The Efficacy and Mechanism of Action of HDAC Inhibitors in Triple Negative Breast Cancer
Histone deacetylase inhibitors (HDACis) are a class of chemically heterogeneous anticancer agents which consist of a hydrophobic capping group, linker region, and Zn 2+ chelating pharmacophore. Manipulation of these three regions, such as the incorporation of a pyridine capping group, and hydroxythiopyridone Zn 2+ binding group has yielded Jazz-90, and M1S and M2S, respectively. Alongside vorinostat, these HDACis were tested for their cytotoxicity and activity in triple negative MDA-MB-231 and MDA-MB-468 breast cancer cells. The novel compound Jazz-90 displayed the greatest cytotoxic potency in MDA-MB-231 cells (EC 50 1.14 μM) while M1S had an EC 50 of 1.66 μM in MDA-MB-468 cells. Time-course cytotoxicity studies showed the compounds to have a cytostatic effect at the EC 50 concentration (1X) and a cytotoxic effect at four-fold the EC 50 concentration (4X). Furthermore, the effect on protein expression was determined using Western blotting at 24 h post-treatment. Relative to their cytotoxicity, the EC 50 of vorinostat (2.10 μM) showed the greatest increase in acetylated histone-H3 protein expression followed by 1X Jazz-90, 1472% and 1041%, respectively, compared to control in MDA-MB-231 cells; a similar trend was seen in MDA- MB-468 cells. Compared to vehicle control, only 4X of M1S (6.66 μM) and M2S (17.17 μM) significantly increased acetylated histone-H3 expression in MDA-MB-468 cells, 11404% and 10795%, respectively. Cell cycle protein analysis revealed that compared to control 1X of vorinostat, M1S, and M2S significantly downregulated cyclin B1 expression in MDA-MB-231 cells by 56%, 65%, and 29%, respectively, suggesting the compounds to induce G 2 /M-phase arrest. Interestingly, 1X of M1S and M2S were found to significantly increase cyclin D1 expression in MDA-MB-231 cells by 122% and 97%, respectively, suggesting that the compounds promote progression through the G 1 -phase. No change in total cyclin E2, Hsp90, or NF-κB p65 protein expression was seen in either cell line, p53 in MDA-MB-231 cells, or nuclear Hsp90 and nuclear NF-κB p65 in MDA-MB-468 cells. Thus, our findings provide evidence of the cytotoxic potency of Jazz-90, M1S, and M2S, but limited HDAC inhibitory effects in TNBC cells in vitro, hence warranting further modulation of the capping and Zn 2+ binding moieties to enhance their HDAC inhibitory profile.
Advisor: Rosengren, Rhonda
Degree Name: Master of Science
Degree Discipline: Pharmacology
Publisher: University of Otago
Keywords: HDAC; TNBC; Triple; Negative; Breast; Cancer; Curcumin; Pharmacology
Research Type: Thesis