PROP1 Mutations Research Articles

Screening a Large Pediatric Cohort with GH Deficiency for Mutations in Genes Regulating Pituitary Development and GH Secretion: Frequencies, Phenotypes and Growth Outcomes

Background: Pituitary development and GH secretion are orchestrated by multiple genes including GH1, GHRHR, GLI2, HESX1, LHX3, LHX4, PROP1, POU1F1, and SOX3. We aimed to assess their mutation frequency and clinical relevance in children with severe GH deficiency (GHD). Methods:The Genetics and Neuroendocrinology of Short Stature International Study (GeNeSIS) was a prospective, open-label, observational research program for pediatric patients receiving GH treatment, conducted in 30 countries between 1999 and 2015. The study included a sub-study to investigate mutations in the genes listed above. PCR products from genomic blood cell DNA were analyzed by Sanger sequencing. DNA variants were classified as pathogenic according to the recommendations of the American College of Medical Genetics and Genomics. Demographic, auxologic, and endocrine data at baseline and during GH treatment were documented and related to the genotyping results. Findings: The analysis comprised 917 patients. In 92 patients (10%) 33 mutations were found, 16 previously described and 17 novel (52%). Mutation carriers were significantly younger, shorter, and more slowly growing than non-carriers. In general, their peak values in GH stimulation tests were very low; however, in 15/77 (20%) patients with GH1, PROP1, and SOX3 mutations they were only moderately diminished (3- 6µg/L). Two patients with a GH1 mutation developed TSH deficiency and one ADH deficiency. Using logistic multi-regression analysis, significant indicators of a mutation were combined pituitary hormone deficiency, greater patient-parent height difference (SDS), low GH peak, and young age. Final height SDS gain in mutation carriers (mean±SD 3.4±1.4) was greater than in non-carriers (2.0±1.4; P<0.001) and in patients with nonGHD short stature. Interpretation: DNA testing for mutations in children with severe GHD shows a positive finding in approximately 10%. Phenotypes of mutation carriers can be variable. The benefit for clinical practice justifies DNA testing as an important component in the diagnostic work-up of patients with severe GHD. Clinical Trial Number: (GeNeSIS; Clinical Trial Registry Number: NCT01088412) Funding Statement: Eli Lilly and Company, Indianapolis, IN, USA. Declaration of Interests: GeNeSIS was sponsored by Eli Lilly and Company (Lilly, Indianapolis, IN, USA). The sponsor funded all aspects of study design, data collection, genetic analyses and statistical analyses, but did not impose any impediment, directly or indirectly, on the publication of the study results. CJC and CJ are employees and stockholders of Lilly, while WFB, AGZ, and CAQ are former employees and stockholders of Lilly, GBC is a former employee of Lilly. WFB also reports he is a consultant for Ammonett Pharma, Lilly Germany and Merck KGaA Darmstadt. CLD, JL, RGR, JSP and RWP were members of the GeNeSIS International Scientific Advisory Board and received consulting and speaker fees from Lilly. The laboratories of SA and RWP received fees and grants from Lilly for setting up and conducting the genetic analyses. JK, HMP, ML. M-L S, and M-PL have no conflicting interests to report. Ethics Approval Statement: The study was conducted in accordance with the Declaration of Helsinki and approved by ethics review boards of the participating institutions. Written informed consent for data collection, data processing, DNA testing and publication was obtained from the parents or legal guardians, in accordance with national regulatory requirements.

SCREENING OF PROP-1, LHX2 AND POU1F1 MUTATIONS IN PATIENTS WITH ECTOPIC POSTERIOR PITUITARY GLAND.

Ectopic posterior pituitary gland (EPP) is usually characterized by an abnormal pituitary stalk and hypoplasia of the anterior hypophysis. The genetic mechanisms involved in the development of EPP remain uncertain. The aim of this study is to determine whether mutations in the three genes, PROP-1, LHX2, and POU1F1, are associated with the risk for and the characteristics of EPP. In the Endocrinology Outpatient Clinic of "Dr. Behcet Uz" Children's Hospital, 27 patients with EPP were submitted to sequencing analyses of the PROP-1, LHX2, and POU1F1 genes. Growth hormone, thyrotropin, corticotropin, gonadotropin, and vasopressin deficiency were observed in 22 (81.5%), 23 (85.2%), 17 (63%), 14 (51.9%), and two (7.4%) patients. Thirteen patients (48.1%) presented with hyperprolactinemia. Fourteen patients (51%) had a history of birth dystocia, and 12 cases (42.1%) had a history of breech presentation. Central nervous system abnormalities included five cases with corpus callosum agenesis, one case with schizencephaly, and one case with Chiari type 1 malformation. We identified a homozygous p.S109* mutation in exon 2 in one male patient with EPP and two different PROP1 gene polymorphisms (A142T or c.109+3 G>A polymorphism) in thirteen patients. Our results suggest that PROP1 gene abnormalities might explain the genetic mechanisms involved in the development of EPP.

