Tumour microenvironment factors and the promotion of epithelial to mesenchymal transition in MDA-MB-468 breast cancer cells [Poster]

Introduction: Epithelial to mesenchymal transition (EMT) in breast cancer cells involves the acquisition of a more invasive phenotype and is believed to be important in the development of metastasis. EMT results in the upregulation of mesenchymal cell markers including vimentin, N-cadherin, Twist, and the ratio of the stem cell markers CD44/CD24. Inducers of EMT include growth factors and conditions present in the tumour microenvironment such as epidermal growth factor (EGF). A recent study using conditioned media from wounded MDA-MB-468 breast cancer cells suggests that some tumour microenvironment factors may augment EGF- mediated increases in vimentin protein expression in MDA-MB-468 cells. However, effects on other EMT markers are unknown.

Aims: To determine whether conditioned media from wounded MDA-MB-468 breast cancer cells increases the sensitivity of MDA-MB-468 breast cancer cells to EGF-induced changes in EMT markers.

Methods: MDA-MB-468 cells were plated in 96 well plates and allowed to attach overnight. Cells were serum starved (24 h) before treatment with EGF (0 - 50 ng/mL) in the presence of conditioned media (from a wounded monolayer of MDA-MB-468 cells) or control media (from an intact monolayer of MDA-MB-468 cells). RNA was isolated 24 h after cell treatments. Real-time RT-PCR was used to compare vimentin, N-cadherin, Twist and CD44/CD24 mRNA levels in all samples.

Results: EGF treatment of MDA-MB-468 cells produced increases in the mRNA levels of the EMT markers vimentin, N-cadherin and Twist, and resulted in an increase in the CD44/CD24 mRNA ratio. EGF-mediated increases in vimentin mRNA levels appeared to be promoted by wound-conditioned media at submaximal concentrations of EGF (3 ng/mL). Some markers of EMT appeared to be insensitive to this induction (e.g. N- cadherin).

Discussion: These studies suggest that some markers of EMT may be differentially sensitive to agents that promote EGF-induced EMT in MDA-MB-468 breast cancer cells.