RESUMO
Our study characterized the neurodevelopmental spectrum of individuals with PTEN Hamartoma Tumor Syndrome (PHTS), a syndrome that predisposes to both neurodevelopmental phenotypes and cancer risk. We aim to better understand life-impacting neurodevelopmental features of PHTS. Our study recruited 20 children/adolescents with PHTS, who were then administered assessments for autism spectrum disorder (ASD) and other neurocognitive measures, including assessment of IQ, executive and adaptive functioning, and health-related quality of life. Thirteen individuals (65%) were identified as having ASD, of which five were newly diagnosed during the study. Of those, ASD symptom severity was in the mild-moderate range for 77%. Overall, IQ was in the average range, with a mean of 92.61 (SD 24.45, p = 0.5), though there was a non-statistically significant trend toward individuals without ASD having a higher mean IQ (102.7 vs 82.3; p = 0.1). Subjects had significant impairment in processing speed (mean 75.38, SD 24.75, p < 0.05), decreased adaptive functioning skills across all domains, and a trend toward having more executive functioning problems. Individuals with PHTS are at increased risk of neurodevelopmental disorders, including ASD and impaired executive and adaptive functioning. Although clear guidelines exist for cancer surveillance for individuals with PHTS, additional guidelines and screening for neurodevelopmental disorders are warranted.
Assuntos
Transtorno do Espectro Autista , Síndrome do Hamartoma Múltiplo , PTEN Fosfo-Hidrolase , Fenótipo , Humanos , Masculino , Feminino , Criança , Adolescente , Síndrome do Hamartoma Múltiplo/genética , Síndrome do Hamartoma Múltiplo/patologia , Síndrome do Hamartoma Múltiplo/diagnóstico , Transtorno do Espectro Autista/genética , Transtorno do Espectro Autista/patologia , Transtorno do Espectro Autista/epidemiologia , Transtorno do Espectro Autista/diagnóstico , PTEN Fosfo-Hidrolase/genética , Pré-Escolar , Qualidade de VidaRESUMO
Peutz-Jeghers syndrome (PJS) is a childhood-onset cancer predisposition syndrome that is associated with oral freckling and gastrointestinal polyposis. Male patients with PJS are at risk for large-cell calcifying Sertoli cell tumors in childhood. These tumors are estrogen-producing and can cause symptoms of precocious puberty, gynecomastia, and growth acceleration. Here we discuss our experience with spontaneous resolution or stabilization of breast enlargement without medical intervention in three patients with PJS and gynecomastia. These cases indicate that a watchful waiting approach can be considered in the management of gynecomastia in male children with PJS.
Assuntos
Ginecomastia , Síndrome de Peutz-Jeghers , Humanos , Síndrome de Peutz-Jeghers/complicações , Síndrome de Peutz-Jeghers/terapia , Síndrome de Peutz-Jeghers/patologia , Síndrome de Peutz-Jeghers/genética , Ginecomastia/terapia , Ginecomastia/etiologia , Masculino , Criança , Tratamento Conservador , Adolescente , Pré-EscolarRESUMO
High-risk solid tumors continue to pose a tremendous therapeutic challenge due to multidrug resistance. Biological mechanisms driving chemoresistance in high-risk primary and recurrent disease are distinct: in newly diagnosed patients, non-response to therapy is often associated with a higher level of tumor "stemness" paralleled by overexpression of the ABCG2 drug efflux pump, whereas in tumors relapsing after non-curative therapy, poor drug sensitivity is most commonly linked to the dysfunction of the tumor suppressor protein, p53. In this study, we used preclinical models of aggressive neuroblastoma featuring these characteristic mechanisms of primary and acquired drug resistance to experimentally evaluate a macromolecular prodrug of a structurally enhanced camptothecin analog, SN22, resisting ABCG2-mediated export, and glucuronidation. Together with extended tumor exposure to therapeutically effective drug levels via reversible conjugation to Pluronic F-108 (PF108), these features translated