Keywords

4.1 Introduction

Pheochromocytoma (PHEO) and paraganglioma (PGL) are currently considered the tumors with the greatest genetic determinism: in addition to hereditary PHEO associated with neurofibromatosis, Von Hippel Lindau syndrome and multiple endocrine neoplasia type 2 syndrome, many other susceptibility genes have been discovered, identifying PHEO and PGL (PPGL) as tumors with high genetic heterogeneity. The application of genetic screening to all PPGL patients, irrespective of family history or syndromic features, allows detection of a germline mutation in 40% of subjects, with a germline mutation frequency of about 10–12% also in patients with sporadic presentation. Predisposition to PPGL is mostly transmitted in an autosomal dominant fashion, even if large variability in penetrance does not always permit recognition of the traces of hereditability, and one or more generations appear to be skipped. Bilateral PHEO or multifocal PGL, recurrent or malignant disease along with a young age of onset (<45 years) are all possible signs of inherited disease; however, due to the high prevalence of germline mutations also in apparently sporadic disease, the application of genetic screening is currently recommended for all patients with PPGL [1, 2]. The identification of a specific hereditary form has implications for the correct management/follow-up of the patient and also allows extension of genetic analysis to relatives and implementation of presymptomatic surveillance in mutation carriers.

The current availability of high-throughput gene sequencing techniques (next generation sequencing, NGS) allows for the simultaneous study of many genes, overcoming the difficulties associated with the study of a disease with high genetic heterogeneity [3]. Sequencing detects small intragenic insertion or deletion, missense, no sense and splice variants, but whole gene deletion and large intragenic deletion or duplication are not detected; for this analytical purpose other methods must be used, such as quantitative C-reactive protein, multiple ligation-dependent probe amplification, or gene-targeted microarray, according to laboratory preference.

NGS panels available for the study of PPGL susceptibility genes are usually customized with the genes most solidly associated with the development of the disease; the identification of a pathologic variant in one of these genes makes it possible to propose to the patient a surveillance program according to the risk of relapse, malignancy, and involvement of other organs. Larger panels, including more recently identified genes, are available in clinical research centers, though the characteristics of penetrance and phenotype expressivity for mutations in these genes are not yet defined, and the clinical interpretation in many cases cannot be conclusive.

In this chapter, the genetics of PPGL will be described starting from the first recognized syndromic forms up to the recently identified susceptibility genes, with a focus on the specific clinical implications associated with the different genes involved (Table 4.1).

Table 4.1 Clinical phenotype of genetic pheochromocytoma/paraganglioma (PPGL) syndromes
Full size table

4.2 NF1 Gene

NF1 gene is a tumor suppressor gene responsible for neurofibromatosis type 1 (NF1), an autosomal dominant disease. The prevalence of PPGL in NF1 patients is lower than 3%. Frequently the diagnosis is incidental. In most cases the tumor is a single PHEOs, extra-adrenal PGLs are rarer. The mean age at diagnosis is 40 years, similar to sporadic disease, even though cases in young subjects (before 20 years of age) have also been described. Bilateral disease occurs in about 15% of patients, and this relevant information must be kept in mind to favor adrenal-sparing surgery. About 10–12% of NF1-PPGL are malignant.

NF1 is a large gene, and it is not routinely comprised in PPGL-NGS panels: as the penetrance of disease is virtually complete by the age of 7, an appropriate anamnesis and clinical examination are sufficient for the diagnosis.

4.3 RET Gene

The RET gene is a protooncogene, which encodes a transmembrane tyrosine kinase: gain of function mutations activate RET kinase activity conferring oncogenic properties. RET is the causative gene of multiple neoplasia endocrine syndrome type 2 (Men2), a disease with autosomal dominant transmission. In the Men2 syndrome, PHEO, together with medullary thyroid cancer, is one of the two main clinical manifestations, and has a penetrance of about 20–50%. In 10% of Men2, PHEO is the first clinical manifestation. It usually presents at the age of 30–40 years, but it can also develop in infancy by the age of 11. PHEO may be bilateral at presentation or may recur in the contralateral gland, about 50–70% of patients develop bilateral disease; extra-adrenal PGL are very rare, but have been described. They are very rarely malignant (fewer than 5%). Due to the high risk of recurrence, adrenal-sparing surgery is advocated.

