Clinical relevance and prognostic significance of PD-1/PD-Ls in non-metastatic bladder cancer: A role for PD-L2.

BACKGROUND Bladder cancer (BC) can be successfully treated by manipulating immune responses with intravesical Bacillus Calmette-Guerin instillation or targeting the PD-1/PD-L signaling pathway. In the present study we investigated the prognostic significance of the immune checkpoint inhibitor PD-1 and its ligands PD-L1 and PD-L2 on tumor cells and infiltrating lymphocytes, in the tumor microenvironment and draining lymph nodes in patients with non-metastatic BC. PATIENTS AND METHODS Cells were mechanically isolated from tissues and draining lymph nodes from 58 patients, and surface-stained for CD45, PD-1, PD-L1 and PD-L2. The cells were then analyzed with a flow cytometric method. RESULTS Approximately 2% of CD45-negative tumor and stromal cells expressed PD-L1. Expression was not associated with the main clinicopathological characteristics of the disease or with survival. However, as tumors progressed the frequency of PD-L1+CD45hi cells and the mean expression of PD-1 on CD45hi cells increased remarkably on immune cells in tumor tissues and draining lymph nodes. In addition, frequency analysis showed that cell percentages as well as mean expression of PD-L2 on total CD45+ lymphocytes and their CD45hi subpopulation in tumor-draining lymph nodes was significantly associated with cancer-related death (P < 0.05). Multiple Cox regression also revealed that while CD45+ (hazard ratio: 0.596, 95 % CI 0.439-0.809, P = 0.001) was associated with improved survival, CD45neg (HR: 0.615, 95 % CI 0.454-0.831, P = 0.002), and PD-L2+CD45+ cells (hazard ratio: 1.472, 95 % CI 1.023-2.120, P = 0.038) in draining lymph nodes were associated with lower survival. CONCLUSION Our results suggest that in patients with BC, PD-1 and PD-L expression on immune cells, especially in draining lymph nodes, is valuable for predicting prognosis and survival, and possibly responsiveness to immunotherapy. However, expression of the inhibitor molecule or its ligands on tumor cells was not associated with prognosis. The results highlight the significance of PD-L2 as a second important suppressive molecule in tumors.