into rapid tumor regression and long-term survival in models of both ABCG2-overexpressing and p53-mutant high-risk neuroblastomas, in contrast to a marginal effect of the clinically used camptothecin derivative, irinotecan. Our results demonstrate that pharmacophore enhancement, increased tumor uptake, and optimally stable carrier-drug association integrated into the design of the hydrolytically activatable PF108-[SN22]2 have the potential to effectively combat multiple mechanisms governing chemoresistance in newly diagnosed (chemo-naïve) and recurrent forms of aggressive malignancies. As a macromolecular carrier-based delivery system exhibiting remarkable efficacy against two particularly challenging forms of high-risk neuroblastoma, PF108-[SN22]2 can pave the way to a robust and clinically viable therapeutic strategy urgently needed for patients with multidrug-resistant disease presently lacking effective treatment options.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Neuroblastoma/tratamento farmacológico , Pró-Fármacos/uso terapêutico , Inibidores da Topoisomerase I/uso terapêutico , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Camptotecina/análogos & derivados , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos Nus , Camundongos SCID , Poloxâmero/química , Pró-Fármacos/administração & dosagem , Pró-Fármacos/química , Inibidores da Topoisomerase I/químicaRESUMO
Juvenile polyposis syndrome (JPS) is a childhood polyposis syndrome with up to a 50% lifetime risk of gastrointestinal cancer. Germline pathogenic variants in BMPR1A and SMAD4 are responsible for around 40% of cases of JPS, but for the majority of individuals, the underlying genetic cause is unknown. We identified a family for which polyposis spanned four generations, and the proband had a clinical diagnosis of JPS. Next-generation sequencing was conducted, followed by Sanger sequencing confirmation. We identified an internal deletion of the FOCAD gene in all family members tested that altered the reading frame and is predicted to be pathogenic. We conclude that inactivation of the FOCAD gene is likely to cause JPS in this family.
Assuntos
Polipose Intestinal , Síndromes Neoplásicas Hereditárias , Criança , Neoplasias Gastrointestinais , Mutação em Linhagem Germinativa , Humanos , Polipose Intestinal/congênito , Polipose Intestinal/genética , Síndromes Neoplásicas Hereditárias/genéticaRESUMO
Despite the use of intensive multimodality therapy, the majority of high-risk neuroblastoma (NB) patients do not survive. Without significant improvements in delivery strategies, anticancer agents used as a first-line treatment for high-risk tumors often fail to provide clinically meaningful results in the settings of disseminated, recurrent, or refractory disease. By enhancing pharmacological selectivity, favorably shifting biodistribution, strengthening tumor cell killing potency, and overcoming drug resistance, nanocarrier-mediated delivery of topoisomerase I inhibitors of the camptothecin family has the potential to dramatically improve treatment efficacy and minimize side effects. In this study, a structurally enhanced camptothecin analog, SN22, reversibly coupled with a redox-silent tocol derivative (tocopheryl oxamate) to allow its optimally stable encapsulation and controlled release from PEGylated sub-100 nm nanoparticles (NP), exhibited strong NB cell growth inhibitory activity, translating into rapid regression and durably suppressed regrowth of orthotopic, MYCN-amplified NB tumors. The robust antitumor effects and markedly extended survival achieved in preclinical models recapitulating different phases of high-risk disease (at diagnosis vs. at relapse with an acquired loss of p53 function after intensive multiagent chemotherapy) demonstrate remarkable potential of SN22 delivered in the form of a hydrolytically cleavable superhydrophobic prodrug encapsulated in biodegradable nanocarriers as an experimental strategy for treating refractory solid tumors in high-risk cancer patients.
