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1.
Int J Cancer ; 147(10): 2801-2810, 2020 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-32875553

RESUMO

BRAF V600E mutations have been reported as a marker of sporadic microsatellite instability (MSI) colorectal cancer (CRC). Current international diagnostic guidelines recommend BRAF mutation testing in MSI CRC patients to predict low risk of Lynch syndrome (LS). We evaluated the age-specific performance of BRAF testing in LS diagnostics. We systematically compared the prevalence of BRAF mutations in LS-associated CRCs and unselected MSI CRCs in different age groups as available from published studies, databases and population-based patient cohorts. Sensitivity/specificity analysis of BRAF testing for exclusion of LS and cost calculations were performed. Among 969 MSI CRCs from LS carriers in the literature and German HNPCC Consortium, 15 (1.6%) harbored BRAF mutations. Six of seven LS patients with BRAF-mutant CRC and reported age were <50 years. Among 339 of 756 (44.8%) of BRAF mutations detected in unselected MSI CRC, only 2 of 339 (0.6%) BRAF mutations were detected in patients <50 years. The inclusion of BRAF testing led to high risk of missing LS patients and increased costs at age <50 years. BRAF testing in patients <50 years carries a high risk of missing a hereditary cancer predisposition and is cost-inefficient. We suggest direct referral of MSI CRC patients <50 years to genetic counseling without BRAF testing.


Assuntos
Substituição de Aminoácidos , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Detecção Precoce de Câncer/métodos , Marcadores Genéticos , Proteínas Proto-Oncogênicas B-raf/genética , Fatores Etários , Idoso , Neoplasias Colorretais Hereditárias sem Polipose/genética , Feminino , Humanos , Masculino , Instabilidade de Microssatélites , Pessoa de Meia-Idade , Taxa de Mutação , Sensibilidade e Especificidade
2.
J Med Genet ; 48(8): 513-9, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21712435

RESUMO

BACKGROUND: A positive family history, germline mutations in DNA mismatch repair genes, tumours with high microsatellite instability, and loss of mismatch repair protein expression are the hallmarks of hereditary non-polyposis colorectal cancer (Lynch syndrome). However, in ~10-15% of cases of suspected Lynch syndrome, no disease-causing mechanism can be detected. METHODS: Oligo array analysis was performed to search for genomic imbalances in patients with suspected mutation-negative Lynch syndrome with MLH1 deficiency in their colorectal tumours. RESULTS AND CONCLUSION: A deletion in the LRRFIP2 (leucine-rich repeat flightless-interacting protein 2) gene flanking the MLH1 gene was detected, which turned out to be a paracentric inversion on chromosome 3p22.2 creating two new stable fusion transcripts between MLH1 and LRRFIP2. A single-nucleotide polymorphism in MLH1 exon 8 was expressed from both alleles, initially pointing to appropriate MLH1 function at least in peripheral cells. In a second case, an inherited duplication of the MLH1 gene region resulted in constitutional MLH1 promoter methylation. Constitutional MLH1 promoter methylation may therefore in rare cases be a heritable disease mechanism and should not be overlooked in seemingly sporadic patients.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias Colorretais Hereditárias sem Polipose/genética , Metilação de DNA/genética , DNA Complementar/genética , Rearranjo Gênico/genética , Proteínas Nucleares/genética , Polimorfismo de Nucleotídeo Único/genética , Regiões Promotoras Genéticas , Alelos , Sequência de Bases , Inversão Cromossômica/genética , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Análise Mutacional de DNA , Éxons/genética , Família , Feminino , Duplicação Gênica/genética , Testes Genéticos , Genoma Humano/genética , Humanos , Masculino , Dados de Sequência Molecular , Proteína 1 Homóloga a MutL , Linhagem
3.
Genes Chromosomes Cancer ; 48(2): 143-54, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18973139

