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1.
N Engl J Med ; 384(5): 428-439, 2021 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-33471991

RESUMO

BACKGROUND: Genetic testing for breast cancer susceptibility is widely used, but for many genes, evidence of an association with breast cancer is weak, underlying risk estimates are imprecise, and reliable subtype-specific risk estimates are lacking. METHODS: We used a panel of 34 putative susceptibility genes to perform sequencing on samples from 60,466 women with breast cancer and 53,461 controls. In separate analyses for protein-truncating variants and rare missense variants in these genes, we estimated odds ratios for breast cancer overall and tumor subtypes. We evaluated missense-variant associations according to domain and classification of pathogenicity. RESULTS: Protein-truncating variants in 5 genes (ATM, BRCA1, BRCA2, CHEK2, and PALB2) were associated with a risk of breast cancer overall with a P value of less than 0.0001. Protein-truncating variants in 4 other genes (BARD1, RAD51C, RAD51D, and TP53) were associated with a risk of breast cancer overall with a P value of less than 0.05 and a Bayesian false-discovery probability of less than 0.05. For protein-truncating variants in 19 of the remaining 25 genes, the upper limit of the 95% confidence interval of the odds ratio for breast cancer overall was less than 2.0. For protein-truncating variants in ATM and CHEK2, odds ratios were higher for estrogen receptor (ER)-positive disease than for ER-negative disease; for protein-truncating variants in BARD1, BRCA1, BRCA2, PALB2, RAD51C, and RAD51D, odds ratios were higher for ER-negative disease than for ER-positive disease. Rare missense variants (in aggregate) in ATM, CHEK2, and TP53 were associated with a risk of breast cancer overall with a P value of less than 0.001. For BRCA1, BRCA2, and TP53, missense variants (in aggregate) that would be classified as pathogenic according to standard criteria were associated with a risk of breast cancer overall, with the risk being similar to that of protein-truncating variants. CONCLUSIONS: The results of this study define the genes that are most clinically useful for inclusion on panels for the prediction of breast cancer risk, as well as provide estimates of the risks associated with protein-truncating variants, to guide genetic counseling. (Funded by European Union Horizon 2020 programs and others.).


Assuntos
Neoplasias da Mama/genética , Predisposição Genética para Doença/genética , Variação Genética , Mutação de Sentido Incorreto , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Modelos Logísticos , Pessoa de Meia-Idade , Razão de Chances , Risco , Análise de Sequência de DNA , Adulto Jovem
2.
Nucleic Acids Res ; 2020 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-32990755

RESUMO

The knowledge of the genetic variability of the local population is of utmost importance in personalized medicine and has been revealed as a critical factor for the discovery of new disease variants. Here, we present the Collaborative Spanish Variability Server (CSVS), which currently contains more than 2000 genomes and exomes of unrelated Spanish individuals. This database has been generated in a collaborative crowdsourcing effort collecting sequencing data produced by local genomic projects and for other purposes. Sequences have been grouped by ICD10 upper categories. A web interface allows querying the database removing one or more ICD10 categories. In this way, aggregated counts of allele frequencies of the pseudo-control Spanish population can be obtained for diseases belonging to the category removed. Interestingly, in addition to pseudo-control studies, some population studies can be made, as, for example, prevalence of pharmacogenomic variants, etc. In addition, this genomic data has been used to define the first Spanish Genome Reference Panel (SGRP1.0) for imputation. This is the first local repository of variability entirely produced by a crowdsourcing effort and constitutes an example for future initiatives to characterize local variability worldwide. CSVS is also part of the GA4GH Beacon network. CSVS can be accessed at: http://csvs.babelomics.org/.

