Molecular Subtype And Immunotherapy Evaluation Of Bladder Cancer Based On Hypoxia-related Genes

Objective: Bladder cancer is one of the most common genitourinary malignancies,with a risk of invasiveness,metastasis and recurrence.Immunotherapy has made breakthroughs in the treatment of bladder cancer in recent years,and several immune checkpoint inhibitors have been approved for the clinical treatment of bladder cancer.Hypoxia is one of the hallmarks of malignancy that induces an immunosuppressive tumor microenvironment and hinders the use of immunotherapy in bladder cancer.Therefore,we aimed to identify reliable predictive biomarkers of immunotherapy and provide new directions for clinical immunotherapy in bladder cancer.Methods: 1.Download data such as RNA expression profiles of408 bladder cancer cases from the TCGA bladder cancer cohort and obtain hypoxia-related genes from the Gene Cards database.A scale-free gene co-expression network of bladder cancer based on hypoxia-related genes was constructed using weighted gene co-expression network analysis,and gene function annotation analysis was performed on the genes in key modules using GO and KEGG analysis.2.Consensus clustering algorithm was used to construct the molecular subtype of bladder cancer based on hypoxia-related genes,and GO,KEGG and GSEA analysis were used to perform gene function annotation analysis for different bladder cancer subtypes,and the R package GSVA was used to calculate the enrichment scores of gene signatures related to bladder cancer.R packages maftools and GISTIC 2.0 were used to evaluate the genomic variation of different subtypes,and ss GSEA,MCPcounter,EPIC,ESTIMATE and TIMER algorithms were used to quantify the level of immune cell infiltration of different subtypes in bladder cancer.3.Based on the expression levels of hypoxia-related genes,hypoxia score was constructed using principal component analysis.The ss GSEA method was used to evaluate the level of tumor immune cell infiltration in different hypoxia score groups.The combined analysis based on the IMvigor210 cohort and the Sub Map algorithm were used to predict immunotherapy responsiveness in patients with different hypoxia scores.4.Use Cytoscape to screen hub genes,GSCALite and TIMER are used for pan-cancer analysis and immune analysis of hub genes.Results: 1.The brown module screened based on hypoxia-related genes has a strong correlation with immune cells.The results of gene function and pathway analysis show that the hypoxia-related genes in the brown module are mainly enriched in immune-related pathways.2.Molecular subtype of bladder cancer was constructed according to the expression levels of hypoxia-related genes in the brown modules,and the patients were divided into Cluster1 and Cluster2.2.The differentially expressed genes between the two subtypes are mainly enriched in immune-related pathways,and there are significant differences between the two subtypes in terms of genomic variation and gene signatures enrichment scores in bladder cancer,the level of tumor immune cell infiltration and the expression of immune markers in Cluster 1 was significantly higher than that of Cluster2.3.According to the expression levels of hypoxia-related genes in the brown module,hypoxia score was constructed,and bladder cancer patients were divided into high hypoxia score group and low hypoxia score group.The abundance of tumor immune cell infiltration in the high hypoxia score group was higher than that in the low hypoxia score group.The immunotherapy response prediction of two score groups based on the IMvigor210 cohort and Sub Map algorithm showed that the high hypoxia score group had higher response to immunotherapy.4.The immune infiltration analysis of 10 hub genes FCER1 G,ANXA6,CYBB,TLR8,SPI1,C5AR1,COL6A1,COL6A2,ITGB2 and TIMP2 was performed,and it was found that their genomic variations were related to the immune infiltration of bladder cancer.Conclusions: In this study,we constructed molecular subtype and hypoxia score in bladder cancer based on hypoxia-related genes.Molecular subtype of bladder cancer allows assessment of immune cell infiltration in the tumor microenvironment of different patients.The hypoxia score can effectively predict the responsiveness to immunotherapy in bladder cancer patients.The hub genes FCER1 G,ANXA6,CYBB,TLR8,SPI1,C5AR1,COL6A1,COL6A2,ITGB2 and TIMP2 screened in this paper are also expected to become new targets for bladder cancer treatment.