Study On Identifying The Clinical Value Of Plasma Exosome MiRNA And Its Potentially Regulated Immune-related Genes In Prostate Cancer

Objective:The exosome miRNAs(exo-miRNA)with clinical value in prostate cancer(PCa)was screened and identified.Combined with bioinformatics analysis,to further explore the clinical value of immune-related genes which potentially regulated in the primary tumor microenvironment.Methods:Patients with benign prostatic hyperplasia(BPH),which prostate specific antigen(PSA)> 4ng/ml,and PCa were selected from the affiliated Hospital of Guizhou Medical University,in total 31 cases.Collected venous blood samples from 2 patients with PCa before and after operation,and then performed exosome miRNA sequencing.Combined data GSE136321 from Gene Expression Omnibus(GEO),which include PCa and castrated resistant prostate(CRPC)patients’ plasma exo-miRNA sequencing data,to extracted the differentially expressed exosome miRNAs.the expression of screened exo-miRNAs was detected in BPH group(n = 13)and PCa group(n = 16),and the diagnostic efficacy was evaluated by ROC curve.Using exosomes database(EVmiRNA),the source of exo-miRNA was highly associated with cancer cell and fibroblasts.The exo-miRNA sequencing data of PC-3 cell line and primary cultured fibroblasts of PCa were used to further verify the expression of screened exo-miRNAs.Combined with the sequencing data of PCa samples in tumor gene map database(TCGA),the infiltration of 67 cell classes in tumor tissue were evaluated by x Cell algorithm.The correlation analysis showed that the correlation between immune cells and screened exo-miRNAs expression was the highest.Therefore,the immune-related genes potentially regulated by screened exo-miRNAs were further extracted by combining predicting miRNA binding genes with the immune-related gene set obtained from the immunology database(Imm Port).The genes associated with PCa recurrence were screened by univariate Cox regression and Lasso regression analysis,and then the prognostic risk model was constructed.The prediction performance of risk model was verified by using TCGA and GSE116918 dataset.Finally,the risk score and clinical characterize(TNM stage,PSA,Gleason score)collected from TCGA data were incorporated into univariate and multivariate COX regression analysis to establish a Nomograph for predicting PCa recurrence.In addition,the potential clinical value of immune-related genes regulated by screened exo-miRNAs were further discussed by analyzing the correlation between risk model and PCa tumor mutation,drug resistance and suppressive immune microenvironment.Results:In this study,three differentially expressed plasma exo-miRNAs.The verification shown that the expression of miRNA(hsa-miR-9-5p,P<0.001;hsa-miR-200a-3p,P<0.05)in plasma exosomes of PCa patients were significantly higher than BPH patients,and the diagnostic efficacy of them were better than that of PSA in the 29 Patients,which included in this study(hsa-miR-9-5p,AUC=0.865;hsa-miR-200a-3p,AUC=1.760;PSA,AUC=0.0678).Combined with the exosome miRNA database and the exosome miRNA sequencing data of PC-3 cell lines and primary cultured fiberbiasts cells of PCa tissue,it was found that PCa cells were the potential source of exo-miRNAs(hsa-miR-9-5p,hsa-miR-200a-3p),and the expression in PCa tumor tissues were positively correlated with immune cell infiltration.The prognostic risk score model and normogram based on exo-miRNA(hsa-miR-9-5p,hsa-miR-200a-3p)potentially regulated immune-related genes could well predicted the recurrence of PCa.Further analysis showed that the high-risk cohort had higher frequency of TP53 and AR mutations,Higher effective half inhibitory concentration of bicalutamide,and lower response rate to immunotherapy.Correlation analysis showed that the risk score was positive related to the tumor mutation burden and inhibitory immune microenvironment of PCa.In addition,GSEA enrichment analysis showed that the high-risk cohort was significantly enriched in cell cycle,homologous recombination,oxidative phosphorylation,primary immunodeficiency,and positive related to P27 pathway and SUMO pathway.Conclusions:1.Plasma exo-miRNA(hsa-miR-9-5p,hsa-miR-200a-p)was differentially expressed in patients with PCa and BPH.In the cases of abnormal PSA,it has a certain ability to distinguish between tumor and BPH patients,and has the potential to be used as a diagnostic marker marker of PCa.2.PCa cells are the potential source of exo-miRNA(hsa-miR-9-5p,hsa-miR-200ap).The expression of hsa-miR-9-5p and hsa-miR-200a-p in tumor tissues were related to immune cell infiltration and might be involved in the communication between PCa cells and immune cells.3.The prognostic model based on immune genes potentially regulated by exomiRNA(hsa-miR-9-5p,hsa-miR-200a-p)could predict the recurrence of PCa and is related to PCa drug resistance and tumor immune escape,might have clinical value.However,the mechanism of immune genes regulation mediated by exo-miRNAs still needs to be verified by in vivo and vitro experiments.