DNA Methylation In Colorectal Cancer Research Articles

Association of DNA methyltransferases expression with global and gene-specific DNA methylation in colorectal cancer cells.

There are conflicting reports regarding the association between DNA methyltransferases (DNMTs) expression and global or gene-specific DNA methylation in colorectal cancer (CRC) cells. To correlate DNMTs expression with DNA methylation, we quantified DNMT1, DNMT3A and DNMT3B mRNA levels in five CRC cell lines (HCT116, LS180, HT29/219, Caco2 and SW742) by real-time reverse-transcriptase polymerase chain reaction (PCR) assay. In addition, we examined the global 5-methyl cytosine levels and the methylation patterns of 12 CpG islands in these CRC cells by enzyme-linked immunosorbent assay and methylation-specific PCR methods, respectively. The average expression levels of three DNMTs in HCT116, Caco2, HT29/219 and SW742, relative to the expression level in LS180 (taken to be 1), were 90.1, 31.6, 2.66 and 1.86. Our data indicated that overall about 1.45%, 1.03%, 0.98%, 0.86% and 0.85% of the cytosines were methylated in the genome of HCT116, Caco2, HT29/219, SW742 and LS180 cells, respectively. The 5-mC percentages were positively correlated with the relative cellular DNMTs expression in five CRC cell lines as verified by Pearson correlation test. However, we found no positive correlation between mRNA expression of DNMTs and gene promoter hypermethylation in these cells. Our results suggest that cellular DNMT expression is positively correlated with global DNA methylation level but not with regional DNA hypermethylation at each locus.

Application of artificial neural networks to link genetic and environmental factors to DNA methylation in colorectal cancer.

We applied artificial neural networks (ANNs) to understand the connections among polymorphisms of genes involved in folate metabolism, clinico-pathological features and promoter methylation levels of MLH1, APC, CDKN2A(INK4A), MGMT and RASSF1A in 83 sporadic colorectal cancer (CRC) tissues, and to link dietary and lifestyle factors with gene promoter methylation. Promoter methylation was assessed by means of methylation-sensitive high-resolution melting and genotyping by PCR-RFLP technique. Data were analyzed with the Auto Contractive Map, a special kind of ANN able to define the strength of the association of each variable with all the others and to visually show the map of the main connections. We observed a strong connection between the low methylation levels of the five CRC genes and the MTR 2756AA genotype. Several other connections were revealed, including those between dietary and lifestyle factors and the methylation levels of CRC genes. ANNs revealed the complexity of the interconnections among factors linked to DNA methylation in CRC.

Heterogeneous DNA methylation contributes to tumorigenesis through inducing the loss of coexpression connectivity in colorectal cancer.

Increasing evidence indicates the high heterogeneity of cancer cells. Recent studies have revealed distinct subtypes of DNA methylation in colorectal cancer (CRC); however, the mechanism of heterogeneous methylation remains poorly understood. Gene expression is a natural, intermediate quantitative trait that bridges genotypic and phenotypic features. In this work, we studied the role of heterogeneous DNA methylation in tumorigenesis via gene expression analyses. Specifically, we integrated methylation and expression data in normal and tumor tissues, and examined the perturbations in coexpression patterns. We found that the heterogeneity of methylation leads to significant loss of coexpression connectivity in CRC; this finding was validated in an independent cohort. Functional analyses showed that the lost coexpression partners participate in important cancer-related pathways/networks, such as ErbB and mitogen-activated protein kinase (MAPK) signaling pathways. Our analyses suggest that the loss of coexpression connectivity induced by methylation heterogeneity might play an important role in CRC. To our knowledge, this is the first study to interpret methylation heterogeneity in cancer from the perspective of coexpression perturbation. Our results provide new perspectives in tumor biology and may facilitate the identification of potential biomedical therapies for cancer treatment.

Prognostic potential of DNA methylation and transcript levels of HIF1A and EPAS1 in colorectal cancer.