Znaczenie mutacji genu PROP 1 w diagnostyce i leczeniu niedoczynności przysadki – prezentacja przypadku

Znaczenie mutacji genu PROP 1 w diagnostyce i leczeniu niedoczynności przysadki – prezentacja przypadku

Molecular analysis of brazilian patients with combined pituitary hormone deficiency and orthotopic posterior pituitary lobe reveals eight different PROP1 alterations with three novel mutations.

Mutations in PROP1, HESX1 and LHX3 are associated with combined pituitary hormone deficiency (CPHD) and orthotopic posterior pituitary lobe (OPP). To identify mutations in PROP1, HESX1 and LHX3 in a large cohort of patients with CPHD and OPP (35 Brazilian, two Argentinian). We studied 23 index patients with CPHD and OPP (six familial and 17 sporadic) as well as 14 relatives. PROP1 was sequenced by the Sanger method in all except one sporadic case studied using a candidate gene panel. Multiplex ligation-dependent probe amplification (MLPA) was applied to one familial case in whom PROP1 failed to amplify by PCR. In the 13 patients without PROP1 mutations, HESX1 and LHX3 were sequenced by the Sanger method. We identified PROP1 mutations in 10 index cases. Three mutations were novel: one affecting the initiation codon (c.1A>G) and two affecting splicing sites, c.109+1G>A and c.342+1G>C. The known mutations, c.150delA (p.Arg53Aspfs*112), c.218G>A (p.Arg73His), c.263T>C (p.Phe88Ser) and c.301_302delAG (p.Leu102Cysfs*8), were also detected. MLPA confirmed complete PROP1 deletion in one family. We did not identify HESX1 and LHX3 mutations by Sanger. PROP1 mutations are a prevalent cause of congenital CPHD with OPP, and therefore, PROP1 sequencing must be the first step of molecular investigation in patients with CPHD and OPP, especially in populations with a high frequency of PROP1 mutations. In the absence of mutations, massively parallel sequencing is a promising approach. The high prevalence and diversity of PROP1 mutations is associated with the ethnic background of this cohort.

Lessons from monogenic causes of growth hormone deficiency

Through the multicentric international GENHYPOPIT network, 10 transcription factor genes involved in pituitary development have been screened in more than 1200 patients with constitutional hypopituitarism over the past two decades. The present report summarizes the main lessons learned from this phenotype-based genetic screening: (1) genetically determined hypopituitarism does not necessarily present during childhood; (2) constitutional hypopituitarism may be characterized by a pure endocrine phenotype or by various combinations of endocrine deficits and visceral malformations; (3) syndromic hypopituitarism may also be observed in patients with POU1F1 or PROP1 mutations; (4) in cases of idiopathic hypopituitarism, extensive genetic screening identifies gene alterations in a minority of patients; (5) functional studies are imperfect in determining the involvement of an allelic variant in a specific pituitary phenotype.

Identification of Novel PROP1 and POU1F1 Mutations in Patients with Combined Pituitary Hormone Deficiency.

Growth hormone deficiency (GHD) results from variations affecting the production and release of growth hormone (GH) and is of 2 types: isolated growth hormone deficiency (IGHD) and combined pituitary hormone deficiency (CPHD). IGHD results from mutations in GH1 and GHRHR while CPHD is associated with defects in transcription factor genes PROP1, POU1F1, and HESX1. The present study reports on screening of POU1F1, PROP1, and HESX1 in CPHD patients and the novel variations identified. Fifty-one CPHD patients from 49 unrelated families clinically diagnosed on the basis of biochemical and imaging investigations along with 100 controls were enrolled. Detailed family history was noted from all participants and 5 ml blood samples drawn were processed for DNA isolation followed by direct sequencing of POU1F1, PROP1, and HESX1genes. Of the 51 patients, 8 were females and 43 were males. Mean height standard deviation score (SDS) and weight SDS were -5.50 and -2.76, respectively. Thirty-six of the 51 patients underwent MRI of which 9 (25%) had normal pituitary structure and morphology while 27 (75%) showed abnormalities. Molecular analysis revealed 10 (20%) patients to have POU1F1 and PROP1 mutations/variations of which 5 were novel and 2 previously reported. No mutations were identified in HESX1. The novel variations identified were absent in the 100 healthy individuals screened and the control database Exome Aggregation Consortium (ExAC). Reported POU1F1 and PROP1 mutation hotspots were absent in our patients. Instead, novel POU1F1 changes were identified suggesting existence of a distinct mutation spectrum in our population.

