The Role And Potential Mechanism Of EPB41L3 In The Prognosis And Diagnosis Of Glioma

Background:In recent years,the incidence and mortality of malignant tumors have continued to rise in China.Glioma,as the most common primary nervous system tumor,has an annual incidence of about 3-8/100,000 people.The prognosis of glioma tends to depend on WHO grade,the higher the grade,the worse the prognosis and the higher the recurrence rate of the patient Therefore,there is an increasing need for biomarkers that can accurately assess prognosis and treatment efficacy,and predict tumor recurrence.With the deepening understanding of the tumor immune microenvironment,how to reverse glioma immunosuppressive microenvironment and expand the application of immunotherapy in glioma is an urgent problem to be solved at present Erythrocyte membrane protein ligand 4.1-like 3(EPB41L3),as a membrane protein,has been confirmed to be involved in the development and progression of multiple tumors.However,there is a lack of studies on EPB41L3 and glioma,and the role of EPB41L3 in glioma remains to be clarified.Methods:(1)RNAseq data and clinical data were obtained from the UCSC XENA database and TCGA database,and the data were collated and analyzed by R software.The mRNA expression level of EPB41L3 in glioma was investigated using bioinformatics analysis and validated in GEO and CGGA databases.The survival analysis of OS and PFI was performed by dividing the median value of EPB41L3 into high and low expression groups.Subsequently,univariate Cox regression and multivariable Cox regression analyses were performed to analyze the relationship between EPB41L3 expression,WHO grade,glioma survival prognosis,etc.(2)The correlation between the expression level of EPB41L3 and methylation and the relationship between methylation and prognosis were analyzed by methylation site data of TCGA.Enrichment analysis was performed according to differentially expressed genes associated with high and low EPB41L3 expression to explore the molecular mechanisms and regulatory pathways associated with glioma prognosis.To investigate whether EPB41L3 impacts the tumor immune microenvironment by investigating its association with immune function,immune cells,and immune checkpoints.(3)Finally,the expression and effect of EPB41L3 in glioma were further verified by cell function tests,immunohistochemistry,and other experiments.Results:(1)EPB41L3 was lower expressed in glioma compared with normal tissues.The results of subgroup analysis showed that the expression level of EPB41L3 was lower with a higher WHO grade.Further validation was performed in the GEO data accessibility CGGA database.The expression level of EPB41L3 in gliomas was correlated with overall survival and progression-free time interval,and the higher the expression level,the better the prognosis.(2)The expression of EPB41L3 was negatively correlated with methylation,and most of the methylation sites were associated with poor OS in gliomas.Gene enrichment analysis showed that the expression of EPB41L3 was mostly related to cell membrane transport,neural synapses,cell cycle,and other mechanisms and pathways.In addition,our study suggests that EPB41L3 is associated with a variety of immune functions,immune cells,and immune checkpoints.(3)EPB41L3 overexpressing cells were constructed by using lentivirus,and successful overexpression was verified by WB and RT-qPCR.CCK8,colony formation assay,transwell,and scratch assay showed that EPB41L3 inhibited glioma cell proliferation and migration,respectively.The immunohistochemical results showed that the expression of EPB41L3 gradually decreased with the increase of WHO grade.Conclusion:EPB41L3 expression is lower and methylation level is increased in glioma,which not only plays a role in tumor suppression but also is related to prognosis and tumor immune microenvironment EPB41L3 can be used as a biomarker of glioma to provide value for the diagnosis,treatment,and prognosis of glioma.