Investigating sexual dimorphism in the tumour immune microenvironment of non-muscle invasive bladder cancer

Introduction The incidence of urothelial carcinoma of the bladder is four times higher in men compared to women. However, women with bladder cancer present with more aggressive disease and do not respond as well to Bacille Calmette-Guerin immunotherapy, the gold-standard treatment for non-muscle invasive bladder cancer (NMIBC). Based on the established evidence on sexual dimorphism in physiologic immune responses in bladder mucosa, we hypothesized that sex associated bladder immune physiology may influence the tumour immune microenvironment (TIME) and clinical outcomes. Methods We interrogated the expression patterns of genes associated with immune cell phenotypes (T cells, B cells and macrophages) and immune checkpoint pathways using tumour whole transcriptome profiles from male (n=376) and female (n=109) NMIBC patients. In an independent cohort of 390 tumours (n=85 females and n=305 males) on a tissue microarray, we conducted multiplexed immunofluorescence to evaluate the density and spatial distribution of CD163+ M2 tumour associated macrophages, CD3+CD8- (T helper), CD8+ (T cytotoxic), CD79a+ (B), CD103+ (T resident) cells and the immune checkpoint proteins PD-1 and PD-L1. Results High-grade tumours from female patients exhibited significantly increased expression of CD274 (PDL1), CTLA4 and IDO1 immune checkpoint genes. Significantly higher infiltration of CD163+ M2 macrophages in the epithelial and stromal compartments was observed in high-grade tumours from females. Significantly increased PD-L1 protein expression was also observed in the epithelial compartment of high-grade tumours from females compared to those from males. Higher infiltration of CD163+ M2 macrophages and CD79a+ B cells significantly associated with shorter recurrence-free survival in high-grade tumours irrespective of patient sex. Conclusions Findings from this study provide the first evidence supporting sexual dimorphism in the TIME of NMIBC. Future mechanistic studies are warranted to determine the potential roles of M2 tumour associated macrophages in promoting an aggressive disease behaviour and inferior clinical outcomes, specifically in female patients.