RESUMO

Genomic alterations of cyclin-dependent kinase inhibitors have been demonstrated in a variety of tumor types including brain tumors. Among them, the cyclin-dependent kinase inhibitor 2A (CDKN2A or p16(INK4a)) gene has been shown to be frequently deleted or inactivated in astrocytic tumors. The CDKN2C (p18(INK4c)) gene is functionally related to CDKN2A. Moreover, mice with targeted disruption of CDKN2C alone or combined CDKN2C and cyclin-dependent kinase inhibitor 1B (CDKN1B or p27(Kip1)), or CDKN2C and TP53 gene disruption develop pituitary adenomas (PA) at high frequencies. The purpose of our study was to investigate genetic alterations of the CDKN2C gene by analysis of loss of heterozygosity (LOH), screening for mutations, analysis of promoter methylation, and protein expression in 38 PAs. In addition, genomic alterations and protein expression of the cell cycle genes CDKN2A and its alternatively spliced form, p14(ARF), as well as the retinoblastoma RB1 gene were investigated. LOH at the CDKN2C gene locus was detected in 25% of pituitary adenomas, whereas the RB1 and CDKN2A loci were altered in only 10%. No mutations were detected within the coding regions of the CDKN2C gene. However, 39.5% of adenomas displayed CDKN2C promoter methylation. The absence of CDKN2C protein was correlated with LOH of the CDKN2C locus on chromosome 1 and with methylation of the CDKN2C promoter. This is the first report to describe that the tumor suppressor gene CDKN2C is frequently targeted by genomic alterations in pituitary adenoma. The most common genetic alteration was promoter methylation suggesting that inactivation of CDKN2C by this mechanism may play an important role in pituitary adenoma development. Additional Supporting Information may be found in the online version of this article.


Assuntos
Adenoma/genética , Inibidor de Quinase Dependente de Ciclina p18/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hipofisárias/genética , Adenoma/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Distribuição de Qui-Quadrado , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor de Quinase Dependente de Ciclina p18/metabolismo , Metilação de DNA/genética , Feminino , Humanos , Imuno-Histoquímica , Lactente , Perda de Heterozigosidade , Masculino , Pessoa de Meia-Idade , Hipófise/metabolismo , Neoplasias Hipofisárias/metabolismo , Regiões Promotoras Genéticas , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Análise de Sequência de DNA , Proteína Supressora de Tumor p14ARF/genética , Proteína Supressora de Tumor p14ARF/metabolismo
4.
Immunol Invest ; 38(3-4): 297-310, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19811440

RESUMO

The aim of the study was to assess whether genotypes in the Toll-like receptor 4 gene and in the promoter of the interleukin-18 gene are associated with the susceptibility to chronic periodontitis. 108 chronic periodontitis patients and 76 controls were genotyped for c.896A>G/1196C>T (TLR4 gene) and for c.-368G>C/ c.-838C>A (IL-18 promoter). There were no significant differences in genotype and allele distributions between the study groups. Periodontitis severity in patients with TLR4 c.896AG/1196CT genotype was significantly higher than wildtype carriers. The percentage of teeth with clinical attachment loss > or = 5 mm was 77.3% and 58.8%, respectively (p < or = 0.006, t-test). All subjects were further classified into carriers and non-carriers of at least one variant of each gene. A logistic regression analysis adjusted for gender, smoking, and age showed no association between gene variant carrier status and periodontitis (OR = 1.98, 95% CI 0.61-6.39). The results did not show that IL-18 and TLR4 variants have an effect on the susceptibility to chronic periodontitis. Considering the low number of periodontitis patients carrying TLR4 variants (11%), a comparison of the periodontitis severity depending on the genotype has to be interpreted cautiously.


Assuntos
Periodontite Crônica/genética , Periodontite Crônica/patologia , Predisposição Genética para Doença , Interleucina-18/genética , Receptor 4 Toll-Like/genética , Estudos de Casos e Controles , Feminino , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas
5.
Eur J Hum Genet ; 16(7): 804-11, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18301449

RESUMO

Germline mutations in mismatch repair (MMR) genes, tumours with high microsatellite instability (MSI-H) and loss of MMR protein expression are the hallmarks of HNPCC (Lynch syndrome). While somatic MLH1 promoter hypermethylation is generally accepted in the tumorigenesis of sporadic tumours, abnormal MLH1 promoter methylation in normal body cells is controversially discussed as a mechanism predisposing patients to HNPCC. In all 94 patients suspected of HNPCC-syndrome with a mean age of onset of 45.5 years, MLH1-deficiency in their tumours but no germline mutation, underwent methylation-specific PCR-screening for MLH1 promoter methylation. In peripheral blood cells of 12 patients an MLH1 promoter methylation, in seven informative cases allele-specific, was found. Normal colonic tissue, buccal mucosa, and tumour tissue available from three patients also presented abnormal methylation in the MLH1 promoter. The heredity of aberrant methylation is questionable. Pro: MLH1 promoter methylation was found in a patient and his mother giving evidence for a familial predisposition for an epimutation in MLH1. Contra: a de novo set-up of methylation in one patient, a mosaic or incomplete methylation pattern in six patients, and no evidence for inheritance of MLH1 promoter methylation in the remaining families. Our findings provide strong evidence that MLH1 promoter methylation in normal body cells mimics HNPCC and constitutes a pathogenic pre-lesion in MLH1. The identification of hypermethylation as an epigenetic defect has important implications for surveillance recommendations, as these patients should be treated like Lynch syndrome patients, whereas the heritability of methylation is still under investigation.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Células Sanguíneas/patologia , Neoplasias Colorretais Hereditárias sem Polipose/genética , Metilação de DNA , Padrões de Herança/genética , Mutação/genética , Proteínas Nucleares/genética , Regiões Promotoras Genéticas/genética , Alelos , Sequência de Bases , Ilhas de CpG/genética , Análise Mutacional de DNA , DNA Complementar/genética , DNA de Neoplasias/genética , Família , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Proteína 1 Homóloga a MutL , Fenótipo , Sulfitos/metabolismo
6.
J Clin Periodontol ; 35(12): 1020-6, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18983635

RESUMO

AIM: We aimed to assess the association of different genotypes with increased aggressive periodontitis susceptibility by studying functional relevant variants in the pathogen-recognition receptor Toll-like receptor 4 (TLR4) and variants in the promoter region of the pro-inflammatory cytokine interleukin-18 (IL-18). MATERIAL AND METHODS: One hundred and eleven patients with aggressive periodontitis and 80 periodontally healthy controls were genotyped for four functional variants in the TLR4 gene (c.896A>G and c.1196C>T) and in the IL-18 promoter (c.-368G>C and c.-838C>A). The genotype and allele frequencies, as well as the frequency of combined genotypes were compared between study groups. RESULTS: There were no statistical differences in genotype and allele frequencies within the four variants between the groups. All study subjects were further classified into carriers and non-carriers of at least one variant of both genes. The logistic regression analysis adjusted for gender and smoking showed no association between carrier status of at least one variant of both genes and periodontal status (OR=1.41, 95% CI: 0.43-4.70). CONCLUSIONS: Our results reject the hypothesis that functionally relevant IL-18 and TLR4 gene mutations have a major effect on aggressive periodontitis susceptibility alone or in combination.


Assuntos
Periodontite Agressiva/genética , Interleucina-18/genética , Receptor 4 Toll-Like/genética , Adulto , Periodontite Agressiva/imunologia , Estudos de Casos e Controles , Feminino , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , População Branca
7.
J Clin Periodontol ; 35(4): 311-6, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18294227

RESUMO

AIM: The autosomal-recessive Papillon-Lefèvre syndrome (PLS) is characterized by severe aggressive periodontitis, combined with palmoplantar hyperkeratosis, and is caused by mutations in the Cathepsin C (CTSC) gene. This study aimed to identify CTSC mutations in different PLS phenotypes, including atypical forms and isolated pre-pubertal aggressive periodontitis (PAP). MATERIAL AND METHODS: Thirteen families with different phenotypes were analysed by direct sequencing of the entire coding region and the regulatory regions of CTSC. The function of novel mutations was tested with enzyme activity measurements. RESULTS: In 11 of 13 families, 12 different pathogenic CTSC mutations were found in 10 typical PLS patients, three atypical cases and one PAP patient. Out of four novel mutations, three result in protein truncation and are thus considered to be pathogenic. The homozygous c.854C>T nucleotide exchange (p.P285L) was associated with an almost complete loss of enzyme activity. The observed phenotypic heterogeneity could not be associated with specific genotypes. CONCLUSIONS: The phenotypic variability of the PLS associated with an identical genetic background may reflect the influence of additional genetic or environmental factors on disease characteristics. CTSC mutation analyses should be considered for differential diagnosis in all children suffering from severe aggressive periodontitis.


Assuntos
Periodontite Agressiva/enzimologia , Periodontite Agressiva/genética , Catepsina C/genética , Doença de Papillon-Lefevre/enzimologia , Doença de Papillon-Lefevre/genética , Adulto , Criança , Códon sem Sentido , Análise Mutacional de DNA , Mutação da Fase de Leitura , Humanos , Elastase de Leucócito/genética , Mutação de Sentido Incorreto , Fenótipo
8.
Oncogene ; 24(32): 5079-88, 2005 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-15897887

