Marta Barontini

MAX mutations cause hereditary and sporadic pheochromocytoma and paraganglioma.

Purpose: Pheochromocytomas (PCC) and paragangliomas (PGL) are genetically heterogeneous neural crest–derived neoplasms. Recently we identified germline mutations in a new tumor suppressor susceptibility gene, MAX (MYC-associated factor X), which predisposes carriers to PCC. How MAX mutations contribute to PCC/PGL and associated phenotypes remain unclear. This study aimed to examine the prevalence and associated phenotypic features of germline and somatic MAX mutations in PCC/PGL. Design: We sequenced MAX in 1,694 patients with PCC or PGL (without mutations in other major susceptibility genes) from 17 independent referral centers. We screened for large deletions/duplications in 1,535 patients using a multiplex PCR-based method. Somatic mutations were searched for in tumors from an additional 245 patients. The frequency and type of MAX mutation was assessed overall and by clinical characteristics. Results: Sixteen MAX pathogenic mutations were identified in 23 index patients. All had adrenal tumors, including 13 bilateral or multiple PCCs within the same gland (P < 0.001), 15.8% developed additional tumors at thoracoabdominal sites, and 37% had familial antecedents. Age at diagnosis was lower (P = 0.001) in MAX mutation carriers compared with nonmutated cases. Two patients (10.5%) developed metastatic disease. A mutation affecting MAX was found in five tumors, four of them confirmed as somatic (1.65%). MAX tumors were characterized by substantial increases in normetanephrine, associated with normal or minor increases in metanephrine. Conclusions: Germline mutations in MAX are responsible for 1.12% of PCC/PGL in patients without evidence of other known mutations and should be considered in the genetic work-up of these patients. Clin Cancer Res; 18(10); 2828–37. ©2012 AACR.

Spectrum and prevalence of FP/TMEM127 gene mutations in pheochromocytomas and paragangliomas.

CONTEXT Pheochromocytomas and paragangliomas are genetically heterogeneous neural crest-derived neoplasms. We recently identified germline mutations of the novel transmembrane-encoding gene FP/TMEM127 in familial and sporadic pheochromocytomas consistent with a tumor suppressor effect. OBJECTIVES To examine the prevalence and spectrum of FP/TMEM127 mutations in pheochromocytomas and paragangliomas and to test the effect of mutations in vitro. DESIGN, SETTING, AND PARTICIPANTS We sequenced the FP/TMEM127 gene in 990 individuals with pheochromocytomas and/or paragangliomas, including 898 previously unreported cases without mutations in other susceptibility genes from 8 independent worldwide referral centers between January 2009 and June 2010. A multiplex polymerase chain reaction-based method was developed to screen for large gene deletions in 545 of these samples. Confocal microscopy of 5 transfected mutant proteins was used to determine their subcellular localization. MAIN OUTCOME MEASURES The frequency and type of FP/TMEM127 mutation or deletion was assessed and correlated with clinical variables; the subcellular localization of 5 overexpressed mutants was compared with wild-type FP/TMEM127 protein. RESULTS We identified 19 potentially pathogenic FP/TMEM127 germline mutations in 20 independent families, but no large deletions were detected. All mutation carriers had adrenal tumors, including 7 bilateral (P = 2.7 × 10(-4)) and/or with familial disease (5 of 20 samples; P = .005). The median age at disease onset in the FP/TMEM127 mutation group was similar to that of patients without a mutation (41.5 vs 45 years, respectively; P = .54). The most common presentation was that of a single benign adrenal tumor in patients older than 40 years. Malignancy was seen in 1 mutation carrier (5%). Expression of 5 novel FP/TMEM127 mutations in cell lines revealed diffuse localization of the mutant proteins in contrast with the discrete multiorganelle distribution of wild-type TMEM127. CONCLUSIONS Germline mutations of FP/TMEM127 were associated with pheochromocytoma but not paraganglioma and occurred in an age group frequently excluded from genetic screening algorithms. Disease-associated mutations disrupt intracellular distribution of the FP/TMEM127 protein.

Peripheral Administration of an Angiotensin II AT1 Receptor Antagonist Decreases the Hypothalamic-Pituitary-Adrenal Response to Isolation Stress

