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1.
Klin Onkol ; 32(Supplementum2): 36-50, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31409080

RESUMO

BACKGROUND: Hereditary mutations in the CHEK2 gene (which encodes CHK2 kinase) contribute to a moderately increased risk of breast cancer (BC) and other cancers. Large variations in the frequency of CHEK2 mutations and the occurrence of variants of unknown clinical significance (VUS) complicate estimation of cancer risk in carriers of germline CHEK2 mutations. PATIENTS AND METHODS: We performed mutation analysis of 1,526 high-risk Czech BC patients and 3,360 Czech controls. Functional analysis was performed for identified VUS using a model system based on a human RPE1-CHEK2-KO cell line harboring biallelic inactivation of endogenous CHEK2. RESULTS: The frequency of ten truncating CHEK2 variants differed markedly between BC patients (2.26%) and controls (0.11%; p = 4.1 × 1012). We also found 23 different missense variants in 4.5% patients and in 4.0% of controls. The most common was p.I157T, which was found in patients and controls with the same frequency. Functional analysis identified nine functionally deleterious VUS, another nine functionally neutral VUS, and four intermediate VUS (including p.I157T). We found that carriers of truncating CHEK2 mutations had a high BC risk (OR 8.19; 95% CI 4.11-17.75), and that carriers of functionally deleterious missense variants had a moderate risk (OR 4.06; 95% CI, 1.37-13.39). Carriers of these mutations developed BC at 44.4 and 50.7 years, respectively. Functionally neutral and functionally intermediate missense variants did not increase the BC risk. BC in CHEK2 mutation carriers was frequently ER-positive and of higher grade. Notably, carriers of CHEK2 mutations developed second cancers more frequently than BRCA1/BRCA2/PALB2/p53 or mutation non-carriers. CONCLUSION: Hereditary CHEK2 mutations contribute to the development of hereditary BC. The associated cancer risk in mutation carriers increases with the number of affected individuals in a family. Annual follow-up with breast ultrasound, mammography, or magnetic resonance imaging is recommended for asymptomatic mutation carriers from the age of 40. Surgical prevention and specific follow-up of other tumors should be considered based on family cancer history. The work was supported by grants from the Czech Health Research Council of the Ministry of Health of the Czech Republic NR 15-28830A, 16-29959A, NV19-03-00279, projects of the PROGRES Q28/LF1, GAUK 762216, SVV2019 / 260367, PRIMUS/17/MED/9, UNCE/MED/016, Progress Q26, LQ1604 NPU II and project AVČR Qualitas. The analysis of a set of unselected controls was made possible by the existence and support of the scientific infrastructure of the National Center for Medical Genomics (LM2015091) and its project aimed at creating a reference database of genetic variants of the Czech Republic (CZ.02.1.01/0.0/0.0/16_013/0001634). The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers. Submitted: 2. 4. 2019 Accepted: 14. 5. 2019.

2.
Klin Onkol ; 32(Supplementum2): 72-78, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31409082

RESUMO

BACKGROUND: Ovarian cancer is a disease with high mortality. Approximately 1,000 women are diagnosed with ovarian cancer in the Czech Republic annually. Women harboring a mutation in cancer-predisposing genes face an increased risk of tumor development. Mutations in BRCA1, BRCA2, BRIP1, and Lynch syndrome genes (RAD51C, RAD51D, and STK11) are associated with a high risk of ovarian cancer, and mutations in ATM, CHEK2, NBN, PALB2, and BARD1 appear to increase the risk. Our aim was to examine the frequency of mutations in cancer-predisposing genes in the Czech Republic. MATERIALS AND METHODS: We analyzed 1,057 individuals including ovarian cancer patients and 617 non-cancer controls using CZECANCA panel next-generation sequencing on the Illumina platform. Pathogenic mutations in high-risk genes, including CNVs, were detected in 30.6% of patients. The mutation frequency reached 25.0% and 18.2% in subgroups of unselected ovarian cancer patients and patients with a negative family cancer history, respectively. The most frequently mutated genes were BRCA1 and BRCA2. The overall frequency of mutations in non-BRCA genes was comparable to that in BRCA2. The mutation frequency in ovarian cancer patients aged >70 years was three times higher than that in patients diagnosed before the age of 30. CONCLUSION: Ovarian cancer is a heterogeneous disease with a high proportion of hereditary cases. The lack of efficient screening for early diagnosis emphasizes the importance of identifying carriers of mutations in ovarian cancer-predisposing genes; this is because proper follow-up and prevention strategies can reduce overall ovarian cancer-related mortality. This work was supported by grants AZV 15-27695A, SVV2019/260367, PROGRES Q28/LF1. The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers. Submitted: 7. 3. 2019 Accepted: 24. 4. 2019.