RESUMO

Pancreatic ductal adenocarcinoma is the eighth most common cancer with the lowest overall 5-year relative survival rate of any tumor type today. Expression profiling using microarrays has been widely used to identify genes associated with pancreatic cancer development. To extract maximum value from the available gene expression data, we applied a meta-analysis to search for commonly differentially expressed genes in pancreatic ductal adenocarcinoma. We obtained data sets from four different gene expression studies on pancreatic cancer. We selected a consensus set of 2984 genes measured in all four studies and applied a meta-analysis approach to evaluate the combined data. Of the genes identified as differentially expressed, several were validated using RT-PCR and immunohistochemistry. Additionally, we used a class discovery algorithm to identify a gene expression signature. Our meta-analysis revealed that the pancreatic cancer gene expression data sets shared a significant number of up- and downregulated genes, independent of the technology used. This interstudy crossvalidation approach generated a set of 568 genes that were consistently and significantly dysregulated in pancreatic cancer. Of these, 364 (64.1%) were upregulated and 204 (35.9%) were downregulated in pancreatic cancer. Only 127 (22%) were described in the published individual analyses. Functional annotation of the genes revealed that genes presumably associated with the cell adhesion-mediated drug resistance pathway are frequently overexpressed in pancreatic cancer. Meta-analysis is an important tool for the identification and validation of differentially expressed genes. These could represent good candidates for novel diagnostic and therapeutic approaches to pancreatic cancer.


Assuntos
Regulação Neoplásica da Expressão Gênica , Análise de Sequência com Séries de Oligonucleotídeos , Neoplasias Pancreáticas/genética , Adenocarcinoma/genética , Carcinoma Ductal Pancreático/genética , Humanos , Neoplasias Pancreáticas/mortalidade , Reprodutibilidade dos Testes , Taxa de Sobrevida
9.
Clin Cancer Res ; 11(3): 1259-67, 2005 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-15709197

RESUMO

PURPOSE: Cerebral metastases represent the most common type of brain tumors. This study investigated the effects of endogenous endostatin on hematogenous cerebral melanoma metastases. EXPERIMENTAL DESIGN: Murine K1735 melanoma cells were transfected with the mouse endostatin cDNA. Experimental tumors were induced either by s.c. injection, intracerebral implantation, or via injection into the internal carotid artery to simulate hematogenous metastatic spread. The effects of endostatin expression on tumor incidence, growth pattern, and vascularity were analyzed. RESULTS: In vitro secretion of endostatin by 2.5 x 10(5) cells within 24 hours was 0.12 +/- 0.03 ng, 4.35 +/- 0.4, and 1.18 +/- 0.7 ng/mL for wild type and two endostatin-transfected K1735 clones termed K1735-endo/2 and K1735-endo/8, respectively. Tumor inhibition in vivo correlated with endogenous endostatin production. Within 25 days, growth of s.c. K1735-endo/2 tumors was <20% compared with wild-type controls. Following intracerebral implantation the average survival time of mice was 27.8 +/- 2.6 versus 13.3 +/- 3.7 days in the K1735-endo/2 versus the wild-type group, respectively. Intracarotid injection of 1 x 10(5) wild-type cells killed the mice within 24 +/- 1.8 days. In contrast, endostatin expression prevented macroscopic metastatic tumor growth in 11 of 12 mice, although viable microscopic tumor pockets were detectable in all animals. CONCLUSION: Endostatin inhibits tumor progression of multiple cerebral metastases in vivo. Hematogenous micrometastases are more efficiently suppressed than tumors resulting from high focal cell numbers which may be due to a higher angiogenic signaling exerted by massive cell deposits. Endostatin may prevent solid tumor growth more effectively by inhibition of early angiogenesis.


Assuntos
Neoplasias Encefálicas/secundário , Endostatinas/metabolismo , Melanoma/patologia , Animais , Apoptose , Linhagem Celular Tumoral , Endostatinas/genética , Feminino , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Injeções Intraventriculares , Injeções Subcutâneas , Antígeno Ki-67/análise , Melanoma/genética , Melanoma/metabolismo , Camundongos , Camundongos Endogâmicos , Transplante de Neoplasias , Neovascularização Patológica/patologia , Transfecção
10.
Neoplasia ; 6(5): 611-22, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15548371

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) remains an important cause of malignancy-related death and is the eighth most common cancer with the lowest overall 5-year relative survival rate. To identify new molecular markers and candidates for new therapeutic regimens, we investigated the gene expression profile of microdissected cells from 11 normal pancreatic ducts, 14 samples of PDAC, and 4 well-characterized pancreatic cancer cell lines using the Affymetrix U133 GeneChip set. RNA was extracted from microdissected samples and cell lines, amplified, and labeled using a repetitive in vitro transcription protocol. Differentially expressed genes were identified using the significance analysis of microarrays program. We found 616 differentially expressed genes. Within these, 140 were also identified in PDAC by others, such as Galectin-1, Galectin-3, and MT-SP2. We validated the differential expression of several genes (e.g., CENPF, MCM2, MCM7, RAMP, IRAK1, and PTTG1) in PDAC by immunohistochemistry and reverse transcription polymerase chain reaction. We present a whole genome expression study of microdissected tissues from PDAC, from microdissected normal ductal pancreatic cells and pancreatic cancer cell lines using high-density microarrays. Within the panel of genes, we identified novel differentially expressed genes, which have not been associated with the pathogenesis of PDAC before.


