The Expression And The Significance Of Circular RNA Hsa_circ₀01988 In Colorectal Cancer

Background:Colorectal cancer （CRC） is one of the leading causes of cancer-related death and a common malignant tumor of the gastrointestinal tract. There were about 1.40 million new cases of colorectal cancer worldwide, ranking third in malignant tumors in 2012,posing a great threat to people’s health. While the diagnostic performance of existing diagnostic indicators is limited. It is necessary to find new tumor biomarkers and exploring the potential biotherapy target. All of those can help to improve early diagnosis rate of colorectal cancer.With the development of second generation sequencing technology and bioinformatics, circular RNA which was found many years ago re-entered the field of our vision, it is a new type of non-linear RNA which formed by back-splicing with 5’-end and 3’-end. Although the circular RNA has been reported for decades,it was considered as by-product of splicing. So this kind of RNA did not arouse the interest of scientists. Now more and more studies have found that circular RNA was widely present in eukaryotic cells, involving a variety of biological functions of cells,meanwhile circular RNA is engaged in a variety of tumor development mechanism. Its other biological roles are also drawing increasing attention. Thus we explored the role of circular RNA in colorectal cancer.Purpose:1. Identify differential expression of circular RNA between colorectal cancer tissues and adjacent normal mucosa by the second-generation sequencing and bioinformatics methods. Then verify the differentially expressed circular RNA through PCR.2. Real-time fluorescence quantitative PCR was performed to compare the expression of cancer tissue and adjacent normal mucosa3. Combined with the patient’s various clinical information, one-way analysis of variance was conducted to explore whether the target circular RNA is related to clinical factors.4. According to the data of real-time fluorescence quantitative PCR, the ROC curve was constructed for the target circular RNA.Method:1. Two pairs of samples （cancer tissue and adjacent normal mucosa） were selected for second-generation sequencing analysis and different expression of circular RNAs were indentified by bioinformatics. Divergent primers were designed and the relative expression of circular RNA was determined by real time fluorescence quantitative PCR.Meanwhile, the PCR product was sequenced to further conform the existence of some of those circular RNA.2. The expression data of cancer and paracancer were analyzed statistically and various clinical information was combined to find out if certain circular RNA was related to those clinical factors.Result:1. The multiple circular RNAs were screened by second-generation sequencing and data analysis in colorectal cancer. Among them, hsa_circ₀01988 was selected to confirm the existence of circular RNA hsa_circ₀01988.2. The relative expression data from real time fluorescence PCR showed that hsa circ 001988 is differentially expressed in cancer and adjacent tissues. The expression level in cancer tissue was lower than that in adjacent normal tissue.3. Statistical analysis of 31-patients clinical data showed that the expression of circular RNA hsa_circ₀01988 was related to the degree of tumor differentiation and peripheral nerve invasion.4. By constructing the ROC curve, the results showed circular RNA hsa_circ₀01988 has certain diagnostic value （AUC= 0.788, 95%CI=0.68-0.90,P<0.001）.Conclusion:1. The existence of circular RNA hsa_circ₀01988 was validated in colorectal cancer. Expression of hsa_circ₀01988 in colorectal cancer and adjacent tissue is different. Moreover, data showed that the expression of cancer tissue was lower than the adjacent tissue.2. Circular RNA of hsa_circ₀01988 may participate in the differentiation of tumor cells and the biological behavior of peripheral nerve invasion.3. Circular RNA of hsa_circ₀01988 may have potential to become a new tumor biomarkers and therapeutic target.