Investigation the Molecular Mechanism of Long Non-Coding Rna-Crnde-Regulated Gene Expression and Identification Inhibitors for Crnde or for Molecules Involved Crnde-Regulated Signaling in Colorectal Cancer Cells

Activation of β-catenin due to an APC gene mutation was linked to the initiation of colorectal cancer (CRC) tumorigenesis. However, our recent study showed that the low mutation rate (~42%) of the APC gene was found in Taiwanese CRC patients, which is close to the reported level in Asia. These results indicated that another factors exist to contribute CRC tumorigenesis for low mutation rate of APC gene of CRC patients. Several studies have highlighted the role of long non-coding RNAs (lncRNAs) in the development of CRC. Our recent study showed that ColoRectal Neoplasia Differentially Expressed (CRNDE) was top 20 upregulated genes in CRC clinical tissues to compare with colorectal normal tissues by analysis GEO dataset (GSE21815) which was increased by 29-fold in CRC tissues compared with normal colorectal tissues. However, the functions and detailed signaling pathways of CRNDE in CRC tumorigenesis remain to be clarified. To further characterize the clinical relevance of CRNDE in cancers, we performed a pan-cancer analysis for the expressions of CRNDE gene in normal and cancerous tissues by analysis Oncomine database. Our preliminary result shown that CRNDE was frequently overexpressed in various cancer datasets. In addition, significant high expression level of CRNDE was found in TCGA-CRC dataset. To evaluate mRNA expression levels of CRNDE in different status of APC gene of CRC cell lines, mRNA expression levels of CRNDE were assessed. Surprisingly, we found that that high expression level of CRNDE mRNA was detected in APCMut. of CRC cell lines to compare with APCWT of CRC cell lines. Moreover, we further examined CRNDE mRNA expression profiles in 35 NT pairs tissues of Taiwanese CRC tissues. We found that there was no significant difference for CRNDE mRNA expression levels in 35 NT pairs tissues of Taiwanese CRC patients with low mutation rate of APC gene. The association of lincRNAs with Polycomb Repressive Complex 2 (PRC2) has been demonstrated to provide regulatory specificity to the complexes by localizing them to genomic DNA targets and modulate PRC2-specific targeting of genes to repress genes expression. Our preliminary results revealed that 85 genes were overlapped between two gene sets of siRNA-mediated knockdown of CRNDE and PRC2 to show that these genes identified enrichment in cell cycle and DNA replication pathways. To identify specific inhibitor to target CRNDE, the GEO dataset (GSE16226) was analyzed and queried to Enrichr and CMap databases. Our preliminary result show that the clinical use of antifungal agent, Ciclopirox (CPX), has high potential to inhibit CRNDE expression and has the ability similar to Hsp90 inhibitor but less toxicity in human ocular cells. Thus, four specific aims will be achieved in this proposal: Aim 1: To establish the correlation between CRNDE and APC gene in CRC cells Aim 2: To investigate the function of CRNDE in CRC cells Aim 3: To investigate the interaction between CRNDE and Polycomb repressive complex 2 (PCR2) in HT29 cells with ZnCl2-inducible APC expression (APC/HT29 cells) Aim 4: To investigate the effect of CPX in CRC cells in vitro and in vivo