Division of Tumor Biochemistry, German Cancer Research Center, D-69120 Heidelberg, Germany.
Drug Metab Dispos. 2010 Jul;38(7):1054-63. doi: 10.1124/dmd.110.032664. Epub 2010 Apr 1.
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.
鉴定负责摄取和外排核苷及其代谢物的转运蛋白，能够使它们的载体转运特性得到表征，从而更好地理解它们的吸收、分布和消除。已知人类浓缩核苷转运蛋白（hCNTs/SLC28A）以钠离子依赖和单向的方式介导天然核苷和一些核苷类似物进入细胞的转运。另一方面，几种人类多药耐药蛋白[人 ATP 结合盒转运蛋白，亚家族 C（ABCC）]对核苷类似物产生耐药性，并以 ATP 依赖的方式将磷酸化核苷衍生物转运出细胞。为了综合分析这些化合物的摄取和外排，我们建立了一个稳定表达人摄取转运蛋白 hCNT3 的双转染 Madin-Darby 犬肾 (MDCK) II 细胞系，hCNT3 位于细胞顶膜，人外排泵 ABCC4 位于基底外侧膜。转运方向是从顶侧向基底外侧，这与两种重组蛋白在 MDCKII 细胞中的单向转运和定位一致。重组 hCNT3 介导了几种已知核苷底物的转运，我们发现 5-氮杂胞苷是 hCNT3 的一个新底物。有趣的是，两种转运蛋白的共表达在胰腺腺癌中得到了证实，这是核苷类似物治疗应用的一个重要临床指征。因此，我们的结果建立了一个新的细胞系统，用于研究核苷及其类似物从顶侧向基底外侧的载体转运。这些结果有助于更好地理解核苷药物的细胞转运特性。
