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1.
PLoS Comput Biol ; 15(7): e1007095, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31329578

RESUMO

Alternative transcript isoforms are common in tumors and act as potential drivers of cancer. Mechanisms determining altered isoform expression include somatic mutations in splice regulatory sites or altered splicing factors. However, since DNA methylation is known to regulate transcriptional isoform activity in normal cells, we predicted the highly dysregulated patterns of DNA methylation present in cancer also affect isoform activity. We analyzed DNA methylation and RNA-seq isoform data from 18 human cancer types and found frequent correlations specifically within 11 cancer types. Examining the top 25% of variable methylation sites revealed that the location of the methylated CpG site in a gene determined which isoform was used. In addition, the correlated methylation-isoform patterns classified tumors into known subtypes and predicted distinct protein functions between tumor subtypes. Finally, methylation-correlated isoforms were enriched for oncogenes, tumor suppressors, and cancer-related pathways. These findings provide new insights into the functional impact of dysregulated DNA methylation in cancer and highlight the relationship between the epigenome and transcriptome.

2.
Nat Immunol ; 19(12): 1403-1414, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30397350

RESUMO

Repair of tissue damaged during inflammatory processes is key to the return of local homeostasis and restoration of epithelial integrity. Here we describe CD161+ regulatory T (Treg) cells as a distinct, highly suppressive population of Treg cells that mediate wound healing. These Treg cells were enriched in intestinal lamina propria, particularly in Crohn's disease. CD161+ Treg cells had an all-trans retinoic acid (ATRA)-regulated gene signature, and CD161 expression on Treg cells was induced by ATRA, which directly regulated the CD161 gene. CD161 was co-stimulatory, and ligation with the T cell antigen receptor induced cytokines that accelerated the wound healing of intestinal epithelial cells. We identified a transcription-factor network, including BACH2, RORγt, FOSL2, AP-1 and RUNX1, that controlled expression of the wound-healing program, and found a CD161+ Treg cell signature in Crohn's disease mucosa associated with reduced inflammation. These findings identify CD161+ Treg cells as a population involved in controlling the balance between inflammation and epithelial barrier healing in the gut.

3.
Atherosclerosis ; 278: 278-285, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30347343

RESUMO

BACKGROUND AND AIMS: High density lipoprotein cholesterol (HDL-C) is associated with risk of cardiovascular disease (CVD); however, therapeutic manipulations of HDL-C have failed to reduce CVD events. This suggests that HDL-C and the atheroprotective capacity of HDL are not directly linked. The goal of this study was to evaluate the relationships between HDL-bound proteins and measures of atherosclerosis burden and HDL function. METHODS: The HDL proteome was analyzed using mass spectrometry in 126 human subjects, who had undergone coronary computed tomography angiography (CCTA) to quantify calcified (CB) and non-calcified (NCB) atherosclerosis burden. Partial least squares regression analysis was used to evaluate associations between HDL-bound proteins and CB, NCB, or cholesterol efflux capacity (CEC). RESULTS: Significant overlap was found among proteins associated with NCB and CEC. Proteins that were associated with NCB displayed an inverse relationship with CEC, supporting a link between this protective function of HDL and clinical plaque burden. CB was associated with a set of proteins mostly distinct from NCB and CEC. When CVD risk factors were evaluated, BMI had a stronger influence on important HDL proteins than gender, age, or HDL-C. Most HDL proteins associated with function or atherosclerosis burden were not significantly correlated with HDL-C. CONCLUSIONS: These findings indicate that the HDL proteome contains information not captured by HDL- C and, therefore, has potential for future development as a biomarker for CVD risk. Additionally, the proteome effects detected in this study may provide HDL compositional goals for evaluating new and existing HDL-modification therapies.