Hypoxic conditions during the formation of colorectal cancer may support the development of more aggressive tumors. Hypoxia-inducible factor (HIF) is a heterodimeric complex, composed of oxygen-induced HIFα and constitutively expressed HIFβ subunits, which mediates the primary transcriptional response to hypoxic stress. Among HIFα isoforms, HIF1α (HIF1A) and endothelial PAS domain-containing protein 1 (EPAS1) are able to robustly activate hypoxia-responsive gene signatures. Although posttranslational regulation of HIFα subunits is well described, less is known about their transcriptional regulation. Here, molecular analysis determined that EPAS1 mRNA was significantly reduced in primary colonic adenocarcinoma specimens compared with histopathologically nonneoplastic tissue from 120 patients. In contrast, no difference in HIF1A mRNA levels was observed between cancerous and noncancerous tissue. Bisulfite DNA sequencing and high-resolution melting analysis identified significant DNA hypermethylation in the EPAS1 regulatory region from cancerous tissue compared with nonneoplastic tissue. Importantly, multivariate Cox regression analysis revealed a high HR for patients with cancer with low EPAS1 transcript levels (HR, 4.91; 95% confidence interval, CI, 0.42-56.15; P = 0.047) and hypermethylated EPAS1 DNA (HR, 33.94; 95% CI, 2.84-405.95; P = 0.0054). Treatment with a DNA methyltransferase inhibitor, 5-Aza-2'-deoxycytidine (5-aza-dC/Decitabine), upregulated EPAS1 expression in hypoxic colorectal cancer cells that were associated with DNA demethylation of the EPAS1 regulatory region. In summary, EPAS1 is transcriptionally regulated by DNA methylation in colorectal cancer. DNA methylation and mRNA status of EPAS1 have novel prognostic potential for colorectal cancer.

Sa1926 Mucin O-Glycans As Potential Prognosis and Recurrence Markers of Colorectal Cancer

Background Micro-array analysis of promoter hypermethylation provides insight into the role and extent of DNA methylation in colorectal cancer (CRC) development, and allows direct comparison with DNA mutations. Methods Colonic biopsy samples were obtained endoscopically from 10 normal, 23 adenoma (17 low grade (LGD) and 6 high grade dysplasia (HGD)), and 8 ulcerative colitis patients. CRC samples were obtained from 24 patients (17 primary, 7 metastatic (mCRC)). Field effects were analyzed in tissues 1cm(n=5) and 10cm(n= 5) from CRC margins. Tissue materials were analyzed for DNA methylation status using a 96 gene panel and expression levels were assayed using whole genomie mRNA arrays. SFRP1 was examined by immunohistochemistry. 5-aza treatment was performed on HT29 cells to analyze reversal of DNA methylation. Results 10 genes (including SFRP1, MAL, SLIT2, SEPTIN9) showed hypermethylation in more than 85% of tumor samples. The number of methylated gene alterations were: HGD(56) > LGD(40) > CRC(37) > mCRC(12). 2 genes (MAL and SFRP1) distinguished precancerous and cancerous lesions from inflammed and healthy tissue (p<0.05). Methylation influenced mRNA and protein expression. The mRNA alteratons caused by the systematic methylation could be reversed by demethylation treatment. Conclusion Systematic changes in methylation patterns were observed early in CRC carcinogenesis, occuring in precursor lesions and all classes of CRC, often prior to the appearance of sporadic mutations. DNA hypermethylation is an early and systematic event reflected in mRNA and protein alterations in colorectal carcinogenesis, and can be potentially reversed by systematic demethylation therapy.

Sa1925 Early and Universal Aberrant DNA Hypermethylation Changes in Colorectal Adenomas and Cancers