Assuntos
Camptotecina/análogos & derivados , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos , Nanopartículas/química , Neuroblastoma/tratamento farmacológico , Pró-Fármacos/uso terapêutico , Tocoferóis/uso terapêutico , Camptotecina/química , Camptotecina/farmacologia , Camptotecina/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Neuroblastoma/patologia , Fatores de Risco , Análise de Sobrevida , Tocoferóis/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: MYCN amplification (MYCN-A) is an established adverse prognostic factor in neuroblastoma. The extent to which the prognostic impact of MYCN-A depends on other factors has not been fully characterized. PATIENTS AND METHODS: Using the International Neuroblastoma Risk Group database, we constructed Cox models of overall survival (OS) to obtain hazard ratios of the effect of MYCN-A within subgroups defined by other prognostic factors. Cox models assessed the degree to which the prognostic impact of MYCN-A was modulated by each other covariate. We used absolute hazard ratio (HR) differences to construct classification trees to identify subgroups with greatest differential prognostic effect of MYCN-A. RESULTS: In a cohort of 6223 patients with known MYCN status, the OS hazard ratio associated with MYCN-A was 6.3 (95% confidence interval 5.7-7.0, P < .001). Age at diagnosis conferred the largest HR absolute difference for MYCN-A between subgroups (HR absolute difference 16.6; HRs for MYCN-A of 19.6 for <18 months, 3.0 for ≥18 months). MYCN-A remained significantly prognostic of OS after controlling for other factors, abrogating their prognostic strength. Patients whose outcome was most impacted by MYCN status were those who were <18 months, had high mitosis karrhyohexis index (MKI) and low ferritin. CONCLUSION: The prognostic strength of MYCN-A varies depending on which patient subgroup defined by other neuroblastoma risk factors is examined, with greatest strength in patients with otherwise favorable features. MYCN-A has little effect within some subgroups, aiding clinical decision-making if MYCN status cannot be assessed. Subgroups where MYCN-A has large effect may be prioritized for agents targeting Myc family proteins.
Assuntos
Biomarcadores Tumorais/análise , Neoplasias da Medula Óssea/secundário , Neoplasias Ósseas/secundário , Amplificação de Genes , Proteína Proto-Oncogênica N-Myc/genética , Neuroblastoma/patologia , Neoplasias da Medula Óssea/genética , Neoplasias da Medula Óssea/terapia , Neoplasias Ósseas/genética , Neoplasias Ósseas/terapia , Estudos de Coortes , Feminino , Ferritinas/sangue , Seguimentos , Humanos , Lactente , Masculino , Mitose , Neuroblastoma/genética , Neuroblastoma/terapia , Prognóstico , Fatores de Risco , Taxa de SobrevidaRESUMO
Gastrointestinal polyps are mucosal overgrowths that, if unchecked, can undergo malignant transformation. Although relatively uncommon in the pediatric age group, they can be the harbingers of multiorgan cancer risk and require close management and follow-up. Additionally, as many polyposis syndromes are inherited, appropriate genetic testing and management of relatives is vital for the health of the entire family. In this review, we discuss both common and uncommon childhood gastrointestinal polyposis syndromes in terms of clinical presentation, management, and surveillance. We also detail any additional malignancy risk and surveillance required in the pediatric age group (<21 years old). Through this review, we provide a framework for gastroenterologists to manage the multifaceted nature of pediatric polyposis syndromes.
