Tumour microenvironment and pituitary tumour behaviour.

The pituitary tumour microenvironment encompasses a spectrum of non-tumoural cells, such as immune, stromal or endothelial cells, as well as enzymes and signalling peptides like cytokines, chemokines and growth factors, which surround the tumour cells and may influence pituitary tumour behaviour and tumourigenic mechanisms. Recently, there has been intensive research activity in this field describing various pituitary tumour-infiltrating immune and stromal cell subpopulations, and immune- and microenvironment-related pathways. Key changes in oncological therapeutic avenues resulted in the recognition of pituitary as a target of adverse events for patients treated with immune checkpoint regulators. However, these phenomena can be turned into therapeutic advantage in severe cases of pituitary tumours. Therefore, unravelling the pituitary tumour microenvironment will allow a better understanding of the biology and behaviour of pituitary tumours and may provide further developments in terms of diagnosis and management of patients with aggressively growing or recurrent pituitary tumours.

Novel AIP mutation in exon 6 causing acromegaly in a German family.

The most frequent genetic alteration of familial isolated growth hormone producing pituitary neuroendocrine tumors is a germline mutation of the aryl hydrocarbon receptor-interacting protein (AIP) gene. Various AIP mutations are already known; however, an AIP mutation in exon 6 (c.811_812del; p.Arg271Glyfs*16) has not been reported yet. Here, we report a German family with two identical twins who were both affected by acromegaly and carried the above-mentioned novel AIP mutation. The father was found to be an unaffected carrier, while the paternal aunt most likely suffered from acromegaly as well and died from metastatic colorectal cancer. Apart from reporting a novel AIP mutation, this study does not only highlight the different clinical and histological features of the AIP mutated growth hormone producing pituitary neuroendocrine tumors but also confirms the poor responsiveness of dopamine agonists in AIP mutated acromegaly. Furthermore, it highlights the increased mortality risk of comorbidities typically associated with acromegaly.

Fasting and postprandial liver glycogen content in patients with type 1 diabetes mellitus after successful pancreas-kidney transplantation with systemic venous insulin delivery.

BACKGROUND: In patients with type 1 diabetes mellitus (T1DM), insulin is usually replaced systemically (subcutaneously) and not via the physiological portal route. According to previous studies, the liver's capacity to store glycogen is reduced in T1DM patients, but it remains unclear whether this is due to hyperglycaemia, or whether the route of insulin supply could contribute to this phenomenon. T1DM patients after successful pancreas-kidney transplantation with systemic venous drainage (T1DM-PKT) represent a suitable human model to further investigate this question, because they are normoglycaemic, but their liver receives insulin from the pancreas transplant via the systemic route. MATERIALS AND METHODS: In nine T1DM-PKT, nine controls without diabetes (CON) and seven patients with T1DM (T1DM), liver glycogen content was measured at fasting and after two standardized meals employing (13) C-nuclear-magnetic-resonance-spectroscopy. Circulating glucose and glucoregulatory hormones were measured repeatedly throughout the study day. RESULTS: The mean and fasting concentrations of peripheral plasma glucose, insulin, glucagon and C-peptide were comparable between T1DM-PKT and CON, whereas T1DM were hyperglycaemic and hyperinsulinaemic (P < 0·05 vs T1DM-PKT and CON). Total liver glycogen content at fasting and after breakfast did not differ in the three groups. After lunch, T1DM-PKT and T1DM had a 14% and 21% lower total liver glycogen content than CON (P < 0·02). CONCLUSION: In spite of normalized glycaemic control, postprandial liver glycogen content was reduced in T1DM-PKT with systemic venous drainage. Thus, not even optimized systemic insulin substitution is able to resolve the defect in postprandial liver glycogen storage seen in T1DM patients.

MicroRNAs: Suggested role in pituitary adenoma pathogenesis.

MicroRNAs (miRNAs) are small non-coding RNA molecules that represent a major class of molecular regulators. miRNAs have been implicated in the pathogenesis of several human tumors, including pituitary adenomas. Altered expression of miRNAs has been described in pituitary adenomas, and specific miRNA signatures are related to clinical and therapeutic characteristics of the tumors. The data suggest that miRNAs influence various genes known to be associated with the pathogenesis of pituitary adenomas and in this review we summarize these currently available studies focusing on miRNAs in pituitary adenomas.

Familial isolated pituitary adenomas: an emerging clinical entity.

