Construction Of An Immune Prognostic Model And Exploration Of The Biological Function Of MiR-143-3p In Breast Cancer

Objective The incidence of breast cancer in the female population has always been the top one,and it is also a major cause of cancer deaths.Currently,emerging immune markers are urgently needed to promote the development of breast cancer immunotherapy.The expression level of Mi R-143-3p in breast cancer is reduced,and it exhibits a tumor suppressor effect,showing great potential value in the treatment of breast cancer.Therefore,the purpose of this study is to:1.Searching for immune genes with prognostic value in breast cancer,and construct a prognostic risk assessment model in order to make a more accurate assessment of the prognosis of breast cancer patients.In addition,it provides potential molecular markers and therapeutic targets for breast cancer immunotherapy.2.Explore the effects of miR-143-3p on the occurrence and development of breast cancer,and the genomic changes of breast cancer MCF7 cells caused by the overexpression of miR-143-3p.Finding the target genes regulated by miR-143-3p and the potential ce RNA regulatory network,and providing more theoretical basis for miR-143-3p as a potential therapeutic target for breast cancer.Methods1.Using bioinformatics analysis methods,the breast cancer data in the TCGA database was used as the training set,screen out immune gene sets related to survival-related clinical traits through WGCNA,and use gene enrichment analysis to explore the potential biological functions of this gene set.KM survival analysis and Lasso regression analysis were used to further screen out immune genes with survival value.Then,multivariate Cox regression analysis was used to construct a prognostic risk assessment model for breast cancer.According to the formula: Risk score(RS)= ∑coef(i)*exp(i),calculate the RS of each sample and group them according to the median value of RS.Using KM survival curve,C-index and ROC curve to evaluate the predictive performance of the model,and construct a nomograph for clinical use.Then,the two breast cancer data sets GSE20685 and GSE31448 in the GEO database were used as validation sets to verify the prognosis model externally.Finally,construct a transcription factor(TF)regulatory network of breast cancer immune-related prognostic genes to identify key transcription factors that regulate the immune microenvironment of breast cancer.GSEA enrichment analysis was used to explore the differences in KEGG signaling pathways between high and low risk groups,and ESTIMATE and TIMER algorithms were used to evaluate the relationship between RS and tumor immune microenvironment.2.First,the breast cancer MCF7 cell line overexpressing miR-143-3p was constructed through lentiviral transfection,and overexpression was verified by q PCR.Cell function experiments such as CCK8,Transwell,and wound healing were used for biological function verification.Whole-genome high-throughput sequencing to explore differences in genomic expression caused by miR-143 overexpression,and combine the results of differential analysis and target gene prediction to screen potential target genes of miR-143-3p,and construct miR-143-3p related ce RNA regulatory networks.The miR-143-3p inhibitor was transiently transfected to construct a miR-143-3p knockout MCF7 cell line,and the CCK8 cells function verification test was performed,and q PCR was used to verify the expression regulation relationship of the target gene.Results1.Screened out 10 immune genes(CCL5,CCR7,CSF2 RA,ESRRA,HLA-DOB,HSPA2,NFKBIA,NR1H3,S100 B and SEMA3B)that are significantly related to the prognosis of breast cancer,and successfully constructed an immune related prognosis model for breast cancer.The model has accurate predictive value and independent predictive performance.In addition,7 transcription factors related to the immune prognosis of breast cancer were identified.Multiple signal pathways that inhibit tumor progression were significantly enriched in the low-risk group,and RS was significantly negative with the degree of immune cell infiltration and immune checkpoint gene expression.2.The MCF7 cell line overexpressing miR-143-3p was successfully constructed,and the proliferation,migration and invasion experiments further confirmed that miR-143-3p can inhibit the growth of breast cancer cell lines.Whole genome sequencing analysis of overexpression cell lines showed that miR-143-3p may inhibit tumor cell proliferation and metastasis mainly by enhancing the adhesion between tumor cells.BIRC5,NACC1,FN1,RABIF,TSPAN13,BGN,GPRC5 A and LINC00511 are potential target genes for miR-143-3p to perform biological functions.Among them,BIRC5,GPRC5 A,NACC1 and LINC00511 were successfully verified in miR-143-3p knockout cell lines.Conclusions1.This model is not only an independent prognostic indicator of breast cancer,but also has higher prediction accuracy than traditional tumor pathological grades,and has important clinical value.Moreover,this model is closely related to the degree of immune infiltration,and these 10 immune-related prognostic genes may be potential immunotherapy markers for breast cancer.2.BIRC5,GPRC5 A,NACC1 and LINC00511 are important target genes for miR-143-3p to perform biological functions.Mi R-143-3p inhibits the proliferation and metastasis of tumor cells by regulating the expression of target genes and enhancing the adhesion between cells.