RESUMO
Normal brain tissue from 28 individuals and 50 glioma samples were analyzed by real-time Quantitative Methylation-Specific PCR (QMSP). Data from this analysis were compared with results obtained on the same samples by MSP. QMSP analysis demonstrated a statistically significant difference in both methylation level (P = .000009 Mann Whitney Test) and frequencies (P = .0000007, Z-test) in tumour samples as compared with normal brain tissues. Although QMSP and MSP showed similar sensitivity, the specificity of QMSP analysis was significantly higher (93%; CI95%: 84%-100%) as compared with MSP (64%; 95%CI: 46%-82%). Our results suggest that QMSP analysis may represent a powerful tool to identify glioma patients that will benefit from alkylating agents chemotherapy.
Assuntos
Metilação de DNA , Metilases de Modificação do DNA/genética , Enzimas Reparadoras do DNA/genética , Glioma/genética , Reação em Cadeia da Polimerase/métodos , Regiões Promotoras Genéticas , Proteínas Supressoras de Tumor/genética , Área Sob a Curva , Encéfalo/metabolismo , Química Encefálica , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Curva ROC , Sensibilidade e Especificidade , Estatísticas não ParamétricasRESUMO
In a series of invasive ductal breast carcinoma, we investigated the status of chromosomal and intrachromosomal instability by fluorescence in situ hybridisation and determined the level of mRNA expression for two genes involved in the mitotic spindle checkpoint pathway, BUB1B and MAD2L1. All breast cancers demonstrated higher chromosomal instability rates in tumor samples (average: 56.86%, range: 36.24-76.78%) than in controls (average: 11.54%, range: 9.91-14.84%) (P<0.0001). As well as intrachromosomal instability rates were elevated in tumor (average: 18.45% range: 8.34-35.8%) as compared with controls (average: 4.18% range: 3.47-4.81%) (P<0.0001). An increase in BUB1B and MAD2L1 transcripts was demonstrated in the majority of the tumor tested. BUB1B mRNA levels but not MAD2L1 levels correlated with intrachromosomal instability (r=0.722, P=0.018).
Assuntos
Neoplasias da Mama/genética , Proteínas de Ligação ao Cálcio/genética , Carcinoma Ductal/genética , Proteínas de Ciclo Celular/genética , Instabilidade Cromossômica , Instabilidade Genômica , Proteínas Serina-Treonina Quinases/genética , Proteínas Repressoras/genética , Adulto , Idoso , Neoplasias da Mama/patologia , Neoplasias da Mama/cirurgia , Carcinoma Ductal/patologia , Carcinoma Ductal/cirurgia , Cromossomos Humanos Par 13 , Cromossomos Humanos Par 17 , Cromossomos Humanos Par 8 , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Hibridização in Situ Fluorescente , Proteínas Mad2 , Pessoa de Meia-Idade , RNA Mensageiro/genéticaRESUMO
The HIC1 gene is a transcriptional regulator commonly methylated in a variety of human cancer. Thirty-three invasive ductal carcinomas of the breast and 21 matched normal breast tissues were analysed for HIC1 promoter methylation, and allelic loss of a 700 kb region spanning the gene locus. At least one genetic or epigenetic abnormality was found in 27 of the carcinomas tested (82%). Promoter methylation was demonstrated in 21 carcinomas (64%), and nine normal tissues (43%), whereas 18 malignant tumors (54%) showed allelic loss. Concomitant loss of heterozigosity and promoter hypermethylation in the region spanning HIC1 was detected in eight carcinomas (24%) suggesting that in this subset of tumors both copies of the gene are functionally lost. These observations support a role for the HIC1 gene in the pathogenesis of breast ductal carcinomas.