3.
JAMA Dermatol ; 156(9): 987-991, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32756886

RESUMO

Importance: Persistent chemotherapy-induced alopecia (pCIA) has been recently described in patients with breast cancer and in its most severe form occurs in up to 10% of these patients. Genetic risk factors associated with pCIA have not been adequately explored. Objective: To identify genetic variants associated with pCIA. Design, Setting, and Participants: In this genetic association study, 215 women with breast cancer treated with docetaxel-based chemotherapy with a follow-up of 1.5 to 10 years after the end of the treatment were recruited retrospectively through 3 hospital oncology units across Spain between 2005 and 2018. Severe pCIA was defined as lack of scalp hair recovery (Common Terminology Criteria for Adverse Events, version 3.0, grade 2) 18 months or more after the end of treatment. Patients with grade 2 pCIA were selected as cases, and those with no sign of residual alopecia 12 months after the end of docetaxel treatment were selected as controls. A genome-wide association study in a discovery phase was conducted, and logistic regression was used to identify variants associated with the risk to develop this adverse effect. The validity of the association was addressed through a replication phase. Exposures: Docetaxel-based chemotherapy. Main Outcomes and Measures: Genotypes of single-nucleotide variants associated with pCIA. Results: In total, 215 women with breast cancer (median age, 51.6 years; interquartile range, 44-60 years) were recruited (173 patients for the discovery phase and 42 patients for the replication phase). In the discovery phase, ABCB1 genetic variants were associated with risk to develop pCIA. In particular, single-nucleotide variation rs1202179, a regulatory variant located in an enhancer element that interacts with the ABCB1 promoter, was associated with the occurrence of pCIA. This finding was validated in the replication cohort (combined odds ratio, 4.05; 95% CI, 2.46-6.67; P = 3.946 × 10-8). This variant is associated with ABCB1 mRNA expression, and the risk allele was associated with decreased ABCB1 expression levels (P = 1.64 × 10-20). Conclusions and Relevance: This is the first study, to our knowledge, that identifies an association between a regulatory variant in the ABCB1 gene and the occurrence of pCIA in patients with breast cancer who were treated with docetaxel-based therapies. This finding suggests an important insight into the biological mechanisms underlying pCIA and opens the opportunity to explore personalized treatment of these patients.

4.
Clin Pharmacol Ther ; 2020 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-32757270

RESUMO

Capecitabine-induced hand-foot syndrome (CiHFS) is a common dermatological adverse reaction affecting around 30% of patients with capecitabine-treated cancer, and the main cause of dose reductions and chemotherapy delays. To identify novel genetic factors associated with CiHFS in patients with cancer, we carried out an extreme-phenotype genomewide association study in 166 patients with breast and colorectal capecitabine-treated cancer with replication in a second cohort of 85 patients. We discovered and replicated a cluster of four highly correlated single-nucleotide polymorphisms associated with susceptibility to CiHFS at 20q13.33 locus (top hit = rs6129058, hazard ratio = 2.40, 95% confidence interval = 1.78-3.20; P = 1.2 × 10-8 ). Using circular chromosome conformation capture sequencing, we identified a chromatin contact between the locus containing the risk alleles and the promoter of CDH4, located 90 kilobases away. The risk haplotype was associated with decreased levels of CDH4 mRNA and the protein it encodes, R-cadherin (RCAD), which mainly localizes in the granular layer of the epidermis. In human keratinocytes, CDH4 downregulation resulted in reduced expression of involucrin, a protein of the cornified envelope, an essential structure for skin barrier function. Immunohistochemical analyses revealed that skin from patients with severe CiHFS exhibited low levels of RCAD and involucrin before capecitabine treatment. Our results uncover a novel mechanism underlying individual genetic susceptibility to CiHFS with implications for clinically relevant risk prediction.

5.
Cancers (Basel) ; 12(1)2019 Dec 25.
Artigo em Inglês | MEDLINE | ID: mdl-31881643

RESUMO

Pediatric Central Nervous System (CNS) tumors are the most fatal cancer diseases in childhood. Due to their localization and infiltrative nature, some tumor resections or biopsies are not feasible. In those cases, the use of minimally invasive methods as diagnostic, molecular marker detection, prognostic or monitoring therapies are emerging. The analysis of liquid biopsies which contain genetic information from the tumor has been much more widely explored in adults than in children. We compare the detection of BRAF V600E targetable mutation by digital-PCR from cell-free-DNA and EV-derived DNA (ctDNA) in serum, plasma and cerebrospinal fluid (CSF) isolated from a cohort of 29 CNS pediatric patients. Here we demonstrate that ctDNA isolated from serum and plasma could be successfully analyzed to obtain tumor genetic information which could be used to guide critical treatment decisions.

6.
Sci Rep ; 8: 46928, 2018 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-29308784

RESUMO

This corrects the article DOI: 10.1038/srep25749.

