Abstract
IntroductionTumor-initiating cells (TICs), aka “cancer stem cells”, are believed to fuel tumors and to sustain therapy resistance and systemic metastasis. Breast cancer is the first human carcinoma in which a subpopulation of cells displaying a specific CD44+/CD24-/low/ESA+ antigenic phenotype was found to have TIC properties. However, CD44+/CD24-/low/ESA+ is not a universal marker phenotype of TICs in all breast cancer subtypes. The aim of this study was to identify novel antigens with which to isolate the TIC population of the basal-A/basal-like breast cancer cell lines.MethodsWe used polychromatic flow-cytometry to characterize the cell surface of several breast cancer cell lines that may represent different tumor molecular subtypes. We next used fluorescence-activated cell sorting to isolate the cell subpopulations of interest from the cell lines. Finally, we explored the stem-like and tumorigenic properties of the sorted cell subpopulations using complementary in vitro and in vivo approaches: mammosphere formation assays, soft-agar colony assays, and tumorigenic assays in NOD/SCID mice.ResultsThe CD44+/CD24+ subpopulation of the BRCA1-mutated basal-A/basal-like cell line HCC1937 is enriched in several stemness markers, including the ABCG2 transporter (i.e., the CD338 antigen). Consistently, CD338-expressing cells were also enriched in CD24 expression, suggesting that coexpression of these two antigenic markers may segregate TICs in this cell line. In support of ABCG2 expression in TICs, culturing of HCC1937 cells in ultra-low adherent conditions to enrich them in precursor/stem-cells resulted in an increase in CD338-expressing cells. Furthermore, CD338-expressing cells, unlike their CD338-negative counterparts, displayed stemness and transformation potential, as assessed in mammosphere and colony formation assays. Lastly, CD338-expressing cells cultured in ultra-low adherent conditions maintained the expression of CD326/EpCAM and CD49f/α6-integrin, which is a combination of antigens previously assigned to luminal progenitors.ConclusionCollectively, our data suggest that CD338 expression is specific to the tumor-initiating luminal progenitor subpopulation of BRCA1-mutated cells and is a novel antigen with which to sort this subpopulation.Electronic supplementary materialThe online version of this article (doi:10.1186/1476-4598-13-213) contains supplementary material, which is available to authorized users.
Highlights
Tumor-initiating cells (TICs), aka “cancer stem cells”, are believed to fuel tumors and to sustain therapy resistance and systemic metastasis
To identify novel antigens that can improve the power of the CD44/CD24 antigenic phenotype in order to isolate TICs, we measured the expression of 28 surface antigens reported to be essential for cell adhesion, migration, apoptosis, cell signaling or stemness (Table 1 and Additional file 1: Table S1) in two basal A/basal-like cell lines, namely BT20 and HCC1937 (BRCA1−/−) and the basal B/claudin-low Hs578T cell line
No significant differences were observed in the expression of the examined antigens between the CD44+/CD24+ and CD44+/CD24-/low cell subpopulations in the BT20 and Hs578T cell lines, while several of them were significantly enriched in the CD44+/CD24+ population of HCC1937, including the stemness markers CD10, CD133 and CD338/ATP-Binding Cassette G2 (ABCG2) [25,31,32]
Summary
Tumor-initiating cells (TICs), aka “cancer stem cells”, are believed to fuel tumors and to sustain therapy resistance and systemic metastasis. CD44+/CD24-/low/ESA+ does not constitute a universal antigenic phenotype of TICs in all breast cancer subtypes [16,17,18] Rather, it marks a heterogeneous mix of cells in normal mammary gland [19] and is a profile associated with cell commitment to an EMT program [20]. It is necessary to better define this antigenic phenotype by combining CD44 and CD24 with additional as yet unidentified markers or activity, as previously shown with aldehyde dehydrogenase [21] In this context, the similar distribution of the gene expression profiles of breast cancer cell lines and primary tumors in the five subtypes identified [2,22] suggests that the cell line diversity reflects the tumor heterogeneity. The aim of this study was to identify novel antigens that are able to isolate the TIC population of basal-A/basal-like breast cancer cell lines
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