Expression of microRNA and their gene targets are dysregulated in preinvasive breast cancer

Abstract

INTRODUCTION: microRNA (miRNA) are short, noncoding RNA that negatively regulate gene expression and may play a causal role in invasive breast cancer. Since many genetic aberrations of invasive disease are detectable in early stages, we hypothesized that miRNA expression dysregulation and the predicted changes in gene expression might also be found in early breast neoplasias. METHODS: Expression profiling of 365 miRNA by real-time quantitative polymerase chain reaction assay was combined with laser capture microdissection to obtain an epithelium-specific miRNA expression signature of normal breast epithelium from reduction mammoplasty (RM) (n = 9) and of paired samples of histologically normal epithelium (HN) and ductal carcinoma in situ (DCIS) (n = 16). To determine how miRNA may control the expression of codysregulated mRNA, we also performed gene expression microarray analysis in the same paired HN and DCIS samples and integrated this with miRNA target prediction. We further validated several target pairs by modulating the expression levels of miRNA in MCF7 cells and measured the expression of target mRNA and proteins. RESULTS: Thirty-five miRNA were aberrantly expressed between RM, HN and DCIS. Twenty-nine miRNA and 420 mRNA were aberrantly expressed between HN and DCIS. Combining these two data sets with miRNA target prediction, we identified two established target pairs (miR-195:CCND1 and miR-21:NFIB) and tested several novel miRNA:mRNA target pairs. Overexpression of the putative tumor suppressor miR-125b, which is underexpressed in DCIS, repressed the expression of MEMO1, which is required for ErbB2-driven cell motility (also a target of miR-125b), and NRIP1/RIP140, which modulates the transcriptional activity of the estrogen receptor. Knockdown of the putative oncogenic miRNA miR-182 and miR-183, both highly overexpressed in DCIS, increased the expression of chromobox homolog 7 (CBX7) (which regulates E-cadherin expression), DOK4, NMT2 and EGR1. Augmentation of CBX7 by knockdown of miR-182 expression, in turn, positively regulated the expression of E-cadherin, a key protein involved in maintaining normal epithelial cell morphology, which is commonly lost during neoplastic progression. CONCLUSIONS: These data provide the first miRNA expression profile of normal breast epithelium and of preinvasive breast carcinoma. Further, we demonstrate that altered miRNA expression can modulate gene expression changes that characterize these early cancers. We conclude that miRNA dysregulation likely plays a substantial role in early breast cancer development.