Immune senescence and antigen presentation defects define a pre-existing state of BCG unresponsiveness in bladder cancer

Background: Bacillus Calmette-Guérin (BCG) is standard therapy for high-risk non-muscle invasive bladder cancer (NMIBC), yet a substantial proportion of patients fail to respond. Although T-cell dysfunction and impaired antigen presentation have been implicated in BCG resistance, their pre-treatment presence and clinical relevance remain unclear. We aimed to define a reproducible, treatment-specific transcriptomic signature of BCG unresponsiveness centered on antigen presentation machinery (APM) deficiency. Methods: We performed single-cell RNA sequencing (scRNA-seq) on 10 pre-treatment NMIBC tumors to identify cellular programs associated with BCG outcomes. Tumor-intrinsic APM activity and interferon pathway status were quantified at the independent bulk transcriptomic cohorts (UROMOL and BRS). Pre-treatment urine samples were assessed for a targeted chemokine panel (IL-6, IL-8, IFN-gamma, CXCL10, CXCL13, CCL5, CCL21) to determine concordance between transcriptomic IFN/APM activity and protein-level immune signaling. Results: Single-cell profiling revealed that BCG-unresponsive tumors exhibited markedly reduced epithelial APM expression accompanied by suppression of type I/II interferon signaling and loss of plasmacytoid dendritic cell activity. These transcriptomic features were reflected at the protein level: non-responders displayed significantly lower urine chemokine levels prior to BCG instillation. Bulk transcriptomic validation across >350 patients demonstrated that high APM activity robustly predicted improved recurrence-free survival in BCG-treated patients (p < 0.05). Conclusions: BCG-unresponsive NMIBC is defined by a pre-existing antigen presentation defect coupled with suppressed interferon signaling, detectable at both the transcriptomic and urine protein levels. The APM signature exhibits strong, treatment-specific predictive value across multi-cohort datasets and provides a biologically compelling rationale for APM-restoring or IFN-enhancing intravesical immunotherapies as alternatives for patients unlikely to benefit from BCG. Acknowledgments : The results shown here are in part based upon data generated by the TCGA Research Network: https://www.cancer.gov/tcga. We thank the staff of the Department of Urology and Laboratory Medicine at Yonsei University College of Medicine for their technical assistance. We also thank all the patients who participated in this study. This study was supported by grants from the National Research Foundation of Korea (NRF) grants funded by the Korean government (MSIT) (grant numbers: 2022R1A2C2003831). This work was also supported by the National Research Foundation of Korea (NRF) funded by the Ministry of Education (RS-2018-NR031072).