Downregulation of miRNAs in Pluripotent Cancer Stem Cells Is Mirrored in Advanced Ovarian Serous Carcinoma
RJ Flavin, M Gallagher, S Elbaruni, CC Heffron, S O'Toole, P Smyth, A Laios, S Aherne, C Martin, K Lao, O Sheils, JJ O'Leary. Trinity College Dublin, Dublin, Ireland; Applied Biosystems, Foster City, CA
Background: Cancer stem cells (CSCs) can self-renew, differentiate and regenerate tumours and have now been described in ovarian malignancy. Description of de novo tumourigenesis in SCID mice from a single CSC has fuelled the belief that CSCs drive primary tumourigenesis and that their persistence post-intervention contributes to metastasis and recurrence. We have previously reported independent identification of sets of microRNAs (miRNAs) specifically expressed during early differentiation of pluripotent Ntera2 CSCs. Despite their dissimilar origins, an overall comparison of CSC and tumour-specific data sets identified striking similarities with respect to downregulated miRNAs.
Design: Pluripotent Ntera2 cells were maintained in the self-renewal state, passaged via cell scrapping, and differentiation was facilitated by addition of 10mM retinoic acid (RA) for 3 days. Self-renewing and 3 days RA-differentiated samples were then harvested. Total RNA was extracted from Ntera2 cells and 6 fresh frozen serous carcinomas of high grade and advanced FIGO stage using mirVANA miRNA isolation kit. A novel Applied Biosystems 330 multiplex stemloop RT/PCR kit was used for miRNA gene expression profiling. Analysis of relative miRNA expression data was performed using the 2-Ct method. miRNAs were subsequently analysed for clustering, predicted targets and pathway associations using the miRBase, miRGen and miRNApath web-based resources.
Results: Of 26 miRNAs upregulated in self-renewing CSCs, only one, miR-301, was similarly upregulated in tumour samples. However, of 56 miRNAs downregulated in self-renewing CSCs, 31 (55%) were similarly downregulated in tumour samples. Seven of the top ten downregulated miRNAs were mirrored in tumour samples, two of which, miRs-509 and -188, were in the top ten downregulated miRs in tumour samples. These miRs target key developmental, malignancy and stemness signalling pathway genes.
Conclusions: Groups of miRs with roles in CSC self-renewal and differentiation function in CSCs alone and throughout tumourigenesis. State-specific miRs are likely to facilitate extensive self-renewal and differentiation to facilitate tumourigenesis while the constant repression of miR targets may facilitate key cancer processes such as avoidance of the immune system and internal growth regulation.
Wednesday, March 11, 2009 9:30 AM
Poster Session V # 142, Wednesday Morning