Presence of CD8+ cytotoxic T-cells in proximity to tumor cells, TIM3, CTLA-4 and prognostic implications for muscle-invasive urothelial cancer.

Abstract

4582 Background: Although it is well known that the quantity of specific immune cell types can provide prognostic information in bladder cancer, only little is known about the prognostic relevance of the localization of immune cells within the tumor microenvironment. Methods: To study the spatial interplay of different immune cell subpopulations and its prognostic role in bladder cancer, 2463 urothelial bladder carcinomas were immunostained with 21 antibodies (i.e., TIM-3, PD-L1, PD-1, CTLA4, CD3, CD8, CD4, FOXP3, CD20, CD68, CD163, CD11c, panCK, Ki67, CD31, Vimentin, HLA-DRa, Myosin-11, Desmoglein 3, PAX-8, and CDH16) using BLEACH&STAIN multiplex fluorescence immunohistochemistry in a tissue microarray format and analyzed using a framework of neuronal networks for image analysis. Thus, Spatial immune parameters were compared with histopathological parameters and overall survival data. Results: The identification of more than 300 different immune cell subpopulations and the characterization of their spatial relationship resulted in numerous spatial interaction patterns. 32 immune parameters showed prognostic significance in univariate analyses of which 16 were independent from pT, pN, and histologic grade in muscle-invasive bladder cancer. The strongest association to clinical outcome was identified for intraepithelial CD8+ cytotoxic T-cells (AUC: 0.70) while stromal CD8+ cells were less relevant (AUC: 0.64, p=0.016). A favorable prognosis of inflamed cancers with high levels of “exhaustion markers” suggests that TIM3, PD-L1, PD-1, and CTLA-4 on inflammatory cells do not hinder anti-tumoral immune response in tumors rich of tumor infiltrating tumor cells. Conclusions: The density of intraepithelial CD8+ T-cells was the strongest prognostic feature in muscle-invasive bladder cancer. Given that tumor cell killing by CD8+ cytotoxic T-lymphocytes through direct cell-to-cell-contacts represents the “terminal end route” of anti-tumor immunity, the quantity of “tumor-cell adjacent CD8+ T-cells” may constitute a surrogate for the efficiency of cancer recognition by the immune system that can be measured straightaway using conventional brightfield IHC in routine pathology as the CD8 labelling index.