7.
PLoS One ; 11(12): e0168265, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27973562

RESUMO

Contradictory data about the impact of the rs738409 steatosis-related polymorphism within PNPLA3 gene on liver fibrosis progression in HIV/hepatitis C virus (HIV/HCV)-coinfected patients have been reported. Our objective was to test whether this, and other polymorphisms previously related to fatty liver disease in HIV infection linked to SAMM50 or LPPR4 genes, influence liver fibrosis progression in HIV/HCV-coinfected individuals. Three hundred and thirty two HIV/HCV-coinfected patients who consecutively attended four Spanish university hospitals from November 2011 to July 2013 were included. A liver stiffness cut-off of 14.6 kPa, as determined by transient elastography, was used to diagnose cirrhosis. Liver stiffness progression was studied in 171 individuals who had two available LS determinations without anti-HCV treatment between them. Moreover, 28 HIV/HCV-coinfected patients who underwent liver transplant, as well as 19 non-cirrhotic coinfected individuals used as controls, were included in an additional study. Only rs738409 was associated with cirrhosis: 45 (29.6%) of 152 G allele carriers versus 36 (20.0%) of 180 CC carriers showed cirrhosis (multivariate p = 0.018; adjusted odds ratio = 1.98; 95% confidence interval = 1.12-3.50). Also, 21 (30.4%) of 69 G allele carriers versus 16 (15.7%) of 102 CC patients showed significant liver stiffness progression (adjusted p-value = 0.015; adjusted odds ratio = 2.89; 95% confidence interval = 1.23-6.83). Finally, the proportion of rs738409 G allele carriers was significantly higher in transplanted individuals than in controls (p = 0.044, odds ratio = 3.43; 95% confidence interval = 1.01-11.70). Our results strongly suggest that the rs738409 polymorphism is associated with liver fibrosis progression in HIV/HCV-coinfected patients.


Assuntos
Infecções por HIV/genética , Infecções por HIV/fisiopatologia , Hepatite C Crônica/genética , Hepatite C Crônica/fisiopatologia , Lipase/genética , Cirrose Hepática/genética , Proteínas de Membrana/genética , Adulto , Alelos , Coinfecção/virologia , Estudos Transversais , Progressão da Doença , Técnicas de Imagem por Elasticidade , Fígado Gorduroso/genética , Feminino , Marcadores Genéticos , Variação Genética , Humanos , Cirrose Hepática/diagnóstico , Cirrose Hepática/virologia , Transplante de Fígado , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Razão de Chances , Polimorfismo Genético , Estudos Retrospectivos
8.
Sci Rep ; 6: 25749, 2016 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-27167008

RESUMO

Large-scale RNAseq has substantially changed the transcriptomics field, as it enables an unprecedented amount of high resolution data to be acquired. However, the analysis of these data still poses a challenge to the research community. Many tools have been developed to overcome this problem, and to facilitate the study of miRNA expression profiles and those of their target genes. While a few of these enable both kinds of analysis to be performed, they also present certain limitations in terms of their requirements and/or the restrictions on data uploading. To avoid these restraints, we have developed a suite that offers the identification of miRNA, mRNA and circRNAs that can be applied to any sequenced organism. Additionally, it enables differential expression, miRNA-mRNA target prediction and/or functional analysis. The miARma-Seq pipeline is presented as a stand-alone tool that is both easy to install and flexible in terms of its use, and that brings together well-established software in a single bundle. Our suite can analyze a large number of samples due to its multithread design. By testing miARma-Seq in validated datasets, we demonstrate here the benefits that can be gained from this tool by making it readily accessible to the research community.


Assuntos
MicroRNAs/genética , RNA Mensageiro/genética , RNA/genética , Análise de Sequência de RNA/métodos , MicroRNAs/metabolismo , RNA/metabolismo , RNA Circular , RNA Mensageiro/metabolismo
9.
J Infect ; 71(5): 571-7, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26212868

RESUMO

OBJECTIVES: To assess the incidence of hepatitis C virus (HCV) reinfections after therapy-induced clearance in HIV-coinfected patients with prior chronic hepatitis C. METHODS: Eighty-four HIV-infected subjects, who had previously achieved sustained virological response (SVR) after being treated of chronic hepatitis C, were analyzed. In all of them, at least yearly HCV RNA determinations were carried out during a median (range) of 34 (12-146) months. RESULTS: Seventy-two (86%) subjects had been people who inject drugs (PWID), of whom 11 (15%) continued to use snorted or injected drugs during the follow-up. Four (4.76%) patients showed HCV reinfection (incidence 1.21 [95% confidence interval: 0.3-3.09] cases per 100 person-years). These patients maintained risk factors for HCV infection. In three cases, HCV genotype switched. Phylogenetic analysis of the remaining case suggested reinfection from his sexual partner. CONCLUSION: The incidence of HCV reinfection in the overall population of HIV-coinfected patients who achieved SVR after being treated against chronic hepatitis C is low. A low frequency of risk behavior is the main factor accounting for this modest rate of reinfection. The possibility of reinfection should not be considered a reason against treatment of HCV infection with direct acting antivirals in PWID.


