Browsing by Author "Prangley, Charne"
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- ItemThe cellular response of triple-negative breast cancer to short-term starvation: implications for chemosensitivity(Stellenbosch : Stellenbosch University, 2021-03) Prangley, Charne; Engelbrecht, Anna-Mart; Davis, Tanja; Stellenbosch University. Faculty of Science. Dept. of Physiological Sciences.ENGLISH ABSTRACT: Introduction: Breast cancer is currently the most common cancer among women globally. Triplenegative breast cancer (TNBC) is an aggressive and often drug-resistant sub-type of breast cancer that is correlated with poor patient outcomes. As a result, adjuvant therapies that may improve drug sensitivity are currently being sought. Due to the unique metabolic hallmarks of cancer, metabolic adjuvant therapies have become an area of increasing interest. We therefore set out to investigate the effect of short-term starvation (STS) on the growth, viability, and metabolism of TNBC cells and a benign breast epithelial cell line. We also investigated the effect of STS on chemotherapyinduced cytotoxicity in these cells to determine whether STS may enhance the effect of doxorubicin in TNBC. Methods: Three cell lines were utilised for this study: a benign breast epithelial cell line (MCF- 12A), and two triple-negative breast cancer cell lines (BT-549 and MDA-MB-231). Western blotting was employed to determine the effect of starvation over time on growth and proliferation signalling pathways (PI3K/Akt) and markers of autophagy (Atg5, p62 and LC3-II). Immunocytochemistry was utilised to quantify autophagic puncta. Cell cycle progression and viability were assessed using flow cytometry and a WST1 assay, respectively. The effect of STS on chemosensitivity was then established by incubating cells in standard or starvation-mimicking media for 24 hours, whereafter they received doxorubicin at a concentration of 2.5 μM. Chemosensitivity was then established in terms of live cell number, cell death and viability, and cell cycle progression. Results and Discussion: In response to STS, the MCF-12A cells downregulated pro-growth signalling pathways, while the MDA-MB-231 cells showed significant upregulation. A 24-hour starvation period had no significant effects on these pathways or on autophagic flux in the BT-549 cells. Both the MCF-12A and MDA-MB-231 cell lines significantly upregulated autophagic flux in response to STS, with the latter achieving the most significant effect at 24-hours. This may have offered protection to these cells, as a period of starvation prior to drug administration reduced doxorubicin-induced G2/M arrest. Additionally, STS had no other significant effects on chemosensitivity in these cells. In the BT-549 cells, however, starvation was able to significantly increase the percentage of dead cells in the group that received STS prior to doxorubicin treatment. As autophagy was not significantly increased in this cell line during starvation, this suggests that autophagy may indeed play a role in drug resistance. Conclusion: In summary, the cell lines which displayed an upregulation of autophagy at 24 hours of starvation were not sensitised to doxorubicin in terms of cell death, and also experienced amelioration of doxorubicin-induced G2/M arrest. This supports the notion that autophagic upregulation may protect cancer cells from doxorubicin-induced cytotoxicity and contribute to drug resistance. However, to gain a more thorough understanding of this phenomenon, future studies investigating the mechanisms by which autophagy promotes chemoprotection are recommended.