Azab Samar S, Salama Salama A, Abdel-Naim Ashraf B, Khalifa Amani E, El-Demerdash Ebtehal, Al-Hendy Ayman
Department of Obstetrics and Gynecology, University of Texas-Medical Branch, Galveston, TX, USA.
Breast Cancer Res Treat. 2009 Jan;113(1):9-19. doi: 10.1007/s10549-008-9898-3. Epub 2008 Jan 29.
2-Methoxyestradiol (2ME), a natural derivative of estradiol, is currently evaluated in clinical trials for breast cancer. The current study aims to evaluate the modulatory effects of 2ME on regulation of multidrug resistance (MDR) in doxorubicin (Dox) resistant breast cancer cells (MCF-7/Dox) and its underlying mechanisms. The chemosensitizing effect of 2ME on Dox cytotoxicity is tested by MTT assay. RT(2) Profiler PCR Array was used to identify differentially expressed genes in Dox and/or 2ME treatment groups, based on significance of results 4 genes were selected: MDR1, Bcl2, P53 and Cyclin D1. The expression of these genes was confirmed using western blotting. Lastly, functions of these genes were examined by studying p-glycoprotein (p-gp) function, caspase 3 activity and flowcytometric cell cycle assays respectively. 2ME significantly increased sensitivity of the resistant MCF-7/Dox cells to the cytotoxic effect of Dox by 2.9-folds. The array and western blotting showed that Bcl2 and Cyclin D1 expression were down regulated; P53 expression was not affected while MDR1 was over expressed by combination of 2ME with Dox. 2ME increased p-gp function by 24+/-7.05%, compared to control. Addition of 2ME to Dox increased caspase activity by 27-folds. Combination of 2ME to Dox arrested the cell cycle in G(1) and S phases, compared to Dox. In conclusion, 2ME chemosensitizes resistant breast cancer cells to Dox cytotoxicity by down regulating expression of Bcl2 and Cyclin D1, augmenting caspase 3 activity as well as inducing cell cycle block in G(1) and S phases.
2-甲氧基雌二醇（2ME）是雌二醇的一种天然衍生物，目前正在乳腺癌的临床试验中接受评估。本研究旨在评估2ME对多药耐药（MDR）的调节作用，该调节作用存在于对阿霉素（Dox）耐药的乳腺癌细胞（MCF-7/Dox）中及其潜在机制。通过MTT法检测2ME对Dox细胞毒性的化学增敏作用。基于结果的显著性，使用RT(2) Profiler PCR Array来鉴定Dox和/或2ME处理组中差异表达的基因，选择了4个基因：MDR1、Bcl2、P53和细胞周期蛋白D1。使用蛋白质印迹法确认这些基因的表达。最后，分别通过研究P-糖蛋白（p-gp）功能、半胱天冬酶3活性和流式细胞术细胞周期分析来检测这些基因的功能。2ME显著提高了耐药性MCF-7/Dox细胞对Dox细胞毒性的敏感性，提高了2.9倍。基因芯片和蛋白质印迹分析表明，Bcl2和细胞周期蛋白D1的表达下调；P53的表达不受影响，而2ME与Dox联合使用时MDR1过表达。与对照组相比，2ME使p-gp功能增加了24±7.05%。将2ME添加到Dox中可使半胱天冬酶活性增加27倍。与单独使用Dox相比，2ME与Dox联合使用使细胞周期停滞在G(1)期和S期。总之，2ME通过下调Bcl2和细胞周期蛋白D1的表达、增强半胱天冬酶3活性以及诱导细胞周期在G(1)期和S期阻滞，使耐药乳腺癌细胞对Dox细胞毒性产生化学增敏作用。
