Aberrant Methylation Of CpG Islands Research Articles

E3 ubiquitin ligase RNF180 prevents excessive PCDH10 methylation to suppress the proliferation and metastasis of gastric cancer cells by promoting ubiquitination of DNMT1

BackgroundDownregulation of certain tumor-suppressor genes (TSGs) by aberrant methylation of CpG islands in the promoter region contributes a great deal to the oncogenesis and progression of several cancers, including gastric cancer (GC). Protocadherin 10 (PCDH10) is a newly identified TSG in various cancers and is downregulated in GC; however, the specific mechanisms of PCDH10 in GC remain elusive. Here, we elucidated a novel epigenetic regulatory signaling pathway involving the E3 ubiquitin ligase RNF180 and DNA methyltransferase 1 (DNMT1), responsible for modulating PCDH10 expression by affecting its promoter methylation.ResultsWe revealed that PCDH10 was downregulated in GC cells and tissues, and low PCDH10 expression was correlated with lymph node metastasis and poor prognosis in patients with GC. Additionally, PCDH10 overexpression suppressed GC cell proliferation and metastasis. Mechanistically, DNMT1-mediated promoter hypermethylation resulted in decreased expression of PCDH10 in GC tissues and cells. Further analysis revealed that RNF180 can bind directly to DNMT1 and was involved in DNMT1 degradation via ubiquitination. Additionally, a positive correlation was found between RNF180 and PCDH10 expression and an inverse association between DNMT1 and PCDH10 expression showed considerable prognostic significance.ConclusionOur data showed that RNF180 overexpression upregulated PCDH10 expression via ubiquitin-dependent degradation of DNMT1, thus suppressing GC cell proliferation, indicating that the RNF180/DNMT1/PCDH10 axis could be a potential therapeutic target for GC treatment.

25P Relationship between aberrant methylation of CpG islands of microRNA gene promoters and changes in their expression in epithelial ovarian cancer

25P Relationship between aberrant methylation of CpG islands of microRNA gene promoters and changes in their expression in epithelial ovarian cancer

Investigation of Isocitrate Dehydrogenase 1 and 2 Mutations in Acute Leukemia Patients in Saudi Arabia.

Different forms of human cancer show mutations for isocitrate dehydrogenases 1 and 2 (IDH1/2). Mutation of these genes can cause aberrant methylation of the genome CpG islands (CGIs), which leads to an increase of suppressed oncogenes transcription or repression of active tumor suppressor gene transcription. This study aimed to identify the prevalence of IDH1/2 mutations in acute leukemia patients. The study cohort included 43 AML patients and 30 childhood ALL patients, from whom DNA bone marrow samples were taken. The alteration hotspots in codons IDH1 (R132) and IDH2 (R172 and R140) were examined via direct sequencing. Mutations in IDH1 were detected in 7 out of 43 (16.2%) AML patients; 5 of them occurred at codon R132. The other two mutations included a single-nucleotide polymorphism, which affected codon G105 in one patient. However, no mutation was detected in the IDH2 in any of the patients. Moreover, no mutations were detected in either IDH1 or IDH2 in ALL patients. The dominance of IDH1 mutations in AML, which was 16%, emphasizes the existence of the mutation in our population. On the other hand, IDH2 mutation was observed to be less frequent in both illnesses. Due to the limitation of using a small sample size, larger cohort screening is recommended to determine their usefulness as prognostic indicators.

Promoter hypomethylation as potential confounder of Ras gene overexpression and their clinical significance in subsets of urothelial carcinoma of bladder.