Long-Term Outcomes, Genetics, and Pituitary Morphology in Patients with Isolated Growth Hormone Deficiency and Multiple Pituitary Hormone Deficiencies: A Single-Centre Experience of Four Decades of Growth Hormone Replacement

Background: Growth hormone (GH) has been used to treat children with GH deficiency (GHD) since 1966. Aims: Using a combined retrospective and cross-sectional approach, we explored the long-term outcomes of patients with GHD, analysed factors influencing therapeutic response, determined persistence into adulthood, investigated pituitary morphology, and screened for mutations in causative genes. Methods: The files of 96 GH-deficient children were reviewed. In a subset of 50 patients, re-assessment in adulthood was performed, including GHRH-arginine testing, pituitary magnetic resonance imaging (MRI), and mutational screening for the growth hormone-1 gene (GH1) and the GHRH receptor gene (GHRHR) in isolated GHD (IGHD), and HESX1, PROP1, POU1F1, LHX3, LHX4, and GLI2 in multiple pituitary hormone deficiency (MPHD) patients. Results: GH was started at a height SDS of -3.2 ± 1.4 in IGHD patients and of -4.1 ± 2.1 in MPHD patients. Relative height gain was 0.3 SDS/year, absolute gain 1.6 SDS, and 1.2/2.6 SDS in IGHD/MPHD, respectively. Mid-parental target height was reached in 77%. Initial height SDS, bone age retardation and duration of GH replacement were correlated with height SDS gain. GHD persisted into adulthood in 19 and 89% of subjects with IGHD and MPHD, respectively. In 1/42 IGHD patients a GH1 mutation was detected; PROP1 mutations were found in 3/7 MPHD subjects. Anterior pituitary hypoplasia, combined with posterior pituitary ectopy and pituitary stalk invisibility on MRI, was an exclusive finding in MPHD patients. Conclusions: GH replacement successfully corrects the growth deficit in children with GHD. While the genetic aetiology remains undefined in most cases of IGHD, PROP1 mutations constitute a major cause for MPHD. Persistence of GHD into adulthood is related to abnormal pituitary morphology.

PROP1 triggers epithelial-mesenchymal transition-like process in pituitary stem cells.

Mutations in PROP1 are the most common cause of hypopituitarism in humans; therefore, unraveling its mechanism of action is highly relevant from a therapeutic perspective. Our current understanding of the role of PROP1 in the pituitary gland is limited to the repression and activation of the pituitary transcription factor genes Hesx1 and Pou1f1, respectively. To elucidate the comprehensive PROP1-dependent gene regulatory network, we conducted genome-wide analysis of PROP1 DNA binding and effects on gene expression in mutant mice, mouse isolated stem cells and engineered mouse cell lines. We determined that PROP1 is essential for stimulating stem cells to undergo an epithelial to mesenchymal transition-like process necessary for cell migration and differentiation. Genomic profiling reveals that PROP1 binds to genes expressed in epithelial cells like Claudin 23, and to EMT inducer genes like Zeb2, Notch2 and Gli2. Zeb2 activation appears to be a key step in the EMT process. Our findings identify PROP1 as a central transcriptional component of pituitary stem cell differentiation.

All Hormone-Producing Cell Types of the Pituitary Intermediate and Anterior Lobes Derive From Prop1-Expressing Progenitors.

