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The circular RNA hsa_circ_000780 as a potential molecular diagnostic target for gastric cancer

ABSTRACT: The circular RNA hsa_circ_000780 as a potential molecular diagnostic target for gastric cancer

PROVIDER: PRJNA766718 | ENA |

REPOSITORIES: ENA

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The circular RNA hsa_circ_000780 as a potential molecular diagnostic target for gastric cancer

Project description:The present study aimed to identify a specific circular RNA (circRNA) for early diagnosis of gastric cancer (GC).

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Project description:Peritoneal carcinomatosis with malignant ascites is associated with dismal prognosis in gastric cancer. Malignant ascites is the most relevant body fluid in which to seek diagnostic biomarkers for peritoneal carcinomatosis. We aimed to identify and validate ascites-derived circulating microRNAs (miRNAs) that are differentially expressed between liver cirrhosis-associated benign ascites (LC-ascites) and gastric cancer-associated malignant ascites (GC-ascites). MiRNA expression levels were investigated in three independent cohorts. Overall, 165 ascites samples (73 LC-ascites and 92 GC-ascites) were obtained from the National Biobank of Korea. Initially, microarrays were used to screen the expression levels of 2,006 miRNAs in the discovery cohort (n = 22). Subsequently, quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) analyses were used to validate the expression levels of selected miRNAs in the training (n = 70) and validation (n= 73) cohorts. In addition, the levels of carcinoembryonic antigen (CEA), a commonly used tumor marker, were determined in the ascites samples. Expression levels of miR-574-3p, miR-181b-5p, miR-4481, and miR-181d were significantly lower in the GC-ascites samples than in the LC-ascites samples, and miR-181b-5p showed the best diagnostic performance for GC-ascites (area under the curve [AUC] = 0.798 and 0.846 for the training and validation cohorts, respectively). The diagnostic performance of CEA for GC-ascites was improved if CEA and miR-181b-5p were analyzed together (AUC = 0.981 and 0.946 for the training and validation cohorts, respectively). Overall, we identified ascites-derived circulating miRNAs capable of differentiating non-malignant ascites and GC-ascites, and demonstrated that the combined use of miR-181b-5p and CEA produces the optimal diagnostic yield.

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Search for a novel diagnostic marker for liver cancer.

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Project description:MicroRNA (miRNA) sponges containing miRNA complementary binding sites constitute a potentially useful strategy for miRNA-inhibition therapeutics in cancer patients. Recently, naturally occurring circular RNAs (circRNAs) have been revealed to function as efficient microRNA sponges. We hypothesized that synthetic circRNA sponges targeting oncomiRs could be constructed and used to achieve potentially therapeutic microRNA loss of function. In this study, linear RNA molecules containing five miR-21 binding sites were transcribed in vitro. After dephosphorylation by calf intestinal phosphatase and phosphorylation by T4 polynucleotide kinase, circRNA sponges were circularized using 5’-3’ end ligation by T4 RNA ligase 1. Synthetic circular sponge stability was assayed in the presence of RNase R or fetal bovine serum. Luciferase reporter and cell proliferation assays were performed to assess competitive inhibition of miR-21 activity by circRNA sponges in NCI-N87 gastric cancer cells. Tandem Mass Tag (TMT) labeling proteomics analysis and Western blotting were performed to delineate effects of circRNA sponges on miR-21 downstream targeted proteins. Our experiments revealed that artificial circRNA sponges can be synthesized using enzymatic ligation. These synthetic circRNA sponges are more resistant than their linear RNA counterparts to nuclease degradation in vitro. They effectively suppress the activity of miR-21 on its downstream protein targets, including the important cancer protein DAXX. Finally, they also inhibit gastric cancer cell proliferation. Our results suggest that synthetic circRNA sponges represent a rapid, effective, convenient strategy to achieve loss of miRNA function in vitro, with potential future therapeutic application in vivo.

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