Assuntos
Polipose Adenomatosa do Colo/genética , Neoplasias Gastrointestinais/genética , Predisposição Genética para Doença , Síndrome de Peutz-Jeghers/genética , Adolescente , Criança , Aconselhamento Genético , Testes Genéticos , Humanos , Guias de Prática Clínica como AssuntoRESUMO
Neuroblastomas are characterized by heterogeneous clinical behavior, from spontaneous regression or differentiation into a benign ganglioneuroma, to relentless progression despite aggressive, multimodality therapy. Indeed, neuroblastoma is unique among human cancers in terms of its propensity to undergo spontaneous regression. The strongest evidence for this comes from the mass screening studies conducted in Japan, North America and Europe and it is most evident in infants with stage 4S disease. This propensity is associated with a pattern of genomic change characterized by whole chromosome gains rather than segmental chromosome changes but the mechanism(s) underlying spontaneous regression are currently a matter of speculation. There is evidence to support several possible mechanisms of spontaneous regression in neuroblastomas: (1) neurotrophin deprivation, (2) loss of telomerase activity, (3) humoral or cellular immunity and (4) alterations in epigenetic regulation and possibly other mechanisms. It is likely that a better understanding of the mechanisms of spontaneous regression will help to identify targeted therapeutic approaches for these tumors. The most easily targeted mechanism is the delayed activation of developmentally programmed cell death regulated by the tropomyosin receptor kinase A (TrkA) pathway. Pan-Trk inhibitors are currently in clinical trials and so Trk inhibition might be used as the first line of therapy in infants with biologically favorable tumors that require treatment. Alternative approaches consist of breaking immune tolerance to tumor antigens but approaches to telomere shortening or epigenetic regulation are not easily druggable. The different mechanisms of spontaneous neuroblastoma regression are reviewed here, along with possible therapeutic approaches.
Assuntos
Neuroblastoma/terapia , Detecção Precoce de Câncer , Genômica , Humanos , Imunoterapia , Neuroblastoma/diagnóstico , Neuroblastoma/genética , Neuroblastoma/patologia , Indução de RemissãoRESUMO
Beckwith-Wiedemann syndrome (BWS) is a genetic syndrome associated with overgrowth and cancer predisposition, including predisposition to Wilms tumor (WT). Patients with BWS and BWS spectrum are screened from birth to age 7 years for BWS-associated cancers. However, in some cases a BWS-associated cancer may be the first recognized manifestation of the syndrome. We describe 12 patients diagnosed with BWS after presenting with a WT. We discuss the features of BWS in these patients and hypothesize that earlier detection of BWS by attention to its subtler manifestations could lead to earlier detection of children at risk for associated malignancies.
Assuntos
Síndrome de Beckwith-Wiedemann/complicações , Síndrome de Beckwith-Wiedemann/diagnóstico , Neoplasias Renais/genética , Tumor de Wilms/genética , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , MasculinoRESUMO
Neuroblastoma is a childhood cancer derived from cells of neural crest origin. The hallmarks of its enigmatic character include its propensity for spontaneous regression under some circumstances and its association with paraneoplastic opsoclonus, myoclonus, and ataxia. The neurodevelopmental underpinnings of its origins may provide important clues for development of novel therapeutic and preventive agents for this frequently fatal malignancy and for the associated paraneoplastic syndromes. Ann Neurol 2016;80:13-23.
Assuntos
Crista Neural/patologia , Neuroblastoma/etiologia , Transtornos do Neurodesenvolvimento/etiologia , Progressão da Doença , Humanos , Terapia de Alvo Molecular , Regressão Neoplásica Espontânea , Crista Neural/crescimento & desenvolvimento , Neuroblastoma/complicações , Neuroblastoma/tratamento farmacológico , Neuroblastoma/patologia , Transtornos do Neurodesenvolvimento/complicações , Transtornos do Neurodesenvolvimento/patologia , Síndrome de Opsoclonia-Mioclonia/imunologia , Síndromes Paraneoplásicas do Sistema Nervoso/complicações , Síndromes Paraneoplásicas do Sistema Nervoso/etiologiaAssuntos
Síndrome do Hamartoma Múltiplo , Hemangiossarcoma , Neuroblastoma , Neoplasias Cutâneas , Criança , Síndrome do Hamartoma Múltiplo/complicações , Síndrome do Hamartoma Múltiplo/genética , Síndrome do Hamartoma Múltiplo/patologia , Humanos , Neuroblastoma/genética , PTEN Fosfo-Hidrolase/genética , Neoplasias Cutâneas/genéticaRESUMO
Beckwith-Wiedemann syndrome (BWS) is a genetic overgrowth and cancer predisposition syndrome, associated with both benign and malignant adrenal findings. Literature review and an institutional case series elucidate the wide spectrum of adrenal findings in BWS patients. The altered expression of the 11p15 region is likely related to adrenal gland hyperplasia and growth dysregulation. Given the absence of guidelines for managing adrenal findings in BWS, we propose a systematic approach to adrenal findings in BWS patients, to allow for maximum detection of potentially malignant pathology without posing additional risk to patients.
