scholarly journals Downregulation of STAT3, β-Catenin, and Notch-1 by Single and Combinations of siRNA Treatment Enhance Chemosensitivity of Wild Type and Doxorubicin Resistant MCF7 Breast Cancer Cells to Doxorubicin

2019 ◽  
Vol 20 (15) ◽  
pp. 3696 ◽  
Author(s):  
Walhan Alshaer ◽  
Dana A. Alqudah ◽  
Suha Wehaibi ◽  
Duaa Abuarqoub ◽  
Malek Zihlif ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Signaling Pathways ◽  
Breast Cancer Cells ◽  
Therapeutic Potential ◽  
Therapeutic Modality ◽  
Wild Type ◽  
Notch 1 ◽  
Sirna Treatment ◽  
Antitumor Chemotherapy

Combinatorial therapeutic strategies using siRNA and small molecules to eradicate tumors are emerging. Targeting multiple signaling pathways decreases the chances of cancer cells switching and adapting new signaling processes that may occur when using a single therapeutic modality. Aberrant functioning of Notch-1, Wnt/β-catenin, and STAT3 proteins and their crosstalk signaling pathways have been found to be involved in tumor survival, drug resistance, and relapse. In the current study, we describe a therapeutic potential of single and combinations of siRNA designed for silencing Notch-1, Wnt/β-catenin, and STAT3 in MCF7_DoxS (wild type) and MCF7_DoxR (doxorubicin resistant) breast cancer cells. The MCF7_DoxR cells were developed through treatment with a gradual increase in doxorubicin concentration, the expression of targeted genes was investigated, and the expression profiling of CD44/CD24 of the MCF7_DoxS and MCF7_DoxR cells were detected by flow cytometry. Both MCF7_DoxS and MCF7_DoxR breast cancer cells were treated with single and combinations of siRNA to investigate synergism and were analyzed for their effect on cell proliferation with and without doxorubicin treatment. The finding of this study showed the overexpression of targeted genes and the enrichment of the CD44−/CD24+ phenotype in MCF7_DoxR cells when compared to MCF7_DoxS cells. In both cell lines, the gene silencing efficacy showed a synergistic effect when combining STAT3/Notch-1 and STAT3/Notch-1/β-catenin siRNA. Interestingly, the chemosensitivity of MCF7_DoxS and MCF7_DoxR cells to doxorubicin was increased when combined with siRNA treatment. Our study shows the possibility of using single and combinations of siRNA to enhance the chemosensitivity of cancer cells to conventional antitumor chemotherapy.

scholarly journals Role of Sialyl-O-Acetyltransferase CASD1 on GD2 Ganglioside O-Acetylation in Breast Cancer Cells

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1468
Author(s):  
Sumeyye Cavdarli ◽  
Larissa Schröter ◽  
Malena Albers ◽  
Anna-Maria Baumann ◽  
Dorothée Vicogne ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Cancer Cell Line ◽  
Wild Type ◽  
Cellular Models ◽  
Promising Therapeutic Target ◽  
Plasmid Transfection ◽  
Ganglioside Species

The O-acetylated form of GD2, almost exclusively expressed in cancerous tissues, is considered to be a promising therapeutic target for neuroectoderm-derived tumors, especially for breast cancer. Our recent data have shown that 9-O-acetylated GD2 (9-OAcGD2) is the major O-acetylated ganglioside species in breast cancer cells. In 2015, Baumann et al. proposed that Cas 1 domain containing 1 (CASD1), which is the only known human sialyl-O-acetyltransferase, plays a role in GD3 O-acetylation. However, the mechanisms of ganglioside O-acetylation remain poorly understood. The aim of this study was to determine the involvement of CASD1 in GD2 O-acetylation in breast cancer. The role of CASD1 in OAcGD2 synthesis was first demonstrated using wild type CHO and CHOΔCasd1 cells as cellular models. Overexpression using plasmid transfection and siRNA strategies was used to modulate CASD1 expression in SUM159PT breast cancer cell line. Our results showed that OAcGD2 expression was reduced in SUM159PT that was transiently depleted for CASD1 expression. Additionally, OAcGD2 expression was increased in SUM159PT cells transiently overexpressing CASD1. The modulation of CASD1 expression using transient transfection strategies provided interesting insights into the role of CASD1 in OAcGD2 and OAcGD3 biosynthesis, and it highlights the importance of further studies on O-acetylation mechanisms.

