Bernhard U. Bender

The impact of molecular genetic analysis of the VHL gene in patients with haemangioblastomas of the central nervous system

OBJECTIVES Haemangioblastoma of the CNS occurs as a sporadic entity and as a manifestation of the autosomal dominant von Hippel-Lindau disease with the major additional components retinal angioma, renal cancer, and pheochromocytoma. Genetic testing for germline mutations predisposing to von Hippel-Lindau disease has been available since identification of the VHL tumour suppressor gene. The impact of this testing was evaluated in patients with haemangioblastomas seen in this centre. METHODS A register and database of patients with symptomatic haemangioblastomas for the last 15 years was evaluated. The VHL gene was analysed by the SSCP method for all exons and Southern blotting for mutations and deletions of the gene. RESULTS 141 patients with haemangioblastoma of the CNS were registered. In 81 patients (57%) there was a disease predisposing germline mutation including eight novel mutations. Population related calculation of patients from the administrative district of Freiburg disclosedVHL germline mutations in 22% of the patients with haemangioblastoma. Analysis of mutation carriers for clinical information suggestive of the syndrome showed (1) a positive family history of a brain tumour in 50%, (2) a history for the patient of extracranial manifestations in 36% (retinal angioma 30%, pheochromocytoma 6%), and (3) 19% presenting with multiple brain tumours when first admitted. By genetic testing of haemangioblastoma patients without any indications of von Hippel-Lindau disease mutation carriers were identified in 14%. Sensitivity of VHL germline testing was 86%. CONCLUSIONS DNA analysis for VHL germline mutations is clearly superior to clinical information in the diagnosis of von Hippel-Lindau disease. Although the percentage of von Hippel-Lindau disease associated haemangioblastoma decreases after the fourth decade of life and is infrequent in patients without other symptomatic lesions and a negative family history, it is recommended that every patient with CNS haemangioblastoma should be screened for von Hippel-Lindau disease germline mutations. This provides the key information and enables screening for extraneurological tumours of the patients and investigations of the patient′s family to ameliorate management of von Hippel-Lindau disease.

Reconsideration of biallelic inactivation of the VHL tumour suppressor gene in hemangioblastomas of the central nervous system.

OBJECTIVES Cerebellar haemangioblastoma occurs sporadically or as a component tumour of autosomal dominant von Hippel-Lindau disease. Biallelic inactivation of the VHL tumour suppressor gene, which is located on chromosome 3p, has been shown to be involved in the pathogenesis of both tumour entities. Mechanisms ofVHL inactivation are intragenic mutations, mitotic recombination events, and hypermethylation of the promoter region. The systematic and complete examination of these genetic and epigenetic phenomena in large series of von Hippel-Lindau disease related and sporadic hemangioblastomas has, thus far, not been performed. METHODS In the largest series to date, 29 von Hippel-Lindau disease associated and 13 sporadic haemangioblastomas were investigated for all suggested inactivating mechanisms of theVHL gene using single strand conformational polymorphism (SSCP), loss of heterozygosity (LOH), and methylation analyses. Additionally, corresponding blood samples of all patients were screened for VHL germline mutations by SSCP and Southern blotting. RESULTS Germline mutations were identified in 94% of patients with von Hippel-Lindau disease and their tumours and 62% of these hemangioblastomas showed LOH of chromosome 3p. Of the 13 sporadic tumours, 23% showed a single somatic mutation of theVHL gene that was not present in the germline. 3p LOH was identified in 50% of informative sporadic tumours. No von Hippel-Lindau disease related or sporadic tumour demonstrated VHLpromoter hypermethylation. CONCLUSIONS For most von Hippel-Lindau disease related haemangioblastomas, the inactivation or loss of both alleles of theVHL gene, as predicted by the Knudson two hit theory, is required. However, in a subset of tumours including most sporadic haemangioblastomas, the genetic pathways involved in tumorigenesis have yet to be defined and may represent alterations of a different pathway or pathways.

