Fascin Is Critical for the Maintenance of Breast Cancer Stem Cell Pool Predominantly via the Activation of the Notch Self-Renewal Pathway

Abstract

An emerging dogma shows that tumors are initiated and maintained by a subpopulation of cancer cells that hijack some stem cell features and thus referred to as "cancer stem cells" (CSCs). The exact mechanism that regulates the maintenance of CSC pool remains largely unknown. Fascin is an actin-bundling protein that we have previously demonstrated to be a major regulator of breast cancer chemoresistance and metastasis, two cardinal features of CSCs. Here, we manipulated fascin expression in breast cancer cell lines and used several in vitro and in vivo approaches to examine the relationship between fascin expression and breast CSCs. Fascin knockdown significantly reduced stem cell-like phenotype (CD44 /CD24 and ALDH ) and reversal of epithelial to mesenchymal transition. Interestingly, expression of the embryonic stem cell transcriptional factors (Oct4, Nanog, Sox2, and Klf4) was significantly reduced when fascin expression was down-regulated. Functionally, fascin-knockdown cells were less competent in forming colonies and tumorspheres, consistent with lower basal self-renewal activity and higher susceptibility to chemotherapy. Fascin effect on CSC chemoresistance and self-renewability was associated with Notch signaling. Activation of Notch induced the relevant downstream targets predominantly in the fascin-positive cells. Limiting-dilution xenotransplantation assay showed higher frequency of tumor-initiating cells in the fascin-positive group. Collectively, our data demonstrated fascin as a critical regulator of breast CSC pool at least partially via activation of the Notch self-renewal signaling pathway and modification of the expression embryonic transcriptional factors. Targeting fascin may halt CSCs and thus presents a novel therapeutic approach for effective treatment of breast cancer. Stem Cells 2016;34:2799-2813 Video Highlight: https://youtu.be/GxS4fJ_Ow-o.

β1 Integrin is essential for fascin‐mediated breast cancer stem cell function and disease progression

“…10 Spontaneous or chemotherapy-mediated cell apoptosis was analyzed using Annexin V staining kit (Life Technologies) as previously described. 10 Spontaneous or chemotherapy-mediated cell apoptosis was analyzed using Annexin V staining kit (Life Technologies) as previously described.…”

“…10 Adhesion assay A plate (96-well) was coated overnight at 4 C with different concentration of ECM substrates including; Matrigel (Corning), 40 μg of Collagen IV (Millipore), 5 μg of Fibronectin (Millipore) and 60 μg of Laminin (Millipore) as previously described. Fascin-positive or -negative cells that express or lack ITGB1 were seeded at 100 cell/well in 3 ml of complete DMEM media for 10-12 days at 37 ο C, 5% CO 2 humidified incubator.…”

“…4,5 Increased cell motility that enhances the metastatic potential of cells was shown to be dependent on active actin cytoskeletal rearrangements. 10 Therefore, a better understanding of the mechanism by which fascin regulates breast CSCs to confer chemoresistance and metastasis will encourage the development of more therapeutic targets and would impact the success rate of treatment. 8 In a subsequent study, we have illustrated that fascin expression in breast cancer cells confers resistance to chemotherapy through activation of the PI3K/Akt signaling pathway.…”

Targeting Signaling Pathways in Cancer Stem Cells for Cancer Treatment

“…For instance, Barnawi et al reported that fascin (an actin-bundling protein) effectively regulates breast CSCs at least partially through Notch pathway [78]. Fascin knockdown significantly reduced breast stem cell-like phenotype (downregulation of stem cell pluripotent genes such as Oct4, Nanog, Sox2, and Klf4), and the cells became less competent in forming colonies and tumorspheres.…”

“…Fascin knockdown significantly reduced breast stem cell-like phenotype (downregulation of stem cell pluripotent genes such as Oct4, Nanog, Sox2, and Klf4), and the cells became less competent in forming colonies and tumorspheres. Conversely, activation of Notch signaling induced the relevant downstream targets predominantly in the fascin-positive cells, and fascin-positive CSCs showed stronger tumorigenesis [78]. In another study, immunohistochemical analysis of 115 breast tumor tissues from primary lesions was performed, and results showed that Notch positive tissues were significantly associated with a CSC marker aldehyde dehydrogenase 1 family member A1 levels [79].…”