Mutations in PROP1, the most common known cause of combined pituitary hormone deficiency in humans, can result in the progressive loss of all hormones of the pituitary anterior lobe. In mice, Prop1 mutations result in the failure to initiate transcription of Pou1f1 (also known as Pit1) and lack somatotropins, lactotropins, and thyrotropins. The basis for this species difference is unknown. We hypothesized that Prop1 is expressed in a progenitor cell that can develop into all anterior lobe cell types, and not just the somatotropes, thyrotropes, and lactotropes, which are collectively known as the PIT1 lineage. To test this idea, we produced a transgenic Prop1-cre mouse line and conducted lineage-tracing experiments of Prop1-expressing cells. The results reveal that all hormone-secreting cell types of both the anterior and intermediate lobes are descended from Prop1-expressing progenitors. The Prop1-cre mice also provide a valuable genetic reagent with a unique spatial and temporal expression for generating tissue-specific gene rearrangements early in pituitary gland development. We also determined that the minimal essential sequences for reliable Prop1 expression lie within 10 kilobases of the mouse gene and demonstrated that human PROP1 can substitute functionally for mouse Prop1. These studies enhance our understanding of the pathophysiology of disease in patients with PROP1 mutations.

Two coexisting heterozygous frameshift mutations in PROP1 are responsible for a different phenotype of combined pituitary hormone deficiency.

The role of genetic background in childhood-onset combined pituitary hormone deficiency (CPHD) has been extensively studied. The major contributors are the PROP1, POU1F1, LHX3, LHX4 and HESX1 genes coding transcription factors implicated in pituitary organogenesis. The clinical consequences of mutations encompass impaired synthesis of a growth hormone (GH) and one or more concurrent pituitary hormones (i.e. LH, FSH, TSH, PRL). Manifestation of the disorder may vary due to various mutation impacts on the final gene products or an influence of environmental factors during pituitary organogenesis. We describe the clinical and molecular characteristics of two brothers aged 47 and 39 years presenting an uncommon manifestation of congenital hypopituitarism. Sequencing of the PROP1, POU1F1, LHX3, LHX4 and HESX1 genes was performed to confirm the genetic origin of the disorder. A compound heterozygosity in the PROP1 gene has been identified for both probands. The first change represents a mutational hot spot (c.150delA, p.R53fsX164), whereas the second is a novel alteration (p.R112X) that leads to protein disruption. Based on precise genetic diagnosis, an in silico prediction of a p.R112X mutation on protein architecture was performed. The resulting clinical phenotype was surprisingly distinct compared to most patients with genetic alterations in PROP1 reported in the current literature. This may be caused by a residual activity of a newly identified p.R112X protein that preserves over 70 % of the homeodomain structure. This examination may confirm a key role of a DNA-binding homeodomain in maintaining PROP1 functionality and suggests a conceivable explanation of an unusual phenotype.

Novel Mutations in HESX1 and PROP1 Genes in Combined Pituitary Hormone Deficiency

Background/Aims: The HESX1 gene is essential in forebrain development and pituitary organogenesis, and its mutations are the most commonly identified genetic cause of septo-optic dysplasia (SOD). The PROP1 gene is involved in anterior pituitary cell lineage specification and is commonly implicated in non-syndromic combined pituitary hormone deficiency (CPHD). We aimed to assess the involvement of HESX1 and PROP1 mutations in a cohort of patients with SOD and CPHD. Methods: Six patients with sporadic SOD and 16 patients with CPHD from 14 pedigrees were screened for mutations in HESX1 and PROP1 genes by exon sequencing. Half of the CPHD patients had variable associated clinical characteristics, such as hearing loss, orofacial cleft, kidney disorder or developmental delay. Novel variants were evaluated in silico and verified in SNP databases. Results: A novel heterozygous p.Glu102Gly mutation in the HESX1 gene and a novel homozygous p.Arg121Thr mutation in the PROP1 gene were detected in 2 pedigrees with CPHD. A small previously reported deletion in PROP1 c.301_302delAG was detected in a separate patient with CPHD, in heterozygous state. No mutations were identified in patients with SOD. Conclusions: Our results expand the spectrum of mutations implicated in CPHD. The frequency of 15% of the PROP1 mutations in CPHD was low, likely due to the clinical heterogeneity of the cohort.

