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Adrenal myelolipomas (AML) are composed of mature adipose and hematopoietic components. They represent approximately 3 percent of adrenal tumors and are commonly found in patients with congenital adrenal hyperplasia (CAH). CAH provides a unique environment to explore AML pathogenesis. We aimed to evaluate the role of the immune system and hormones that accumulate in poorly controlled CAH in the development of AML. When compared to normal adrenal tissue, CAH-affected adrenal tissue and myelolipomas showed an increased expression of inflammatory cells (CD68, IL2Rbeta), stem cells (CD117) B cells (IRF4), and adipogenic markers (aP2/FABP4, AdipoQ, PPARγ, Leptin, CideA), and immunostaining showed nodular lymphocytic accumulation. Immunohistochemistry staining revealed a higher density of inflammatory cells (CD20, CD3, CD68) in CAH compared to non-CAH myelolipomas. In vitro RNA-sequencing studies using NCI-H295R adrenocortical cells with exogenous exposure to ACTH, testosterone, and 17-hydroxyprogesterone hormones, showed the differential expression of genes involved in cell cycle progression, phosphorylation, and tumorigenesis. Migration of B-lymphocytes was initiated after the hormonal treatment of adrenocortical cells using the Boyden chamber chemotaxis assay, indicating a possible hormonal influence on triggering inflammation and the development of myelolipomas. These findings demonstrate the important role of inflammation and the hormonal milieu in the development of AML in CAH.
Assuntos
Neoplasias das Glândulas Suprarrenais , Hiperplasia Suprarrenal Congênita , Leucemia Mieloide Aguda , Lipoma , Mielolipoma , Humanos , Mielolipoma/patologia , Neoplasias das Glândulas Suprarrenais/genéticaRESUMO
CONTEXT: High childhood obesity rates coincide with increased incidence of nonalcoholic fatty liver disease (NAFLD) and other comorbidities. Understanding the genetics of susceptibility to obesity and its comorbidities could guide intervention. The cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) signaling pathway regulates energy balance, glucose homeostasis, and lipid metabolism. OBJECTIVE: We hypothesized that PKA-related gene variants may be associated with obesity or associated metabolic conditions. METHODS: We included 457 youths from the Yale Obesity Clinic into the Pathogenesis of Youth-Onset Diabetes cohort (NCT01967849); a variety of clinical tests were performed to characterize NAFLD. Exon sequencing of 54 PKA pathway genes was performed. Variants were confirmed by Sanger sequencing. Clinical data were analyzed, correcting for NAFLD status and body mass index z-score with adjustments for multiple comparisons. Fluorescence resonance energy transfer (FRET) and PKA enzymatic assays were performed in HEK293 cells transfected with the PRKAR1B p.R115K construct. In silico structural analysis for this variant was done. RESULTS: We identified the variant PRKAR1B p.R115K in 4 unrelated, African American patients. Analyses compared this variant group to other African American patients in the cohort. PRKAR1B p.R115K was associated with favorable circulating lipoprotein levels. Analysis of FRET and PKA enzymatic assay showed stronger interaction between the R1ß mutant and PKA catalytic subunit Cα and decreased basal PKA activity compared with the wildtype (Pâ <â .0001). Structural analysis revealed that p.R115K may hinder conformational changes resulting from cAMP binding at cAMP binding domain A. CONCLUSION: Data suggest PRKAR1B p.R115K affects cAMP signaling and may favorably modulate lipoprotein profile in African American youth, protecting them from some adverse metabolic outcomes.
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Malignant pheochromocytomas (PCs) and paragangliomas (PGLs) are rare neuroendocrine neoplasms defined by the presence of distant metastases. There is currently a relatively paucity of data regarding the natural history of PCs/PGLs and the optimal approach to their treatment. We retrospectively analyzed the clinical, biochemical, imaging, genetic and histopathological characteristics of fourteen patients with metastatic PCs/PGLs diagnosed over 15 years, along with their response to treatment. Patients were followed-up for a median of six years (range: 1-14 years). Six patients had synchronous metastases and the remaining developed metastases after a median of four years (range 2-10 years). Genetic analysis of seven patients revealed that three harbored succinate dehydrogenase subunit B/D gene (SDHB/D) mutations. Hormonal hypersecretion occurred in 70% of patients; normetanephrine, either alone or with other concomitant hormones, was the most frequent secretory component. Patients were administered multiple first and subsequent treatments including surgery (n = 12), chemotherapy (n = 7), radionuclide therapy (n = 2) and radiopeptides (n = 5). Seven patients had stable disease, four had progressive disease and three died. Ectopic hormonal secretion is rare and commonly encountered in benign PCs. Ectopic secretion of interleukin-6 in one of our patients, prompted a literature review of ectopic hormonal secretion, particularly from metastatic PCs/PGLs. Only four cases of metastatic PC/PGLs with confirmed ectopic secretion of hormones or peptides have been described so far.