Assuntos
Carcinoma Ductal Pancreático/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pancreáticas/genética , Idoso , Carcinoma Ductal Pancreático/diagnóstico , Carcinoma Ductal Pancreático/patologia , Proteínas de Ciclo Celular/análise , Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/análise , Proteínas Cromossômicas não Histona/genética , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/genética , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Microdissecção , Proteínas dos Microfilamentos , Pessoa de Meia-Idade , Componente 2 do Complexo de Manutenção de Minicromossomo , Componente 7 do Complexo de Manutenção de Minicromossomo , Proteínas Nucleares/análise , Proteínas Nucleares/genética , Análise de Sequência com Séries de Oligonucleotídeos , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/patologia , Regulação para Cima
11.
Oral Oncol ; 38(5): 486-90, 2002 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-12110344

RESUMO

We investigated the association of polymorphisms of drug-metabolizing enzymes and susceptibility to oral cavity cancer. Polymerase chain reaction (PCR)-based analyses were performed on genomic DNA of 94 Caucasian patients in Germany and 92 healthy German controls to determine genotypes of polymorphisms in CYP1A1, GSTM1 and NAT2. For CYP1A1, the homozygous mutant genotype Val/Val did not occur. The heterozygous genotype Ile/Val (6.5% cases versus 4.3% controls) and the homozygous wild-type Ile/Ile (95.7% cases versus 93.5% controls) showed no statistically significant differences between groups (X(2)=0.47; P=0.534, Fisher's exact test, two-sided). The GSTM1 homozygous null genotype occurred more frequently in cancer patients (59.6%) compared to controls (53.3%) but this difference remained insignificant in X(2)-analysis (X(2)=1.07; P=0.587). Almost identical genotype distributions between cases and controls were found for all three NAT2 acetylators. Hence, these three genetic polymorphisms are unlikely to be associated with oral cavity cancer in the population studied.


Assuntos
Arilamina N-Acetiltransferase/genética , Carcinoma de Células Escamosas/genética , Predisposição Genética para Doença , Glutationa Transferase/genética , Neoplasias Bucais/genética , Proteínas de Neoplasias/genética , Polimorfismo Genético , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/enzimologia , Citocromo P-450 CYP1A1/genética , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/enzimologia
12.
Dtsch Arztebl Int ; 110(3): 32-8, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23413378

RESUMO

BACKGROUND: Hereditary nonpolyposis colorectal cancer HNPCC, Lynch syndrome) is a genetic disease of autosomal dominant inheritance. It is caused by a mutation in one of four genes of the DNA mismatch repair system and confers a markedly increased risk for various types of cancer, particularly of the colon and the endometrium. Its prevalence in the general population is about 1 in 500, and it causes about 2% to 3% of all colorectal cancers. Lynch syndrome is diagnosed in two steps: If it is suspected (because a patient develops cancer at an unusually young age or because of familial clustering), the tumor tissue is analyzed for evidence of deficient mismatch repair (microsatellite instability, loss of mismatch repair protein expression). If such evidence is found, a genetic mutation is sought. The identification of a pathogenic mutation confirms the diagnosis in the patient and enables predictive testing of other family members. Diagnostic evaluations for Lynch syndrome should be carried out with appropriate genetic counseling. METHOD: Selective literature review. RESULTS: Prospective cohort studies from Germany, Finland and the Netherlands have shown that colorectal cancers detected by systematic colonoscopic surveillance tend to be at an earlier stage than those that are discovered after the patients present with symptoms. The Finnish study also showed an overall reduction in cancer risk from colonoscopic polypectomy at regular intervals. CONCLUSION: The studies conducted so far have not yet clearly documented the putative benefit of an individualized, risk-adapted surveillance strategy. Until this is done, patients with Lynch syndrome and healthy carriers of causative mutations should be monitored with annual colonoscopy and (for women) annual gynecological examination.