4.
PLoS Comput Biol ; 13(11): e1005840, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29125844

RESUMO

Recent evidence shows that mutations in several driver genes can cause aberrant methylation patterns, a hallmark of cancer. In light of these findings, we hypothesized that the landscapes of tumor genomes and epigenomes are tightly interconnected. We measured this relationship using principal component analyses and methylation-mutation associations applied at the nucleotide level and with respect to genome-wide trends. We found that a few mutated driver genes were associated with genome-wide patterns of aberrant hypomethylation or CpG island hypermethylation in specific cancer types. In addition, we identified associations between 737 mutated driver genes and site-specific methylation changes. Moreover, using these mutation-methylation associations, we were able to distinguish between two uterine and two thyroid cancer subtypes. The driver gene mutation-associated methylation differences between the thyroid cancer subtypes were linked to differential gene expression in JAK-STAT signaling, NADPH oxidation, and other cancer-related pathways. These results establish that driver gene mutations are associated with methylation alterations capable of shaping regulatory network functions. In addition, the methodology presented here can be used to subdivide tumors into more homogeneous subsets corresponding to underlying molecular characteristics, which could improve treatment efficacy.


Assuntos
Metilação de DNA/genética , Mutação/genética , Neoplasias/genética , Transdução de Sinais/genética , Biologia Computacional , Ilhas de CpG/genética , Estudos de Associação Genética , Genoma/genética , Humanos , Análise de Componente Principal
5.
Hum Mutat ; 38(9): 1266-1276, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28544481

RESUMO

The advent of next-generation sequencing has dramatically decreased the cost for whole-genome sequencing and increased the viability for its application in research and clinical care. The Personal Genome Project (PGP) provides unrestricted access to genomes of individuals and their associated phenotypes. This resource enabled the Critical Assessment of Genome Interpretation (CAGI) to create a community challenge to assess the bioinformatics community's ability to predict traits from whole genomes. In the CAGI PGP challenge, researchers were asked to predict whether an individual had a particular trait or profile based on their whole genome. Several approaches were used to assess submissions, including ROC AUC (area under receiver operating characteristic curve), probability rankings, the number of correct predictions, and statistical significance simulations. Overall, we found that prediction of individual traits is difficult, relying on a strong knowledge of trait frequency within the general population, whereas matching genomes to trait profiles relies heavily upon a small number of common traits including ancestry, blood type, and eye color. When a rare genetic disorder is present, profiles can be matched when one or more pathogenic variants are identified. Prediction accuracy has improved substantially over the last 6 years due to improved methodology and a better understanding of features.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequenciamento Completo do Genoma/métodos , Área Sob a Curva , Predisposição Genética para Doença , Projeto Genoma Humano , Humanos , Fenótipo , Locos de Características Quantitativas
6.
World J Gastrointest Oncol ; 9(3): 105-120, 2017 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-28344746

RESUMO

Over the last two decades, cancer-related alterations in DNA methylation that regulate transcription have been reported for a variety of tumors of the gastrointestinal tract. Due to its relevance for translational research, great emphasis has been placed on the analysis and molecular characterization of the CpG island methylator phenotype (CIMP), defined as widespread hypermethylation of CpG islands in clinically distinct subsets of cancer patients. Here, we present an overview of previous work in this field and also explore some open questions using cross-platform data for esophageal, gastric, and colorectal adenocarcinomas from The Cancer Genome Atlas. We provide a data-driven, pan-gastrointestinal stratification of individual samples based on CIMP status and we investigate correlations with oncogenic alterations, including somatic mutations and epigenetic silencing of tumor suppressor genes. Besides known events in CIMP such as BRAF V600E mutation, CDKN2A silencing or MLH1 inactivation, we discuss the potential role of emerging actors such as Wnt pathway deregulation through truncating mutations in RNF43 and epigenetic silencing of WIF1. Our results highlight the existence of molecular similarities that are superimposed over a larger backbone of tissue-specific features and can be exploited to reduce heterogeneity of response in clinical trials.

