RESUMO
BACKGROUND: BRCA1 and BRCA2 germline mutations predispose heterozygous carriers to hereditary breast/ovarian cancer. However, unclassified variants (UVs) (variants with unknown clinical significance) and missense polymorphisms in BRCA1 and BRCA2 genes pose a problem in genetic counseling, as their impact on risk of breast and ovarian cancer is still unclear. The objective of our study was to identify UVs and missense polymorphisms in Algerian breast/ovarian cancer patients and relatives tested previously for BRCA1 and BRCA2 genes germline mutations analysis. METHODS: We analyzed 101 DNA samples from 79 breast/ovarian cancer families. The approach used is based on BRCA1 and BRCA2 sequence variants screening by SSCP or High-Resolution Melting (HRM) curve analysis followed by direct sequencing. In silico analyses have been performed using different bioinformatics programs to individualize genetics variations that can disrupt the BRCA1 and BRCA2 genes function. RESULTS: Among 80 UVs and polymorphisms detected in BRCA1/2 genes (33 BRCA1 and 47 BRCA2), 31 were new UVs (10 BRCA1 and 21 BRCA2), 7 were rare UVs (4 BRCA1 and 3 BRCA2) and 42 were polymorphic variants (19 BRCA1 and 23 BRCA2). Moreover, 8 new missense UVs identified in this study: two BRCA1 (c.4066C>A/p.Gln1356Lys, c.4901G>T/p.Arg1634Met) located respectively in exons 11 and 16, and six BRCA2 (c.1099G>A/p.Asp367Asn, c.2636C>A/p.Ser879Tyr, c.3868T>A/p.Cys1290Ser, c.5428G>T/p.Val1810Phe, c.6346C>G/p.His2116Asp and c.9256G>A/p.Gly3086Arg) located respectively in exons 10, 11 and 24, show a damaging PSIC score yielded by PolyPhen2 program and could be pathogenic. In addition, 5 new BRCA} missense UVs out of six that were found to be damaging by PolyPhen2 program, also were deleterious according to SIFT program. The rare BRCA1 UV c.5332G>A/p.Asp1778Asn was found here for the first time in co-occurrence in trans with the deleterious BRCA1 mutation c.798_799delTT/p.Ser267LysfsX19 in young breast cancer patient. Moreover, 10 new identified intronic variants with unknown clinical significance (3 BRCA1 and 7 BRCA2) in the present study, could be considered as benign, because GeneSplicer, SpliceSiteFinder and MaxEntScan prediction programs show no splice site alteration for these variants. Several missense polymorphisms of BRCA1 c.2612C>T/p.Pro871Leu, c.3548A>G/p.Lys1183Arg, c.4837A>G/p.Ser1613Gly and BRCA2 c.865A>C/p.Asn289His, c.1114A>C/p.Asn372His, c.2971A>G/p.Asn991Asp, c.7150C>A/p.Gly2384Lys have been identified with high frequency in patients who were tested negative for BRCA1 and BRCA2 mutations. These missense polymorphisms could have a role as susceptibility breast cancer markers in Algerian breast/ovarian cancer families where pathological BRCA1 and BRCA2 mutations were not present. CONCLUSIONS: For the first time, UVs and missense polymorphisms in BRCA1 and BRCA2 genes have been identified in Algerian breast/ovarian cancer families. Evaluation of breast/ovarian cancer risk induced by the eight new missense UVs and common polymorphisms detected in our present work is on going in a larger study.
