The effect of HER2 overexpression on breast epithelium subpopulations

Abstract

The human mammary gland is a dynamic organ that undergoes major developmental stages which lead to the differentiation of the cells into two main cell types: luminal and myoepithelial. Because of the cell heterogeny within the gland, it is predisposed to tumorigenesis. Normal tissue expresses EGFR receptors to a certain degree, but dysregulation can cause cancer. HER2, a member of the EGFR receptor family, is overexpressed or amplified in 15-20% of breast cancers. Although treatment is available, these tumors are aggressive, and patients often regress or present with metastasis. However, the origin of these tumors is not well understood. The main hypothesis of the study was how HER2 affects the phenotype of the two mammary gland cell lines and whether the HER2 overexpression leads them to adapt a similar phenotype.To analyze the effect of HER2 overexpression on phenotypes in mammary gland cells, we used two previously established cell lines, SI28 and D382. SI28 represents basal/myoepithelial cells and D382 represents luminal epithelial cells. The master’s student before me successfully overexpressed the HER2 protein in these cell lines which were then used as the test group compared to the original empty cells. The cells were cultured in 2D and 3D and RNA lysate collected at two time points, 2 hours and 48 hours after feeding, and the lysate then sequenced. Comparing of the control versus the test group gave us differential expression data directly relating to HER2 overexpression. We analyzed the proliferation capabilities of the cell lines in medium both supplemented with EGF and depleted for 9 days. The D382 cells grew similarly in both mediums while both SI28 empty and HER2 preferred the EGF medium. This led us to believe that HER2 did not factor into the proliferation in relation to EGF in the luminal D382 cells.To analyze the RNA sequencing data, the Sleuth R package was used. We had to optimize our methods of approaching the data for proper analysis and began by reducing the number of transcripts expressed by discarding data with p>0.05, from which we got lists that were manageable. The lists expressed various values that could be construed as upregulation and downregulation, specifically beta estimates that were based on transcripts per million values.There were some discrepancies between expressions in cell types which indicates that the cell lines respond differently to HER2. Both cell lines upregulated genes that assist in motility, survival, and proliferation, all hallmarks of cancer. The results indicate that the cell types adapt a phenotype unspecific to the cell lines upon overexpressing HER2.