Elsevier

The Lancet Oncology

Volume 10, Issue 8, August 2009, Pages 764-771
The Lancet Oncology

Fast track — Articles
An immunohistochemical procedure to detect patients with paraganglioma and phaeochromocytoma with germline SDHB, SDHC, or SDHD gene mutations: a retrospective and prospective analysis

https://doi.org/10.1016/S1470-2045(09)70164-0Get rights and content

Summary

Background

Phaeochromocytomas and paragangliomas are neuro-endocrine tumours that occur sporadically and in several hereditary tumour syndromes, including the phaeochromocytoma–paraganglioma syndrome. This syndrome is caused by germline mutations in succinate dehydrogenase B (SDHB), C (SDHC), or D (SDHD) genes. Clinically, the phaeochromocytoma–paraganglioma syndrome is often unrecognised, although 10–30% of apparently sporadic phaeochromocytomas and paragangliomas harbour germline SDH-gene mutations. Despite these figures, the screening of phaeochromocytomas and paragangliomas for mutations in the SDH genes to detect phaeochromocytoma–paraganglioma syndrome is rarely done because of time and financial constraints. We investigated whether SDHB immunohistochemistry could effectively discriminate between SDH-related and non-SDH-related phaeochromocytomas and paragangliomas in large retrospective and prospective tumour series.

Methods

Immunohistochemistry for SDHB was done on 220 tumours. Two retrospective series of 175 phaeochromocytomas and paragangliomas with known germline mutation status for phaeochromocytoma-susceptibility or paraganglioma-susceptibility genes were investigated. Additionally, a prospective series of 45 phaeochromocytomas and paragangliomas was investigated for SDHB immunostaining followed by SDHB, SDHC, and SDHD mutation testing.

Findings

SDHB protein expression was absent in all 102 phaeochromocytomas and paragangliomas with an SDHB, SDHC, or SDHD mutation, but was present in all 65 paraganglionic tumours related to multiple endocrine neoplasia type 2, von Hippel–Lindau disease, and neurofibromatosis type 1. 47 (89%) of the 53 phaeochromocytomas and paragangliomas with no syndromic germline mutation showed SDHB expression. The sensitivity and specificity of the SDHB immunohistochemistry to detect the presence of an SDH mutation in the prospective series were 100% (95% CI 87–100) and 84% (60–97), respectively.

Interpretation

Phaeochromocytoma–paraganglioma syndrome can be diagnosed reliably by an immunohistochemical procedure. SDHB, SDHC, and SDHD germline mutation testing is indicated only in patients with SDHB-negative tumours. SDHB immunohistochemistry on phaeochromocytomas and paragangliomas could improve the diagnosis of phaeochromocytoma–paraganglioma syndrome.

Funding

The Netherlands Organisation for Scientific Research, Dutch Cancer Society, Vanderes Foundation, Association pour la Recherche contre le Cancer, Institut National de la Santé et de la Recherche Médicale, and a PHRC grant COMETE 3 for the COMETE network.

Introduction

Phaeochromocytomas and paragangliomas are rare, usually benign, highly vascularised tumours that both originate from neural-crest-derived chromaffin cells. The term phaeochromocytoma is reserved for intra-adrenal tumours, whereas similar but extra-adrenal tumours are termed paragangliomas. Paragangliomas are subdivided into sympathetic and parasympathetic paragangliomas, depending on their location and catecholamine production. Parasympathetic paragangliomas are located in the head and neck region, and usually do not produce catecholamines, whereas sympathetic paragangliomas are situated along the sympathetic trunk in the abdomen, and usually produce catecholamines.1

Phaeochromocytomas and paragangliomas occur sporadically and in the context of several inherited tumour syndromes, including multiple endocrine neoplasia type 2 (MEN2, with RET gene germline mutations), von Hippel–Lindau (VHL) disease (caused by germline mutations in the VHL gene), neurofibromatosis type 1 (NF1, with NF1 gene germline mutations), and the phaeochromocytoma–paraganglioma syndrome.2, 3 The latter syndrome is the most frequent hereditary condition with manifestation of paragangliomas, and is caused by germline mutations in the SDHB, SDHC, or SDHD genes. The syndrome is characterised by the familial occurrence of phaeochromocytomas or paragangliomas, usually at a young age, and often by multifocal disease with an increased risk of recurrence and an increased frequency of malignancy in the case of SDHB mutations.4 SDHB, SDHC, and SDHD encode three of four subunits of mitochondrial complex II, the succinate-ubiquinone oxidoreductase (succinate dehydrogenase) enzyme located at the crossroads between the mitochondrial aerobic electron transport chain and the tricarboxylic acid cycle.5 Recent studies showed that SDH inactivation induces angiogenesis and tumorigenesis through the inhibition of hypoxia-inducible factors (HIF)-prolyl hydroxylase.6 The SDHB, SDHC, and SDHD genes are bona fide tumour-suppressor genes, as biallelic inactivation is found in phaeochromocytoma–paraganglioma-syndrome tumours (inherited inactivating germline mutation and acquired inactivating mutation of the corresponding wild-type allele in the tumour).7

With the exception of the NF1 syndrome, where the cutaneous café-au-lait spots are characteristic,8 patients with inherited phaeochromocytomas and paragangliomas often go without clinical detection. In large published series of patients with phaeochromocytomas and paragangliomas, it has been shown that 25–30% of patients have an inherited form and 12% of patients with an apparently sporadic phaeochromocytoma and paraganglioma have unexpected germline mutations in VHL, SDHB, or SDHD genes.3, 7, 8, 9 The underdiagnosis of patients with inherited phaeochromocytoma and paraganglioma is the result of a combination of factors, including lack of family information, overlap in age distribution between hereditary and sporadic cases, de-novo mutations, incomplete penetrance (SDHB), parent-of-origin effects on penetrance (SDHD), phenotypic heterogeneity of the disease, and insufficient awareness of clinicians. There is controversy among experts as to whether RET, VHL, SDHB, SDHC, and SDHD genetic testing should be done in all patients with phaeochromocytoma and paraganglioma. Many experts have advocated that molecular genetic testing should be targeted in patients fulfilling specific clinical criteria.4, 10, 11, 12 However, reliable clinical indicators for the presence of SDHB, SDHC, and SDHD germline mutations in patients with phaeochromocytoma and paraganglioma are often absent.

