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Rare Germline Mutations Identified by Targeted Next-Generation Sequencing of Susceptibility Genes in Pheochromocytoma and Paraganglioma
Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences.
Karolinska Institute, Sweden; Karolinska University Hospital Solna, Sweden; Karolinska University Hospital, Sweden.
Karolinska Institute, Sweden; Karolinska University Hospital Solna, Sweden; Karolinska University Hospital, Sweden.
University of Wisconsin, WI 53715 USA.
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2014 (English)In: Journal of Clinical Endocrinology and Metabolism, ISSN 0021-972X, E-ISSN 1945-7197, Vol. 99, no 7, E1352-E1360 p.Article in journal (Refereed) Published
Abstract [en]

Context: Pheochromocytomas and paragangliomas have a highly diverse genetic background, with a third of the cases carrying a germline mutation in 1 of 14 identified genes. Objective: This study aimed to evaluate next-generation sequencing for more efficient genetic testing of pheochromocytoma and paraganglioma and to establish germline and somatic mutation frequencies for all known susceptibility genes. Design: A targeted next-generation sequencing approach on an Illumina MiSeq instrument was used for a mutation analysis in 86 unselected pheochromocytoma and paraganglioma tumor samples. The study included the genes EGLN1, EPAS1, KIF1B beta, MAX, MEN1, NF1, RET, SDHA, SDHB, SDHC, SDHD, SDHAF2, TMEM127, and VHL. Results were verified in tumor and constitutional DNA with Sanger sequencing. Results: In all cases with clinical syndromes or known germline mutations, a mutation was detected in the expected gene. Among 68 nonfamilial tumors, 32 mutations were identified in 28 of the samples (41%), including germline mutations in EGLN1, KIF1B beta, SDHA, SDHB, and TMEM127 and somatic mutations in EPAS1, KIF1B beta, MAX, NF1, RET, and VHL, including one double monoallelic EPAS1 mutation. Conclusions: Targeted next-generation sequencing proved to be fast and cost effective for the genetic analysis of pheochromocytoma and paraganglioma. More than half of the tumors harbored mutations in the investigated genes. Notably, 7% of the apparently sporadic cases carried germline mutations, highlighting the importance of comprehensive genetic testing. KIF1B beta, which previously has not been investigated in a large cohort, appears to be an equally important tumor suppressor as MAX and TMEM127 and could be considered for genetic testing of these patients.

Place, publisher, year, edition, pages
Endocrine Society , 2014. Vol. 99, no 7, E1352-E1360 p.
National Category
Clinical Medicine
URN: urn:nbn:se:liu:diva-111616DOI: 10.1210/jc.2013-4375ISI: 000342341000026PubMedID: 24694336OAI: diva2:758480

Funding Agencies|University of Linkoping; LiU Cancer Network; Swedish Research Council; Cancer Society in Stockholm; Swedish Cancer Society; StratCan at Karolinska Institutet; Genetics Services Unit at the Wisconsin National Primate Research Center [P51RR000167/P51OD011106]

Available from: 2014-10-27 Created: 2014-10-27 Last updated: 2015-03-04
In thesis
1. Genetic Alterations in Pheochromocytoma and Paraganglioma
Open this publication in new window or tab >>Genetic Alterations in Pheochromocytoma and Paraganglioma
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Pheochromocytomas and paragangliomas are neuroendocrine tumors that arise from neural crest-derived cells of the adrenal medulla and the extra-adrenal paraganglia. They cause hypertension due to an abnormally high production of catecholamines (mainly adrenaline and noradrenaline), with symptoms including recurrent episodes of headache, palpitations and sweating, and an increased risk of cardiovascular disease. Malignancy in the form of distant metastases occurs in 10-15% of the patients. The malignant cases are difficult to predict and cure, and have a poor prognosis. About a third of pheochromocytomas and paragangliomas are caused by hereditary mutations in a growing list of known susceptibility genes. However, the cause of the remaining, sporadic, tumors is still largely unknown. The aim of this thesis project has been to further characterize the genetic background of pheochromocytomas and paragangliomas, with a focus on the sporadic tumors.

First, we investigated the role of the genes known from the familial tumors in the sporadic form of the disease. By studying mutations, copy number variations, DNA methylation and gene expression, we found that many of the known susceptibility genes harbor somatic alterations in sporadic pheochromocytomas. Particularly, we found that the NF1 gene, which plays an important role in suppressing cell growth and proliferation by regulating the RASMAPK pathway, was inactivated by mutations in more than 20% of the cases. The mutations occurred together with deletions of the normal allele and were associated with a reduced NF1 gene expression and a specific hormone profile. We also detected activating mutations in the gene EPAS1, which encodes HIF-2α, in a subset of sporadic cases. Microarray analysis of gene expression showed that several genes involved in angiogenesis and cell metabolism were upregulated in EPAS1-mutated tumors, which is in agreement with the role of HIF-2α in the cellular response to hypoxia. In order to comprehensively investigate all the known susceptibility genes in a larger patient cohort, we designed a targeted next-generation sequencing approach and could conclude that it was fast and cost-efficient for genetic testing of pheochromocytomas and paragangliomas. The results showed that about 40% of the sporadic cases had mutations in the tested genes. The majority of the mutations were somatic, but some apparently sporadic cases in fact carried germline mutations. Such knowledge of the genetic background can be of importance to facilitate early detection and correct treatment of pheochromocytomas, paragangliomas and potential co-occurring cancers, and also to identify relatives that might be at risk. By sequencing all the coding regions of the genome, the exome, we then identified recurrent activating mutations in a novel gene, in which mutations have previously only been reported in subgroups of brain tumors. The identified mutations are proposed to cause constitutive activation of the encoded receptor tyrosine kinase, resulting in the activation of downstream kinase signaling pathways that promote cell growth and proliferation.

In summary, the studies increase our biological understanding of pheochromocytoma and paraganglioma, and possibly also co-occurring cancers in which the same genes and pathways are involved. Together with the findings of other scientific studies, our results may contribute to the development of future treatment options.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2015. 64 p.
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1439
Pheochromocytoma; paraganglioma; genetics; sequencing; cancer; mutation
National Category
Clinical Medicine
urn:nbn:se:liu:diva-114806 (URN)10.3384/diss.diva-114806 (DOI)978-91-7519-145-4 (print) (ISBN)
Public defence
2015-04-09, Berzeliussalen, Campus US, Linköping, 13:00 (English)
Available from: 2015-03-04 Created: 2015-03-04 Last updated: 2016-04-07Bibliographically approved

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Welander, JennyGimm, OliverSöderkvist, Peter
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Division of Cell BiologyFaculty of Health SciencesDivision of Clinical SciencesDepartment of Surgery in LinköpingDepartment of Clinical Pathology and Clinical Genetics
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