Immune characterization of urothelial bladder carcinoma integrating transurethral resection of bladder tumor (TURBT) samples and serum: A feasibility study.

538 Background: Urothelial bladder cancer (UBC) is characterized by high immunogenicity. Multiple immunotherapies both in the perioperative and advanced setting are being developed. However, we lack good predictive biomarkers of response to immunotherapy in UBC. We aimed to characterize the immune profile of UBC patients using TURBT and serum samples. Methods: 13 UBC biopsies from TURBT samples were analyzed along with sera of these patients. Biopsy samples were divided in two equal parts. One part was formalin-fixed paraffin embedded and stained for immune histochemistry [IHC] for the following markers (CD3, CD4, CD8, CD20, CD68 and PD-L1 using SP142 antibody) and results expressed in percentages. The other part was homogenized and processed for immune cell isolation (human tumor isolation Miltenyi Kit). Tumor cells were stained with PHK and double positive CD8 and PHK were isolated using a MACSTM procedure. Different immune cell populations were analyzed by flow cytometry for the following markers (CD3, CD4, CD8, CD20 and CD68) and results expressed in percentages. Cytokines were examined in sera using the Luminex multiparametric procedure and results were expressed as pg/mL. Results: IHC and flow cytometry data were comparable and identified three groups according to the immune cells associated with the tumors (Table). Groups 1 and 2 showed high and moderate levels of tumor activated CD8+ T cells (1-4%) along with a higher expression of PD-L1 (40-20%). Group 3 lacked tumor activated CD8+ T cells and show lower expression of PD-L1 (15%). All groups showed significant levels of Th2 cytokines (IL-4, IL-6 or IL-10) while very low or lack of Th1 cytokines (IFN or IL-2) was observed except in group 2. Conclusions: TURBT biopsies and serum analysis of cytokines could represent a valid approach to further evaluate potential predictive biomarkers of response to immunotherapy in UBC. Different immune populations subgroups were identified in our series and could potentially predict for diverse responses to treatment. Further prospective validation in larger series is ongoing to confirm these results.[Table: see text]