Liu Nan-nan, Xi Yue, Callaghan Michael U, Fribley Andrew, Moore-Smith Lakisha, Zimmerman Jacquelyn W, Pasche Boris, Zeng Qinghua, Li Yu-lin
The Key Laboratory of Pathobiology, Ministry of Education, Jilin University, Changchun, 130021, China,
Tumour Biol. 2014 Jan;35(1):641-50. doi: 10.1007/s13277-013-1088-1. Epub 2013 Aug 24.
SMAD4 is a downstream mediator of transforming growth factor beta. While its tumor suppressor function has been investigated as a prognostic biomarker in several human malignancies, its role as a prognostic marker in breast carcinoma is still undefined. We investigated SMAD4 expression in breast carcinoma samples of different histologic grades to evaluate the association between SMAD4 and outcome in breast cancer. We also investigated the role of SMAD4 expression status in MDA-MB-468 breast cancer cells in responding to TGF-β stimulation. SMAD4 expression was assessed in 53 breast ductal carcinoma samples and in the surrounding normal tissue from 50 of the samples using immunohistochemistry, Western blot, and real-time PCR. TGF-β-SMAD and non-SMAD signaling was assessed by Western blot in MDA-MB-468 cells with and without SMAD4 restoration. SMAD4 expression was reduced in ductal breast carcinoma as compared to surrounding uninvolved ductal breast epithelia (p < 0.05). SMAD4 expression levels decreased from Grade 1 to Grade 3 ductal breast carcinoma as assessed by immunohistochemistry (p < 0.05). Results were recapitulated by tissue array. In addition, immunohistochemistry results were further confirmed at the protein and mRNA level. We then found that non-SMAD MEK/MAPK signaling was significantly different between SMAD4 expressing MDA-MB-468 cells and SMAD4-null MDA-MB-468 cells. This is the first study indicating that SMAD4 plays a key role in shifting MAPK signaling. Further, we have demonstrated that SMAD4 has a potential role in the development of breast carcinoma and SMAD4 was a potential prognostic marker of breast carcinoma. Our findings further support the role of SMAD4 in breast carcinoma development. In addition, we observed an inverse relationship between SMAD4 levels and breast carcinoma histological grade. Our finding indicated that SMAD4 expression level in breast cancer cells played a role in responding non-SMAD signaling but not the canonic SMAD signaling. Further mechanistic studies are necessary to establish the role of SMAD4 in breast carcinoma prognosis and potential specific targeting.
SMAD4是转化生长因子β的下游介质。虽然其肿瘤抑制功能已在多种人类恶性肿瘤中作为预后生物标志物进行了研究，但其在乳腺癌中作为预后标志物的作用仍不明确。我们研究了不同组织学分级的乳腺癌样本中SMAD4的表达，以评估SMAD4与乳腺癌预后之间的关联。我们还研究了SMAD4表达状态在MDA-MB-468乳腺癌细胞对TGF-β刺激反应中的作用。使用免疫组织化学、蛋白质印迹和实时PCR评估了53例乳腺导管癌样本以及其中50例样本的周围正常组织中SMAD4的表达。通过蛋白质印迹评估了有和没有SMAD4恢复的MDA-MB-468细胞中的TGF-β-SMAD和非SMAD信号传导。与周围未受累的乳腺导管上皮相比，乳腺导管癌中SMAD4的表达降低（p<0.05）。通过免疫组织化学评估，从1级到3级乳腺导管癌，SMAD4表达水平降低（p<0.05）。组织芯片重现了结果。此外，免疫组织化学结果在蛋白质和mRNA水平上得到了进一步证实。然后我们发现，表达SMAD4的MDA-MB-468细胞和缺失SMAD4的MDA-MB-468细胞之间的非SMAD MEK/MAPK信号传导存在显著差异。这是第一项表明SMAD4在改变MAPK信号传导中起关键作用的研究。此外，我们已经证明SMAD4在乳腺癌的发生发展中具有潜在作用，并且SMAD4是乳腺癌的潜在预后标志物。我们的发现进一步支持了SMAD4在乳腺癌发生发展中的作用。此外，我们观察到SMAD4水平与乳腺癌组织学分级之间呈负相关。我们的发现表明，乳腺癌细胞中SMAD4的表达水平在对非SMAD信号传导而非经典SMAD信号传导的反应中起作用。需要进一步的机制研究来确定SMAD4在乳腺癌预后中的作用以及潜在的特异性靶向作用。