Background Micro-array analysis of promoter hypermethylation provides insight into the role and extent of DNA methylation in colorectal cancer (CRC) development, and allows direct comparison with DNA mutations. Methods Colonic biopsy samples were obtained endoscopically from 10 normal, 23 adenoma (17 low grade (LGD) and 6 high grade dysplasia (HGD)), and 8 ulcerative colitis patients. CRC samples were obtained from 24 patients (17 primary, 7 metastatic (mCRC)). Field effects were analyzed in tissues 1cm(n=5) and 10cm(n= 5) from CRC margins. Tissue materials were analyzed for DNA methylation status using a 96 gene panel and expression levels were assayed using whole genomie mRNA arrays. SFRP1 was examined by immunohistochemistry. 5-aza treatment was performed on HT29 cells to analyze reversal of DNA methylation. Results 10 genes (including SFRP1, MAL, SLIT2, SEPTIN9) showed hypermethylation in more than 85% of tumor samples. The number of methylated gene alterations were: HGD(56) > LGD(40) > CRC(37) > mCRC(12). 2 genes (MAL and SFRP1) distinguished precancerous and cancerous lesions from inflammed and healthy tissue (p<0.05). Methylation influenced mRNA and protein expression. The mRNA alteratons caused by the systematic methylation could be reversed by demethylation treatment. Conclusion Systematic changes in methylation patterns were observed early in CRC carcinogenesis, occuring in precursor lesions and all classes of CRC, often prior to the appearance of sporadic mutations. DNA hypermethylation is an early and systematic event reflected in mRNA and protein alterations in colorectal carcinogenesis, and can be potentially reversed by systematic demethylation therapy.

Gene promoter methylation in colorectal cancer and healthy adjacent mucosa specimens

We evaluated the promoter methylation levels of the APC, MGMT, hMLH1, RASSF1A and CDKN2A genes in 107 colorectal cancer (CRC) samples and 80 healthy adjacent tissues. We searched for correlation with both physical and pathological features, polymorphisms of folate metabolism pathway genes (MTHFR, MTRR, MTR, RFC1, TYMS, and DNMT3B), and data on circulating folate, vitamin B12 and homocysteine, which were available in a subgroup of the CRC patients. An increased number of methylated samples were found in CRC respect to adjacent healthy tissues, with the exception of APC, which was also frequently methylated in healthy colonic mucosa. Statistically significant associations were found between RASSF1A promoter methylation and tumor stage, and between hMLH1 promoter methylation and tumor location. Increasing age positively correlated with both hMLH1 and MGMT methylation levels in CRC tissues, and with APC methylation levels in the adjacent healthy mucosa. Concerning gender, females showed higher hMLH1 promoter methylation levels with respect to males. In CRC samples, the MTR 2756AG genotype correlated with higher methylation levels of RASSF1A, and the TYMS 1494 6bp ins/del polymorphism correlated with the methylation levels of both APC and hMLH1. In adjacent healthy tissues, MTR 2756AG and TYMS 1494 6bp del/del genotypes correlated with APC and MGMT promoter methylation, respectively. Low folate levels were associated with hMLH1 hypermethylation. Present results support the hypothesis that DNA methylation in CRC depends from both physiological and environmental factors, with one-carbon metabolism largely involved in this process.

Efficacy and Feasibility of the Epithelial Cell Adhesion Molecule (EpCAM) Immunomagnetic Cell Sorter for Studies of DNA Methylation in Colorectal Cancer

The aim of this work was to assess the impact on measurements of methylation of a panel of four cancer gene promoters of purifying tumor cells from colorectal tissue samples using the epithelial cell adhesion molecule (EpCAM)-immunomagnetic cell enrichment approach. We observed that, on average, methylation levels were higher in enriched cell fractions than in the whole tissue, but the difference was significant only for one out of four studied genes. In addition, there were strong correlations between methylation values for individual samples of whole tissue and the corresponding enriched cell fractions. Therefore, assays on whole tissue are likely to provide reliable estimates of tumor-specific methylation of cancer genes. However, tumor cell tissue separation using immunomagnetic beads could, in some cases, give a more accurate value of gene promoter methylation than the analysis of the whole cancer tissue, although relatively expensive and time-consuming. The efficacy and feasibility of the immunomagnetic cell sorting for methylation studies are discussed.

