Human Transporters

OAT2

OAT2 (organic Anion Transporter 2)

Alias: NLT
Gene name: Solute carrier family 22 member 7 (SLC22A7)

Summary

Human OAT2 is primarily expressed on the basolateral side of hepatocytes and kidney proximal tubule epithelial cells. Most likely acting as an organic anion/dicarboxylate exchanger, OAT2 mediates transport of endogenous molecules including dicarboxylic acids, nucleotides and uric acid, lipid hormones and mediators, and creatinine, as well as xenobiotics such as anticancer drugs, antivirals, and antibiotics. OAT2 is thought to be the dominant drug transporter in the eye, and its expression predicts response to combination chemotherapy in some cases of colorectal cancer. As no polymorphisms or drug interactions with clinical relevance have been reported yet, OAT2 is currently not recommended by FDA or EMA for in vitro testing.

Localization

Similar to its rodent counterpart, OAT2 is highly expressed in the liver and, to a lesser extent, in the kidney. In the rat liver, Oat2 localizes to the sinusoidal membrane of hepatocytes, and OAT2 is thought to behave similarly in the human liver, although interspecific differences cannot be excluded [1, 2]. In the kidney, subcellular localization of OAT2 is known to be species-dependent: in humans, OAT2 was seen in the basolateral membrane of proximal tubule cells [3-5], and the same was reported in cynomolgus monkeys, while in rats the transporter was observed in the apical membrane [2]. In both human and monkey kidneys, OAT2 was located along the basolateral infoldings of the cell membrane, but an apical localization was also visible [2]. Together with OAT1, OAT3 and URAT1, OAT2 was shown to be expressed in both ductal and serous acinar cells of salivary glands, where it may play a role in the transport of organic anions and uric acid [6]. As one of the main transporters responsible for ocular homeostasis and drug disposition, OAT2 is also expressed in the corneal epithelium [7]. Additionally, high OAT2 expression was observed in a subset of gastric and colorectal cancer samples [8-10].
In rats, the abundance of Oat2 in the kidney and the liver was found to be sex-dependent, with higher rOat2 expression in the female kidney and the male liver [11]. Such sex differences have not been confirmed in humans.
Although the structures of OATs are not available, hydropathy analysis suggests that OAT2 contains 12 transmembrane domains (TMD), with a large extracellular loop between TMD1 and TMD2, and a second large intracellular loop between TMD6 and TMD7. The large extracellular loop carries N-glycosylation sites important for targeting the protein to the plasma membrane, and the large intracellular loop carries consensus phosphorylation sites [12, 13].