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Carney complex is a rare, autosomal dominant, multiple endocrine neoplasia and lentiginosis syndrome, caused in most patients by defects in the PRKAR1A gene, which encodes the regulatory subunit type 1α of protein kinase A. Inactivating defects of PRKAR1A lead to aberrant cyclic-AMP-protein kinase A signaling. Patients may develop multiple skin abnormalities and a variety of endocrine and non-endocrine tumors. Endocrine manifestations include primary pigmented nodular adrenocortical disease, that may cause Cushing syndrome, growth-hormone secreting pituitary adenoma or pituitary somatotropic hyperplasia which can result in acromegaly, as well as gonadal and thyroid tumors. Non-endocrine tumors associated with Carney complex include myxomas of the heart, breast, and other sites, psamommatous melanotic schwannomas, breast ductal adenomas, osteochondromyxomas, and a predisposition to a number of malignancies from adrenal to pancreatic and liver cancer.
Assuntos
Complexo de Carney , Complexo de Carney/complicações , Complexo de Carney/genética , Complexo de Carney/metabolismo , Complexo de Carney/patologia , HumanosRESUMO
CONTEXT: Carney complex (CNC) is a syndrome characterized by hyperplasia of endocrine organs and may present with clinical features of Cushing syndrome and acromegaly due to functional adrenal and pituitary gland tumors. CNC has been linked to mutations in the regulatory subunit of protein kinase A type I-alpha (PRKAR1A) gene. DESIGN: Tissue samples were taken from the hypothalamus or thalamus or tumors of patients with pituitary adenomas seen and operated on by neurosurgeon Harvey Cushing between 1913 and 1932. Following DNA extraction, sequencing for genes of interest was attempted, including PRKAR1A, AIP, USP8, GNAS1, and GPR101, to explore the possibility that these mutations associated with acromegaly, CNC, and Cushing syndrome have been conserved over time. RESULTS: We report a patient described by Dr. Cushing in 1914 with a clinical presentation and postmortem findings suggestive of CNC. Genetic sequencing of the hypothalamus and pituitary adenoma revealed a germline heterozygous p.Arg74His mutation in the PRKAR1A gene, a codon previously described as mutated in CNC, but with a novel amino acid change. CONCLUSIONS: This patient is, to our knowledge, the first molecularly confirmed individual with CNC. This case demonstrates the power of modern genetics in studying archived tissues and the importance of recording detailed clinical notes in the diagnosis of disease.
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Ovarian cancer (OC) is the eighth most common cancer among women in Brazil and seventh in the world population. OC has a high mortality rate and is difficult to diagnose. Currently, OC detection most often occurs at an advanced stage of the disease due to its silent progression, which contributes to the high mortality rate. Available genetic markers are not considered specifically enough for an initial and definite diagnosis. The association with new genes involved with OC can provide a better understanding of this pathology as well as contribute to the development of a marker scenario, providing an improvement in the treatment and survival of patients. The aim of this study was to examine the potential association between the PARK2 gene and epithelial ovarian cancer (EOC). Accordingly, we conducted a study for which 25 patients and 87 controls were recruited. Linkage disequilibrium analysis showed that the four studied tag SNPs (rs2803073, rs6930532, rs1040079, and rs2276201) were independent. Our results using the multivariate analysis between the additive and dominant model demonstrated that tag SNP rs2803073 of PARK2 is associated with susceptibility to EOC (p = 0.018, OR = 0.42). These findings suggest that hereditary variation in the PARK2 gene could influence EOC development mechanisms.
