Identification Of Abnormal Expression Of MRNA,microRNA And Aberrant DNA Methylation Patterns In Rectal Adenocarcinoma Based On TCGA Database

Objective:Colorectal cancer is one of the most common tumors and thesecondleadingcause of cancer mortality in China.Although fecal occult blood test and colonoscopy can significantly improve the detection rate of precancerous lesions and early stage colorectal cancer,and thus improve the prognosis of patients,the poor compliance of patients with colonoscopy seriously inhibit the promotion of colon cancer screening.Therefore,we still need to find a convenient and accurate biomarker for colorectal cancer.Although a large number of studies have identified various markers for early diagnosis,prognosis prediction and treatment evaluation for colorectal cancer,most of them have small sample size.Different studies included different pathological types and stage of patients,which make the results inconsistent.Therefore,based on the reactal adenocarcinoma(READ)cohort in TCGA database,the present study aims to analyze the expression profiles of mRNA,microRNA and DNA methylation data comprehensively,in order to lay a theoretical foundation for the identification of new biomarkers and elucidation of the mechanism of development and progression of rectal adenocarcinoma.Method:Firstly,by compared the profile of mRNA,miRNAand DNA methylation level between tumor tissues and normal adjacent tissues of 155 patients of TCGA READ cohort,differentially expressed mRNAs/miRNAs/methylated regions were identified.Then,the predictive target genes of differentially expressed miRNA,differentially expressed mRNAs and abnormal methylation genes were compared through bioinformatics methods,and the differentially expressed genes possibly regulated by miRNAs or methylation were identified.The biological functions of the differentially expressed genes were analyzed by pathway enrichment analysis Next,in order to further validate the results of TCGA READ cohort,five paired samples of tumor tissues and adjacent normal tissues were used to verify the expression of 9 randomly selected differentially expressed genes by qRT-PCR.Then,295 specimens of READ tumor tissues and adjacent normal tissues were collected from two datasets(GSE75970 and GSE20842)in GEO database and the aboved results were further validated.Result:Based on the data of mRNA,miRNAs and methylation in TCGA READ cohort,1192 dysregulated mRNA 27 dysregulated miRNAs and 6403 aberrant methylation CpG sites were screened.Then,a total of 668 differentially expressed genes were identified as target genes of 27 miRNAs.50 genes with abnormal methylation sites were identified.Pathway enrichment analysis showed that these 50 genes involved in 9 KEGG pathways,including cAMP signaling pathway,circadian entrainment,glutamatergic synapse,nicotine addiction,amphetamine addiction,morphine addiction,melanogenesisretrograde endocannabinoid signaling and cholinergic synapse.Subsequently,we randomly selected 9 genes from the 50 genes for validation by using 5 pairs of cancer and paracancerous tissues.Compared with paired paracancerous tissues,ATP2B4,ROR1 and PKCB were significantly down-regulated,while TCF7,SLC6A6,PDPN,WNT2 and ONECUT2 were significantly up-regulated in READ,which were consistent with TCGA READ cohort.In addition,295 cases of READ from two datasets(GSE75970 and GSE20842)in GEO database were.The analysis showed that the expression trend of 33 differentially expressed genes screened from GEO datasets were identical with the results of TCGA READ cohort.Conclusion:In this study,differentially expressed genes,miRNAs and abnormal methylation sites in READ were screened by analyzing the TCGA READcohort.A regulatory network of-miRNA-mRNA-methylation in READ was constructed.These results lay a theoretical foundation for screening new markers of rectal adenocarcinoma.