Abstract:
2-Methoxyestradiol (2ME), a 17β-estradiol metabolite, exerts anticancer properties
however, the compound was found to possess low bioavailability. This resulted in the
in silico-design of 2ME analogues with a sulphamoyl moiety which made them more
potent than the parent compound. Sulphamoylated 2ME analogues are suspected to
induce the antitumourigenic effects through the induction of reactive oxygen species.
However, the exact role of oxidative stress in the activity exerted by these
compounds remains elusive.
In the current study, 2-ethyl-13-methyl-17-oxo-7,8,9,11,12,13,14,15,16,17-
decahydro-6-cyclopenta[a]phenanthrane-3 sulphamate (ESE-one) was chosen as a
sulphamoylated estradiol analogue representative to investigate the role of reactive
oxygen species (ROS) in the effects exerted by these sulphamoylated compounds
on cell proliferation, morphology, cell cycle progression, antioxidant activity and
mitochondrial membrane potential in estrogen receptor positive breast epithelial
adenocarcinoma (MCF-7) cells and estrogen receptor negative breast epithelial
adenocarcinoma (MDA-MB-231) cells.
Fluorescent microscopy data revealed that sulphamoylated estradiol analogues
induced more ROS production compared to their non-sulphamoylated counterparts
in both MCF-7- and MDA-MB-231 cells. Crystal violet staining demonstrated a
significant growth inhibition in cells exposed to sulphamoylated estradiol analogues
compared to cells exposed to the non-sulphamoylated compounds. ESE-one
exposure resulted in a ROS-dependent growth inhibition which was repressed by
tiron (superoxide inhibitor), trolox (peroxyl inhibitor) and DMTU (hydrogen peroxide
inhibitor). ESE-one exposure to MCF-7- and MDA-MB-231 cells resulted in an
accumulation of cells in G2/M phase after 24 hours and sub-G1 phase after 48 hours.
The effect induced after 24 hours exposure was inhibited by tiron and trolox, and that
induced after 48 hours exposure was inhibited by tiron, trolox and DMTU.
Proliferation data was confirmed by morphology studies. Tiron, trolox and DMTU significantly decreased the number of rounded cells,
shrunken cells and apoptotic bodies in MCF-7 and MDA-MB-231 cells induced by
ESE-one exposure; cell density was recuperated indicating the rescue effects of
ROS inhibitors. Antioxidant activity data demonstrated that ESE-one induced cell
rounding and antiproliferative effects via ROS evident in the reduced catalase protein
concentration in MCF-7 cells which was opposed by tiron and DMTU and in MDAMB-
231 cells, inhibited by tiron and trolox. Reduction in mitochondrial membrane
potential was inhibited by tiron in MCF-7 cells and DMTU in MDA-MB-231 cells.
This in vitro study suggests that ESE-one induces growth inhibition, cell rounding,
cell cycle arrest, catalase inhibition and depolarization of the mitochondrial
membrane by production of superoxide anion, peroxyl radical and hydrogen peroxide
which culminates in apoptosis. This study contributes to targeted therapy based on
ROS-dependent cell death pathways in tumourigenic breast cells.
