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Metastatic pheochromocytoma and paraganglioma (PPGL) have poor prognosis and limited therapeutic options. The recent advent of immunotherapies showing remarkable clinical efficacies against various cancer types offers the possibility of novel opportunities also for metastatic PPGL. Most PPGLs are pathogenically linked to inactivating mutations in genes encoding different succinate dehydrogenase (SDH) subunits. This causes activation of the hypoxia-inducible factor 2 (HIF2)-mediated transcriptional program in the absence of decreased intratumoral oxygen levels, a phenomenon known as pseudohypoxia. Genuine hypoxia in a tumor creates an immunosuppressive tumor microenvironment. However, the impact of pseudohypoxia in the immune landscape of tumors remains largely unexplored. In this study, tumoral expression of programmed death-ligand 1 (PD-L1) and HIF2α and tumor infiltration of CD8 T lymphocytes (CTLs) were examined in PPGL specimens from 102 patients. We assessed associations between PD-L1, CTL infiltration, HIF2α expression, and the mutational status of SDH genes. Our results show that high PD-L1 expression levels in tumor cells and CTL tumor infiltration were more frequent in metastatic than nonmetastatic PPGL. However, this phenotype was negatively associated with SDH mutations and high HIF2α protein expression. These data were validated by analysis of mRNA levels of genes expressing PD-L1, CD8, and HIF2α in PPGL included in The Cancer Genome Atlas database. Further, PD-L1 and CD8 expression was lower in norepinephrine than epinephrine-secreting PPGL. This in silico analysis also revealed the low PD-L1 or CD8 expression levels in tumors with inactivating mutations in VHL or activating mutations in the HIF2α-coding gene, EPAS1, which, together with SDH-mutated tumors, comprise the pseudohypoxic molecular subtype of PPGL. These findings suggest that pseudohypoxic tumor cells induce extrinsic signaling toward the immune cells promoting the development of an immunosuppressive environment. It also provides compelling support to explore the differential response of metastatic PPGL to immune checkpoint inhibitors. © 2022 The Authors. The Journal of Pathology published by John WileySons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Hypoxia-inducible factors (HIF) 2α and 1α are the major oxygen-sensing molecules in eukaryotic cells. HIF2α has been pathogenically linked to paraganglioma and pheochromocytoma (PPGL) arising in sympathetic paraganglia or the adrenal medulla (AM), respectively. However, its involvement in the pathogenesis of paraganglioma arising in the carotid body (CB) or other parasympathetic ganglia in the head and neck (HNPGL) remains to be defined. Here, we retrospectively analyzed HIF2α by immunohistochemistry in 62 PPGL/HNPGL and human CB and AM, and comprehensively evaluated the HIF-related transcriptome of 202 published PPGL/HNPGL. We report that HIF2α is barely detected in the AM, but accumulates at high levels in PPGL, mostly (but not exclusively) in those with loss-of-function mutations in VHL and genes encoding components of the succinate dehydrogenase (SDH) complex. This is associated with upregulation of EPAS1 and the HIF2α-regulated genes COX4I2 and ADORA2A. In contrast, HIF2α and HIF2α-regulated genes are highly expressed in CB and HNPGL, irrespective of VHL and SDH dysfunctions. We also found that HIF2α and HIF1α protein expressions are not correlated in PPGL nor HNPGL. In addition, HIF1α-target genes are almost exclusively overexpressed in VHL-mutated HNPGL/PPGL. Collectively, the data suggest that involvement of HIF2α in the physiology and tumor pathology of human paraganglia is organ-of-origin-dependent and HIF1α-independent.