Preliminary Exploration Of CHIT1 As A Biomarker For Immunotherapy

Research Background:Tumor is one of the major chronic diseases that seriously threaten the lives and health of people.With the approval of immune checkpoint inhibitors(ICIs)such as programmed cell death 1(PD-1),programmed cell death-ligand 1(PD-L1),and cytotoxic T lymphocyte-associated antigen-4(CTLA-4)for clinical application,tumor clinical treatment is entering the era of immunotherapy.Immune checkpoints play a crucial role in maintaining self-tolerance,and during the body’s infection process,they can effectively avoid immune system damage to body tissues and prevent the occurrence of autoimmune diseases.Various immune checkpoints have been discovered that play a key role in regulating the activation and inhibition of immune pathways,forming a complex and extensive immune regulation network in the body.For tumor growth,how to evade immune system killing is a key issue for its survival and continued growth.Immune checkpoints provide a good opportunity for tumor immune escape,and they can also be used as drug targets in clinical practice,providing effective treatment options for cancer patients.Currently,the two commonly used targets are PD-1and CTLA-4,blocking these receptors in tumor-bearing animal models can effectively slow down tumor growth.PD-1 and CTLA-4 have different and complementary functions in the activation process of T cells.In clinical practice,combining the above two can promote T cell infiltration,increase the number of CD8~+T cells,and strengthen their function,leading to the release of more Interferon-γ(IFN-γ)and tumor necrosis factor-α(TNF-α).However,the reality of clinical application is that most patients do not benefit from ICI treatment,as evidenced by limited objective response rates(ORR),unavoidable immune-related adverse events(ir AEs),and the emergence of resistance and hyperprogression in some patients.Although the use of existing biomarkers,such as PD-L1expression of tumor tissue,mismatch repair defects,microsatellite instability,and tumor mutation burden,increased the ORR of ICIs to about 50%,almost half of the patients still do not benefit.These clinical realities prompt us to further explore the tumor immune microenvironment,search for new targets to enhance immune therapy,and discover new ICIs efficacy prediction biomarkers.Research objective:Based on the current publicly available ICIs treatment cohorts and The Cancer Genome Atlas(TCGA)database,screen for target genes that significantly prolonged the survival of patients treated with ICIs.Collecting clinical samples to verify the relationship between the protein level of the gene and the progression of ICIs treatment.Further analyze the relationship between the gene and the tumor immune microenvironment at both the overall transcriptional level and spatial transcriptional level,and explore the immune-related mechanisms by which it extends the survival of patients treated with ICIs.This gene is expected to provide candidate targets for population selection,prognosis prediction,and new drug development in ICIs treatment,thus providing a theoretical basis for improving clinical treatment efficacy and patient prognosis.Research Methods1.Data Download:1.1 Collect and organize transcriptome data and clinical data of 33 different types of cancer patients from TCGA database;1.2 Collect and organize transcriptome data and clinical data of 7 ICIs treatment cohorts;1.3 Collect and organize 30 publicly available and usable spatial transcriptome data.2.Differential Gene Analysis:In the public ICIs treatment cohorts,we used the median overall survival(OS)as the grouping standard,divided them into long survival group and short survival group,and used the edge R package(DOI:10.18129/B9.bioc.edge R)in R(4.1.0)to perform differential gene analysis,screening for target genes associated with long survival.3.Survival analysis:We divided the patients into high and low expression groups based on the median expression level of the target gene,calculated the survival curves of the ICIs treatm ent cohorts using the product limit method.We also calculated the hazard ratio of the target gene using the Cox proportional hazards model.4.Clinical sample validation:We enrolled 26 patients who received anti-PD-1 antibody treatment at Xinqiao Hospital and collected serum samples before ICIs treatment.The chitotriosidase concentration in the peripheral blood of the patients were detected by enzyme-linked immunosorbent assay(ELISA),and survival analysis was performed based on the patients’progression-free survival(PFS).5.Gene enrichment analysis:We collected the activated CD8~+T cell gene set,the classical hallmark inflammatory response gene set,the classical hallmark interferon gamma response gene set,and the IFN-gamma-related gene set.The genes were divided into two groups based on the median expression of the target gene,and gene set enrichment analysis(GSEA)was used to calculate the enrichment of the four gene sets in the public cohorts.Single-sample GSEA(ss GSEA)was used to calculate the scores of each gene set in the cohort,and Spearman correlation analysis was used to evaluate the correlation between the target gene and immune-related pathways.6.Spatial transcriptome analysis:We used the bivariate Moran’s I and local indicators of spatial association(LISA)to evaluate the spatial correlation between the target gene and the hot-cold tumor immune microenvironment in 30 public spatial transcriptome datasets.Research Results:1.Differential gene analysis of the discovery cohorts(Van Allen cohort and Liu cohort)identified 5 genes positively correlated with long-term survival.2.Survival analysis results showed that in the discovery cohorts,only grouping by high or low CHIT1 expression had a statistically significant difference i n OS,with patients in the high expression group having longer OS.3.In the multi-tumor validation cohorts,grouping by high or low CHIT1 expression showed a statistically significant difference in OS and PFS between the two groups.Analysis using a Cox proportional hazards model revealed that the hazard ratio(HR)of CHIT1 was less than 1 in the ICIs treatment cohort,suggesting that CHIT1 is a protective factor.4.Serum chitotriosidase concentration of the Xin Qiao cohort patients was detected by ELISA.The cohort was then divided into high and low concentration groups based on a cutoff of 20ng/m L.Survival analysis showed that patients in the high concentration group had longer PFS and better survival prognosis.5.GSEA analysis of the tumor immune microenvironment revealed that the gene sets related to hot tumor immune microenvironments were enriched in the CHIT1 high concentration group of the ICIs treatment cohort and TCGA database.6.Spatial transcriptomic analysis revealed a spatial positive correl ation between CHIT1 expression and the hot tumor immune microenvironment represented by the IFN-γrelated gene set.Conclusions:1.In the ICIs-treated discovery cohort,CHIT1 was significantly positively correlated with prolonged survival.2.In the multi-tumor validation cohort,patients with high CHIT1 expression had significantly prolonged survival.3.CHIT1 was positively correlated with hot tumor immune microenvironment in immunotherapy and pan-cancer cohorts.