Assuntos
Usuários de Drogas , Infecções por HIV/complicações , Hepatite C Crônica/complicações , Hepatite C Crônica/tratamento farmacológico , Hepatite C/epidemiologia , Hepatite C/virologia , Adulto , Antivirais/uso terapêutico , Coinfecção/tratamento farmacológico , Coinfecção/imunologia , Coinfecção/virologia , Feminino , Seguimentos , Hepacivirus/genética , Hepacivirus/imunologia , Hepacivirus/fisiologia , Hepatite C/complicações , Hepatite C Crônica/epidemiologia , Hepatite C Crônica/imunologia , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Filogenia , RNA Viral/genética , Recidiva , Fatores de Risco , Espanha/epidemiologia
10.
J Clin Microbiol ; 52(11): 4027-9, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25143567

RESUMO

The accuracy of LiPA 2.0 for hepatitis C virus 1 (HCV-1) subtype classification was analyzed. LiPA 2.0 genotype results from 101 HCV-1-infected patients were compared to genotype findings determined by direct core sequencing. Eleven (11%) samples were misclassified. Given the influence of the HCV-1-subtype in the anti-HCV therapy response, an alternative classification method is warranted.


Assuntos
Antivirais/uso terapêutico , Hepacivirus/classificação , Hepacivirus/genética , Hepatite C Crônica/virologia , Adulto , Feminino , Genótipo , Hepacivirus/isolamento & purificação , Hepatite C Crônica/tratamento farmacológico , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , RNA Viral/genética , Análise de Sequência de DNA
11.
PLoS One ; 8(1): e54043, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23342068

RESUMO

Hirschsprung disease (HSCR) is a congenital malformation of the hindgut resulting from a disruption of neural crest cell migration during embryonic development. It has a complex genetic aetiology with several genes involved in its pathogenesis. PHOX2B plays a key function in the development of neural crest derivatives, and heterozygous mutations cause a complex dysautonomia associating HSCR, Congenital Central Hypoventilation Syndrome (CCHS) and neuroblastoma (NB) in various combinations. In order to determine the role of PHOX2B in isolated HSCR, we performed a mutational screening in a cohort of 207 Spanish HSCR patients. Our most relevant finding has been the identification of a de novo and novel deletion (c.393_410del18) in a patient with HSCR. Results of in silico and functional assays support its pathogenic effect related to HSCR. Therefore our results support that PHOX2B loss-of-function is a rare cause of HSCR phenotype.


Assuntos
Doença de Hirschsprung/genética , Doença de Hirschsprung/patologia , Proteínas de Homeodomínio/genética , Fatores de Transcrição/genética , Estudos de Casos e Controles , Biologia Computacional , Análise Mutacional de DNA , Feminino , Humanos , Masculino
12.
Orphanet J Rare Dis ; 7: 103, 2012 Dec 28.
Artigo em Inglês | MEDLINE | ID: mdl-23270508

RESUMO

Finding gene associations in rare diseases is frequently hampered by the reduced numbers of patients accessible. Conventional gene-based association tests rely on the availability of large cohorts, which constitutes a serious limitation for its application in this scenario. To overcome this problem we have used here a combined strategy in which a pathway-based analysis (PBA) has been initially conducted to prioritize candidate genes in a Spanish cohort of 53 trios of short-segment Hirschsprung's disease. Candidate genes have been further validated in an independent population of 106 trios. The study revealed a strong association of 11 gene ontology (GO) modules related to signal transduction and its regulation, enteric nervous system (ENS) formation and other HSCR-related processes. Among the preselected candidates, a total of 4 loci, RASGEF1A, IQGAP2, DLC1 and CHRNA7, related to signal transduction and migration processes, were found to be significantly associated to HSCR. Network analysis also confirms their involvement in the network of already known disease genes. This approach, based on the study of functionally-related gene sets, requires of lower sample sizes and opens new opportunities for the study of rare diseases.


