caa a22 4500 477127231 CHVBK 20180405111653.0 cr unu---uuuuu 170330e19971001xx s 000 0 eng 10.1023/A:1012148016794 doi (NATIONALLICENCE)springer-10.1023/A:1012148016794 Transient Expression of a Purine-Selective Nucleoside Transporter (SPNTint) in a Human Cell Line (HeLa) [Elektronische Daten] [Marci Schaner, Juan Wang, Shoshana Zevin, Karin Gerstin, Kathleen Giacomini] Purpose. The goal of this study was to develop a mammalian expression system for the cloned rat intestinal, Na+-dependent, purine-selective nucleoside transporter (SPNTint) and to study the interactions of nucleosides and nucleoside analogs with this transporter. Methods. Lipofection was used to transfect HeLa cells with a mammalian expression vector (pcDNA3) containing the cDNA insert encoding SPNTint. Nucleoside transport activity was measured using [3H] inosine, [3H]uridine, [3H]-dideoxyinosine (ddl), and [3H]-2-chloro-2′-deoxyadenosine (2CdA) as model substrates. Results. Expression of SPNTint was observed between 36 and 90 h post-transfection, with maximal expression at 66 h. At 66 h, Na+-stimulated uptake of [3H]inosine in cells transiently transfected with SPNTint was approximately threefold greater than that in cells transfected with empty vector (p < 0.05). The Na+-stimulated uptake of both inosine and uridine was saturable (Km = 28.1 ± 7.1 μM and 20.6 ± 5.6 μM, respectively) in the transfected cells and was significantly inhibited by the naturally occurring nucleosides (1 mM) inosine and uridine and to a lesser extent by thymidine. The nucleoside analogs ddl (IC50 = 46 μM) and 2CdA (IC50 =.13 μM) also significantly inhibited the Na+-stimulated uptake of [3H]inosine. A Na+-stimulated uptake of [3H]2CdA was observed suggesting that 2CdA is also a permeant of SPNTint. Conclusions. HeLa cells transiently transfected with SPNTint represent a useful tool to study the kinetics and interactions of drugs with SPNTint. Plenum Publishing Corporation, 1997 nucleosides nationallicence transporters nationallicence HeLa nationallicence mammalian expression systems nationallicence nucleoside analogs nationallicence Schaner Marci Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut Wang Juan Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut Zevin Shoshana Division of Clinical Pharmacology and Experimental Therapeutics, University of California San Francisco, 94143, San Francisco, California aut Gerstin Karin Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut Giacomini Kathleen Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut Pharmaceutical Research Kluwer Academic Publishers-Plenum Publishers 14/10(1997-10-01), 1316-1321 0724-8741 14:10<1316 1997 14 11095 https://doi.org/10.1023/A:1012148016794 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1023/A:1012148016794 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Schaner Marci Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut NATIONALLICENCE 700 1- Wang Juan Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut NATIONALLICENCE 700 1- Zevin Shoshana Division of Clinical Pharmacology and Experimental Therapeutics, University of California San Francisco, 94143, San Francisco, California aut NATIONALLICENCE 700 1- Gerstin Karin Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut NATIONALLICENCE 700 1- Giacomini Kathleen Department of Biopharmaceutical Sciences, University of California San Francisco, Box 0446, 94143, San Francisco, California aut NATIONALLICENCE 773 0- Pharmaceutical Research Kluwer Academic Publishers-Plenum Publishers 14/10(1997-10-01), 1316-1321 0724-8741 14:10<1316 1997 14 11095 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer