Renal Cell Carcinoma Programmed Death-ligand 1, a New Direct Target of Hypoxia-inducible Factor-2 Alpha, is Regulated by von Hippel-Lindau Gene Mutation Status

Background: Clear cell renal cell carcinomas (ccRCC) frequently display a loss of function of the von Hippel-Lindau (VHL) gene. Objective: To elucidate the putative relationship between VHL mutation status and immune checkpoint ligand programmed death-ligand 1 (PD-L1) expression. Design, setting, and participants: A series of 32 renal tumors composed of 11 VHL tumor-associated and 21 sporadic RCCs were used to evaluate PD-L1 expression levels after sequencing of the three exons and exon-intron junctions of the VHL gene. The 786-O, A498, and RCC4 cell lines were used to investigate the mechanisms of PD-L1 regulation. Outcome measurements and statistical analysis: Fisher's exact test was used for VHL mutation and Kruskal-Wallis test for PD-L1 expression. If no covariate accounted for the association of VHL and PD-L1, then a Kruskal-Wallis test was used; otherwise Cochran-Mantel-Haenzsel test was used. We also used the Fligner-Policello test to compare two medians when the distributions had different dispersions. Results and limitations: We demonstrated that tumors from ccRCC patients with VHL biallelic inactivation (ie, loss of function) display a significant increase in PD-Ll expression compared with ccRCC tumors carrying one VHL wild-type allele. Using the inducible VHL 786-O-derived cell lines with varying hypoxia-inducible factor-2 alpha (HIF-2 alpha) stabilization levels, we showed that PD-Ll expression levels positively correlate with VHL mutation and HIF-2 alpha expression. Targeting HIF-2 alpha decreased PD-L1, while HIF-2 alpha overexpression increased PD-Ll mRNA and protein levels in ccRCC cells. Interestingly, chromatin immunoprecipitation and luciferase assays revealed a direct binding of HIF-2 alpha to a transcriptionally active hypoxia-response element in the human PD-Ll proximal promoter in 786-O cells. Conclusions: Our work provides the first evidence that VHL mutations positively correlate with PD-Ll expression in ccRCC and may influence the response to ccRCC anti-PD-Ll/PD-1 immunotherapy. Patient summary: We investigated the relationship between von Hippel-Lindau mutations and programmed death-ligand 1 expression. We demonstrated that von Hippel-Lindau mutation status significantly correlated with programmed death-ligand 1 expression in clear cell renal cell carcinomas. (C) 2015 European Association of Urology. Published by Elsevier B.V. All rights reserved.