Versantvoort C H, Broxterman H J, Lankelma J, Feller N, Pinedo H M
Department of Medical Oncology, Free University Hospital, Amsterdam, The Netherlands.
Biochem Pharmacol. 1994 Sep 15;48(6):1129-36. doi: 10.1016/0006-2952(94)90149-x.
In several multidrug resistant tumor cell lines without overexpression of P-glycoprotein (non-Pgp MDR), a decreased accumulation of drugs has been shown to contribute to resistance. We have recently reported that daunorubicin (DNR) accumulation was decreased in the multidrug resistance-associated protein overexpressing GLC4/ADR non-Pgp MDR small cell lung cancer cell line due to an enhanced energy-dependent efflux which could be inhibited by the isoflavonoid genistein. The purpose of this work was 2-fold: (i) to investigate the mechanism by which genistein inhibits the DNR efflux in the GLC4/ADR cells; and (ii) to characterize the dependence of DNR transport on ATP concentration in intact GLC4/ADR cells. The active transport of DNR in GLC4/ADR cells appeared to be a saturable process with an apparent Km of DNR of 1.4 +/- 0.4 microM. Genistein increased the apparent Km value of DNR, suggesting that this agent is a competitive inhibitor of DNR transport. These data provide additional evidence that energy-dependent DNR transport in GLC4/ADR cells is a protein-mediated process. In addition, genistein decreased cellular ATP concentration in a dose-dependent manner in sensitive as well as in resistant cells. Marked inhibition of DNR transport activity in intact GLC4/ADR cells was found when cellular ATP concentration was decreased below 2 mM by sodium azide or 2-deoxy-D-glucose. Thus, since DNR transport in intact GLC4/ADR is already inhibited at modest cellular ATP depletion, a limitation in ATP supply might open ways to make MDR cells more susceptible to drug toxicity.
在几种未过度表达P-糖蛋白的多药耐药肿瘤细胞系（非Pgp MDR）中，药物蓄积减少被证明与耐药性有关。我们最近报道，在多药耐药相关蛋白过表达的GLC4/ADR非Pgp MDR小细胞肺癌细胞系中，柔红霉素（DNR）蓄积减少是由于能量依赖性外排增强，而异黄酮染料木黄酮可抑制这种外排。这项工作的目的有两个：（i）研究染料木黄酮抑制GLC4/ADR细胞中DNR外排的机制；（ii）表征完整GLC4/ADR细胞中DNR转运对ATP浓度的依赖性。DNR在GLC4/ADR细胞中的主动转运似乎是一个可饱和过程，DNR的表观Km为1.4±0.4μM。染料木黄酮增加了DNR的表观Km值，表明该药物是DNR转运的竞争性抑制剂。这些数据提供了额外的证据，证明GLC4/ADR细胞中能量依赖性DNR转运是一个蛋白质介导的过程。此外，染料木黄酮在敏感细胞和耐药细胞中均以剂量依赖性方式降低细胞ATP浓度。当用叠氮化钠或2-脱氧-D-葡萄糖使细胞ATP浓度降至2 mM以下时，发现完整GLC4/ADR细胞中的DNR转运活性受到显著抑制。因此，由于在适度的细胞ATP消耗时完整GLC4/ADR中的DNR转运就已被抑制，ATP供应的限制可能为使MDR细胞对药物毒性更敏感开辟途径。
