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PURPOSE: Examine the association between bulky DNA adduct levels in colon mucosa and colorectal adenoma prevalence, and explore the correlation between adduct levels in leukocytes and colon tissue. METHODS: Bulky DNA adduct levels were measured using 32P-postlabelling in biopsies of normal-appearing colon tissue and blood donated by 202 patients. Multivariable logistic regression was used to examine associations between DNA adducts, and interactions of DNA adduct-DNA repair polymorphisms, with the prevalence of colorectal adenomas. Correlation between blood and tissue levels of DNA adducts was evaluated using Spearman's correlation coefficient. RESULTS: An interaction between bulky DNA adduct levels and XPA rs1800975 on prevalence of colorectal adenoma was observed. Among individuals with lower DNA repair activity, increased DNA adduct levels were associated with increased colorectal adenoma prevalence (OR = 1.41 per SD increase, 95%CI: 0.92-2.18). Conversely, among individuals with normal DNA activity, an inverse association was observed (OR = 0.60 per SD increase, 95%CI: 0.34-1.07). Blood and colon DNA adduct levels were inversely correlated (ρ = -0.20). CONCLUSIONS: Among genetically susceptible individuals, higher bulky DNA adducts in the colon was associated with the prevalence of colorectal adenomas. The inverse correlation between blood and colon tissue measures demonstrates the importance of quantifying biomarkers in target tissues.
Assuntos
Neoplasias Colorretais/etiologia , Adutos de DNA/análise , Mucosa Intestinal/química , Adenoma/etiologia , Adulto , Feminino , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo Genético , Prevalência , Proteína de Xeroderma Pigmentoso Grupo A/genéticaRESUMO
BACKGROUND: The methylation of DNA is recognized as a key epigenetic mechanism and evidence for its role in the development of several malignancies is accumulating. We evaluated the relationship between global methylation in DNA derived from normal appearing colon mucosal tissue and blood leukocytes, and colorectal adenoma risk. METHODS: Patients, aged 40 to 65, scheduled for a screening colonoscopy were recruited. During the colonoscopy, two pinch biopsies of healthy, normal appearing mucosa were obtained from the descending colon. A fasting blood sample was also collected. The methylation status of LINE-1 (long interspersed nuclear element-1) repetitive sequences, as a surrogate measure of global methylation, was quantified in DNA extracted from normal colon mucosa and blood leukocytes. Statistical analysis of the relationship between global DNA methylation and adenoma risk was conducted on 317 participants, 108 subjects with at least one pathologically confirmed adenoma and 209 subjects with a normal colonoscopy. RESULTS: A statistically significant inverse relationship was observed between LINE-1 methylation in colon tissue DNA and adenoma risk for males and for both sexes combined for the lowest methylation quartile compared to the highest (adjusted ORs = 2.94 and 2.26 respectively). For blood, although the overall pattern of odds ratio estimates was towards an increase in risk for lower methylation quartiles compared to the highest methylation quartile, there were no statistically significant relationships observed. A moderate correlation was found between LINE-1 methylation levels measured in tissue and blood (Pearson correlation 0.36). CONCLUSIONS: We observed that lower levels of LINE-1 DNA methylation in normal appearing background colon mucosa were associated with increased adenoma risk for males, and for both sexes combined. Though these findings provide some support for a relationship between LINE-1 DNA methylation in colon mucosal tissue and adenoma risk, large prospective cohort studies are needed to confirm results. Until such investigations are done, the clinical usefulness of LINE-1 methylation as a biomarker of increased adenoma risk is uncertain. Regardless, this study contributes to a better understanding of the role of global DNA methylation as an early event in CR carcinogenesis with implications for future etiologic research.