Assuntos
Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Neoplasias Colorretais Hereditárias sem Polipose/genética , Aconselhamento Genético/métodos , Marcadores Genéticos/genética , Predisposição Genética para Doença/genética , Testes Genéticos/métodos , Polimorfismo de Nucleotídeo Único/genética , Diagnóstico Diferencial , Humanos
13.
Fam Cancer ; 10(2): 273-84, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21404117

RESUMO

Missense mutations of the DNA mismatch repair gene MLH1 are found in a significant fraction of patients with Lynch syndrome (hereditary nonpolyposis colorectal cancer, HNPCC) and their pathogenicity often remains unclear. We report here all 88 MLH1 missense variants identified in families from the German HNPCC consortium with clinical details of these patients/families. We investigated 23 MLH1 missense variants by two functional in vivo assays in yeast; seven map to the ATPase and 16 to the protein interaction domain. In the yeast-2-hybrid (Y2H) assay three variants in the ATPase and twelve variants in the interaction domain showed no or a reduced interaction with PMS2; seven showed a normal and one a significantly higher interaction. Using the Lys2A (14) reporter system to study the dominant negative mutator effect (DNE), 16 variants showed no or a low mutator effect, suggesting that these are nonfunctional, three were intermediate and four wild type in this assay. The DNE and Y2H results were concordant for all variants in the interaction domain, whereas slightly divergent results were obtained for variants in the ATPase domain. Analysis of the stability of the missense proteins in yeast and human embryonic kidney cells (293T) revealed a very low expression for seven of the variants in yeast and for nine in human cells. In total 15 variants were classified as deleterious, five were classified as variants of unclassified significance (VUS) and three were basically normal in the functional assays, P603R, K618R, Q689R, suggesting that these are neutral.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias Colorretais Hereditárias sem Polipose/genética , Mutação de Sentido Incorreto , Proteínas Nucleares/genética , Adenosina Trifosfatases/química , Adenosina Trifosfatases/genética , Células Cultivadas , Reparo de Erro de Pareamento de DNA , Humanos , Proteína 1 Homóloga a MutL , Estrutura Terciária de Proteína , Técnicas do Sistema de Duplo-Híbrido , Leveduras/genética
14.
PLoS One ; 5(8): e12243, 2010 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-20808857

RESUMO

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) remains an important cause of cancer death. Changes in apoptosis signaling in pancreatic cancer result in chemotherapy resistance and aggressive growth and metastasizing. The aim of this study was to characterize the apoptosis pathway in pancreatic cancer computationally by evaluation of experimental data from high-throughput technologies and public data bases. Therefore, gene expression analysis of microdissected pancreatic tumor tissue was implemented in a model of the apoptosis pathway obtained by computational protein interaction prediction. METHODOLOGY/PRINCIPAL FINDINGS: Apoptosis pathway related genes were assembled from electronic databases. To assess expression of these genes we constructed a virtual subarray from a whole genome analysis from microdissected native tumor tissue. To obtain a model of the apoptosis pathway, interactions of members of the apoptosis pathway were analysed using public databases and computational prediction of protein interactions. Gene expression data were implemented in the apoptosis pathway model. 19 genes were found differentially expressed and 12 genes had an already known pathophysiological role in PDAC, such as Survivin/BIRC5, BNIP3 and TNF-R1. Furthermore we validated differential expression of IL1R2 and Livin/BIRC7 by RT-PCR and immunohistochemistry. Implementation of the gene expression data in the apoptosis pathway map suggested two higher level defects of the pathway at the level of cell death receptors and within the intrinsic signaling cascade consistent with references on apoptosis in PDAC. Protein interaction prediction further showed possible new interactions between the single pathway members, which demonstrate the complexity of the apoptosis pathway. CONCLUSIONS/SIGNIFICANCE: Our data shows that by computational evaluation of public accessible data an acceptable virtual image of the apoptosis pathway might be given. By this approach we could identify two higher level defects of the apoptosis pathway in PDAC. We could further for the first time identify IL1R2 as possible candidate gene in PDAC.


Assuntos
Apoptose , Biologia Computacional/métodos , Neoplasias Pancreáticas/patologia , Transdução de Sinais , Linhagem Celular Tumoral , Perfilação da Expressão Gênica , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Neoplasias Pancreáticas/genética , Fenótipo
15.
Gynecol Oncol ; 102(2): 189-94, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16476474

RESUMO

BACKGROUND AND OBJECTIVE: Hereditary nonpolyposis colorectal cancer (HNPCC) is the most frequent form of hereditary colorectal cancer. In addition to the high lifetime risk for colorectal cancer in mutation carriers, there is also a remarkably increased risk for endometrial cancer (EC). METHODS: In this retrospective study, clinical and molecular approach to the individual decision making as to whether or not to perform a prophylactic hysterectomy in a subset of HNPCC patients is discussed. 147 female patients meeting at least one criterion of the Bethesda guidelines were included in this analysis between 1995 and 2003. After clinical and genetic counseling, patients gave informed written consent and microsatellite analysis, immunohistochemistry and sequencing of the mismatch repair genes MLH1, MSH2 and MSH6 was performed. RESULTS: 11 of the analyzed patients had a personal history of EC and had undergone previous hysterectomy at ages 26 to 62 years. Prophylactic hysterectomy with oophorectomy was considered in postmenopausal women meeting the Amsterdam criteria and/or carrying a disease causing mismatch repair gene mutation who were operated on because of diagnosed colorectal cancer in our center for hereditary cancer. This procedure was performed in 4 patients. None of them had shown any symptoms of a gynecologic malignancy. Preoperative gynecological examination showed no evidence for EC or ovarian cancer in these patients. Postoperative histological examination showed EC stage T1b N0 M0 in 2 patients. CONCLUSIONS: Since the efficiency of gynecological surveillance is uncertain, prophylactic hysterectomy could be an option for a subset of HNPCC patients and mutation carriers.


