A New Compound Heterozygosis For Inactivating Mutations In The Glucokinase Gene As Cause Of Permanent Neonatal Diabetes Mellitus (pndm) In Double-first Cousins

Background: Permanent neonatal diabetes mellitus (PNDM) is a rare disorder, characterized by uncontrolled hyperglycemia diagnosed during the first 6 months of life. In general, PNDM has a genetic origin and most frequently it results from heterozygous mutations in KCNJ11, INS and ABCC8 genes. Homozygous or compound heterozygous inactivating mutations in GCK gene as cause of PNDM are rare. In contrast, heterozygosis for GCK inactivating mutations is frequent and results in the maturity-onset diabetes of young (MODY), manifested by a mild fasting hyperglycemia usually detected later in life. Therefore, as an autosomal recessive disorder, GCK-PNDM should be considered in families with history of glucose intolerance or MODY in first relatives, especially when consanguinity is suspected. Results: Here we describe two patients born from non-consanguineous parents within a family. They presented low birth weight with persistent hyperglycemia during the first month of life. Molecular analyses for KCNJ11, INS, ABCC8 did not show any mutation. GCK gene sequencing, however, revealed that both patients were compound heterozygous for two missense combined in a novel GCK-PNDM genotype. The p.Asn254His and p.Arg447Gly mutations had been inherited from their mothers and fathers, respectively, as their mothers are sisters and their fathers are brothers. Parents had been later diagnosed as having GCK-MODY. Conclusions: Mutations' in silico analysis was carried out to elucidate the role of the amino acid changes on the enzyme structure. Both p.Asn254His and p.Arg447Gly mutations appeared to be quite damaging. This is the first report of GCK-PNDM in a Brazilian family.

Phenotypic Variability In A Family With X-linked Adrenoleukodystrophy Caused By The P.trp132ter Mutation.

X-linked adrenoleukodystrophy (X-ALD) is an inherited disease with clinical heterogeneity varying from presymptomatic individuals to rapidly progressive cerebral ALD forms. This disease is characterized by increased concentration of very long chain fatty acids (VLCFAs) in plasma and in adrenal, testicular and nervous tissues. Affected individuals can be classified in different clinical settings, according to phenotypic expression and age at onset of initial symptoms. Molecular defects in X-ALD individuals usually result from ABCD1 gene mutations. In the present report we describe clinical data and the ABCD1 gene study in two boys affected with the childhood cerebral form that presented with different symptomatic manifestations at diagnosis. In addition, their maternal grandfather had been diagnosed with Addison's disease indicating phenotypic variation for X-ALD within this family. The mutation p.Trp132Ter was identified in both male patients; additionally, three females, out of eleven family members, were found to be heterozygous after screening for this mutation. In the present report, the molecular analysis was especially important since one of the heterozygous females was in first stages of pregnancy. Therefore, depending on the fetus outcome, if male and p.Trp132Ter carrier, storage of the umbilical cord blood should be recommended as hematopoietic stem cell transplantation could be considered as an option for treatment in the future.54738-4

A New Compound Heterozygosis For Inactivating Mutations In The Glucokinase Gene As Cause Of Permanent Neonatal Diabetes Mellitus (pndm) In Double-first Cousins.

Permanent neonatal diabetes mellitus (PNDM) is a rare disorder, characterized by uncontrolled hyperglycemia diagnosed during the first 6 months of life. In general, PNDM has a genetic origin and most frequently it results from heterozygous mutations in KCNJ11, INS and ABCC8 genes. Homozygous or compound heterozygous inactivating mutations in GCK gene as cause of PNDM are rare. In contrast, heterozygosis for GCK inactivating mutations is frequent and results in the maturity-onset diabetes of young (MODY), manifested by a mild fasting hyperglycemia usually detected later in life. Therefore, as an autosomal recessive disorder, GCK-PNDM should be considered in families with history of glucose intolerance or MODY in first relatives, especially when consanguinity is suspected. Here we describe two patients born from non-consanguineous parents within a family. They presented low birth weight with persistent hyperglycemia during the first month of life. Molecular analyses for KCNJ11, INS, ABCC8 did not show any mutation. GCK gene sequencing, however, revealed that both patients were compound heterozygous for two missense combined in a novel GCK-PNDM genotype. The p.Asn254His and p.Arg447Gly mutations had been inherited from their mothers and fathers, respectively, as their mothers are sisters and their fathers are brothers. Parents had been later diagnosed as having GCK-MODY. Mutations' in silico analysis was carried out to elucidate the role of the amino acid changes on the enzyme structure. Both p.Asn254His and p.Arg447Gly mutations appeared to be quite damaging. This is the first report of GCK-PNDM in a Brazilian family.710

Long-term Follow-up Of An 8-year-old Boy With Insulinoma As The First Manifestation Of A Familial Form Of Multiple Endocrine Neoplasia Type 1.

