Study Of Pan-Cancer Small Non Coding RNA In Digestive Tract Based On Sequencing Data And Construction Of Prognosis Molecular Pathology Model

Background:In April 5,2018,the world's top academic journal,cell,published 10 papers on the findings of the Pan-Cancer Atlas(TCGA)universal cancer map(Pan-Cancer Atlas).Pan-Cancer Atlas is the integration of TCGA data based on different tumor types and different platforms.It not only focuses on the analysis of tumor gene sequencing data,but also analyzes data of other types of data,such as protein group,clinical and imaging data.Gastrointestinal cancer is a common disease that affects human health and life.The National Central Cancer Registry of China(NCCR)has published a total of 2,089,500 new cancers in the digestive system in China in 2015,accounted for 48.68% of all new cancers,and 1,565,500 deaths,accounted for 55.63%of the total cancer deaths.The incidence and mortality of colorectal cancer,gastric cancer and esophageal cancer were among the top five in China.Small non-coding RNA(sncRNA),especially snoRNA and snRNA,used to be considered to be stable in vivo and widely used as an internal reference gene.In recent years,these sncRNA have been found to be involved in the occurrence and development of tumors.Some snoRNA have also been found to be associated with tumor clinical characteristics and prognosis.Gastrointestinal cancer has a high mortality rate and poor postoperative prognosis.There is a high recurrence rate and a poor five year survival rate.At present,few studies on prognosis are based on single or a few close proximity medical units.These studies have shortcomings such as small sample size,lack of follow-up data,low accuracy,poor reliability and single prognostic indicators.It is urgent to have a molecular pathological model for prognosis of patients with digestive tract cancer based on large sample size,good reliability and high prediction accuracy.Molecular studies of single digestive tract tumors have been widely reported,but comprehensive reports on Pan-Cancer in digestive tract are relatively rare.The molecular mechanism and role of the tumor and the clinical value of snoRNA and snRNA in the small molecule non coded RNA is not clear.The sequencing data provides a rich sequence of high-throughput gene expression data,as well as the survival time,and other clinicopathological information.The purpose of this study is to analyze the molecular role of small molecular noncoding RNA(snoRNA,snRNA and miRNA)in the digestive tract adenocarcinoma and to construct a prognostic model for predicting the molecular and clinical prognostic value of adenocarcinoma of the digestive tract by using the gene expression data of the digestive tract adenocarcinoma in the sequencing data.Through the construction of the regulation network,protein interaction network and signal pathway to find the general law of tumor development,it opens new ideas for the research of tumor molecules.The expression of miRNA in the adenocarcinoma of the digestive tract was analyzed in the early stage,and the expression of miR-7 was up-regulated in 4 kinds of digestive tract tumors.Some studies of colorectal cancer indicate that the upregulation of miR-7 in tumor tissues has the effect of promoting cell proliferation and tumor growth.Other studies indicate that miR-7 downregulation in colorectal cancer plays a role in tumor suppressor gene.In this study,we detected the expression of miR-7-5p in tumor tissues by the fluorescent RT-PCR test of colorectal cancer paraffin tissue,and explored its cell function through in vitro cell experiments,to provide a new clue for the diagnosis and treatment of colorectal cancer.Materials and methods:This study included the level 3 expression data and survival time and other clinical pathological information of small intestinal non coding RNA and mRNA in the TCGA database in 2017,including 4 kinds of tumors,including esophageal cancer,gastric cancer,colon cancer and rectal cancer.The differentially expressed miRNA,snoRNA,snRNA and mRNA of 4 kinds of tumors were screened using the edgeR package of the R language.Univariate and multivariate COX risk regression models were used to screen prognostic factors and construct prognostic molecular pathological models.The target gene of differential miRNA was predicted by miwalk2.0 online software and the mRNA with the difference of the common expressionwas intersected.The miRNA-mRNA regulatory network was constructed,and the genes in the network were analyzed by PPI protein interaction network,GO and KEGG signaling pathway analysis and gene radar analysis.A total of 80 paraffin tissue samples from patients with colorectal or rectal cancer from July 2017 to November 2017 were collected to verify the expression of miRNA.In the tissue wax blocks of colon and rectal cancer,the level of expression was verified by fluorescence RT-PCR.The cell function experiment used the human colon cancer RKO cell line provided by the Shanghai Academy of life sciences of the Chinese Academy of Sciences.We use RT-PCR validation and functional study of miR-7-5p,which was highly expressed in colorectal cancer.After colon cancer cells were transfected with miR-7-5p inhibitor by lentivirus,MTT and flow cytometry and apoptosis assay were used to study the effect of colon cancer cells on the proliferation of colon cancer cells.Bioinformatics methods were used to analyze the molecular biological roles of miR-7-5p in colon cancer.Results:Based on a comprehensive study of gene expression profiles and clinical parameters of the digestive tract Pan-Cancer,we constructed a risk assessment model of snoRNA(SNORD14E,SNORD67)in colon cancer.3 common expressions of snRNA(RNU1-106 P,RNU6-850 P and RNU6-529P)and the prognostic value of snRNA(RNVU1-4 and RNU6-101P)were obtained in 4 kinds of digestive tract adenocarcinoma.A miRNA-mRNA regulatory network with 3 distinct miRNA(miR-552,miR-490 and miR-133a-2)as the core of the digestive tract adenocarcinoma was constructed,and the PPI protein interaction network analysis,GO and KEGG Pathway enrichment and gene radar detection were carried out on the genes in the network.PI3K-Akt signaling pathway and other signaling pathways are found in the network.In addition,miRNA(miR-7-3,miR-328 and miR-323a)with prognostic value were also screened out.Sequencing data showed that miR-7-3 was highly expressed in 4 of the digestive tract adenocarcinoma.The expression level of miR-7-5p was verified by fluorescent RT-PCR,which showed that it was upregulated in colon cancer and rectal cancer tissues.In vitro studies on the function of colon cancer cells showed that the proliferation ability of colon cancer cells decreased and the G2/M cell cycle was blocked after inhibition of miR-7-5p expression.The regulatory network of miR-7-5p and core target genes was constructed by bioinformatics.Conclusions:1.There was a large number of common abnormal expressions of miRNA,snoRNA,snRNA and mRNA molecules in 4 kinds of digestive tract adenocarcinoma in sequencing data.These common abnormally expressed biomolecules may have a common mechanism in gastrointestinal adenocarcinoma.2.In colon cancer,we successfully constructed a molecular pathological risk score prediction model consisting of snoRNA(SNORD14E and SNORD67).3.miR-7-5p played the role of oncogenein colon cancer.