Abstract LB-252: Synergy of epithelial and mesenchymal cells under anoikis-inducing conditions.
Authors: Grosse-Wilde Anne , Ertaylan Gokhan , West Robert , Walters Kathie , Ozinsky Adrian 
Published: August 2014

Tumor metastasis is the ultimate cause for death of cancer patients. In epithelial tumors such as breast cancer the early steps of the metastatic cascade require tumor cells to migrate, invade and survive detachment from the extracellular matrix in suspension which in epithelial (E) cells results in cell death (anoikis). Therefore, all of these earlier steps appear to be accomplished by mesenchymal (M) cells. In a process called epithelial to mesenchymal transition (EMT) E tumor cells can become M. However, a central paradox of this model is that in most patients the metastatic tumor has an E morphology just as the primary tumor. Analogous to primary tumors and their corresponding metastases, in transformed and immortalized mammary epithelial HMLER cells we frequently found E cells in cultures of replated derivatives of HMLER cells grown in suspension, as well as in suspended mammospheres. In order to disentangle the role of E and M cells under suspension culture conditions in the heterogeneously E and M HMLER cells, we generated mammospheres of HMLER cells, and of isolated E or M HMLER clones. As expected, only the heterogeneous HMLER cells and the M clones were able to form mammospheres, and in gene expression arrays we detected a significant upregulation of genes encoding the pluripotency factor OCT4 and its targets. Importantly, the M cell clone significantly upregulated E genes during suspension conditions. Intriguingly, amongst those were many secreted growth factors and inflammatory cytokines such as SLPI, IL1B, MMP7, S100 proteins and CXCL1, several of which have previously been described to enhance aggressive cancer growth and suppress apoptosis at the metastatic site. In order to directly test the effect of E cells on M cells in suspended mammospheres we generated suspension cultures of isolated E and M cells or of E and M co-cultures. We found that i) E cells can survive in suspension only in conjunction with M cells, and ii) co-culture of E and M cells in suspension resulted in significantly larger and higher number of mammospheres than culture of isolated M cells. Moreover, treatment of M cells with supernatant of E cells could mimic the coculture effect of increased and enlarged mammosphere formation, suggesting that soluble E factors are responsible for enhanced proliferation of M HMLER cells in suspension. In summary, in stem cell-enriched mammospheres of M cells we found high induction of secreted growth and survival factors which are constitutively high expressed in adherent as well as in suspended E cells.

Indeed, co-culture of suspended E and M cells resulted in a synergistic effect of enhanced proliferation of M cells in mammospheres and survival of E cells. Synergy of E and M cells in suspension can explain how epithelial cells could be retained under anoikis-inducing conditions in the vasculature and why metastatic tumors are E like their primary counterparts.

Citation Format: Anne Grosse-Wilde, Gokhan Ertaylan, Robert West, Kathie Walters, Adrian Ozinsky. Synergy of epithelial and mesenchymal cells under anoikis-inducing conditions. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-252. doi:10.1158/1538-7445.AM2013-LB-252