Function, physiology, and clinically relevant polymorphisms

OAT2 was originally cloned from the rat liver. The newly discovered transporter first became known as novel liver-specific transporter, or NLT, because its mRNA levels were significantly higher in the liver than in the kidney [1]. Later, Sekine et al. recognized that NLT was able to transport organic anions with different chemical structures, and as it showed 42% homology with rat Oat1 they renamed it to Oat2 [14]. Human OAT2 shares 79% identity with rat Oat2. The homology of OAT2 with OAT1 and OAT3 is 39 and 38%, respectively [15].
In the liver and kidney, OAT2 mediates transport of various endogenous compounds. While OAT1 and OAT3 are known to operate as organic anion/dicarboxylate exchangers, the transport mechanism of OAT2 is not well characterized, and existing results are contradictory. It seems most likely that OAT2 also works as an organic anion/dicarboxylate exchanger with a preference for 4-carbon dicarboxylates [12]. OAT2 transported α-ketoglutarate, although α-ketoglutarate was unable to inhibit PAH uptake [15] or cGMP transport. No trans-stimulation effect was observed between glutarate or α-ketoglutarate and cGMP, while suberate slightly inhibited cGMP uptake. OAT2-mediated cGMP uptake was also found to be pH- and chloride-independent [16]. Fumarate and succinate, but not glutarate, have been reported to trans-stimulate ES uptake [17]. Both hOAT2 and rOat2 were able to transport orotic acid, and hOAT2 transported glutamate in both directions (uptake and efflux). In rOat2-overexpressing cells glutamate had an effect on the efflux of orotic acid. It has been proposed that OAT2 may function as a glutamate efflux transporter in hepatocytes, and OAT2 may play an active role in the hepatic release of glutamate into blood [18].
An important group of OAT2 substrates comprises pyrimidine bases, nucleosides, and nucleotides. Specifically, OAT2 was shown to transport adenine, cytidine, adenosine, inosine, guanidine (but not thymine, thymidine and cytosine), as well as cAMP, cGMP, guanosine nucleotides, and 2-deoxyguanosine [15, 19]. The major purine nucleotide metabolite uric acid is also transported by OAT2 [16]; thus, OAT2 may play a role in the first step of the tubular secretion of uric acid [20].
Some results show that OAT2 can transport hormones and endogenous prostaglandins, namely, dehydroepiandrosterone sulfate (DHEAS), estrone 3-sulfate (ES) and prostaglandin E2 [17] in a time-dependent manner [21], while Sun et al. failed to observe OAT2-mediated DHEAS transport [15]. Enomoto et al. measured OAT2-mediated prostaglandin F2α transport, which was inhibited with probenecid  [3]. Creatinine is mainly eliminated by glomerular filtration, which is complemented by active tubular secretion via OCT2. Some investigations have shown that OAT2 is able to transport creatinine more efficiently than other renal transporters and may contribute to creatinine clearance. OAT2-dependent creatinine uptake was inhibited by indomethacine and bromsulphthalein [22, 23]. The difference in localization between rat and human OAT2 in the kidney may be related to species-specific creatinine handling [2].
Multiple splice variants of OAT2 have been identified. Sun et al. cloned 2 splice variants (546-aa and 538-aa) from a liver cDNA library, with variation in the C-terminal segment [15]. An 548-aa-long isoform, identified by Cropp et al., exhibited a complete loss of cGMP transport capability, and while the GFP-tagged hOAT2-546-aa localized to the plasma membrane, GFP-tagged hOAT2-548-aa remained trapped intracellularly [19]. A 539-aa splice variant was cloned from human kidney cDNA library using rat Oat2 cDNA as a probe [3]. 539-aa shares 97% sequence identity with other splice variants, the main difference affecting the C-terminal end [24].
Out of four single nucleotide polymorphisms (SNPs) identified by Xu et al., three were nonsynonymous (C329T, G571A, and G1520A), all heterozygous at their respective loci in this study. The synonymous SNP C1269T was found across all ethnic groups [25]. Shin et al. identified 6 intronic and 2 exonic SNPs, two of them (4207T>C, intronic, and T513T, coding) being novel at the time [26].
In HepaRG cells and human hepatocytes, OAT2 expression was repressed by all-trans retinoid acid, an effect which was abrogated by the simultaneous siRNA-mediated knock-down of RARα and RXRα in HepaRG cells [27]. The expression of multiple transporters including OAT2 was reduced in HepaRG cells when compared to hepatocytes, and in monolayer-cultured versus freshly isolated human hepatocytes [28, 29].

Clinical significance

OAT2 transports different types of clinically important drugs, and plays a role in their disposition. Therapeutic drugs may also act as inhibitors of OAT2-mediated transport. In a study conducted with a panel of antineoplastic drugs, alkylating agents (melphalan, bendamustine, chlorambucil, and busulfan), antimetabolites (methotrexate, gemcitabine, and fluoroadenine), the topoisomerase inhibitors irinotecan and etoposide, and the mitotic inhibitor paclitaxel interfered with OAT2-mediated cGMP transport, while bendamustine and irinotecan were also substrates of OAT2 [30]. Some authors were able to detect 5-fluorouracil (5-FU) transport, and this result indicated the contribution of OAT2 to the hepatic uptake of 5-FU, but others failed to observe inhibition of cGMP transport by 5-FU [17, 30]. High OAT2 expression was observed in alpha-fetoprotein-producing gastric cancer [8]. In metastatic colorectal cancer, the effectiveness of the commonly used FOLFOX (5-fluorouracil/leucovorin/oxaliplatin) and uracil/ftorafur plus leucovorin (UFT/LV) regimens is dependent on transporter-associated drug accumulation in tumor cells. High expression of OAT2 and OCT2 could predict effectiveness of FOLFOX-based therapy [9]. Leucovorin is transported by RFC1 (reduced folate carrier 1), and in patients with high expression levels of both OAT2 and RFC1, the two transporters synergized through increased uptake of both 5-FU and leucovorin to yield a better therapeutic outcome [10].
Diuretics are secreted in the proximal tubule cells, which is important for the drug action. It is mainly organic anions that are subject to this elimination route. The diuretic drug bumetanide was reported to be an OAT2 substrate in one study [17], while others only detected inhibition of ES uptake by bumetanide but not direct uptake of the drug itself [31]. The elimination pathway for antiviral drugs is renal excretion, with active tubular secretion. Some guanine-containing antiviral drugs like acyclovir, ganciclovir, and penciclovir are substrates of OAT2 [4], and since OAT2 is localized to the corneal epithelial layer it is thought to transport acyclovir to the anterior chamber of the eye [7]. An earlier report found the thymidine nucleoside analogue zidovudine, but not acyclovir and ganciclovir, to be transported by OAT2; however, ganciclovir inhibited OAT2-mediated PGF2α uptake [32]. OAT2 may also be required for the effective treatment of hepatitis B virus infection with entenavir, an important nucleoside analogue, since it mediates hepatic uptake of the drug [33].
OAT2 is implicated in the transport of antibiotics and nonsteroidal anti-inflammatory drugs (NSAID), too. OAT2 is a transporter of both erythromycin and theophylline, the latter being widely used in asthma and chronic pulmonary disease therapy. In case of co-administration, e.g. during an episode of pyogenic infection in a chronic asthmatic patient, OAT2 may at least in part account for the interaction of these drugs in the liver [34]. OAT2 mediates tetracycline uptake to proximal tubular cells, and various tetracyclines, but not doxycycline, inhibited OAT2-mediated PGF2α uptake [35]. NSAIDs are primarily regarded as inhibitors of OAT2; reports of OAT2-mediated NSAID uptake are sparse [36]. Diclofenac, as an exception, appears to be a specific OAT2 substrate, while its major metabolite diclofenac acyl-β-D-glucuronide is a common substrate of multiple OATs as well as other transporters [37].

