Li Guangliang, Zhang Jing, Xu Zhenzhen, Li Zhongqi
Institute of Cancer Research and Basic Medicine (ICBM), Chinese Academy of Sciences, Department of Medical Oncology (Breast), Cancer Hospital of University of Chinese Academy of Sciences, Zhejiang Cancer Hospital, Hangzhou, Zhejiang, 310022, People's Republic of China.
Department of Surgical Oncology, The 1st Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang 310003, People's Republic of China.
Cancer Manag Res. 2020 Jan 14;12:265-275. doi: 10.2147/CMAR.S226410. eCollection 2020.
Acquired tamoxifen resistance is one of the major barriers to the successful treatment of breast cancer. Recently, overexpression of ERα36 was demonstrated to be a potential mechanism for the generation of acquired tamoxifen resistance. This study aims to evaluate the possibility of ERα36 being a therapeutic target for tamoxifen-resistant breast cancer.
A tamoxifen-resistant cell subline (MCF-7/TAM) was established by culturing MCF-7 cells in medium plus 1 μM tamoxifen over 6 months. Colony-forming assay was used to determine the sensitivity of MCF-7/TAM cells to tamoxifen. Stable transfection was used to knockdown ERα36 expression in MCF-7/TAM cells. MTT assay and Transwell migration assay were used to assess the in vitro proliferation and migration, respectively. Nude mouse tumorigenicity assay was used to evaluate in vivo tumorigenicity. Western blot analysis and quantitative real-time PCR (qRT-PCR) were used to examine the expression of ERα36, ERα, EGFR and phosphorylated ERK1/2. The dual-luciferase reporter assay was used to determine the effect of ERα36 on the activity of -promotor.
MCF-7/TAM cells possessed greatly increased ERα36 expression and EGFR expression and exhibited significantly increased in vitro proliferation and migration ability, as well as increased in vivo tumor growth ability, compared to parental MCF-7 cells. Knockdown of ERα36 expression inhibited in vitro proliferation and migration, as well as in vivo tumor growth ability of MCF-7/TAM cells. ERα36 regulated EGFR expression at the transcriptional level, and knockdown of ERα36 in MCF-7/TAM cells downregulated EGFR expression and then blocked EGFR/ERK signaling pathway.
Knockdown of ERα36 inhibits the growth of MCF-7/TAM cells in vitro and in vivo by blocking EGFR/ERK signaling pathway. ERα36 may be a candidate therapeutic target for the treatment of tamoxifen-resistant breast cancer.
获得性他莫昔芬耐药是乳腺癌成功治疗的主要障碍之一。最近，有研究表明雌激素受体α36（ERα36）的过表达是产生获得性他莫昔芬耐药的一种潜在机制。本研究旨在评估ERα36作为他莫昔芬耐药乳腺癌治疗靶点的可能性。
通过在含有1μM他莫昔芬的培养基中培养MCF-7细胞6个月以上，建立他莫昔芬耐药细胞亚系（MCF-7/TAM）。采用集落形成试验测定MCF-7/TAM细胞对他莫昔芬的敏感性。通过稳定转染敲低MCF-7/TAM细胞中ERα36的表达。分别采用MTT试验和Transwell迁移试验评估体外增殖和迁移能力。采用裸鼠成瘤试验评估体内成瘤性。采用蛋白质免疫印迹分析和定量实时聚合酶链反应（qRT-PCR）检测ERα36、雌激素受体α（ERα）、表皮生长因子受体（EGFR）和磷酸化细胞外信号调节激酶1/2（ERK1/2）的表达。采用双荧光素酶报告基因试验确定ERα36对启动子活性的影响。
与亲本MCF-7细胞相比，MCF-7/TAM细胞的ERα36表达和EGFR表达显著增加，体外增殖和迁移能力以及体内肿瘤生长能力显著增强。敲低ERα36表达可抑制MCF-7/TAM细胞的体外增殖和迁移以及体内肿瘤生长能力。ERα36在转录水平调节EGFR表达，敲低MCF-7/TAM细胞中的ERα36可下调EGFR表达，进而阻断EGFR/ERK信号通路。
敲低ERα36通过阻断EGFR/ERK信号通路抑制MCF-7/TAM细胞的体外和体内生长。ERα36可能是治疗他莫昔芬耐药乳腺癌的候选治疗靶点。
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