Loss of Rho GDIα and Resistance to Tamoxifen via Effects on Estrogen Receptor α
Loss of Rho GDIα and Resistance to Tamoxifen via Effects on Estrogen Receptor α
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DOI：
10.1093/jnci/djr058
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发表时间：
2011-04-01
影响因子：
10.3
通讯作者：
Fuqua, Suzanne A. W.
中科院分区：
文献类型：
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作者：
Barone, Ines;Brusco, Lauren;Fuqua, Suzanne A. W.
背景雌激素受体(ER)α是乳腺癌成功的治疗靶点，但患者最终会对他莫昔芬等抗雌激素药物产生耐药性。方法为了确定与他莫昔芬耐药和转移相关的基因表达，我们使用微阵列比较了4例乳腺癌未复发的他莫昔芬治疗患者的原发肿瘤和5例在他莫昔芬辅助治疗期间肿瘤进展的转移肿瘤的基因表达。由于Rho鸟嘌呤解离抑制物(GDI)α在三苯氧胺耐药组中低表达，我们用编码短发夹状(Sh)RNA的质粒稳定地转染ERα阳性的MCF-7乳腺癌细胞以沉默Rho GDIα的表达。我们使用免疫印迹和转录分析来检测Rho GDIα在ER相关信号和细胞生长中的作用，以及作为裸鼠(每组8-9只)异种移植的作用，以检测Rho GDIα阻断对激素反应和转移行为的影响。在接受治疗的小鼠中，检测了从随机化到总肿瘤体积比基线增加三倍所需的几周内肿瘤增加三倍的时间。在来自患者的微阵列数据中，研究了Rho、GDIα和MTA2水平与他莫昔芬耐药性的关系。结果Rho GDIα在Era阳性肿瘤中的表达水平低于在他莫昔芬治疗期间复发的Era阳性肿瘤。与对照细胞相比，沉默Rho GDIa表达的MCF-7乳腺癌细胞对他莫昔芬耐药，Rho GTP酶和p21激活的激酶1活性增加，ERα丝氨酸305位的磷酸化水平增加，他莫昔芬诱导的ERα转录活性增强。Rho GDIα表达沉默的Mcf-7细胞在作为肿瘤移植瘤生长时转移频率很高。当小鼠接受雌激素或雌激素撤除治疗时，来自Rho GDIα沉默的细胞的异种移植的三倍时间与含有载体的对照细胞的三倍时间相似；但是，当他莫昔芬治疗小鼠时，三倍时间显著快于对照小鼠(携带Rho GDIα小干扰RNA的肿瘤的中位数三倍时间=2.34周；对于未达到的对照肿瘤，危险比=4.13，95%可信区间=1.07至15.96，P=.040[经多次比较调整，P=.119])。Rho GDIα沉默后，转移相关蛋白MTA2的水平也增加，并且Rho GDIα和MTA2的联合水平与他莫昔芬治疗的250例患者的复发相关。结论Rho GDIα的缺失通过影响ERα阳性乳腺癌模型和三苯氧胺治疗患者的肿瘤，增强了肿瘤的转移和对他莫昔芬的耐药性。
Background Estrogen receptor (ER) alpha is a successful therapeutic target in breast cancer, but patients eventually develop resistance to antiestrogens such as tamoxifen.Methods To identify genes whose expression was associated with the development of tamoxifen resistance and metastasis, we used microarrays to compare gene expression in four primary tumors from tamoxifen-treated patients whose breast cancers did not recur vs five metastatic tumors from patients whose cancers progressed during adjuvant tamoxifen treatment. Because Rho guanine dissociation inhibitor (GDI) alpha was underexpressed in the tamoxifen-resistant group, we stably transfected ER alpha-positive MCF-7 breast cancer cells with a plasmid encoding a short hairpin (sh) RNA to silence Rho GDI alpha expression. We used immunoblots and transcription assays to examine the role of Rho GDI alpha in ER-related signaling and growth of cells in vitro and as xenografts in treated nude mice (n = 8-9 per group) to examine the effects of Rho GDI alpha blockade on hormone responsiveness and metastatic behavior. The time to tumor tripling as the time in weeks from randomization to a threefold increase in total tumor volume over baseline was examined in treated mice. The associations of Rho GDI alpha and MTA2 levels with tamoxifen resistance were examined in microarray data from patients. All statistical tests were two-sided.Results Rho GDI alpha was expressed at lower levels in ERa-positive tumors that recurred during tamoxifen treatment than in ERa-positive tamoxifen-sensitive primary tumors. MCF-7 breast cancer cells in which Rho GDIa expression had been silenced were tamoxifen-resistant, had increased Rho GTPase and p21-activated kinase 1 activity, increased phosphorylation of ER alpha at serine 305, and enhanced tamoxifen-induced ER alpha transcriptional activity compared with control cells. MCF-7 cells in which Rho GDI alpha expression was silenced metastasized with high frequency when grown as tumor xenografts. When mice were treated with estrogen or estrogen withdrawal, tripling times for xenografts from cells with Rho GDI alpha silencing were similar to those from vector-containing control cells; however, tripling times were statistically significantly faster than control when mice were treated with tamoxifen (median tripling time for tumors with Rho GDI alpha small interfering RNA = 2.34 weeks; for control tumors = not reached, hazard ratio = 4.13, 95% confidence interval = 1.07 to 15.96, P = .040 [ adjusted for multiple comparisons, P = .119]). Levels of the metastasis-associated protein MTA2 were also increased upon Rho GDI alpha silencing, and combined Rho GDI alpha and MTA2 levels were associated with recurrence in 250 tamoxifen-treated patients.Conclusion Loss of Rho GDI alpha enhances metastasis and resistance to tamoxifen via effects on both ER alpha and MTA2 in models of ER alpha-positive breast cancer and in tumors of tamoxifen-treated patients.