Liu Dan, Deng Que, Sun Limin, Wang Tao, Yang Zhengyan, Chen Hongyu, Guo Liang, Liu Yanjun, Ma Yuanfang, Guo Ning, Shi Ming
Institute of Basic Medical Sciences, Beijing, 100850, P.R. China.
307 Hospital of People's Liberation Army, Beijing, 100071, P.R. China.
BMC Cancer. 2015 Nov 2;15:832. doi: 10.1186/s12885-015-1869-6.
Our previous studies show that β2-adrenergic receptor (β2-AR) is highly expressed in most Her2-overexpressing breast cancers. However, the mechanisms underlying upregulation of the β2-AR expression in Her2-overexpressing breast cancer cells are not fully understood. The clinical significance of the β2-AR overexpression in breast cancer is unclear.
Human breast cancer cells MCF-7 and MCF-7/Her2 were transfected with the let-7 mimics or inhibitors. The expression of β2-AR was analyzed by Western blot. The β2-AR status in primary and metastatic sites of breast cancer and the human breast cancer tissue microarrays containing 49 primary tumors and 50 metastatic lymph node tissues was analyzed by immunohistochemistry. The correlation of lymph node metastasis with the β2-AR level was determined in 59 primary tumor tissues from the patients with Her2-positive breast cancer. The clinical prognostic significance of the β2-AR overexpression in the patients with Her2-positive breast cancers was evaluated by a retrospective study.
The let-7f level in Her2-overexpressing breast cancer cells SKBR3 and BT474 was significantly lower than that in MCF-7 cells, which express low level of Her2. Ectopic expression of Her2 in MCF-7 cells (MCF-7/Her2) represses the expression of microRNA let-7f, which is previously identified to regulate baseline β2-AR expression. The treatment with MEK1/2 inhibitors PD98059 or PD184352 effectively restored the let-7f level, suggesting that Her2-overexpression-mediated ERK constitutive activation inhibited let-7f, leading to the upregulation of the β2-AR expression. The transfection with the let-7f mimics markedly downregulated the β2-AR level, whereas the let-7 inhibitor significantly upregulated the β2-AR expression in both parental MCF-7 and MCF-7/Her2 cells. In addition, treatment of MCF-7/Her2 cells with isoproterenol resulted in a concentration-dependent reduction of the let-7f expression, demonstrating that the inhibitory effect of Her2 overexpression on let-7f can be reinforced by agonist-triggered β2-AR activation. We further demonstrate that high level of β2-AR associates with lymph node metastasis and poor outcome in the patients with Her2-positive breast cancer.
The mutual and reciprocal interaction between Her2, β2-AR, and let-7f may maintain a high level of β2-AR in breast cancer cells. Our data suggest that β2-AR may be a new useful biomarker for predicting prognosis in Her2-positive breast cancer and may also be a promising selective therapeutic target for the aggressive subtype of breast cancer.
我们之前的研究表明，β2肾上腺素能受体（β2-AR）在大多数人表皮生长因子受体2（Her2）过表达的乳腺癌中高表达。然而，Her2过表达的乳腺癌细胞中β2-AR表达上调的潜在机制尚未完全阐明。β2-AR在乳腺癌中过表达的临床意义尚不清楚。
用let-7模拟物或抑制剂转染人乳腺癌细胞MCF-7和MCF-7/Her2。通过蛋白质免疫印迹法分析β2-AR的表达。采用免疫组织化学方法分析乳腺癌原发灶和转移灶以及包含49个原发性肿瘤和50个转移淋巴结组织的人乳腺癌组织芯片中的β2-AR状态。在59例Her2阳性乳腺癌患者的原发性肿瘤组织中确定淋巴结转移与β2-AR水平的相关性。通过回顾性研究评估β2-AR在Her2阳性乳腺癌患者中过表达的临床预后意义。
Her2过表达的乳腺癌细胞SKBR3和BT474中的let-7f水平显著低于Her2表达水平低的MCF-7细胞。MCF-7细胞（MCF-7/Her2）中Her2的异位表达抑制了微小RNA let-7f的表达，let-7f先前已被确定可调节β2-AR的基础表达。用丝裂原活化蛋白激酶激酶1/2（MEK1/2）抑制剂PD98059或PD184352处理可有效恢复let-7f水平，这表明Her2过表达介导的细胞外信号调节激酶（ERK）组成性激活抑制了let-7f，导致β2-AR表达上调。用let-7f模拟物转染可显著下调β2-AR水平，而let-7抑制剂则显著上调亲本MCF-7和MCF-7/Her2细胞中的β2-AR表达。此外，用异丙肾上腺素处理MCF-7/Her2细胞导致let-7f表达呈浓度依赖性降低，表明Her2过表达对let-7f的抑制作用可被激动剂触发的β2-AR激活增强。我们进一步证明，β2-AR高水平与Her2阳性乳腺癌患者的淋巴结转移和不良预后相关。
Her2、β2-AR和let-7f之间的相互作用可能维持乳腺癌细胞中β2-AR的高水平。我们的数据表明，β2-AR可能是预测Her2阳性乳腺癌预后的一种新的有用生物标志物，也可能是侵袭性乳腺癌亚型的一个有前景的选择性治疗靶点。