Angiotensin II, which stimulates AT(1) receptors, is a brain and peripheral stress hormone. We pretreated rats with the AT(1) receptor antagonist candesartan for 13 d via sc-implanted osmotic minipumps, followed by 24-h isolation in individual metabolic cages. We measured angiotensin II receptor-type binding and mRNAs and tyrosine hydroxylase mRNA by quantitative autoradiography and in situ hybridization, catecholamines by HPLC, and hormones by RIA. Isolation increased AT(1) receptor binding in hypothalamic paraventricular nucleus as well as anterior pituitary ACTH, and decreased posterior pituitary AVP. Isolation stress also increased AT(1) receptor binding and AT(1B) mRNA in zona glomerulosa and AT(2) binding in adrenal medulla, adrenal catecholamines, tyrosine hydroxylase mRNA, aldosterone, and corticosterone. Candesartan blocked AT(1) binding in paraventricular nucleus and adrenal gland; prevented the isolation-induced alterations in pituitary ACTH and AVP and in adrenal corticosterone, aldosterone, and catecholamines; abolished the increase in AT(2) binding in adrenal medulla; and substantially decreased urinary AVP, corticosterone, aldosterone, and catecholamines during isolation. Peripheral pretreatment with an AT(1) receptor antagonist blocks brain and peripheral AT(1) receptors and inhibits the hypothalamic-pituitary-adrenal response to stress, suggesting a physiological role for peripheral and brain AT(1) receptors during stress and a possible beneficial effect of AT(1) antagonism in stress-related disorders.

Very early detection of RET proto-oncogene mutation is crucial for preventive thyroidectomy in multiple endocrine neoplasia type 2 children: presence of C-cell malignant disease in asymptomatic carriers.

Multiple endocrine neoplasia type 2 (MEN 2) is an inherited disease caused by germline mutations in the RET proto‐oncogene, and is responsible for the development of endocrine neoplasia. Its prognosis is dependent on the appearance and spread of medullary thyroid carcinoma (MTC). Relatives at risk can be identified before clinical or biochemical signs of the disease become evident.

Risk Profiles and Penetrance Estimations in Multiple Endocrine Neoplasia Type 2A Caused by Germline RET Mutations Located in Exon 10

Multiple endocrine neoplasia type 2 is characterized by germline mutations in RET. For exon 10, comprehensive molecular and corresponding phenotypic data are scarce. The International RET Exon 10 Consortium, comprising 27 centers from 15 countries, analyzed patients with RET exon 10 mutations for clinical‐risk profiles. Presentation, age‐dependent penetrance, and stage at presentation of medullary thyroid carcinoma (MTC), pheochromocytoma, and hyperparathyroidism were studied. A total of 340 subjects from 103 families, age 4–86, were registered. There were 21 distinct single nucleotide germline mutations located in codons 609 (45 subjects), 611 (50), 618 (94), and 620 (151). MTC was present in 263 registrants, pheochromocytoma in 54, and hyperparathyroidism in 8 subjects. Of the patients with MTC, 53% were detected when asymptomatic, and among those with pheochromocytoma, 54%. Penetrance for MTC was 4% by age 10, 25% by 25, and 80% by 50. Codon‐associated penetrance by age 50 ranged from 60% (codon 611) to 86% (620). More advanced stage and increasing risk of metastases correlated with mutation in codon position (609→620) near the juxtamembrane domain. Our data provide rigorous bases for timing of premorbid diagnosis and personalized treatment/prophylactic procedure decisions depending on specific RET exon 10 codons affected. Hum Mutat 31:1–8, 2010.

Characteristics of pheochromocytoma in a 4- to 20-year-old population.

Abstract:  Hypertension in children and adolescents has become a major health problem recently recognized, and in a significant number of patients it is due to an endocrine tumor. The aim of this study was to establish the characteristics of pheochromocytoma in a population of 58 patients between 4 and 20 years of age studied at our Center. They represented a 23% of the total population of 255 pheochromocytoma patients studied. In the younger group (under 20 years of age), there was a marked predominance of severe sustained hypertension (93%), only 7% presented paroxysmal hypertension and none of them was normotensive. The youngsters studied showed a higher incidence of bilateral adrenal pheochromocytoma (34%) and extra‐adrenal pheochromocytoma (22%). Malignancy was found in 12% of these patients. In addition, the incidence of familial pheochromocytoma was elevated in these patients (39%). Surprisingly, in contrast with the adult population where the most frequent familial pheochromocytomas were multiple endocrine neoplasia (MEN) type 2A (15%), the younger population showed a higher predominance of von Hippel–Lindau (VHL) (28%) and lower incidence of MEN 2A, MEN 2B, neurofibromatosis (NF), and succinate dehydrogenase subunit B (SDHB). In the VHL group, only two patients belonging to one family, showed the R167W mutation, while the others showed novel mutations in conserved amino acids. It may be speculated that the high incidence of VHL in youngsters may account for the biochemical and clinical features they usually present.

