Renes J, de Vries E G, Nienhuis E F, Jansen P L, Müller M
Groningen Institute for Drug Studies, Division of Gastroenterology and Hepatology, University Hospital Groningen, The Netherlands.
Br J Pharmacol. 1999 Feb;126(3):681-8. doi: 10.1038/sj.bjp.0702360.
The present study was performed to investigate the ability of the multidrug resistance protein (MRPI) to transport different cationic substrates in comparison with MDR1-P-glycoprotein (MDR1). Transport studies were performed with isolated membrane vesicles from in vitro selected multidrug resistant cell lines overexpressing MDR1 (A2780AD) or MRP1 (GLC4/Adr) and a MRP1-transfected cell line (S1(MRP)). As substrates we used 3H-labelled derivatives of the hydrophilic monoquaternary cation N-(4',4'-azo-in-pentyl)-21-deoxy-ajmalinium (APDA), the basic drug vincristine and the more hydrophobic basic drug daunorubicin. All three are known MDR1-substrates. MRP1 did not mediate transport of these substrates per se. In the presence of reduced glutathione (GSH), there was an ATP-dependent uptake of vincristine and daunorubicin, but not of APDA, into GLC4/Adr and S1(MRP) membrane vesicles which could be inhibited by the MRP1-inhibitor MK571. ATP- and GSH-dependent transport of daunorubicin and vincristine into GLC4/Adr membrane vesicles was inhibited by the MRP1-specific monoclonal antibody QCRL-3. MRP1-mediated daunorubicin transport rates were dependent on the concentration of GSH and were maximal at concentrations > or = 10 mM. The apparent KM value for GSH was 2.7 mM. Transport of daunorubicin in the presence of 10 mM GSH was inhibited by MK571 with an IC50 of 0.4 microM. In conclusion, these results demonstrate that MRP1 transports vincristine and daunorubicin in an ATP- and GSH-dependent manner. APDA is not a substrate for MRP1.
本研究旨在比较多药耐药蛋白（MRPI）与多药耐药1 - P - 糖蛋白（MDR1）转运不同阳离子底物的能力。使用从体外筛选出的过表达MDR1（A2780AD）或MRP1（GLC4/Adr）的多药耐药细胞系以及MRP1转染细胞系（S1(MRP)）分离得到的膜囊泡进行转运研究。我们将亲水性单季铵阳离子N - (4',4'-偶氮-戊基)-21 - 脱氧-阿马林（APDA）的3H标记衍生物、碱性药物长春新碱以及疏水性更强的碱性药物柔红霉素作为底物。这三种都是已知的MDR1底物。MRP1本身并不介导这些底物的转运。在还原型谷胱甘肽（GSH）存在的情况下，GLC4/Adr和S1(MRP)膜囊泡中存在ATP依赖的长春新碱和柔红霉素摄取，但APDA没有，这种摄取可被MRP1抑制剂MK571抑制。柔红霉素和长春新碱向GLC4/Adr膜囊泡的ATP和GSH依赖转运被MRP1特异性单克隆抗体QCRL - 3抑制。MRP1介导的柔红霉素转运速率取决于GSH浓度，在浓度≥10 mM时达到最大值。GSH的表观KM值为2.7 mM。在10 mM GSH存在下，柔红霉素的转运被MK571抑制，IC50为0.4 μM。总之，这些结果表明MRP1以ATP和GSH依赖的方式转运长春新碱和柔红霉素。APDA不是MRP1的底物。
