Challenges in genetic screening for inherited endocrinopathy affecting the thyroid, parathyroid and adrenal glands in Singapore.

Significant progress has been made in the understand-ing of many human diseases, especially cancers, which has contributed to improved and increased survival. The Human Genome Project and The Cancer Genome Atlas project brought about a new era, with an understanding of inherited diseases at a molecular level, which subsequently facilitated the option of precision medicine. Precision medicine has helped tailor treatment decisions at an individual level, for instance in terms of surgical treatments or targeted therapies in advanced diseases. Despite the increasing advances in genetic-lead precision medicine, this has not translated into increasing uptake among patients. Reasons for this may be potential knowledge gaps among clinicians; on reasons for poor uptake of genetic testing such as for cultural, religious or personal beliefs; and on financial implications such as lack of support from insurance companies. In this review, we look at the current scenario of genetic screening for common inherited endocrine conditions affecting the thyroid, parathyroid and adrenal glands in Singapore, and the implications associated with it.

Monoclonal/polyclonal PAX-8, PTH and GATA3 immunohistochemistry in parathyroid lesions.

BACKGROUND: Parathyroid gland (PG) is an endocrine organ which may display different immunohistochemical stainings with chief cells and oxyphilic cells in normal as well as hyperplasic/tumoral lesions. PURPOSE: In this study, we aimed to identify the demographic properties and diagnostic value of the GATA3 antibody, which is a transcription factor in addition to PTH, and of PAX-8 (monoclonal and polyclonal) antibody. METHODS: We have analyzed in detail the cellular components and staining intensities of 46 adenomas all of which contained parathyroid rims, 12 hyperplasia and 5 adjacent non-neoplastic thyroidectomy materials (63 patients, 114 tissues). RESULTS: While no staining was identified in the thyroid tissue, cytoplasmic PTH immunoreactivity was observed in all (100%) normal parathyroid tissues, rim of PGs and hyperplasia, and in 43/46 cases (93.4%) of adenomas. Adenoma and hyperplasia were less stained than normal PG (p < 0.05). We detected GATA3 staining in all cases except for the thyroid (100%). Weak positivity (1+) was most apparent in adenoma cases (p < 0.05). Monoclonal PAX-8 immunoreactivity was not identified in any normal parathyroid tissue and rim of PG but positive immunoreactivity was detected in 83.3% of hyperplasia cases (10/12), 84.8% of adenoma (39/46) and 100% of thyroid tissues (5/5) (p < 0.05). However, polyclonal PAX-8 immunoreactivity was detected in one normal parathyroid tissue (1/5) and seven (7/46) rim of PGs. In cases of hyperplasia and adenoma, positive immunoreactivity was 75% (9/12) and 74% (34/46), respectively. CONCLUSION: In conclusion, we have observed that PTH and GATA3 constitute a much more reliable and sensitive marker for parathyroid and are stained less in adenomas. While monoclonal PAX-8 (MRQ-50) never stains normal parathyroid and rim of PGs, it may help in the differential diagnosis of proliferated parathyroid lesions as a considerably sensitive and relatively specific marker by staining hyperplasic parathyroid, adenomas and the thyroid.

Hereditary Parathyroid Disease: Sometimes Pathologists Do Not Know What They Are Missing.

Parathyroid gland excision specimens are common and sometimes underestimated cases that many surgical pathologists encounter regularly. In the vast majority of cases, these will be spot diagnoses of sporadic primary parathyroid adenomas or, perhaps, hyperplasias commonly in the setting of renal failure. However, a small but significant number of parathyroid gland excisions may be due to heritable disease. In most cases, hereditary disease is suspected by the referring clinicians. Nevertheless, a subset of these are undetected which is significant, particularly in the setting of the multiple endocrine neoplasia (MEN), and the hyperparathyroidism jaw tumour (HPT-JT) syndromes. There have been recent advances in recognition of the morphological and immunohistochemical characteristics of these tumours and hyperplasias. While hereditary kindreds are over-represented at specialist referral centres, with awareness of the characteristic clinical and morphological features, the general surgical pathologist is frequently able to suggest the possibility of hereditary parathyroid disease. We therefore provide a succinct guide for pathologists to increase the recognition of hereditary parathyroid disease.