3.
Int J Cancer ; 145(7): 1782-1797, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31050813

RESUMO

Germline mutations in checkpoint kinase 2 (CHEK2), a multiple cancer-predisposing gene, increase breast cancer (BC) risk; however, risk estimates differ substantially in published studies. We analyzed germline CHEK2 variants in 1,928 high-risk Czech breast/ovarian cancer (BC/OC) patients and 3,360 population-matched controls (PMCs). For a functional classification of VUS, we developed a complementation assay in human nontransformed RPE1-CHEK2-knockout cells quantifying CHK2-specific phosphorylation of endogenous protein KAP1. We identified 10 truncations in 46 (2.39%) patients and in 11 (0.33%) PMC (p = 1.1 × 10-14 ). Two types of large intragenic rearrangements (LGR) were found in 20/46 mutation carriers. Truncations significantly increased unilateral BC risk (OR = 7.94; 95%CI 3.90-17.47; p = 1.1 × 10-14 ) and were more frequent in patients with bilateral BC (4/149; 2.68%; p = 0.003), double primary BC/OC (3/79; 3.80%; p = 0.004), male BC (3/48; 6.25%; p = 8.6 × 10-4 ), but not with OC (3/354; 0.85%; p = 0.14). Additionally, we found 26 missense VUS in 88 (4.56%) patients and 131 (3.90%) PMC (p = 0.22). Using our functional assay, 11 variants identified in 15 (0.78%) patients and 6 (0.18%) PMC were scored deleterious (p = 0.002). Frequencies of functionally intermediate and neutral variants did not differ between patients and PMC. Functionally deleterious CHEK2 missense variants significantly increased BC risk (OR = 3.90; 95%CI 1.24-13.35; p = 0.009) and marginally OC risk (OR = 4.77; 95%CI 0.77-22.47; p = 0.047); however, carriers low frequency will require evaluation in larger studies. Our study highlights importance of LGR detection for CHEK2 analysis, careful consideration of ethnicity in both cases and controls for risk estimates, and demonstrates promising potential of newly developed human nontransformed cell line assay for functional CHEK2 VUS classification.

4.
Chemosphere ; 213: 568-577, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30268053

RESUMO

Nano-scale zero-valent iron (nZVI) began attracting research attention in remediation practice in recent decades as a prospective nanomaterial applicable to various contaminated matrices. Despite concerns about the negative effects of nanomaterials on ecosystems, the number of reliable toxicity tests is limited. We have developed a test based on the evaluation of oxidative stress (OS). The test employed the analysis of a typical OS marker (malondialdehyde, MDA), after exposure of six bacterial strains to the tested nanomaterial. We also attempted to use other OS and cell membrane damage assays, including the determination of glutathione and lactate dehydrogenase, respectively. However, we found that the components of these assays interfered with nZVI; therefore, these tests were not applicable. The MDA assay was tested using nZVI and three newly engineered oxide shell nZVI materials with different oxide thicknesses. Six different bacterial species were employed, and the results showed that the test was fully applicable for the concentrations of nanomaterials used in remediation practice (0.1-10 g/L). MDA was produced in a dose-response manner, and the bacteria showed a similar response toward pure pyrophoric nZVI, reaching EC50 values of 0.3-1.1 g/L. We observed different responses in the absolute production of MDA; however, the MDA concentrations were correlated with the cell membrane surfaces of the individual strains (R > 0.75; P < 0.09). Additionally, the EC50 values correlated with the thickness of the oxide shells (except for Escherichia coli: R > 0.95; P < 0.05), documenting the reliability of the assay, where reactivity was confirmed to be an important factor for reactive oxygen species production.


