James W. Freeman, Alakesh Bera, Kolaparthi Venkata Subba Rao and Muthu Saravanan Manoharan
Accepted Abstracts: J Cancer Sci Ther
Recent studies link cancer cells that possess an epithelial to mesenchymal transition (EMT) phenotype and cancer stem cell (CSC) properties with chemoresistance and metastatic potential. Here, we further investigated the molecular bases of the CRMP with emphasis on identifying a miRNA signature that may play role in maintenance of CRMP and whether this signature is found in advanced pancreatic cancers. We developed a gemcitabine resistant cell line model from BxPC3. This gemcitabine resistant cell line (BxPC3-GZR) showed a pronounced mesenchymal phenotype and expressed high levels of CD44 a marker found on stem cells and showed a highly significant increase in expression of nine miRNAs and decrease in eight miRNAs. Based on relevance to cancer and on validation by q-RT-PCR we chose six of the miRNAs (four up regulated and two down regulated) as a signature to determine whether they were similarly deregulated in 43 advanced PDAC tumor specimens previously analyzed as part of the Total Cancer Genome Atlas (TCGA). A strong correlation was observed for five of the six miRNAs in the tumor specimens compared to normal pancreas tissue. To begin to assess the functional relevance we focused on miRNA-125b for which target genes are known. Knockdown of miRNA 125b in BxPC3-GZR cells caused a partial reversal of the mesenchymal phenotype and enhanced response to gemcitabine. Moreover, a screen of additional PDAC cell lies revealed an association of miRNA-125b expression with expression of the mesenchymal marker vimentin and CD44 and with loss in expression of E-cadherin an epithelial marker. Moreover, RNA seq data from each of the 43 PDAC tumor specimens revealed a negative correlation between the expression of miRNA-125b and five potential miRNA-125b target genes (BAP1, BBC3, NEU1, BCL2, STARD13). The results of this study suggests that CRMP may be caused in part by deregulation of a specific set of miRNAs and that these miRNAs are also deregulated in advanced PDAC. These miRNAs and their molecular targets may serve as targets to enhance sensitivity to chemotherapy and reduce metastatic spread.