Construction Of CeRNA Network In Breast Cancer With Bone Metastasis And Transcriptome Study Of Bone Metastases In Breast Cancer

Part 1:Bioinformatics analysis to explore the biological process of bone metastasis of breast cancerBackground:Compared with other types of malignant tumors,the survival peroid of patients with breast cancer is longer.Bone metastasis is particularly common in patients with advanced breast cancer,which often leads to skeletal related events and serious decline in quality of life.So far,the potential molecular mechanism of bone metastasis in breast cancer has not been fully elucidated.Therefore,it is of great significance to explore the pathogenesis of bone metastatic breast cancer.Objective:To explore the expression characteristics of related genes in breast cancer metastasis by using whole gene expression profile combined with EMT-related and BMP-related genes,and to establish the molecular regulation,immune infiltration and clinical prediction characteristics of breast cancer bone metastasis based on the ceRNA network,immune infiltration pattern and prognostic model.Methods:Three sets of gene expression profile data from GEO were downloaded,and EMT-related differentially expressed genes were screened by combining with dbEMT database.We downloaded two sets of GEO datasets related to bone metastasis of breast cancer and analyzed the differential expression of BMP-related genes.The expression profiles of mRNAs,lncRNAs,and miRNAs in 1091 cases of primary breast cancer were obtained from TCGA database,of which 58 cases had bone metastasis and 1033 cases had no bone metastasis.We constructed the ceRNA network based on the different expression between two groups.Cibersort was used to analyze the composition and clinical significance of 22 kinds of immune cells in breast cancer samples.We analyzed the genes in the ceRNA network in batch,explored the genes related to clinical prognosis,constructed the related gene prognostic model and evaluated its value.Finally,the biological verification of some differentially expressed molecules found in our previous study was completed.Results:A total of 304 EMT-related differentially expressed genes were screened by analyzing the expression profiles downloaded from three GEO datasets(GSE20685,GSE12276,GSE16446)and combining with the dbEMT database.These genes are mainly related to the signaling pathways of PI3K/Akt and TGF-beta signaling pathway.To analyze the BMP-related differential genes in breast cancer bone metastasis,GEO datasets(GSE2034 and GSE124647)were downloaded.These genes are mainly related to PI3K/Akt,TGF-beta and other signaling pathways.BMPR2,BMP2,and GREM1 were differentially expressed in both datasets.From TCGA database,we found that there were significant differences in two lncRNAs,18 miRNAs and 20 mRNAs between the two groups.Four PCGs(GJB3,CAMKV,PTPRZ1,FBN3)in the ceRNA network had clinical significance in Kaplan-Meier analysis.There were significant differences in plasma cell and follicular helper T cell between the two groups.The proportion of mast cell,gamma delta T cell and plasma cell in breast cancer samples with bone metastasis has a certain correlation with TNM stage.The samples with high proportion of follicular helper T cell had better survival status(P=0.025);the samples with low proportion of eosinophilic had better survival status(P=0.01).ROC curves based on six genes in ceRNA network and tumor infiltrating immune cells showed satisfactory accuracy.It is worth noting that based on the co-expression pattern between ceRNA network and immune cells,we found that DLX6-AS1,WNT6,GABBR2 and a variety of immune cells may be associated with bone metastasis of breast cancer.Finally,qRT-PCR was used to detect the expression of hsa-miR-132-3p and WNT6 in three cases with bone metastases of breast cancer and three cases without bone metastases of breast cancer.Conclusions:This study identified the expression of EMT-related genes in metastatic breast cancer and the expression features of BMP-related genes in breast cancer with bone metastasis.Based on the ceRNA network and infiltration pattern,the characteristics of the initial stage of biological process of bone metastasis of breast cancer are explored.This study provides an effective bioinformatics basis for exploring the molecular mechanism and establishing the clinical prognosis model of bone metastasis of breast cancer.Part 2:Molecular biological characteristics and regulatory network of bone metastases in breast cancer by whole transcriptome sequencingBackground:Bone metastasis of breast cancer is a serious threat to the physical and mental health of patients and can shorten the survival time.At present,the international research mainly focuses on how to monitor and treat breast cancer in situ,however,the diagnosis,monitoring and treatment of breast cancer bone metastases are sometimes ignored.Objective:The purpose of this study is to clarify the molecular biological characteristics of bone metastasis in breast cancer,to form the molecular regulatory network of bone metastasis in breast cancer.Methods:Whole transcriptome sequencing was used to analyze the expression characteristics and differential expression of lncRNAs,mRNAs,circRNAs and miRNAs in bone metastases of breast cancer.In the subsequent study,we used bioinformatics analysis methods to predict biological function and correlation analysis,draw WTS network and ceRNA network of bone metastasis of breast cancer.Results:We found 174 differentially expressed lncRNAs(81 up-regulated and 93 down-regulated)in breast cancer tissues compared with peritumoral tissues.The functional enrichment results showed that the differentially expressed lncRNAs were mainly associated with TNF signaling pathway and estrogen signaling pathway.There were 13 differentially expressed mRNAs(3 up-regulated and 10 down-regulated),which were mainly associated with protein digestion and absorption,PI3K/Akt signaling pathway,and focal pathway.There were 149 differentially expressed circRNAs(89 up-regulated and 60 down-regulated),which were mainly related to B cell receptor signaling pathway,ErbB signaling pathway,and T cell receptor signaling pathway.There were 83 differentially expressed miRNAs(57 up-regulated and 26 down-regulated),which were mainly related to pathways in cancer and MAPK signaling pathway.We further performed WTS network and ceRNA network construction of breast cancer with bone metastasis.Conclusions:The information of lncRNAs,mRNAs,circRNAs and miRNAs differentially expressed in bone metastases and peritumoral tissues of breast cancer was found by whole transcriptome sequencing analysis.GO and KEGG analysis were performed to predict the characteristics of differentially expressed RNAs and to construct the regulatory network of ceRNA.We found circRNA₀069718/miR-483-3p/COL11A2 axis may be involved in the formation of bone metastases in breast cancer,and we hope to provide a theoretical basis for the etiology,pathogenesis,clinical diagnosis and treatment of bone metastasis of breast cancer.