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Adenoma/diagnóstico , Adenoma/genética , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Metilação de DNA , Adenoma/sangue , Adulto , Neoplasias Colorretais/sangue , Estudos Transversais , Epigênese Genética , Feminino , Humanos , Elementos Nucleotídeos Longos e Dispersos , Masculino , Pessoa de Meia-IdadeRESUMO
INTRODUCTION: Meat consumption is a risk factor for colorectal cancer. This research investigated the relationship between meat-derived carcinogen exposure and bulky DNA adduct levels, a biomarker of DNA damage, in colon mucosa. METHODS: Least squares regression was used to examine the relationship between meat-derived carcinogen exposure (PhIP and meat mutagenicity) and bulky DNA adduct levels in normal-appearing colon tissue measured using 32P-postlabelling among 202 patients undergoing a screening colonoscopy. Gene-diet interactions between carcinogen exposure and genetic factors relevant to biotransformation and DNA repair were also examined. Genotyping was conducting using the MassARRAY® iPLEX® Gold SNP Genotyping assay. RESULTS: PhIP and higher meat mutagenicity exposures were not associated with levels of bulky DNA adducts in colon mucosa. The XPC polymorphism (rs2228001) was found to associate with bulky DNA adduct levels, whereby genotypes conferring lower DNA repair activity were associated with higher DNA adduct levels than the normal activity genotype. Among individuals with genotypes associated with lower DNA repair (XPD, rs13181 and rs1799179) or detoxification activity (GSTP1, rs1695), higher PhIP or meat mutagenicity exposures were associated with higher DNA adduct levels. Significant interactions between the XPC polymorphism (rs2228000) and both dietary PhIP and meat mutagenicity on DNA adduct levels was observed, but associations were inconsistent with the a priori hypothesized direction of effect. CONCLUSION: Exposure to meat-derived carcinogens may be associated with increased DNA damage occurring directly in the colon among genetically susceptible individuals.
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Carcinógenos/toxicidade , Colo/efeitos dos fármacos , Adutos de DNA/metabolismo , Imidazóis/metabolismo , Imidazóis/toxicidade , Mucosa Intestinal/efeitos dos fármacos , Carne/análise , Adulto , Idoso , Biotransformação , Carcinógenos/metabolismo , Carcinógenos/farmacocinética , Colo/metabolismo , Colo/patologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Colonoscopia , Estudos Transversais , Adutos de DNA/genética , Dieta , Feminino , Interação Gene-Ambiente , Humanos , Imidazóis/farmacocinética , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Síndrome do Intestino Irritável/metabolismo , Síndrome do Intestino Irritável/patologia , Masculino , Pessoa de Meia-Idade , Polimorfismo GenéticoRESUMO
Polymorphisms in DNMT3B and MTHFR have been implicated in cancer etiology; however, it is increasingly clear that gene-specific DNA methylation also affects gene expression. A cross-sectional study (N = 272) investigated the main and joint effects of polymorphisms and DNA methylation in DNMT3B and MTHFR on colorectal adenoma risk. Polymorphisms examined included DNMT3B c.-6-1045G > T, and MTHFR c.665C > T and c.1286A > C. DNA methylation of 66 and 28 CpG sites in DNMT3B and MTHFR, respectively, was quantified in blood leukocytes using Sequenom EpiTYPER®. DNA methylation was conceptualized using two approaches: (1) average methylation and (2) unsupervised principal component analysis to identify variables that represented methylation around the transcription start site and the gene coding area of both genes. Logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) associated with the main and joint effects of polymorphisms and DNA methylation. DNA methyltransferase 3B (DNMT3B) TT versus GG/GT genotypes was associated with increased colorectal adenoma risk (OR = 2.12; 95% CI: 1.03-4.34). In addition, increasing DNA methylation in the gene-coding area of DNMT3B was associated with higher risk of colorectal adenomas (OR = 1.34; 95% CI: 1.01-1.79 per SD). In joint effect analyses, synergistic effects were observed among those with both the DNMT3B TT genotype and higher DNMT3B methylation levels compared to those with GT/GG genotypes and lower methylation levels (OR = 4.19; 95% CI: 1.45-12.13 for average methylation; OR = 4.26; 95%CI: 1.31-13.87 for methylation in the transcription start site). This research provides novel evidence that genetic and epigenetic variations contribute to colorectal adenoma risk, precursor to the majority of colorectal cancer (CRC).
