Ambrosio, Maria Rosaria (2015) Glucose-induced gene expression changes in breast cancer cells: a putative role in Tamoxifen responsiveness. [Tesi di dottorato]

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Diabetes is one of the most challenging and growing health problems worldwide. Diabetes (mainly type 2; T2D) is an independent risk factor for development of several types of cancer. A great number of epidemiological studies have reported that T2D patients have an increased risk of breast cancer, and up to 16 % of breast cancer patients suffer from T2D or impaired glucose tolerance. The presence of T2D, as cancer-associated comorbidity, is linked to poorer prognosis and survival in breast cancer post-menopausal women. Hyperglycemia, the most important feature of T2D, is known to affect breast cancer cell proliferation. Little is known about the contribution of glucose on breast cancer cell drug resistance. Besides acting on cancer cells, glucose may affect surrounding cells as well as distant cells, which in turn may interfere with anti-cancer drug response. Breast cancer cell growth and/or metastasis predominantly occur as a result of the adipocyte-rich microenvironment in which breast cancer cells are embedded, and reflect a role for adipocytes in tumour maintenance and progression. Adipocytes secrete a variety of adipokines and proinflammatory cytokines that may contribute to drug resistance. Therefore, the aim of this work is to investigate whether and how hyperglycaemia and adipose-derived factors may affect cellular response of breast cancer cell to Tamoxifen, an antagonist of the estrogen receptor (ER), widely used as hormonal treatment of estrogen dependent breast cancer. It was observed that Tamoxifen reduced MCF7 breast cancer (ER+) cell viability by about 50% when cells were cultured in glucose concentration corresponding to normal fasting glucose levels in humans (5.5 mM; Low Glucose-LG). Conversely, MCF7 cell sensitivity to Tamoxifen of was 2-fold reduced when cells were cultured in glucose concentration resembling hyperglycaemia in humans (25 mM; High Glucose-HG). Interestingly, shifting MCF7 cells from HG to LG medium restored their drug sensitivity, whereas the shift from LG to HG medium reduced their responsiveness to Tamoxifen. In addition, MCF7 cell response to Tamoxifen in HG medium was worsened in presence of adipocyte-released factors. Notably, RNA-Sequencing revealed that glucose significantly deregulated gene expression and that 70 cell cycle-related genes were significantly down-regulated when MCF7 cells were shifted from HG to LG medium. Among those genes, CTGF and CYR61 were significantly reduced. Consistently, experimental evidences showed that both genes were overexpressed in MCF7 exposed to HG concentrations or to adipocytes released factors in HG concentrations. On the opposite, CTGF and CYR61 gene expression was reduced in MCF7 cells shifted from HG to LG concentrations. Interestingly, CTGF, while not CYR61, gene silencing significantly increased Tamoxifen sensitivity of MCF7 cells in HG medium. Hence, CTGF may be a novel diabetes associated predictive marker for chemo-sensitivity and may represent a potential therapeutic target to overcome Tamoxifen resistance, improving survival of T2D patients affected by breast cancer.

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