Fox Emily M, Bernaciak Teresa M, Wen Jie, Weaver Amanda M, Shupnik Margaret A, Silva Corinne M
Department of Pharmacology, University of Virginia Health Sciences Center, Charlottesville, Virginia 22908, USA.
Mol Endocrinol. 2008 Aug;22(8):1781-96. doi: 10.1210/me.2007-0419. Epub 2008 Jun 11.
17beta-Estradiol (E2) acts through the estrogen receptor alpha (ERalpha) to stimulate breast cancer proliferation. Here, we investigated the functional relationship between ERalpha and signal transducer and activator of transcription (STAT)5b activity in ER+ MCF-7 and T47D human breast cancer cells after specific knockdown of STAT5b. STAT5b small interfering RNA (siRNA) inhibited E2-induced bromodeoxyuridine (BrdU) incorporation in both cell lines, as well as the E2-induced increase in MCF-7 cell number, cyclin D1 and c-myc mRNA, and cyclin D1 protein expression, indicating that STAT5b is required for E2-stimulated breast cancer proliferation. E2 treatment stimulated STAT5b tyrosine phosphorylation at the activating tyrosine Y699, resulting in increased STAT5-mediated transcriptional activity, which was inhibited by a Y669F STAT5b mutant. E2-induced STAT5-mediated transcriptional activity was inhibited by overexpressing a kinase-defective epidermal growth factor receptor (EGFR), or the EGFR tyrosine kinase inhibitor tyrphostin AG1478, indicating a requirement for EGFR kinase activity. Both E2-induced STAT5b tyrosine phosphorylation and STAT5-mediated transcription were also inhibited by the ER antagonist ICI 182,780 and the c-Src inhibitor PP2, indicating additional requirements for the ER and c-Src kinase activity. EGFR and c-Src kinase activities were also required for E2-induced cyclin D1 and c-myc mRNA. Together, these studies demonstrate positive cross talk between ER, c-Src, EGFR, and STAT5b in ER+ breast cancer cells. Increased EGFR and c-Src signaling is associated with tamoxifen resistance in ER+ breast cancer cells. Here we show that constitutively active STAT5b not only increased basal DNA synthesis, but also conferred tamoxifen resistance. Because STAT5b plays an integral role in E2-stimulated proliferation and tamoxifen resistance, it may be an effective therapeutic target in ER+ breast tumors.
17β-雌二醇（E2）通过雌激素受体α（ERα）发挥作用，刺激乳腺癌细胞增殖。在此，我们研究了在ER⁺ MCF-7和T47D人乳腺癌细胞中特异性敲低信号转导和转录激活因子（STAT）5b后，ERα与STAT5b活性之间的功能关系。STAT5b小干扰RNA（siRNA）抑制了两种细胞系中E2诱导的溴脱氧尿苷（BrdU）掺入，以及E2诱导的MCF-7细胞数量增加、细胞周期蛋白D1和c-myc mRNA以及细胞周期蛋白D1蛋白表达增加，表明STAT5b是E2刺激的乳腺癌细胞增殖所必需的。E2处理刺激了STAT5b在激活酪氨酸Y699处的酪氨酸磷酸化，导致STAT5介导的转录活性增加，而Y669F STAT5b突变体可抑制该活性。过表达激酶缺陷型表皮生长因子受体（EGFR）或EGFR酪氨酸激酶抑制剂 tyrphostin AG1478可抑制E2诱导的STAT5介导的转录活性，表明需要EGFR激酶活性。ER拮抗剂ICI 182,780和c-Src抑制剂PP2也抑制了E2诱导的STAT5b酪氨酸磷酸化和STAT5介导的转录，表明还需要ER和c-Src激酶活性。E2诱导的细胞周期蛋白D1和c-myc mRNA也需要EGFR和c-Src激酶活性。总之，这些研究证明了ER⁺乳腺癌细胞中ER、c-Src、EGFR和STAT5b之间存在正向相互作用。EGFR和c-Src信号增强与ER⁺乳腺癌细胞对他莫昔芬耐药有关。在此我们表明，组成型激活的STAT5b不仅增加了基础DNA合成，还赋予了对他莫昔芬的耐药性。由于STAT5b在E2刺激的增殖和他莫昔芬耐药中起不可或缺的作用，它可能是ER⁺乳腺肿瘤的一个有效治疗靶点。
