BackgroundCTCs expressing variable levels of epithelial and mesenchymal markers in breast cancer have previously been reported. However, no information exists for keratin expression levels of CTCs in association with disease status, whereas assays for the characterization of transitional EMT phenotypes of CTCs in breast cancer are rather lacking. We investigated the correlation between keratin expression of CTCs and patients’ outcome and characterized the EMT status of CTCs via the establishment of a numerical “ratio” value of keratin and vimentin expression levels on a single cell basis.MethodsKeratin expression was evaluated in 1262 CTCs from 61 CTC-positive patients with metastatic breast cancer, using analysis of images obtained through the CellSearch System. For the determination of vimentin/keratin (vim/K) ratios, expression levels of keratin and vimentin were measured in cytospin preparations of luminal (MCF-7 and T47D) and basal (MDA.MB231 and Hs578T) breast cancer cell lines and 110 CTCs from 5 CTC-positive patients using triple immunofluorescence laser scanning microscopy and image analysis.ResultsMCF-7 and T47D displayed lower vim/K ratios compared to MDA.MB231 and Hs578T cells, while MCF-7 cells that had experimentally undergone EMT were characterized by varying intermediate vim/K ratios. CTCs were consisted of an heterogeneous population presenting variable vim/K values with 46% of them being in the range of luminal breast cancer cell lines. Keratin expression levels of CTCs detected by the CellSearch System correlated with triple negative (p = 0.039) and ER-negative (p = 0.025) breast cancer, and overall survival (p = 0.038).ConclusionsKeratin expression levels of CTCs correlate with tumor characteristics and clinical outcome. Moreover, CTCs display significant heterogeneity in terms of the degree of EMT phenotype that probably reflects differential invasive potential. The assessment of the vim/K ratios as a surrogate marker for the EMT status of CTCs merits further investigation as a prognostic tool in breast cancer.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1386-7) contains supplementary material, which is available to authorized users.

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Breast cancer stem cells (BCSCs) represent a heterogeneous subpopulation of rare cells within breast cancer tumors, displaying an enhanced tumor initiating capability and underlying disease progression and therapy resistance. Unraveling their phenotypic, biological and functional profile is a major challenge in the context of diminishing patient mortality. In this review, following a brief description on how cancer stem cells (CSCs) and their microenvironment contribute to tumor preservation and heterogeneity, we summarize the current literature regarding the molecular signature of BCSCs either localized in the primary tumor or circulating in the blood of breast cancer patients. We present recent data on specific stem and epithelial-to-mesenchymal transition (EMT) markers designating the BCSC subpopulation and underline their pathogenic significance. The molecular characterization of BCSCs has promoted the design of novel therapeutic approaches targeting the BCSC subpopulation which are currently being experimentally and clinically evaluated. We highlight recent advances on the development of novel BCSC-targeting therapeutic strategies including the inhibition of cell signaling pathways, differentiation therapy, metabolic interference and nucleotide-, bio- and nano-technology based approaches. Eliminating the chemo- and radio-resistance properties of breast cancer tumor cells via BCSC-directed therapies, combined to conventional therapeutic approaches, will augment the effectiveness of breast cancer treatment and improve the clinical outcome of breast cancer patients.