| Part ?DNA methylation analysis of breast cancer based on TCGA databaseAIM:In recent years,DNA methylation has become one of the hot spots in biomedi-cal research,especially its relationship with tumors.DNA methylation may be used as a detection method to diagnose tumors and even predict the occurrence of tumors.In this study,methylation data of breast cancer patients and healthy controls in the pub-lic database TCGA were used to analyze the differences between DNA methylation profiles and normal subjects in breast cancer patients.Through further analysis and screening,Select the methylation genes that are helpful for the diagnosis and moni-toring of breast cancer.STUDY DESIGN:Illumina Human Methylase 450-chip methylation data from breast cancer samples and paracancerous tissue samples was downloaded from TCGA and the differential methylation sites in breast cancer tissue samples and paracancer-ous tissue samples were screened using the programming language R and with refer-ence to Illumina Human Methylation 450 hg19 chip annotation file for gene annota-tion.The DAVID online tools were then used to perform GO(Gene Ontology)en-richment analysis and KEGG(Kyoto Encyclopedia of Genes and Genomes)pathway analysis.Finally,the clinical data of the downloaded data was used to cox regression analysis of selected genes to screen for the survival-related methylation genes.RESULTS:A total of 791 breast cancer tissue samples and 96 paracancerous samples were sequenced and the corresponding clinical data were obtained at TCGA.A total of 28710 differentially methylated Cite(DMS)sites were obtained through R analysis.After that,we got a total of 6,623 genes related to these 28,710 DMS through gene annotation.We annotated and annotated the loci and genes structurally.As a result,we can see that most of the DMS are located on CpG islands and are hypermethylated;while the gene body is dominated by hypomethylation.Similarly,we used the DAVID tool to perform KEGG pathway analysis on the 6623 breast cancer methyla-tion-related genes previously obtained.The genes involved in the DMS involved a total of 61 pathways,and we extracted the genes from common tumor pathways Twenty-two genes were obtained by the intersection with genes containing more than 10 DMS.After further filtering according to the mean value of ?,Cox regression analysis showed that there were 7 genes related to survival time,including CTNNA2,APC,RASSF1,SOX17,TNXB,EDNRB,CREB5.CONCLUSIONS:1.Based on data sources such as bioinformatics databases such as TCGA,genetic di-versity analysis and survival analysis of diseases can be performed.2.DNA methylation profiles of breast cancer tissues and paracancerous tissues were significantly different.DNA hypermethylation in breast cancer tissues mainly oc-curred in CpG islands,while hypomethylation occurred in gene body.3.Breast cancer DNA methylation involves a variety of biological processes,we com-bined with common tumor molecular pathways and TCGA downloaded clinical in-formation files,we screened CTNNA2,APC,RASSF1,SOX 17,EDNRB,TNXB,CREB5 seven genes,Which has the potential as a methylation marker for breast cancer diagnosis and monitoring.Part ?Detection of circulating free DNA in breast cancer patients using high-throughput methylation sequencing.AIM:In our study,by collecting and extracting circulating free DNA in the peripher-al blood of patients with breast cancer and using high-precision genome-wide meth-ylation sequencing to map the methylation patterns of cfDNA in breast cancer pa-tients to explore its potential Characterization.This part of the study is the first part of the extension of the research,development and validation,based on the second gen-eration of sequencing,breast cancer patients with DNA and cfDNA methylation dif-ferences.And to examine the value of cfDNA methylation in the early diagnosis of adjuvant breast cancer,in order to find potential methylation markers in peripheral blood for screening breast cancer.STUDY DESIGN:Choose from March 2017 to July 2017 in Southern Medical Uni-versity Southern Hospital,breast cancer hospitalized 10 cases of early breast cancer female patients into the study,an additional 10 healthy women as a control group.About 10ml venous blood was taken before surgery and serum was separated.After extracting cfDNA by magnetic bead method,genome-wide methylation library was constructed,and then the captured cfDNA was sequenced by Illumina sequencing platform.RESULTS:In the process of extracting cfDNA,we found there was no significant difference in cfDNA concentration between breast cancer group and healthy control group(0.908 ng/ul in breast cancer group and 0.883 ng/ul in healthy control group,p=0.399).In addition,There was no significant correlation between cfDNA concen-tration and age,tumor size,stage and lymph node metastasis.By constructing a full-genome methylation-based high-throughput sequencing library and using full-fledged,high-throughput sequencing techniques to accurately obtain ge-nome-wide methylation site information,we found that 4052 Differentially methyl-ated sites,and by lateral comparison with the first part of the tissue DNA methylation data shows that the methylation status of cfDNA of breast cancer is different from that of breast cancer tissue DNA.In clinical correlation analysis,we found that DUSP5P1 contains 11 methylation sites related to tumor staging,including 3 hyper-methylation sites and 8 hypomethylation sites.CONCLUSIONS:1.There was no significant difference in cfDNA concentrations between breast cancer women and healthy women.In addition,cfDNA concentration in breast cancer pa-tients was not significantly correlated with patient's age,tumor size,stage and lymph node metastasis.2.The breast cancer patients cfDNA methylation is different from breast cancer DNA methylation.3.The abnormal methylation of cfDNA in breast cancer patients mainly focused on the gene body.The frequency of methylation up-regulation and methylation down-regulation was almost the same except for the CpG island gene region,while CpG island was hypermethylated to the Lord. |
PDF Full Text Request