Genomic and functional analysis of breast cancer

Breast cancer is the second leading cause of cancer death in women in the United States, effecting women of all races and ethnicities. The widespread use of the DNA microarray in the study of breast cancer has yielded novel cancer genes and uncovered global changes that influence disease progression. Both array comparative genomic hybridization and gene expression analysis represent genomic techniques which have advanced the study of breast cancer by measuring both the genetic alterations and the transcription programs that contribute to tumorigenesis. Here these techniques were applied systematically to address varying aspects of breast cancer. Utilizing the microarray platform we were able to identify recurrent copy number alterations harboring putative oncogenes and to develop gene expression signatures of the epithelial to mesenchymal transition in breast cancer.;DNA amplifications in breast cancer are frequent on chromosome 11q, where multiple driver oncogenes likely reside in addition to CCND1 (cyclin DI). One such candidate, the scaffolding adapter protein GAB2, functions in ErbB signaling and was recently shown to enhance mammary epithelial cell proliferation, and metastasis of ERBB2 (HER2/neu)-driven murine breast cancer. However, the amplification status and function of GAB2 in the context of amplification remain undefined. Here, by genomic profiling of 172 breast tumors, and fluorescence in situ hybridization (FISH) validation, we observed focal amplification spanning GAB2 (11q14.1) independent of CCND1 (11q13.2) amplification, consistent with a driver role. Further, small interfering RNA (siRNA)-mediated knockdown of GAB2 in breast cancer lines with GAB2 amplification revealed a dependency on GAB2 for cell proliferation, cell-cycle progression, survival, and invasion, likely mediated in part through altered AKT and MAPK signaling. GAB2 knockdown also reduced proliferation in a cell line with ERBB2 amplification, consistent with the possibility that GAB2 can function downstream of ERBB2. Thus, focal amplification of GAB2 in breast carcinogenesis promotes oncogene dependency, and underscores scaffolding/adapter proteins as potential new points of therapeutic intervention.;Epithelial-mesenchymal transition (EMT), a switch of polarized epithelial cells to a migratory, fibroblastoid phenotype, is considered a key process driving tumor cell invasiveness and metastasis. Breast cancer cells in culture exhibit varying degrees of EMT. Using this model system, we sought to discover gene-expression signatures of EMT with clinical and mechanistic relevance. Supervised analysis of expression profiles was done to build an EMT signature distinguishing breast cancer lines with "epithelial-like" and "fibroblast-like" phenotypes, and in comparison to normal breast fibroblasts. Expression of a select signature gene was analyzed by immunohistochemistry on a tissue microarray, and function characterized by RNA-interference knockdown in breast cancer cells in culture. Analysis of cell line profiles defined a 200-gene EMT signature that was prognostic across multiple breast cancer cohorts. Expression of LYN, a top-ranked signature gene, was associated with significantly shorter overall survival and correlated with the basal-like phenotype. Knockdown of LYN inhibited cell migration and invasion, but not proliferation. Dasatinib, a dual-specificity tyrosine kinase inhibitor, also blocked invasion at concentrations comparable to LYN inhibition, suggesting that LYN is a likely target and invasion a relevant endpoint for dasatinib therapy. Our findings define a prognostically-relevant EMT signature in breast cancer, and implicate LYN as a mediator of invasion and possible new therapeutic target with particular relevance to clinically-aggressive basal-like breast cancer.;Taken together these studies underlie the advances genomics has provided cancer research. By combining the power of RNA inhibition technology with extensive array comparative genomic hybridization and gene expression breast cancer datasets, we were able to identify GAB2, a breast cancer oncogene driving amplification of 11q14.1 and LYN, a kinase that mediates the metastatic transition of cancer cells.