Analysis Of Key Molecules And Regulatory Network Of T Cell Exhaustion From Multiple Tumor Microenvironment Based On Single Cell Transcriptome Data

Tumor immune escape refers that tumor cells can escape the recognition and attack of the immune system to survive and proliferate in the body,T cell exhaustion is one of the main reasons which results in a significant obstacle to the antitumor immunity.Single cell RNA sequencing technology can quantify the expression at the level of single cell,rather than the average expression value of all cells of traditional tissue samples.Using single cell transcriptome data of multiple cancers to study the key molecules and regulatory network of T cell exhaustion is conducive to more comprehensively reveal the common mechanism of T cell exhaustion in pan-cancer.Therefore,based on the single cell transcriptome data from peripheral blood,adjacent normal and tumor tissues of colorectal cancer,liver cancer and non small cell lung cancer,using the bioinformatics methods including Seurat and Monocle package,differential expression analysis,KEGG and GO pathway enrichment,protein-protein interaction analysis,transcription factor regulation analysis,gene set variation analysis(GSVA),weighted gene co-expression network analysis and survival analysis,this research aims to explore the potential function of key molecules and common regulatory networks of T cell exhaustion of multiple tumor microenvironment.Firstly,using the single cell transcriptome data of T cells from peripheral blood,adjacent normal and tumor tissue of colorectal cancer,liver cancer and non small cell lung cancer in GEO database,different T cell subtypes of three cancers were obtained by dimension reduction clustering: CD8 + T cells included na(?)ve cells,effector cells,pre＿exhausted cells,exhausted cells,tissue resident memory cells and mucosal-associated invariant T cells.CD4 + T cells included na(?)ve cells,effector cells,exhausted cells,tissue resident memory cells,regulatory T cells,T helper cells 1,follicular helper T cells.Among them,the naive and effector cells were mainly from peripheral blood,pre＿exhausted and exhausted cells were mainly from tumor tissue.The GO analysis revealed that exhausted T cells were associated with negative regulation of immune progress.Secondly,the analysis of the effector,pre＿exhausted and exhausted cells in CD8 + T cells showed that there were 162 up-regulated genes in the exhausted cells.RGS1 showed a large fold change in the pre＿exhausted and exhausted cells,with high expression and poor prognosis.The result of the weighted gene co-expression network analysis identified 35 genes which can be enriched in the exhausted cells.The gene set can distinguish the exhausted cells obviously and had poor prognosis in many cancers.In addition to the reported exhausted markers,CD69 and IL2RB also contained more connection nodes.Transcription factors regulating CD8+exhausted T cells mainly included IRF4 and BATF,which can be combined with the promoter of PDCD1 to regulate their transcription.In addition,GSVA scores of the main pathways were calculated,showing that the exhausted cells were significantly up-regulated in the process of hypoxia,glycolysis,cholesterol,reactive oxygen,and endoplasmic reticulum stress.Combined with the above results,the potential regulatory network of the exhausted cells was speculated.Thirdly,the analysis of effector and exhausted cells in CD4 + T cells showed that there were 86 genes up regulated in the exhausted cells,49 of them were also found in CD8+exhausted cells.RNF19 A encodes a ubiquitin ligase,which showed a greater fold change in CD4+ exhausted cells.Both this research and verification datasets showed higher expression and poor prognosis.Considering that the gene from the same family participated in the T cell exhaustion,the role of RNF19 A was suggested to be involved in the formation of exhausted cells.The protein-protein interaction showed that AHR and ANXA5 were enriched in the functional modules including inhibitory receptors with immunosuppressive effect.The transcription factors that regulated CD4+ exhausted cells,including PRDM1,EGR2 and FOXO1,can promote the expression of PDCD1.The results showed that CD4+ exhausted cells were up regulated in hypoxia,glycolysis and cholesterol,but not in the reactive oxygen response and endoplasmic reticulum stress,suggesting that CD4+ and CD8+ exhausted cells were different.In conclusion,the genes related to T cell exhaustion were analyzed by bioinformatics methods,and some key genes,such as RGS1 and RNF19 A,can be candidate targes to solve T cell exhaustion,a gene set including 35 genes can distinguish the exhausted cells significantly,providing theoretical basis for research and immunotherapy of exhausted cells.