Assuntos
Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias da Mama/genética , Genes Neoplásicos , Mutação de Sentido Incorreto , Neoplasias Ovarianas/genética , Adolescente , Adulto , Idoso , Argélia/epidemiologia , Neoplasias da Mama/epidemiologia , Biologia Computacional , Éxons , Feminino , Predisposição Genética para Doença , Testes Genéticos , Genoma Humano , Mutação em Linhagem Germinativa , Projeto HapMap , Humanos , Pessoa de Meia-Idade , Neoplasias Ovarianas/epidemiologia , Linhagem , Polimorfismo de Nucleotídeo Único , Adulto JovemRESUMO
The c.156_157insAlu BRCA2 mutation has so far only been reported in hereditary breast/ovarian cancer (HBOC) families of Portuguese origin. Since this mutation is not detectable using the commonly used screening methodologies and must be specifically sought, we screened for this rearrangement in a total of 5,443 suspected HBOC families from several countries. Whereas the c.156_157insAlu BRCA2 mutation was detected in 11 of 149 suspected HBOC families from Portugal, representing 37.9% of all deleterious mutations, in other countries it was detected only in one proband living in France and in four individuals requesting predictive testing living in France and in the USA, all being Portuguese immigrants. After performing an extensive haplotype study in carrier families, we estimate that this founder mutation occurred 558 ± 215 years ago. We further demonstrate significant quantitative differences regarding the production of the BRCA2 full length RNA and the transcript lacking exon 3 in c.156_157insAlu BRCA2 mutation carriers and in controls. The cumulative incidence of breast cancer in carriers did not differ from that of other BRCA2 and BRCA1 pathogenic mutations. We recommend that all suspected HBOC families from Portugal or with Portuguese ancestry are specifically tested for this rearrangement.
Assuntos
Neoplasias da Mama/epidemiologia , Neoplasias da Mama/genética , Genes BRCA2 , Mutação , Neoplasias Ovarianas/epidemiologia , Neoplasias Ovarianas/genética , Sequência de Aminoácidos , Feminino , Efeito Fundador , Predisposição Genética para Doença , Testes Genéticos , Genética Populacional , Humanos , Repetições de Microssatélites , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Portugal/epidemiologia , RNA Mensageiro/análise , Fases de Leitura/genética , Deleção de SequênciaRESUMO
BACKGROUND: Breast cancer is the leading cause of cancer death in women in Algeria. The contribution of BRCA1 and BRCA2 mutations to hereditary breast/ovarian cancer in Algerian population is largely unknown. Here, we describe analysis of BRCA1 and BRCA2 genes in 86 individuals from 70 families from an Algerian cohort with a personal and family history suggestive of genetic predisposition to breast cancer. METHODS: The approach used is based on BRCA1 and BRCA2 mutations screening by High-Resolution Melting (HRM) curve analysis followed by direct sequencing. All samples for which no pathogenic mutation was found were analyzed by MLPA for large deletions or duplications. RESULTS: Three distinct pathogenic mutations c.83_84delTG, c.181T>G, c.798_799delTT and two large rearrangements involving deletion of exon 2 and exon 8 respectively, were detected in BRCA1 gene. Moreover 17 unclassified variants and polymorphisms were detected in BRCA1 gene (6 described for the first time). Two pathogenic mutations, c.1310_1313delAAGA and c.5722_5723delCT and 40 unclassified variants and polymorphisms (14 never described before) were identified in BRCA2 gene. CONCLUSIONS: For the first time, we used HRM and MLPA to identify BRCA1 and BRCA2 mutations in Algerian patients with a personal and family history suggestive of genetic predisposition to breast cancer. The implications of these new findings in regard to genetic testing and counseling are substantial for the Algerian population.
Assuntos
Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias da Mama/genética , Mutação em Linhagem Germinativa/genética , Programas de Rastreamento , Neoplasias Ovarianas/genética , Adulto , Argélia , Proteínas Reguladoras de Apoptose , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/epidemiologia , DNA de Neoplasias/genética , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neoplasias Ovarianas/diagnóstico , Neoplasias Ovarianas/epidemiologia , Reação em Cadeia da Polimerase , Polimorfismo Genético , Prognóstico , Deleção de Sequência , Taxa de SobrevidaRESUMO
Background Around half of familial breast cancer cases are caused by germ-line mutations in genes which are critically involved in the maintenance of genome stability. Mutations in related genes functioning in DNA repair may account for currently unattributed cases. Two such genes, RAP80 and Abraxas, have recently been identified to be in a complex with BRCA1, and are required for the localization of BRCA1 to DNA damage foci. Methods RAP80 and Abraxas variants were screened for in a cohort of 95 high risk, non-BRCA1/2 breast cancer cases of varying ethnicity: those of Ashkenazi Jewish (n = 35), mixed Canadian (n = 34) and Swiss descent (n = 26). Results We have identified four missense variants, four silent SNPs, three SNPs in the UTRs and seven intronic variants in RAP80. Two of the previously reported RAP80 variants were further investigated. In Abraxas, we have identified two missense, nine intronic and two variants in the 3' UTR. Conclusions Overall, it seems unlikely that moderate to highly penetrant alleles of either RAP80 or Abraxas, confer a significantly high relative risk of breast cancer.