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Predisposição Genética para Doença/genética , Neoplasias Epiteliais e Glandulares/genética , Neoplasias Ovarianas/genética , Ubiquitina-Proteína Ligases/genética , Brasil , Carcinoma Epitelial do Ovário , Estudos de Casos e Controles , Feminino , Mutação em Linhagem Germinativa , Humanos , Desequilíbrio de Ligação , Polimorfismo de Nucleotídeo ÚnicoRESUMO
CONTEXT: We have recently reported somatic mutations in the ubiquitin-specific protease USP8 gene in a small series of adenomas of patients with Cushing's disease. OBJECTIVE: To determine the prevalence of USP8 mutations and the genotype-phenotype correlation in a large series of patients diagnosed with Cushing's disease. DESIGN: We performed a retrospective, multicentric, genetic analysis of 134 functioning and 11 silent corticotroph adenomas using Sanger sequencing. Biochemical and clinical features were collected and examined within the context of the mutational status of USP8, and new mutations were characterized by functional studies. PATIENTS: A total of 145 patients who underwent surgery for an ACTH-producing pituitary adenoma. MAIN OUTCOMES MEASURES: Mutational status of USP8. Biochemical and clinical features included sex, age at diagnosis, tumor size, preoperative and postoperative hormonal levels, and comorbidities. RESULTS: We found somatic mutations in USP8 in 48 (36%) pituitary adenomas from patients with Cushing's disease but in none of 11 silent corticotropinomas. The prevalence was higher in adults than in pediatric cases (41 vs 17%) and in females than in males (43 vs 17%). Adults having USP8-mutated adenomas were diagnosed at an earlier age than those with wild-type lesions (36 vs 44 y). Mutations were primarily found in adenomas of 10 ± 7 mm and were inversely associated with the development of postoperative adrenal insufficiency. All the mutations affected the residues Ser718 or Pro720, including five new identified alterations. Mutations reduced the interaction between USP8 and 14-3-3 and enhanced USP8 activity. USP8 mutants diminished epidermal growth factor receptor ubiquitination and induced Pomc promoter activity in immortalized AtT-20 corticotropinoma cells. CONCLUSIONS: USP8 is frequently mutated in adenomas causing Cushing's disease, especially in those from female adult patients diagnosed at a younger age.
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Adenoma Hipofisário Secretor de ACT/genética , Adenoma/genética , Endopeptidases/genética , Complexos Endossomais de Distribuição Requeridos para Transporte/genética , Mutação , Hipersecreção Hipofisária de ACTH/genética , Ubiquitina Tiolesterase/genética , Adenoma Hipofisário Secretor de ACT/epidemiologia , Adenoma/epidemiologia , Adolescente , Adulto , Animais , Células COS , Criança , Chlorocebus aethiops , Análise Mutacional de DNA , Feminino , Frequência do Gene , Estudos de Associação Genética , Células HeLa , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Hipersecreção Hipofisária de ACTH/epidemiologia , Estudos Retrospectivos , Células Tumorais Cultivadas , Adulto JovemRESUMO
Familial isolated pituitary adenoma (FIPA) occurs in families and is unrelated to multiple endocrine neoplasia type 1 and Carney complex. Mutations in AIP account only for 15-25% of FIPA families. CDKN1B mutations cause MEN4 in which affected patients can suffer from pituitary adenomas. With this study, we wanted to assess whether mutations in CDKN1B occur among a large cohort of AIP mutation-negative FIPA kindreds. Eighty-eight AIP mutation-negative FIPA families were studied and 124 affected subjects underwent sequencing of CDKN1B. Functional analysis of putative CDKN1B mutations was performed using in silico and in vitro approaches. Germline CDKN1B analysis revealed two nucleotide changes: c.286A>C (p.K96Q) and c.356T>C (p.I119T). In vitro, the K96Q change decreased p27 affinity for Grb2 but did not segregate with pituitary adenoma in the FIPA kindred. The I119T substitution occurred in a female patient with acromegaly. p27(I119T) shows an abnormal migration pattern by SDS-PAGE. Three variants (p.S56T, p.T142T, and c.605+36C>T) are likely nonpathogenic because In vitro effects were not seen. In conclusion, two patients had germline sequence changes in CDKN1B, which led to functional alterations in the encoded p27 proteins in vitro. Such rare CDKN1B variants may contribute to the development of pituitary adenomas, but their low incidence and lack of clear segregation with affected patients make CDKN1B sequencing unlikely to be of use in routine genetic investigation of FIPA kindreds. However, further characterization of the role of CDKN1B in pituitary tumorigenesis in these and other cases could help clarify the clinicopathological profile of MEN4.
