The mechanism of paraganglioma tumorigenesis

Paragangliomas (PGL) of the head and neck are rare, slow-growing and usually
benign tumors. Surgical removal of these tumors is hindered by the growth and
imbedding of the tumor in a rich system of blood vessels and nerves, damage to
which leads to significant morbidity. A significant number of patients with
paraganglioma carry a hereditary DNA change (mutation) in genes located on
chromosome 11, and show a pattern of autosomal inheritance with *imprinting*,
meaning that paragangliomas almost invariably arise only if the mutation is
transmitted via the paternal line. We have previously identified, together with
colleagues, two genes responsible for hereditary paraganglioma: the SDHD gene
codes for one of the two membrane anchor proteins of the mitochondrial
respiratory complex II, and the SDHAF2 gene codes for an SDHA-related cofactor.
Mutations in the SDHA, SDHB and SDHC genes, all coding for components of
respiratory complex II, are also causative in paraganglioma.

The main goal of this project is the elucidation of the mechanism underlying
the paternal inheritance seen in SDHD- and SDHAF2-related paraganglioma. The
results of this project should aid us in the identification of the *modifier*
gene, essential to the early formation of paragangliomas. In addition to this
research, we will also focus on new genes and mechanisms related to
paraganglioma. This research could lead to important new insights into the
genesis of paraganglioma, and results may be important not only to the
understanding of SDHD and SDHAF2-dependent tumors, but also to other related
tumors. In the longer term, these developments may permit the improvement of
current clinical treatment strategies.

A. The involvement of patients in the identification of the paraganglioma
*modifier* gene and other new paraganglioma genes will be limited to the use of
DNA and tumor material. DNA and RNA will be isolated from frozen and
formalin-fixed tumor tissues, and coded samples (without name or date of birth,
but still traceable) will be investigated. DNA studies will include mutation
analysis of paraganglioma genes and the typing of polymorphisms in blood and
tumor DNA, inclusief candidate modifier gene expression analysis in tumor DNA.
B. In addition, we will study the role of chromosome 11 in paragangliomas by
typing DNA polymorphisms. DNA and RNA will be isolated from frozen and
formalin-fixed tumor tissues. Patients will be asked to give informed consent
for the use of this material. DNA will be used to establish LOH and copy number
changes in tumors. Pedigree analysis, required to establish disease status,
will be conducted by the ENT doctor or clinical geneticist. Newly identified
mutation carriers may be asked to participate in normal ENT and
endocrinological clinical protocols (addendum 2).

None: the withdrawal of venous blood is a standard procedure involving limited
discomfort and a very low risk. The investigations of tumor tissue will use
remnant material remaining after normal surgical procedures, and thus involve
no additional burden for the patient. The standard ENT and endocrinological
investigations of clinical parameters (addendum 2) do not represent any
additional burden for the patient as they occur only when dictated by normal
clinical protocols. These may include MRI to detect occult tumors and the
withdrawal of blood for the determination of catecholamine levels. These
procedures will yield data that will be also used to provide optimal clinical
care for the patient.

Addendum 2:
Following DNA testing (with informed consent), mutation carriers at a high risk
of tumors will be referred to the outpatient clinics of the Departments of
Endocrinology and Otorhinolaryngology of the LUMC, a tertiary referral center
for paragangliomas. Patients will be seen following standardized clinical
protocols and in cases with suspected actively secreting tumors, urine will be
collected in duplicate, over 24 hours, under strict dietary regulations and
after stopping medication for several weeks or changing antihypertensive
medication to doxazosine. In cases showing excessive catecholamine secretion,
MIBG scanning and additional whole-body MRI and/or CT imaging will be used to
identify the source of catecholamine overproduction. Independent of urine
analysis results, whole-body MRI and/or CT imaging will be performed at least
every two years. The Ear-Nose-Throat (ENT)-surgeon will examine the head and
neck region, with special attention to the lower cranial nerves, annually, and
if a paraganglioma is present, patients will also undergo MRI annually. In
mutation carriers without evidence for head and neck paragangliomas, MRI of the
head and neck region will be performed every 3-5 years.