屈洛昔芬逆转人乳腺癌耐药细胞株MCF7/ADR多药耐药性的研究

Reversal of Droloxifene on Multidrug Resistance in Drug-resistant Human Breast Cancer Cell Line MCF7/ADR

目的:研究屈洛昔芬(DRO)对乳腺癌耐药细胞株(MCF7/ADR)多药耐药性(MDR)的影响。方法:应用MTT、半定量RT-PCR和免疫细胞化学染色,研究DRO对MCF7/ADR药物敏感性、耐药相关基因MDR1、谷胱甘肽S转移酶Pi(GST-π)、拓扑异构酶Ⅱα(TopoⅡα)和多药耐药相关蛋白(MRP)表达的影响。流式细胞技术观察细胞内药物积累。结果:在20、10和5μmol/L浓度时DRO对MCF7/ADR耐药性的逆转倍数分别为16、8和3倍,逆转强度与同类化合物三苯氧胺相当。DRO处理后,MDR1和GSTπ基因表达水平明显降低。DRO使ADR在细胞内的积累分别增加到2.5(20μmol/L)、2.0(10μmol/L)和1.5(5μmol/L)倍。结论:DRO具有较强的逆转MCF7/ADR耐药性的作用,其逆转机制是多途径的。

Objective To study the reversal effects of droloxifene (DRO) on multidrug resistance (MDR) in MCF7/ADR and its mechanism. Methods Using tetrazolium dye (MTT) assay, effects of various concentrations of DRO on MDR in MCF7/ADR were studied. Expression of MDR related genes MDR1, glutathoine S-transferase Pi (GST-π), Topoisomerase Ⅱα (Topo Ⅱα) and MDR related protein (MRP) were assayed by reverse transcription-polymerase chain reaction ( RT-PCR) , immunocytochemistry assay. Using flow cytometry (FCM) , intracellular ADR concentration were observed. Results DRO significantly reversed MDR in MCF7/ ADR. After 20,10 and 5 μmol/L of DRO treatment,the chemosensitivity to ADR increased to 16,8 and 3 times respectively.The reversal activity of DRO was comparable to similar compound tamaxifen (TAM). After 10 μmol/L DRO treatment, both MDR1 and GST-π mRNA genes expression significantly declined on the third day ( both P < 0.01). The expression of MRP and Topo Ⅱα gene had no significant changes. Changes of P-gp and GST-π protein expressions were similar to those of their mRNA expressions.Two hours after 20,10 and 5 μmol/ L DRO treatment, intracellular ADR concentration increased to 2.5,2.0 and 1.5 times respectively. Conclu-SionS DRO has significant reversal effects on MDR in MCF7/ADR. The reversal is via different routes, i. e. , down regulating mRNA and protein expression levels of MDR1 and GST-π and increasing intracellular drug concentration.The reversal mechanism needs to be further studied.The low toxicity may provide prospects for DRO in clinical application.