Role of DNA Methylation in Altering Gene Expression During the Early Stages of Human Breast Cancer Progression in the MCF10AT Xenograft Model

摘要

We hypothesize that simple interpretable patterns can be derived by following changes in breast cell from a single individual as they undergo progression from normal appearing ductular forms. We propose to: (1) Collect microdissected tissue representative of each of the morphologically different stages of early breast cancer progression in the MCFlOAT model to obtain RNA and DNA for miroarray analysis of gene expression and PCR- amplification for analysis of global and gene specific CpG island methylation, (2) Compare methylation patterns of candidate genes in tissue with those of the cultured MCFlOAT derived lines used for xenografts, (3) Prepare a CpG island methylation and a gene expression profile for each of the tissue types and cell lines. This year, we completed examination of DNA methylation and mRNA levels of 7 candidate genes by bisulfite sequencing and quantitive methylation specific PCR in 6 lines including MCFlOCA lines with differing metastatic potential. Methylation status of 3 genes was determined in microdessected DCIS formed by these lines. No major differences were detected between cell lines but significant changes in methylation occurred in tumors. Microarray analysis of DNA methylation and expression to be carried out in the next year should aid interpretation of these differences.