Assuntos
Doenças das Glândulas Suprarrenais/diagnóstico , Doenças das Glândulas Suprarrenais/etiologia , Doenças das Glândulas Suprarrenais/terapia , Síndrome de Beckwith-Wiedemann/complicações , HumanosRESUMO
BACKGROUND: MYCN gene amplification (MNA) is a hallmark of aggressive neuroblastoma. This study was performed to determine univariate and multivariate predictors of tumor MNA. METHODS: Data from the International Neuroblastoma Risk Group were analyzed for a subset of 7102 patients with known MYCN status. Chi-square testing and logistic regression were used to identify univariate and multivariate predictors of MYCN status. Recursive partitioning was used to identify groups of patients with maximal differences in rates of MNA. RESULTS: All clinical features (age ≥ 18 months, high ferritin levels, high lactate dehydrogenase [LDH] levels, International Neuroblastoma Staging System stage 4, and adrenal sites) and pathological/biological features (DNA index ≤ 1, high mitosis-karyorrhexis index [MKI], undifferentiated/poorly differentiated grade, unfavorable histology according to the International Neuroblastoma Pathology Classification, and segmental chromosomal aberrations [SCAs]) were significantly associated with MNA. LDH (odds ratio [OR], 8.4; P < .001) and chromosomal 1p loss of heterozygosity (OR, 19.8; P < .001) were the clinical and biological variables, respectively, most strongly associated with MNA. In logistic regression, all variables except chromosome 17q aberration and pooled SCAs were independently predictive of MNA. Recursive partitioning identified subgroups with disparate rates of MNA, including subgroups with 85.7% MNA (patients with high LDH levels who had poorly differentiated adrenal tumors with chromosome 1p deletion) and 0.6% MNA (localized tumors having hyperdiploidy and low MKIs and lacking chromosome 1p aberrations). CONCLUSIONS: MNA is strongly associated with other clinical and biological variables in neuroblastoma. Recursive partitioning has identified subgroups of neuroblastoma patients with highly disparate rates of MNA. These findings can be used to inform investigations of molecular mechanisms of MNA.
Assuntos
Amplificação de Genes , Neuroblastoma/diagnóstico , Neuroblastoma/genética , Proteínas Nucleares/genética , Proteínas Oncogênicas/genética , Fatores Etários , Aberrações Cromossômicas , Feminino , Humanos , Lactente , Modelos Logísticos , Perda de Heterozigosidade , Masculino , Proteína Proto-Oncogênica N-Myc , Gradação de Tumores , Estadiamento de Neoplasias , Neuroblastoma/patologia , Neuroblastoma/secundário , Valor Preditivo dos Testes , PrognósticoRESUMO
Eukaryotic gene expression is developmentally regulated, in part by chromatin remodelling, and its dysregulation has been linked to cancer. CHD5 (chromodomain helicase DNA-binding protein 5) is a tumour suppressor gene (TSG) that maps to a region of consistent deletion on 1p36.31 in neuroblastomas (NBs) and other tumour types. CHD5 encodes a protein with chromatin remodelling, helicase and DNA-binding motifs that is preferentially expressed in neural and testicular tissues. CHD5 is highly homologous to CHD3 and CHD4, which are the core subunits of nucleosome remodelling and deacetylation (NuRD) complexes. To determine if CHD5 forms a similar complex, we performed studies on nuclear extracts from NBLS, SY5Y (both with endogenous CHD5 expression), NLF (CHD5 null) and NLF cells stably transfected with CHD5 cDNA (wild-type and V5-histidine-tagged). Immunoprecipitation (IP) was performed with either CHD5 antibody or antibody to V5/histidine-tagged protein. We identified NuRD components both by GST-FOG1 (Friend Of GATA1) pull-down and by IP. We also performed MS/MS analysis to confirm the presence of CHD5 or other protein components of the NuRD complex, as well as to identify other novel proteins. CHD5 was clearly associated with all canonical NuRD components, including metastasis-associated protein (MTA)1/2, GATA zinc finger domain containing 2A (GATAD2A), histone deacetylase (HDAC)1/2, retinoblastoma-binding protein (RBBP)4/7 and methyl DNA-binding domain protein (MBD)2/3, as determined by Western blotting and MS/MS. Our data suggest CHD5 forms a NuRD complex similar to CHD4. However, CHD5-NuRD may also have unique protein associations that confer functional specificity and may contribute to normal development and to tumour suppression in NB and other cancers.