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Adenoma/genética , Neoplasias Colorretais/genética , DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Polimorfismo Genético , Adenoma/diagnóstico , Adenoma/epidemiologia , Adulto , Idoso , Colo/metabolismo , Colo/patologia , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/epidemiologia , Estudos Transversais , Epigênese Genética , Feminino , Predisposição Genética para Doença , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Razão de Chances , Reto/metabolismo , Reto/patologia , Fatores de Risco , DNA Metiltransferase 3BRESUMO
Among 272 patients undergoing a screening colonoscopy, DNA methylation of DNMT3B and MTHFR, genes encoding enzymes critical to one-carbon metabolism, was quantified in blood leukocytes using Sequenom EpiTYPER®. DNA methylation was quantified in 66 and 28 CpG sites of DNMT3B and MTHFR respectively, and conceptualized using two approaches. First, measures representing average methylation across all CpG sites were created. Second, unsupervised principal component (PC) analysis was used as a pattern derivation and data-reduction approach, to develop two summary variables (PC1 and PC2). These two summary variables represented methylation around the transcription start site (PC1) and in the gene-coding area (PC2) for both DNMT3B and MTHFR. The data contained in this article presents the variation of methylation levels for individual CpG sites within the DNMT3B and MTHFR genes and possible correlations uncovered using PC analysis. The data are related to the research article "Gene-specific DNA methylation of DNMT3B and MTHFR and colorectal adenoma risk" in Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis.
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DNA methyltransferase 3B (DNMT3B) and methylenetetrahydrofolate reductase (MTHFR) are genes which encode enzymes critical to one-carbon metabolism. Polymorphisms in these genes have been implicated in colorectal cancer etiology; however, epigenetic modifications such as gene-specific DNA methylation also affect gene expression. DNA methylation of DNMT3B and MTHFR was quantified in blood leukocytes using Sequenom EpiTYPER® among 272 participants undergoing a screening colonoscopy. DNA methylation was quantified in 66 and 28CpG sites of DNMT3B and MTHFR respectively, and conceptualized using two approaches. First, measures representing average methylation across all CpG sites were created. Second, unsupervised principal component (PC) analysis was used to identify summary variables representing methylation around the transcription start site and in the gene-coding area for both DNMT3B and MTHFR. Logistic regression was used to compare methylation levels between participants diagnosed with colorectal adenoma(s) versus those with a normal colonoscopy via the estimation of odds ratios (ORs) and 95% confidence intervals (95% CIs) for the risk of colorectal adenomas. No association was observed between average DNA methylation of either DNMT3B or MTHFR and colorectal adenoma risk. For DNMT3B, increasing DNA methylation of CpG sites in the gene-coding area was associated with a higher risk of colorectal adenomas (OR=1.34; 95% CI: 1.01-1.79 per SD). This research provides preliminary evidence that methylation of DNMT3B may have functional significance with respect to colorectal adenomas, precursors to the vast majority of colorectal cancers.
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Adenoma/genética , Neoplasias Colorretais/genética , DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Adenoma/patologia , Adulto , Idoso , Colonoscopia , Neoplasias Colorretais/patologia , Ilhas de CpG/genética , Estudos Transversais , Epigênese Genética , Feminino , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Risco , DNA Metiltransferase 3BRESUMO
Exposure to heterocyclic aromatic amines (HAAs), carcinogens produced when meat is cooked at high temperatures, is an emerging risk factor for colorectal cancer (CRC). In a cross-sectional study of 342 patients undergoing a screening colonoscopy, the role of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (DiMeIQx), the three most abundant HAAs found in cooked meats, and total mutagenic activity in cooked meats were examined in relation to colorectal adenoma risk. Given that genetic differences in the ability to biotransform HAAs and repair DNA are postulated to modify the HAA-CRC relationship, gene-diet interactions were also examined. Among the total study population, no relationships were observed between dietary HAAs or meat mutagenicity, and colorectal adenoma risk; however, in males, positive associations between dietary HAAs/meat mutagenicity exposures and adenoma risk were suggestive of a relationship. In a separate analysis, polymorphisms in CYP1B1 were found to be associated with colorectal adenoma risk. Additionally, gene-diet interactions were observed for dietary PhIP and polymorphisms in CYP1B1 and XPD, dietary DiMeIQx and XPD polymorphisms, and meat mutagenicity exposure and CYP1B1 polymorphisms. Overall, increased colorectal adenoma risk was observed with higher HAA/meat mutagenicity exposures among those with polymorphisms which confer greater activity to biotransform HAAs and/or lower ability to repair DNA. This research supports the link between dietary HAAs and genetic susceptibility in colorectal adenoma etiology. The vast majority of CRCs arise from colorectal adenomas; thus, the results of this study suggest that changes in meat preparation practices limiting the production of HAAs may be beneficial for CRC prevention.
