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MP63-17 MITOMYCIN C TRIGGERS IMMUNOGENIC CELL DEATH IN BLADDER CANCER CELLS

INTRODUCTION AND OBJECTIVES: Despite preventing recurrence of non-muscle invasive bladder cancer (BC), the chemotherapeutic drug Mitomycin C(MMC) is effective only in a proportion of patients. We speculated that treatment success might not only rely on MMC cytotoxic potential, but also on its ability to trigger immunogenic cell death (ICD). In this study, we assessed MMC-induced ICD in a panel of BC cell lines and patient-derived samples. Moreover, we investigated the molecular changes in ICD-responsive versus not-responsive BC cells to identify potential underlying mechanisms of MMC efficacy. METHODS: Human BC cell lines and cancer samples (stage Ta, T1 retrieved during transurethral resection- ICH-MIM-01) were treated or not with MMC for 1h and expression of ICD markers was then assessed by flow cytometry and enzyme-linked immunosorbent assay. The ability of dendritic cells (DCs) to phagocytose tumor cells and undergo maturation was analyzed after 2h and 24h coculture by flow cytometry. Illumina mRNA sequencing was used to screen for differentially expressed genes between ICD-responsive and not responsive BC cell lines. RESULTS: Treatment with MMC fostered ICD in BC cells by inducing endoplasmic reticulum stress response, increased secretion of the damage signal high-mobility-group box 1 (HMGB1), and augmented DC-mediated phagocytic clearance of treated tumor cells compared to untreated cells. By scoring MMC-induced ICD responses in a panel of different BC cell lines we identified ICD-responsive and ICD-not responsive cells, which we analyzed by mRNA sequencing. We found an upregulation of apoptosis and TNF-signaling pathways and a downregulation of protein processing in ICD-responsive cells, indicating that pro-apoptotic signals and local inflammation are necessary for MMC-induced ICD. Consistently, ex-vivo treatment of patient-derived tumors with MMC induced expression of ICD markers in some of the samples. Importantly, only BC cells and tumor tissues undergoing ICD upon MMC treatment, as measured by increased secretion of HMGB1 and pro-inflammatory molecules, could promote DC maturation, a crucial step for proper priming of adaptive immunity. CONCLUSIONS: Overall, we showed a novel immune-related mechanism of action of MMC that may be exploited to optimize bladder cancer management and identify possible ICD-related biomarkers. Figure. No caption available. Source of Funding: Ricerca Finalizzata, Italian Ministry of Health