Assuntos
Antígenos CD/metabolismo , Caderinas/metabolismo , Montagem e Desmontagem da Cromatina , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/metabolismo , Neuroblastoma/metabolismo , Nucleossomos/metabolismo , Western Blotting , Núcleo Celular/metabolismo , Cromatografia Líquida , Imunofluorescência , Humanos , Técnicas Imunoenzimáticas , Imunoprecipitação , Neuroblastoma/patologia , Espectrometria de Massas em Tandem , Células Tumorais CultivadasRESUMO
BACKGROUND: Chromodomain-helicase DNA binding protein 5 (CHD5) is an important tumor suppressor gene deleted from 1p36.31 in neuroblastomas (NBs). High CHD5 expression is associated with a favorable prognosis, but deletion or low expression is frequent in high-risk tumors. We explored the role of CHD5 expression in the neuronal differentiation of NB cell lines. METHODS: NB cell lines SH-SY5Y (SY5Y), NGP, SK-N-DZ, IMR5, LAN5, SK-N-FI, NB69 and SH-EP were treated with 1-10 µM 13-cis-retinoic acid (13cRA) for 3-12 days. qRT-PCR and Western blot analyses were performed to measure mRNA and protein expression levels, respectively. Morphological differences were examined by both phase contrast and immunofluorescence studies. RESULTS: Treatment of SY5Y cells with 13cRA caused upregulation of CHD5 expression in a time- and dose-dependent manner (1, 5, or 10 µM for 7 or 12 days) and also induced neuronal differentiation. Furthermore, both NGP and SK-N-DZ cells showed CHD5 upregulation and neuronal differentiation after 13cRA treatment. In contrast, 13cRA treatment of IMR5, LAN5, or SK-N-FI induced neither CHD5 expression nor neuronal differentiation. NB69 cells showed two different morphologies (neuronal and substrate adherent) after 12 days treatment with 10 µM of 13cRA. CHD5 expression was high in the neuronal cells, but low/absent in the flat, substrate adherent cells. Finally, NGF treatment caused upregulation of CHD5 expression and neuronal differentiation in SY5Y cells transfected to express TrkA (SY5Y-TrkA) but not in TrkA-null parental SY5Y cells, and both changes were blocked by a pan-TRK inhibitor. CONCLUSIONS: Treatment with 13cRA induces neuronal differentiation only in NB cells that upregulate CHD5. In addition, NGF induced CHD5 upregulation and neuronal differentiation only in TrkA expressing cells. Together, these results suggest that CHD5 is downstream of TrkA, and CHD5 expression may be crucial for neuronal differentiation induced by either 13cRA or TrkA/NGF signaling.
