Mechanistic Study Of MicroRNA X And MicroRNA185Suppress Cell Invasion And Migration In Triple Negative Breast Cancer And Esophageal Squamous Cell Carcinoma

MicroRNAs(miRNAs) are evolutionarily conserved, endogenous, small, single-stranded noncoding RNA molecules. MiRNAs are generally19-25nucleotides in length that widely exsits in eukaryotic organisms. MiRNAs can bind to the3’ untranslated region (3’UTR) of the target mRNA to trigger mRNA degradation and/or inhibit gene translation and negatively regulate the gene expression. Accordingly, miRNAs influence numerous processes, such as proliferation, cell cycle control, apoptosis, differentiation, migration and metabolism. Recent studies show that miRNAs regulate a significant number of oncogenes, tumor suppressor genes, and genes associated with invasion, dissemination and chemoresistace of tumors.Breast cancer with triple negative phenotype (TNBC) are characterized by the absence of the estrogen receptor alpha (ER-a), the progesterone receptor (PgR), and the human epidermal growth factor receptor-2(HER-2) expression. The characteristics of TNBC are prone to early recurrence, distant metastasis and poor clinical prognosis, which remains a clinical challenge. In the first part of dissertation, we showed that miR-X was downregulated in TNBC cell compared with other breast cancer cell by miRNA microarray analysis. By miR-X mimic transfection, miR-X potently repressed proliferation, migration and invasion of TNBC cell in cell culture. Using the luciferase-reporter assay, we showed that miR-X expression significantly suppressed the cdc42and Six13’UTR activities, and the inhibitory effect was lost when the putative target sites for miR-X were mutated. Therefore, both cdc42and Sixl are the target gene of miR-X. Furthermore, we found that overexpression of miR-X could downregulate cdc42and Six1, which could arrest cell cycle and inhibit epithelial mesenchymal transition in TNBC cell. Intriguingly, we also demonstrated that the expression of miR-X was downregulated in40invasive breast cancers compared with adjacent normal tissues (P=0.015). Conclusion as a result, miR-X can inhibit proliferation, migration and invasion through its target genes cdc42and Sixl by regulating cell cycle and EMT in TNBC.MiR-185is an important member of miRNA family, which has been shown to be downregulated in various tumors, and it acts as a tumor suppressor gene, but its function in ESCC remains to be defined. In the second part of dissertation, we showed that the expression level of miR-185was downregulated in ESCC compared with adjacent normal tissue (P=0.016). Performing the luciferase-reporter assay, we verified that Sixl was the target gene of miR-185in ESCC. By miR-185mimic transfection, miR-185significantly suppressed proliferation, migration and invasion of ESCC cell, and the expression level of cyclin Al that was the downstream gene of Sixl was decreased. By miR-185inhibitor transfection, the expression of E-cadhrein that was the downstream gene of Six1was decreased. In summary, miR-185negatively regulated proliferation, migration and invasion in ESCC by repressing the expression of its target:Sixl.This newly identified miR-X-cdc42/Sixl nexus regulating TNBC aggression suggests that the loss of miR-X expression along with cdc42/Six1are likely targets for TNBC therapy. Our findings indicate that miR-185may be a tumor suppressor in tumor proliferation, invasion and migration by downregulation of Six1, therefore miR-185is a new therapeutic target in ESCC.