Regulatory requirements

As no polymorphisms or drug interactions with clinical relevance have been reported yet, OAT2 is currently not recommended by FDA or EMA for in vitro testing.

Location Endogenous substrates In vitro substrates used experimentally Substrate drugs Inhibitors
liver, kidney, eye PAH, cGMP,cAMP, ES, orotic acid, DHEAS, prostaglandins uric acid, creatinine cGMP, uric acid bumetadine, bendamustine, irinotecan, 5-FU, acyclovir, ganciclovir, penciclovir, entenavir, tetracycline, diclofenac, zidovudine indomethacine, bromsulphthalein, probenecid, irinotecan, etoposide, paclitaxel

 

References

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2.    Shen, H., et al., Characterization of Organic Anion Transporter 2 (SLC22A7): A Highly Efficient Transporter for Creatinine and Species-Dependent Renal Tubular Expression. Drug Metab Dispos, 2015. 43(7): p. 984-93.
3.    Enomoto, A., et al., Interaction of human organic anion transporters 2 and 4 with organic anion transport inhibitors. J Pharmacol Exp Ther, 2002. 301(3): p. 797-802.
4.    Cheng, Y., et al., Expression of organic anion transporter 2 in the human kidney and its potential role in the tubular secretion of guanine-containing antiviral drugs. Drug Metab Dispos, 2012. 40(3): p. 617-24.
5.    Breljak, D., et al., Distribution of organic anion transporters NaDC3 and OAT1-3 along the human nephron. Am J Physiol Renal Physiol, 2016. 311(1): p. F227-38.
6.    Ikarashi, R., K. Shibasaki, and A. Yamaguchi, Immunohistochemical studies of organic anion transporters and urate transporter 1 expression in human salivary gland. Acta Odontol Scand, 2013. 71(2): p. 312-6.
7.    Dahlin, A., et al., Gene expression profiling of transporters in the solute carrier and ATP-binding cassette superfamilies in human eye substructures. Mol Pharm, 2013. 10(2): p. 650-63.
8.    Shimakata, T., et al., Immunohistochemical expression profiles of solute carrier transporters in alpha-fetoprotein-producing gastric cancer. Histopathology, 2016. 69(5): p. 812-821.
9.    Tashiro, A., et al., High expression of organic anion transporter 2 and organic cation transporter 2 is an independent predictor of good outcomes in patients with metastatic colorectal cancer treated with FOLFOX-based chemotherapy. Am J Cancer Res, 2014. 4(5): p. 528-36.
10.    Nishino, S., et al., Immunohistochemical analysis of organic anion transporter 2 and reduced folate carrier 1 in colorectal cancer: Significance as a predictor of response to oral uracil/ftorafur plus leucovorin chemotherapy. Mol Clin Oncol, 2013. 1(4): p. 661-667.
11.    Buist, S.C., et al., Gender-specific and developmental influences on the expression of rat organic anion transporters. J Pharmacol Exp Ther, 2002. 301(1): p. 145-51.
12.    Shen, H., Y. Lai, and A.D. Rodrigues, Organic Anion Transporter 2: An Enigmatic Human Solute Carrier. Drug Metab Dispos, 2017. 45(2): p. 228-236.
13.    Tanaka, K., et al., Role of glycosylation in the organic anion transporter OAT1. J Biol Chem, 2004. 279(15): p. 14961-6.
14.    Sekine, T., et al., Identification of multispecific organic anion transporter 2 expressed predominantly in the liver. FEBS Lett, 1998. 429(2): p. 179-82.
15.    Sun, W., et al., Isolation of a family of organic anion transporters from human liver and kidney. Biochem Biophys Res Commun, 2001. 283(2): p. 417-22.
16.    Henjakovic, M., et al., Human organic anion transporter 2 is distinct from organic anion transporters 1 and 3 with respect to transport function. Am J Physiol Renal Physiol, 2015. 309(10): p. F843-51.
