Role of organic cation transporters in the renal secretion of nucleosides

doi:10.1016/S0006-2952(00)00334-8

Biochemical Pharmacology

Volume 60, Issue 2, 15 July 2000, Pages 215-219

https://doi.org/10.1016/S0006-2952(00)00334-8 Get rights and content

Abstract

The mammalian kidney eliminates toxic substances from the body, in part via secretion by the organic cation transporters (OCT) or organic anion transporters. Nucleosides are nitrogenous heterocycles that are often secreted by human and other animal kidneys. Previous experiments have shown that 2′-deoxytubercidin (7-deazadeoxyadenosine, dTub) is secreted by the mouse kidney via a cimetidine-sensitive OCT (Nelson et al., Biochem Pharmacol 32: 2323–2327, 1983). Experiments reported herein demonstrated that the cloned rat kidney rOCT1 transports dTub, cytosine arabinoside, 2-chlorodeoxyadenosine, and azidothymidine when expressed in the Xenopus laevis oocyte translation system. Although rOCT2 is 67% identical with rOCT1 in its amino acid sequence, rOCT2 does not mediate the uptake of these nucleosides. Uptake of dTub mediated by rOCT1 was pH-dependent in a manner suggesting that the positive charged moiety of dTub may be the true substrate. Protons acted as competitive inhibitors for the rOCT1-mediated uptake of dTub or tetraethylammonium (TEA), with K_i values corresponding to a pH of about 6.1. TEA and dTub mutually inhibited the uptake of one another by rOCT1, competitively, with K_i values approximately the same as their respective K_m values. These findings suggest that protons, dTub, and TEA act at a common site on rOCT1, and that rOCT1 participates in the renal secretion of dTub and other nucleosides.

Section snippets

Materials

[¹⁴C]TEA (2.4 or 3.36 mCi/mmol) was purchased from DuPont NEN. [³H]dTub (9.3 Ci/mmol) was prepared by Moravek Biochemicals Inc. using dTub synthesized in our laboratory as previously described [8]. [³H]araC (23.3 Ci/mmol), [³H]araG (2.5 Ci/mmol), [³H]AZT (33 Ci/mmol), and [³H]CldAdo (6.5 Ci/mmol) were purchased from Moravek Biochemicals, Inc. Other chemicals were obtained from the Sigma Chemical Co.

X. laevis oocyte injection and transport measurements

Cloned rOCT1 [6] and rOCT2 [7] cDNAs contained in the pSPORT vector (Life Technologies, Inc.)

Transport of radioactively labeled dTub and TEA by rOCT1 and rOCT2 in X. laevis oocytes

TEA is a classical substrate for putative and identified OCTs, i.e. it has served as a model for organic cation secretion in various species for many years. As shown in Fig. 1 B, the uptake of [¹⁴C]TEA into oocytes previously injected with the cRNAs from rOCT1 and rOCT2 was enhanced, similar to the reports of others 6, 7. In contrast to the enhanced uptake of TEA by both OCTs, the uptake of dTub was increased by rOCT1 but not by rOCT2 (Fig. 1A). The enhanced uptake of dTub by rOCT1 and of TEA

Discussion

The apparent secretion of dAdo and reabsorption of Ado were first observed in a child lacking adenosine deaminase and in cancer patients treated with the deaminase inhibitor deoxycoformycin [1]. Secretion of dAdo was confirmed in mice, and the analogue dTub was used subsequently to determine the potential mechanism(s) for the secretion [4]. Although there are at least five known nucleoside transporters 14, 15, renal secretion of dTub was proven to occur by a Cim-sensitive OCT using classical

Acknowledgements

We thank Dr. William P. Dubinsky, Dr. V. Ganapathy, and Dr. John B. Pritchard for their critical review of the manuscript and provision of the materials. Ms. Bih-fang Pan and Ms. Rosalind Alexander provided excellent assistance. This work was supported by National Institutes of Health Grants DK41606, Institute for Digestive Diseases and Kidney, and 2-P30-CA16672, a National Cancer Center Core Grant.

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Molecular and cellular pharmacologyRole of organic cation transporters in the renal secretion of nucleosides

Abstract

Section snippets

Materials

X. laevis oocyte injection and transport measurements

Transport of radioactively labeled dTub and TEA by rOCT1 and rOCT2 in X. laevis oocytes

Discussion

Acknowledgements

Biochem Pharmacol

Biochem Pharmacol

Biochem Biophys Res Commun

Trends Biochem Sci

J Biol Chem

Renal handling of 2′-deoxyadenosine and adenosine in humans and mice

Cancer Chemother Pharmacol

Pharmacokinetics of cladribine (2-chlorodeoxyadenosine) in children with acute leukemia

Cancer Res

Pharmacokinetics of zidovudine (AZT) and its metabolite (G-AZT) in healthy subjects and in patients with kidney failure

Therapie

Drug excretion mediated by a new prototype of polyspecific transporter

Nature

Nucleic acid related compounds. 42. A general procedure for the efficient deoxygenation of secondary alcohols. Regiospecific and stereoselective conversion of ribonucleosides to 2′-deoxynucleosides

J Am Chem Soc

Molecular and cellular pharmacology
Role of organic cation transporters in the renal secretion of nucleosides