Assuntos
Adenoma/genética , Inibidor de Quinase Dependente de Ciclina p27/genética , Neoplasias Hipofisárias/genética , Linhagem Celular Tumoral , Família , Feminino , Variação Genética , Genótipo , Células HeLa , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , MutaçãoAssuntos
Antineoplásicos Fitogênicos/uso terapêutico , Orquiectomia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/cirurgia , Taxoides/uso terapêutico , Inibidores da Aromatase/uso terapêutico , Docetaxel , Estrogênios/metabolismo , Humanos , Masculino , Neoplasias da Próstata/genética , Receptores de Estrogênio/efeitos dos fármacos , Receptores de Estrogênio/genética , Fatores de RiscoRESUMO
BACKGROUND: Carney complex (CNC) is an autosomal dominant multiple neoplasia, caused mostly by inactivating mutations of the regulatory subunit 1A of the protein kinase A (PRKAR1A). Primary pigmented nodular adrenocortical disease (PPNAD) is the most frequent endocrine manifestation of CNC with a great inter-individual variability. Germline, protein-truncating mutations of phosphodiesterase type 11A (PDE11A) have been described to predispose to a variety of endocrine tumors, including adrenal and testicular tumors. OBJECTIVES: Our objective was to investigate the role of PDE11A as a possible gene modifier of the phenotype in a series of 150 patients with CNC. RESULTS: A higher frequency of PDE11A variants in patients with CNC compared with healthy controls was found (25.3 vs. 6.8%, P < 0.0001). Among CNC patients, those with PPNAD were significantly more frequently carriers of PDE11A variants compared with patients without PPNAD (30.8 vs. 13%, P = 0.025). Furthermore, men with PPNAD were significantly more frequently carriers of PDE11A sequence variants (40.7%) than women with PPNAD (27.3%) (P < 0.001). A higher frequency of PDE11A sequence variants was also found in patients with large-cell calcifying Sertoli cell tumors (LCCSCT) compared with those without LCCSCT (50 vs. 10%, P = 0.0056). PDE11A variants were significantly associated with the copresence of PPNAD and LCCSCT in men: 81 vs. 20%, P < 0.004). The simultaneous inactivation of PRKAR1A and PDE11A by small inhibitory RNA led to an increase in cAMP-regulatory element-mediated transcriptional activity under basal conditions and after stimulation by forskolin. CONCLUSIONS: We demonstrate, in a large cohort of CNC patients, a high frequency of PDE11A variants, suggesting that PDE11A is a genetic modifying factor for the development of testicular and adrenal tumors in patients with germline PRKAR1A mutation.
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Neoplasias das Glândulas Suprarrenais/genética , Complexo de Carney/genética , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/genética , Diester Fosfórico Hidrolases/genética , Tumor de Células de Sertoli/genética , Neoplasias Testiculares/genética , 3',5'-GMP Cíclico Fosfodiesterases , Adolescente , Adulto , Criança , Pré-Escolar , Feminino , Variação Genética , Células HEK293 , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Fenótipo , RNA Interferente Pequeno , Fatores SexuaisRESUMO
CONTEXT: Among the genomic loci harboring potential candidate genes for prostatic cancer (PCa) is the 2q31-33 chromosomal region that harbors the gene encoding phosphodiesterase 11A (PDE11A). In addition, the combined cancer genome expression metaanalysis datasets included PDE11A among the top 1% down-regulated genes in PCa. OBJECTIVE: In the present study, we screened 50 unrelated PCa patients of Brazilian descent for PDE11A coding defects. DESIGN: The study consisted of PDE11A sequencing, in vitro functional assays, and immunostaining analysis. RESULTS: We identified eight different sequence alterations in 15 patients (30%): one stop-codon and seven missense mutations. Three of the variants (R202C, Y658C, and E840K) were novel, and the remaining five (Y727C, R804H, R867G, M878V, and R307X) have been associated with predisposition to adrenal or testicular tumors. The overall prevalence of PDE11A-inactivating sequence variants among PCa patients was significantly higher than in 287 healthy controls (0.16 vs. 0.051, respectively, P < 0.001, odds ratio 3.81, 95% confidence interval 1.86-7.81) and the R202C, Y658C, and E840K substitutions were not found in controls. All missense mutations led to decreased PDE11A activity in human embryonic kidney 293 and PC3M cells and immunostaining of PCa samples with sequence changes showed decreased PDE11A protein expression. CONCLUSION: Our data suggest that, like in the adrenal cortex and the testicular germ cells, PDE11A-inactivating genetic alterations may play a role in susceptibility to PCa.