Assuntos
Neoplasias da Mama/genética , Carcinoma Ductal de Mama/genética , Metilação de DNA , Regiões Promotoras Genéticas , Fatores de Transcrição/genética , Deleção Cromossômica , Cromossomos Humanos Par 17/genética , Análise Mutacional de DNA , DNA de Neoplasias/análise , Proteínas de Ligação a DNA , Feminino , Humanos , Fatores de Transcrição Kruppel-Like , Repetições de MicrossatélitesRESUMO
PURPOSE: In an effort to additionally determine the global patterns of CpG island hypermethylation in sporadic breast cancer, we searched for aberrant promoter methylation at 10 gene loci in 54 primary breast cancer and 10 breast benign lesions. EXPERIMENTAL DESIGN: Genomic DNA sodium bisulfate converted from benign and malignant tissues was used as template in methyl-specific PCR for BRCA1, p16, ESR1, GSTP1, TRbeta1, RARbeta2, HIC1, APC, CCND2, and CDH1 genes. RESULTS: The majority of the breast cancer (85%) showed aberrant methylation in at least 1 of the loci tested with half of them displaying 3 or more methylated genes. The highest frequency of aberrant promoter methylation was found for HIC1 (48%) followed by ESR1 (46%), and CDH1 (39%). Similar methylation frequencies were detected for breast benign lesions with the exception of the CDH1 gene (P = 0.02). The analysis of methylation distribution indicates a statistically significant association between methylation of the ESR1 promoter, and methylation at CDH1, TRbeta1, GSTP1, and CCND2 loci (P < 0.03). Methylated status of the BRCA1 promoter was inversely correlated with methylation at the RARbeta2 locus (P < 0.03). CONCLUSIONS: Our results suggest a nonrandom distribution for promoter hypermethylation in sporadic breast cancer, with tumor subsets characterized by aberrant methylation of specific cancer-related genes. These breast cancer subgroups may represent separate biological entities with potential differences in sensitivity to therapy, occurrence of metastasis, and overall prognosis.
Assuntos
Neoplasias da Mama/genética , Metilação de DNA , Genes Supressores de Tumor , Regiões Promotoras Genéticas/genética , Análise de Variância , Neoplasias da Mama/patologia , DNA de Neoplasias/genética , DNA de Neoplasias/isolamento & purificação , Fosfatos de Dinucleosídeos/genética , Feminino , Humanos , Reação em Cadeia da PolimeraseRESUMO
Aberrant promoter methylation of several known or putative tumor suppressor genes occurs frequently during carcinogenesis, and this epigenetic change has been considered as a potential molecular marker for cancer. We examined the methylation status of nine genes (APC, CDH1, CTNNB1, TIMP3, ESR1, GSTP1, MGMT, THBS1, and TMS1), by quantitative methylation specific PCR. Synchronous preinvasive lesions (atypical ductal hyperplasia and/or ductal carcinoma in situ) and invasive ductal breast carcinoma from 52 patients, together with pure lesions from 24 patients and 12 normal tissues paired to tumor and 20 normal breast distant from tumor were analyzed. Aberrant promoter methylation was detected in both preinvasive and invasive lesions for genes APC, CDH1, CTNNB1, TIMP3, ESR1, and GSTP1. However, hierarchical mixed model and Generalized Estimating Equations model analyses showed that only APC, CDH1, and CTNNB1 promoter regions showed a higher frequency and methylation levels in pathologic samples when compared with normal breast. Whereas APC and CTNNB1 did not show differences in methylation levels or frequencies, CDH1 showed higher methylation levels in invasive tumors as compared with preinvasive lesions (P < 0.04, Mann-Whitney test with permutation correction). The analysis of APC, CDH1, and CTNNB1 methylation status was able to distinguish between normal and pathologic samples with a sensitivity of 67% (95% confidence interval, 60-71%) and a specificity of 75% (95% confidence interval, 69-81%). Our data point to the direct involvement of APC, CDH1, and CTNNB1 promoter methylation in the early stages of breast cancer progression and suggest that they may represent a useful tool for the detection of tumor cells in clinical specimens.