Annals of Diagnostic Paediatric Pathology

Annals of Diagnostic Paediatric Pathology 2007, 11(1–2):15–19
© Copyright by Polish Paediatric Pathology Society Annals of
Pheochromocytoma in children and adolescents based
on Polish Pheochromocytoma Registry
Mariola Peczkowska 1 , Andrzej Januszewicz 1 , Barbara Jarz¹b 3 , Hartmut Neumann 4 ,
Agata Kubaszek 1 , Hanna Janaszek-Sitkowska 1 , Mieczys³aw Litwin 5 , Jolanta Antoniewicz 5 ,
Ewa Aksamit-Bialoszewska 6 , El¿bieta Ros³onowska 7 , Aleksander Prejbisz 1 ,
Magdalena Januszewicz 8 , Ilona Micha³owska 2 , Jaros³aw Æwik³a 9 , Mariusz Furmanek 9 ,
Jerzy Walecki 9
Diagnostic
Paediatric
Pathology
1
Department of Hypertension
2
Department of Radiology
Institute of Cardiology
Warsaw, Poland
3
Nuclear Medicine and Endocrine Oncology Department
Maria Sklodowska−Curie Memorial Institute
Gliwice, Poland
4
Medizinische Universitatsklinik
Abteilung IV
Freiburg, Germany
5
Department of Nephrology, Kidney Transplantation and
Arterial Hypertension
The Children's Memorial Health Institute
Warsaw, Poland
6
Endocrinology and Diabetology Ward
General District Hospital
Olsztyn, Poland
7
Department of Endocrinology
Centre for Postgraduate Medical Education
Warsaw, Poland
8
II Department of Clinical Radiology
Medical Academy in Warsaw
Poland
9
Department of Radiology
Central Clinical Hospital of the Ministry
of Internal Affairs and Administration
Warsaw, Poland
Abstract
Pheochromocytomas and paragangliomas occur as sporadic tumors or in a familial context. It has been shown
that approximately 25% of patients with pheochromocytoma and paraganglioma carry germline mutations in
one of 4 susceptibility genes. Peripheral blood from unrelated, registry patients with pheochromocytoma was
tested for mutations of RET, VHL, SDHD, and SDHB. Clinical data at first presentation and follow-up were
evaluated. Among 136 patients (93 female and 43 male, 68,4% and 31,6% respectively; mean age, 45±14
years) who presented with pheochromocytoma, 34 (24,8%) were found to have mutations. Younger age,
multifocal tumors, and extra-adrenal tumors were significantly associated with the presence of a mutation.
There were 16 patients with pheochromocytoma under 20 years of age (9 female and 7 male, mean age
15,9±4,6 years, age range 5-20 years). Among them, we identified 15 patients with germ-line mutations (94
percent); 1 had M918T mutation of RET proto-oncogene resulting in MEN 2B syndrome, 6 had mutations of
VHL gene, 5 mutations of SDHD gene, and 3 mutations of SDHB gene. Only two patients had positive family
history at presentation. One fourth of patients with pheochromocytoma are carriers of mutations. Incidence
of hereditary syndromes in patients under 20 years is 94%. Paraganglioma – pheochromocytoma syndrome
resulting from SDHD or SDHB gene mutations (PGL1 and PGL4 syndrome, respectively) seems to be most
common hereditary syndrome in Polish population of patients with pheochromocytoma under 20 years of age.
Second major hereditary syndrome in this group was von Hippel-Lindau disease.
Key words: Hereditary syndromes, paraganglioma, pheochromocytoma
Address for correspondence
Mariola Peczkowska, MD, PhD phone: 022 34 34 338
Department of Hypertension fax: 022 34 34 517
Institute of Cardiology
e-mail: mpeczkowska@ikard.pl
04-628 Warsaw
Alpejska 42

16
Introduction
Familial pheochromocytomas
Pheochromocytomas are rare tumors of the adrenal gland
that arise from chromaffin cells in the adrenal medulla.
Paragangliomas arise from extra – adrenal chromaffin
cells and can originate in either the sympathetic nervous
system or parasympathetic ganglia. Paragangliomas that arise
from sympathetic nervous system occur most frequently
in the retroperitoneum and are traditionally
termed extra – adrenal pheochro-
Table 1
mocytoma. Paragangliomas have been
broadly categorized into two groups:
those in the head and neck region with
carotid body as the major site and those
located elsewhere, with adrenal medulla
as the major site. Tumors in the
head and neck are anatomically associated
with parasympathetic nervous system
and are located close to the major
arteries and nerves, whereas the adrenal
medulla and other paraganglia below
neck and head are closer associated
with the sympathetic nervous system.
