Correlation Between Extracellular Matrix Protein COL6A1 And Malignant Progression,Prognosis And Immunotherapy Efficacy Of Bladder Cancer

Objective:Recent studies have shown that the interaction between tumor microenvironment and tumor cells is a necessary condition for malignant tumor progression and metastasis.Tumor microenvironment is a local internal environment composed of extracellular matrix,soluble molecules and tumor stromal cells.Among them,extracellular matrix(ECM)is an important framework of tumor microenvironment,it plays an important role in the growth and metastasis of cancer cells.Among them,human collagen VI(COL6A1)is a major extracellular matrix(ECM)protein,which plays an important role in the occurrence and metastasis of various solid tumors.However,the role of COL6A1 in the development and prognosis of bladder cancer is still lacking.Therefore,the aim of this study was to explore the correlation between extracellular matrix protein COL6A1 and malignant progression,prognosis and immunotherapy efficacy of bladder cancer.Methods:The clinicopathological information of bladder tumor patients were downloaded from the public database of The Cancer Genome Atlas(TCGA),ECM related differentially expressed genes were screened and determined from MSig DB molecular characteristic database.Liquid chromatography-mass spectrometry(LC-MS/MS analysis)was used to carry out proteomic analysis of tumor tissue specimens and normal bladder tissues of 10 bladder cancer patients in the Affiliated Hospital of Qingdao University,and further screen ECM related differentially expressed genes from protein level.Univariate Cox regression analysis was performed using R software to determine the prognosis related genes.Multivariate Cox regression analysis and Lasso regression analysis were used to establish the prognosis prediction signature based on ECM differentially expressed genes,draw the ROC curve,set and verify the area under curve(AUC),and detect the predictive value of the signature for clinicopathological features and prognosis.Meanwhile,we constructed a protein-protein interaction(PPI)network to screen the hub gene(COL6A1).We verified the correlation between the hub gene’s(COL6A1)expression in bladder cancer tissues and normal tissues and the prognosis by TCGA database and Gene Expression Omnibus(GEO)database.The clinical information of 71 patients receiving bladder cancer surgery in the Affiliated Hospital of Qingdao University were retrospectively analyzed.Immunohistochemistry was used to detect the expression of COL6A1 protein in 71 cases of bladder cancer and 10 cases of normal bladder tissue.The correlation between COL6A1 expression and clinicopathological features and prognosis of bladder cancer was analyzed.At the same time,a retrospective analysis of 30 patients with advanced bladder cancer who received immunotherapy with PD-1 immune checkpoint inhibitor Tislelizumab Injection was carried out in the Affiliated Hospital of Qingdao University.The efficacy of PD-1inhibitor immunotherapy was evaluated.Immunohistochemistry was used to detect the expression of COL6A1 protein in 30 cases of bladder cancer.The correlation between COL6A1 expression and efficacy of bladder cancer patients receiving PD-1 inhibitor immunotherapy was analyzed.Results:From the public database,225 ECM related genes of bladder cancer were screened out.Combined with proteomic analysis,71 ECM related differentially expressed genes of bladder cancer were finally identified.Univariate Cox regression analysis showed that a total of 24 ECM related differentially expressed genes had significant prognostic value in bladder cancer.The prognosis signature was constructed using R software,including 4ECM differentially expressed genes(MAP1B,FBN1,COL6A1,MFAP5).K-M survival curve analysis showed that the survival rate of patients in the high-risk group was significantly lower than that in the low-risk group.ROC curve analysis showed that AUC was 0.641,indicating that the prediction accuracy of the signature for survival results was good.The correlation between ECM risk signature and clinical characteristics was shown to be closely related to T stage,N stage and pathological grade of bladder cancer.Univariate Cox regression analysis showed that the age,grade,T stage,N stage and signature score were significantly correlated with survival rate of bladder cancer.Multivariate Cox regression analysis showed that the signature was an independent prognostic factor for bladder cancer.PPI confirmed that COL6A1 is an important prognostic risk gene for bladder cancer,the expression of COL6A1 and its relationship with prognosis was confirmed in TCGA bladder cancer cohort and GEO bladder cancer cohort.COL6A1 is recognized as an important prognostic risk gene in bladder cancer,and is closely related to the malignant progression and prognosis of bladder cancer.We further validated the expression of COL6A1 in bladder cancer tissues by immunohistochemistry.It was found that the expression level of COL6A1 in BC patients was significantly high,and the positive rate was 70.42%(50/71).Further analysis of the correlation between the expression level of COL6A1 and clinical pathological characteristics and prognosis of the patients in the cohort showed that there was no significant difference in age,gender,T stage and N stage,but was closely related to the metastasis of patients.The positive rate of COL6A1 expression in patients with metastatic bladder cancer was significantly higher than that in non-metastatic group.Meanwhile,KM survival analysis suggested that the overall survival of the high COL6A1 expression group was lower than that of the low expression group,suggesting that COL6A1 is closely related to the prognosis of bladder cancer patients.We also retrospectively analyzed the clinical data in 30 patients with advanced bladder cancer receiving immunotherapy.Among the 30 patients,2 achieved complete response(CR),7 partial response(PR),13 stable disease(SD)and 8 progressive disease(PD).In the low expression group of COL6A1,ORR was 60% and DCR was 90%,and in the COL6A1 high expression group,ORR was15% and DCR was 65%.In the 21 patients of group SD+PD,COL6A1 was highly expressed in 17 cases and low in 4 cases,suggesting that the expression level of COL6A1 is closely related to the reactivity and efficacy of immunotherapy in patients with advanced bladder cancer,and can be used as a potential biomarker for bladder cancer immunotherapy.Conclusion:COL6A1 is closely related to the malignancy progression,prognosis and immunotherapy efficacy of bladder cancer patients.It is a potential biomarker and deserves further study.