Short communication: Validation of 4 candidate causative trait variants in 2 cattle breeds using targeted sequence imputation

Most association studies for pinpointing trait-associated variants are performed within breed. The availability of sequence data from key ancestors of several cattle breeds now enables immediate assessment of the frequency of trait-associated variants in populations different from the mapping population and their imputation into large validation populations. The objective of this study was to validate the effects of 4 putatively causative variants on milk production traits, male fertility, and stature in German Fleckvieh and Holstein-Friesian animals using targeted sequence imputation. We used whole-genome sequence data of 456 animals to impute 4 missense mutations in DGAT1, GHR, PRLR, and PROP1 into 10,363 Fleckvieh and 8,812 Holstein animals. The accuracy of the imputed genotypes exceeded 95% for all variants. Association testing with imputed variants revealed consistent antagonistic effects of the DGAT1 p.A232K and GHR p.F279Y variants on milk yield and protein and fat contents, respectively, in both breeds. The allele frequency of both polymorphisms has changed considerably in the past 20 yr, indicating that they were targets of recent selection for milk production traits. The PRLR p.S18N variant was associated with yield traits in Fleckvieh but not in Holstein, suggesting that it may be in linkage disequilibrium with a mutation affecting yield traits rather than being causal. The reported effects of the PROP1 p.H173R variant on milk production, male fertility, and stature could not be confirmed. Our results demonstrate that population-wide imputation of candidate causal variants from sequence data is feasible, enabling their rapid validation in large independent populations.

Molecular screening of a large cohort of Moroccan patients with congenital hypopituitarism.

Congenital hypopituitarism is a rare disease which, for most patients, has no identified molecular cause. We aimed to document the molecular basis of growth retardation in a Moroccan cohort. 80 index cases [54 with isolated growth hormone deficiency (IGHD), 26 with combined pituitary hormone deficiency (CPHD)] were screened for molecular defects in GH1 (including LCR-GH1), GHRHR, GHSR, GHRH, PROP1, POU1F1, HESX1, LHX3, LHX4 and SOX3. Five different deleterious mutations were identified in 14 patients from eight families. In the IGHD group, three genes were found to be involved: GH1, GHRHR and GHSR. In the CPHD group, PROP1 was the only mutated gene. In addition, two heterozygous variations whose deleterious effect remains to be demonstrated were identified (in GH1 and LHX4), and two polymorphisms (missense variations) were detected (in LHX3 and in GHSR). The prevalence of mutations in this Moroccan GHD cohort was 10% (8/80), 11·1% (6/54) in the IGHD group and 7·7% (2/26) in the CPHD group. This is the first molecular screening of congenital GHD in a Moroccan population and, like other studies, mutations were preferentially identified in familial cases (75%); mutations in genes such as POU1F1, HESX1, SOX3, LHX3 and LHX4 are extremely rare. The p.R73C PROP1 mutation was the most frequent mutation in CPHD; this should be the first one to screen in this population. Our results should contribute to a better diagnosis and management of this heterogeneous disease condition.

Molecular analysis of PROP1, POU1F1, LHX3, and HESX1 in Turkish patients with combined pituitary hormone deficiency: a multicenter study.

To investigate the specific mutations in PROP1, POU1F1, LHX3, and HESX1 genes in patients with combined pituitary hormone deficiency (CPHD) in Turkey. Seventy-six patients with CPHD were included in this study. Based on clinical, hormonal, and neuro-radiological data, relevant transcription factor genes were evaluated by Sanger sequencing and multiplex ligation-dependent probe amplification. Total frequency of mutations was 30.9 % in patients with CPHD. Frequency was significantly higher in familial patients (p = 0.001). Three different types of mutations in PROP1 gene (complete gene deletion, c.301-302delAG, a novel mutation; IVS1+2T>G) were found in 12 unrelated patients (21.8 %). Mutations in PROP1 gene were markedly higher in familial than in sporadic cases (58.8 vs. 5.3 %, p < 0.001). Homozygous complete gene deletion was the most common mutation in PROP1 gene (8/12) and was identified in six familial patients. Four different homozygous mutations [p.Q4X, novel mutations; exons 1-2 deletion, p.V153F, p.I244S] were detected in POU1F1 gene. Central precocious puberty was firstly observed in a sporadic-male patient with homozygous POU1F1 (p.I244S) mutation. A homozygous mutation in HESX1 gene (p.R160H) was detected in one patient. This study is the first to investigate specific mutations in CPHD patients in Turkey. Complete deletion in PROP1 gene was the most common mutation encountered in patients with CPHD. We believe that the results of this study will contribute to the establishment of genetic screening strategies in Turkey, as well as to the studies on phenotype-genotype correlations and early diagnosis of CPHD patients.

