Prognosis Of HTERT In Melanoma And Gene Regulation Network Based On Bioinformatics Analysis

Objective: 1)Detect hTERT promoter mutation and its protein expression in paraffin-embedded tissues of malignant melanoma and nevus,analyze the site distribution of hTERT promoter mutation,and analyze the relationship between mutation and protein expression.The known BRAF V600 E mutation that is closely related to the prognosis of melanoma was introduced to detect the BRAF V600 E mutation and analyze its relationship with hTERT promoter mutation;analyze the relationship between hTERT promoter and BRAF V600 E mutation and the clinicopathological features of malignant melanoma.2)The 5-year survival of 77 melanoma patients was analyzed to determine whether hTERT promoter mutation was an independent prognostic factor for melanoma,and the effect of hTERT promoter mutation with BRAF V600 E on the prognosis of melanoma patients was further analyzed.3)The relationship between hTERT promoter mutation and prognosis of melanoma was meta-analyzed,and our survival analysis results were included in the meta-analysis as a study.4)Through bioinformatics technology,the hTERT gene network and its interacting genes were analyzed to further explore the potential prognostic value of hTERT-related genes and explore the hTERT-related genomic changes and functional networks.Methods: 1)The hTERT promoter and BRAF mutations in the paraffin-embedded tissues of malignant melanoma and nevus were detected by qPCR and sanger sequencing;Immunohistochemistry was used to detect the expression of hTERT gene in paraffin-embedded tissues of malignant melanoma and nevus.2)5-year survival of 77 cases of cutaneous melanoma was analyzed using Cox proportional risk regression model for univariate and multivariate regression analysis.3)327 papers published by Pub Med,Web of Science and Embase were retrieved from the electronic database.By the end of December 2019,two reviewers independently screened the papers according to inclusion and exclusion criteria to extract relevant data and assess their quality.4)The hTERT-mrna-related differentially expressed genes in melanoma were analyzed using Linked Omics database,and GO functional enrichment analysis and KEGG pathway enrichment analysis were performed to detect the target network of kinase,mirna and transcription factors.The online Target Scan software and Starbase database were used to predict the target genes of mir-497-5p.The differentially expressed genes related to hTERT mRNA were intermingled with the target genes of mir-497-5p.Then,the gene groups were enriched by GO function and KEGG pathway,and the genes were interacted by PPI network to grab the core genes.The gene changes,survival prognosis and differential expression of core genes in melanoma were further analyzed using the STRING database,c-bioportal database and Oncomine database.Results: 1)The mutation site of hTERT promoter is C228 T C250T,and mutation is more frequent with C250 T.Mutation of hTERT promoter was significantly correlated with protein expression,and the consistency was high.The alone or combined mutation of the hTERT promoter and BRAF V600 E was associated with invasive lesions in melanoma patients,and the difference was statistically significant.2)77 cases of melanoma 5 years survival analysis results indicate: Gender,age group,population distribution,subtype of melanoma and tumor location were not related to the prognosis of DFS and OS.The mitosis rate of tumor cells had prognostic significance in the univariate analysis of DFS and OS,and the independent prognostic significance was lost after COX regression analysis was included.Breslow thickness and the presence or absence of ulcer were independent prognostic factors for melanoma DFS and OS.hTERT promoter mutation alone is an independent prognostic factor for DFS OS in melanoma patients.The combined mutation of hTERT promoter and BRAF V600 E is an independent prognostic factor for OS in melanoma.3)The results of Meta analysis suggested that hTERT promoter mutation was an independent prognostic factor for poor prognosis of melanoma DFS and OS.hTERT promoter and BRAF V600 E combined mutation is an important independent prognostic factor for OS in melanoma patients.However,a larger prospective randomized controlled trial with a larger sample size is needed to demonstrate whether co-mutants are synergistic in survival outcomes for melanoma.4)hTERT is specifically associated with several tumor-related kinases(such as ATR)and transcription factors(such as E2F1 and E2F4).These kinases and transcription factors regulate genomic stability mitosis and the cell cycle.hTERT and CDK4 were directly correlated in protein interaction,and the mRNA expression of both was significantly correlated(P<0.05).CDK4 gene mutation and amplification are independent prognostic factors for DFS and OS in melanoma patients From the perspective of bioxin analysis,this result verified that hTERT promoter mutation is an important factor affecting the occurrence and development of cutaneous melanoma.Conclusion: 1)hTERT promoter mutation plays an important role in the development of melanoma and is an independent prognostic factor in patients with melanoma.2)hTERT promoter and BRAF V600 E combined mutation is an important independent prognostic factor for OS in melanoma patients.However,a larger prospective randomized controlled trial with a larger sample size is needed to demonstrate whether co-mutants are synergistic in survival outcomes for melanoma.