7.
JAMA Psychiatry ; 73(6): 590-7, 2016 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-27120077

RESUMO

IMPORTANCE: Complex disorders, such as bipolar disorder (BD), likely result from the influence of both common and rare susceptibility alleles. While common variation has been widely studied, rare variant discovery has only recently become feasible with next-generation sequencing. OBJECTIVE: To utilize a combined family-based and case-control approach to exome sequencing in BD using multiplex families as an initial discovery strategy, followed by association testing in a large case-control meta-analysis. DESIGN, SETTING, AND PARTICIPANTS: We performed exome sequencing of 36 affected members with BD from 8 multiplex families and tested rare, segregating variants in 3 independent case-control samples consisting of 3541 BD cases and 4774 controls. MAIN OUTCOMES AND MEASURES: We used penalized logistic regression and 1-sided gene-burden analyses to test for association of rare, segregating damaging variants with BD. Permutation-based analyses were performed to test for overall enrichment with previously identified gene sets. RESULTS: We found 84 rare (frequency <1%), segregating variants that were bioinformatically predicted to be damaging. These variants were found in 82 genes that were enriched for gene sets previously identified in de novo studies of autism (19 observed vs. 10.9 expected, P = .0066) and schizophrenia (11 observed vs. 5.1 expected, P = .0062) and for targets of the fragile X mental retardation protein (FMRP) pathway (10 observed vs. 4.4 expected, P = .0076). The case-control meta-analyses yielded 19 genes that were nominally associated with BD based either on individual variants or a gene-burden approach. Although no gene was individually significant after correction for multiple testing, this group of genes continued to show evidence for significant enrichment of de novo autism genes (6 observed vs 2.6 expected, P = .028). CONCLUSIONS AND RELEVANCE: Our results are consistent with the presence of prominent locus and allelic heterogeneity in BD and suggest that very large samples will be required to definitively identify individual rare variants or genes conferring risk for this disorder. However, we also identify significant associations with gene sets composed of previously discovered de novo variants in autism and schizophrenia, as well as targets of the FRMP pathway, providing preliminary support for the overlap of potential autism and schizophrenia risk genes with rare, segregating variants in families with BD.


Assuntos
Transtorno Bipolar/genética , Exoma/genética , Análise de Sequência de DNA , Alelos , Transtorno Autístico/genética , Transtorno Autístico/psicologia , Transtorno Bipolar/diagnóstico , Transtorno Bipolar/psicologia , Estudos de Casos e Controles , Proteína do X Frágil de Retardo Mental/genética , Heterogeneidade Genética , Predisposição Genética para Doença/genética , Variação Genética/genética , Estudo de Associação Genômica Ampla , Humanos , Esquizofrenia/genética , Psicologia do Esquizofrênico
8.
Cancer Discov ; 6(2): 166-75, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26658419

RESUMO

UNLABELLED: Pancreatic cancer is projected to become the second leading cause of cancer-related death in the United States by 2020. A familial aggregation of pancreatic cancer has been established, but the cause of this aggregation in most families is unknown. To determine the genetic basis of susceptibility in these families, we sequenced the germline genomes of 638 patients with familial pancreatic cancer and the tumor exomes of 39 familial pancreatic adenocarcinomas. Our analyses support the role of previously identified familial pancreatic cancer susceptibility genes such as BRCA2, CDKN2A, and ATM, and identify novel candidate genes harboring rare, deleterious germline variants for further characterization. We also show how somatic point mutations that occur during hematopoiesis can affect the interpretation of genome-wide studies of hereditary traits. Our observations have important implications for the etiology of pancreatic cancer and for the identification of susceptibility genes in other common cancer types. SIGNIFICANCE: The genetic basis of disease susceptibility in the majority of patients with familial pancreatic cancer is unknown. We whole genome sequenced 638 patients with familial pancreatic cancer and demonstrate that the genetic underpinning of inherited pancreatic cancer is highly heterogeneous. This has significant implications for the management of patients with familial pancreatic cancer.


