Testing new susceptibility genes in the cohort of apparently sporadic phaeochromocytoma/paraganglioma patients with clinical characteristics of hereditary syndromes.

BACKGROUND: Phaeochromocytoma (PCC) and paraganglioma (PGL) can occur sporadically or as a part of familial cancer syndromes. Red flags of hereditary syndromes are young age and multifocal tumours. We hypothesized that such patients are candidates for further molecular diagnosis in case of normal results in 'classical' genes. MATERIAL AND METHODS: We selected patients with PCC/PGL under the age of 40 and/or with multiple tumours. First, we tested the genes RET, VHL, NF1, SDHB, SDHC and SDHD. Patients without mutations in these genes were tested for mutations in MAX, TMEM127 and SDHAF2. RESULTS: In 153 patients included, mutations were detected in the classical genes in 72 patients (47%) [RET-22 (14%), VHL-13 (9%), NF1-3 (2%), SDHB-13 (9%), SDHC-3 (2%), SDHD-16 (11%), SDHB large deletions- 2 (1%)]. One patient with MAXc.223C>T (p.R75X) mutation was detected. It was a male with bilateral, metachronous phaeochromocytomas diagnosed in 36 and 40 years of age. Remarkably, he showed in the period before the MAX gene was detected, a RET p. Y791F variant. During 10-year follow-up, we did not find any thyroid abnormalities. LOH examination of tumour tissue showed somatic loss of the wild-type allele of MAX. CONCLUSION: Analysis of the MAX gene should be performed in selected patients, especially those with bilateral adrenal phaeochromocytoma in whom mutations of the classical genes are absent. Our study provides with further support that Y791F RET is a polymorphism.

Pathogenicity of DNA variants and double mutations in multiple endocrine neoplasia type 2 and von Hippel-Lindau syndrome.

CONTEXT: Cancer genetics is fundamental for preventive medicine, in particular in pheochromocytoma-associated syndromes. Variants in two susceptibility genes, SDHC and RET, were found in a kindred with head and neck paraganglioma. This observation of coincident DNA variants, both reported as pathogenic, in two known susceptibility genes prompted the question of their pathogenic relevance. OBJECTIVE: Our objective was to elucidate the pathogenic role of the detected variants and study the prevalence of such variants. PATIENTS: Patients were registrants from the European-American Pheochromocytoma-Paraganglioma and German von Hippel-Lindau Disease Registries. DESIGN: Analysis of germline mutation screening results for all pheochromocytoma-paraganglioma susceptibility genes, including RET [multiple endocrine neoplasia type 2 (MEN 2)] and VHL [von Hippel-Lindau disease (VHL)]. Cases in which more than one DNA variant was found were clinically reevaluated, and cosegregation of the disease with the variant was analyzed within the registrants' families. A total of 1000 controls were screened for the presence of detected variants, and in silico analyses were performed. RESULTS: Three variants were identified, RET p.Tyr791Phe and p.Ser649Leu and VHL p.Pro81Ser. The frequencies of RET p.Ser649Leu (0.07%) and p.Tyr791Phe (0.9%) compared with controls excluded the two variants' role in the etiology of MEN 2 and VHL. None of the carriers of the RET variants who underwent prophylactic thyroidectomy showed medullary thyroid carcinoma. Clinical reinvestigation of 18 variant carriers excluded MEN 2. VHL variant p.Pro81Ser, also previously described as a mutation, did not segregate with the VHL in one family. In silico analyses for these variants predicted unmodified protein function. CONCLUSIONS: RET p.Tyr791Phe and p.Ser649Leu and VHL p.Pro81Ser are definitely not pathogenic mutations for VHL and MEN 2. Misinterpretation results in irreversible clinical consequences.

Serum CD40/CD40L system in Graves' disease and Hashimoto's thyroiditis related to soluble Fas, FasL and humoral markers of autoimmune response.

Activated CD4 T cells' express CD40 ligand (CD154) interacting with CD40 on the B cells surface, protecting them from Fas-mediated apoptosis and in this study, influence humoral response. The aim of the study was to assess soluble CD40 and CD154 in Graves' disease (GD) and Hashimoto's thyroiditis (HT) in relation to Fas and FasL and to the markers of humoral response: aTPO, aTG and aTSHR. The study was carried out in 5 groups of subjects: 1/14 patients with GD in euthyreosis on methimazol (euGD), 2/20 patients with hyperthyroid GD (hrGD), 3/15 patients with HT in euthyreosis on levothyroxine (euHT), 4/16 patients with hypothyroid Ht (hoHT), 5/12 healthy volunteers, age and sex-matched to groups 1-4. The serum levels of CD40, CD154, Fas and FasL, aTPO and aTG were determined by ELISA and aTSHR was determined by the RIA method. CD40 serum concentration was significantly higher in hoHT individuals: 55.8 (24.0-83.2) pg/ml (p<0.01) and euHT patients: 51.2 (20.0-80.1) (p<0.05) as compared to the controls. Also sCD40L values were significantly increased in euHT individuals: 5.1 (1.0-11.8) (p<0.05) and hoHT patients: 3.9 (0.7-11.2) ng/ml (p<0.05) as compared to the controls. There was a positive correlation between sCD40 and sCD154 in the patients studied (r=0.36, p<0.001). In HT patients we found positive correlations between sCD40 and aTPO (r=0.45, p<0.001) and sFas (r=0.36, p<0.05) as well as a negative correlation between sCD40 and FasL (r=-0.24, p<0.05). In GD patients there was a positive correlation between sCD40 and aTSHR (r=0.28, p<0.05). In summary, our results suggest that CD40/CD154 interaction plays an important role in the regulation of autoimmune humoral response, both in Hashimoto's thyroiditis and Graves' disease. Fas-mediated apoptosis seems to be involved in this process especially in Hashimoto thyroiditis. Soluble CD40 may serve as a marker of the active stage of autoimmune thyroid disease.