Hidden heredity is most pronounced for patients with apparently sporadic parasympathetic paragangliomas, with up to 34% of cases having a germline mutation in SDHD.13 Clinical indications with high specificity but low sensitivity for the detection of phaeochromocytoma–paraganglioma syndrome (family history of phaeochromocytoma or paraganglioma, multifocal disease, younger age at onset, and malignant tumours) are insufficient for correct diagnosis of the syndrome. The detection of inherited phaeochromocytoma–paraganglioma syndrome is of major importance for patients with phaeochromocytoma and paraganglioma, as well as for their family members, since they are at an increased risk of developing multiple, various, and malignant neoplasms.4, 14, 15, 16 Additionally, after identification of an SDHB, SDHC, or SDHD germline mutation, surveillance can be offered to the individual patient with the paraganglionic tumour and to any family members who carry the mutation. Mutation analysis of SDHB, SDHC, and SDHD has been advocated to diagnose phaeochromocytoma–paraganglioma syndrome in all cases of phaeochromocytoma and paraganglioma where there are no clear clinical or family indications for the syndrome.16 Although SDH-mutation carriers will be identified frequently by mutation analysis of all patients with phaeochromocytomas and paragangliomas, most cases will be without mutation, making this genetic-screening strategy a labour-intensive and financially demanding procedure. Phaeochromocytoma–paraganglioma syndrome tumours differ from sporadic phaeochromocytomas and paragangliomas by the presence of SDHB, SDHC, or SDHD mutations, which are, except for a few incidental cases,17, 18 not found in truly sporadic phaeochromocytomas and paragangliomas. Despite this genotypic difference, no reliable phenotypic discrimination between sporadic phaeochromocytomas and paragangliomas, and phaeochromocytoma–paraganglioma syndrome-related tumours, is possible at present. In the present study we determined the value of SDHB immunohistochemistry for discriminating between SDH-related and non-SDH-related phaeochromocytomas and paragangliomas in large retrospective and prospective series in two different centres.

Section snippets

Patients

Two retrospective series of phaeochromocytomas and paragangliomas were investigated by SDHB immunohistochemistry (Erasmus MC, Rotterdam, Netherlands, 110 cases; Hôpital Européen Georges Pompidou and Hôpital Cochin, Paris, France, 65 cases). These series consisted of phaeochromocytomas diagnosed at Erasmus MC between 1982 and 2007, and diagnosed at INSERM U970 between 1995 and 2007, and of paragangliomas diagnosed in Erasmus MC between 1993 and 1998, and in INSERM U970 between 1993 and 2008. The

Results

Immunohistochemical staining was done on all 220 tumour samples. Of these tumours, 102 had a germline SDH mutation (36 SDHB, five SDHC and 61 SDHD) and all were negative for SDHB immunohistochemistry (figure 1A–C). In four SDH-mutated tumours (SDHB p.Cys98Arg and p.Pro197Arg, and SDHD p.Asp92Tyr and c.169_169+9delTGTATGTTCT) a weak and diffuse cytoplasmic SDHB immunoreactivity was seen in the tumour cells, clearly distinct from the strong speckled pattern present in normal cells of the

Discussion

The results of this study show that SDHB immunohistochemistry on routine FFPE paragangliomas and phaeochromocytomas can reveal the presence of SDHB, SDHC, and SDHD germline mutations with a high degree of reliability. The absence of SDHB staining in tumour cells was found irrespective of whether SDHB, SDHC, or SDHD is mutated, and regardless of the type of mutation, whether missense, nonsense, splice site, or frameshift. The SDHB protein-expression results obtained by immunohistochemistry using

References (33)

  • HP Neumann et al.

    Germ-line mutations in nonsyndromic phaeochromocytoma

    N Engl J Med

    (2002)
  • DE Benn et al.

    Genetic testing in phaeochromocytoma- and paraganglioma-associated syndromes

    Ann N Y Acad Sci

    (2006)
  • AP Gimenez-Roqueplo et al.

    Phaeochromocytoma, new genes and screening strategies

    Clin Endocrinol (Oxf)

    (2006)
  • HP Neumann et al.

    Clinical predictors for germline mutations in head and neck paraganglioma patients: cost reduction strategy in genetic diagnostic process as fall-out

    Cancer Res

    (2009)
  • H Dannenberg et al.

    Frequent germ-line succinate dehydrogenase subunit D gene mutations in patients with apparently sporadic parasympathetic paraganglioma

    Clin Cancer Res

    (2002)
  • DE Benn et al.

    Clinical presentation and penetrance of phaeochromocytoma/paraganglioma syndromes

    J Clin Endocrinol Metab

    (2006)
  • Cited by (438)

    • Succinate Dehydrogenase Mutations as Familial Pheochromocytoma Syndromes

      2023, Surgical Oncology Clinics of North America
    • Methylation and hydroxymethylation in cancer

      2023, Epigenetic Cancer Therapy, Second Edition
    View all citing articles on Scopus
    *

    These authors contributed equally to the paper

    Dr Verhofstad died in March, 2008

    These authors should be considered equal last authors

    View full text