Familial pituitary tumors are increasingly recognized. While some of these cases are related to wellknown syndromic conditions such as multiple endocrine neoplasia type 1 (MEN1) or Carney complex, others belong to the familial isolated pituitary adenoma (FIPA) patient group. The discovery of heterozygous, loss-of-function germline mutations in the gene encoding the aryl hydrocarbon receptor interacting protein (AIP) in 2006 has subsequently enabled the identification of a mutation in this gene in 20% of FIPA families and 20% of childhood-onset simplex soma- totroph adenomas. The exact mechanism by which the lack of AIP leads to pituitary adenomas is not clear. AIP mutations cause a low penetrance autosomal dominant disease with often a distinct phenotype characterized by young-onset, aggressive, large GH, mixed GH and PRL or PRL-secreting adenomas. This review aims to summarize currently available clinical data on AIP mutation-positive and negative FIPA patients.

A genetic study of the ghrelin and growth hormone secretagogue receptor (GHSR) genes and stature.

Growth and nutrition are interrelated and influenced by multiple genetic and environmental factors. We studied whether common variants in ghrelin and ghrelin receptor (GHSR) genes could play a role in stature variation in the general population and in families ascertained for obesity. Selected tagging SNPs in the ghrelin and GHSR genes were genotyped in 263 Caucasian families recruited for childhood obesity (1,275 subjects), and in 287 families from a general population (1,072 subjects). We performed familial testing for associations in the entire population and in a sub-set of the samples selected for a case-control study. In the case-control study for height (cases were selected from the obese cohort with mean ZH = 3.17 +/- 0.15 confidence interval (CI) versus controls with mean ZH 0.14 +/- 0.09), we found an association with a 2 base-pair intronic deletion in the GHSR gene (rs10618418) (p = 0.006, odds ratio (OR) 1.86, 95% CI [1.26;2.74] under additive model), although when adjusting for BMI, the association disappeared (p = 0.06). Individuals carrying no deletion or who were heterozygous were significantly more frequent among the tall obese population (52% vs. 36% in controls, p = 0.007, OR 1.97, 95%CI [1.22;3.18]). However, the association was not maintained after correcting for multiple testing. Familial association testing of the ghrelin and GHSR genes and their interaction testing failed to show that any combination of SNPs had any significant effect. Thus, our results suggest that common variants of the ghrelin and GHSR genes are not major contributors to height variation in a French population.

The current landscape of European registries for rare endocrine conditions.

Objective To identify cross-border international registries for rare endocrine conditions that are led from Europe and to understand the extent of engagement with these registries within a network of reference centres (RCs) for rare endocrine conditions. Methods Database search of international registries and a survey of RCs in the European Reference Network for rare endocrine conditions (Endo-ERN) with an overall response rate of 82%. Results Of the 42 conditions with orphacodes currently covered within Endo-ERN, international registries exist for 32 (76%). Of 27 registries identified in the Orphanet and RD-Connect databases, Endo-ERN RCs were aware of 11 (41%). Of 21 registries identified by the RC, RD-Connect and Orphanet did not have a record of 10 (48%). Of the 29 glucose RCs, the awareness and participation rate in an international registry was highest for rare diabetes at 75 and 56% respectively. Of the 37 sex development RCs, the corresponding rates were highest for disorders of sex development at 70 and 52%. Of the 33 adrenal RCs, the rates were highest for adrenocortical tumours at 68 and 43%. Of the 43 pituitary RCs, the rates were highest for pituitary adenomas at 43 and 29%. Of the 31 genetic tumour RCs, the rates were highest for MEN1 at 26 and 9%. For the remaining conditions, awareness and participation in registries was less than 25%. Conclusion Although there is a need to develop new registries for rare endocrine conditions, there is a more immediate need to improve the awareness and participation in existing registries.

A new variation in the promoter region, the -604 C>T, and the Leu72Met polymorphism of the ghrelin gene are associated with protection to insulin resistance.