4.4 VHL Gene

VHL is an oncosuppressor gene and is the causative gene of Von Hippel Lindau syndrome. This hereditary neoplastic syndrome, with autosomal dominant transmission, is characterized by multiorgan involvement; tumors which most impact on the clinical course of disease are central nervous system and retinal hemangioblastomas, clear cell carcinoma, neuroendocrine tumors and PPGL. Penetrance for PPGL is estimated to be 20–24%, sometimes with a very young age of onset (18–30 years); VHL gene mutations are in fact identified in about 40% of pediatric PHEOs. Tumors are mostly adrenal and are often bilateral (50%), frequently even at presentation. Malignant disease is rare (5–8%). Due to the high risk of bilateral/recurrent disease, adrenal surgery should always be preferred.

4.5 SDHx Genes and SDHAF2 Gene

The SDHx genes (SDHA, SDHB, SDHC, and SDHD) codify for the four subunits of the succinate dehydrogenase (SDH), a mitochondrial enzyme involved in the transfer of electrons in the mitochondrial respiratory chain; SDHAF2 is a mitochondrial protein required for the activation of the SDH complex. SDHx and SDHAF2 are oncosuppressor genes: mutations in these genes were identified in the 2000s as responsible for nearly 50% of hereditary PPGL. SDHx mutations may also predispose to kidney cancer and GIST (wild c-kit, wild PDGFRA). Some particularities for each of these genes are briefly described below.

4.5.1 SDHD Gene (PGL1 Syndrome)

Mutations in the SDHD gene are characterized by high penetrance, about 86% for the age of 50; this explains why most of the subjects with SDHD-related PPGL have familial antecedents [4]. The model of hereditary transmission is autosomal dominant, with maternal imprinting: this means that the phenotype is expressed mostly with paternal transmission, while only 5% of maternal inherited SDHD mutations express the phenotype [5]. The typical SDHD-related phenotype is head and neck PGL (HNPGL), even though thoracic-abdominal or pelvic PGL have been described. The mean age of onset is 36 years, and in more than half of subjects PGLs are multiple and/or bilateral. Malignancy is described in 15–29% of cases [6].

4.5.2 SDHAF2 Gene (PGL2 Syndrome)

Mutations in the SDHAF2 gene are a very rare cause of familial PGL. As for SDHD mutations, the transmission is autosomal dominant with a parent-of-origin effect, so cancer susceptibility is expressed only when the mutation is inherited from the father [7]. The usual phenotype is multiple HNPGLs, with a high penetrance [8], even though only a few families have been described and these data need to be confirmed. The mean age of onset is 33 years, and no malignancy has been described so far.

4.5.3 SDHC Gene (PGL3 Syndrome)

Mutations in the SDHC are a rare cause of hereditary PGL, transmission is autosomal dominant. Patients develop mostly HNPGLs, even though PHEO and extra-adrenal PGLs have been described. Penetrance is not defined, only 25% of patients have a suggestive family history [4], assuming a lower penetrance compared to SDHD mutations. Average age at diagnosis is 38 years, the risk of malignancy is low.

4.5.4 SDHB Gene (PGL4 Syndrome)

Mutations in SDHB account, together with mutations in SDHD, for most SDHx-related PPGLs. SDHB mutation carriers develop abdominal, pelvic, thoracic, or cervical PGLs, less frequently PHEO. The average age of onset of PPGL is 25–30 years, although the disease may appear at very young age (6–7 years). The most relevant clinical feature of the SDHB gene is the association with malignant disease occurring in more than 50% of SDHB-related PGL; it is estimated that about 36% of all malignant PPGL are due to SDHB mutations [9]. Penetrance is lower compared to SDHD/SDHC mutations and is estimated to be about 25–40%.

4.5.5 SDHA Gene (PGL5 Syndrome)

The SDHA gene encodes the catalytic subunit A of the SDH complex. Biallelic mutations of the SDHA gene are responsible for Leigh’s syndrome, a severe early-onset and progressive neurometabolic disorder.

Association of the SDHA gene with PPGL has been more recently identified: heterozygous mutations in SDHA are in fact a rare cause of PPGL and have been identified in subjects with abdominal, pelvic, thoracic, thyroid or cervical PGL. The model of inheritance is autosomal dominant, penetrance is low (about 10% for the age of 70), mean age of onset is 43 years [10]. SDHA mutations have been associated with malignant disease in more than 30% of cases [11].