Assuntos
Carcinoma/genética , Predisposição Genética para Doença , Mutação em Linhagem Germinativa , Neoplasias Pancreáticas/genética , Análise de Sequência de DNA/métodos , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteína BRCA2/genética , Inibidor p16 de Quinase Dependente de Ciclina/genética , Humanos , Mutação Puntual
9.
PLoS Comput Biol ; 10(9): e1003825, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25188385

RESUMO

Genetic screening is becoming possible on an unprecedented scale. However, its utility remains controversial. Although most variant genotypes cannot be easily interpreted, many individuals nevertheless attempt to interpret their genetic information. Initiatives such as the Personal Genome Project (PGP) and Illumina's Understand Your Genome are sequencing thousands of adults, collecting phenotypic information and developing computational pipelines to identify the most important variant genotypes harbored by each individual. These pipelines consider database and allele frequency annotations and bioinformatics classifications. We propose that the next step will be to integrate these different sources of information to estimate the probability that a given individual has specific phenotypes of clinical interest. To this end, we have designed a Bayesian probabilistic model to predict the probability of dichotomous phenotypes. When applied to a cohort from PGP, predictions of Gilbert syndrome, Graves' disease, non-Hodgkin lymphoma, and various blood groups were accurate, as individuals manifesting the phenotype in question exhibited the highest, or among the highest, predicted probabilities. Thirty-eight PGP phenotypes (26%) were predicted with area-under-the-ROC curve (AUC)>0.7, and 23 (15.8%) of these were statistically significant, based on permutation tests. Moreover, in a Critical Assessment of Genome Interpretation (CAGI) blinded prediction experiment, the models were used to match 77 PGP genomes to phenotypic profiles, generating the most accurate prediction of 16 submissions, according to an independent assessor. Although the models are currently insufficiently accurate for diagnostic utility, we expect their performance to improve with growth of publicly available genomics data and model refinement by domain experts.


Assuntos
Predisposição Genética para Doença/genética , Genoma/genética , Genômica/métodos , Modelos Estatísticos , Análise de Sequência de DNA/métodos , Teorema de Bayes , Estudo de Associação Genômica Ampla , Projeto Genoma Humano , Humanos , Fenótipo
10.
PLoS Genet ; 9(1): e1003224, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23358228

RESUMO

In the past few years, case-control studies of common diseases have shifted their focus from single genes to whole exomes. New sequencing technologies now routinely detect hundreds of thousands of sequence variants in a single study, many of which are rare or even novel. The limitation of classical single-marker association analysis for rare variants has been a challenge in such studies. A new generation of statistical methods for case-control association studies has been developed to meet this challenge. A common approach to association analysis of rare variants is the burden-style collapsing methods to combine rare variant data within individuals across or within genes. Here, we propose a new hybrid likelihood model that combines a burden test with a test of the position distribution of variants. In extensive simulations and on empirical data from the Dallas Heart Study, the new model demonstrates consistently good power, in particular when applied to a gene set (e.g., multiple candidate genes with shared biological function or pathway), when rare variants cluster in key functional regions of a gene, and when protective variants are present. When applied to data from an ongoing sequencing study of bipolar disorder (191 cases, 107 controls), the model identifies seven gene sets with nominal p-values < 0.05, of which one MAPK signaling pathway (KEGG) reaches trend-level significance after correcting for multiple testing.


Assuntos
Estudos de Associação Genética , Quinases de Proteína Quinase Ativadas por Mitógeno , Modelos Genéticos , Transdução de Sinais/genética , Estudos de Casos e Controles , Simulação por Computador , Exoma , Genoma Humano , Humanos , Funções Verossimilhança , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Modelos Teóricos , Polimorfismo de Nucleotídeo Único , Probabilidade
11.
ISRN Bioinform ; 2012: 139842, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-25969743

RESUMO

In this postgenomic era, a huge volume of information derived from expressed sequence tags (ESTs) has been constructed for functional description of gene expression profiles. Comparative studies have become more and more important to researchers of biology. In order to facilitate these comparative studies, we have constructed a user-friendly EST annotation pipeline with comparison tools on an integrated EST service website, Bio301. Bio301 includes regular EST preprocessing, BLAST similarity search, gene ontology (GO) annotation, statistics reporting, a graphical GO browsing interface, and microarray probe selection tools. In addition, Bio301 is equipped with statistical library comparison functions using multiple EST libraries based on GO annotations for mining meaningful biological information.

