Identification of Novel Mutations and Expressions of EPAS1 in Phaeochromocytomas and Paragangliomas

Abstract

Endothelial PAS domain-containing protein 1 (EPAS1) is an oxygen-sensitive component of the hypoxia-inducible factors (HIFs) having reported implications in many cancers by inducing a pseudo-hypoxic microenvironment. However, the molecular dysregulation and clinical significance of EPAS1 has never been investigated in depth in phaeochromocytomas/paragangliomas. This study aims to identify EPAS1 mutations and alterations in DNA copy number, mRNA and protein expression in patients with phaeochromocytomas/paragangliomas. The association of molecular dysregulations of EPAS1 with clinicopathological factors in phaeochromocytomas and paragangliomas were also analysed. High-resolution melt-curve analysis followed by Sanger sequencing was used to detect mutations in EPAS1. EPAS1 DNA number changes and mRNA expressions were examined by polymerase chain reaction (PCR). Immunofluorescence assay was used to study EPAS1 protein expression. In phaeochromocytomas, 12% (n = 7/57) of patients had mutations in the EPAS1 sequence, which includes two novel mutations (c.1091A > T; p.Lys364Met and c.1129A > T; p.Ser377Cys). Contrastingly, in paragangliomas, 7% (n = 1/14) of patients had EPAS1 mutations and only the c.1091A > T; p.Lys364Met mutation was detected. In silico analysis revealed that the p.Lys364Met mutation has pathological potential based on the functionality of the protein, whereas the p.Ser377Cys mutation was predicted to be neutral or tolerated. The majority of the patients had EPAS1 DNA amplification (79%; n = 56/71) and 53% (n = 24/45) patients shown mRNA overexpression. Most of the patients with EPAS1 mutations exhibited aberrant DNA changes, mRNA and protein overexpression. In addition, these alterations of EPAS1 were associated with tumour weight and location. Thus, the molecular dysregulation of EPAS1 could play crucial roles in the pathogenesis of phaeochromocytomas and paragangliomas.

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

“…PPGL are hereditary in about 40% of cases affecting, among other genes, activators of the oxygen-sensing pathways, mainly VHL and genes encoding for components of the succinate dehydrogenase (SDH) complex ( SDHB , SDHD , SDHC , SDHA and SDHAF2 abbreviated hereafter as SDH ) [ 13 ]. In addition, gain-of-function mutations in EPAS1 have been recently identified [ 14 , 15 , 16 , 17 , 18 ]. The activation of the HIF-signaling pathway has been reported to occur in PPGL with SDH , VHL or EPAS1 mutations, a condition that has been termed pseudohypoxia, given that it occurs in highly vascularized, non-hypoxic tumors.…”

The underlying molecular mechanisms and biomarkers between periodontitis and COVID- 19

“…EPAS1, also known as Hypoxia inducible factor 2 alpha (HIF2α), play an important role in the transcription of many hypoxia-responsive genes 42 . In addition, EPAS1 has associated with various diseases, such as non-small cell lung cancer 43 , paraganglioma and pheochromocytoma 44 , and chronic mountain sickness, which can regulate proliferation of erythroblasts 45 . In general, consistent with previous ndings, these TFs play an extremely important role in immune in ammatory response.…”

Cytoplasmic HIF-2α correlates to proliferation and predicts worse outcome in sympathetic paraganglioma

“…Two novel mutations were found in PGL patients, associated with increased protein half-life and HIF-2α activity 15 . Since then, a number of EPAS1 mutations have been detected in PPGL [16][17][18][19][20][21][22] . Fliedner et al further defined EPAS1 mutated tumors as a subcluster of the pseuodohypoxic cluster 1 PGLs, separated from other hereditary pseudohypoxic PGLs driven by VHL or SDHx genes 23 .…”