A NOVEL MICRO-RNA FAMILY AS MOLECULAR DETERMINANT OF MAMMARY STEM CELLS

A novel microRNA family as molecular determinant in mammary stem cells MicroRNAs (miRNAs) are an evolutionarily conserved class of small (18-22 nucleotides) noncoding RNAs involved in the regulation of a variety of cellular and developmental processes. MiRNAs have recently emerged as key regulators of transcriptional programs that control self-renewal and the cell-fate of stem cells (SCs). In the breast compartment, forced expression of some miRNAs (e.g. miR-200, let-7 and miR-93) was shown to inhibit normal and/or cancer SCs by silencing self-renewal determinants. In fact, these miRNAs are poorly or not expressed in the SC compartment, and their stem-inhibiting function is mainly achieved through the induction of differentiation. The aim of this project is to identify miRNAs specifically expressed in mammary SCs isolated from normal and cancer samples that could act as markers and/or as regulators of stem cell biology. We will also evaluate their potential as novel therapeutic targets in breast cancer. Using an innovative technique developed in our lab for the isolation of quasi-pure SCs/CSCs, we identified a miRNA family as being highly enriched in the SC compartment, in both primary normal and tumor samples. Endogenous levels of miRNAs of this family can stratify distinct sub-populations with different SC abilities, and in human cancer high miRNA levels correlate with a basal tumor subtype and with an elevated CSC content. Furthermore, depletion of these miRNAs by a synthetic sponge impair self-renewal of SCs/CSCs, as measured by a serial mammosphere propagation assay in human cancer cell lines and in mouse primary epithelial cells. In vivo, miRNA loss reduced the frequency of tumor formation, as well as the mean tumor volume and the frequency of CSCs measured with either an established human basal-like cancer cell line or human patient-derived xenografts (PDXs). To determine the molecular mechanisms underlying the regulation of SCs/CSCs behavior, we employed an unbiased approach assessing the effects of miRNA loss on global gene expression profile, coupled with the prediction of miRNA targets involved in stem cell biology. We identified 91 stem cell genes that are putatively targeted by this miRNA family, belonging to pathways critically involved in the regulation or in the mainteinance of stem cell traits. In conclusion, our results suggest that this SC-specific miRNA family could potentially be used as novel therapeutic targets for breast cancer treatment. This study also provides insights into the mechanisms that sustain CSCs growth in the breast, involving complex molecular circuits of coding as well as noncoding RNAs.