Assuntos
Bactérias/efeitos dos fármacos , Ferro/toxicidade , Peroxidação de Lipídeos , Nanopartículas Metálicas/toxicidade , Malondialdeído , Nanoestruturas/toxicidade , Estresse Oxidativo , Estudos Prospectivos , Espécies Reativas de Oxigênio/metabolismo , Reprodutibilidade dos Testes , Testes de Toxicidade/métodos
5.
Hum Mutat ; 39(12): 2025-2039, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30204945

RESUMO

The widespread use of next generation sequencing for clinical testing is detecting an escalating number of variants in noncoding regions of the genome. The clinical significance of the majority of these variants is currently unknown, which presents a significant clinical challenge. We have screened over 6,000 early-onset and/or familial breast cancer (BC) cases collected by the ENIGMA consortium for sequence variants in the 5' noncoding regions of BC susceptibility genes BRCA1 and BRCA2, and identified 141 rare variants with global minor allele frequency < 0.01, 76 of which have not been reported previously. Bioinformatic analysis identified a set of 21 variants most likely to impact transcriptional regulation, and luciferase reporter assays detected altered promoter activity for four of these variants. Electrophoretic mobility shift assays demonstrated that three of these altered the binding of proteins to the respective BRCA1 or BRCA2 promoter regions, including NFYA binding to BRCA1:c.-287C>T and PAX5 binding to BRCA2:c.-296C>T. Clinical classification of variants affecting promoter activity, using existing prediction models, found no evidence to suggest that these variants confer a high risk of disease. Further studies are required to determine if such variation may be associated with a moderate or low risk of BC.

6.
PLoS One ; 13(4): e0195761, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29649263

RESUMO

BACKGROUND: Carriers of mutations in hereditary cancer predisposition genes represent a small but clinically important subgroup of oncology patients. The identification of causal germline mutations determines follow-up management, treatment options and genetic counselling in patients' families. Targeted next-generation sequencing-based analyses using cancer-specific panels in high-risk individuals have been rapidly adopted by diagnostic laboratories. While the use of diagnosis-specific panels is straightforward in typical cases, individuals with unusual phenotypes from families with overlapping criteria require multiple panel testing. Moreover, narrow gene panels are limited by our currently incomplete knowledge about possible genetic dispositions. METHODS: We have designed a multi-gene panel called CZECANCA (CZEch CAncer paNel for Clinical Application) for a sequencing analysis of 219 cancer-susceptibility and candidate predisposition genes associated with frequent hereditary cancers. RESULTS: The bioanalytical and bioinformatics pipeline was validated on a set of internal and commercially available DNA controls showing high coverage uniformity, sensitivity, specificity and accuracy. The panel demonstrates a reliable detection of both single nucleotide and copy number variants. Inter-laboratory, intra- and inter-run replicates confirmed the robustness of our approach. CONCLUSION: The objective of CZECANCA is a nationwide consolidation of cancer-predisposition genetic testing across various clinical indications with savings in costs, human labor and turnaround time. Moreover, the unified diagnostics will enable the integration and analysis of genotypes with associated phenotypes in a national database improving the clinical interpretation of variants.


Assuntos
Biomarcadores Tumorais , Sequenciamento de Nucleotídeos em Larga Escala , Síndromes Neoplásicas Hereditárias/genética , Biologia Computacional/métodos , Variações do Número de Cópias de DNA , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Testes Genéticos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequenciamento de Nucleotídeos em Larga Escala/normas , Humanos , Mutação INDEL , Masculino , Mutação , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
7.
Gene ; 637: 41-49, 2017 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-28919163