Assuntos
Estudo de Associação Genômica Ampla/métodos , Doença de Hirschsprung/genética , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Masculino
13.
PLoS One ; 7(5): e36524, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22574178

RESUMO

Hirschsprung disease (HSCR, OMIM 142623) is a developmental disorder characterized by the absence of ganglion cells along variable lengths of the distal gastrointestinal tract, which results in tonic contraction of the aganglionic gut segment and functional intestinal obstruction. The RET proto-oncogene is the major gene for HSCR with differential contributions of its rare and common, coding and noncoding mutations to the multifactorial nature of this pathology. Many other genes have been described to be associated with the pathology, as NRG1 gene (8p12), encoding neuregulin 1, which is implicated in the development of the enteric nervous system (ENS), and seems to contribute by both common and rare variants. Here we present the results of a comprehensive analysis of the NRG1 gene in the context of the disease in a series of 207 Spanish HSCR patients, by both mutational screening of its coding sequence and evaluation of 3 common tag SNPs as low penetrance susceptibility factors, finding some potentially damaging variants which we have functionally characterized. All of them were found to be associated with a significant reduction of the normal NRG1 protein levels. The fact that those mutations analyzed alter NRG1 protein would suggest that they would be related with HSCR disease not only in Chinese but also in a Caucasian population, which reinforces the implication of NRG1 gene in this pathology.


Assuntos
Variação Genética/genética , Doença de Hirschsprung/genética , Neuregulina-1/genética , Animais , Células COS , Chlorocebus aethiops , Análise Mutacional de DNA , Sistema Nervoso Entérico/metabolismo , Feminino , Frequência do Gene/genética , Haplótipos/genética , Humanos , Masculino , Polimorfismo de Nucleotídeo Único/genética
14.
Am J Med Genet A ; 158A(4): 816-20, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22344793

RESUMO

X-linked hydrocephalus (XLH) has an incidence of 1/30,000 male births and is characterized by intellectual disability, spastic paraplegia, adducted thumbs, and agenesis of corpus callosum, and/or corticospinal tract. The great proportion of cases is ascribed to loss of function mutations of L1CAM gene. Hirschsprung disease (HSCR) is characterized by the absence of ganglion cells in the submucosal and myenteric plexuses along a variable portion of the intestinal tract and has incidence of about 1/5,000. Although with several genes involved in its pathogenesis, the major HSCR gene is the RET proto-oncogene. To date only a few patients have been reported with both phenotypes and mutations in the L1CAM gene. In this report, we describe a new patient with concurrent XLH and HSCR. L1CAM mutational screening showed the presence of the G698R hemizygous mutation, which is a deleterious substitution affecting a key residue necessary for the correct folding of the protein. Moreover, the patient also carried the transcriptional enhancer RET mutation (c.73 + 9277T > C) in heterozygosis. We speculate that both the RET enhancer variant, and the L1CAM mutation may act in combination to produce the enteric phenotype, probably with the participation of other still unidentified molecular events. While it is obvious that additional studies are necessary to further delineate the association between XLH and HSCR in the presence of L1CAM mutations, the documentation of this new patient reinforces the role of this gene acting either in a direct or indirect way in the pathogenesis of Hirschsprung disease.


Assuntos
Doenças Genéticas Ligadas ao Cromossomo X/genética , Doença de Hirschsprung/genética , Hidrocefalia/genética , Molécula L1 de Adesão de Célula Nervosa/genética , Anormalidades Múltiplas/genética , Sequência de Bases , Aqueduto do Mesencéfalo/anormalidades , Aqueduto do Mesencéfalo/patologia , Corpo Caloso/patologia , Análise Mutacional de DNA , Doenças Genéticas Ligadas ao Cromossomo X/diagnóstico , Doenças Genéticas Ligadas ao Cromossomo X/patologia , Doença de Hirschsprung/diagnóstico , Doença de Hirschsprung/patologia , Humanos , Hidrocefalia/diagnóstico , Hidrocefalia/patologia , Lactente , Masculino , Análise de Sequência de DNA
15.
BMC Med Genet ; 12: 138, 2011 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-21995290

