Pharmacological analysis and molecular cloning of the canine equilibrative nucleoside transporter 1

We studied the binding of [H-3]nitrobenzylthioinosine (NBMPR) and the uptake [H-3]formycin B by the es (equilibrative inhibitor-sensitive) nucleoside transporter of Madin Darby Canine Kidney (MDCK) cells. NBMPR inhibited [H-3]formycin B uptake with a K-i of 2.7 +/- 0.6 nM, and [H-3]NBMPR had a K-D of 1.3 +/- 0.3 nM for binding to these cells; these values are significantly higher than those obtained in human and mouse cell models. In contrast, other recognized es inhibitors, such as dipyridamole, were significantly more effective as inhibitors of [H-3]NBMPR binding and [H-3]formycin B uptake by MDCK cells relative to that seen for human cells. We isolated a cDNA encoding the canine es nucleoside transporter (designated cENTI), and assessed its function by stable expression in nucleoside transport deficient PK15NTD cells. The PK15-cENTI cells displayed inhibitor sensitivities that were comparable to those obtained for the endogenous es nucleoside transporter in MDCK cells. These data indicate that the dog es/ENTI transporter has distinctive inhibitor binding characteristics, and that these characteristics are a function of the protein structure as opposed to the environment in which it is expressed. (C) 2004 Elsevier B.V. All rights reserved.