RESUMO

Alternative pre-mRNA splicing increases transcriptome plasticity by forming naturally-occurring alternative splicing variants (ASVs). Alterations of splicing processes, caused by DNA mutations, result in aberrant splicing and the formation of aberrant mRNA isoforms. Analyses of hereditary cancer predisposition genes reveal many DNA variants with unknown clinical significance (VUS) that potentially affect pre-mRNA splicing. Therefore, a comprehensive description of ASVs is an essential prerequisite for the interpretation of germline VUS in high-risk individuals. To identify ASVs in a gene of interest, we have proposed an approach based on multiplex PCR (mPCR) amplification of all theoretically possible exon-exon junctions and subsequent characterization of size-selected and pooled mPCR products by next-generation sequencing (NGS). The efficiency of this method is illustrated by a comprehensive analysis of BRCA1 ASVs in human leukocytes, normal mammary, and adipose tissues and stable cell lines. We revealed 94 BRCA1 ASVs, including 29 variants present in all tested samples. While differences in the qualitative expression of BRCA1 ASVs among the analyzed human tissues were minor, larger differences were detected between tissue and cell line samples. Compared with other ASV analysis methods, this approach represents a highly sensitive and rapid alternative for the identification of ASVs in any gene of interest.


Assuntos
Processamento Alternativo , Proteína BRCA1/genética , Neoplasias da Mama/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Reação em Cadeia da Polimerase Multiplex/métodos , Mutação , Biologia Computacional , Feminino , Humanos , Isoformas de RNA
8.
Gene ; 587(2): 169-72, 2016 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-27150568

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is the sixth most frequent cancer type in the Czech Republic with a poor prognosis that could be improved by an early detection and subsequent surgical treatment combined with chemotherapy. Genetic factors play an important role in PDAC risk. We previously identified one PDAC patient harboring the Slavic founder deleterious mutation c.657del5 in the NBN gene, using a panel next-generation sequencing (NGS). A subsequent analysis of 241 unselected PDAC patients revealed other mutation carriers. The overall frequency of c.657del5 in unselected PDAC patients (5/241; 2.07%) significantly differed from that in non-cancer controls (2/915; 0.2%; P=0.006). The result indicates that the NBN c.657del5 variant represents a novel PDAC-susceptibility allele increasing PDAC risk (OR=9.7; 95% CI: 1.9 to 50.2). The increased risk of PDAC in follow-up recommendations for NBN mutation carriers should be considered if other studies also confirm an increased frequency of c.657del5 carriers in PDAC patients from other populations.


Assuntos
Adenocarcinoma/genética , Carcinoma Ductal Pancreático/genética , Proteínas de Ciclo Celular/genética , Deleção de Genes , Proteínas Nucleares/genética , Neoplasias Pancreáticas/genética , Tchecoslováquia , Feminino , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Linhagem
9.
Small ; 12(11): 1432-9, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26797691

RESUMO

Chemically functional core/shell microtubes made of biodegradable polymers are fabricated using coaxial electrospinning. The luminal walls are chemically functionalized, allowing for regioselective chemical binding or adsorption inside the microtube. Attaching catalytic nanoparticles or enzymes to the luminal walls converts the microtubes into bubble-propelled microrockets. Upon exposure to ultrasound, the microtubes undergo shape shifting, transforming them into picoliter-scale containers.


Assuntos
Nanopartículas/química , Nanotecnologia/métodos , Polímeros/química , Ácido Láctico/química , Microscopia Confocal , Nanopartículas/ultraestrutura , Polietilenoglicóis/química , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Prata/química , Imagem com Lapso de Tempo
10.
Food Chem Toxicol ; 85: 20-30, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26518667

RESUMO

Commercially manufactured nanomaterials are used massively for modification of products of everyday use, including products intended for children. Therefore their potential risks have to be ultimately studied. Aside from toxicity of nanomaterials with known specific parameters, the end-consumer is potentially endangered by materials with unknown specification. Commercially available products are not usually accompanied by parameter/specification sheet providing the consumer with sufficient chemico-physical parameters allowing the evaluation of possible toxic effects. The aim of this work was to evaluate the declared parameters of commercially available TiO2 and Ag NPs employing chemico-physical methods and consequently in vitro cytotoxicity and genotoxicity tests performed on non-cancer cell lines. Based on the results of our complex study we can conclude that the data provided by the producers are not in good agreement with the performed measurements. Furthermore, all tested NPs penetrated into the SVK14 cells and all NPs had significant effect on the kinetics of ROS production in all cell lines (note: the ROS production has not been established as the major mechanism of cell damage elicited by Ag NPs). The study revealed greater cytotoxic potential of Ag NPs in comparison with TiO2 NPs and all of the studied NPs caused significant DNA damage.