RESUMO

BACKGROUND: RET is the major gene associated to Hirschsprung disease (HSCR) with differential contributions of its rare and common, coding and noncoding mutations to the multifactorial nature of this pathology. In the present study, we have performed a comprehensive study of our HSCR series evaluating the involvement of both RET rare variants (RVs) and common variants (CVs) in the context of the disease. METHODS: RET mutational screening was performed by dHPLC and direct sequencing for the identification of RVs. In addition Taqman technology was applied for the genotyping of 3 RET CVs previously associated to HSCR, including a variant lying in an enhancer domain within RET intron 1 (rs2435357). Statistical analyses were performed using the SPSS v.17.0 to analyze the distribution of the variants. RESULTS: Our results confirm the strongest association to HSCR for the "enhancer" variant, and demonstrate a significantly higher impact of it in male versus female patients. Integration of the RET RVs and CVs analysis showed that in 91.66% of cases with both kinds of mutational events, the enhancer allele is in trans with the allele bearing the RET RV. CONCLUSIONS: A gender effect exists on both the transmission and distribution of rare coding and common HSCR causing mutations. In addition, these RET CVs and RVs seem to act in a synergistic way leading to HSCR phenotype.


Assuntos
Doença de Hirschsprung/genética , Mutação , Proteínas Proto-Oncogênicas c-ret/genética , Alelos , Estudos de Casos e Controles , Análise Mutacional de DNA , Elementos Facilitadores Genéticos , Éxons , Feminino , Frequência do Gene , Estudos de Associação Genética , Teste de Complementação Genética , Variação Genética , Mutação em Linhagem Germinativa , Humanos , Íntrons , Masculino , Polimorfismo de Nucleotídeo Único , Caracteres Sexuais , Espanha
16.
PLoS One ; 6(8): e23475, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21858136

RESUMO

BACKGROUND: The enteric nervous system (ENS) is entirely derived from neural crest and its normal development is regulated by specific molecular pathways. Failure in complete ENS formation results in aganglionic gut conditions such as Hirschsprung's disease (HSCR). Recently, PROKR1 expression has been demonstrated in mouse enteric neural crest derived cells and Prok-1 was shown to work coordinately with GDNF in the development of the ENS. PRINCIPAL FINDINGS: In the present report, ENS progenitors were isolated and characterized from the ganglionic gut from children diagnosed with and without HSCR, and the expression of prokineticin receptors was examined. Immunocytochemical analysis of neurosphere-forming cells demonstrated that both PROKR1 and PROKR2 were present in human enteric neural crest cells. In addition, we also performed a mutational analysis of PROKR1, PROKR2, PROK1 and PROK2 genes in a cohort of HSCR patients, evaluating them for the first time as susceptibility genes for the disease. Several missense variants were detected, most of them affecting highly conserved amino acid residues of the protein and located in functional domains of both receptors, which suggests a possible deleterious effect in their biological function. CONCLUSIONS: Our results suggest that not only PROKR1, but also PROKR2 might mediate a complementary signalling to the RET/GFRα1/GDNF pathway supporting proliferation/survival and differentiation of precursor cells during ENS development. These findings, together with the detection of sequence variants in PROKR1, PROK1 and PROKR2 genes associated to HSCR and, in some cases in combination with RET or GDNF mutations, provide the first evidence to consider them as susceptibility genes for HSCR.


Assuntos
Doença de Hirschsprung/genética , Mutação , Células-Tronco Neurais/metabolismo , Receptores Acoplados a Proteínas-G/genética , Receptores de Peptídeos/genética , Células Cultivadas , Pré-Escolar , Estudos de Coortes , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/metabolismo , Feminino , Hormônios Gastrointestinais/genética , Hormônios Gastrointestinais/metabolismo , Expressão Gênica , Predisposição Genética para Doença/genética , Fator Neurotrófico Derivado de Linhagem de Célula Glial/genética , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/genética , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Doença de Hirschsprung/metabolismo , Doença de Hirschsprung/patologia , Humanos , Imuno-Histoquímica , Lactente , Recém-Nascido , Masculino , Microscopia Confocal , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Proteínas Proto-Oncogênicas c-ret/genética , Proteínas Proto-Oncogênicas c-ret/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Receptores de Peptídeos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Fator de Crescimento do Endotélio Vascular Derivado de Glândula Endócrina/genética , Fator de Crescimento do Endotélio Vascular Derivado de Glândula Endócrina/metabolismo
17.
BMC Med Genet ; 11: 137, 2010 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-20860806

