Investigating the impact of regulatory B cells and regulatory B cell-related genes on bladder cancer progression and immunotherapeutic sensitivity.

BACKGROUND: Regulatory B cells (Bregs), a specialized subset of B cells that modulate immune responses and maintain immune tolerance in malignant tumors, have not been extensively investigated in the context of bladder cancer (BLCA). This study aims to elucidate the roles of Bregs and Breg-related genes in BLCA. METHODS: We assessed Breg infiltration levels in 34 pairs of BLCA and corresponding paracancerous tissues using immunohistochemical staining. We conducted transwell and wound healing assays to evaluate the impact of Bregs on the malignant phenotype of SW780 and T24 cells. Breg-related genes were identified through gene sets and transcriptional analysis. The TCGA-BLCA cohort served as the training set, while the IMvigor210 and 5 GEO cohorts were used as external validation sets. We employed LASSO regression and random forest for feature selection and developed a risk signature using Cox regression. Primary validation of the risk signature was performed through immunohistochemical staining and RT-qPCR experiments using the 34 local BLCA samples. Additionally, we employed transfection assays and flow cytometry to investigate Breg expansion ability and immunosuppressive functions. RESULTS: Breg levels in BLCA tissues were significantly elevated compared to paracancerous tissues (P < 0.05) and positively correlated with tumor malignancy (P < 0.05). Co-incubation of SW780 and T24 cells with Bregs resulted in enhanced invasion and migration abilities (all P < 0.05). We identified 27 Breg-related genes, including CD96, OAS1, and CSH1, which were integrated into the risk signature. This signature demonstrated robust prognostic classification across the 6 cohorts (pooled HR = 2.25, 95% CI = 1.52-3.33). Moreover, the signature exhibited positive associations with advanced tumor stage (P < 0.001) and Breg infiltration ratios (P < 0.05) in the local samples. Furthermore, the signature successfully predicted immunotherapeutic sensitivity in three cohorts (all P < 0.05). Knockdown of CSH1 in B cells increased Breg phenotype and enhanced suppressive ability against CD8 + T cells (all P < 0.05). CONCLUSIONS: Bregs play a pro-tumor role in the development of BLCA. The Breg-related gene signature established in this study holds great potential as a valuable tool for evaluating prognosis and predicting immunotherapeutic response in BLCA patients.

Development and experimental validation of an M2 macrophage and platelet-associated gene signature to predict prognosis and immunotherapy sensitivity in bladder cancer.

Platelets and M2 macrophages both play crucial roles in tumorigenesis, but their relationship and the prognosis value of the relative genes in bladder cancer (BLCA) remain obscure. In the present study, we found that platelets stimulated by BLCA cell lines could promote M2 macrophage polarization, and platelets were significantly associated with the infiltration of M2 macrophages in BLCA samples. Through the bioinformatic analyses, A2M, TGFB3, and MYLK, which were associated with platelets and M2 macrophages, were identified and verified in vitro and then included in the predictive model. A platelet and M2 macrophage-related gene signature was constructed to evaluate the prognosis and immunotherapeutic sensitivity, helping to guide personalized treatment and to disclose the underlying mechanisms.

Construction and experimental validation of a B cell-related gene signature to predict the prognosis and immunotherapeutic sensitivity in bladder cancer.

BACKGROUND: B cells are essential components of tumor microenvironment and exert important functions in anti-tumor immune response. However, the prognosis value of B cell-related genes in bladder cancer (BLCA) remains obscure. MATERIALS AND METHODS: The infiltrating levels of B cells were measured via the CD20 staining in the local samples and the computational biology analyses in the TCGA-BLCA cohort. The single-cell RNA sequencing analysis, gene-pair strategy, LASSO regression, random forest, and Cox regression were used for B cell-related signature construction. TCGA-BLCA cohort was chosen as the training cohort, and three independent cohorts from GEO and the local cohort were used for external validation. 326 B cells were adopted to explore the association between the model and B cells' biological processes. TIDE algorithm and two BLCA cohorts receiving anti-PD1/PDL1 treatment were utilized to detect its predictive ability to the immunotherapeutic response. RESULTS: High infiltration levels of B cells heralded favorable prognosis, both in the TCGA-BLCA cohort and the local cohort (all P < 0.05). A 5-gene-pair model was established and served as a significant prognosis predictor across multiple cohorts (pooled hazard ratio = 2.79, 95% confidence interval = 2.22-3.49). The model could evaluate the prognosis effectively in 21 of 33 cancer types (P < 0.05). The signature was negatively associated with B cells' activation, proliferation, and infiltrating levels, and could serve as a potential predictor of immunotherapeutic outcomes. CONCLUSIONS: A B cell-related gene signature was constructed to predict the prognosis and immunotherapeutic sensitivity in BLCA, helping to guide the personalized treatment.

Construction and experimental validation of a B cell senescence-related gene signature to evaluate prognosis and immunotherapeutic sensitivity in bladder cancer.

Senescent B cells exhibit reduced antibody production and enhanced proinflammatory cytokine and chemokine secretion, exerting non-negligible functions in antitumor immunity. This study aims to clarify the prognosis value of B cell senescence-related genes in bladder cancer (BLCA). Twelve B cell senescence-related genes were identified based on previous studies and the single-cell RNA sequencing of a BLCA sample from Gene Expression Omnibus (GEO). The Cancer Genome Atlas BLCA cohort was used as the training dataset. Three cohorts from GEO, 35 clinical samples from the local hospital, and in vitro cell experiments were used for validation. The unsupervised clustering based on the 12 genes was associated with the prognosis and the tumor immunity. Through least absolute shrinkage and selection operator regression and random forest algorithm, G protein subunit gamma 11 (GNG11) and inhibitor of DNA binding 1 (ID1) of the 12 genes were determined as significant prognosis predictors and then included in the multivariate Cox regression model. The model was a reliable and robust prognosis biomarker across multiple large-scale cohorts (pooled HR = 1.76, 95% CI = 1.41-2.20). The tight association between the model and BLCA malignant degree was demonstrated in the local cohort (P < 0.01). The model could also predict the immunotherapeutic sensitivity, which was confirmed by the tumor immune dysfunction and exclusion algorithm (P < 0.0001) and IMvigor210 cohort (P < 0.0001). At last, in vitro cell experiments in IM-9 and GM12878 B cells indicated that GNG11 and ID1 were involved in the cellular aging process. Collectively, a B cell senescence-related gene signature was constructed to evaluate the prognosis and immunotherapeutic response in BLCA, providing novel insights into the biological mechanisms.