Poster Presentation 14th International Biennial Conference on Metastasis Research 2012

DISCOVERY OF BREAST CANCER EPITHELIAL MESENCHYMAL PLASTICITY-ASSOCIATED MICRORNAS USING BIOINFORMATICS AND NEXT GENERATION SEQUENCING (#124)

Eliza T.L Soo 1 2 , Tony Blick 1 , Mark Waltham 1 , Gregory J Goodall 3 , Angels Fabra-Fres 4 , Izhak Haviv 5 , Kaylene J Simpson 6 , Erik W Thompson 1 2
  1. St Vincent's Institute, Fitzroy, VIC, Australia
  2. Surgery, University of Melbourne, Melbourne, Victoria, Australia
  3. Center for Cancer Biology, SA Pathology, Adelaide, South Australia, Victoria
  4. IDIBELL, Barcelona, Spain
  5. Bar Ilan University, Ramat Gan, Israel
  6. Victorian Centre for Functional Genomics, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
MicroRNAs (miRs) are a class of small RNA molecules (18-25 nucleotides) that regulate gene transcript stability and processing by binding to discreet motifs in the 3’ and 5’ UTRs of mRNAs. They play important roles in development, including epithelial-to-mesenchymal transition (EMT) and mesenchymal-to-epithelial transition (MET), which have been associated with cancer metastasis. Epithelial mesenchymal plasticity (EMP) encompasses the dynamic interconversion between epithelial and mesenchymal states. Mesenchymal PMC42-ET (ET) human breast cancer cells and its’ epithelial-like PMC42-LA (LA) sub-line provide an ideal tool to study miR expression profiles associated with EMP. miR profiling of control and EMT-induced ET and LA cells using the mirVana probe set V1 and Next Generation Sequencing (miRSeq) was undertaken. Several miRs were reproducibly up- or down-regulated between untreated ET and LA cells, as well as in response to EGF. Variations in miR expression were also assessed bioinformatically in public data from >50 human breast cancer cell lines. Whilst a number of these have already been implicated in cancer (e.g. miR-200 family), other novel miRs consistently associated with EMP were also identified. The expression levels of >20 miRs were validated using TaqMan® miR assays in 6 breast cancer cell lines. Stable over-expression of miRs in cell lines with low endogenous expression was achieved by lentiviral transduction. Stable breast cancer cell lines over-expressing the miRs of interest were produced and examined by qRT-PCR for EMP-associated gene expression. Following this, miR over-expressing cell lines with EMP-related gene signatures were tested for changes in in vitro migratory potential using a monolayer wound healing assay on the Cellomics platform. miRs with the most compelling functional changes will be examined in the MDA-MB-468 xenograft model of in vivo EMP.