Genetic Disorders of Parathyroid Development and Function.

Hypoparathyroidism is characterized by hypocalcemia and hyperphosphatemia and is due to insufficient levels of circulating parathyroid hormone. Hypoparathyroidism may be an isolated condition or a component of a complex syndrome. Although genetic disorders are not the most common cause of hypoparathyroidism, molecular analyses have identified a growing number of genes that when defective result in impaired formation of the parathyroid glands, disordered synthesis or secretion of parathyroid hormone, or postnatal destruction of the parathyroid glands.

Genetics of parathyroids disorders: Overview.

Several familial forms of primary hyperparathyroidism (PHTP) have been discovered over the past 25 years, and molecular test for their risk assessment has been widely increasing. These syndromic and non-syndromic forms have received benefits from the identification of the responsible genes whose mutations account for the genetic susceptibility to develop parathyroid tumours as also other endocrine and nonendocrine tumours. In recent years, care options have been made available to patients and families with hereditary PHPT, and the process of systematically assessing the genetic risk has been becoming increasingly important. The aim of this review is to help health providers not frequently dealing with genetic testing use, introducing general concepts with regard to genetic diagnosis issues. The role and the practical usefulness of DNA-based diagnosis in patients affected by different forms of "congenital" PHPT is described, closely looking on why, when and how genetic testing should be performed in these subjects and their relatives. Moreover, this review will provide some practical suggestions and recommendations concerning on how to deal with a suspected or known case of familial PHPT.

Oxyphil cell metaplasia in the parathyroids is characterized by somatic mitochondrial DNA mutations in NADH dehydrogenase genes and cytochrome c oxidase activity-impairing genes.

Oxyphil cell transformation of epithelial cells due to the accumulation of mitochondria occurs often during cellular aging. To understand the pathogenic mechanisms, we studied mitochondrial DNA (mtDNA) alterations in the three cell types of the parathyroids using multiplex real-time PCR and next-generation sequencing. mtDNA was analyzed from cytochrome c oxidase (COX)-positive and COX-negative areas of 19 parathyroids. Mitochondria-rich pre-oxyphil/oxyphil cells were more prone to develop COX defects than the mitochondria-poor clear chief cells (P < 0.001). mtDNA increased approximately 2.5-fold from clear chief to oxyphil cells. In COX deficiency, the increase was even more pronounced, and COX-negative oxyphil cells had approximately two times more mtDNA than COX-positive oxyphil cells (P < 0.001), illustrating the influence of COX deficiency on mtDNA biosynthesis, probably as a consequence of insufficient ATP synthesis. Next-generation sequencing revealed a broad spectrum of putative pathogenic mtDNA point mutations affecting NADH dehydrogenase and COX genes as well as regulatory elements of mtDNA. NADH dehydrogenase gene mutations preferentially accumulated in COX-positive pre-oxyphil/oxyphil cells and, therefore, could be essential for inducing oxyphil cell transformation by increasing mtDNA/mitochondrial biogenesis. In contrast, COX-negative cells predominantly harbored mutations in the MT-CO1 and MT-CO3 genes and in regulatory mtDNA elements, but only rarely NADH dehydrogenase mutations. Thus, multiple hits in NADH dehydrogenase and COX activity-impairing genes represent the molecular basis of oxyphil cell transformation in the parathyroids.

Parathyroid hormone-related protein: an update.

