Differential HIF2α Protein Expression in Human Carotid Body and Adrenal Medulla under Physiologic and Tumorigenic Conditions

Abstract

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.

Pseudohypoxia in paraganglioma and pheochromocytoma is associated with an immunosuppressive phenotype

“…Analysis of HIF2α protein expression in 15 MPPGL tumor samples allowed us to confirm our previously reported data [ 7 ] showing that HIF2α protein expression was positively associated with SDH mutations ( P = 0.006) (Figure 5 ). We also found that the prevalence of tumors expressing PD‐L1 or with CTL infiltration was higher among tumors with negative than positive HIF2α immunostaining (Figure 5 ): 6/7 (85.7%) versus 1/9 (11%) expressed PD‐L1 ( P = 0.006) and 5/7 (71.4%) versus 1/7 (14.3%) had immunosuppressive phenotype ( P = 0.051) (Figure 5 ).…”

“…Increased EPAS1 mRNA levels are a feature of pseudohypoxic PPGL [ 7 ]. We found that high levels of expression of CD274 or CD8A genes, encoding PD‐L1 and CD8, respectively, were associated with low EPAS1 (encoding HIF2α) mRNA levels ( P < 0.0001 and P = 0.009, respectively) (Figure 6 ).…”

“…Although the role of hypoxia and HIF1α as a driving force of immunosuppressive tumor phenotypes is being resolved, it is unknown whether pseudohypoxic tumor cells also suppress tumor immunity. This is especially intriguing in PPGL where, as indicated earlier, activation of pseudohypoxia is mediated by HIF2α, rather than HIF1α [ 7 , 11 ]. Moreover, in contrast to truly hypoxic tumors, the HIF‐activated transcriptional program in PPGL occurs in tumor cells harboring genetic inactivation of VHL or SDH genes but not in the infiltrated immune cells.…”

“…PPGL are also pathogenically associated with somatic gain‐of‐function mutations of EPAS1 , the HIF2α encoding gene [ 6 ]. In addition, HIF2α, but not HIF1α, is overexpressed in pseudohypoxic PPGL [ 7 ] and in Sdhb knockout‐immortalized mouse chromaffin cells [ 8 ]. Thus, although the exact mechanism of PPGL tumor formation remains to be elucidated, proposed models include HIF2α‐ rather than HIF1α‐dependent pseudohypoxic drive [ 9 , 10 , 11 , 12 ].…”

Exploratory Analysis of the Gene Expression Matrix Based on Dual Conditional Dimensionality Reduction

“…The resulting dataset contains 157 samples of pheochromocytoma and paraganglioma (PCPG), some carrying mutations in hypoxia related genes such as VHL, SDH and EPAS1, 221 of kidney clear cell carcinoma (KIRC), and 71 of normal renal tissue. For each sample, we selected the expression levels of 129 miRNA and 442 hypoxia-related genes (those available after data curation from the list of 446 described in [24]), including the so-called canonical hypoxia genes, and others with special functions (angiogenesis, extracellular matrix, etc.).…”