Rajesh V. Thakker

Gastroenteropancreatic neuroendocrine tumours

Gastroenteropancreatic (GEP) neuroendocrine tumours (NETs) are fairly rare neoplasms that present many clinical challenges. They secrete peptides and neuroamines that cause distinct clinical syndromes, including carcinoid syndrome. However, many are clinically silent until late presentation with mass effects. Investigation and management should be highly individualised for a patient, taking into consideration the likely natural history of the tumour and general health of the patient. Management strategies include surgery for cure (which is achieved rarely) or for cytoreduction, radiological intervention (by chemoembolisation and radiofrequency ablation), chemotherapy, and somatostatin analogues to control symptoms that result from release of peptides and neuroamines. New biological agents and somatostatin-tagged radionuclides are under investigation. The complexity, heterogeneity, and rarity of GEP NETs have contributed to a paucity of relevant randomised trials and little or no survival increase over the past 30 years. To improve outcome from GEP NETs, a better understanding of their biology is needed, with emphasis on molecular genetics and disease modeling. More-reliable serum markers, better tumour localisation and identification of small lesions, and histological grading systems and classifications with prognostic application are needed. Comparison between treatments is currently very difficult. Progress is unlikely to occur without development of centers of excellence, with dedicated combined clinical teams to coordinate multicentre studies, maintain clinical and tissue databases, and refine molecularly targeted therapeutics.

Guidelines for the management of gastroenteropancreatic neuroendocrine (including carcinoid) tumours (NETs)

These guidelines update previous guidance published in 2005. They have been revised by a group who are members of the UK and Ireland Neuroendocrine Tumour Society with endorsement from the clinical committees of the British Society of Gastroenterology, the Society for Endocrinology, the Association of Surgeons of Great Britain and Ireland (and its Surgical Specialty Associations), the British Society of Gastrointestinal and Abdominal Radiology and others. The authorship represents leaders of the various groups in the UK and Ireland Neuroendocrine Tumour Society, but a large amount of work has been carried out by other specialists, many of whom attended a guidelines conference in May 2009. We have attempted to represent this work in the acknowledgements section. Over the past few years, there have been advances in the management of neuroendocrine tumours, which have included clearer characterisation, more specific and therapeutically relevant diagnosis, and improved treatments. However, there remain few randomised trials in the field and the disease is uncommon, hence all evidence must be considered weak in comparison with other more common cancers.

HRPT2, encoding parafibromin, is mutated in hyperparathyroidism-jaw tumor syndrome.

We report here the identification of a gene associated with the hyperparathyroidism–jaw tumor (HPT–JT) syndrome. A single locus associated with HPT–JT (HRPT2) was previously mapped to chromosomal region 1q25–q32. We refined this region to a critical interval of 12 cM by genotyping in 26 affected kindreds. Using a positional candidate approach, we identified thirteen different heterozygous, germline, inactivating mutations in a single gene in fourteen families with HPT–JT. The proposed role of HRPT2 as a tumor suppressor was supported by mutation screening in 48 parathyroid adenomas with cystic features, which identified three somatic inactivating mutations, all located in exon 1. None of these mutations were detected in normal controls, and all were predicted to cause deficient or impaired protein function. HRPT2 is a ubiquitously expressed, evolutionarily conserved gene encoding a predicted protein of 531 amino acids, for which we propose the name parafibromin. Our findings suggest that HRPT2 is a tumor-suppressor gene, the inactivation of which is directly involved in predisposition to HPT–JT and in development of some sporadic parathyroid tumors.

A Familial Syndrome of Hypocalcemia with Hypercalciuria Due to Mutations in the Calcium-Sensing Receptor

BACKGROUND The calcium-sensing receptor regulates the secretion of parathyroid hormone in response to changes in extracellular calcium concentrations, and mutations that result in a loss of function of the receptor are associated with familial hypocalciuric hypercalcemia. Mutations involving a gain of function have been associated with hypocalcemia in two kindreds. We examined the possibility that the latter type of mutation may result in a phenotype of familial hypocalcemia with hypercalciuria. METHODS We studied six kindreds given a diagnosis of autosomal dominant hypoparathyroidism on the basis of their hypocalcemia and normal serum parathyroid hormone concentrations, a combination that suggested a defect of the calcium-sensing receptor. The hypocalcemia was associated with hypercalciuria, and treatment with vitamin D resulted in increased hypercalciuria, nephrocalcinosis, and renal impairment. Mutations in the calcium-sensing-receptor gene were identified by DNA-sequence analysis and expressed in human embryonic kidney cells (HEK-293). RESULTS Five heterozygous missense mutations (Asn118Lys, Phe128Leu, Thr151Met, Glu191Lys, and Phe612Ser) were detected in the extracellular domain of the calcium-sensing-receptor gene and shown to cosegregate with the disease. Analysis of the functional expression of three of the mutant receptors in HEK-293 cells demonstrated shifts in the dose-response curves so that the extracellular calcium concentrations needed to produce half-maximal increases in total inositol phosphate in the cells were significantly (P=0.02 to P<0.001) lower than those required for the wild-type receptor. CONCLUSIONS Gain-of-function mutations in the calcium-sensing receptor are associated with a familial syndrome of hypocalcemia with hypercalciuria that needs to be distinguished from hypoparathyroidism.