Abstract PL05-01: Inflammatory mediators, tumor microenvironment, and progression of colorectal cancer

Abstract Population studies and experimental observations indicate that chronic inflammation and diet are risk factors for colorectal cancer (CRC). The mechanisms by which these factors contribute to the development of cancer are poorly understood. A large body of work has shown that use of nonsteroidal anti-inflammatory drugs (NSAIDs) reduce the relative risk for developing colorectal cancer (CRC) by up to 40-50%. NSAIDs exert some of their anti-inflammatory and anti-tumor effects by targeting cyclooxygenase enzymes (COX-1 and COX-2). Prostaglandin E2 (PGE2) is a pro-inflammatory mediator that promotes colorectal cancer progression. The aim of our present study was to determine the underlying mechanisms by which PGE2 affects progression of CRC. We have found a novel function of pro-inflammatory PGE2 in regulating DNA methylation in colorectal cancer. Our in vitro and in vivo studies demonstrated that PGE2 treatment silenced certain tumor suppressors and DNA repair genes such as CDKN2B (p15), CNR1, MGMT, and MLH1 by enhancing CpG island methylation of their promoter via inducing DNMT1 and DNMT3B expression. We also found that a demethylating agent inhibited the effect of PGE2 on promoting tumor growth in a mouse model of colorectal cancer, indicating that PGE2 accelerates tumor growth via regulating DNA methylation. Furthermore, combined treatment of Apcmin/+ mice with inhibitors of COX-2 and DNA methylation has more effectively anti-tumor effects than either single agent alone. To further determine the clinical relevance of our in vitro and in vivo results, we examine the relationship between COX-2 expression, PGE2 levels, DNMT1/3b expression, CpG island methylation status and expression of these tumor suppressors and DNA repair genes in human colorectal carcinomas. The levels of COX-2 and PGE2 positively correlated with DNMT1 and DNMT3B expression in these human colorectal cancer specimens. Importantly, the PGE2 levels as well as COX-2, DNMT1, and DNMT3B expression are also positively associated with the CpG island methylation in the promoters of these tumor suppressors and DNA repair genes. We also observed a specific type of immune cells (MDSCs) attracted to the tumor microenvironment by the presence of PGE2. Citation Format: Raymond N. DuBois. Inflammatory mediators, tumor microenvironment, and progression of colorectal cancer. [abstract]. In: Proceedings of the Twelfth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2013 Oct 27-30; National Harbor, MD. Philadelphia (PA): AACR; Can Prev Res 2013;6(11 Suppl): Abstract nr PL05-01.

Genome-Wide Screening for Understanding the Role of DNA Methylation in Colorectal Cancer

DNA methylation analysis methods have undergone an impressive revolution over the past 15 years. Regarding colorectal cancer (CRC), the localization and distribution of several differently methylated genes have been determined by genome-wide DNA methylation assays. These genes do not just influence the pathogenesis of CRC, but can be used further as diagnostic or prognostic markers. Moreover, the identified four DNA methylation-based subgroups of CRC have important clinical and therapeutic merit. Since genome-wide DNA methylation analyzes result in a large amount of data, there is a need for complex bioinformatic and pathway analysis. Future challenges in epigenetic alterations of CRC include the demand for comprehensive identification and experimental validation of gene abnormalities. By introduction of genome-wide DNA methylation profiling into clinical practice not only the patients' risk stratification but development of targeted therapies will also be possible.

Genome-scale analysis of DNA methylation in colorectal cancer using Infinium HumanMethylation450 BeadChips

Illumina’s Infinium HumanMethylation450 BeadChip arrays were used to examine genome-wide DNA methylation profiles in 22 sample pairs from colorectal cancer (CRC) and adjacent tissues and 19 colon tissue samples from cancer-free donors. We show that the methylation profiles of tumors and healthy tissue samples can be clearly distinguished from one another and that the main source of methylation variability is associated with disease status. We used different statistical approaches to evaluate the methylation data. In general, at the CpG-site level, we found that common CRC-specific methylation patterns consist of at least 15,667 CpG sites that were significantly different from either adjacent healthy tissue or tissue from cancer-free subjects. Of these sites, 10,342 were hypermethylated in CRC, and 5,325 were hypomethylated. Hypermethylated sites were common in the maximum number of sample pairs and were mostly located in CpG islands, where they were significantly enriched for differentially methylated regions known to be cancer-specific. In contrast, hypomethylated sites were mostly located in CpG shores and were generally sample-specific. Despite the considerable variability in methylation data, we selected a panel of 14 highly robust candidates showing methylation marks in genes SND1, ADHFE1, OPLAH, TLX2, C1orf70, ZFP64, NR5A2, and COL4A. This set was successfully cross-validated using methylation data from 209 CRC samples and 38 healthy tissue samples from The Cancer Genome Atlas consortium (AUC = 0.981 [95% CI: 0.9677–0.9939], sensitivity = 100% and specificity = 82%). In summary, this study reports a large number of loci with novel differential methylation statuses, some of which may serve as candidate markers for diagnostic purposes.