Assuntos
Neoplasias Colorretais Hereditárias sem Polipose/cirurgia , Neoplasias do Endométrio/cirurgia , Adulto , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/patologia , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/prevenção & controle , Feminino , Aconselhamento Genético , Predisposição Genética para Doença , Humanos , Histerectomia , Pessoa de Meia-Idade , Estudos Retrospectivos
16.
J Neurooncol ; 72(2): 95-102, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15925987

RESUMO

Two metachronous anaplastic oligoastrocytomas with different cerebral locations were analyzed in a 51-year-old patient with an extended recurrence-free interval of 6 years and an a long survival of 9 years. Remarkably, the patient had not undergone adjuvant chemotherapy. Different cytogenetic and molecular techniques were performed including comparative genomic hybridization (CGH), fluorescence in situ hybridization (FISH), allelic loss analysis, sequencing of p53, p16(INK4a)/CDKN2A and p14(ARF), EGFRamplification studies, investigation of the DNA mismatch repair system as well as tumor clonality. Using CGH and FISH a profile of low accumulation of cytogenetic aberrations was found in the second tumor, with no significant increase in the percentage of hyperdiploid nuclei. Microsatellite analysis showed a common pattern of allelic losses at 1p36, 19q13 and 9p21. Both specimens were also similar in that they retained heterozygosity at 10q23-q24 and 13q14 and that they harbor neither EGFR amplification nor mutations of p53, p16(INK4a)/CDKN2A or p14(ARF). The only further alteration in the second tumor was an allelic loss at p53. The X-chromosome inactivation (HUMARA) analysis revealed a polyclonal pattern in both samples. Our data strongly suggest that the second anaplastic oligoastrocytoma developed as a distant relapse of the first tumor. Whether the paucity of accumulation of the observed genetic alterations might be associated with the unusually extended relapse-free time of the patient remains to be elucidated.


Assuntos
Neoplasias Encefálicas/genética , Segunda Neoplasia Primária/genética , Oligodendroglioma/genética , Neoplasias Encefálicas/radioterapia , Neoplasias Encefálicas/cirurgia , Análise Mutacional de DNA , Intervalo Livre de Doença , Feminino , Amplificação de Genes , Genes erbB-1/genética , Humanos , Hibridização in Situ Fluorescente , Perda de Heterozigosidade , Repetições de Microssatélites , Pessoa de Meia-Idade , Segunda Neoplasia Primária/radioterapia , Segunda Neoplasia Primária/cirurgia , Oligodendroglioma/radioterapia , Oligodendroglioma/cirurgia , Análise de Sequência de DNA
17.
Int J Colorectal Dis ; 19(5): 401-13, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-14745573

RESUMO

BACKGROUND: Pancreatic ductal adenocarcinoma has an extremely poor prognosis. To improve the prognosis, novel molecular markers and targets for earlier diagnosis and adjuvant and/or neoadjuvant treatment need to be identified. One of the key techniques that has been developed to achieve this goal is DNA microarray profiling, which is used to identify the mechanisms of deregulated molecular functions in pancreatic carcinoma cells. OBJECTIVE: As several studies using microarrays have already been published, this review attempts to compare published data and crossvalidate the results. In addition, the applied techniques are discussed.


Assuntos
Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Perfilação da Expressão Gênica , Análise de Sequência com Séries de Oligonucleotídeos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/terapia , DNA de Neoplasias/análise , Humanos , Terapia Neoadjuvante , Neoplasias Pancreáticas/terapia , Prognóstico
18.
Acta Neuropathol ; 103(3): 255-66, 2002 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-11907806