17.    Kobayashi, Y., et al., Transport mechanism and substrate specificity of human organic anion transporter 2 (hOat2 [SLC22A7]). J Pharm Pharmacol, 2005. 57(5): p. 573-8.
18.    Fork, C., et al., OAT2 catalyses efflux of glutamate and uptake of orotic acid. Biochem J, 2011. 436(2): p. 305-12.
19.    Cropp, C.D., et al., Organic anion transporter 2 (SLC22A7) is a facilitative transporter of cGMP. Mol Pharmacol, 2008. 73(4): p. 1151-8.
20.    Sato, M., et al., Renal secretion of uric acid by organic anion transporter 2 (OAT2/SLC22A7) in human. Biol Pharm Bull, 2010. 33(3): p. 498-503.
21.    Kimura, H., et al., Human organic anion transporters and human organic cation transporters mediate renal transport of prostaglandins. J Pharmacol Exp Ther, 2002. 301(1): p. 293-8.
22.    Ciarimboli, G., et al., Proximal tubular secretion of creatinine by organic cation transporter OCT2 in cancer patients. Clin Cancer Res, 2012. 18(4): p. 1101-8.
23.    Lepist, E.I., et al., Contribution of the organic anion transporter OAT2 to the renal active tubular secretion of creatinine and mechanism for serum creatinine elevations caused by cobicistat. Kidney Int, 2014. 86(2): p. 350-7.
24.    Hotchkiss, A.G., L. Berrigan, and R.M. Pelis, Organic anion transporter 2 transcript variant 1 shows broad ligand selectivity when expressed in multiple cell lines. Front Pharmacol, 2015. 6: p. 216.
25.    Xu, G., et al., Analyses of coding region polymorphisms in apical and basolateral human organic anion transporter (OAT) genes [OAT1 (NKT), OAT2, OAT3, OAT4, URAT (RST)]. Kidney Int, 2005. 68(4): p. 1491-9.
26.    Shin, H.J., et al., Identification of genetic polymorphisms of human OAT1 and OAT2 genes and their relationship to hOAT2 expression in human liver. Clin Chim Acta, 2010. 411(1-2): p. 99-105.
27.    Le Vee, M., et al., Differential regulation of drug transporter expression by all-trans retinoic acid in hepatoma HepaRG cells and human hepatocytes. Eur J Pharm Sci, 2013. 48(4-5): p. 767-74.
28.    Le Vee, M., et al., Polarized expression of drug transporters in differentiated human hepatoma HepaRG cells. Toxicol In Vitro, 2013. 27(6): p. 1979-86.
29.    Le Vee, M., et al., Polarized location of SLC and ABC drug transporters in monolayer-cultured human hepatocytes. Toxicol In Vitro, 2015. 29(5): p. 938-46.
30.    Marada, V.V., et al., Interaction of human organic anion transporter 2 (OAT2) and sodium taurocholate cotransporting polypeptide (NTCP) with antineoplastic drugs. Pharmacol Res, 2015. 91: p. 78-87.
31.    Hasannejad, H., et al., Interactions of human organic anion transporters with diuretics. J Pharmacol Exp Ther, 2004. 308(3): p. 1021-9.
32.    Takeda, M., et al., Human organic anion transporters and human organic cation transporters mediate renal antiviral transport. J Pharmacol Exp Ther, 2002. 300(3): p. 918-24.
33.    Furihata, T., et al., Human organic anion transporter 2 is an entecavir, but not tenofovir, transporter. Drug Metab Pharmacokinet, 2017. 32(1): p. 116-119.
34.    Kobayashi, Y., et al., Possible involvement of organic anion transporter 2 on the interaction of theophylline with erythromycin in the human liver. Drug Metab Dispos, 2005. 33(5): p. 619-22.
35.    Babu, E., et al., Human organic anion transporters mediate the transport of tetracycline. Jpn J Pharmacol, 2002. 88(1): p. 69-76.
36.    Burckhardt, G., Drug transport by Organic Anion Transporters (OATs). Pharmacol Ther, 2012. 136(1): p. 106-30.
37.    Zhang, Y., et al., Diclofenac and Its Acyl Glucuronide: Determination of In Vivo Exposure in Human Subjects and Characterization as Human Drug Transporter Substrates In Vitro. Drug Metab Dispos, 2016. 44(3): p. 320-8.

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