4.6 TMEM127 Gene

TMEM127 mutations predispose to PHEO. Bilateral disease has been described, but most of the subjects have a solitary non-metastatic PHEO. Penetrance is unknown, probably low, and usually family history is not evocative.

4.7 MAX Gene

MAX mutations predispose to PHEO and abdominal PGL. Half of the patients develop bilateral disease. Penetrance is probably higher compared to TMEM mutations and about 40% of patients have a suggestive family history [12].

4.8 FH Gene

Mutations in FH genes were known to be associated with hereditary leiomyomatosis and papillary renal cell carcinoma; only recently mutations in this gene have been identified in PPGL. FH mutations predispose to multiple and/or malignant PPGL.

4.9 Other Genes

Application of exome whole-exome sequencing to PPGLs patients has led to identification of germline mutations in several genes; among them, some are of particular interest since they are involved in the hypoxia pathway (EPAS1 and EGLN1), in mitochondrial metabolism (MDH2, GOT2, SLC25A11, DLST, KIF1Bβ), in the MAP kinase pathway (MET and MERKT) or in DNA methylation (H3F3A, DNMT3A) [6]. Currently, we have no clear knowledge about the real pathogenic role of these genes in the predisposition to PPGL, nor even elements to hypothesize penetrance and associated clinical phenotype.

4.10 Molecular Biology

Comprehensive genomic and transcriptome analysis has achieved the identification of three distinguishing molecular signatures in PPGL, each of them having germinal and/or somatic mutations in susceptibility genes leading to the activation of a specific oncogenic signaling pathway: the pseudo hypoxic, kinase, and Wnt signaling pathways. These three molecular clusters differ in phenotype and clinical behavior and, above all, they allow identification of molecular predictors of response to therapy, promising the application of personalized genetic-driven therapy in PPGL patients [6].

Cluster 1 is characterized by activation of the hypoxic pathway, and includes tumors with germinal/somatic mutations in the Krebs cycle-associated genes (SDHx, FH, MDH2, GOT2, SLC25A11, DLST, IDH1) and VHL/EPAS1-related genes (PHD1/2, EGLN1, EPAS1, IRP1). Mutations in these genes induce stabilization of HIF2α which in turn determines activation of angiogenesis, cell proliferation and migration; these tumors may have an aggressive phenotype, more than half of the patients with metastatic PPGL carry cluster 1 mutations. Cluster 1 tumors are mostly extra-adrenal and have a preference for noradrenaline secretion; SDHx-related PPGLs intensely express the somatostatin receptor 2 (SSTR2), then 68Ga-DOTATATE PET/CT is considered the most sensitive functional imaging; on the contrary, VHL/EPAS-related tumors have lower expression of SSTR2 and the most sensitive functional imaging is probably 18F-DOPA PET/CT. New inhibitors of hypoxia-inducible factor 2-α represent an extraordinary opportunity for the therapy of metastatic forms.

Cluster 2 is characterized by activation of the tyrosine kinase-linked signaling pathway. It includes tumors with germline/somatic mutations in NF1, RET, TMEM127, MAX, MET, MERTK, HRAS, FGFR1, B-RAF. Mutations in these genes induce activation of phosphatidylinositol-3-kinase (PI3K)/AKT, mTORC and RAS/RAF/ERK signaling pathways leading to tumor proliferation, chromatin remodeling and angiogenesis. Cluster 2 tumors are mostly PHEO, have frequently an adrenergic phenotype and a less aggressive behavior compared to cluster 1. 18F-DOPA/PET/TC is the preferable functional imaging due to the high uptake by the tumoral tissue compared to normal adrenal gland, achieving the detection of multiple lesions within the adrenal parenchyma. Tyrosine kinase inhibitors, PI3K/AKT/mTORC1 inhibitors and RAF/MEK/ERK inhibitors are all promising drugs for the systemic treatment in these tumors.

Cluster 3 is clearly not characterized from a molecular point of view, so it is much more difficult to identify unifying elements regarding clinical aspects. Until now only somatic driver mutations (MAML3 fusion gene and CSDE1 gene mutation) have been identified leading to overactivation of the Wnt signaling and β-catenin, which induces molecular events involved in carcinogenesis. Therapies targeting Wnt signaling are potentially suitable for tumors belonging to cluster 3.