Parasympathetic tumors are usually
present as an asymptomatic slow growing
mass, lacking endocrine activity,
whereas sympathetic tumors are hormonally
active and secret excess amount of
catecholamines. Pheochromocytomas
and paragangliomas occur as sporadic
tumors or may be associated with hereditary
syndromes [2]. Several genetic
syndromes are known to be associated
with increased risk for pheochromocytoma,
including von Hippel – Lindau
(VHL) syndrome, multiple endocrine
neoplasia typ 2 (MEN 2) and neurofibromatosis
type 1 (NF1), occurring as
a result of germline mutations in VHL,
RET, or NF1 genes, respectively. More
recently, nuclear genes encoding mitochondrial
complex II subunit proteins
have been associated with the development
of pheochromocytomas and paragangliomas
and newly discovered paraganglioma/pheochromocytoma
syndrome
(PPS) was added to the list of
hereditary syndrome in pheochromocytoma
and paraganglioma. Neumann et
al. showed that close to 24% of patients
with pheochromocytomas and paragangliomas
carry a germline mutation [8].
This was confirmed by Amar et al. who
even found 27% carrying germline mutations
[1]. Therefore, the historically
established rule of tens’, stating that approximately
10% of pheochromocytomas
are hereditary, 10% are malignant, 10% are bilateral,
10% are extra-adrenal, 10% are not associated with hypertension
and 10% occur in children, is no longer valid concerning
genetics. Molecular medicine makes it possible to
differentiate sporadic from hereditary disease, which will
affect medical management not only for the patient but also
for the family. Familial syndromes of pheochromocytoma
are presented in Table 1.
Syndrome and prevalence Gene Gene locus
of pheochromocytomas
Multiple endocrine neoplasia Type 2A Ret-protooncogene 10q11.2
(30–60%)
Medullary thyroid carcinoma
Pheochromocytoma
Hyperparathyroidism
Multiple endocrine neoplasia Type 2B Ret-protooncogene 10q11.2
(MEN2B) (30-60%)
Medullary thyroid carcinoma
Pheochromocytoma
Marfanoid habitus
Skeletal deformation
Mucosal neuromas
Ganglioneuromatosis of the intestinal tract
Von Hippel-Lindau disesase (VHL) VHL-tumor 3p25-26
(15–20%) type 2 suppressor gene
A: Retinal and CNS
haemangioblastomas
Pheochromocytomas
Endolymphatic sac tumors
Epididymal cystadenomas
B: As in VHL type 2A +
renal cell cysts and carcinomas
As in VHL type 2A + pancreatic
neoplasmas and cysts
C: Pheochromocytomas only
Neurofibromatosis type 1 (NF1) NF-1-gene 1 7q11.2
(3–5%)
Multiple fibromas on skin
Cafe au lait spots
Lisch nodules of the iris
Pheochromocytoma-paraganglioma SDHB-gene 1p35-36
syndrome (PGL, 70-80%) SDHD-gene 11q21-23
Head and neck tumors
Extra – adrenal and adrenal pheochromocytomas

17
In the present study, we analyzed the known susceptibility
genes for pheochromocytoma – VHL, RET, SDHD,
and SDHB – in a large, unselected series of patients collected
in Polish Registry of Pheochromocytoma who presented
with this tumor in order to classify them as having either
sporadic or hereditary disease. In addition, we analyzed hereditary
syndrome prevalence and clinical characteristic in
young pheochromocytoma patients under 20 years of age.
Material and methods
The population based registry for pheochromocytoma was
conducted in Poland. Patients were recruited from thirty five
centres and clinical, radiological and demographical data
was collected. One hundred thirty-six consecutive, unrelated
patients with histologically confirmed pheochromocytoma
from whom blood-leukocyte DNA was available were enrolled.
All patients provided written or oral informed consent.
For classification we used term paraganglioma for tumors of
location in the head and neck area whereas those of adrenal,
extraadrenal abdominal and thoracic location were named
pheochromocytomas.
All eight exons of SDHB, all four exons of SDHD, all
three exons of VHL, and exons 10, 11, and 13 through 16 of
RET were examined by analysis of single-strand conformation
polymorphisms and direct sequencing, as previously described
[5].
