Abstract: Objective To investigate the role and clinical significance of the loss of  Y chromosome (LOY) in bladder cancer. Methods  Tissue samples and clinical data were collected from 44 male patients with bladder cancer undergoing radical cystectomy at the First Affiliated Hospital of Guangxi Medical University between January 2018 and October 2019. Whole⁃transcriptome sequencing data were analyzed and scored based on Y chromosome transcriptional signature (YchrS） using single⁃sample gene set enrichment analysis (ssGSEA). Patients were stratified into high and low YchrS score groups. The prognosis of the two groups were evaluated using the Kaplan⁃Meier method, and the potential molecular characteristics were revealed via differential expression analysis and gene set enrichment analysis （GSEA）. The diversity of immune cells in the two groups was analyzed using cytometry by time of flight (CyTOF). Single⁃cell transcriptomic sequencing data from a public database (HRA000212) was used to characterize LOY epithelial cells through cell differentiation trajectories and cell communication analysis. The CMap  database was utilized to identify potential therapeutic agents. Results  Patients with bladder cancer in the low YchrS score group exhibited shorter overall survival (P=0.006) and higher tumor grade (P=0.036). Upregulated genes in low YchrS score group were enriched in biological processes related to the cell cycle regulation, DNA replication, and chromosome segregation (P<0.05), showing elevated scores for p53⁃repressed genes and reduced scores for p53⁃induced genes (all P<0.001). Compared with other epithelial cells, LOY epithelial cells displayed activation of immune⁃related signaling pathways and stronger interactions with myeloid cells and T cells. CyTOF analysis revealed an enrichment of regulatory T (Treg) cells and upregulated expression of CD279 (PD⁃1) on CD4+ T cells in the low YchrS score group. CMap analysis indicated that cyclin⁃dependent kinase (CDK) inhibitors (e.g., Palbociclib) and histone deacetylase (HDAC) inhibitors were potential therapeutic agents for bladder cancer patients in the low YchrS score group. Conclusions LOY is strongly associated with poor prognosis in bladder cancer patients, with its tumorigenic mechanism may involve p53 suppression and immune suppression. CDK inhibitors and HDAC inhibitors serve as potential therapeutic agents for the LOY bladder cancer patients.

Key words: Bladder cancer, Loss of Y chromosome, Tumor microenvironment, Regulatory T cells