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Predisposição Genética para Doença , Variação Genética , Diester Fosfórico Hidrolases/genética , Neoplasias da Próstata/genética , 3',5'-GMP Cíclico Fosfodiesterases , Brasil , Células HEK293 , Humanos , Imuno-Histoquímica , Masculino , Mutação , Diester Fosfórico Hidrolases/metabolismo , Neoplasias da Próstata/metabolismoRESUMO
PRKAR1A encodes the regulatory subunit type 1-alpha (RIalpha) of the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA). Inactivating PRKAR1A mutations are known to be responsible for the multiple neoplasia and lentiginosis syndrome Carney complex (CNC). To date, at least 117 pathogenic variants in PRKAR1A have been identified (online database: http://prkar1a.nichd.nih.gov). The majority are subject to nonsense mediated mRNA decay (NMD), leading to RIalpha haploinsufficiency and, as a result, activated cAMP signaling. Recently, it became apparent that CNC may be caused not only by RIalpha haploinsufficiency, but also by the expression of altered RIalpha protein, as proven by analysis of expressed mutations in the gene, consisting of amino acid substitutions and in-frame genetic alterations. In addition, a new subgroup of mutations that potentially escape NMD and result in CNC through altered (rather than missing) protein has been analyzed-these are frame-shifts in the 3' end of the coding sequence that shift the stop codon downstream of the normal one. The mutation detection rate in CNC patients is recently estimated at above 60%; PRKAR1A mutation-negative CNC patients are characterized by significant phenotypic heterogeneity. In this report, we present a comprehensive analysis of all presently known PRKAR1A sequence variations and discuss their molecular context and clinical phenotype.
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Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/genética , Mutação/genética , Polimorfismo de Nucleotídeo Único/genética , Processamento Alternativo/genética , Complexo de Carney/diagnóstico , Complexo de Carney/enzimologia , Complexo de Carney/genética , Mutação da Fase de Leitura/genética , Estudos de Associação Genética , Humanos , Penetrância , Deleção de Sequência/genéticaRESUMO
OBJECTIVE: The present study evaluated the association between lactotransferrin (LTF) gene polymorphism (exon 2, A/G, Lys/Arg) and dental caries. MATERIAL AND METHODS: A convenience sample of 110 individuals, 12 years old, was divided into: group 1, 48 individuals without caries experience (DMFT=0), and group 2, 62 subjects with caries experience (DMFT>1). DNA was obtained from a mouthwash with 3 percent glucose solution, followed by a scrapping of the oral mucosa. After DNA purification, polymerase chain reaction (PCR), single strand conformation polymorphism (SSCP) was performed to access the study polymorphism. The LTF A/G (Lys/Arg) polymorphism had been previously reported as located in exon 1. RESULTS: Allele 1 of the study polymorphism was associated with low DMFT index and showed a protective effect against caries experience (OR=0.16, IC=0.03-0.76, p=0.01). CONCLUSIONS: Lactotransferrin A/G (exon 2, Lys/Arg) polymorphism was associated with susceptibility to dental caries in 12-year-old students.
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Criança , Humanos , Suscetibilidade à Cárie Dentária/genética , Cárie Dentária/genética , Lactoferrina/genética , Alelos , Substituição de Aminoácidos , Arginina , Sequência de Bases , Estudos de Casos e Controles , Índice CPO , Análise Mutacional de DNA , Frequência do Gene , Lisina , Dados de Sequência Molecular , Polimorfismo de Nucleotídeo Único , Polimorfismo Conformacional de Fita Simples , SalivaRESUMO
Inactivating germline mutations in phosphodiesterase 11A (PDE11A) have been implicated in adrenal tumor susceptibility. PDE11A is highly expressed in endocrine steroidogenic tissues, especially the testis, and mice with inactivated Pde11a exhibit male infertility, a known testicular germ cell tumor (TGCT) risk factor. We sequenced the PDE11A gene-coding region in 95 patients with TGCT from 64 unrelated kindreds. We identified 8 nonsynonymous substitutions in 20 patients from 15 families: four (R52T, F258Y, G291R, and V820M) were newly recognized, three (R804H, R867G, and M878V) were functional variants previously implicated in adrenal tumor predisposition, and one (Y727C) was a known polymorphism. We compared the frequency of these variants in our patients to unrelated controls that had been screened and found negative for any endocrine diseases: only the two previously reported variants, R804H and R867G, known to be frequent in general population, were detected in these controls. The frequency of all PDE11A-gene variants (combined) was significantly higher among patients with TGCT (P = 0.0002), present in 19% of the families of our cohort. Most variants were detected in the general population, but functional studies showed that all these mutations reduced PDE activity, and that PDE11A protein expression was decreased (or absent) in TGCT samples from carriers. This is the first demonstration of the involvement of a PDE gene in TGCT, although the cyclic AMP signaling pathway has been investigated extensively in reproductive organ function and their diseases. In conclusion, we report that PDE11A-inactivating sequence variants may modify the risk of familial and bilateral TGCT.