OBJECTIVE: Previous studies suggested that polymorphisms in the coding region of the preproghrelin were involved in the etiology of obesity and might modulate glucose-induced insulin secretion. We evaluated the association of a new variation, -604C>T, in the promoter region of the ghrelin gene, of Leu72Met (247C>A) and of Gln90Leu (265A>T), all haplotype-tagging single nucleotide polymorphisms (SNPs), with measures of insulin sensitivity in 1420 adult individuals. RESEARCH METHODS: The three SNPs were genotyped using ABI PRISM 7900 HT Sequence Detection System. We used multiple linear regression analysis for quantitative traits and THESIAS software for haplotype analysis. RESULTS: We observed a protective effect exerted by Met72 variant of Leu72Met SNP on insulin resistance parameters; a significant decreasing trend from Leu/Leu to Leu/Met and to Met/Met homozygous subjects in triglycerides, fasting insulin levels and HOMA-IR index (P=0.02, 0.01 and 0.003, respectively), and, consistently, an increase in ghrelin levels (P=0.003) was found. A significant decrease from CC to TC and to TT genotypes in insulin levels and HOMA-IR index was also detected (P=0.00l for both), but only in subjects homozygous for Leu72, where the protective effect of Met72 was not present. The haplotype analysis results supported the data obtained by the evaluation of each single SNP, showing the highest value of insulin levels and HOMA-IR index in the -604(c)247(c) haplotype intermediate value in -604(T)247(C) and lowest value in -604(C)247(A). CONCLUSION: Our observations suggest a protective role of the Met72 variant and of -604 T allele in modulating insulin resistance. These SNPs or an unknown functional variant in linkage disequilibrium could increase ghrelin levels and probably insulin sensitivity.

De novo HNF1 homeobox B mutation as a cause for chronic, treatment-resistant hypomagnesaemia.

29-year-old female presenting with an 8-year history of unexplained hypomagnesaemia, which was severe enough to warrant intermittent inpatient admission for intravenous magnesium. Urinary magnesium was inappropriately normal in the context of hypomagnesaemia indicating magnesium wasting. Ultrasound imaging demonstrated unilateral renal cysts and computed tomography of kidneys, ureters and bladder showed a bicornuate uterus. Referral to genetic services and subsequent testing revealed a de novo HNF1B deletion. LEARNING POINTS: HNF1B loss-of-function mutations are one of the most common monogenic causes of congenital anomalies of the kidney and urinary tract.Those with HNF1B mutations may have some of a constellation of features (renal and hepatic cysts, deranged liver function tests, maturity onset diabetes of the young type 5 (MODY5), bicornuate uterus, hyperparathyroidism, hyperuricaemic gout, but presenting features are highly heterogeneous amongst patients and no genotype/phenotype correlation exists. HNF1B mutations are inherited in an autosomal dominant pattern but up to 50% of cases are de novo.HNF1B mutations can be part of the Chr17q12 deletion syndrome, a contiguous gene deletion syndrome.Inorganic oral magnesium replacements are generally poorly tolerated with side effects of diarrhoea. Organic magnesium compounds, such as magnesium aspartate, are better absorbed oral replacement therapies.

Imprinting of the G(s)alpha gene GNAS1 in the pathogenesis of acromegaly.

Approximately 40% of growth hormone-secreting pituitary adenomas have somatic mutations in the GNAS1 gene (the so-called gsp oncogene). These mutations at codon 201 or codon 227 constitutively activate the alpha subunit of the adenylate cyclase-stimulating G protein G(s). GNAS1 is subject to a complex pattern of genomic imprinting, its various promoters directing the production of maternally, paternally, and biallelically derived gene products. Transcripts encoding G(s)alpha are biallelically derived in most human tissues. Despite this, we show here that in 21 out of 22 gsp-positive somatotroph adenomas, the mutation had occurred on the maternal allele. To investigate the reason for this allelic bias, we also analyzed GNAS1 imprinting in the normal adult pituitary and found that G(s)alpha is monoallelically expressed from the maternal allele in this tissue. We further show that this monoallelic expression of G(s)alpha is frequently relaxed in somatotroph tumors, both in those that have gsp mutations and in those that do not. These findings imply a possible role for loss of G(s)alpha imprinting during pituitary somatotroph tumorigenesis and also suggest that G(s)alpha imprinting is regulated separately from that of the other GNAS1 products, NESP55 and XLalphas, imprinting of which is retained in these tumors.

Sporadic pituitary adenomas: the role of germline mutations and recommendations for genetic screening.

INTRODUCTION: Although most pituitary adenomas occur sporadically, these common tumors can present in a familial setting in approximately 5% of cases. Germline mutations in several genes with autosomal dominant (AIP, MEN1, CDKN1B, PRKAR1A, SDHx) or X-linked dominant (GPR101) inheritance are causative of familial pituitary adenomas. Due to variable disease penetrance and occurrence of de novo mutations, some patients harboring germline mutations have no family history of pituitary adenomas (simplex cases). Areas covered: We summarize the recent findings on the role of germline mutations associated with familial pituitary adenomas in patients with sporadic clinical presentation. Expert commentary: Up to 12% of patients with young onset pituitary adenomas (age at diagnosis/onset ≤30 years) and up to 25% of simplex patients with gigantism carry mutations in the AIP gene, while most cases of X-linked acrogigantism (XLAG) due to GPR101 duplication are simplex female patients with very early disease onset (<5 years). With regard to the syndromes of multiple endocrine neoplasia (MEN), MEN1 mutations can be identified in a significant proportion of patients with childhood onset prolactinomas. Somatotroph and lactotroph adenomas are the most common pituitary adenomas associated with germline predisposing mutations. Genetic screening should be considered in patients with young onset pituitary adenomas.