PTHrP was identified as a cause of hypercalcemia in cancer patients 25 yr ago. In the intervening years, we have learned that PTHrP and PTH are encoded by related genes that are part of a larger "PTH gene family." This evolutionary relationship permits them to bind to the same type 1 PTH/PTHrP receptor, which explains why humoral hypercalcemia of malignancy resembles hyperparathyroidism. This review will outline basic facts about PTHrP biology and its normal physiological functions, with an emphasis on new findings of the past 5-10 yr. The medical and research communities first became aware of PTHrP because of its involvement in a common paraneoplastic syndrome. Now, research into the basic biology of PTHrP has suggested previously unrecognized connections to a variety of disease states such as osteoporosis, osteoarthritis, and breast cancer and has highlighted how PTHrP itself might be used in therapy for osteoporosis and diabetes. Therefore, the story of this remarkable protein is a paradigm for translational research, having gone from bedside to bench and now back to bedside.

Molecular genetics of parathyroid disease.

BACKGROUND: Primary hyperparathyroidism (HPT) is often caused by a benign parathyroid tumor, adenoma; less commonly by multiglandular parathyroid disease/hyperplasia; and rarely by parathyroid carcinoma. Patients with multiple tumors require wider exploration to avoid recurrence and have increased risk for hereditary disease. Secondary HPT is a common complication of renal failure. Improved knowledge of the molecular background of parathyroid tumor development may help select patients for appropriate surgical treatment and can eventually provide new means of treatment. The present contribution summarizes more recent knowledge of parathyroid molecular genetics. METHODS: A literature search and review was made to evaluate the level of evidence concerning molecular biology and genetics of primary, secondary, and familial HPT according to criteria proposed by Sackett, with recommendation grading by Heinrich et al. RESULTS: Most parathyroid adenomas and hyperplastic glands are monoclonal lesions. Cyclin D1 gene (CCND1) translocation and oncogene action occur in 8% of adenomas; cyclin D1 overexpression is seen in 20% to 40% of parathyroid adenomas and in 31% of secondary hyperplastic glands. Somatic loss of one MEN1 allele is seen in 25% to 40% of sporadic parathyroid adenomas, half of which have inactivating mutation of the remaining allele. Inactivating somatic HRPT2 mutations are common in parathyroid carcinoma, often causing decreased expression of the protein parafibromin involved in cyclin D1 regulation. Aberrant regulation of Wnt/beta-catenin signaling may be important for parathyroid tumor development. CONCLUSIONS: Molecular genetic studies of parathyroid tumors are well designed basic experimental studies providing strong level III evidence, with data frequently confirmed by subsequent studies.

Molecular alterations in hereditary and sporadic thyroid and parathyroid diseases.

Thyroid and parathyroid diseases are fairly common and can be either hereditary or sporadic in nature. Tumors and tumor-like processes account for the majority of surgical pathology specimens in both of these endocrine organs. Molecular alterations are well known to occur in both the hereditary and the sporadic settings, and include alterations in tumor suppressor genes and oncogenes. The genetic pathways of tumors of parathyroid and thyroid are beginning to be well understood and are proving to be useful diagnostic, prognostic, and potential therapeutic targets. The molecular alterations in parathyroid and thyroid tumors and tumor-like processes are reviewed, with a focus on the potentially clinically useful diagnostic markers.

Familial risks for hospitalization with endocrine diseases.