Missense mutations were diagnosed if the DNA variants
were absent in 100 healthy controls. All molecular examinations
were performed in Department of Nephrology and
Hypertension, Albert Ludwigs University, Freiburg, Germany.
Results
A total of 136 patients (93 female and 43 male, 68,4% and
31,6% respectively; mean age, 45±14 years) with pheochromocytoma
were enrolled in the study. We identified 34 patients
with deleterious germ-line mutations (24,8%). Only
3 patients had family history at the presentation. Among the
34 patients with mutations, 32% had mutations of RET, 18%
had germ-line mutations of VHL, 12% had clinical signs of
NF1 syndrome, and 18 and 21% presented mutations of two
newly identified genes, SDHD and SDHB.
The age at the onset of symptoms was statistically lower
in all carriers of mutations than in patients with sporadic
disease. Multiple, extra – adrenal, and bilateral adrenal
pheochromocytomas as well as recurrences of the disease
were statistically more frequent among patients with mutations
than among patients without mutations. Incidence of
malignant pheochromocytoma did not differ between study
groups (Table 2).
The most frequent symptom of the disease was hypertension
(86%), then palpitations (67%), sweating (56%), headache
(40%), blanching of the skin (40%), severe cardiovascular
complications (myocardial infarction, stroke, severe
arrhythmia, etc.) (7,4%). Asymptomatic clinical course was
observed in 8,8% of patients (Table 3).
Table 2
Comparison between hereditary pheochromocytoma and
sporadic pheochromocytoma groups
Hereditary Sporadic p
Age (years) 34,3±13,2 48,7±13,4 p

18
types were diagnosed in participating patients. A medullary
thyroid carcinoma was present in one patient with MEN 2B
syndrome. Renal cell carcinoma (RCC) occurred in two patients
– in SDHB mutation carrier and in patient with von
Hippel – Lindau disease. Pancreatic islet tumors and haemangioblastoma
of IV brain ventricle were found in patient
with VHL. In the cohort presented here, there were also two
reports of multiple head and neck paraganglioma in SDHD
gene mutation carriers [Table 4].
Table 4
Clinical characteristic of patients with hereditary syndromes
of pheochromocytoma
Characteristics of pheochromocytoma patients with
hereditary syndromes under 20 years of age
Multiple tumors – 12 patients (80%)
Extra – adrenal tumors – 7 patients (47%)
Bilateral adrenal tumors – 6 patients (40%)
Thoracic localization of tumors – 2 patients (13%) – only in
SDHD gene mutations
Recurrence of the disease – 5 patients (33%)
Malignant – 0
Associated tumors:
! Renal cell carcinoma – 2 patients: VHL and SDHB
! Pancreatic islet tumors – 1 patient – VHL
! haemangioblastoma of IV brain ventricle
! Multiple head and neck paraganglioma – 2 patients –
SDHD
! Medullary thyroid carcinoma – 1 patient – MEN 2B
Multiple or extraadrenal or bilateral adrenal pheochromocytoma
– 13 patients (87%)
Discussion
Our clinical and molecular evaluation of 136 unrelated
patients who presented with pheochromocytoma revealed
that 34 patients (24,8%) had a hereditary predisposition
to von Hippel–Lindau disease, MEN-2, neurofibromatosis
type 1, or the syndromes associated with pheochromocytoma
and paraganglioma. Among the 34 patients with mutations,
32% had mutations of RET, 18% had germ-line mutations
of VHL, 12 percent had clinical signs of NF1 syndrome,
and 18 and 21% had mutations of two newly
identified genes, SDHD and SDHB. Patients with hereditary
syndromes were younger that those with sporadic disease
and multiple and extra- adrenal tumors were more frequent
in this group. Our results are consistent with other population-based
studies [2, 4, 7, 8, 9]. In our registry almost one
half of patients with hereditary pheochromocytoma was under
20 years of age. We can state that 87% of all multifocal
(including bilateral tumors) and extra – adrenal tumors in
patients with onset of the disease at the age of 20 years or
younger were found to be hereditary. All these findings po-
inted out, that extra – adrenal and/or multiple pheochromocytomas
as well as young age at presentation may be striking
features of hereditary disease [9]. Interestingly, 91% of
probands found to have hereditary disease with the use of
molecular testing had no associated signs and symptoms at
presentation. Only three (9%) of them had positive family
history at presentation. A partial explanation for the high
frequency of hereditary pheochromocytoma without family
history of disease might include spontaneous mutation in
one of the susceptibility genes, decreased penetrance, and
maternal imprinting [9]. In our registry, spontaneous mutations
in one of four susceptibility genes occurred in two patients
with hereditary von Hippel–Lindau disease, one with
MEN 2B syndrome, one with SDHD gene mutation and one
with SDHB mutation (Table 5).