High Prevalence ofPROP1Defects in Lithuania: Phenotypic Findings in an Ethnically Homogenous Cohort of Patients With Multiple Pituitary Hormone Deficiency

PROP1 gene mutations cause multiple pituitary hormone deficiency (MPHD). We sought to expand experience with PROP1 mutation carriers by studying a large cohort of Lithuanian patients. Sixty-seven MPHD patients were tested for PROP1 defects. Perinatal and postnatal data were obtained from medical records. Hormonal investigations, pituitary imaging, and GH therapy were provided in a single center in Kaunas, Lithuania. A biallelic PROP1 gene mutation was found in 47 subjects (70.1%), of which 46 were homozygous for 296delGA. Positive finding rate among MPHD and population prevalence of PROP1 defects in Lithuania (15.8 per million) were the highest reported to date. Patients' birth lengths/weights were normal. Testicular retention was noted in 31% of boys. Median height SD scores declined over years 1-5: -1.56, -2.34, -3.43, -3.52, and -3.70. Mid-parental height predicted severity of growth retardation at diagnosis (r2=0.30; P=.0001). Deficiencies of GH, TSH, ACTH, and FSH/LH were diagnosed in 44/44, 44/44, 19/44, and 22/44 subjects at median age of 5.5, 5.6, 13.1, and 15.0 years, respectively. Pituitary height ranged from 16.6 mm (+20.2 SD) to 1.4 mm (-15.5 SD) and declined with age (r2=0.27, P=.001). GH replacement (dose 0.027 mg/kg/d) led to height velocities 12.2; 9.1; 6.9; 6.8; 6.7; 5.6; and 5.7 cm/y (medians) at years 1-7 and final height SD scores (17 patients) -0.98±1.77 (-1.04±1.41 below target height; P=.008 vs 0). High prevalence of PROP1 defects in Lithuania is due to 296delGA mutation, suggesting a founder effect.

Suprasellar mass mimicking a hypothalamic glioma in a patient with a complete PROP1 deletion

Mutations in PROP1 are the most frequent defect detected in patients with combined pituitary hormone deficiency (MIM #262600), characterized by a clinical phenotype of proportionate growth deficit due to impaired production of growth hormone in combination with deficiency of one or more of the additional anterior pituitary hormones. Approximately one third of patients with PROP1 inactivating mutations present with abnormal development of the anterior lobe of the pituitary gland as revealed by MRI. We report on the clinical and molecular characterization of the fourth complete PROP1 deletion in a girl with proportional short stature, combined pituitary hormone deficiency and a suprasellar mass mimicking a hypothalamic glioma. The proband, born to consanguineous parents, presented with proportional growth failure (height 108.8 cm, −3.48 SDS), combined pituitary hormone deficiency (GH, TSH, PRL and gonadotropins) and a suprasellar mass with optic chiasm invasion, compatible with a diagnosis of chiasmatic hypothalamic glioma, as revealed by MRI. PROP1 mutation screening by PCR and MLPA detected a homozygous deletion of the entire PROP1. The deletion was delimited to at least 7.7 kb upstream of PROP1 and more finely to ∼541–74 bp downstream from PROP1 by aCGH and PCR mapping.We describe the fourth case with a complete PROP1 deletion in homozygosis. The apparent location of the respective 5′ (within a highly repetitive region, rich in Alu sequences) and 3′ (within an Alu sequence) breakpoints, suggests that the deletion may have arisen through homologous recombination. The differentiation between PROP1 mutation associated pituitary enlargements from craniopharyngioma, pituitary adenoma, dys-germinoma, or Rathke's pouch cyst, is critical for the correct patient management. It is important to recognize that PROP1 mutations can present associated with evolving pituitary masses and/or other MRI alterations of the pituitary during early childhood and that surgery is not indicated in these patients. Therefore, in the presence of combined pituitary hormone deficiency and a pituitary or hypothalamic mass, PROP1 analysis should be considered before referring the patient to a neurosurgeon.

