Intravesical Bacillus Calmette-Guérin vaccine (BCG) is an established immunotherapeutic in bladder cancer (BlCa), provoking inflammation leading to tumour-specific immunity. Immune checkpoint blockers such as anti-PD-L1 have potential for enhancing tumour-specific lymphocyte-mediated cytotoxicity in BCG-refractive or advanced disease. In both cases, Interferon-gamma (IFNγ) plays a central role. We investigated the transcriptomic response of normal human urothelium to IFNγ to disentangle mechanisms of BCG and anti-PD-L1 therapy failure. Exposure of differentiated human urothelium to IFNγ resulted in upregulated MHC class I and class II and de novo expression of CXCL9-11 chemokine genes. Normal urothelium expressed only immuno-inhibitory B7 family members: PD-L1 expression was induced by IFNγ, whereas VISTA was expressed constitutively. A urothelial IFNγ response gene set was derived and used for unsupervised clustering of tumours, which predicted longer recurrence-free survival in non-muscle invasive bladder cancer (NMIBC). In muscle invasive bladder cancer (MIBC), the IFNγ-signature split the basal/squamous consensus subtype, with significantly worse overall survival when weak/absent. Normal urothelium has few resident lymphocytes. Tumour cell killing requires recruitment and activation of IFNγ-secreting pro-inflammatory/cytotoxic lymphocytes while surmounting both innate (VISTA) and upregulated (PD-L1) inhibitory mechanisms. This study offers supportive evidence for strategies to enhance immunotherapy via the IFNγ and VISTA/PD-L1 nexus.

The authors have declared no competing interest.

This study was funded by York Against Cancer.

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The cell lines were established as described [1] using anonymous discarded tissue from renal transplant surgery, with UK NHS approval (REC reference 99/095) from the Leeds East Research Ethics Committee.

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