RESUMO

BACKGROUND: Hirschsprung disease (HSCR) is a neurocristopathy characterized by the absence of parasympathetic intrinsic ganglion cells in the submucosal and myenteric plexuses along a variable portion of the intestinal tract. In approximately 18% of the cases HSCR also presents with multiple congenital anomalies including recognized syndromes. METHODS: A combination of MLPA and microarray data analysis have been undertaken to refine a duplication at the Xq28 region. RESULTS: In this study we present a new clinical association of severe neonatal encephalopathy (Lubs syndrome) and HSCR, in a male patient carrying a duplication at the Xq28 region which encompasses the MECP2 and L1CAM genes. CONCLUSIONS: While the encephalopathy has been traditionally attributed to the MECP2 gene duplication in patients with Lubs syndrome, here we propose that the enteric phenotype in our patient might be due to the dosage variation of the L1CAM protein, together with additional molecular events not identified yet. This would be in agreement with the hypothesis previously forwarded that mutations in L1CAM may be involved in HSCR development in association with a predisposing genetic background.


Assuntos
Encefalopatias/genética , Cromossomos Humanos X , Duplicação Gênica , Doença de Hirschsprung/genética , Encefalopatias/complicações , Encefalopatias/congênito , Análise Mutacional de DNA , Estudo de Associação Genômica Ampla , Doença de Hirschsprung/complicações , Humanos , Recém-Nascido , Masculino , Linhagem , Índice de Gravidade de Doença , Síndrome
18.
BMC Med Genet ; 11: 71, 2010 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-20459765

RESUMO

BACKGROUND: Hirschsprung disease is characterized by the absence of intramural ganglion cells in the enteric plexuses, due to a fail during enteric nervous system formation. Hirschsprung has a complex genetic aetiology and mutations in several genes have been related to the disease. There is a clear predominance of missense/nonsense mutations in these genes whereas copy number variations (CNVs) have been seldom described, probably due to the limitations of conventional techniques usually employed for mutational analysis. In this study, we have looked for CNVs in some of the genes related to Hirschsprung (EDNRB, GFRA1, NRTN and PHOX2B) using the Multiple Ligation-dependent Probe Amplification (MLPA) approach. METHODS: CNVs screening was performed in 208 HSCR patients using a self-designed set of MLPA probes, covering the coding region of those genes. RESULTS: A deletion comprising the first 4 exons in GFRA1 gene was detected in 2 sporadic HSCR patients and in silico approaches have shown that the critical translation initiation signal in the mutant gene was abolished. In this study, we have been able to validate the reliability of this technique for CNVs screening in HSCR. CONCLUSIONS: The implemented MLPA based technique presented here allows CNV analysis of genes involved in HSCR that have not been not previously evaluated. Our results indicate that CNVs could be implicated in the pathogenesis of HSCR, although they seem to be an uncommon molecular cause of HSCR.


Assuntos
Variações do Número de Cópias de DNA , Doença de Hirschsprung/genética , Técnicas de Amplificação de Ácido Nucleico , Sondas de DNA , Feminino , Humanos , Masculino
19.
BMC Med Genet ; 10: 119, 2009 Nov 19.
Artigo em Inglês | MEDLINE | ID: mdl-19925665

RESUMO

BACKGROUND: Hirschsprung disease (HSCR) is a congenital malformation of the hindgut produced by a disruption in neural crest cell migration during embryonic development. HSCR has a complex genetic etiology and mutations in several genes, mainly the RET proto-oncogene, have been related to the disease. There is a clear predominance of missense/nonsense mutations in these genes whereas copy number variations (CNVs) have been seldom described, probably due to the limitations of conventional techniques usually employed for mutational analysis. METHODS: In this study we have aimed to analyze the presence of CNVs in some HSCR genes (RET, EDN3, GDNF and ZFHX1B) using the Multiple Ligation-dependent Probe Amplification (MLPA) approach. RESULTS: Two alterations in the MLPA profiles of RET and EDN3 were detected, but a detailed inspection showed that the decrease in the corresponding dosages were due to point mutations affecting the hybridization probes regions. CONCLUSION: Our results indicate that CNVs of the gene coding regions analyzed here are not a common molecular cause of Hirschsprung disease. However, further studies are required to determine the presence of CNVs affecting non-coding regulatory regions, as well as other candidate genes.


Assuntos
Variações do Número de Cópias de DNA/genética , Doença de Hirschsprung/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Endotelina-3/genética , Feminino , Humanos , Masculino , Mutação , Proteínas Proto-Oncogênicas c-ret/genética , Sequências Reguladoras de Ácido Nucleico , Espanha
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