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CRISPR/cas9-mediated Generation of an EMT Reporter Cell Line for Metastatic Breast Cancer Drug Discovery and Development

Blue and red mcf-7 human breast cells.

AACR Annual Meeting 2019

Atlanta, Georgia, United States

March 30, 2019


Among women, breast cancer continues to be the most common cancer, with metastasis being the leading cause of mortality in patients around the world. Epithelial to mesenchymal transition (EMT)—the process by which epithelial cells shift to the mesenchymal state—has been implicated in many aspects of breast cancer tumorigenesis, metastasis, and drug resistance. However, despite the extensive accumulation of data on the association of EMT with cancer over the years, EMT has not been an active target for therapeutic development. This is due in part to the lack of appropriate in vitro models. Here, we have exploited some of the basic biology of EMT to create an advanced in vitro metastatic breast cancer reporter cell line model for use in basic research and the discovery of new EMT inhibitors.

During EMT, E-cadherin (ECAD) protein expression is downregulated in cancer cells in association with the loss of cell-to-cell adhesion and apico-basal polarity and changes to spindle-shaped morphology. By installing an emerald green fluorescent protein (EmGFP) tag on the C-terminus of the ECAD gene in the epithelial BT-474 (ATCC HTB-20) breast cancer cell line via CRISPR/Cas9 genome-editing technology, end-point or real-time EMT status of cells can be tracked under defined conditions. The EMT reporter cell line was verified at the nucleic acid (genomic and mRNA) and protein levels as well as in cell-based assays. Functional evaluation of the BT-474 ECAD EmGFP cell line can be monitored by both EmGFP expression and the invasive capacity of the cells. Our results demonstrate that these cells respond to EMT induction by showing decreased ECAD EmGFP expression along with an increased invasive capacity. Further, this EMT reporter cell line shows sensitivity to the MEK1/2 inhibitor U0126, thereby providing the basis for the use of this cell line in high-throughput screening (HTS) applications such as the identification of new anti-EMT drugs for metastatic breast cancer. Further, the BT-474 ECAD EmGFP reporter cell line is also a convenient and sensitive model for basic science research on the mechanisms of metastasis.

Download the poster to explore the development of an advanced in vitro metastatic breast cancer reporter cell line model.


Watch the poster presentation