Association of leucocyte levels in urine with tissue PD-L1 status and immune infiltration into basal bladder cancer subtype in the prospective real-world clinicopathological register trial Bladder BRIDGister.

Abstract

560 Background: The objective of the present study was to evaluate whether preoperative standard urine parameters such as leucocyte count in urine samples dominate in particular molecular subtypes of bladder cancer and reflect the immune infiltration status in the tissue in matched urine and TUR biopsy samples from patients being suspicious of bladder cancer and undergoing first TURB within the prospective Real World Experience registry trial "BRIDGister". Methods: For this pilot study paraffin fixed pretreatment tissue samples from the first TURB of 48 pts participating in the BRIDGister trial and matched urine samples were prospectively collected and analyzed. RNA from FFPE tissues were extracted by commercial kits- Relative gene expression of subtyping markers (KRT5, KRT20) by standardized RT-qPCR systems for PD-1, PD-L1, CTLA4 (STRATIFYER Molecular Pathology GmbH, Cologne). In addition urine samples were analyzed for leucocyte and erythrocyte count. Spearman correlation, Kruskal-Wallis, MannWhitney and Sensitivity/Specificity tests were done by JMP 9.0.0 (SAS software). Results: The pilot cohort of the BRIDGister trial consisted of 48 patients (median age: 77, male 65% vs. female 35%) of diverse clinical stages (Benign lesions/no tumor 38%, pTa 23%, pT1 20%, pT2 19%) and WHO 1973 grade (G1 11%, G2 43%, G3 23%). Presence of leucocytes but not erythrocytes was negatively associated with KRT20 (r=-0.4249, p=0.0216) but positive with KRT5 (r=0.3704, p=0.0479). Moreover high levels of leucocytes in urine were positively associated with tumor expression of PD-L1 (r=0.5081, p=0.0041), PD-1 (r=0.5342, p=0.00024) and CTLA4 (r=0.4244, p=0.0194). In contrast there was no significant association of tumor PDL-1, PD-1 and CTLA4 expression with erythrocyte count in urine. Conclusions: Presence of leucotytes in urine is strongly associated with basal subtype and immune cell infiltration into early bladder cancer. As the presence of erythrcytes did not reveal these significant associations, the presence of leucocytes is not due to simple bleeding or tissue vulnerability. Moreover, the strong tumor subtype specificity further demonstrate the basal tumor specificity of urine leucocytes and therefore may be helpful for detecting and monitoring basal bladder cancer in a non invasive fashion. Given the prognostic value of tissue determination of PD-L1, PD-1 and CTLA4 quantitation on mRNA level, these results warrant further investigation to conclude on its impact on outcome prediction (BCG responsiveness, recurrence, progression, etc.), which will be prospectively analyzed in the framework of the ongoing multicenter BRIDGister Real World Experience trial.