A PROP1-binding factor, AES cloned by yeast two-hybrid assay represses PROP1-induced Pit-1 gene expression

PROP1 mutation causes combined pituitary hormone deficiency (CPHD). Several mutations are located in a transactivation domain (TAD) of Prop1, and the loss of TAD binding to cofactors is likely the cause of CPHD. PROP1 cofactors have not yet been identified. In the present study, we aimed to identify the PROP1-interacting proteins from the human brain cDNA library. Using a yeast two-hybrid assay, we cloned nine candidate proteins that may bind to PROP1. Of those nine candidates, amino-terminal enhancer of split (AES) was the most abundant, and we analyzed the AES function. AES dose-dependently decreased the PROP1-induced Pit-1 reporter gene expression. An immunoprecipitation assay revealed the relationship between AES and PROP1. In a mammalian two-hybrid assay, a leucine zipper-like motif of the AES Q domain was identified as a region that interacted with TAD. These results indicated that AES was a corepressor of PROP1.

Single‐nucleotide variants in two Hedgehog genes, SHH and HHIP, as genetic cause of combined pituitary hormone deficiency

Combined pituitary hormone deficiency (CPHD) is characterized by deficiencies of two or more anterior pituitary hormones. Its genetic cause is unknown in the majority of cases. The Hedgehog (Hh) signalling pathway has been implicated in disorders associated with pituitary development. Mutations in Sonic Hedgehog (SHH) have been described in patients with holoprosencephaly (with or without pituitary involvement). Hedgehog interacting protein (HHIP) has been associated with variations in adult height in genome wide association studies. We investigated whether mutations in these two genes of the Hh pathway, SHH and HHIP, could result in 'idiopathic' CPHD. We directly sequenced the coding regions and exon - intron boundaries of SHH and HHIP in 93 CPHD patients of the Dutch HYPOPIT study in whom mutations in the classical CPHD genes PROP1, POU1F1, HESX1, LHX3 and LHX4 had been ruled out. We compared the expression of Hh genes in Hep3B transfected cells between wild-type proteins and mutants. We identified three single-nucleotide variants (p.Ala226Thr, c.1078C>T and c.*8G>T) in SHH. The function of the latter was severely affected in our in vitro assay. In HHIP, we detected a new activating variant c.-1G>C, which increases HHIP's inhibiting function on the Hh pathway. Our results suggest involvement of the Hedgehog pathway in CPHD. We suggest that both SHH and HHIP are investigated as a second screening in CPHD, after mutations in the classical CPHD genes have been ruled out.

Clinical Case Seminar. Peculiar prolactinomas in patients with pituitary developmental gene mutations: from an adult endocrinologist perspective

Congenital hypopituitarism is a syndrome which is associated with single or multiple pituitary hormone deficiencies. Mutations in a number of developmental genes have been linked to combined pituitary hormone deficiencies, the most common being mutation in the pituitary homeobox protein prophet of the Pit 1 gene (PROP1). PROP1 exhibits DNA-binding and transcriptional activities. On magnetic resonance imaging, most patients with PROP1 mutation have a hypoplastic pituitary gland. Occasionally, transient pituitary enlargement before definite involution is reported. Kallmann syndrome (KS) is a human developmental genetic disorder which is a clinically (isolated hypogonadotropic hypogonadism-IHH) and genetically heterogeneous disease. Routine neuroimaging in classical IHH is thought to be of limited clinical value and normal anatomy of the hypothalamic-pituitary region is often reported. For neither disorder are there many reports on imaging during adulthood. Nor are there any guidelines concerning long-term imaging follow-up in patients with developmental pituitary disorders. Our aim was to present unusual endocrine and imaging abnormalities which developed in adulthood in two patients with developmental pituitary disorders. We report a female with combined pituitary hormone deficiencies (GH, TSH, gonadotropin and ACTH), except for prolactin, as a consequence of PROP1 mutation, and a male with KS (anosmia and IHH) due to Kal 2 gene (fibroblast growth factor receptor 1- FGFR1) mutation, both of whom in adulthood presented with prolactinomas. Both patients with developmental gene mutations, after long-term correction of their sex steroid status, developed prolactinomas. Although the exact mechanism of pituitary tumorigenesis is not known, we speculate that sex steroids may have facilitated prolactinoma development from the prolactin cell pool which underwent uncontrolled proliferation in the setting of a developmental disorder.

Genetic causes of combined pituitary hormone deficiencies in humans

Congenital hypopituitarism is a rare disease, usually induced by mutations of genes coding for transcription factors involved in pituitary development. PROP1 mutations represent the first cause of identified congenital hypopituitarism. Current techniques only identify 10–20% of congenital hypopituitarism etiologies, suggesting that new techniques are needed to improve this ratio. This should lead to a better management and follow-up of patients presenting with combined pituitary hormone deficiencies.