Today, more than 200 different VHL mutations have
been identified which appear to be equally distributed throughout
the gene. De novo mutations at hypermutable sequences
result in most of the recurrent mutations. In large German
registry of von Hippel–Lindau disease, frequency of
spontaneous mutation in VHL is thirteen percent of all cases
[9, 10]. In MEN 2B syndrome, which is the most distinct and
aggressive of the MEN 2 variants, de novo mutations are very
often and reach fifty percent [12]. A little is known about
spontaneous mutations in SDHD and SDHB genes because
they are newly identified genes.
Penetrance is known to be relatively high (approximately
70 percent by the age of 70) among patients with MEN-
2 and von Hippel–Lindau disease, overall [9, 10]. Benn et al.
reported a statistically significant age-related penetrance difference
for SDHB and SDHD mutation carriers. By age 30
yr, 29% of SDHB mutation carriers and 48% of SDHD mutation
carriers were diagnosed with paraganglioma; by age
40 yr, 45% of SDHB mutation carriers and 73% of SDHD
mutation carriers were diagnosed with paraganglioma [3,
13]. As discussed by authors, SDHD and SDHB mutations
have an age-related penetrance, and the lifetime risk of developing
paraganglioma (s)/pheochromocytoma (s) approaches
100% by age 70 yr [3, 13].
In patients with SDHD mutations, penetrance depends
on whether the individual inherited the mutation from
the mother or the father [11, 13]. The disease is not manifested
when the mutation is inherited from the mother, but is
highly penetrant when inherited from the father. This phenomenon
is known as maternal imprinting.
Overall, therefore, pheochromocytomas in patients
without family histories are due to spontaneous mutations,
decreased penetrance, or maternal imprinting, although other
causes such as gene–gene interactions and gene–environment
interactions may be possible. Genetic testing can be
a powerful aid to the identification of a syndrome in such cases.
Since disease is likely to develop in virtually all patients
with a family specific mutation, it seems reasonable to subject
such patients to lifelong surveillance.
This report highlights the value of genetic testing for
affected patients and at-risk asymptomatic family members.
Genetic testing should be considered in all patients with pa-

19
Table 5
Germ-line mutations in the four genes detected in the series of patients with pheochromocytoma under 20 years of age
Independent cases Gene Exon cDNA nucleotide Amino-acid change Mutation type Family history
1. SDHB 3 402 C>T R90X Stopcodon hereditary
2. 6 721 G>A C196Y Missense unknown
3. 6 847-50delTCTC F238frameshift Frameshift spontaneous
4. SDHD 1 33 C>A C11X Stopcodon spontaneous
5. 1 33 C>A C11X Stopcodon hereditary
6. 1 33 C>A C11X Stopcodon unknown
7. 1 33 C>A C11X Stopcodon hereditary
8. 1 33 C>A C11X Stopcodon hereditary
9. VHL 3 712 C>T A167Y Missense spontaneous
10. 3 764 T>A L184H Missense unknown
11. 1 451A>G S80G Missense hereditary
12. 1 451A>G S80G Missense hereditary
13. 1 451A>G S80G Missense hereditary
14. 1 404G>C R64P Missense spontaneous
15. RET 16 918 ATG>ACG M918T Missense spontaneous
raganglioma and/or pheochromocytoma [6, 9, 13]. Familial
pheochromocytoma/paraganglioma is inherited in an autosomal
dominant manner; thus, an affected person has a 50%
chance of passing the mutation on to each child.
The surveillance strategy for a germline mutation-positive
asymptomatic patient should include: annual history
and physical examination by a clinician experienced with
pheochromocytoma and paraganglioma; annual biochemical
testing, e. g. 24-h urine for metanephrines and catecholamines
and computed tomography or magnetic resonance imaging
every 2 yr taking into consideration the tumour locations
typically associated with the mutated gene [6, 13]. Given
the observation that children of female SDHD mutation
carriers do not manifest the disease when the mutation has
been inherited, rigorous clinical surveillance is probably not
warranted for these offspring.
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