12.
PLoS One ; 6(5): e20242, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21637803

RESUMO

The Neuregulin-ErbB4 pathway plays a crucial role in brain development and constitutes one of the most biologically plausible signaling pathways implicated in schizophrenia and, to a lesser extent, in bipolar disorder (BP). However, recent genome-wide association analyses have not provided evidence for common variation in NRG1 or ERBB4 influencing schizophrenia or bipolar disorder susceptibility. In this study, we investigate the role of rare coding variants in ERBB4 in BP cases with mood-incongruent psychotic features, a form of BP with arguably the greatest phenotypic overlap with schizophrenia. We performed Sanger sequencing of all 28 exons in ERBB4, as well as part of the promoter and part of the 3'UTR sequence, hypothesizing that rare deleterious variants would be found in 188 cases with mood-incongruent psychosis from the GAIN BP study. We found 42 variants, of which 16 were novel, although none were non-synonymous or clearly deleterious. One of the novel variants, present in 11.2% of cases, is located next to an alternative stop codon, which is associated with a shortened transcript of ERBB4 that is not translated. We genotyped this variant in the GAIN BP case-control samples and found a marginally significant association with mood-incongruent psychotic BP compared with controls (additive model: OR = 1.64, P-value = 0.055; dominant model: OR = 1.73. P-value = 0.039). In conclusion, we found no rare variants of clear deleterious effect, but did uncover a modestly associated novel variant that could affect alternative splicing of ERBB4. However, the modest sample size in this study cannot definitively rule out a role for rare variants in bipolar disorder and studies with larger sample sizes are needed to confirm the observed association.


Assuntos
Transtorno Bipolar/genética , Receptores ErbB/genética , Éxons/genética , Predisposição Genética para Doença , Humanos , Polimorfismo de Nucleotídeo Único/genética , Receptor ErbB-4 , Análise de Sequência de DNA
13.
J Biol Chem ; 286(11): 9726-36, 2011 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-21233208

RESUMO

The pathogenesis of dengue hemorrhagic fever and dengue shock syndrome (DHF/DSS), both serious complications of dengue virus (DV) infection, remains unclear. In this study, we found that anti-DV NS1 (nonstructural protein 1) polyclonal antibodies cross-reacted with human umbilical vein endothelial cells (HUVECs). We further identified a complex-specific mAb, DB16-1, which could recognize DV NS1 and cross-react with HUVECs and human blood vessels. The target protein of DB16-1 was further purified by immunoaffinity chromatography. LC-MS/MS analysis and co-immunoprecipitation revealed that the target protein of DB16-1 was human LYRIC (lysine-rich CEACAM1 co-isolated). Our newly generated anti-LYRIC mAbs bound to HUVECs in a pattern similar to that of DB16-1. The B-cell epitope of DB16-1 displayed a consensus motif, Lys-X-Trp-Gly (KXWG), which corresponded to amino acid residues 116-119 of DV NS1 and mimicked amino acid residues 334-337 in LYRIC. Moreover, the binding activity of DB16-1 in NS1 of DV-2 and in LYRIC disappeared after the KXWG epitope was deleted in each. In conclusion, DB16-1 targeted the same epitope in DV NS1 and LYRIC protein on human endothelial cells, suggesting that it might play a role in the pathogenesis of DHF/DSS. Future studies on the role of the anti-NS1 antibody in causing vascular permeability will undoubtedly be performed on sera collected from individuals before, during, and after the endothelial cell malfunction phase of a dengue illness.


