Involvement of CUL4A in Regulation of Multidrug Resistance to P-gp Substrate Drugs in Breast Cancer Cells

Yunshan Wang,Taomei Yang,Pengju Zhang,Yangyang Xu,Xiuquan He,Yuli Wang,Mingxin Wen,Guangwei Wei,Panpan Zhan,Qin Wang,Guangxin Ma

doi:10.3390/molecules19010159

Abstract

CUL4A encodes a core component of a cullin-based E3 ubiquitin ligase complex that regulates many critical processes such as cell cycle progression, DNA replication, DNA repair and chromatin remodeling by targeting a variety of proteins for ubiquitination and degradation. In the research described in this report we aimed to clarify whether CUL4A participates in multiple drug resistance (MDR) in breast cancer cells. We first transfected vectors carrying CUL4A and specific shCUL4A into breast cancer cells and corresponding Adr cells respectively. Using reverse transcription polymerase chain reactions and western blots, we found that overexpression of CUL4A in MCF7 and MDA-MB-468 cells up-regulated MDR1/P-gp expression on both the transcription and protein levels, which conferred multidrug resistance to P-gp substrate drugs, as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. On the other hand, silencing CUL4A in MCF7/Adr and MDA-MB-468/Adr cells led to the opposite effect. Moreover, ERK1/2 in CUL4A-overexpressing cells was highly activated and after treatment with PD98059, an ERK1/2-specific inhibitor, CUL4A-induced expression of MDR1/P-gp was decreased significantly. Lastly, immunohistochemistry in breast cancer tissues showed that P-gp expression had a positive correlation with the expression of CUL4A and ERK1/2. Thus, these results implied that CUL4A and ERK1/2 participated in multi-drug resistance in breast cancer through regulation of MDR1/P-gp expression.

Highlights

Breast cancer is the most frequently diagnosed cancer in woman and one of the leading causes of cancer death worldwide [1,2]
Our results suggest that overexpression of CUL4A in breast cancer is responsible for the required resistant to P-gp substrate drugs through regulation of multidrug resistance 1 gene (MDR1)/P-gp expression
To establish cell lines with ectopic or silencing CUL4A expression, MCF7 and MDA-MB-468 and their Adr cells were stably transfected with retroviruses expressing pBabe-CUL4A and pSuper-shCUL4A

Summary

Introduction

Breast cancer is the most frequently diagnosed cancer in woman and one of the leading causes of cancer death worldwide [1,2]. Multiple drug resistance (MDR) plays a major role in the failure of breast cancer therapy [3,4,5]. The development of MDR, in which tumor cells become resistant to a wide spectrum of anti-cancer agents with different structures or different target sites, severely limits the success of chemotherapy in breast cancer [3,4,6,7]. Overexpression of MDR1/P-gp results in an active efflux of anticancer agents from cells, lowering intracellular drug concentrations and inducing cancer cells to resist to chemotherapeutic drugs, especially P-gp substrate anti-cancer drugs, such as doxorubicin and paclitaxel [8,9,10,11]. Several strategies have been developed to restore chemotherapeutic sensitivity in MDR cells with limited success [7,12]

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Results

Conclusion