VHL c.505 T>C mutation confers a high age related penetrance but no increased overall mortality

BACKGROUND Germline mutations of the VHL gene cause von Hippel-Lindau syndrome (VHL). In southern Germany, a specific mutation in this gene, c.505 T>C, is one of the most frequent alterations owing to a founder effect. METHODS This study was conducted to evaluate morbidity, specific clinical risk profile, and mortality among a series of VHL c.505 T/C mutation carriers. A total of 125 eligible subjects carryingVHL c.505 T/C underwent ophthalmoscopy and gadolinium enhanced magnetic resonance imaging of the brain, the spinal cord, and the abdomen. Age related penetrance, morbidity, and mortality were assessed. RESULTS Frequently observed lesions were phaeochromocytoma (47%), retinal angiomas (36%), haemangioblastoma of the spine (36%), and haemangioblastoma of the brain (16%). Four patients developed renal cell carcinoma. VHL was symptomatic in 47% of subjects; 30% were asymptomatic despite the presence of at least one VHL related tumour and 23% of the carriers had no detectable VHL lesion. Of the 19 patients who had died (15%), 10 died of symptomatic VHL lesions. Overall penetrance by cumulative incidence functions is estimated at 48% by 35 years and 88% by 70 years. In contrast to the only existing published report based on patients with presumably unselected VHLgermline mutations, the mortality rate for c.505 T/C mutation carriers is comparable to that of the general population of Germany. CONCLUSIONS Our results are an important example that a specific genotype, at least in the case ofVHL c.505 T/C, can favourably impact on mortality despite a high age related penetrance. Our study also indirectly provides objective data which might be useful to the life and health insurance industry; it would appear that c.505 T>C mutation positive subjects have similar disease specific mortality to that of the general population owing to a combination of phenotype and timely detection of mutation carrier status followed by aggressive clinical screening and, if necessary, treatment.

Monogenetic Hypertension and Pheochromocytoma

Hypertension attributable to pheochromocytoma is a very attractive model for the elucidation of the pathogenesis of hypertension. Sixteen different point mutations in the RET proto-oncogene and 30 mutations in the Von Hippel-Lindau (VHL) tumor suppressor gene have been identified so far associated with expression of pheochromocytoma. Each of these mutations initiates either the syndrome of multiple endocrine neoplasia type 2 (MEN 2) (MEN 2A and MEN 2B) or the VHL disease. Certain mutations in both genes are associated with the presence of pheochromocytoma. In general, these pheochromocytomas produce catecholamines that result in hypertension. Therefore, analysis for germline mutations in these genes are of practical value, because susceptibility to these diseases can be predicted in as yet clinically unaffected relatives.

A novel frameshift mutation of the lecithin:cholesterol acyltransferase (LCAT) gene associated with renal failure in familial LCAT deficiency

Abstract Background: The lecithin:cholesterol acyltransferase (LCAT) gene is located on the long arm of chromosome 16 and encodes a highly conserved enzyme that catalyzes the formation of most plasma lipoprotein cholesteryl esters. Two autosomal recessive disorders, familial LCAT deficiency (FLD) and fish eye disease, are associated with germline LCAT mutations. Patients with FLD and fish-eye disease frequently present with corneal opacity, anemia and renal failure with proteinuria. Methods: We clinically and biochemically characterized a German patient with classical FLD and used molecular genetic analysis to identify a novel homozygous LCAT mutation within codon 178. Results: The insertion of adenine identified is located in one of the two motifs that resemble sequences found in several lipases, and results in a frameshift with a stop codon at residue 214. Therefore, the mutation alters a large portion of the LCAT enzyme, including both protein regions with putative lipase activity. Clinically, the female patient presented with corneal opacity, mild anemia and a slow deterioration in kidney function that led to a requirement for hemodialysis until she received a renal transplant. Conclusions: The present data provide additional insights into the genotype/phenotype correlations of FLD and thus may improve the genetic diagnosis of this interesting inborn error of metabolism. Clin Chem Lab Med 2007;45:483–6.

