Background:The intratumoural heterogeneity, often driven by epithelial-to-mesenchymal transition (EMT), significantly contributes to chemoresistance and disease progression in adenocarcinomas. Methods:We introduced a high-throughput screening platform to identify surface antigens that associate with epithelial–mesenchymal plasticity in well-defined pairs of epithelial cell lines and their mesenchymal counterparts. Using multicolour flow cytometry, we then analysed the expression of 10 most robustly changed antigens and identified a 10-molecule surface signature, in pan-cytokeratin-positive/EpCAM-positive and -negative fractions of dissociated breast tumours. Results:We found that surface CD9, CD29, CD49c, and integrin ß5 are lost in breast cancer cells that underwent EMT in vivo. The tetraspanin family member CD9 was concordantly downregulated both in vitro and in vivo and associated with epithelial phenotype and favourable prognosis. Conclusions:We propose that overall landscape of 10-molecule surface signature expression reflects the epithelial–mesenchymal plasticity in breast cancer.

Center of Biomolecular and Cellular Engineering International Clinical Research Center St Anne's University Hospital Brno Pekařská 53 Brno 656 91 Czech Republic

Department of Comprehensive Cancer Care Masaryk Memorial Cancer Institute Žlutý kopec 7 Brno 656 53 Czech Republic

Department of Experimental Biology Faculty of Science Masaryk University Kamenice 5 Brno 625 00 Czech Republic

Department of Oncological Pathology Masaryk Memorial Cancer Institute Žlutý kopec 7 Brno 656 53 Czech Republic

Institute of Biophysics of the Czech Academy of Sciences Královopolská 135 Brno 612 65 Czech Republic

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Al-Hajj M, Wicha MS, Benito-Hernandez A, Morrison SJ, Clarke MF (2003) Prospective identification of tumorigenic breast cancer cells. Proc Natl Acad Sci USA 100(7): 3983–3988. PubMed PMC

Anderson RC, Elder JB, Brown MD, Mandigo CE, Parsa AT, Kim PD, Senatus P, Anderson DE, Bruce JN (2002) Changes in the immunologic phenotype of human malignant glioma cells after passaging in vitro. Clin Immunol 102(1): 84–95. PubMed

Andreu Z, Yanez-Mo M (2014) Tetraspanins in extracellular vesicle formation and function. Front Immunol 5: 442. PubMed PMC

Berditchevski F (2001) Complexes of tetraspanins with integrins- more than meets the eye. J Cell Sci 114(23): 4143–4151. PubMed

Camphausen K, Purow B, Sproull M, Scott T, Ozawa T, Deen DF, Tofilon PJ (2005) Influence of in vivo growth on human glioma cell line gene expression- Convergent profiles under orthotopic conditions. Proc Natl Acad Sci USA 102(23): 8287–8292. PubMed PMC

Criscitiello C, Azim HA Jr., Schouten PC, Linn SC, Sotiriou C (2012) Understanding the biology of triple-negative breast cancer. Ann Oncol 23(Suppl 6): vi13–vi18. PubMed

Goswami C, Nakshatri H (2013) PROGgene - gene expression based survival analysis web application for multiple cancers. J Clin Bioinforma 3(1): 22. PubMed PMC

Gyorffy B, Schafer R (2009) Meta-analysis of gene expression profiles related to relapse-free survival in 1,079 breast cancer patients. Breast Cancer Res Treat 118(3): 433–441. PubMed

Halo TL, McMahon KM, Angeloni NL, Xu Y, Wang W, Chinen AB, Malin D, Strekalova E, Cryns VL, Cheng C, Mirkin CA, Thaxton CS (2014) NanoFlares for the detection, isolation, and culture of live tumor cells from human blood. Proc Natl Acad Sci USA 111(48): 17104–17109. PubMed PMC

Malladi S, Macalinao DG, Jin X, He L, Basnet H, Zou Y, de Stanchina E, Massague J (2016) Metastatic latency and immune evasion through autocrine inhibition of WNT. Cell 165(1): 45–60. PubMed PMC

Marusyk A, Almendro V, Polyak K (2012) Intra-tumour heterogeneity: a looking glass for cancer? Nat Rev Cancer 12(5): 323–334. PubMed

Medema JP (2013) Cancer stem cells: the challenges ahead. Nat Cell Biol 15(4): 338–344. PubMed

Mimori K, Kataoka A, Yoshinaga K, Ohta M, Sagara Y, Yoshikawa Y, Ohno S, Barnard GF, Mori M (2005) Identification of molecular markers for metastasis-related genes in primary breast cancer cells. Clin Exp Metastasis 22(1): 59–67. PubMed

Rappa G, Green TM, Karbanova J, Corbeil D, Lorico A (2015) Tetraspanin CD9 determines invasiveness and tumorigenicity of human breast cancer cells. Oncotarget 6(10): 7970–7991. PubMed PMC

Scheel C, Eaton EN, Li SH, Chaffer CL, Reinhardt F, Kah KJ, Bell G, Guo W, Rubin J, Richardson AL, Weinberg RA (2011) Paracrine and autocrine signals induce and maintain mesenchymal and stem cell states in the breast. Cell 145(6): 926–940. PubMed PMC

Schneck H, Gierke B, Uppenkamp F, Behrens B, Niederacher D, Stoecklein NH, Templin MF, Pawlak M, Fehm T, Neubauer H Disseminated Cancer Cell Network D (2015) EpCAM-independent enrichment of circulating tumor cells in metastatic breast cancer. PLoS ONE 10(12): e0144535. PubMed PMC

Slabakova E, Kharaishvili G, Smejova M, Pernicova Z, Suchankova T, Remsik J, Lerch S, Strakova N, Bouchal J, Kral M, Culig Z, Kozubik A, Soucek K (2015) Opposite regulation of MDM2 and MDMX expression in acquisition of mesenchymal phenotype in benign and cancer cells. Oncotarget 6(34): 36156–36171. PubMed PMC

Tsai JH, Yang J (2013) Epithelial-mesenchymal plasticity in carcinoma metastasis. Genes Dev 27(20): 2192–2206. PubMed PMC

Ye X, Weinberg RA (2015) Epithelial-mesenchymal plasticity: a central regulator of cancer progression. Trends Cell Biol 25(11): 675–686. PubMed PMC

Zhang C, Lodish H (2005) Murine hematopoietic stem cells change their surface phenotype during ex vivo expansion. Blood 105(11): 4314–4320. PubMed PMC

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