Received July 30, 2018; Accepted November 11, 2018; Published December 15, 2018
http://dx.doi.org/10.18081/2333-5106/018-349-361
Yu-Chun Lin; Dong-Qing Chin; Eva Wang; Hao Cheng
Abstract
The overall objective of this study is to determine whether EGFR suppression and Erk1/2 disruption limit brain metastasis and suppress invasiveness of MDA-MB 231 and T47D cells, switch their phenotype from mesenchymal to epithelial and EMT, and have no negative side effects. The first task was to create stable MDA-MB 231 cells with suppression of the EGFR gene (G33M.231, G29M.231) and MCF-10A mammary epithelial cells (G16M) with EGFR suppression, for comparison of cell proliferation and survival, clonogeny, tumorigenicity, phenotypic and aggressiveness characteristics of cells, adhesion properties to the endothelium. Cells of the panel were genotyped by PCR to check the homozygosity of the modified alleles. Protein expression was verified by immunoblotting. Next, the invasive and adhesive properties of tumor cells were examined. It was shown that the level of ERK1/2, SRC, MET, TGFb1, β-catenin phosphorylation in G33M.231, G29M.231, and G16M cells decreased. These keys showed slower proliferation and tumor size in mouse xenografts. In studies of the kinetic properties of migration, adhesion, and invasion, it was shown that the suppression of the EGFR gene changed the phenotype of the cells from mesenchymal to epithelial. However, G29M.231 metastasis when xenografting tumor cells into mice did not affect. Further analysis showed that the cell survival/apoptosis rate and tumor neovascularization were not cell-specific characteristics. The regulation of E-cadherin, p53, albumin transcription pathways plays a central role in G33M.231 and G16M target cell regulation, while the EGFR and FGFR3/ErbB14 were the main ones in equilibrium. G33M.231, G29M.231 deletion in peptide sequence or Erk1/2 inhibition trigger EMT and breast cancer stem cell expansion. The cell reprogramming inhibitors peptide KEPP and BKM120 do not suppress the migration, invasion, adhesion, proliferation, and survival of G33M, G29M.231, and G16M cells.
Keywords: Breast cancer; ERK1/2/MMP-9; RT-PCR; Metastatic breast cancer
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