Molekulare Grundlagen von Erkrankungen der Nebenniere

Die Nebennieren wurden das erste Mal von Eustachio, Professor der Anatomie in Rom, 1564 beschrieben. Es dauerte allerdings bis ins 19. Jahrhundert, bis durch Cuvier (l805) die Unterscheidung von Nebennierenrinde und Nebennierenmark folgte. 1866 beschrieb Arnold die verschiedenen histologischen Zonen der Nebennierenrinde. Die Bedeutung der Nebennierenrinde fur das Uberleben wurde von Thomas Addison 1855 erkannt. 1856/57 wies auserdem Edouard BraunSequard tierexperimentell die Lebensnotwendigkeit der Nebennieren nach. Beide Autoren fanden allerdings in der Folgezeit keine Anerkennung. So konnte es geschehen, dass Ioseph Hyrtl, Professor der Anatomie in Wien, in seinem weit verbreiteten Lehrbuch noch um die Jahrhundertwende schrieb: „Die unbekannte Funktion der Nebenniere sichert dieses Organ vor lastigen Nachfragen der Heilwissenschaft“

Clinical and genetic screening in patients with capillary retinal angioma

SummaryCapillary retinal angiomas are rare vascular tumors that frequently occur in von Hippel-Lindau syndrome (vHL) but may also be sporadic. In all patients presenting with this tumor a thorough search for other vHL-associated lesions must be performed. After identification of the vHL gene on the short arm of chromosome 3 (3p25–26), the diagnosis is supported by molecular genetic analysis. Patients: In 20 patients with retinal angioma a clinical search for other manifestations of vHL was performed. In 5 patients only one angioma was present. In all patients molecular genetic tests for a mutation of the vHL gene were performed by SSCP and direct sequencing. Results: In 16 (80 %) patients vHL was present, and in 15 it could be diagnosed by clinical findings or a positive familiy history. Organ lesions in vHL patients were CNS hemangioblastoma in 10 (63 %), pancreatic cysts in 7 (43 %) and renal cysts in 7 (43 %) patients. In two patients (13 %) renal carcinoma could be detected; in one patient a pheochromcytoma was present. A mutation could be detected in all 15 patients with clinically confirmed vHL. In three patients a new mutation of vHL disease was diagnosed genetically. In one of these patients a single retinal angioma was the only sign of vHL. Conclusion: In patients presenting with capillary retinal angioma a careful search for other vHL lesions has to be performed. A mutation of the vHL gene can be detected in the majority of patients; thus, moleculargenetic testing is a powerful tool for diagnosis and detection of asymptomatic gene- carriers.ZusammenfassungHintergrund: Kapilläre retinale Angiome treten in der Regel im Rahmen eines von Hippel-Lindau-Syndroms (vHL), selten sporadisch auf. Bei allen Patienten mit einem solchen Tumor ist eine Untersuchung auf das Vorliegen weiterer Organmanifestationen eines vHL unerläßlich. Nachdem das auf dem kurzen Arm des Chromosoms 3 (3p25–26) lokalisierte vHL-Gen identifiziert wurde, kann die molekulargenetische Diagnostik die Diagnose bestätigen. Patienten und Methode: Bei 20 Patienten mit retinalen Angiomen wurden klinische Untersuchungen für weitere Organmanifestationen eines vHL durchgeführt. Bei 5 dieser Patienten bestand ein solitäres Angiom. Bei allen Patienten wurde eine molekulargenetische Untersuchung auf das Vorliegen einer Mutation des vHL-Gens durchgeführt. Ergebnisse: Bei 16 (80 %) Patienten bestand ein vHL, welches bei 15 Patienten klinisch oder familienanamnestisch nachweisbar war. Bei 10 (63 %) dieser Patienten bestanden zentralnervöse Hämangioblastome. Pankreaszysten und Nierenzysten fanden sich bei jeweils 7 Patienten (44 %), bei 2 Patienten (13 %) wurde ein Nierenzellkarzinom gefunden. Bei einem Patienten bestand ein Phäochromozytom. Molekulargenetisch konnte bei allen 15 Patienten mit klinisch gesichertem vHL eine Mutation des vHL-Gens nachgewiesen werden. Bei 3 Patienten lag wahrscheinlich eine Neumutation vor, bei einem dieser Patienten bestand lediglich ein solitäres retinales Angiom als einziges Zeichen des Syndroms. Schlußfolgerung: Bei Patienten mit kapillären retinalen Angiomen muß eine sorgfältige Suche nach weiteren Organmanifestationen eines von Hippel-Lindau-Syndroms erfolgen. Da eine Mutation des vHL-Gens bei den meisten erkrankten Patienten nachgewiesen werden kann, ist die molekulargenetische Diagnostik zur Sicherung der Diagnose und zur Untersuchung nicht erkrankter Verwandter auf Genträgerschaft geeignet.