Assuntos
Anticorpos Monoclonais Murinos/imunologia , Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , Autoanticorpos/imunologia , Moléculas de Adesão Celular/imunologia , Vírus da Dengue/imunologia , Dengue/imunologia , Células Endoteliais/imunologia , Mimetismo Molecular/imunologia , Veias Umbilicais/imunologia , Proteínas não Estruturais Virais/imunologia , Animais , Autoantígenos/imunologia , Permeabilidade Capilar/imunologia , Células Cultivadas , Dengue/complicações , Mapeamento de Epitopos , Epitopos/imunologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C
14.
J Med Chem ; 53(5): 2309-13, 2010 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-20148562

RESUMO

A series of aroylquinoline derivatives were synthesized and evaluated for anticancer activity. 5-Amino-6-methoxy-2-aroylquinoline 15 showed more potent antiproliferative activity (IC(50) values ranging from 0.2 to 0.4 nM) as compared to 1a (combretastatin A-4) (IC(50) = 1.9-835 nM) against various human cancer cell lines and a MDR-resistant cancer cell line. Compound 15 (IC(50) = 1.6 microM) exhibited more potent inhibition of tubulin polymerization than 1a (IC(50) = 2.1 microM) and showed strong binding property to the colchicine binding site of microtubules.


Assuntos
Antineoplásicos/síntese química , Quinolinas/síntese química , Moduladores de Tubulina/síntese química , Tubulina (Proteína)/fisiologia , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Concentração Inibidora 50 , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Quinolinas/química , Quinolinas/farmacologia , Relação Estrutura-Atividade , Moduladores de Tubulina/química , Moduladores de Tubulina/farmacologia
15.
J Control Release ; 142(3): 416-21, 2010 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-19919845

RESUMO

A lipid coated calcium phosphate (LCP) nanoparticle (NP) formulation was developed for efficient delivery of small interfering RNA (siRNA) to a xenograft tumor model by intravenous administration. Based on the previous formulation, liposome-polycation-DNA (LPD), which was a DNA-protamine complex wrapped by cationic liposome followed by post-insertion of PEG, LCP was similar to LPD NP except that the core was replaced by a biodegradable nano-sized calcium phosphate precipitate prepared by using water-in-oil micro-emulsions in which siRNA was entrapped. We hypothesized that after entering the cells, LCP would de-assemble at low pH in the endosome, which would cause endosome swelling and bursting to release the entrapped siRNA. Such a mechanism was demonstrated by the increase of intracellular Ca(2+) concentration as shown by using a calcium specific dye Fura-2. The LCP NP was further modified by post-insertion of polyethylene glycol (PEG) with or without anisamide, a sigma-1 receptor ligand for systemic administration. Luciferase siRNA was used to evaluate the gene silencing effect in H-460 cells which were stably transduced with a luciferase gene. The anisamide modified LCP NP silenced about 70% and 50% of luciferase activity for the tumor cells in culture and those grown in a xenograft model, respectively. The untargeted NP showed a very low silencing effect. The new formulation improved the in vitro silencing effect 3-4 folds compared to the previous LPD formulation, but had a negligible immunotoxicity.


Assuntos
Fosfatos de Cálcio/química , Materiais Revestidos Biocompatíveis/química , Portadores de Fármacos/química , Lipídeos/química , Nanopartículas/química , RNA Interferente Pequeno/administração & dosagem , Animais , Cálcio/metabolismo , Fosfatos de Cálcio/toxicidade , Linhagem Celular Tumoral , Materiais Revestidos Biocompatíveis/toxicidade , Citocinas/imunologia , Portadores de Fármacos/toxicidade , Composição de Medicamentos , Feminino , Inativação Gênica/efeitos dos fármacos , Injeções Intravenosas , Lipídeos/toxicidade , Luciferases/genética , Camundongos , Camundongos Nus , Nanopartículas/toxicidade , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Polietilenoglicóis/química , Polietilenoglicóis/toxicidade , RNA Interferente Pequeno/genética , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Mol Ther ; 16(1): 163-9, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17923843