Assuntos
Neoplasias da Mama/genética , Proteínas de Transporte/genética , Predisposição Genética para Doença , Proteínas Nucleares/genética , Neoplasias da Mama/etnologia , Análise Mutacional de DNA , Proteínas de Ligação a DNA , Feminino , Genes BRCA1 , Genes BRCA2 , Chaperonas de Histonas , Humanos , Judeus/genética , Masculino , Linhagem , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Polimorfismo Conformacional de Fita Simples , Fatores de RiscoRESUMO
ATM gene alterations have been described in various lymphoproliferative malignancies suggesting that ATM contributes to lymphomagenesis. Using multiplex ligation-dependant probe amplification (MLPA), we screened 61 childhood lymphoid malignancies for ATM genomic deletion/duplication. Five samples were found to have a complete deletion or duplication. All the three deletions were found in B-precursor ALL (15%), two were submicroscopic, not detected by standard cytogenetic studies. These observations indicate that as in adult ALL, complete ATM submicroscopic deletion is frequent in childhood B-precursor ALL. As previously hypothesized, these results suggest that ATM may act as a tumor suppressor gene in the pathogenesis of childhood B-precursor ALL.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Ligação a DNA/genética , Deleção de Genes , Duplicação Gênica , Linfoma/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Supressoras de Tumor/genética , Proteínas Mutadas de Ataxia Telangiectasia , Criança , Amplificação de Genes , Dosagem de Genes , Humanos , IoimbinaRESUMO
BRCA1 and BRCA2 are the major genes predisposing to breast-ovarian cancer (i.e., breast or ovarian cancer or both). Since 1994, hundreds of distinct germline alterations have been reported in these two genes. Besides pathogenic mutations resulting in loss of function of the protein, an increased number of variants of unknown clinical significance have been described. In a cohort of 350 Swiss breast-ovarian cancer families, the systematic search for BRCA1/BRCA2 germline mutations was carried out using denaturating high-performance liquid chromatography as the first screening procedure. The screening strategy resulted in the identification of 23 alterations not previously reported: 9 in BRCA1 and 14 in BRCA2. By using the available tools to assign a functional role to newly identified sequence variations, 5 (22%) of these were classified as new disease-causing mutations, 5 (22%) were classified as benign polymorphisms, and the remaining 13 (56%) alterations were considered as unclassified variants. These data illustrate the major challenge for clinical oncologists currently facing the interpretation of alterations identified in BRCA1 or BRCA2. The key points are to classify these genetic variations as pathogenic mutations, benign polymorphisms, or variants of unknown clinical significance and to adequately use this information for the management of high-risk individuals and their families.
