title: Vectorial transport of nucleoside analogs from the apical to the basolateral membrane in double-transfected cells expressing the human concentrative nucleoside transporter hCNT3 and the export pump ABCC4.
abstract: The identification of the transport proteins responsible for the uptake and the efflux of nucleosides and their metabolites enables the characterization of their vectorial transport and a better understanding of their absorption, distribution, and elimination. Human concentrative nucleoside transporters (hCNTs/SLC28A) are known to mediate the transport of natural nucleosides and some nucleoside analogs into cells in a sodium-dependent and unidirectional manner. On the other hand, several human multidrug resistance proteins [human ATP-binding cassette transporter, subfamily C (ABCC)] cause resistance against nucleoside analogs and mediate transport of phosphorylated nucleoside derivatives out of the cells in an ATP-dependent manner. For the integrated analysis of uptake and efflux of these compounds, we established a double-transfected Madin-Darby canine kidney (MDCK) II cell line stably expressing the human uptake transporter hCNT3 in the apical membrane and the human efflux pump ABCC4 in the basolateral membrane. The direction of transport was from the apical to the basolateral compartment, which is in line with the unidirectional transport and the localization of both recombinant proteins in the MDCKII cells. Recombinant hCNT3 mediated the transport of several known nucleoside substrates, and we identified 5-azacytidine as a new substrate for hCNT3. It is of interest that coexpression of both transporters was confirmed in pancreatic adenocarcinomas, which represent an important clinical indication for the therapeutic use of nucleoside analogs. Thus, our results establish a novel cell system for studies on the vectorial transport of nucleosides and their analogs from the apical to the basolateral compartment. The results contribute to a better understanding of the cellular transport characteristics of nucleoside drugs.
authors.affiliation: Division of Tumor Biochemistry, German Cancer Research Center, D-69120 Heidelberg, Germany. m.rius@dkfz.de
authors.forename: Maria
authors.initials: M
authors.lastname: Rius
authors.forename: Daniela
authors.initials: D
authors.lastname: Keller
authors.forename: Manuela
authors.initials: M
authors.lastname: Brom
authors.forename: Johanna
authors.initials: J
authors.lastname: Hummel-Eisenbeiss
authors.forename: Frank
authors.initials: F
authors.lastname: Lyko
authors.forename: Dietrich
authors.initials: D
authors.lastname: Keppler
mesh_terms: D000818:Animals; D001374:Azacitidine / Q000378:metabolism; D001692:Biological Transport / Q000235:genetics*; D002461:Cell Line, Transformed; D004285:Dogs; D006801:Humans; D026901:Membrane Transport Proteins / Q000096:biosynthesis* / Q000235:genetics; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000096:biosynthesis* / Q000235:genetics; D009705:Nucleosides / Q000378:metabolism*; D010179:Pancreas / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection
chemical_list: D001374:Azacitidine; D026901:Membrane Transport Proteins; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009705:Nucleosides; C073492:ABCC4 protein, human; C065171:cif nucleoside transporter