Long-term systemic hypertension in children after successful repair of coarctation of the aorta☆

The mechanisms responsible for long-term hypertension in children after successful repair of coarctation of the aorta have not yet been determined. We measured plasma renin activity and aldosterone, adrenalin, and noradrenalin concentrations both under basal conditions and in response to standing and treadmill exercise in 24 normal normotensive children, 16 normotensive postcoarctectomy children, eight hypertensive postcoarctectomy children, and seven children with essential hypertension. Exercise-induced changes in plasma renin activity, aldosterone, adrenalin, and noradrenalin were comparable in the four groups in spite of a significantly greater increase in systolic blood pressure in the children with hypertension. In response to standing, the plasma concentration of noradrenalin increased significantly in normotensive but not in hypertensive children. Hyperresponse of blood pressure to exercise in hypertensive postcoarctectomy children and children with essential hypertension is not related to abnormalities in the sympathetic nervous system or the angiotensin-aldosterone axis. Hypertension could be related to primary baroreceptor alterations, to structural changes in the arterial wall, or both. Twenty percent of normotensive postcoarctectomy children had a blood pressure hyperresponse to exercise and an abnormal noradrenalin response to standing similar to that seen in the hypertensive children. Follow-up of children after coarctectomy may elucidate whether these two abnormalities are indicators of an increased risk of developing long-term recurrent hypertension.

Mechanisms of Hypothalamic-Pituitary-Gonadal Disruption in Polycystic Ovarian Syndrome

Although the pathogenesis of polycystic ovarian syndrome (PCOS) is still controversial, a series of investigations has demonstrated an array of neuroendocrine abnormalities as a major component of the syndrome. From a neuroendocrine perspective, patients with PCOS exhibit an accelerated frequency and/or higher amplitude of LH pulses, augmentation of LH secretory burst mass, and a more disorderly LH release. Elevated in vitro LH bioactivity and a preponderance of basic LH isoforms, which correlate positively with elevated serum 17-hydroxyprogesterone, androstenedione, and testosterone concentrations, also characterize adolescents with PCOS. Heightened GnRH drive of gonadotropin secretion and a steroid-permissive milieu appear to jointly promote elevated secretion of basic LH isoforms. Positive feedback is implied, because hypersecretion of highly bioactive LH in PCOS probably contributes to inordinate androgen output. However, the precise nature of feedback disruption remains uncertain. Indeed, recent data suggest that PCOS is marked by anomalies of both feedforward and feedback signaling between GnRH/LH and ovarian androgens. From a single hormone perspective, the individual patterns of LH and androstenedione release are consistently more irregular in patients with PCOS. Bihormonal analysis has disclosed concomitant uncoupling of the pairwise synchrony of LH and testosterone, LH and androstenedione, and testosterone and androstenedione secretion. The foregoing ensemble of findings points to deterioration of both orderly uniglandular and coordinate bihormonal output in PCOS. Additional studies are needed to establish the primary pathophysiologic mechanisms underlying this disorder.

Recurrent mutations of chromatin remodeling genes and kinase receptors in pheochromocytomas and paragangliomas

Purpose: Pheochromocytomas and paragangliomas (PPGL) are genetically heterogeneous tumors of neural crest origin, but the molecular basis of most PPGLs is unknown. Experimental Design: We performed exome or transcriptome sequencing of 43 samples from 41 patients. A validation set of 136 PPGLs was used for amplicon-specific resequencing. In addition, a subset of these tumors was subjected to microarray-based transcription, protein expression, and histone methylation analysis by Western blotting or immunohistochemistry. In vitro analysis of mutants was performed in cell lines. Results: We detected mutations in chromatin-remodeling genes, including histone-methyltransferases, histone-demethylases, and histones in 11 samples from 8 patients (20%). In particular, we characterized a new cancer syndrome involving PPGLs and giant cell tumors of bone (GCT) caused by a postzygotic G34W mutation of the histone 3.3 gene, H3F3A. Furthermore, mutations in kinase genes were detected in samples from 15 patients (37%). Among those, a novel germline kinase domain mutation of MERTK detected in a patient with PPGL and medullary thyroid carcinoma was found to activate signaling downstream of this receptor. Recurrent germline and somatic mutations were also detected in MET, including a familial case and sporadic PPGLs. Importantly, in each of these three genes, mutations were also detected in the validation group. In addition, a somatic oncogenic hotspot FGFR1 mutation was found in a sporadic tumor. Conclusions: This study implicates chromatin-remodeling and kinase variants as frequent genetic events in PPGLs, many of which have no other known germline driver mutation. MERTK, MET, and H3F3A emerge as novel PPGL susceptibility genes. Clin Cancer Res; 22(9); 2301–10. ©2015 AACR.

Peripheral catecholamine alterations in adolescents with polycystic ovary syndrome

Polycystic ovary syndrome (PCO) is one of the most common endocrine disorders affecting women. Several lines of evidence have suggested the involvement of the sympathetic nervous system (SNS) in this condition. The present work was designed to assess neurochemically SNS activity in patients during the early stages of PCO.