Abstract:
INTRODUCTION : Research involving antimitotic compounds identified 2-methoxyestradiol (2ME2), as a promising anticancer
endogenous metabolite. Owing to its low bioavailability, several in silico-designed 2ME2 analogues were synthesized.
Structure-activity relationship studies indicated that an already existing 17-β-estradiol analogue, namely
(8R,13S,14S,17S)-2-ethyl-13-methyl-7,8,9,11,12,13,14,15,16,17-decahydro-6H-cyclopenta[a]phenanthrane-3,17-diyl
bis(sulphamate) (EMBS) to exert potential in vitro anticancer activity.
METHODS : This study investigated the in vitro apoptotic influence of EMBS in an estrogen receptor-positive breast
adenocarcinoma epithelial cell line (MCF-7); an estrogen receptor-negative breast epithelial cell line (MDA-MB-231) and a
non-tumorigenic breast cell line (MCF-12A). Cell cycle progression, a phosphatidylserine flip, caspase 6-, 7- and 8 enzyme
activity levels, Bcl-2 phosphorylation status at serine 70 and Bcl-2- and p53 protein levels were investigated to identify a
possible action mechanism for apoptotic induction.
RESULTS : The xCELLigence real-time label-independent approach revealed that EMBS exerted antiproliferative activity in all
three cell lines after 24 h of exposure. A G2M block was observed and apoptosis induction was verified by means of flow
cytometry using propidium iodide and Annexin V-FITC respectively. EMBS-treated cells demonstrated a reduced
mitochondrial membrane potential. EMBS exposure resulted in a statistically significant increase in p53 protein
expression, decreased Bcl-2 protein expression and a decrease in pBcl-2(s70) phosphorylation status in all three
cell lines. Results support the notion that EMBS induces apoptosis in all three cell lines.
CONCLUSION : This study includes investigation into the apoptotic hallmarks exerted by EMBS after exposure of
three cell lines namely MCF-7-, MDA-MDA-231- and MCF-12A cells. Increased caspase 6-, caspase 7- and caspase
8 activities, upregulation of p53 protein expression and a decrease in phosphorylation status of Bcl-2 at serine 70
in tumorigenic and non-tumorigenic lines were demonstrated.