11.
PLoS One ; 10(6): e0127711, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26057125

RESUMO

Recent studies have conferred that the RAD51C and RAD51D genes, which code for the essential proteins involved in homologous recombination, are ovarian cancer (OC) susceptibility genes that may explain genetic risks in high-risk patients. We performed a mutation analysis in 171 high-risk BRCA1 and BRCA2 negative OC patients, to evaluate the frequency of hereditary RAD51C and RAD51D variants in Czech population. The analysis involved direct sequencing, high resolution melting and multiple ligation-dependent probe analysis. We identified two (1.2%) and three (1.8%) inactivating germline mutations in both respective genes, two of which (c.379_380insG, p.P127Rfs*28 in RAD51C and c.879delG, p.C294Vfs*16 in RAD51D) were novel. Interestingly, an indicative family cancer history was not present in four carriers. Moreover, the ages at the OC diagnoses in identified mutation carriers were substantially lower than those reported in previous studies (four carriers were younger than 45 years). Further, we also described rare missense variants, two in RAD51C and one in RAD51D whose clinical significance needs to be verified. Truncating mutations and rare missense variants ascertained in OC patients were not detected in 1226 control samples. Although the cumulative frequency of RAD51C and RAD51D truncating mutations in our patients was lower than that of the BRCA1 and BRCA2 genes, it may explain OC susceptibility in approximately 3% of high-risk OC patients. Therefore, an RAD51C and RAD51D analysis should be implemented into the comprehensive multi-gene testing for high-risk OC patients, including early-onset OC patients without a family cancer history.


Assuntos
Proteínas de Ligação a DNA/genética , Genes Neoplásicos , Predisposição Genética para Doença , Neoplasias Ovarianas/genética , Proteína BRCA1/genética , Proteína BRCA2/genética , Sequência de Bases , República Tcheca , Análise Mutacional de DNA , DNA Complementar/genética , Éxons/genética , Família , Feminino , Humanos , Íntrons/genética , Dados de Sequência Molecular , Fatores de Risco
12.
Food Chem Toxicol ; 82: 106-15, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25846500

RESUMO

Commercially manufactured nanomaterials are used massively for modification of products of everyday use, including products intended for children. Therefore their potential risks have to be ultimately studied. Aside from toxicity of nanomaterials with known specific parameters, the end-consumer is potentially endangered by materials with unknown specification. Commercially available products are not usually accompanied by parameter/specification sheet providing the consumer with sufficient chemico-physical parameters allowing the evaluation of possible toxic effects. The aim of this work was to evaluate the declared parameters of commercially available TiO2 and Ag NPs employing chemico-physical methods and consequently in vitro cytotoxicity and genotoxicity tests performed on non-cancer cell lines. Based on the results of our complex study we can conclude that the data provided by the producers are not in good agreement with the performed measurements. Furthermore, all tested NPs penetrated into the SVK14 cells and all NPs had significant effect on the kinetics of ROS production in all cell lines (note: the ROS production has not been established as the major mechanism of cell damage elicited by Ag NPs). The study revealed greater cytotoxic potential of Ag NPs in comparison with TiO2 NPs and all of the studied NPs caused significant DNA damage.


Assuntos
Nanopartículas Metálicas/toxicidade , Prata/toxicidade , Titânio/toxicidade , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular/efeitos dos fármacos , Linhagem Celular/metabolismo , Ensaio Cometa , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Nanopartículas Metálicas/química , Camundongos , Microscopia de Força Atômica , Células NIH 3T3/efeitos dos fármacos , Tamanho da Partícula , Espécies Reativas de Oxigênio/metabolismo , Prata/química , Prata/farmacocinética , Espectrofotometria Atômica , Análise Espectral Raman , Titânio/farmacocinética
13.
PLoS One ; 9(8): e103675, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25098570

