Department of Biomedical Sciences, Ontario Veterinary College, University of Guelph, Guelph, Ontario.
Transforming growth factor beta (TGFbeta) is often overexpressed in breast tumors and associates with metastatic disease. TGFbeta’s pro-metastatic role has been linked to its capacity to induce epithelial-to-mesenchymal transition (EMT). Conversely, TGFbeta is also a tumor suppressor in early stage breast tumors and this is mediated, at least in part, by its capacity to promote apoptosis in early- transformed cells. Since TGFbeta’s metastasis-promoting function is the one that predominates in advanced breast cancer, research to date has mainly focused on targeting TGFbeta or its EMT-promoting signaling effectors. Diverging from this classic approach, we are interested in the possibility of rescuing TGFbeta’s tumor suppressor function in tumors, by re-educating the advanced breast cancer cell to respond differently, i.e. by undergoing apoptosis, to the TGFbeta present in the tumor microenvironment.
TGFbeta signals to serine/threonine kinase receptors that activate canonical (Smad) and non-canonical signaling pathways. We have previously identified the non- canonical Par6 pathway as an essential mediator of EMT and metastasis in breast cancer cells. Interestingly, we also found that Par6 activation is necessary for, and cooperates with Smad activation, to induce apoptosis in response to TGFbeta in immortalized mammary cells. Using a combination of monolayer and three- dimensional cultures, immunoblotting and confocal microscopy, here we investigated the effect of Par6 wild type overexpression, which promotes constitutive hyperactivation of Par6, in TGFbeta-induced apoptosis in NMuMG immortalized mammary cells. We found that Par6 overexpression promotes apoptosis in response to TGFbeta under conditions in which these cells are generally resistant to TGFbeta-induced apoptosis, and this effect was associated with downregulation of PI3K/Akt and NFkappa B activation. To investigate whether negative modulation of the PI3K/Akt axis could be a mechanism involved in the positive impact of Par6 overactivation on TGFbeta-induced apoptosis, we used pharmacologic inhibitors of PI3K activity on both immortalized and transformed, metastatic mammary cells. We found that PI3K inhibition synergizes with TGFbeta in promoting apoptosis, but does so more strikingly under conditions that favor Par6 activation. Taken together these results suggest that PI3K inhibition could be a strategy to enhance apoptosis in advanced mammary tumors, particularly those that overexpress Par6 and display active TGFbeta signaling.