The effects of inhibiting the notch signalling pathway in triple negative breast cancer cell lines

Msibi, Thandiwe
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The molecular pathology of triple negative breast cancer (TNBC) is poorly understood, consequently, no successful forms of therapy have been developed. Thus an improvement of knowledge and subsequently the discovery of novel treatments for the disease are imperative. It has been found that deregulation of the Notch signalling pathway promotes tumourigenesis in breast tissue. Therefore, it was of interest here to investigate whether the Notch signalling pathway is deregulated in TNBC and whether its abrogation affects the proliferation and migration of TNBC cell lines. The normal growth characteristics of MCF-7 cells (hormone sensitive) were compared to those of MDA-MB-231 and MDA-MB-436 (both hormone insensitive) cell lines and were determined by real-time cell impedance assays, using the ―xCELLigence‖ instrument. Thereafter, cells were treated with gamma secretase inhibitors (GSI) of the Notch signalling pathway. The MCF-7 cell line proliferated faster than the MDA-MB-231 and MDA-MB-436 cell lines. The proliferation of the MDA-MB-231 and MDA-MB-436 cell lines decreased significantly following treatment with inhibitors. Confocal microscopy was used to assess levels of the Notch intracellular component (a gamma secretase cleavage product) and E-cadherin (a breast tumour suppressor marker), pre- and post- treatment. Prior to drug treatments, confocal microscopy showed that the Notch intracellular component was highly expressed in the MDA-MB-231 cell line, and it was low in the MDA-MB-436 cell line, compared to the MCF-7 cell line. Following drug treatments, confocal microscopy showed a decreased expression of the Notch intracellular component in all three cell lines. Prior to drug treatment only the MCF-7 cell line expressed E-cadherin which was reduced post treatment. Subsequently the cell migration assays revealed that migration is reduced post- drug treatment in all three cell lines, despite no statistical significance. Overall the MDA-MB-231 and MDA-MB-436 cell lines were more significantly sensitive to the gamma secretase inhibitors compared to the MCF-7 cell lines. Therefore these observations suggest that the Notch signalling pathway is a plausible novel therapeutic target in the treatment of TNBC.
A dissertation submitted to the Faculty of Health Sciences, School of Clinical Medicine Department of Internal Medicine, University of The Witwatersrand, Johannesburg in fulfilment of the Degree of Masters of Science in Medicine Johannesburg 2015