RESUMO

The potential for application of any nanoparticles, including silver nanoparticles (AgNPs), is strongly dependent on their stability against aggregation. Therefore, improvement of this parameter is a key task, especially in the case of AgNPs, because a correlation between size and biological activity has been demonstrated. In the present work, a natural stabilizer, gelatin, was investigated for the stabilization of AgNPs in an aqueous dispersion. The particles were prepared via a modified Tollens process, and the gelatin modifier was added prior to the reducing agent. The stability against aggregation of the AgNPs prepared by this method was more than one order of magnitude higher (on the basis of the critical coagulation concentration (CCC)) than that of AgNPs prepared via a similar method but without the assistance of gelatin. Their high stability against aggregation was confirmed over wide pH range (from 2 to 13) in which the particles did not exhibit rapid aggregation; such stability has not been previously reported for AgNPs. Additionally, gelatin not only fulfills the role of a unique stabilizer but also positively influences the modified Tollens process used to prepare the AgNPs. The diameter of the gelatin-modified AgNPs was substantially smaller in comparison to those prepared without gelatin. The polydispersity of the dispersion significantly narrowed. Moreover, the gelatin-stabilized AgNPs exhibited long-term stability against aggregation and maintained high antibacterial activity when stored for several months under ambient conditions.


Assuntos
Antibacterianos , Bactérias/crescimento & desenvolvimento , Gelatina , Nanopartículas Metálicas/química , Prata , Antibacterianos/química , Antibacterianos/farmacologia , Estabilidade de Medicamentos , Gelatina/química , Gelatina/farmacologia , Concentração de Íons de Hidrogênio , Prata/química , Prata/farmacologia , Fatores de Tempo
14.
Cancer Epidemiol Biomarkers Prev ; 22(12): 2323-32, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24136930

RESUMO

BACKGROUND: Several reports indicate that inherited mutations in the PALB2 gene predispose to breast cancer. However, there is little agreement about the clinical relevance and usefulness of mutation screening in this gene. We analyzed the prevalence and spectrum of germline mutations in PALB2 to estimate their contribution to hereditary breast and/or ovarian cancer in the Czech Republic. METHODS: The entire PALB2 coding region was sequenced in 409 breast/ovarian cancer patients negative for BRCA1 and BRCA2 mutations. Testing for large genomic rearrangements (LGR) was performed by multiplex ligation-dependent probe amplification (MLPA) analysis. RESULTS: We have identified 13 different pathogenic alterations including 10 truncating mutations and three LGRs in 16 of 409 patients (3.9%), whereas one truncating mutation was found in a group of 1,226 controls (0.08%; P = 2.6 × 10(-9)). Three novel LGRs included deletions involving exons 7-8 and 9-10, respectively, and a duplication spanning exons 9-11. Five frameshift and two nonsense mutations were novel, whereas three truncating mutations were described previously. The only recurrent mutation was the c.172_175delTTGT detected in four unrelated breast cancer individuals. CONCLUSIONS: Our analyses demonstrated the significant role of the PALB2 gene in breast cancer susceptibility. The highest frequency of PALB2 mutations (comparable with that previously reported for BRCA2) was found in a subgroup of patients with hereditary breast cancer (HBC) (13/235; 5.5%). IMPACT: Our results show that mutation analysis of the PALB2 gene, including the analysis of LGRs, is primarily indicated in patients with HBC in case of their BRCA1 and BRCA2 negativity.


Assuntos
Neoplasias da Mama/genética , Genes BRCA1 , Genes BRCA2 , Proteínas Nucleares/genética , Proteínas Supressoras de Tumor/genética , Sequência de Bases , Neoplasias da Mama/patologia , Estudos de Casos e Controles , Análise Mutacional de DNA/métodos , Proteína do Grupo de Complementação N da Anemia de Fanconi , Feminino , Deleção de Genes , Predisposição Genética para Doença , Humanos , Dados de Sequência Molecular , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Linhagem , Prevalência , Fatores de Risco
16.
Neuromolecular Med ; 13(3): 204-11, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21833744