From pituitary adenoma to pituitary neuroendocrine tumor (PitNET): an International Pituitary Pathology Club proposal.

The classification of neoplasms of adenohypophysial cells is misleading because of the simplistic distinction between adenoma and carcinoma, based solely on metastatic spread and the poor reproducibility and predictive value of the definition of atypical adenomas based on the detection of mitoses or expression of Ki-67 or p53. In addition, the current classification of neoplasms of the anterior pituitary does not accurately reflect the clinical spectrum of behavior. Invasion and regrowth of proliferative lesions and persistence of hormone hypersecretion cause significant morbidity and mortality. We propose a new terminology, pituitary neuroendocrine tumor (PitNET), which is consistent with that used for other neuroendocrine neoplasms and which recognizes the highly variable impact of these tumors on patients.

Cancerous leptomeningitis and familial congenital hypopituitarism.

People are at higher risk of cancer as they get older or have a strong family history of cancer. The potential influence of environmental and behavioral factors remains poorly understood. Earlier population and case control studies reported that upper quartile of circulating IGF-I is associated with a higher risk of developing cancer suggesting possible involvement of the growth hormone (GH)/IGF system in initiation or progression of cancer. Since GH therapy increases IGF-1 levels, there have been concerns that GH therapy in hypopituitarism might increase the risk of cancer. We report a 42-year-old female patient who presented with subacute onset of symptoms of meningitis and with the absence of fever which resulted in death 70 days after the onset of symptoms. The patient together with her younger brother was diagnosed at the age of 5 years with familial congenital hypopituitarism, due to homozygous mutation c.150delA in PROP1 gene. Due to evolving hypopituitarism, she was replaced with thyroxine (from age 5), hydrocortisone (from age 13), GH (from age 13 until 17), and sex steroids in adolescence and adulthood. Her consanguineous family has a prominent history of malignant diseases. Six close relatives had malignant disease including her late maternal aunt with breast cancer. BRCA 1 and BRCA 2 mutational analysis in the patient's mother was negative. Histology after autopsy disclosed advanced ovarian cancer with multiple metastases to the brain, leptomeninges, lungs, heart, and adrenals. Low circulating IGF-1 did not seem to protect this patient from cancer initiation and progression in the context of strong family history of malignancies.

Enhanced protein kinase B/Akt signalling in pituitary tumours.

Pituitary tumours have previously been shown to harbour several abnormalities that cause deregulation of the cell cycle, particularly down-regulation of expression of the cyclin-dependent kinase inhibitor p27. However, it has been unclear whether these are the primary initiating events, or are secondary to other more proximate alterations in signalling pathways. In other cellular systems the Akt signalling pathway has been associated with downstream modulation of cell-cycle control. The aim of the present study was to test the hypothesis that Akt signalling is enhanced in pituitary tumours, and to see if changes in Akt expression are related to previous findings on low expression levels of the nuclear cell-cycle inhibitor p27 in pituitary tumours. We examined normal and adenomatous human pituitary tissue for mRNA and protein expression of Akt1, Akt2 and p27, and the activation of Akt, as well the phosphatase involved in the inactivation of Akt, phosphatase and tensin homologue deleted on chromosome 10 (PTEN). In pituitary adenomas Akt1 and Akt2 mRNA were found to be over-expressed compared with normal pituitary, while PTEN transcripts showed similar levels between the two tissue types. Immunohistochemical expression of phospho-Akt was found to be higher in the tumours than normal pituitaries, while the protein expression of nuclear p27 and PTEN was lower in the adenomas. However, the expression of p27 and Akt were not directly correlated. PTEN sequencing revealed no mutation in the coding region of the gene in pituitary adenomas, and thus we did not locate a cause for the increased phosphorylation of Akt. In summary, we have shown over-expression and activation of the Akt pathway in pituitary tumours, and we speculate that cell-cycle changes observed in such tumours are secondary to these more proximate alterations. Since Akt is a major downstream signalling molecule of growth factor-liganded tyrosine kinase receptors, our data are most compatible with an abnormality at this level as the primary driver of pituitary tumorigenesis.

Growth hormone-releasing peptide and its analogues Novel stimuli to growth hormone release.

