Strategies to Investigate the Role of EpCAM in Cancer Stem Cells of Breast Cancer Cell Lines

Abstract

<p>Based on the cancer stem cell (CSC) model, tumors develop as a hierarchy, much like normal tissue. At the apex, CSCs are capable of self-renewal and differentiating into non-tumorigenic cells that form the bulk of the tumor. CSCs have been isolated for many types of cancer based on cell surface marker expression. For instance in human breast carcinoma, the CD44+ CD24-/low EpCAM+ population are enriched in CSCs. These cells are more resistant to traditional chemo and radiation therapy relative to the bulk of the tumor. As such, many believe CSCs to be responsible for relapse and metastasis events. Hence, a more targeted therapy towards breast CSCs can prove to be very effective. EpCAM has been shown to be a reliable, albeit not exclusive, marker of breast CSCs. A more thorough understanding of EpCAM’s function can provide new angles for designing therapeutic agents. Originally thought to be a mere adhesion molecule, EpCAM is now known to derive a signalling pathway that promotes transcription in its target genes. We aimed to provide further insight into this novel pathway, by manipulating EpCAM’s expression and function in MCF7 breast cancer cell line. We designed a dominant negative allele of the EpCAM protein to hinder the activity of the endogenous EpCAM. However, this transgene offered no significant effect on our cell line. Furthermore, we used RNA interference technology to reduce EpCAM expression. The introduction of the EpCAM short hairpin RNA constructs into our cell line had inhibitory effects on transcription reporter expression, cellular growth, adherent colony and unattached sphere formation. The inhibition of EpCAM in MCF7 spheres was followed by signs of cell death, differentiation and an epithelial mesenchymal transition process. Sphere cultures are used because they are enriched in cancer stem cells. The adverse effects of EpCAM inhibition in MCF7 spheres provides further evidence as to the role of this protein in cancer stem cells.</p>