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There is increasing interest in clarifying the role of global DNA methylation levels in colorectal cancer (CRC) etiology. Most commonly, in epidemiologic studies, methylation is measured in DNA derived from blood leukocytes as a proxy measure of methylation changes in colon tissue. However, little is known about the correlations between global methylation levels in DNA derived from colon tissue and more accessible tissues such as blood or buccal cells. This cross-sectional study utilized DNA samples from a screening colonoscopy population to determine to what extent LINE-1 methylation levels (as a proxy for genome-wide methylation) in non-target tissue (e.g., blood, buccal cells) reflected methylation patterns of colon mucosal tissue directly at risk of developing CRC. The strongest Pearson correlation was observed between LINE-1 methylation levels in buccal and blood leukocyte DNA (r = 0.50; N = 67), with weaker correlations for comparisons between blood and colon tissue (r = 0.36; N = 280), and buccal and colon tissue (r = 0.27; N = 72). These findings of weak/moderate correlations have important implications for interpreting and planning future investigations of epigenetic markers and CRC risk.
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Selective serotonin reuptake inhibitors (SSRIs) are a widely prescribed class of antidepressants. Laboratory and epidemiologic evidence suggests that a prolactin-mediated mechanism secondary to increased serotonin levels at neuronal synapses could lead to a potentially carcinogenic effect of SSRIs. In this population-based case-control study, we evaluated the association between SSRI use and breast cancer risk as a function of their relative degree of inhibition of serotonin reuptake as a proxy for their impact on prolactin levels. Cases were 2,129 women with primary invasive breast cancer diagnosed from 2003 to 2007, and controls were 21,297 women randomly selected from the population registry. Detailed information for each SSRI prescription dispensed was compiled using the Saskatchewan prescription database. Logistic regression was used to evaluate the impact of use of high and lower inhibitors of serotonin reuptake and duration of use, as well as to assess the effect of individual high inhibitors on the risk of breast cancer. Exclusive users of high or lower inhibitors of serotonin reuptake were not at increased risk for breast cancer compared with non-users of SSRIs (OR = 1.01, CI = 0.88-1.17 and OR = 0.91, CI = 0.67-1.25 respectively), regardless of their duration of use or menopausal status. While we cannot rule out the possibility of a clinically important risk increase (OR = 1.83, CI = 0.99-3.40) for long-term users of sertraline (≥24 prescriptions), given the small number of exposed cases (n = 12), the borderline statistical significance, and the wide confidence interval, these results need to be interpreted cautiously. In this large population-based case-control study, we found no conclusive evidence of breast cancer risk associated with the use of SSRIs even after assessing the degree of serotonin reuptake inhibition and duration of use. Our results do not support the serotonin-mediated pathway for the prolactin-breast cancer hypothesis.
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BACKGROUND: The methylation of DNA is recognized as a key mechanism in the regulation of genomic stability and evidence for its role in the development of cancer is accumulating. LINE-1 methylation status represents a surrogate measure of genome-wide methylation. FINDINGS: Using high resolution melt (HRM) curve analysis technology, we have established an in-tube assay that is linear (r > 0.9986) with a high amplification efficiency (90-105%), capable of discriminating between partcipant samples with small differences in methylation, and suitable for quantifying a wide range of LINE-1 methylation levels (0-100%)--including the biologically relevant range of 50-90% expected in human DNA. We have optimized this procedure to perform using 2 µg of starting DNA and 2 ng of bisulfite-converted DNA for each PCR reaction. Intra- and inter-assay coefficients of variation were 1.44% and 0.49%, respectively, supporting the high reproducibility and precision of this approach. CONCLUSIONS: In summary, this is a completely linear, quantitative HRM PCR method developed for the measurement of LINE-1 methylation. This cost-efficient, refined and reproducible assay can be performed using minimal amounts of starting DNA. These features make our assay suitable for high throughput analysis of multiple samples from large population-based studies.