S100P and Ezrin promote trans-endothelial migration of triple negative breast cancer cells
Triple negative breast cancer (TNBC) patients generally have an adverse clinical outcome because their tumors often recur and metastasize to distant sites in the first 3 years after surgery. Therefore, it has become pivotal to identify potential factors associated with metastasis. Here, we focused on the effects of S100P and Ezrin on the trans-endothelial migration (TEM) of TNBC cells, as they have both been suggested to play a role in this process in other malignancies.
The expression of S100P and Ezrin was examined by immunohistochemistry in 58 primary TNBC samples. The mRNA and protein levels of S100P and Ezrin were assessed in breast cancer-derived cell lines using qRT-PCR and Western blotting, respectively. Proliferation and migration assays were performed using TNBC-derived MFM-223 and SUM-185-PE cells transfected with S100P and Ezrin siRNAs. Two different timeframes were employed for TEM assays using TNBC-derived cells and human umbilical vein endothelial-derived cells, respectively. Correlations between the status of EzrinThr-567 expression and various clinicopathological features were analyzed by immunohistochemistry.
We found that S100P and Ezrin double negative TNBC cases were significantly associated with a better disease-free survival. We also found that single and double siRNA-mediated knockdown of S100P and Ezrin in TNBC-derived cells significantly inhibited their TEM and destabilized the intercellular junctions of endothelial cells. In addition, we found that EzrinThr-567 immunoreactivity significantly correlated with vascular invasion in TNBC patients.
From our data we conclude that S100P, Ezrin and EzrinThr-567 are involved in the trans-endothelial migration of TNBC cells and that they may serve as potential targets in TNBC patients.
KeywordsTriple negative breast carcinoma S100P Ezrin Trans-endothelial migration
We would like to acknowledge the support and assistance of the members of the Department of Pathology, Tohoku University School of Medicine. We appreciate Mr. H. Hiranuma (SCRUM Inc., Tokyo, Japan) for his technical assistance regarding xCELLigence. We also acknowledge the technical support of the Biomedical Research Unit of Tohoku University Hospital.
Compliance with ethical standards
This study was approved by Ethics Committee at Tohoku University School of Medicine. Informed consent was obtained from all patients.
Conflict of interest
The authors declare that they have no conflict of interest.
- 3.R. Rouzier, C.M. Perou, W.F. Symmans, N. Ibrahim, M. Cristofanilli, K. Anderson, K.R. Hess, J. Stec, M. Ayers, P. Wagner, P. Morandi, C. Fan, I. Rabiul, J.S. Ross, G.N. Hortobagyi, L. Pusztai, Breast cancer molecular subtypes respond differently to preoperative chemotherapy. Clin Cancer Res 11, 5678–5685 (2005)CrossRefPubMedGoogle Scholar
- 4.C. Liedtke, C. Mazouni, K.R. Hess, F. Andre, A. Tordai, J.A. Mejia, W.F. Symmans, A.M. Gonzalez-Angulo, B. Hennessy, M. Green, M. Cristofanilli, G.N. Hortobagyi, L. Pusztai, Response to neoadjuvant therapy and long-term survival in patients with triple-negative breast cancer. J Clin Oncol 26, 1275–1281 (2008)CrossRefPubMedGoogle Scholar
- 15.M. Komatsu, T. Yoshimura, T. Matsuo, K. Kiyotani, Y. Miyoshi, T. Tanihashi, K. Rokutani, R. Yamaguchi, A. Saito, S. Imoto, S. Miyano, Y. Nakamura, M. Sasa, M. Shimada, T. Katagiri, Molecular features of triple negative breast cancer cells by genome-wide gene expression profiling analysis. Int J Oncol 42, 478–506 (2013)CrossRefPubMedGoogle Scholar
- 29.S. Diederichs, E. Bulk, B. Steffen, P. Ji, L. Tickenbrock, K. Lang, K.S. Zanker, R. Metzger, P.M. Schneider, V. Gerke, M. Thomas, W.E. Berdel, H. Serve, C. Muller-Tidow, S100 family members and trypsinogens are predictors of distant metastasis and survival in early-stage non-small cell lung cancer. Cancer Res 64, 5564–5569 (2004)CrossRefPubMedGoogle Scholar
- 30.J. Austermann, A.R. Nazmi, C. Muller-Tidow, V. Gerke, Characterization of the Ca2+-regulated Ezrin-S100P interaction and its role in tumor cell migration. J Biochem 283, 29331–29340 (2008)Google Scholar
- 33.N. Maishi, Y. Ohba, K. Akiyama, N. Ohga, J. Hamada, H. Nagao-Kitamoto, M.T. Alam, K. Yamamoto, T. Kawamoto, N. Inoue, A. Taketomi, M. Shindoh, Y. Hida, K. Hida, Tumour endothelial cells in high metastatic tumours promote metastasis via epigenetic dysregulation of biglycan. Sci Reports 6, 28039 (2016)CrossRefGoogle Scholar
- 34.J.M. Iglesias, I. Beloqui, F. Garcia-Garcia, O. Leis, A. Vazquez- Martin, A. Eguiara, S. Cufi, A. Pavon, J.A. Menendez, J. Dopazo, A.G. Martin, Mammosphere formation in breast carcinoma cell lines depends upon expression of E-cadherin. PLoS One e77281, 8 (2013)Google Scholar
- 43.A. Chandramouli, M.E. Mercado-Pimentel, A. Hutchinson, A. Gibadulinová, E.R. Olson, S. Dickinson, R. Shañas, J. Davenport, J. Owens, A.K. Bhattacharyya, J.W. Regan, S. Pastorekova, T. Arumugam, C.D. Logsdon, M.A. Nelson, The induction of S100p expression by the Prostaglandin E2 (PGE2 )/EP4 receptor signaling pathway in colon cancer cells. Cancer Biol Ther 10, 1056–1066 (2010)CrossRefPubMedGoogle Scholar
- 46.C. Zhou, Q. Zhong, L.V. Rhodes, I. Townley, M.R. Bratton, Q. Zhang, E.C. Martin, S. Elliott, B.M. Collins-Burow, M.E. Burow, G. Wang, Proteomic analysis of acquired tamoxifen resistance in MCF-7 cells reveals expression signatures associated with enhanced migration. Breast Cancer Res 14, R45 (2012)CrossRefPubMedGoogle Scholar
- 48.Y. Chen, D. Wang, Z. Guo, J. Zhao, B. Wu, H. Deng, T. Zhou, H. Xiang, F. Gao, X. Yu, J. Liao, T. Ward, P. Xia, C. Emenari, X. Ding, W. Thompson, K. Ma, J. Zhu, F. Aikhionbare, K. Dou, S.Y. Cheng, X. Yao, Rho kinase phosphorylation promotes Ezrin-mediated metastasis in hepatocellular carcinoma. Cancer Res 71, 1721–1730 (2011)CrossRefPubMedGoogle Scholar
- 53.S. Hamada, K. Satoh, M. Hirota, W. Fujibuchi, A. Kanno, J. Umino, H. Ito, A. Satoh, K. Kikuta, K. Kume, A. Masamune, T. Shimosegawa, Expression of the calcium-binding protein S100P is regulated by bone morphogenetic protein in pancreatic duct epithelial cell lines. Cancer Sci 100, 103–110 (2008)CrossRefPubMedGoogle Scholar