title: The VHL gene is epigenetically inactivated in pheochromocytomas and abdominal paragangliomas.
abstract: Pheochromocytoma (PCC) and abdominal paraganglioma (PGL) are neuroendocrine tumors that present with clinical symptoms related to increased catecholamine levels. About a third of the cases are associated with constitutional mutations in pre-disposing genes, of which some may also be somatically mutated in sporadic cases. However, little is known about inactivating epigenetic events through promoter methylation in these very genes. Using bisulphite pyrosequencing we assessed the methylation density of 11 PCC/PGL disease genes in 96 tumors (83 PCCs and 13 PGLs) and 34 normal adrenal references. Gene expression levels were determined by quantitative RT-PCR. Both tumors and normal adrenal samples exhibited low methylation index (MetI) in the EGLN1 (PDH2), MAX, MEN1, NF1, SDHB, SDHC, SDHD, SDHAF2 (SDH5), and TMEM127 promoters, not exceeding 10% in any of the samples investigated. Aberrant RET promoter methylation was observed in two cases only. For the VHL gene we found increased MetI in tumors as compared with normal adrenals (57% vs. 27%; P<0.001), in malignant vs. benign tumors (63% vs. 55%; P<0.05), and in PGL vs. PCC (66% vs. 55%; P<0.0005). Decreased expression of the VHL gene was observed in all tumors compared with normal adrenals (P<0.001). VHL MetI and gene expressions were inversely correlated (R = -0.359, P<0.0001). Our results show that the VHL gene promoter has increased methylation compared with normal adrenals (MetI>50%) in approximately 75% of PCCs and PGLs investigated, highlighting the role of VHL in the development of these tumors.
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Adam
authors.initials: A
authors.lastname: Andreasson
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Nimrod B
authors.initials: NB
authors.lastname: Kiss
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Stefano
authors.initials: S
authors.lastname: Caramuta
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Luqman
authors.initials: L
authors.lastname: Sulaiman
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Fredrika
authors.initials: F
authors.lastname: Svahn
authors.affiliation: Department of Molecular Medicine and Surgery; Karolinska Institutet; Stockholm, Sweden; Department of Breast and Endocrine Surgery; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Martin
authors.initials: M
authors.lastname: Bäckdahl
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden; Department of Clinical Pathology and Cytology; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Anders
authors.initials: A
authors.lastname: Höög
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden; Department of Clinical Pathology and Cytology; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: C Christofer
authors.initials: CC
authors.lastname: Juhlin
authors.affiliation: Department of Oncology-Pathology; Karolinska Institutet; Karolinska University Hospital; Stockholm, Sweden.
authors.forename: Catharina
authors.initials: C
authors.lastname: Larsson
mesh_terms: D000008:Abdominal Neoplasms / Q000235:genetics* / Q000378:metabolism; D000310:Adrenal Gland Neoplasms / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D016022:Case-Control Studies; D019175:DNA Methylation; D015536:Down-Regulation; D044127:Epigenesis, Genetic; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008969:Molecular Sequence Data; D010235:Paraganglioma / Q000235:genetics* / Q000378:metabolism; D010673:Pheochromocytoma / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D051794:Von Hippel-Lindau Tumor Suppressor Protein / Q000235:genetics* / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D051794:Von Hippel-Lindau Tumor Suppressor Protein; C497446:VHL protein, human
keywords: DNA promoter methylation; Pheochromocytoma; VHL; gene expression; paraganglioma; pyrosequencing; qRT-PCR