Aberrant septin 9 DNA methylation in colorectal cancer is restricted to a single CpG island

BackgroundThe septin 9 gene (SEPT9) codes for a GTP-binding protein associated with filamentous structures and cytoskeleton formation. SEPT9 plays a role in multiple cancers as either an oncogene or a tumor suppressor gene. Regulation of SEPT9 expression is complex and not well understood; however, hypermethylation of the gene was recently introduced as a biomarker for early detection of colorectal cancer (CRC) and has been linked to cancer of the breast and of the head and neck. Because the DNA methylation landscape of different regions of SEPT9 is poorly understood in cancer, we analyzed the methylation patterns of this gene in distinct cell populations from normal and diseased colon mucosa.MethodsLaser capture microdissection was performed to obtain homogeneous populations of epithelial and stromal cells from normal, adenomatous, and tumorous colon mucosa. Microdissected samples were analyzed using direct bisulfite sequencing to determine the DNA methylation status of eight regions within and near the SEPT9 gene. Septin-9 protein expression was assessed using immunohistochemistry (IHC).ResultsRegions analyzed in SEPT9 were unmethylated in normal tissue except for a methylation boundary detected downstream of the largest CpG island. In adenoma and tumor tissues, epithelial cells displayed markedly increased DNA methylation levels (>80%, p <0.0001) in only one of the CpG islands investigated. SEPT9 methylation in stromal cells increased in adenomatous and tumor tissues (≤50%, p <0.0001); however, methylation did not increase in stromal cells of normal tissue close to the tumor. IHC data indicated a significant decrease (p <0.01) in Septin-9 protein levels in epithelial cells derived from adenoma and tumor tissues; Septin-9 protein levels in stromal cells were low in all tissues.ConclusionsHypermethylation of SEPT9 in adenoma and CRC specimens is confined to one of several CpG islands of this gene. Tumor-associated aberrant methylation originates in epithelial cells; stromal cells appear to acquire hypermethylation subsequent to epithelial cells, possibly through field effects. The region in SEPT9 with disease-related hypermethylation also contains the CpGs targeted by a novel blood-based screening test (Epi proColon®), providing further support for the clinical relevance of this biomarker.

Methylation matters: The importance of DNA methylation in colorectal cancer

Methylation matters: The importance of DNA methylation in colorectal cancer

MicroRNA Mediated Network and DNA Methylation in Colorectal Cancer

MicroRNA Mediated Network and DNA Methylation in Colorectal Cancer

MicroRNA Mediated Network and DNA Methylation in Colorectal Cancer

Colorectal cancer (CRC) is one of the most malignant cancers. A growing number of studies have shown that both genetic and epigenetic play important roles in the etiology of CRC. Both microRNA (miRNA) and DNA methylation belong to the scope of epigenetic and there are complex regulatory mechanisms within miRNA and DNA methylation. We compiled 71 CRC related genes and 134 CRC related miRNAs. Then we identified 417 feed forward loops (FFLs) and 37 feedback loops (FBLs) among these genes, miRNAs and transcription factors (TFs). We constructed a network of miRNAs and TFs mediation for CRC utilizing these FFLs and FBLs. Statistical tests proved that these FFLs were significantly enriched in the CRC comparing to the esophageal cancer, breast cancer and randomly selected CRCmiRNA-gene pairs. Analysis of the network singled out 3 core genes, 2 core miRNAs and 5 core TFs. The KEGG enrichment and GO enrichment for the 2 core miRNA target genes indicated that they were significantly enriched in CRC related pathways. (Ex. MARK pathway, TGFβ pathway and cell cycle) Through the investigation on methylation, we found that most of the CRC related genes and miRNAs were prone to be regulated by methylation. This study sheds lights on the regulatory mechanisms in CRC and we provide some insights on the epigenetic of CRC.