GHRPs are oligopeptides with GH-releasing effects in humans when given by either parenteral or oral routes; in addition, nonpeptide pharmacologic analogues have recently been synthesized. Although the exact mechanism of action of these agents has not been fully established, there is probably a dual site of action on both the pituitary and the hypothalamus, possibly involving regulatory factors in addition to GHRH and somatostatin. GHRPs and their analogues may have a potential role in the treatment of short stature in children or in other situations of GH deficiency, such as adult GH deficiency, obesity, catabolic states, and even normal old age.

Excessive growth hormone expression in male GH transgenic mice adversely alters bone architecture and mechanical strength.

Patients with acromegaly have a higher prevalence of vertebral fractures despite normal bone mineral density (BMD), suggesting that GH overexpression has adverse effects on skeletal architecture and strength. We used giant bovine GH (bGH) transgenic mice to analyze the effects of high serum GH levels on BMD, architecture, and mechanical strength. Five-month-old hemizygous male bGH mice were compared with age- and sex-matched nontransgenic littermates controls (NT; n=16/group). Bone architecture and BMD were analyzed in tibia and lumbar vertebrae using microcomputed tomography. Femora were tested to failure using three-point bending and bone cellular activity determined by bone histomorphometry. bGH transgenic mice displayed significant increases in body weight and bone lengths. bGH tibia showed decreases in trabecular bone volume fraction, thickness, and number compared with NT ones, whereas trabecular pattern factor and structure model index were significantly increased, indicating deterioration in bone structure. Although cortical tissue perimeter was increased in transgenic mice, cortical thickness was reduced. bGH mice showed similar trabecular BMD but reduced trabecular thickness in lumbar vertebra relative to controls. Cortical BMD and thickness were significantly reduced in bGH lumbar vertebra. Mechanical testing of femora confirmed that bGH femora have decreased intrinsic mechanical properties compared with NT ones. Bone turnover is increased in favor of bone resorption in bGH tibia and vertebra compared with controls, and serum PTH levels is also enhanced in bGH mice. These data collectively suggest that high serum GH levels negatively affect bone architecture and quality at multiple skeletal sites.

Ghrelin.

BACKGROUND: The gastrointestinal peptide hormone ghrelin was discovered in 1999 as the endogenous ligand of the growth hormone secretagogue receptor. Increasing evidence supports more complicated and nuanced roles for the hormone, which go beyond the regulation of systemic energy metabolism. SCOPE OF REVIEW: In this review, we discuss the diverse biological functions of ghrelin, the regulation of its secretion, and address questions that still remain 15 years after its discovery. MAJOR CONCLUSIONS: In recent years, ghrelin has been found to have a plethora of central and peripheral actions in distinct areas including learning and memory, gut motility and gastric acid secretion, sleep/wake rhythm, reward seeking behavior, taste sensation and glucose metabolism.

The Krüppel-like transcription factor 6 gene in sporadic pituitary tumours.

The oncogenes and/or tumour suppressor genes which may be involved in the transformation process for the vast majority of pituitary tumours remain unknown. There is substantial evidence for derangement of cell cycle control in such tumours, but cell cycle protein mutations identified in other human malignancies are restricted to only a very small subset of sporadic pituitary neoplasms. Krüppel-like factors are DNA-binding transcriptional regulators with diverse effects including the upregulation of the cell cycle protein p21(WAF1/CIP1). It has been reported that the Krüppel-like transcription factor 6 (KLF6) gene is mutated in a proportion (15-55%) of human prostate cancers, and more recent data are emerging regarding mutated KLF6 in nasopharyngeal carcinomas, astrocytoid gliomas and colorectal cancer. We therefore speculated that other tumours such as pituitary adenomas might also harbour such mutations that may be involved in the control of cell proliferation in the pituitary. The aim of the current study was thus to analyse the KLF6 gene for mutations in sporadic pituitary tumours. We analysed 60 pituitary adenomas (15 GH-, four ACTH-, two PRL-secreting and 39 non-functioning) with direct sequence analysis of exons 2 and 3 of the KLF6 gene, the region where most of the previously described mutations are located. Three non-functioning pituitary adenomas of the 60 pituitary tumours (5%) had two identical sequence changes in exon 2 (missense mutation Val165Met, 523G-->A and a silent substitution in Ser77Ser codon 261C-->T). Analysis of genomic DNA extracted from peripheral lymphocytes in one patient confirmed these changes to be present in the germline and they therefore probably represent polymorphisms, although we cannot exclude the possibility that these are predisposing germline mutations. We conclude that mutations of the KLF6 gene are unlikely to play an important role in sporadic pituitary tumorigenesis.