RESUMO

We have developed a self-assembled nanoparticle (NP) that efficiently delivers small interfering RNA (siRNA) to the tumor by intravenous (IV) administration. The NP was obtained by mixing carrier DNA, siRNA, protamine, and lipids, followed by post-modification with polyethylene glycol and a ligand, anisamide. Four hours after IV injection of the formulation into a xenograft model, 70-80% of injected siRNA/g accumulated in the tumor, approximately 10% was detected in the liver and approximately 20% recovered in the lung. Confocal microscopy showed that fluorescent-labeled siRNA was efficiently delivered into the cytoplasm of the sigma receptor expressing NCI-H460 xenograft tumor by the targeted NPs, whereas free siRNA and non-targeted NPs showed little uptake. Three daily injections (1.2 mg/kg) of siRNA formulated in the targeted NPs silenced the epidermal growth factor receptor (EGFR) in the tumor and induced approximately 15% tumor cell apoptosis. Forty percent tumor growth inhibition was achieved by treatment with targeted NPs, while complete inhibition lasted for 1 week when combined with cisplatin. The serum level of liver enzymes and body weight monitoring during the treatment indicated a low level of toxicity of the formulation. The carrier itself also showed little immunotoxicity (IMT).


Assuntos
Sistemas de Liberação de Medicamentos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/terapia , Nanopartículas/administração & dosagem , Interferência de RNA/fisiologia , RNA Interferente Pequeno/administração & dosagem , Animais , Linhagem Celular Tumoral , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Feminino , Humanos , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Especificidade de Órgãos/genética
17.
Clin Vaccine Immunol ; 14(4): 404-11, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17287314

RESUMO

Dengue virus (DEN), the pathogen behind dengue hemorrhagic fever, remains a public health problem in Asia and South America. In this study, monoclonal antibodies (MAbs) against DEN serotype 1 (DEN-1) were generated by fusing NSI/1-Ag4-1 mouse myeloma cells with lymphocytes from BALB/c mice immunized with DEN-1. Twelve MAbs were found to react specifically to the DENs by enzyme-linked immunosorbent assay, immunofluorescence analysis, and immunoblotting analysis. Five MAbs, namely, DA4-7, DA6-7, DA9-5, DA10-2, and DA11-13, were found to react with envelope proteins of DEN-1. Two serotype-specific MAbs of DEN-1, DA6-7 and DA11-13, were further shown to neutralize DEN-1 infection by a plaque reduction neutralization test. The neutralizing epitopes of these MAbs were further identified from a random peptide library displayed on phage. Immunopositive phage clones reacted specifically with these MAbs and did not react with normal mouse serum. Epitope-based peptide antigens were proved able to detect antibodies in serum samples collected from DEN-1-infected patients but not in those taken from DEN-2-infected patients or healthy controls. We believe that these MAbs and neutralizing epitopes will provide information that will lead to the development of DEN-1 serotype-specific diagnostic reagents and vaccines.


Assuntos
Anticorpos Monoclonais/biossíntese , Antígenos Virais/imunologia , Vírus da Dengue/imunologia , Mapeamento de Epitopos , Peptídeos/imunologia , Animais , Anticorpos Monoclonais/sangue , Anticorpos Monoclonais/química , Antígenos Virais/sangue , Linhagem Celular , Humanos , Camundongos , Peptídeos/sangue
18.
Mol Cell ; 24(3): 341-54, 2006 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-17081986

RESUMO

Small ubiquitin-like modifier (SUMO) modification has emerged as an important posttranslational control of protein functions. Daxx, a transcriptional corepressor, was reported to repress the transcriptional potential of several transcription factors and target to PML oncogenic domains (PODs) via SUMO-dependent interactions. The mechanism by which Daxx binds to sumoylated factors mediating transcriptional and subnuclear compartmental regulation remains unclear. Here, we define a SUMO-interacting motif (SIM) within Daxx and show it to be crucial for targeting Daxx to PODs and for transrepression of several sumoylated transcription factors, including glucocorticoid receptor (GR). In addition, the capability of Daxx SIM to bind SUMO also controls Daxx sumoylation. We further demonstrate that arsenic trioxide-induced sumoylation of PML correlates with a change of endogenous Daxx partitioning from GR-regulated gene promoter to PODs and a relief of Daxx repression on GR target gene expression. Our results provide mechanistic insights into Daxx in SUMO-dependent transcriptional control and subnuclear compartmentalization.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Transporte/metabolismo , Núcleo Celular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/química , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Trióxido de Arsênio , Arsenicais/farmacologia , Células COS , Proteínas de Transporte/química , Cercopithecus aethiops , Dexametasona/farmacologia , Células HeLa , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/química , Camundongos , Dados de Sequência Molecular , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/química , Óxidos/farmacologia , Proteína da Leucemia Promielocítica , Ligação Proteica , Estrutura Terciária de Proteína , Transporte Proteico , Receptores de Glucocorticoides/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Genética/efeitos dos fármacos , Proteínas Supressoras de Tumor/metabolismo
19.
Bioinformatics ; 22(17): 2180-2, 2006 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-16820423