CONTEXT: Familial clustering of a disease is an indicator of a possible heritable cause. In the era of genome scans, the consideration of data on heritability should be important in the assessment of the likely success of the scans. OBJECT: The objective of the study was to carry out a family study on nonthyroid endocrine diseases to search familial clustering of these diseases beyond the known syndromes. DESIGN AND SETTING: The Swedish Multigeneration Register on 0- to 72-yr-old subjects was linked to the Hospital Discharge Register from years 1964 to 2004. MAIN OUTCOME MEASURE: Standardized incidence ratios were calculated for offspring of affected parents and siblings by comparing with those whose relatives had no hospitalization for nonthyroid endocrine diseases. RESULTS: A total of 11,948 hospitalized cases and 443 familial cases were identified. The familial standardized incidence ratios were increased for parathyroid, pituitary, and adrenal hyperfunctions and hypofunctions, some findings consistent with known syndromes, most clearly that for adrenal cortical hypofunction showing recessive inheritance described for autoimmune polyendocrine syndrome 1. The sibling risks were very high for many diseases, but some of these affecting young individual may be due to bias caused by selective hospitalization. A high sibling risk observed for anterior pituitary hypofunction may represent a yet-unknown recessive syndrome. CONCLUSIONS: To our knowledge this is a first population-based study on nonthyroid endocrine diseases. The results call for further studies to sort out the challengingly high sibling risk for many individual nonthyroid endocrine diseases, whether they are due to bias or possible recessive effects.

The calcium-sensing receptor and related diseases.

The calcium-sensing receptor (CASR) adjusts the extracellular calcium set point regulating PTH secretion and renal calcium excretion. The receptor is expressed in several tissues and is also involved in other cellular functions such as proliferation, differentiation and other hormonal secretion. High extracellular calcium levels activate the receptor resulting in modulation of several signaling pathways depending on the target tissues. Mutations in the CASR gene can result in gain or loss of receptor function. Gain of function mutations are associated to Autossomal dominant hypocalcemia and Bartter syndrome type V, while loss of function mutations are associated to Familial hypocalciuric hypercalcemia and Neonatal severe hyperparathyroidism. More than one hundred mutations were described in this gene. In addition to calcium, the receptor also interacts with several ions and polyamines. The CASR is a potential therapeutic target to treatment of diseases including hyperparathyroidism and osteoporosis, since its interaction with pharmacological compounds results in modulation of PTH secretion.

The calcium-sensing receptor and related diseases / O receptor sensor de cálcio e doenças associadas

The calcium-sensing receptor (CASR) adjusts the extracellular calcium set point regulating PTH secretion and renal calcium excretion. The receptor is expressed in several tissues and is also involved in other cellular functions such as proliferation, differentiation and other hormonal secretion. High extracellular calcium levels activate the receptor resulting in modulation of several signaling pathways depending on the target tissues. Mutations in the CASR gene can result in gain or loss of receptor function. Gain of function mutations are associated to Autossomal dominant hypocalcemia and Bartter syndrome type V, while loss of function mutations are associated to Familial hypocalciuric hypercalcemia and Neonatal severe hyperparathyroidism. More than one hundred mutations were described in this gene. In addition to calcium, the receptor also interacts with several ions and polyamines. The CASR is a potential therapeutic target to treatment of diseases including hyperparathyroidism and osteoporosis, since its interaction with pharmacological compounds results in modulation of PTH secretion.

O receptor sensor de cálcio (CASR) ajusta o set point do cálcio extracelular através da regulação da secreção de PTH e da excreção renal de cálcio. O receptor é expresso em diversos tecidos e também está envolvido em outras funções celulares como proliferação, diferenciação e secreção de outros hormônios. Concentrações altas de cálcio extracelular ativam o receptor resultando em modulação de inúmeras vias de sinais intracelulares dependendo do tecido-alvo. Mutações no gene do CASR podem resultar em ganho ou perda de função do receptor. Mutações com ganho de função são associadas à Hipocalcemia autossômica dominante e à Síndrome de Bartter tipo V, enquanto que mutações com perda de função são associadas à Hipercalcemia hipocalciúrica familiar e ao Hiperparatireoidismo neonatal grave. Mais de cem mutações foram descritas neste gene. Além do cálcio, o receptor também interage com inúmeros íons e poliaminas. CASR é um alvo terapêutico potencial para tratamento de doenças incluindo hiperparatireoidismo e osteoporose, pois a sua interação com compostos farmacológicos resulta em modulação da secreção de PTH.

Dermatologic manifestations of parathyroid-related disorders.