RESUMO

The human gene termed LGI1 (leucine-rich gene - glioma inactivated) has been isolated recently, and is supposed to be an additional candidate tumor suppressor gene involved in the formation and progression of glioblastoma multiforme [Chernova et al. (1998) Oncogene 17:2873-2881]. To test this hypothesis and to complete the characterization of the gene, we performed various detailed studies on the genomic structure, the mRNA expression level, the integrity of the cDNA, and retroviral gene transfer into LGI1-deficient cell lines. Two single nucleotide polymorphisms in the promotor region and a highly polymorphic intragenic microsatellite repeat between exon 4 and 5 were found. Phylogenetic sequence analysis techniques were applied, which showed functional relationships between LGI1 and TRK and SLIT protein families that are known to be involved in development and maintenance of the nervous system. Fluorescence in situ hybridization (FISH) analysis showed LGI1 to be present on 10q24 in each of 11 glioma-derived cell lines evaluated. Sequence analysis of the LGI1 transcript did not detect any mutation. Relative amounts of LGI1 mRNA copy numbers as measured by the real-time fluorescence detection LightCycler technology differed more than three orders of magnitude and were significantly reduced in 10 of 11 cell lines. Retroviral gene transfer into LGI1-deficient glioma-derived cell lines could not substantiate any difference to control infected cultures regarding growth rate, S phase transition, and maintenance of marker gene expression. The strong homology to proteins involved in development, differentiation, or maintenance of the nervous system provides evidence for a function of the LGI1 protein in neural tissue. The observation that translocation or deletion of the LGI1 locus or mutation of the coding sequence of the LGI1 mRNA is not a frequent event in malignant glioma cell lines suggests that epigenetic factors lead to substantial differences in the amount of LGI1 mRNA expression. In addition, that the effect is lacking after retroviral gene transfer in cell culture suggests that binding of some kind of a ligand is essential for its biological activity.


Assuntos
Repetições de Dinucleotídeos/genética , Glioblastoma/genética , Regiões Promotoras Genéticas/genética , Proteínas/classificação , Proteínas/genética , RNA Mensageiro/classificação , RNA Mensageiro/genética , DNA Complementar/genética , DNA Complementar/fisiologia , Repetições de Dinucleotídeos/fisiologia , Expressão Gênica/genética , Expressão Gênica/fisiologia , Técnicas de Transferência de Genes , Glioblastoma/fisiopatologia , Humanos , Hibridização in Situ Fluorescente , Peptídeos e Proteínas de Sinalização Intracelular , Regiões Promotoras Genéticas/fisiologia , Proteínas/fisiologia , RNA Mensageiro/fisiologia , Análise de Sequência de DNA , Células Tumorais Cultivadas
19.
Genome Biol ; 5(9): R67, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15345051

RESUMO

BACKGROUND: The ambystomatid salamander, Ambystoma mexicanum (axolotl), is an important model organism in evolutionary and regeneration research but relatively little sequence information has so far been available. This is a major limitation for molecular studies on caudate development, regeneration and evolution. To address this lack of sequence information we have generated an expressed sequence tag (EST) database for A. mexicanum. RESULTS: Two cDNA libraries, one made from stage 18-22 embryos and the other from day-6 regenerating tail blastemas, generated 17,352 sequences. From the sequenced ESTs, 6,377 contigs were assembled that probably represent 25% of the expressed genes in this organism. Sequence comparison revealed significant homology to entries in the NCBI non-redundant database. Further examination of this gene set revealed the presence of genes involved in important cell and developmental processes, including cell proliferation, cell differentiation and cell-cell communication. On the basis of these data, we have performed phylogenetic analysis of key cell-cycle regulators. Interestingly, while cell-cycle proteins such as the cyclin B family display expected evolutionary relationships, the cyclin-dependent kinase inhibitor 1 gene family shows an unusual evolutionary behavior among the amphibians. CONCLUSIONS: Our analysis reveals the importance of a comprehensive sequence set from a representative of the Caudata and illustrates that the EST sequence database is a rich source of molecular, developmental and regeneration studies. To aid in data mining, the ESTs have been organized into an easily searchable database that is freely available online.


Assuntos
Ambystoma/embriologia , Ambystoma/genética , Etiquetas de Sequências Expressas , Biblioteca Gênica , Regeneração/genética , Reprodução Assexuada/genética , Análise de Sequência de DNA/métodos , Animais , Composição de Bases/genética , Ciclo Celular/genética , Proteínas de Ciclo Celular/genética , Diferenciação Celular/genética , Proliferação de Células , Sequência Conservada/genética , Mapeamento de Sequências Contíguas/métodos , Inibidor de Quinase Dependente de Ciclina p21 , Bases de Dados Genéticas , Embrião não Mamífero/química , Embrião não Mamífero/metabolismo , Genes/genética , Genes cdc , Família Multigênica/genética , Crista Neural/química , Crista Neural/metabolismo , Filogenia , RNA/metabolismo , Homologia de Sequência do Ácido Nucleico , Cauda/embriologia
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