Assuntos
DNA Helicases/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas do Tecido Nervoso/genética , Neuroblastoma/genética , Tretinoína/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , DNA Helicases/metabolismo , Humanos , Fator de Crescimento Neural/farmacologia , Proteínas do Tecido Nervoso/metabolismo , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Receptor trkA/genética , Receptor trkA/metabolismo , Regulação para CimaAssuntos
Neoplasias Colorretais/diagnóstico , Detecção Precoce de Câncer/métodos , Síndrome de Li-Fraumeni/diagnóstico , Adolescente , Adulto , Idade de Início , Neoplasias Colorretais/epidemiologia , Neoplasias Colorretais/genética , Bases de Dados Factuais , Predisposição Genética para Doença , Humanos , Incidência , Síndrome de Li-Fraumeni/epidemiologia , Síndrome de Li-Fraumeni/genética , Masculino , Pessoa de Meia-Idade , Mutação , Fenótipo , Philadelphia/epidemiologia , Valor Preditivo dos Testes , Estudos Retrospectivos , Fatores de Risco , Fatores de Tempo , Proteína Supressora de Tumor p53/genética , Adulto JovemRESUMO
Congenital heart defects (CHDs) are the most common major birth defects and the leading cause of death from congenital malformations. The etiology remains largely unknown, though genetic variants clearly contribute. In a previous study, we identified a large copy-number variant (CNV) that deleted 46 genes in a patient with a malalignment type ventricular septal defect (VSD). The CNV included the gene NTRK3 encoding neurotrophic tyrosine kinase receptor C (TrkC), which is essential for normal cardiogenesis in animal models. To evaluate the role of NTRK3 in human CHDs, we studied 467 patients with related heart defects for NTRK3 mutations. We identified four missense mutations in four patients with VSDs that were not found in ethnically matched controls and were predicted to be functionally deleterious. Functional analysis using neuroblastoma cell lines expressing mutant TrkC demonstrated that one of the mutations (c.278C>T, p.T93M) significantly reduced autophosphorylation of TrkC in response to ligand binding, subsequently decreasing phosphorylation of downstream target proteins. In addition, compared with wild type, three of the four cell lines expressing mutant TrkC showed altered cell growth in low-serum conditions without supplemental neurotrophin 3. These findings suggest a novel pathophysiological mechanism involving NTRK3 in the development of VSDs.
Assuntos
Cardiopatias Congênitas/genética , Mutação de Sentido Incorreto , Receptor trkC/genética , Transdução de Sinais , Sequência de Aminoácidos , Western Blotting , Linhagem Celular , Estudos de Coortes , Humanos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Fosforilação , Receptor trkC/química , Receptor trkC/metabolismo , Homologia de Sequência de AminoácidosRESUMO
BACKGROUND: Chromodomain helicase DNA binding protein 5 (CHD5) is a family member of chromatin remodeling factors. The epigenetic silencing mechanisms of CHD5 in colorectal cancer have not been well studied. METHODS: Here we analyzed CHD5 methylation and mRNA expression in vitro and in clinical samples from African American patients. DNA and RNA were isolated from formalin fixed paraffin embedded (FFPE) colon tissues. DNA was tested for methylation using methylation-specific polymerase chain reation (PCR) and bisulfite sequencing. RNA was used for mRNA quantification using qRT-PCR. The RKO cell line was treated with 5-Aza-dC and SAHA. RKO cells were also stably transfected with a CHD5-expressing vector. The transcriptional activity was studied in the 1 kb upstream region of the CHD5 promoter using the dual reporter assay. We performed cell proliferation, migration, and invasion assays using the RKO cell line. RESULTS: In most adenoma samples, CHD5 expression was not detected in contrast to normal tissues. In RKO cells, CHD5 silencing was associated with DNA methylation and repressive histone modifications. CHD5 expression was restored after treatment with 5-Aza-dC and SAHA. CHD5 reactivation reduced cell proliferation, migration, and invasion. The reporter assay indicated that the main regulatory region of the CHD5 promoter is encompassed in the -489 to -823 region with important transcriptional regulatory sites (TCF/LEF, SP1, and AP-2). CONCLUSIONS: The CHD5 gene is repressed in all types of adenomas, either epigenetically or by chromosomal deletion. CHD5 activity is regulated by DNA methylation and repressive histone modifications. CHD5 likely acts as a tumor-suppressor gene in early colorectal carcinogenesis.