Abstract:
BACKGROUND : The estrogen metabolite 2-methoxyestradiol (2ME2) and a number of synthesised derivatives have
been shown to bind to microtubules thereby arresting cancer cells in mitosis which leads to apoptosis. In interphase
cells, microtubules play an important role in the delivery of proteins to subcellular locations including the focal adhesions.
In fact, focal adhesion dynamics and cell migration are in part regulated by microtubules. We hypothesised that
novel 2ME2 derivatives can alter cell migration by influencing microtubule dynamics in interphase cells. In this report
we describe 2ME2 derivatives that display anti-migratory capabilities in a metastatic breast cancer cell line through
their effects on the microtubule network resulting in altered focal adhesion signalling and RhoA activity.
METHODS : Cell migration was assayed using wound healing assays. To eliminate mitosis blockage and cell rounding
as a confounding factor cell migration was also assessed in interphase blocked cells. Fluorescence confocal microscopy
was used to visualise microtubule dynamics and actin cytoskeleton organisation while western blot analysis was
performed to analyse focal adhesion signalling and RhoA activation.
RESULTS : 2ME2 derivatives, ESE-one and ESE-15-one, inhibited cell migration in cycling cells as expected but equally
diminished migration in cells blocked in interphase. While no significant effects were observed on the actin cytoskeleton,
focal adhesion kinase activity was increased while RhoA GTPase activity was inhibited after exposure to either
compound. Microtubule stability was increased as evidenced by the increased length and number of detyrosinated
microtubules while at the same time clear disorganisation of the normal radial microtubule organisation was
observed including multiple foci.
CONCLUSIONS : ESE-15-one and ESE-one are potent migration inhibitors of metastatic breast cancer cells. This ability is
coupled to alterations in focal adhesion signalling but more importantly is associated with severe disorganisation of
microtubule dynamics and polarity. Therefore, these compounds may offer potential as anti-metastatic therapies.
