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01.12.2017 | Research | Ausgabe 1/2017 Open Access

Molecular Cancer 1/2017

Syndecan-1 is a novel molecular marker for triple negative inflammatory breast cancer and modulates the cancer stem cell phenotype via the IL-6/STAT3, Notch and EGFR signaling pathways

Zeitschrift:
Molecular Cancer > Ausgabe 1/2017
Autoren:
Sherif Abdelaziz Ibrahim, Ramy Gadalla, Eslam A. El-Ghonaimy, Omnia Samir, Hossam Taha Mohamed, Hebatallah Hassan, Burkhard Greve, Mohamed El-Shinawi, Mona Mostafa Mohamed, Martin Götte
Wichtige Hinweise

Electronic supplementary material

The online version of this article (doi:10.​1186/​s12943-017-0621-z) contains supplementary material, which is available to authorized users.

Abstract

Background

Inflammatory breast cancer (IBC), a particularly aggressive form of breast cancer, is characterized by cancer stem cell (CSC) phenotype. Due to a lack of targeted therapies, the identification of molecular markers of IBC is of major importance. The heparan sulfate proteoglycan Syndecan-1 acts as a coreceptor for growth factors and chemokines, modulating inflammation, tumor progression, and cancer stemness, thus it may emerge as a molecular marker for IBC.

Methods

We characterized expression of Syndecan-1 and the CSC marker CD44, Notch-1 & -3 and EGFR in carcinoma tissues of triple negative IBC (n = 13) and non-IBC (n = 17) patients using qPCR and immunohistochemistry. Impact of siRNA-mediated Syndecan-1 knockdown on the CSC phenotype of the human triple negative IBC cell line SUM-149 and HER-2-overexpressing non-IBC SKBR3 cells employing qPCR, flow cytometry, Western blotting, secretome profiling and Notch pharmacological inhibition experiments. Data were statistically analyzed using Student’s t-test/Mann-Whitney U-test or one-way ANOVA followed by Tukey’s multiple comparison tests.

Results

Our data indicate upregulation and a significant positive correlation of Syndecan-1 with CD44 protein, and Notch-1 & -3 and EGFR mRNA in IBC vs non-IBC. ALDH1 activity and the CD44(+)CD24(-/low) subset as readout of a CSC phenotype were reduced upon Syndecan-1 knockdown. Functionally, Syndecan-1 silencing significantly reduced 3D spheroid and colony formation. Intriguingly, qPCR results indicate downregulation of the IL-6, IL-8, CCL20, gp130 and EGFR mRNA upon Syndecan-1 suppression in both cell lines. Moreover, Syndecan-1 silencing significantly downregulated Notch-1, -3, -4 and Hey-1 in SUM-149 cells, and downregulated only Notch-3 and Gli-1 mRNA in SKBR3 cells. Secretome profiling unveiled reduced IL-6, IL-8, GRO-alpha and GRO a/b/g cytokines in conditioned media of Syndecan-1 knockdown SUM-149 cells compared to controls. The constitutively activated STAT3 and NFκB, and expression of gp130, Notch-1 & -2, and EGFR proteins were suppressed upon Syndecan-1 ablation. Mechanistically, gamma-secretase inhibition experiments suggested that Syndecan-1 may regulate the expression of IL-6, IL-8, gp130, Hey-1, EGFR and p-Akt via Notch signaling.

Conclusions

Syndecan-1 acts as a novel tissue biomarker and a modulator of CSC phenotype of triple negative IBC via the IL-6/STAT3, Notch and EGFR signaling pathways, thus emerging as a promising therapeutic target for IBC.
Zusatzmaterial
Additional file 1: Table S1. Primers sequences. (DOCX 18 kb)
12943_2017_621_MOESM1_ESM.docx
Additional file 2: Figure S1. A representative image of tumor emboli, a unique feature for tissues of IBC patients showing a positive staining for CD44 and Syndecan-1. (ZIP 2337 kb)
12943_2017_621_MOESM2_ESM.zip
Additional file 3: Figure S2. Flow cytometric analysis of Syndecan-1 expression in control and Syndecan-1 siRNA transfected SUM-149 cells. 500,000 cells were stained for isotype control mouse IgG1-PE and mouse anti-human Syndecan-1 (CD138)-PE and the cells were subjected to flow cytometry. Each plot shows mouse IgG-PE control (dotted line) and CD138-PE-stained cells (solid line). The median fluorescence intensity (MFI) of events is given for each peak. Data are a single experiment representative of three independent experiments. (ZIP 281 kb)
12943_2017_621_MOESM3_ESM.zip
Literatur
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