Clinical practice guidelines for multiple endocrine neoplasia type 1 (MEN1).

OBJECTIVE The aim was to provide guidelines for evaluation, treatment, and genetic testing for multiple endocrine neoplasia type 1 (MEN1). PARTICIPANTS The group, which comprised 10 experts, including physicians, surgeons, and geneticists from international centers, received no corporate funding or remuneration. PROCESS Guidelines were developed by reviews of peer-reviewed publications; a draft was prepared, reviewed, and rigorously revised at several stages; and agreed-upon revisions were incorporated. CONCLUSIONS MEN1 is an autosomal dominant disorder that is due to mutations in the tumor suppressor gene MEN1, which encodes a 610-amino acid protein, menin. Thus, the finding of MEN1 in a patient has important implications for family members because first-degree relatives have a 50% risk of developing the disease and can often be identified by MEN1 mutational analysis. MEN1 is characterized by the occurrence of parathyroid, pancreatic islet, and anterior pituitary tumors. Some patients may also develop carcinoid tumors, adrenocortical tumors, meningiomas, facial angiofibromas, collagenomas, and lipomas. Patients with MEN1 have a decreased life expectancy, and the outcomes of current treatments, which are generally similar to those for the respective tumors occurring in non-MEN1 patients, are not as successful because of multiple tumors, which may be larger, more aggressive, and resistant to treatment, and the concurrence of metastases. The prognosis for MEN1 patients might be improved by presymptomatic tumor detection and undertaking treatment specific for MEN1 tumors. Thus, it is recommended that MEN1 patients and their families should be cared for by multidisciplinary teams comprising relevant specialists with experience in the diagnosis and treatment of patients with endocrine tumors.

GATA3 haplo-insufficiency causes human HDR syndrome

Terminal deletions of chromosome 10p result in a DiGeorge-like phenotype that includes hypoparathyroidism, heart defects, immune deficiency, deafness and renal malformations. Studies in patients with 10p deletions have defined two non-overlapping regions that contribute to this complex phenotype. These are the DiGeorge critical region II (refs 1, 2), which is located on 10p13-14, and the region for the hypoparathyroidism, sensorineural deafness, renal anomaly (HDR) syndrome (Mendelian Inheritance in Man number 146255), which is located more telomeric (10p14–10pter). We have performed deletion-mapping studies in two HDR patients, and here we define a critical 200-kilobase region which contains the GATA3 gene. This gene belongs to a family of zinc-finger transcription factors that are involved in vertebrate embryonic development. Investigation for GATA3 mutations in three other HDR probands identified one nonsense mutation and two intragenic deletions that predicted a loss of function, as confirmed by absence of DNA binding by the mutant GATA3 protein. These results show that GATA3 is essential in the embryonic development of the parathyroids, auditory system and kidneys, and indicate that other GATA family members may be involved in the aetiology of human malformations.

Characterization of Mutations in Patients with Multiple Endocrine Neoplasia Type 1

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder characterized by tumors of the parathyroids, pancreatic islets, and anterior pituitary. The MEN1 gene, on chromosome 11q13, has recently been cloned, and mutations have been identified. We have characterized such MEN1 mutations, assessed the reliability of SSCP analysis for the detection of these mutations, and estimated the age-related penetrance for MEN1. Sixty-three unrelated MEN1 kindreds (195 affected and 396 unaffected members) were investigated for mutations in the 2,790-bp coding region and splice sites, by SSCP and DNA sequence analysis. We identified 47 mutations (12 nonsense mutations, 21 deletions, 7 insertions, 1 donor splice-site mutation, and 6 missense mutations), that were scattered throughout the coding region, together with six polymorphisms that had heterozygosity frequencies of 2%-44%. More than 10% of the mutations arose de novo, and four mutation hot spots accounted for >25% of the mutations. SSCP was found to be a sensitive and specific mutational screening method that detected >85% of the mutations. Two hundred and one MEN1 mutant-gene carriers (155 affected and 46 unaffected) were identified, and these helped to define the age-related penetrance of MEN1 as 7%, 52%, 87%, 98%, 99%, and 100% at 10, 20, 30, 40, 50, and 60 years of age, respectively. These results provide the basis for a molecular-genetic screening approach that will supplement the clinical evaluation and genetic counseling of members of MEN1 families.