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant hereditary cancer syndrome characterized mostly by parathyroid, enteropancreatic, and anterior pituitary tumors. We present a case of an 8-year-old boy referred because of hypoglycemic attacks. His diagnosis was pancreatic insulinoma. Paternal grandmother died due to repeated gastroduodenal ulcerations and a paternal aunt presented similar manifestations. At a first evaluation, the father presented only gastric ulceration but subsequently developed hyperparathyroidism and lung carcinoid tumor. During almost 15 years of follow-up, three brothers and the index case presented hyperparathyroidism and hyperprolactinemia. Molecular study showed a G to A substitution in intron 4, at nine nucleotides upstream of the splicing acceptor site, causing a splicing mutation. All affected members of the family have the same mutation. Paternal grandmother and aunt were not studied and the mother does not carry any mutation. MEN1 is a rare condition that requires permanent medical assistance. Early clinical and genetic identification of affected individuals is essential for their own surveillance and also for genetic counseling.54754-6

Molecular analysis of KCNJ11, ABCC8, INS and GCK genes in patients with neonatal Diabetes Mellitus

Orientadores: Sofia Helena Valente de Lemos Marini, Maricilda Palandi de MelloDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo: O diabetes mellitus neonatal (DMN) é definido pelo surgimento de hiperglicemia com necessidade de tratamento com insulina nos primeiros seis meses de vida. Entretanto, nas crianças com hiperglicemia de início no primeiro ano de vida propõe-se que seja feito o diagnóstico diferencial de diabetes mellitus tipo 1 (DM1) e DMN, especialmente através das dosagens de autoanticorpos (anti-insulina, anti-IA2, anti-GAD e anti-ICA). Dependendo do tempo de duração da doença, o DMN pode ser classificado em transitório (geralmente de início na primeira semana de vida com regressão ao redor dos 18 meses de idade, podendo recorrer na adolescência ou na vida adulta) e permanente (não entra em remissão ao longo da vida). A origem do DMN é heterogênea e pode resultar de uma falha no desenvolvimento pancreático, de uma diminuição da população de células beta-pancreáticas ou de uma disfunção das células beta limitando a secreção de insulina. O conhecimento de genes envolvidos na cascata do desenvolvimento pancreático e nos aspectos relacionados à homeostase da glicose possibilitou entendermos os mecanismos fisiopatológicos envolvidos na origem do DMN e de outros tipos de diabetes classificados de diabetes monogênico, como o MODY por exemplo. Além disso, o diagnóstico molecular trouxe benefícios para o seguimento desses pacientes, permitindo o aconselhamento genético, norteando o prognóstico e até o tratamento em alguns deles. Atualmente são conhecidos 21 genes responsáveis pelo DMN, sendo os mais comuns (em termos de incidência) o gene KCNJ11 (30 a 50% dos casos), o gene INS (16%), o ABCC8 (13%) e mais raramente o gene GCK. Neste trabalho foi realizada a análise molecular dos genes KCNJ11, ABCC8, INS e GCK em quatro pacientes diagnosticados com DMN, três deles com a forma permanente (DMNP) e um com a forma transitória (DMNT). Não foram encontradas mutações nos genes KCNJ11 e INS. O estudo molecular do gene ABCC8 identificou a mutação p.Phe577Leu, presente em heterozigose no único caso diagnosticado com DMNT. A análise estrutural dessa mutação não foi realizada pois não há, até o momento, proteínas com um grau confiável de similaridade que já tenham sido cristalizadas. As análises de predição realizadas através das ferramentas computacionais PolyPhen, SIFT e Align-GVGD confirmaram que se trata de uma mutação deletéria. O estudo molecular do gene GCK confirmou a presença de duas mutações, p.Asn254His e p.Arg447Gly, nos casos 1 e 2, configurando um genótipo de heterozigose composta. A primeira foi herdada das mães dos pacientes, que são irmãs e que foram diagnosticadas com MODY 2 enquanto a segunda mutação foi herdada dos pais dos pacientes, que são irmãos e também foram diagnosticados com MODY 2. No irmão mais novo do caso 1 também foi identificada a mutação p.Arg447Gly herdada do pai cujos exames laboratoriais identificaram leve hiperglicemia de jejum (MODY 2). A análise estrutural das mutações mostrou que ambas são bastante deletérias uma vez que comprometem a flexibilidade da enzima glucoquinase, prejudicando sua função. As análises de predição (PolyPhen, SIFT e Align-GVGD) também mostraram que ambas as mutações são potencialmente deletérias. Dos quatro casos estudados, apenas em um (caso 4) não foi possível estabelecer a origem molecular do DMNP; segundo a literatura, em 40% dos casos de DMN não é possível se estabelecer o diagnóstico molecularAbstract: Neonatal diabetes mellitus (DMN) is defined by the onset of hyperglycemia requiring insulin therapy in the first six months of life. However, the differential diagnosis between diabetes mellitus type 1 (DM1) and DMN in children with early hyperglycemia in the first year of life is necessary by dosage of autoantibodies (IAA, Anti-GAD and ICA). Depending on the duration of the disease, the DMN may be either transient (usually start in the first week of life with regression around 18 months of age, may relapse in adolescence or adulthood) and permanent (does not go into remission lifelong). The etiology of DMN is heterogeneous and may result from a failure in pancreatic development, a reduction in the pancreatic beta cell population, or a dysfunction of beta cells limiting insulin secretion. The knowledge of genes involved in the cascade of pancreatic development and the aspects related to glucose homeostasis allowed us to understand the pathophysiological mechanisms involved in the origin of the DMN and other types of monogenic diabetes such as MODY. In addition, molecular diagnosis has brought benefits to the follow-up of these patients, allowing genetic counseling, guiding prognosis and improving treatment in some of them. Currently, there are 21 genes known as responsible for DMN, and the most common (in terms of incidence) are: KCNJ11 (30 to 50% of cases), INS gene (16%), the ABCC8 (13%), and more rarely the gene GCK. In this work we performed a molecular analysis of genes KCNJ11, ABCC8, INS and GCK in four patients diagnosed with DMN, three of them with permanent (PNDM) and one with the transitory form (DMNT). There were no mutations in KCNJ11 and INS genes. Molecular analysis of ABCC8 revealed the heterozygosis for p.Phe577Leu mutation in the case diagnosed with DMNT. Structural analysis for this mutation was not performed because there are not proteins with a reliable degree of similarity that have already been crystallized. The prediction analyzes through PolyPhen, SIFT and Align-GVGD computational tools confirmed that p.Phe577Leu is a deleterious mutation. Molecular analysis of GCK gene confirmed the compound heterozygosis for p.Asn254His and p.Arg447Gly mutations in cases 1 and 2. The first was inherited from the mothers of the patients, who are sisters and were diagnosed with MODY 2, whereas the second mutation was inherited from fathers, who are brothers and were also diagnosed with MODY 2. The younger brother of case 1, whose laboratory tests revealed impaired fasting glycemia (MODY 2), also carries the p.Arg447Gly mutation. Structural analysis for both mutations showed that they are quite deleterious since they compromise the flexibility of the glucokinase enzyme, damaging its function. The prediction analysis (PolyPhen, SIFT and Align-GVGD) have also shown that both mutations are potentially damaging. Only one (case 4) out of four cases studied, did not have the molecular origin of PNDM established; according to the literature, this can happen in 40% of casesMestradoSaude da Criança e do AdolescenteMestra em Ciência