RESUMO

Ataxia telangiectasia (AT) is a genomic instability syndrome characterised, among others, by progressive cerebellar degeneration, oculocutaneous telangiectases, immunodeficiency, elevated serum alpha-phetoprotein level, chromosomal breakage, hypersensitivity to ionising radiation and increased cancer risk. This autosomal recessive disorder is caused by mutations in the ataxia telangiectasia mutated (ATM) gene coding for serine/threonine protein kinase with a crucial role in response to DNA double-strand breaks. We characterised genotype and phenotype of 12 Slavic AT patients from 11 families. Mutation analysis included sequencing of the entire coding sequence, adjacent intron regions, 3'UTR and 5'UTR of the ATM gene and multiplex ligation-dependent probe amplification (MLPA) for the detection of large deletions/duplications at the ATM locus. The high incidence of new and individual mutations demonstrates a marked mutational heterogeneity of AT in the Czech Republic. Our data indicate that sequence analysis of the entire coding region of ATM is sufficient for a high detection rate of mutations in ATM and that MLPA analysis for the detection of deletions/duplications seems to be redundant in the Slavic population.


Assuntos
Ataxia Telangiectasia/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ligação a DNA/genética , Mutação , Proteínas Serina-Treonina Quinases/genética , Proteínas Supressoras de Tumor/genética , Adolescente , Proteínas Mutadas de Ataxia Telangiectasia , Criança , Pré-Escolar , República Tcheca , Análise Mutacional de DNA , Feminino , Humanos , Masculino
17.
J Cancer Res Clin Oncol ; 137(2): 331-8, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20422428

RESUMO

PURPOSE: Carriers of BRCA1/2 mutations are at high lifetime risk of breast cancer (BC); however, the BC onset broadly vary in individual patients. Recently, polyglutamine (poly-Q) repeat length polymorphism of the amplified in breast cancer 1 (AIB1) gene was analyzed as a risk factor influencing BC onset in BRCA1/2 mutation carriers with contradictory results. METHODS: We genotyped AIB1 poly-Q repeat in 243 BRCA1/2 mutation carriers, 61 patients with familial BC (negatively tested for the presence of BRCA1/2 mutation), 221 patients with sporadic BC, and 176 non-cancer controls using denaturing high-performance liquid chromatography and statistically evaluated the effect of AIB1 poly-Q repeat length polymorphism on BC onset. RESULTS: Having used previously published statistical analyses of AIB1 poly-Q repeat length (≥28 and ≥29 repeat cutpoints or analysis of AIB1 poly-Q repeat length as continuous variable), we did not find any association between AIB1 poly-Q repeat length and BC development in analyzed BC groups. However, the analysis of individual genotypes revealed that AIB1 genotype consisting of 28/28 glutamine repeats served as a protective factor in BRCA1 mutation carriers (HR = 0.64; 95% CI 0.41-0.99; P = 0.045) and as a risk factor in carriers of mutation in exon 11 of the BRCA2 gene (HR = 3.50; 95% CI 1.25-9.78; P = 0.017). CONCLUSIONS: Our results confirm that AIB1 poly-Q repeat length polymorphism does not influence the BC risk in general but suggest that the specific AIB1 genotypes should be considered in patients with BC carrying mutation in the BRCA1/2 genes.


Assuntos
Neoplasias da Mama/genética , Coativador 3 de Receptor Nuclear/genética , Peptídeos/genética , Polimorfismo Genético , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/mortalidade , República Tcheca , Feminino , Genes BRCA1 , Genes BRCA2 , Predisposição Genética para Doença , Genótipo , Heterozigoto , Humanos , Pessoa de Meia-Idade , Mutação , Modelos de Riscos Proporcionais , Medição de Risco , Fatores de Risco , Análise de Sobrevida
18.
Biomaterials ; 30(31): 6333-40, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19698988

