the liver, indicating a difference in tissue distribution of OCT1 between humans and rodents (Gorboulev et al., 1997; Zhang et al., 1997b). OCT2 mRNA was detected predominantly in the kidney in rats and humans (Okuda et al., 1996; Gorboulev et al., 1997). The tissue distribution and subcellular localization of OCT1 and OCT2 have been analyzed by immunohistochemistry in rats and humans (Meyer-Wentrup et al., 1998; Karbach et al., 2000; Sugawara-Yokoo et al., 2000; Motohashi et al., 2002). In the liver, rOCT1 was detected in sinusoidal membranes of hepatocytes around the central veins of the hepatic lobuli. In the kidney, rOCT1 was mainly observed in the pars convoluta (S1) and cortical pars recta (S2) of the proximal tubules, with lower expression in the medullary pars recta (S3), whereas rOCT2 was mainly expressed in the S2 and S3 segments (Meyer-Wentrup et al., 1998; Karbach et al., 2000). By Western analysis, using isolated renal basolateral and brush-border (apical) membranes of proximal tubules, it was shown that both rOCT1 and rOCT2 are localized basolaterally (Urakami et al., 1998; Karbach et al., 2000) (Fig. 2). In addition to the kidney, OCT2 is also expressed in various regions of the brain. By RT-PCR, rOCT2 was detected in dopamine-rich areas of the brain: substantia nigra, nucleus accumbens, and striatum (Gründemann et al., 1997). In humans, hOCT2 was detected by in situ hybridization and immunohistochemistry in the pyramidal cells of the cerebral cortex and hippocampus. In the brain, OCT2 might represent a “background” transporter for the removal of monoamine neurotransmitters that have escaped reuptake by high-affinity monoamine transporters, which are not members of the OCT family (Busch et al., 1998). In contrast to what has been reported by Gründemann et al. (1997), rOCT2 (and rOCT3) was also detected by RT-PCR in the choroid plexus (Sweet et al., 2001). Transfection of intact choroid plexus with an rOCT2-GFP fusion protein construct resulted in strong apiFig. 1. Predicted secondary structure of OCT1. The protein is thought to contain 12 transmembrane domains with both N and C terminus located intracellularly. The first large extracellular loop contains three putative N-linked glycosylation sites (indicated by branches). Cytoplasmic (IN) and extracellular (OUT) orientation are indicated. Organic Cation Transporters OCT1, 2, and 3 3 at A PE T Jornals on July 9, 2017 jpet.asjournals.org D ow nladed from