scholarly journals Glycosylation Modulates Plasma Membrane Trafficking of CD24 in Breast Cancer Cells

2021 ◽  
Vol 22 (15) ◽  
pp. 8165
Author(s):  
Amanda Chantziou ◽  
Kostas Theodorakis ◽  
Hara Polioudaki ◽  
Eelco de Bree ◽  
Marilena Kampa ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Plasma Membrane ◽  
Subcellular Localization ◽  
Cancer Cells ◽  
Membrane Trafficking ◽  
Breast Cancer Cells ◽  
Endocytic Pathway ◽  
Cell Periphery ◽  
Wild Type ◽  
Mcf 7

In breast cancer, expression of Cluster of Differentiation 24 (CD24), a small GPI-anchored glycoprotein at the cell periphery, is associated with metastasis and immune escape, while its absence is associated with tumor-initiating capacity. Since the mechanism of CD24 sorting is unknown, we investigated the role of glycosylation in the subcellular localization of CD24. Expression and localization of wild type N36- and/or N52-mutated CD24 were analyzed using immunofluorescence in luminal (MCF-7) and basal B (MDA-MB-231 and Hs578T) breast cancer cells lines, as well as HEK293T cells. Endogenous and exogenously expressed wild type and mutated CD24 were found localized at the plasma membrane and the cytoplasm, but not the nucleoplasm. The cell lines showed different kinetics for the sorting of CD24 through the secretory/endocytic pathway. N-glycosylation, especially at N52, and its processing in the Golgi were critical for the sorting and expression of CD24 at the plasma membrane of HEK293T and basal B type cells, but not of MCF-7 cells. In conclusion, our study highlights the contribution of N-glycosylation for the subcellular localization of CD24. Aberrant N-glycosylation at N52 of CD24 could account for the lack of CD24 expression at the cell surface of basal B breast cancer cells.

TK1 in Cancer Progression: elucidating its influence on cellular invasion and survival

2019 ◽  
Author(s):  
Eliza E. Bitter ◽  
Michelle H. Townsend ◽  
Kary Y.F. Tsai ◽  
Carolyn I. Allen ◽  
Rachel I. Erickson ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Survival ◽  
Cancer Cells ◽  
Cell Lines ◽  
Cancer Progression ◽  
Breast Cancer Cells ◽  
S Phase ◽  
Wild Type ◽  
Cellular Invasion ◽  
Cancer Pathogenesis

Abstract 1. Background: The salvage pathway enzyme thymidine kinase 1 (TK1) is elevated in the serum of several different cancer types and higher expression is associated with more aggressive tumor grade. As a result, it has potential as a biomarker for diagnosis and prognosis. Recent studies indicate that TK1 may be involved in cancer pathogenesis; however, its direct involvement has not been identified. We propose to evaluate the effects of TK1 on cancer progression in vitro through measuring cellular invasion and survival of breast cancer cells.2.Methods: Breast cancer cells MDA-MB-231, HCC 1806, and MCF7 were cultured according to standard techniques. We employed the use of TK1 target siRNA and a CRISPR-Cas9 TK1 knockout plasmid to compare transfected cell lines to wild type cell lines. Protein factors in survival and invasive pathways were also tested for correlations to TK1 in BRCA RNA-seq patient data (n=1095) using the TIMER program. Cellular invasion was quantified in cell index (factor of impedance) over a 24-hour period. Cell survival was measured by apoptosis under metabolic and DNA stress using flow cytometry. All results were statistically assessed using an ANOVA or t-test in GraphPad PRISM®.3.Results: Cellular invasion assays assessing wild type and TK1 knockdown/knockout (TK1-/-) cell types showed TK1-/- cell lines had increased invasion potential (p= 0.0001). Bioinformatically, we saw a strong overall negative correlation between apoptotic factors and TK1 (p ≤ 0.05). When testing TK1 effects on cell survival we saw a protective affect under DNA stress (p ≤ 0.05), but not under metabolic stress (p= 0.0001).4.Conclusion From cell cycle analysis, we observed a shift towards S phase in TK1-/- cells. This shift to S phase would promote growth and account for the increased cellular invasion and decrease in metabolic induced stress in TK1-/- cells. We propose that cancer cells still may elicit a cancer progressive phenotype based on effects of TK1, but that a system which isolates TK1 is not effective to understand the effects. Instead, identifying protein networks inclusive of TK1 will help to elucidate its effects on cancer progression.

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