RESUMO

The antifungal activity of the silver nanoparticles (NPs) prepared by the modified Tollens process was evaluated for pathogenic Candida spp. by means of the determination of the minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC), and the time-dependency of yeasts growth inhibition. Simultaneously the cytotoxicity of the silver NPs to human fibroblasts was determined. The silver NPs exhibited inhibitory effect against the tested yeasts at the concentration as low as 0.21 mg/L of Ag. The inhibitory effect of silver NPs was enhanced through their stabilization and the lowest MIC equal to 0.05 mg/L was determined for silver NPs stabilized by sodium dodecyl sulfate against Candida albicans II. The obtained MICs of the silver NPs and especially of the stabilized silver NPs were comparable and in some cases even better than MICs of the conventional antifungal agents determined by E-test. The silver NPs effectively inhibited the growth of the tested yeasts at the concentrations below their cytotoxic limit against the tested human fibroblasts determined at a concentration equal to 30 mg/L of Ag. In contrast, ionic silver inhibited the growth of the tested yeasts at the concentrations comparable to the cytotoxic level (approx. 1mg/L) of ionic silver against the tested human fibroblasts.


Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Nanopartículas Metálicas , Prata/farmacologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Nanopartículas Metálicas/uso terapêutico
19.
Nanotechnology ; 20(2): 028001, 2009 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-19417286

RESUMO

The fabrication of nanocomposite materials with a metal-oxide@nobel-metal structure represents a challenge both from the theoretical and experimental points of view. This challenge is connected with a complicated mechanism of formation of these nanocomposite materials. In the paper by Gong et al (2007 Nanotechnology 18 285604), the authors tried to solve this problem but the suggested mechanism is rather improbable. This improbability derived from the found discrepancy in the material balance of reactants/products. Therefore this comment is devoted to evaluation of this discrepancy of the Fe(3)O(4)@Ag nanoparticle preparation and suggestion of another possible mechanism.


Assuntos
Antibacterianos/farmacologia , Óxido Ferroso-Férrico/farmacologia , Ouro/farmacologia , Nanopartículas , Óxido Ferroso-Férrico/química , Ouro/química
20.
Oncol Rep ; 19(6): 1505-10, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18497957

RESUMO

Mutations in the ATM gene are the cause of a rare autosomal recessive syndrome, ataxia-telangiectasia (AT). Of the general population, approximately 0.35-1% has been estimated to be heterozygous for a germline mutation in the ATM gene. The finding that ATM heterozygotes have an increased breast cancer risk was supported by some studies but not confirmed by others. In our study, the entire coding sequence of the ATM gene was prescreened for mutations by the protein truncation test to detect the chain-terminating mutations that are highly predominant in patients with AT. DNA sequencing then characterized 3 (1.9%) pathogenic mutations among 161 high-risk breast cancer patients. The c.5177+1G>A splicing mutation was a novel gene alteration. No mutation was detected in a group of 183 control individuals. Our results suggest that truncating mutations in ATM increase breast cancer risk and contribute to inherited breast cancer. The analysis further uncovered the c.1066-6T>G splicing mutation once among high-risk patients (0.6%) and twice among controls (1.1%) suggesting that this mutation does not confer an increase in breast cancer risk. On the other hand, individuals heterozygous for this truncating variant displayed loss of exon 11 in approximately 50% of ATM transcripts. Immunohistochemistry did not detect the ATM protein in the tumor sample carrying this mutation. Thus, the association of the c.1066-6T>G mutation with familial breast cancer remains uncertain. Loss of the wild-type ATM allele has not been detected in the tumor samples from heterozygous carriers of the ATM mutation. Our experiments did not detect the hypermethylation of the ATM promoter in any of the DNA samples from tumor tissues.


Assuntos
Neoplasias da Mama/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ligação a DNA/genética , Predisposição Genética para Doença , Mutação/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Supressoras de Tumor/genética , Adulto , Idade de Início , Processamento Alternativo , Proteínas Mutadas de Ataxia Telangiectasia , Neoplasias da Mama/epidemiologia , Neoplasias da Mama/patologia , Proteínas de Ciclo Celular/metabolismo , Tchecoslováquia/epidemiologia , Metilação de DNA , Análise Mutacional de DNA , Proteínas de Ligação a DNA/metabolismo , Feminino , Heterozigoto , Humanos , Técnicas Imunoenzimáticas , Perda de Heterozigosidade , Pessoa de Meia-Idade , Regiões Promotoras Genéticas/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Supressoras de Tumor/metabolismo
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