Abstract PL02-03: Inflammatory Mediators and Colorectal Cancer Progression

Abstract Although epidemiologic and experimental observations support the hypothesis that chronic inflammation and diet are risk factors for colorectal cancer, the mechanisms by which chronic inflammation and diet contribute to the development of cancer are poorly understood. Evidence for the link between inflammation and cancer comes from epidemiologic and clinical studies showing that use of nonsteroidal anti-inflammatory drugs (NSAIDs) reduce the relative risk for developing colorectal cancer (CRC) by 40-50%. NSAIDs exert some of their anti-inflammatory and anti-tumor effects by targeting cyclooxygenase enzymes (COX-1 and COX-2). Metabolism of arachidonic acid, a major ingredient in animal fats, by cyclooxygenase enzymes provides one mechanism for the contribution of dietary fats and chronic inflammation to carcinogenesis. COX-2-derived prostaglandin E2 (PGE2) is a pro-inflammatory mediator that promotes tumor progression. Considering the importance of PGE2 signaling in inflammation and colorectal carcinogenesis, the aim of our present study was to determine whether pro-inflammatory PGE2 is a mediator in connection of chronic inflammation to colorectal carcinogenesis. Here we revealed a novel function of proinflammatory PGE2 in regulating DNA methylation in colorectal cancer. Our in vitro and in vivo studies demonstrated that PGE2 treatment silenced certain tumor suppressors and DNA repair genes such as CDKN2B (p15), CNR1, MGMT, and MLH1 by enhancing CpG island methylation of their promoter via inducing DNMT1 and DNMT3B expression. We further found that a demethylating agent inhibited the effect of PGE2 on promoting tumor growth in a mouse model of colorectal cancer, indicating that PGE2 accelerates tumor growth via regulating DNA methylation. Furthermore, combined treatment of Apcmin/+ mice with inhibitors of COX-2 and DNA methylation has more effectively anti-tumor effects than either single agent alone. To further determine the clinical relevance of our in vitro and in vivo results, we examine the relationship between COX-2 expression, PGE2 levels, DNMT1/3b expression, CpG island methylation status and expression of these tumor suppressors and DNA repair genes in human colorectal carcinomas. The levels of COX-2 and PGE2 are positively correlated with DNMT1 and DNMT3B expression in these human colorectal cancer specimens. Importantly, the PGE2 levels as well as COX-2, DNMT1, and DNMT3B expression are also positively associated with the CpG island methylation in the promoters of these tumor suppressors and DNA repair genes. The expression levels of CNR1 (CB1), MGMT, and MLH1 are negatively correlated with their levels of CpG island methylation. Collectively, these findings uncover a previously unrecognized role for PGE2 promotion of tumor formation and progression. Citation Format: Raymond N. DuBois, Dingzhi Wang. Inflammatory mediators and colorectal cancer progression. [abstract]. In: Proceedings of the Eleventh Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2012 Oct 16-19; Anaheim, CA. Philadelphia (PA): AACR; Cancer Prev Res 2012;5(11 Suppl):Abstract nr PL02-03.

Decreased expression of repulsive guidance molecule member A by DNA methylation in colorectal cancer is related to tumor progression

Previous studies have shown decreased expression of repulsive guidance molecule member A (RGMa) in colorectal cancer. However, the relationship between the expression levels and promoter DNA methylation status of RGMa and the clinical characteristics of colorectal cancer has not been previously reported. Here, we investigated the expression of RGMa by immunohistochemistry, real-time PCR and western blotting and analyzed the methylation status of the RGMa promoter using Sequenom's MassARRAY platform in colorectal cancer tissues and adjacent normal colorectal tissues. The results showed that RGMa expression was decreased in cancer tissues compared with adjacent normal tissues (p<0.01). Furthermore, a tendency for decreased expression in tumor tissues was observed from Dukes' stage A to stage D (p<0.01). In addition, significantly higher levels of hypermethylation in promoter regions of RGMa were observed in colorectal cancer tissues, compared with those in adjacent normal colorectal tissues (p<0.01). Moreover, the methylation levels of RGMa in tumor tissues were significantly increased in Dukes' stage C and D compared with Dukes' stage A and B (p<0.01). Our results indicate that RGMa expression and promoter methylation status are closely related to colorectal cancer genesis and progression. Determination of the expression level and methylation frequency of RGMa in colorectal cancer tissues may have benefit for early diagnosis and for evaluating patient prognosis.