Assuntos
Neoplasias da Mama/genética , Genes BRCA1 , Genes BRCA2 , Neoplasias Ovarianas/genética , Adulto , Estudos de Coortes , Feminino , Humanos , Pessoa de Meia-Idade , Mutação de Sentido IncorretoRESUMO
ATM gene alterations and impaired ATM protein expression have been described in various adult lymphoproliferative malignancies, suggesting that ATM contributes to lymphomagenesis. The present study investigated the prevalence of ATM gene and ATM protein expression alterations in sporadic childhood non-Hodgkin lymphoma (NHL). Twenty-seven cases of NHL were screened for ATM mutations by denaturing high-performance liquid chromatography (DHPLC). Direct and indirect criteria, including in silico tools, were used to classify the gene alterations. The methylation status of the ATM promoter CpG island was determined in 25 samples; ATM protein expression was assessed by Western blot in 9 lymphomas. ATM alterations were detected in 12 NHLs (44%). Ten different heterozygous base substitutions were identified in 10 NHLs (37%). Five samples (19%) were found to harbor a gene alteration considered to be a mutation or a rare variant potentially pathogenic. In one case, an ATM mutation was found in the germline. Four NHLs (44%) showed reduced or absent ATM protein expression. Except for one sample, no definite genetic or epigenetic alteration was identified to account for impaired ATM protein expression. These observations document a high prevalence of ATM gene and protein expression alterations, suggesting that ATM is involved in childhood NHL.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Ligação a DNA/genética , Linfoma não Hodgkin/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Supressoras de Tumor/genética , Adolescente , Proteínas Mutadas de Ataxia Telangiectasia , Estudos de Casos e Controles , Criança , Pré-Escolar , Metilação de DNA , Análise Mutacional de DNA , Feminino , Humanos , Masculino , Mutação/genética , Polimorfismo Genético , Regiões Promotoras Genéticas/genéticaRESUMO
Ataxia telangiectasia is a hereditary pleiomorphic syndrome caused by loss of Atm, a phosphoprotein involved in multiple signaling pathways. Here, we propose a novel role for atm in cultured epithelial cells, namely the regulation of cell growth by contact inhibition. We show that atm is upregulated in epithelial cells reaching confluence. Conditional expression of the PI 3-Kinase domain of atm in non-confluent Tac-2 epithelial cells increases the expression of the anti-proliferative gene Tis-21 and downregulates key cell cycle regulator genes, such as cyclins A, B1, B2, E and E2. Finally, we demonstrate that upregulation of atm, and thus Tis-21, in confluent Tac-2 cells can be inhibited by an E-cadherin antibody blocking specifically homophilic E-cadherin interactions between adjacent cell surfaces. Altogether, these results suggest that atm could participate in a molecular pathway linking extracellular signalling to cell cycle control and may help further clarify the role of Atm in epithelial cell biology and carcinogenesis.
Assuntos
Caderinas/metabolismo , Adesão Celular , Proteínas de Ciclo Celular/fisiologia , Proliferação de Células , Inibição de Contato , Proteínas de Ligação a DNA/fisiologia , Células Epiteliais/citologia , Glândulas Mamárias Animais/citologia , Proteínas Serina-Treonina Quinases/fisiologia , Proteínas Supressoras de Tumor/fisiologia , Animais , Ataxia Telangiectasia , Proteínas Mutadas de Ataxia Telangiectasia , Ciclo Celular , Células Cultivadas , Ciclinas/metabolismo , Células Epiteliais/metabolismo , Feminino , Células HeLa/metabolismo , Humanos , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Transcrição GênicaRESUMO
ATM promoter hypermethylation has been recently reported in adult carcinomas, but no information is available concerning the methylation status of ATM gene promoter in childhood B-precursor acute lymphoblastic leukaemia (ALL). Furthermore, involvement of somatic ATM promoter mutations in cancer is not known. We report a complete ATM gene promoter analysis in 74 childhood lymphoid malignancies.
