Department of Biochemistry and Molecular Biology, Third Military Medical University, Chongqing, 400038, China.
Department of Clinical Laboratory, Institute of Surgery Research, Daping Hospital, Third Military Medical University, Chongqing, 400038, China.
J Exp Clin Cancer Res. 2018 Jun 25;37(1):123. doi: 10.1186/s13046-018-0798-z.
ER-α36, a novel ER-α66 variant, has been demonstrated to promote tamoxifen resistance in breast cancer cells. However, the role and mechanisms of ER-α36 in cisplatin resistance of breast cancer cells remain unclear. This study investigates the expression and role of ER-α36 in cisplatin resistance of breast cancer cells and elucidates its underlying mechanisms.
The expression of ER-α36 and the proteins involved in nongenomic estrogen signaling was evaluated by western blot analysis. Cisplatin sensitivity was explored by CCK-8 assay, monolayer colony formation assay and apoptosis assays, respectively. ER-α36 siRNAs/shRNAs and overexpression vector were transfected into cells to down-regulate or up-regulate ER-α36 expression. Loss-and gain-of function assays were performed to investigate the role of ER-α36 in cisplatin sensitivity. The interaction between ER-α36 and EGFR/HER-2 were detected using CoIP. A mouse xenograft model of breast cancer was established to verify the role of ER-α36 in vivo.
ER-α36 is expressed at higher levels in cisplatin-resistant breast cancer cells compared to cisplatin sensitive cells. Cisplatin induced up-regulation of ER-α36 in a dose-dependent manner in breast cancer cells. Overexpression of ER-α36 leaded to cell resistant to cisplatin and knockdown of ER-α36 in cisplatin-resistant breast cancer cells restored cisplatin sensitivity. The up-regulation of ER-α36 resulted in increased activation of nongenomic estrogen signaling, which was responsible for cisplatin resistance. Disruption of ER-α36-mediated nongenomic estrogen signaling with kinase inhibitors significantly inhibited cisplatin-induced expression of ER-α36 and increased cisplatin sensitivity. The in vivo experiment also confirmed that up-regulation of ER-α36 attenuated cisplatin sensitivity in a mouse xenograft model of breast cancer.
The results for the first time demonstrated that ER-α36 mediates cisplatin resistance in breast cancer cells through nongenomic estrogen signaling, suggesting that ER-α36 may serve as a novel target for cisplatin resistance and a potential indicator of cisplatin sensitivity in breast cancer treatment.
已证实新型 ER-α66 变体 ER-α36 可促进乳腺癌细胞对他莫昔芬产生耐药性。然而，ER-α36 在乳腺癌细胞顺铂耐药中的作用和机制尚不清楚。本研究旨在探讨 ER-α36 在乳腺癌细胞顺铂耐药中的表达和作用，并阐明其潜在机制。
通过 Western blot 分析评估 ER-α36 和非基因组雌激素信号转导相关蛋白的表达。分别通过 CCK-8 assay、单层集落形成 assay 和凋亡 assay 探讨顺铂敏感性。用 ER-α36 siRNAs/shRNAs 和过表达载体转染细胞以下调或上调 ER-α36 表达。进行失活和激活功能实验以研究 ER-α36 在顺铂敏感性中的作用。使用 CoIP 检测 ER-α36 与 EGFR/HER-2 的相互作用。建立乳腺癌小鼠异种移植模型以验证 ER-α36 在体内的作用。
与顺铂敏感细胞相比，顺铂耐药乳腺癌细胞中 ER-α36 表达水平更高。顺铂以剂量依赖性方式诱导乳腺癌细胞中 ER-α36 的上调。过表达 ER-α36 导致细胞对顺铂产生耐药性，而在顺铂耐药乳腺癌细胞中敲低 ER-α36 则恢复了顺铂敏感性。ER-α36 的上调导致非基因组雌激素信号的激活增加，从而导致顺铂耐药。用激酶抑制剂阻断 ER-α36 介导的非基因组雌激素信号显著抑制顺铂诱导的 ER-α36 表达并增加顺铂敏感性。体内实验也证实，上调 ER-α36 可减弱乳腺癌小鼠异种移植模型中的顺铂敏感性。
本研究首次证明 ER-α36 通过非基因组雌激素信号介导乳腺癌细胞顺铂耐药，提示 ER-α36 可能成为顺铂耐药的新靶点，并可能成为乳腺癌治疗中顺铂敏感性的潜在指标。
