DNA methylation profiling of breast cancer cell lines along the epithelial mesenchymal spectrum - implications for the choice of circulating tumour DNA methylation markers

Epithelial-mesenchymal plasticity (EMP) is a dynamic process whereby epithelial carcinoma cells reversibly acquire morphological and invasive characteristics typical of mesenchymal cells, which facilitates metastasis. Understanding the methylation differences between epithelial and mesenchymal states may assist in the identification of optimal DNA methylation biomarkers for the blood-based monitoring of cancer. Methylation-sensitive high-resolution melting (MS-HRM) was used to examine the promoter methylation status of a panel of established and novel markers in a range of breast cancer cell lines spanning the epithelial-mesenchymal spectrum. Pyrosequencing was used to validate the MSHRM results. The results indicate an overall distinction in methylation between epithelial and mesenchymal phenotypes. The mesenchymal expression markers VIM, DKK3 and CRABP1 were methylated in the majority of epithelial breast cancer cell lines while methylation of the epithelial expression markers GRHL2, MIR200C and CDH1 was restricted to mesenchymal cell lines. We also examined EMP association of several methylation markers that have been used to assess minimal residual disease. Markers such as AKR1B1 and APC methylation proved to be selective for epithelial breast cell lines, however RASSF1A, RAR{beta}, TWIST1 and SFRP2 methylation was seen in both epithelial and mesenchymal cell lines, supporting their suitability for a multi-marker panel.