RESUMO

UNLABELLED: In this article, we combined EST information from the UniGene database and orthologous relationships from the Ensembl database to construct a ZooDDD database. The primary function of ZooDDD is to mine evolutionary conserved, highly expressed, tissue-specific orthologues in model animals. The candidate genes of interest derived from the ZooDDD database will provide biologists with a good step for comparing the expression, functions and evolution of animal genomes. AVAILABILITY: http://bio301.iis.sinica.edu.tw/~ZooDDDNew/main.php.


Assuntos
Mapeamento Cromossômico/métodos , Bases de Dados Genéticas , Evolução Molecular , Armazenamento e Recuperação da Informação/métodos , Software , Especificidade da Espécie , Interface Usuário-Computador , Sequência de Bases , Gráficos por Computador , Sequência Conservada/genética , Etiquetas de Sequências Expressas , Genoma/genética , Dados de Sequência Molecular , Homologia de Sequência do Ácido Nucleico , Processamento de Sinais Assistido por Computador
20.
J Neurosurg ; 98(2 Suppl): 181-7, 2003 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-12650403

RESUMO

OBJECT: Paraspinal muscle injury is a common but neglected complication of posterior spinal surgery. Evidence suggests that surgical retraction places mechanical and oxidative stress on the paraspinal muscles and that inflammation is a major postoperative pathological finding in the muscles. The roles of cyclooxygenase (COX)-2 and nuclear factor (NF)-kappaB in the inflammatory processes after retraction remain to be clarified. METHODS: In the control group, paraspinal muscles were dissected from the spine via a posterior incision and then laterally retracted. Paraspinal muscle specimens were harvested before as well as at designated time points during and after persistent retraction. The time course of NF-kappaB activation was determined by gel shift assay. Expression of COX-2 was examined using Western blot analysis and immunohistochemistry. The severity of inflammation was evaluated based on histopathology and myeloperoxidase (MPO) activity. The NF-kappaB activation was inhibited by the administration of pyrrolidine dithiolcarbamate (PDTC) in the PDTC-treated group. Retraction induced early activation of NF-kappaB in paraspinal muscle cells. The expression of COX-2 could not be detected until 1 day postoperativley, reaching a peak at 3 days. The time course of COX-2 expression correlated with that of inflammatory responses and MPO activity. Pretreatment with PDTC inhibited intraoperative NF-kappaB activation and greatly downregulated postoperative COX-2 expression and inflammation in the muscles. Postinflammation fibrosis was also abolished by PDTC administration. CONCLUSIONS: Both NF-kappaB-regulated COX-2 expression and inflammation play an important role in the pathogenesis of surgery-associated paraspinal muscle injury. The therapeutic strategy of NF-kappaB inhibition may be applicable to the prevention of such injury.


Assuntos
Isoenzimas/metabolismo , Músculo Esquelético/lesões , NF-kappa B/fisiologia , Procedimentos Neurocirúrgicos/efeitos adversos , Prostaglandina-Endoperóxido Sintases/metabolismo , Coluna Vertebral/cirurgia , Ferimentos e Lesões/etiologia , Animais , Colágeno/metabolismo , Ciclo-Oxigenase 2 , DNA/metabolismo , Masculino , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Peroxidase/metabolismo , Ratos , Ratos Sprague-Dawley , Ferimentos e Lesões/metabolismo , Ferimentos e Lesões/patologia
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