Abstract：
The ergothioneine transporter ETT (formerly OCTN1; human gene symbol SLC22A4) is a powerful and highly specific transporter for the uptake of ergothioneine (ET). Recently, Sparreboom et al. reported that the ETT would transport nucleosides and nucleoside analogues such as cytarabine and gemcitabine with the highest efficiency. In our assay system, we could not detect any such transport. Subsequently, Sparreboom suggested that the intracellular metabolization of the nucleosides occurs so fast that the original compounds cannot be detected by LC-MS/MS after inward transport. Our current experiments with 293 cells disprove this hypothesis. Uptake of gemcitabine was easily detected by LC-MS/MS measurements when we expressed the Na+/nucleoside cotransporter CNT3 (SLC28A3). Inward transport was 1280 times faster than the intracellular production of gemcitabine triphosphate. The deoxycytidine kinase inhibitor 2-thio-2\'-deoxycytidine markedly blocked the production of gemcitabine triphosphate. There was no concomitant surge in intracellular gemcitabine, however. This does not fit the rapid phosphorylation of gemcitabine. Uptake of cytarabine was very slow, but detection by MS was still possible. When the ETT was expressed and incubated with gemcitabine, there was no increase in intracellular gemcitabine triphosphate. We conclude that the ETT does not transport nucleosides.
摘要：
麦角硫因转运蛋白ETT（以前称为OCTN1；人类基因符号SLC22A4）是摄取麦角硫因（ET）的强大且高度特异性的转运蛋白。最近,Spareboom等人。据报道，ETT将以最高的效率运输核苷和核苷类似物，例如阿糖胞苷和吉西他滨。在我们的化验系统中，我们无法检测到任何此类运输。随后，Sparreboom建议核苷的细胞内代谢发生得如此之快，以至于在向内转运后无法通过LC-MS/MS检测到原始化合物。我们目前对293细胞的实验证明了这一假设。当我们表达Na+/核苷共转运蛋白CNT3(SLC28A3)时,通过LC-MS/MS测量容易检测到吉西他滨的摄取。向内运输比吉西他滨三磷酸的细胞内生产快1280倍。脱氧胞苷激酶抑制剂2-硫代-2'-脱氧胞苷显著阻断吉西他滨三磷酸的产生。细胞内吉西他滨没有伴随的激增，however.这不适合吉西他滨的快速磷酸化。阿糖胞苷的摄取非常缓慢，但MS检测仍然是可能的。当ETT表达并与吉西他滨孵育时，细胞内三磷酸吉西他滨没有增加。我们得出结论，ETT不转运核苷。