Dermatologic manifestations of parathyroid-related disorders, although rare in sporadic cases, are not uncommon in familial syndromes. Patients with familial hyperparathyroidism have several types of skin lesions. In multiple endocrine neoplasia 1, patients commonly have angiofibromas (85%) and collagenomas (70%), lesions that show loss of one 11q13 allele, the molecular abnormality in multiple endocrine neoplasia 1. They can also present with lipomas or café-au-lait spots. Cutaneous amyloidosis, an entity that can occur sporadically, has been described in multiple endocrine neoplasia 2a and is usually localized to the interscapular area. Metastatic calcification is an entity commonly encountered in patients with hyperparathyroidism and renal failure. It can be complicated by infections and necrosis. It is best treated by controlling hypercalcemia, hyperphosphatemia, hyperparathyroidism, antibiotics, and analgesia. Parathyroidectomy is reserved for refractory cases. Hypoparathyroidism presenting in the context of polyglandular failure type 1 is characterized by mucocutaneous candidiasis. Pseudohypoparathyroidism, an inherited disorder with end-organ unresponsiveness to parathyroid hormone, is characterized by Albright hereditary osteodystrophy. Patients present with short stature, round facies, brachydactyly, and short fourth or fifth metacarpals.

The spectrum of parathyroid gland dysfunction associated with the microdeletion 22q11.

OBJECTIVE: Clinical features associated with microdeletion of chromosome 22q11 (del(22)(q11)) are highly variable. Increased awareness of this condition is needed among specialists such as endocrinologists to reduce diagnostic delay and improve clinical care. The purpose of this study was to describe the phenotype of patients with del(22)(q11), focusing on parathyroid gland dysfunction. DESIGN AND METHODS: Charts of 19 patients, including one kindred of three, known to have del(22)(q11) diagnosed by fluorescence in situ hybridization (FISH) were reviewed from the register of the department of Medical Genetics. Major clinical features including hypoparathyroidism phenotype were collected. RESULTS: Parathyroid dysfunction was present in 8 out of 16 patients (50%). Six patients were diagnosed with overt hypoparathyroidism. Hypocalcemia manifested as laryngeal stridor within the first days of life (n = 3), seizures in infancy (n = 1) and adolescence (n = 2). The connection between hypoparathyroidism and diagnosis of del(22)(q11) was belated at the median age of 18 years. One patient had presented with transient neonatal hypoparathyroidism, and one patient had latent hypoparathyroidism. Within the kindred family, the phenotype variability including that of parathyroid dysfunction was as marked as between unrelated individuals. Standard karyotype failed to detect the deletion in 15 out of 19 cases. CONCLUSIONS: Abnormal parathyroid function in the del(22)(q11) ranges from severe neonatal hypocalcemia to latent hypoparathyroidism. Del(22)(q11) should be considered as a potential cause of hypocalcemia even in young adult. When suspected, the diagnosis requires investigation by FISH. Furthermore, long-term calcemia follow-up is needed in normocalcemic patients with del(22)(q11) because of the possible evolution to hypocalcemic hypoparathyroidism.

The roles of specific genes implicated as circulating factors involved in normal and disordered phosphate homeostasis: frizzled related protein-4, matrix extracellular phosphoglycoprotein, and fibroblast growth factor 23.

Normal serum phosphate (Pi) concentrations are relatively tightly controlled by endocrine mediators of Pi balance. Recent data involving several disorders of Pi homeostasis have shed new light on the regulation of serum Pi balance. It has been hypothesized that circulating phosphaturic factors, or phosphatonins, exist that, when present at high serum concentrations, directly act on the kidney to induce renal Pi wasting. This review will focus upon recently discovered factors that are overexpressed in tumors associated with tumor-induced osteomalacia and have reported activity consistent with effecting Pi balance in vivo. Currently, the best-characterized group of phosphatonin-like polypeptides includes secreted frizzled related protein-4, matrix extracellular phosphoglycoprotein, and fibroblast growth factor-23. Our understanding of these factors will, in the short term, aid us in understanding normal Pi balance and, in the future, help to design novel therapeutic strategies for disorders of Pi handling.

