Browsing by Author "Cartwright, Meghan Carni"
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- ItemProgestins and breast cancer : significance of progesterone receptor isoforms and their altered ratios(Stellenbosch : Stellenbosch University, 2021-03) Cartwright, Meghan Carni; Africander, Donita; Louw-du Toit, Renate; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.ENGLISH ABSTRACT: Progestins used in menopausal hormonal therapy have been associated with increased incidence of breast cancer. While these synthetic ligands were designed, in four consecutive generations, to mimic the activity of natural progesterone (P4) via the progesterone receptor (PR), the precise mechanism whereby some progestins and/or their metabolites may cause an increase in breast cancer incidence is still mostly unknown. Whether the PR, existing as two isoforms, PR-A and PR-B, plays a role in mediating the effects of progestins on breast cancer is unclear. As the metabolism of a progestin can ultimately influence effects via the PR, ultrahigh performance supercritical fluid chromatography-tandem mass spectrometry was used to investigate the metabolism of P4 and selected progestins in three breast cancer cell lines in the first part of this thesis. Unlike P4 that was rapidly metabolised in all three cell lines, promegestone (R5020), gestodene (GES) and nomegestrol acetate (NOMAC) were not metabolised, while only drospirenone (DRSP) was metabolised in the MDA-MB-231 and T47D cells. Additionally, we showed that P4 metabolism occurred at a similar rate in the MDAMB- 231 and T47D cells, but faster than its metabolism in the MCF-7 BUS cells. In the second part of this study, transactivation and transrepression transcriptional assays showed that the activities of a selected panel of progestins from all four generations are not all similar to each other, P4 or R5020, via PR-A and PR-B. For transactivation, most progestins were more efficacious via PR-B, but more potent via PR-A. We also showed that an increase in PR-A density and excess PR-A relative to PR-B, resulted in decreased efficacies of all progestins for transactivation. While an increase in PR-A density resulted in an increase in the activity of all progestins for transrepression, the activity of only a few progestins were influenced by excess expression of PR-A relative to PR-B. Realtime PCR showed progestin- and gene-specific regulation of endogenous genes known to play a role in breast cancer in T47D breast cancer cells. While the response of some progestins on the selected genes were PR-B mediated, some progestin effects were not mediated by either PR-A or PR-B. In the third part of this thesis, investigations into the effects of the progestins on proliferation, apoptosis, anchorageindependent growth, migration and invasion showed that these processes are differentially influenced by P4 and the selected progestins, and that the responses are also differentially mediated by PR-A or PR-B. Excess expression of PR-A resulted in both positive and/or negative ligand-independent, as well as progestin-induced, effects on these cancer hallmarks. Taken together, the findings of this thesis emphasize the fact that progestins do not always mimic the activities of P4 or each other. The results further highlight the complexity of progestin action via the PR, underscoring the importance of distinguishing progestin activities via PR-A and PR-B, and also considering the PR-A:PR-B ratio when investigating the mechanisms of progestins and the PR in breast cancer. Finally, our results suggest that a progestin such as medroxyprogesterone acetate (MPA) acting via PR-A and/or PR-B may indeed increase breast cancer risk, while others like DRSP may not.