Phytoestrogen coumestrol as an anti-cancer therapy against triple-negative inflammatory breast cancer
Rodriguez-Martir, Keishla M
AdvisorPeterson-Peguero, Esther A
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Coumestrol (Cou) is a phytoestrogen present in soy and clover sprouts which is structurally similar to estrogen (E2) and has a cytotoxic effect in breast cancer cell lines. Nevertheless, the molecular mechanism by which Cou can exert its effect in the most aggressive breast cancer subtype, triple-negative inflammatory breast cancer (TN-IBC), is still unknown. Studies show that TN-IBC cell lines are unresponsive to hormonal therapies, but E2 can activate signaling pathways involved in pro-oncogenic phenotypes such as motility and invasion by activating a rapid estrogen-dependent non-genomic signaling. Therefore, the objective of this study was to determine the anti-cancer effect of Cou in TN-IBC cell lines (using 2D and 3D culture models) and the molecular mechanism Cou exerts for its anti-cancer activity. Dose-response curves of Cou in 2D and 3D culture models of TN-IBC cell lines were generated to determine the half-maximal inhibitory concentration (IC50). Relative cell viability was measured in ER-positive, triple-negative, TN-IBC, and HER2-amplified IBC cell lines. Functional assays were performed in TN-IBC cell lines to determine the effect of E2, Cou, and E2/Cou combination treatments on cell viability, migration, invasion, proliferation, and tumor emboli growth. Additionally, the effect of Cou and E2 treatments on phosphorylation of downstream kinases was analyzed by a proteome profile human phospho-kinase array. RNA-seq was performed in TN-IBC cell lines to identify transcriptome changes after Cou treatment. Our data demonstrated that Cou treatment using the IC50 (13µM in 2D models and 50µM in 3D models) decreased cell viability in TN-IBC cell lines. Similarly, Cou treatment reduces triple-negative non-IBC cell viability. In comparison with E2, Cou decreases migration, invasion, proliferation, and tumor emboli growth in TN-IBC cell lines. Finally, Cou treatment reduces the phosphorylation of kinases MAPK/ERK and PI3K/AKT that promote pro-oncogenic phenotypes and upregulate genes such as TIPARP, recently involved in breast cancer phenotype suppression. In summary, this study guide us in elucidating a signaling pathway affected by Cou’s anti-cancer activity. More importantly, it opens the opportunities to design more effective targeted therapeutic strategies to improve the prognosis and survival rates of IBC patients.