Clinical manifestations of familial medullary thyroid carcinoma.

We conducted a large-scale nation-wide questionnaire survey to ascertain the status of familial medullary thyroid carcinoma (MTC) in Japan in 2002. Out of a total of 271 MTC cases (male to female ratio 1:1.4), multiple endocrine neoplasia (MEN) 2A accounted for 83 cases (30.6%), familial MTC (FMTC) for 14 cases (5.1%), MEN for 11 cases (4.1%), and sporadic MTC for 163 cases (60.1%). Mean age at the time of diagnosis was 35.6 in MEN2A, 34.6 in FMTC, 30.5 in MEN2B, and 47.6 in sporadic MTC. Forty-five percent of MEN2A patients had pheochromocytoma and 11% of MEN2A patients had parathyroid disorders when MTC was diagnosed. Finally, the RET oncogene test yielded the largest number of initial findings that led to diagnosis of familial MTC.

GCMB gene, a master regulator of parathyroid gland development, expression, and regulation in hyperparathyroidism.

BACKGROUND: The glial cell missing gene, GCMB , encodes a transcription factor, which is a master regulator of parathyroid development. We postulated that the GCMB gene might play a role in parathyroid tumorigenesis in hyperparathyroidism. METHODS: We used real-time quantitative reverse transcriptase polymerase chain reaction to study GCMB mRNA expression in parathyroid tissue: normal (n = 3), hyperplastic (n = 16), adenomas (n = 19), and cancers (n = 8). In primary parathyroid culture, the effect of CaCl 2 on parathyroid hormone secretion and GCMB mRNA expression was studied by using enzyme-linked immunosorbent assay and reverse transcriptase polymerase chain reaction, respectively. RESULTS: GCMB mRNA expression was lower in normal (0.4 +/- 0.1, mean +/- standard error of mean) parathyroid glands as compared to adenoma (3.5 +/- 1.7), hyperplasia (3.2 +/- 1.3 primary hyperparathyroidism [n = 11] and 7.6 +/- 4.8 secondary hyperparathyroidism [n = 5]), and cancer (3.6 +/- 1.3) ( P = .001). There was no difference in the level of GCMB mRNA expression between parathyroid adenoma, hyperplasia, and cancer. In primary culture of parathyroid adenoma (n = 9) and hyperplasia (n = 2), parathyroid hormone secretion was increased 2- to 15-fold with low calcium concentration (0.5 to 4.0 mmol/L CaCl 2 from 2 to 6 hours, P < .005). The level of GCMB mRNA expression was down-regulated with lower extracellular CaCl 2 concentration ( P < .005). CONCLUSIONS: GCMB expression is upregulated in abnormal parathyroid glands of hyperparathyroidism and decreases in response to hypocalcemia. The GCMB transcription factor might mediate the effect of calcium on parathyroid cell parathyroid hormone expression/secretion.

[Consequences of clinical genetic analysis of RET proto-oncogene]. / Konsekwencje kliniczne analizy genetycznej protoonkogenu RET.

Preliminary results of treatment of inherited medullary thyroid carcinoma, diagnosed primarily with genetic analysis of mutation of protooncogene RET are presented. Among 16 carriers of mutation identical with mutation diagnosed earlier in proband, there were 4 patients with clinically obvious medullary thyroid carcinoma and 12 asymptomatic carriers. In all patients, in whom calcitonin level was increased preoperatively, its normalization was obtained. The paper summarizes these aspects of cooperation between geneticians and physicians in which diagnostic results influence clinical decisions (indication and time of thyroid and lymph nodes surgery and it's spectrum, range of diagnostic procedures towards pheochromocytoma and parathyroid hyperplasia in relation to the found mutation).