Genome-scale analysis of aberrant DNA methylation in colorectal cancer

Colorectal cancer (CRC) is a heterogeneous disease in which unique subtypes are characterized by distinct genetic and epigenetic alterations. Here we performed comprehensive genome-scale DNA methylation profiling of 125 colorectal tumors and 29 adjacent normal tissues. We identified four DNA methylation-based subgroups of CRC using model-based cluster analyses. Each subtype shows characteristic genetic and clinical features, indicating that they represent biologically distinct subgroups. A CIMP-high (CIMP-H) subgroup, which exhibits an exceptionally high frequency of cancer-specific DNA hypermethylation, is strongly associated with MLH1 DNA hypermethylation and the BRAF(V600E) mutation. A CIMP-low (CIMP-L) subgroup is enriched for KRAS mutations and characterized by DNA hypermethylation of a subset of CIMP-H-associated markers rather than a unique group of CpG islands. Non-CIMP tumors are separated into two distinct clusters. One non-CIMP subgroup is distinguished by a significantly higher frequency of TP53 mutations and frequent occurrence in the distal colon, while the tumors that belong to the fourth group exhibit a low frequency of both cancer-specific DNA hypermethylation and gene mutations and are significantly enriched for rectal tumors. Furthermore, we identified 112 genes that were down-regulated more than twofold in CIMP-H tumors together with promoter DNA hypermethylation. These represent ∼7% of genes that acquired promoter DNA methylation in CIMP-H tumors. Intriguingly, 48/112 genes were also transcriptionally down-regulated in non-CIMP subgroups, but this was not attributable to promoter DNA hypermethylation. Together, we identified four distinct DNA methylation subgroups of CRC and provided novel insight regarding the role of CIMP-specific DNA hypermethylation in gene silencing.

Abstract LB-173: Genome-scale analysis of aberrant DNA methylation in colorectal cancer

Abstract Colorectal cancer (CRC) is a heterogeneous disease in which unique subtypes are characterized by distinct genetic and epigenetic alterations. Here we performed comprehensive genome-scale DNA methylation profiling of 125 colorectal tumors and 29 adjacent normal tissues. We identified four DNA methylation-based subgroups of CRC using model-based cluster analyses. Each subtype shows characteristic genetic and clinical features, indicating that they represent biologically distinct subgroups. A CIMP-high (CIMP-H) subgroup includes all CIMP-positive tumors characterized by the MethyLight five-marker panel, and is strongly associated with MLH1 DNA hypermethylation and the BRAFV600E mutation. A CIMP-low subgroup is enriched for KRAS mutations and characterized by DNA hypermethylation of a subset of CIMP-associated markers rather than a unique group of CpG islands. Non-CIMP tumors are separated into two distinct clusters. One non-CIMP subgroup is distinguished by a significantly higher frequency of TP53 mutations and frequent occurrence in the distal colon, while the tumors that belong to the fourth group exhibit a low frequency of both cancer-specific DNA hypermethylation and gene mutations and are significantly enriched for rectal tumors. Furthermore, we identified 114 genes that were downregulated more than 2-fold in CIMP-H tumors together with promoter DNA hypermethylation. These represent approximately 7% of genes that acquired promoter DNA methylation in CIMP-H tumors. Intriguingly, 48/114 genes were also transcriptionally silent in non-CIMP subgroups, but this was not attributable to promoter DNA hypermethylation. Together, we identified four distinct DNA methylation subgroups of CRC and provided novel insight regarding the role of CIMP-specific DNA hypermethylation in gene silencing. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-173. doi:10.1158/1538-7445.AM2011-LB-173

大腸癌における Homeobox only proteinx (HOPX) 発現抑制による悪性度獲得と癌特異的 HOPX プロモーター DNA メチル化の臨床的意義

大腸癌における Homeobox only proteinx (HOPX) 発現抑制による悪性度獲得と癌特異的 HOPX プロモーター DNA メチル化の臨床的意義