Assuntos
Linfoma de Burkitt/genética , Proteínas de Ciclo Celular/genética , Metilação de DNA , Proteínas de Ligação a DNA/genética , Mutação Puntual , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Regiões Promotoras Genéticas/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Supressoras de Tumor/genética , Adulto , Proteínas Mutadas de Ataxia Telangiectasia , Carcinoma/genética , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Análise de Sequência de DNARESUMO
In 10-30% of patients with classical familial adenomatous polyposis (FAP) and up to 90% of those with attenuated (<100 colorectal adenomas; AFAP) polyposis, no pathogenic germline mutation in the adenomatous polyposis coli (APC) gene can be identified (APC mutation-negative). Recently, biallelic mutations in the base excision repair gene MYH have been shown to predispose to a multiple adenoma and carcinoma phenotype. This study aimed to (i) assess the MYH mutation carrier frequency among Swiss APC mutation-negative patients and (ii) identify phenotypic differences between MYH mutation carriers and APC/MYH mutation-negative polyposis patients. Seventy-nine unrelated APC mutation-negative Swiss patients with either classical (n=18) or attenuated (n=61) polyposis were screened for germline mutations in MYH by dHPLC and direct genomic DNA sequencing. Overall, 7 (8.9%) biallelic and 9 (11.4%) monoallelic MYH germline mutation carriers were identified. Among patients with a family history compatible with autosomal recessive inheritance (n=45), 1 (10.0%) out of 10 classical polyposis and 6 (17.1%) out of 35 attenuated polyposis patients carried biallelic MYH alterations, 2 of which represent novel gene variants (p.R171Q and p.R231H). Colorectal cancer was significantly (p<0.007) more frequent in biallelic mutation carriers (71.4%) compared with that of monoallelic and MYH mutation-negative polyposis patients (0 and 13.8%, respectively). On the basis of our findings and earlier reports, MYH mutation screening should be considered if all of the following criteria are fulfilled: (i) presence of classical or attenuated polyposis coli, (ii) absence of a pathogenic APC mutation, and (iii) a family history compatible with an autosomal recessive mode of inheritance.
Assuntos
Polipose Adenomatosa do Colo/genética , Neoplasias Colorretais/genética , DNA Glicosilases/genética , Genes APC , Predisposição Genética para Doença , Mutação em Linhagem Germinativa , Adulto , Idoso , Estudos de Casos e Controles , Análise Mutacional de DNA , Reparo do DNA , Feminino , Humanos , Padrões de Herança , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , SuíçaRESUMO
4.1R pre-mRNA alternative splicing results in multiple mRNA and protein isoforms that are expressed in virtually all tissues. More specifically, isoforms containing the alternative exon 17a, are exclusively expressed in muscle tissues. In this report, we show that these isoforms are preferentially present in the myoplasm of fast myofibres. 4.1R epitopes are also found at the sarcolemma of both slow and fast myofibres in normal muscle. Interestingly, they are absent from dystrophin-deficient sarcolemma of DMD muscle, and colocalize with partially expressed dystrophin in BMD muscle. We also show that alternative splicing of exons 16 and 17a is regulated during muscle differentiation in an asynchronous fashion, with an early inclusion of exon 16 in forming myotubes, and a late inclusion of exon 17a. Consistently, Western blot analysis led to characterize mainly an approximately 96/98-kDa doublet bearing exons 16-17a-encoding peptide, exclusively occurring in the differentiated muscle.
Assuntos
Processamento Alternativo , Proteínas Sanguíneas/genética , Proteínas Associadas aos Microtúbulos/genética , Músculo Esquelético/metabolismo , Distrofias Musculares/genética , Sequência de Aminoácidos , Sequência de Bases , Proteínas Sanguíneas/metabolismo , Linhagem Celular , Proteínas do Citoesqueleto , DNA Complementar/genética , Humanos , Proteínas de Membrana , Proteínas Associadas aos Microtúbulos/metabolismo , Dados de Sequência Molecular , Distrofias Musculares/metabolismo , Precursores de RNA/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Hereditary ATM gene mutations cause ataxia-telangiectasia, a pleiotropic disorder associated with a high incidence of lymphoid malignancies. Acquired ATM alterations have been described in sporadic lymphoproliferative disorder suggesting that the ATM gene contributes to lymphomagenesis. To assess the prevalence of genomic ATM alterations in childhood acute lymphoblastic leukemias (ALL), we explored a series of 57 sporadic ALL cases (26 B-precursor ALL and 31 T-ALL) using DHPLC (Denaturing High-Performance Liquid Chromatography). We identified 28 distinct genomic ATM alterations in 14 patients (25%). Ten of them were scored as probably biologically significant and appear to be associated with a high risk of relapse (P<0.01). Six alterations of potential biological significance were observed in 5 cases of B-precursor ALL (19%), while 5 were found in 3 cases of T-ALL (10%). In two cases of B-precursor ALL, the ATM alterations were found in the germline, indicating an ATM carrier status. We report here the high prevalence of genomic ATM alterations in childhood ALL. Our observations lend further support to the postulated contribution of ATM in lymphomagenesis.