Association of Parathyroid Tumors in Multiple Endocrine Neoplasia Type 1 with Loss of Alleles on Chromosome 11

Familial multiple endocrine neoplasia type 1 (MEN-1) is an autosomal dominant disorder characterized by the combined occurrence of tumors of the parathyroid glands, the pancreas, and the pituitary gland. Pancreatic tumors have previously been shown to be associated with the loss of alleles on chromosome 11; we therefore looked for similar genetic alterations in specimens of parathyroid tumors, which are the most common feature of MEN-1. We obtained parathyroid tumors and peripheral-blood leukocytes from six patients with MEN-1; 18 cloned human DNA sequences from chromosome 11 were then used to identify restriction-fragment-length polymorphisms. A loss of heterozygosity was detected in parathyroid tumors from three of the six patients with MEN-1; this finding demonstrated that allelic deletions on chromosome 11 are involved in the monoclonal development of parathyroid tumors in patients with MEN-1. In addition, studies of three affected families (with 17 affected members and 51 unaffected members) established linkage with the oncogene INT2 (peak lod score, 3.30, at 0 percent recombination); the MEN-1 gene was thus mapped to the pericentromeric region of the long arm of chromosome 11 (11q13). Our location of the MEN-1 gene at 11q13 is close to the location previously reported. We conclude that a single inherited locus on chromosome 11, band q13, causes MEN-1 and that the monoclonal development of parathyroid and pancreatic tumors in patients with MEN-1 involves similar allelic deletions on chromosome 11.

Multiple Endocrine Neoplasia

Multiple endocrine neoplasia (MEN) is characterized by the occurrence of tumours involving two or more endocrine glands; two major forms, referred to as MEN1 and MEN2, are recognized. MEN1 is characterized by parathyroid, pancreatic islet and anterior pituitary tumours, whilst MEN2 is characterized by medullary thyroid carcinoma (MTC) in association with phaeochromocytoma. There are three clinical variants, referred to as MEN2A, MEN2B and MTC-only. All these forms of MEN may be inherited as autosomal dominant syndromes. The MEN1 gene is on chromosome 11q13 and about 300 MEN1 mutations have been identified. These are of diverse types and are scattered throughout the coding region. There is also a lack of genotype-phenotype correlation. All these findings make it difficult to implement MEN1 mutational analysis in the clinical setting. The situation in MEN2 is more straightforward. The gene causing all three MEN2 variants is located on chromosome 10cen-10q11.2, and is the c-ret proto-oncogene which encodes a tyrosine kinase receptor with cadherin-like and cysteine-rich extracellular domains, and a tyrosine kinase intracellular domain. Specific mutations of c-ret have been identified for each of the three MEN2 variants and mutational analysis has been used in the diagnosis and management of patients and families with the MEN2 variants.

Genetic Contribution to Bone Metabolism, Calcium Excretion, and Vitamin D and Parathyroid Hormone Regulation

A classical twin study was performed to assess the relative contribution of genetic and environmental factors to bone metabolism, calcium homeostasis, and the hormones regulating them. It was examined further whether the genetic effect is menopause dependent. The subjects were 2136 adult twins (98.3% female): 384 monozygotic (MZ) and 684 dizygotic (DZ) twin pairs. The intraclass correlations were calculated, and maximum likelihood model fitting was used to estimate genetic and environmental variance components. The intraclass correlations for all of the variables assessed were higher in MZ twin pairs. The heritabilities (95% CIs) obtained from model fitting for hormones regulating bone metabolism and calcium homeostasis were parathyroid hormone (PTH), 60% (54–65%); 25‐hydroxyvitamin D [25(OH)D]; 43% (28–57%), 1,25‐hydroxyvitamin D [1,25(OH)], 65% (53–74%); and vitamin D binding protein 62% (56–66%). The heritabilities (95% CIs) for markers of bone formation also were assessed; bone‐specific alkaline phosphatase (BSAP), 74% (67–80%), and osteocalcin, 29% (14–44%); marker of bone resorption deoxypyridinoline (DPD), 58% (52–64%); and measure of calcium homeostasis 24 h urine calcium, creatinine (Cr), 52% (41–61%). The magnitude of genetic influence differed with menopause for most variables. This study provides evidence for the importance of genetic factors in determining bone resorption and formation, calcium excretion, and the hormones regulating these processes. It shows for the first time a clear genetic effect on bone resorption in premenopausal women and the regulation of PTH, vitamin D metabolism, and calcium excretion. The genes controlling bone hormones and markers are likely to be useful therapeutic and diagnostic targets.