title: 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.
abstract: 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.
  
  
  
  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.
  
  
  
  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.
  
  
  
  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.
authors.affiliation: Department of Zoology, Faculty of Science, Cairo University, 12613, Giza, Egypt. isherif@sci.cu.edu.eg.
authors.forename: Sherif Abdelaziz
authors.initials: SA
authors.lastname: Ibrahim
authors.affiliation: Department of Zoology, Faculty of Science, Cairo University, 12613, Giza, Egypt.
authors.forename: Ramy
authors.initials: R
authors.lastname: Gadalla
authors.affiliation: Department of Zoology, Faculty of Science, Cairo University, 12613, Giza, Egypt.
authors.forename: Eslam A
authors.initials: EA
authors.lastname: El-Ghonaimy
authors.affiliation: Department of Zoology, Faculty of Science, Cairo University, 12613, Giza, Egypt.
authors.forename: Omnia
authors.initials: O
authors.lastname: Samir
authors.affiliation: Department of Zoology, Faculty of Science, Cairo University, 12613, Giza, Egypt.
authors.forename: Hossam Taha
authors.initials: HT
authors.lastname: Mohamed
authors.affiliation: Department of Zoology, Faculty of Science, Cairo University, 12613, Giza, Egypt.
authors.forename: Hebatallah
authors.initials: H
authors.lastname: Hassan
authors.affiliation: Department of Radiotherapy-Radiooncology, University Hospital Münster, Münster, Germany.
authors.forename: Burkhard
authors.initials: B
authors.lastname: Greve
authors.affiliation: Department of General Surgery, Faculty of Medicine, Ain Shams University, 11566, Cairo, Egypt.
authors.forename: Mohamed
authors.initials: M
authors.lastname: El-Shinawi
authors.affiliation: Department of Zoology, Faculty of Science, Cairo University, 12613, Giza, Egypt.
authors.forename: Mona Mostafa
authors.initials: MM
authors.lastname: Mohamed
authors.affiliation: Department of Gynecology and Obstetrics, Münster University Hospital, Albert-Schweitzer-Campus 1, D11, 48149, Münster, Germany. martingotte@uni-muenster.de.
authors.forename: Martin
authors.initials: M
authors.lastname: Götte
mesh_terms: D000328:Adult; D015415:Biomarkers; D045744:Cell Line, Tumor; D066246:ErbB Receptors / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D058922:Inflammatory Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D015850:Interleukin-6 / Q000378:metabolism; D008875:Middle Aged; D016328:NF-kappa B / Q000378:metabolism; D060787:Neoplasm Grading; D009362:Neoplasm Metastasis; D014411:Neoplastic Stem Cells / Q000378:metabolism* / Q000473:pathology; D010641:Phenotype; D020543:Proteome; D040901:Proteomics / Q000379:methods; D051880:Receptors, Notch / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction; D053668:Syndecan-1 / Q000235:genetics / Q000378:metabolism*; D064726:Triple Negative Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology
chemical_list: D015415:Biomarkers; C497483:CD44 protein, human; D018960:Hyaluronan Receptors; D015850:Interleukin-6; D016328:NF-kappa B; D020543:Proteome; D051880:Receptors, Notch; D050796:STAT3 Transcription Factor; D053668:Syndecan-1; D066246:ErbB Receptors
keywords: Cancer stem cell; EGFR; IL-6/STAT3; Inflammatory breast cancer; Notch; Proteoglycan; Syndecan-1