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A dynamic in vivo model of epithelial-to-mesenchymal transitions in circulating tumor cells and metastases of breast cancer

Oncogene volume 31, pages 3741–3753 (2012)Cite this article

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Abstract

Epithelial-to-mesenchymal transition (EMT) processes endow epithelial cells with enhanced migratory/invasive properties and are therefore likely to contribute to tumor invasion and metastatic spread. Because of the difficulty in following EMT processes in human tumors, we have developed and characterized an animal model with transplantable human breast tumor cells (MDA-MB-468) uniquely showing spontaneous EMT events to occur. Using vimentin as a marker of EMT, heterogeneity was revealed in the primary MDA-MB-468 xenografts with vimentin-negative and vimentin-positive areas, as also observed on clinical human invasive breast tumor specimens. Reverse transcriptase–PCR after microdissection of these populations from the xenografts revealed EMT traits in the vimentin-positive zones characterized by enhanced ‘mesenchymal gene’ expression (Snail, Slug and fibroblast-specific protein-1) and diminished expression of epithelial molecules (E-cadherin, ZO-3 and JAM-A). Circulating tumor cells (CTCs) were detected in the blood as soon as 8 days after s.c. injection, and lung metastases developed in all animals injected as examined by in vivo imaging analyses and histology. High levels of vimentin RNA were detected in CTCs by reverse transcriptase-quantitative PCR as well as, to a lesser extent, Snail and Slug RNA. Von Willebrand Factor/vimentin double immunostainings further showed that tumor cells in vascular tumoral emboli all expressed vimentin. Tumoral emboli in the lungs also expressed vimentin whereas macrometastases displayed heterogenous vimentin expression, as seen in the primary xenografts. In conclusion, our data uniquely demonstrate in an in vivo context that EMT occurs in the primary tumors, and associates with an enhanced ability to intravasate and generate CTCs. They further suggest that mesenchymal-to-epithelial phenomena occur in secondary organs, facilitating the metastatic growth.

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Acknowledgements

The research effort associated with this article was funded in part by the Fonds de la Recherche Scientifique-FNRS (F.R.S.-FNRS, Belgium), the Foundation against Cancer (foundation of public interest, Belgium), the ‘C.G.R.I.-F.N.R.S.-INSERM Coopération’, the Fonds spéciaux de la Recherche (University of Liège), the Centre Anticancéreux près l’Université de Liège, the Fonds Léon Fredericq (University of Liège), the ‘Région Champagne-Ardenne’, the ‘Ligue Contre le Cancer’, the Lions Club of Soissons, Un Euro contre le Cancer and the Fond National pour la Santé ACI 2004-2010 INCa (Cancéropôle Grand-Est project), the US. Army Medical Research and Materiel Command (BC0213201 and BC084667), the Victorian Breast Cancer Research Consortium, Cancer Council Victoria (#509295), the National Breast Cancer Foundation (Australia). CG is a Senior Research Associate from the F.R.S.-FNRS (Belgium). AB is supported by the ‘Région Champagne-Ardenne’. We thank Nathalie Lefin and Benoît Brouwers for their technical assistance.

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Authors and Affiliations

  1. Laboratory of Tumor and Developmental Biology, GIGA-Cancer, University of Liège, Liège, Belgium

    A Bonnomet, L Syne, A Brysse, E Feyereisen, A Noël, J-M Foidart & C Gilles

  2. Department of Surgery, St Vincent's Institute and University of Melbourne, St Vincent's Hospital, Melbourne, Australia

    E W Thompson

  3. Unité INSERM U.903, Laboratoire Pol Bouin, Reims, France

    A Bonnomet, P Birembaut & M Polette

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  1. A Bonnomet
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Correspondence to C Gilles.

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Bonnomet, A., Syne, L., Brysse, A. et al. A dynamic in vivo model of epithelial-to-mesenchymal transitions in circulating tumor cells and metastases of breast cancer. Oncogene 31, 3741–3753 (2012). https://doi.org/10.1038/onc.2011.540

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