Variabilidade fenotípica em uma família com adrenoleucodistrofia ligada ao X causada pela mutação p.Trp132Ter

X-linked adrenoleukodystrophy (X-ALD) is an inherited disease with clinical heterogeneity varying from presymptomatic individuals to rapidly progressive cerebral ALD forms. This disease is characterized by increased concentration of very long chain fatty acids (VLCFAs) in plasma and in adrenal, testicular and nervous tissues. Affected individuals can be classified in different clinical settings, according to phenotypic expression and age at onset of initial symptoms. Molecular defects in X-ALD individuals usually result from ABCD1 gene mutations. In the present report we describe clinical data and the ABCD1 gene study in two boys affected with the childhood cerebral form that presented with different symptomatic manifestations at diagnosis. In addition, their maternal grandfather had been diagnosed with Addison's disease indicating phenotypic variation for X-ALD within this family. The mutation p.Trp132Ter was identified in both male patients; additionally, three females, out of eleven family members, were found to be heterozygous after screening for this mutation. In the present report, the molecular analysis was especially important since one of the heterozygous females was in first stages of pregnancy. Therefore, depending on the fetus outcome, if male and p.Trp132Ter carrier, storage of the umbilical cord blood should be recommended as hematopoietic stem cell transplantation could be considered as an option for treatment in the future.A adrenoleucodistrofia é uma doença genética com padrão de herança ligado ao X (X-ALD) que apresenta heterogeneidade clínica e varia desde a forma infantil cerebral severa até casos de indivíduos pré-sintomáticos. Essa doença é caracterizada pelo acúmulo de ácidos graxos de cadeia muito longa (VLCFA) no plasma, nas adrenais, nos testículos e no sistema nervoso. Indivíduos afetados podem apresentar diferentes formas clínicas, as quais são classificadas de acordo com a expressão fenotípica e a idade de aparecimento dos sintomas iniciais. Alterações moleculares em indivíduos com X-ALD são geralmente mutações no gene ABCD1. No presente trabalho, descrevemos os dados clínicos e a investigação molecular do gene ABCD1 em uma família com duas crianças do sexo masculino afetadas com a forma infantil cerebral, que apresentaram diferenças nas primeiras manifestações sintomáticas para o diagnóstico. Além disso, houve referência ao avô materno diagnosticado com doença de Addison's, indicando a variabilidade fenotípica da X-ALD nessa família. A análise molecular indicou a mutação p.Trp132Ter nos dois pacientes masculinos, e três indivíduos do sexo feminino, entre os onze estudados, mostraram-se heterozigotos para mutação. O conhecimento molecular descrito no presente relato adquiriu maior importância uma vez que uma das portadoras da mutação apresentou-se nos primeiros estágios de gestação. Assim, poderá ser oferecida a possibilidade de armazenamento de sangue de cordão umbilical para que se possa considerar, no futuro, o transplante de células-tronco hematopoiéticas como forma de tratamento, caso a criança seja do sexo masculino e afetada