Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon, Korea.
Br J Pharmacol. 2011 Mar;162(5):1096-108. doi: 10.1111/j.1476-5381.2010.01101.x.
The expression of P-glycoprotein (P-gp), encoded by the multidrug resistance 1 (MDR1) gene, is associated with the emergence of the MDR phenotype in cancer cells. We investigated whether metformin (1,1-dimethylbiguanide hydrochloride) down-regulates MDR1 expression in MCF-7/adriamycin (MCF-7/adr) cells.
MCF-7 and MCF-7/adr cells were incubated with metformin and changes in P-gp expression were determined at the mRNA, protein and functional level. Transient transfection assays were performed to assess its gene promoter activities, and immunoblot analysis to study its molecular mechanisms of action.
Metformin significantly inhibited MDR1 expression by blocking MDR1 gene transcription. Metformin also significantly increased the intracellular accumulation of the fluorescent P-gp substrate rhodamine-123. Nuclear factor-κB (NF-κB) activity and the level of IκB degradation were reduced by metformin treatment. Moreover, transduction of MCF-7/adr cells with the p65 subunit of NF-κB induced MDR1 promoter activity and expression, and this effect was attenuated by metformin. The suppression of MDR1 promoter activity and protein expression was mediated through metformin-induced activation of AMP-activated protein kinase (AMPK). Small interfering RNA methods confirmed that reduction of AMPK levels attenuates the inhibition of MDR1 activation associated with metformin exposure. Furthermore, the inhibitory effects of metformin on MDR1 expression and cAMP-responsive element binding protein (CREB) phosphorylation were reversed by overexpression of a dominant-negative mutant of AMPK.
These results suggest that metformin activates AMPK and suppresses MDR1 expression in MCF-7/adr cells by inhibiting the activation of NF-κB and CREB. This study reveals a novel function of metformin as an anticancer agent.
多药耐药基因 1（MDR1）编码的 P-糖蛋白（P-gp）的表达与癌细胞中多药耐药表型的出现有关。我们研究了二甲双胍（1,1-二甲基双胍盐酸盐）是否下调 MCF-7/阿霉素（MCF-7/adr）细胞中的 MDR1 表达。
用二甲双胍孵育 MCF-7 和 MCF-7/adr 细胞，并在 mRNA、蛋白和功能水平上测定 P-gp 表达的变化。进行瞬时转染实验以评估其基因启动子活性，并进行免疫印迹分析以研究其作用的分子机制。
二甲双胍通过阻断 MDR1 基因转录显著抑制 MDR1 表达。二甲双胍还显著增加了荧光 P-糖蛋白底物罗丹明-123 的细胞内积累。二甲双胍处理降低了核因子-κB（NF-κB）活性和 IκB 降解水平。此外，转染 MCF-7/adr 细胞的 NF-κB p65 亚基诱导 MDR1 启动子活性和表达，而这种作用被二甲双胍减弱。MDR1 启动子活性和蛋白表达的抑制是通过二甲双胍诱导的 AMP 激活蛋白激酶（AMPK）激活介导的。小干扰 RNA 方法证实，降低 AMPK 水平会减弱与二甲双胍暴露相关的 MDR1 激活抑制。此外，过表达 AMPK 的显性失活突变可逆转二甲双胍对 MDR1 表达和 cAMP 反应元件结合蛋白（CREB）磷酸化的抑制作用。
这些结果表明，二甲双胍通过抑制 NF-κB 和 CREB 的激活来激活 AMPK 并抑制 MCF-7/adr 细胞中的 MDR1 表达。本研究揭示了二甲双胍作为抗癌药物的新功能。
