卡马西平对雌激素依赖性乳腺癌细胞增殖的抑制作用及其机制的研究

Inhibitory Effect of Carbamazepine on Proliferation of Estrogen-Dependent Breast Cancer Cells

背景与目的:卡马西平是一种应用多年的抗癫痫药,最近研究证明其具有组蛋白脱乙酰化酶抑制剂(histonedeacetylaseinhibitor,HDI)的性质。而已知的HDI多具有抗肿瘤作用。本实验旨在探讨卡马西平对雌激素受体!(estrogenreceptorα,ERα)阳性乳腺癌细胞增殖的抑制作用及其机制。方法:利用磺酰罗丹明B色度法(sulforhodamineB,SRB)分析不同情况下卡马西平对细胞增殖的影响。Westernblot法检测ERα、CyclinD1等相关蛋白的表达水平;RT-PCR法检测相应mRNA水平的变化;免疫荧光法测定他莫昔芬耐药细胞MCF-7RT中HER-2的表达;免疫沉淀技术检测Hsp90的分子伴侣功能及乙酰化水平。结果:卡马西平处理ERα阳性细胞MCF-7及T47D时,显著抑制雌二醇刺激的细胞增殖效应(P<0.01);卡马西平与4羟基他莫昔芬(4-OHT)联合处理MCF-7细胞,对雌二醇引起的细胞增殖抑制呈相加作用(q=1.00);在MCF-7RT细胞中,卡马西平逆转了4-OHT的刺激增殖作用(P<0.01);卡马西平处理可以降低ERα阳性细胞中ERα、CyclinD1在蛋白及mRNA水平的表达,并降低MCF-7RT细胞中HER-2表达;ERα、CyclinD1的表达降低可以被26s蛋白酶体抑制剂MG132抑制;卡马西平可以促进Hsp90乙酰化并破坏其分子伴侣功能。结论:卡马西平明显抑制ER阳性乳腺癌细胞增殖,该作用可能部分通过促进ERα、CyclinD1的蛋白酶体降解途径实现。卡马西平可以通过降低HER-2表达逆转与HER-2相关的4-OHT耐药。

BACKGROUND ＆ OBJECTIVE： Carbamazepine, which has been used as an anti-epileptic drug in clinic for many years, is currently recognized as a histone deacetylase inhibitor （HDI）, most of which showed anti-tumor characteristics. This study was to investigate the inhibitory effect of carbamazepine on estrogen dependent breast cancer cell lines with estrogen receptor α（ERα） expression and further explore the underlying mechanisms. METHODS. Sulforhodamine B viability assay was used to evaluate the viability of various cells treated with different drugs. Western blot and reverse transcription-polymerase chain reaction （RT-PCR） were performed to detect the protein and mRNA expression of ER~ and Cyclin D1. Immunofluorescence assay was employed to observe HER-2 expression in MCF-7RT cells, which were resistant to tamoxifen. Immunoprecipitation was performed to detect the chaperon function and acetylation level of Hsp90. RESULTS. Carbamazepine treatment could inhibit the proliferation of MCF-7 and T47D cells stimulated by estradiol （P〈0.01）. Carbamazepine and 4-hydroxytamoxifen （4-OHT） demonstrated a synergic effect on the inhibition of proliferation of MCF-7 cells stimulated by estradiol （q=1.00）. Cabamazepine reversed the proliferation of MCF-7RT cells stimulated by 4-hydroxytamoxifen （P〈0.01）. Carbamazepine treatment could decrease the expression of ERα and Cyclin D1 at protein and mRNA level in ERα-positive cells and could reduce HER-2 expression in MCF-7RT cells. The decrease of ERα and Cyclin D1 expression was inhibited by MG132, an inhibitor of 26S proteosome. Carbamazepine treatment elevated the acetylation level of Hsp90 and disruptted its chaperon function. CONCLUSIONS. Carbamazepine shows significant anti-proliferation effect in ERα-positive breast cancer cell lines and this might be due to the enhancement of proteosome-mediated degradation of ERα and Cyclin D1 by carbamazepine. Furthermore, carbamazepine could reverse HER-2 dependent drug resistance to 4-OHT by reducing HER-2 expression.