Abstract 3769: Evaluation of the expression and role of miRNAs-221 and -222 in stem cells derived from breast cancer cell line MCF-7.
Authors: Carneiro Juliana , Iaboni Margherita , Quintavalle Cristina , Roscigno Giuseppina , Martonelli Sara , Condorelli Gerolama 
Published: August 2014

Abstract
Background: A small population of breast tumor cells, named tumor-initiating cells or cancer stem cells (CSCs), is highly tumorigenic when injected into immunodeficient mice. These CSCs, characterized by positive CD44 expression and low or undetectable levels of CD24 (CD44+CD24−/low), contribute to tumor metastasis as well as to drug resistance. Some miRNAs are differentially expressed in stem cells, suggesting they may play a role in stem cell regulation. Aim: This study has shown that miRNAs -221 and -222 are upregulated in breast CSCs when compared to differentiated cells. Our aim is to contribute to the better understanding of the biology of breast cancer cell line-derived CSCs, including the role of miRNAs -221 and -222 in these cells. Methods: Breast cancer cell line MCF-7 were grown as suspension cells in conditions that induce the formation of mammospheres, which are spheroid cell structures enriched with CSCs. Stem cell markers were analysed by Real Time PCR. The percentage of CD44+CD24−/low population was determined by FACS analysis. Overexpression of miRNAs -221 and -222 in mammospheres was done using lentivirus carrying plasmids of interest for overexpression of miR-221 or miR-222 and the control group were transducted by a plasmid with a random sequence. Analysis of mammosphere forming was done through mammosphere formation efficiency (MFE) assay. Placlitaxel resistance was firstly evaluated by MTT assay. Capase 3/7 is being used for apoptosis assay. Results: In this study we show that stem cell properties, such as the expression of OCT-4, NANOG and SOX2 were increased in mammosphere cultures (respectively 14-fold, 20-fold and 7-fold) and an enrichment of CD44+CD24−/low population was observed in mammosphere cultures when compared to adherent cell cultures. Furthermore, we found that miRNA-221 and miRNA-222 were upregulated in mammospheres by approximately 4-fold and 5-fold, respectively, when compared to adherent cells. Overexpression of miRNAs -221 or -222 increases the mammosphere formation efficiency (MFE) and death resistance to paclitaxel when compared to controls. Conclusions: The increased expression of miRNAs -221 and -222 in MFC-7 mammospheres could indicate that these molecules may play a role in regulatory pathways of breast CSCs. Moreover the overexpression of these miRNAs could increase mammosphere-forming capacity and confers paclitaxel resistance in breast CSCs.
Citation Format: Juliana Carneiro, Margherita Iaboni, Cristina Quintavalle, Giuseppina Roscigno, Sara Martonelli, Gerolama Condorelli. Evaluation of the expression and role of miRNAs-221 and -222 in stem cells derived from breast cancer cell line MCF-7. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3769. doi:10.1158/1538-7445.AM2013-3769