Overexpression of normal Ras and its aberrant CpG island methylation in the promoter regions have been shown to direct cells for uncontrolled abnormal growth and bladder tumor formation and therefore, fetched recent attention as a marker of diagnosis and prognosis to predict the biological behavior of urothelial carcinoma of bladder (UCB). Methylation pattern at CpG islands of the promoter regions of rat sarcoma (Ras) gene homologues namely Kristen-Ras (K-Ras), Harvey (H-Ras), and Neuroblastoma (N-Ras) were examined by methylation specific polymerase chain reaction (MSP). Real time-quantitative polymerase chain reaction (RT-qPCR) was done to determine transcriptomic expressions of these Ras isoforms in the prospective series of 42 NMIBC (non-muscle invasive bladder cancer) and 45 MIBC (muscle invasive bladder cancer) biopsies. CpG loci in H-Ras and K-Ras were observed to be more hypomethylated in MIBC, whereas more hypomethylation in N-Ras was noted in NMIBC. Strong association of hypomethylation index with tumor stage, grade, type and size validate them it as marker of diagnosis in UCB patients. Differential overexpression of H-Ras, N-Ras and K-Ras genes in NMIBC and MIBC and their association with patients' demographics identify them as important diagnostic markers in pathogenesis of UCB. Given the reported ability of promoter hypomethylation to activate Ras expression, correlation studies examined positive significant association between hypomethylation index and expression. Study concludes that promoter hypomethylation of N-Ras and K-Ras could be a potential confounder of their increased expression in NMIBC. Biological significance of simultaneous presence of higher expression and promoter hypomethylation of Ras gene isoforms in MIBC is difficult to resolve in a given cohort of patients.

Cathepsin V Mediates the Tazarotene-induced Gene 1-induced Reduction in Invasion in Colorectal Cancer Cells.

Tazarotene-induced gene 1 (TIG1) is a retinoid acid receptor-responsive gene involved in cell differentiation and tumorigenesis. Aberrant methylation of CpG islands in the TIG1 promoter is found in multiple cancers. Currently, the exact mechanism underlying the anticancer effect of TIG1 is unknown. Here, we show that TIG1 interacts with cathepsin V (CTSV), which reduces CTSV stability and subsequently affects the production of activated urokinase-type plasminogen activator (uPA), an epithelial-mesenchymal transition-associated protein. Ectopic expression of CTSV increased the expression of activated uPA and the number of migrated and invaded cells, whereas ectopic TIG1 expression reversed the effects of CTSV on the uPA signaling pathway. Similar patterns in the production of activated uPA and number of migrated and invaded cells were also observed in TIG1-expressing and CTSV-knockdown cells. The results suggest that CTSV may participate in TIG1-regulated uPA activity and the associated downstream signaling pathway.

Aberrant Methylation of Suppressor of Cytokine Signaling 1 (SOCS1) Gene as a Biomarker for Early Prediction and Diagnosis of Bladder Cancer.

SOCS1 protein, the negative regulatory protein of the Janus kinase (JAK)/signal transducers and activators of transcription (STAT) signaling pathway, may inhibit signaling of JAK-STAT pathway by several cytokines and has tumor suppressor activity. Methylation of CpG island in the promoter region of SOCS1 gene has often been shown to inactivate the SOCS1 gene in certain human cancers. However, the precise role of SOCS1 in bladder cancer is unclear. Two hundred forty-seven patients with BCa and 243 healthy controls were enrolled from Tumour Hospital Affiliated to Harbin Medical University, Hongqi Hospital Affiliated to Mudanjiang Medical University, and Mudanjiang Tumour Hospital from September 2013 to March 2019. The methylation rate in the promoter region of the SOCS1 among all participants were detected using the MassARRAY EpiTYPER system. A ROC curve was set out to analyze SOCS1 gene promoter CpG island methylation for BCa diagnosis. There was a significantly higher methylation rate in BCa compared to controls. Then we assessed the methylation rate of different CpG islands in SOCS1 gene among BCa cases and normal controls. Methylation rate was shown to vary among different CpG islands. The methylation rates of CpG islands were shown to vary among different grades. We observed that the methylation rate of different CpG islands vary according to pathological grades. Our study demonstrates that aberrant methylation of CpG island in the promoter region of SOCS1 gene may be involved in occurrence, progression, and prognosis of BCa and, thus, may serve as an independent diagnosis and prognostic biomarker.

Aberrant promoter methylation may be responsible for the control of CD146 (MCAM) gene expression during breast cancer progression.

The CD146 (also known as MCAM, MUC-18, Mel-CAM) was initially reported in 1987, as a protein crucial for the invasiveness of malignant melanoma. Recently, it has been confirmed that CD146 has been involved in progression and poor overall survival of many cancers including breast cancer. Importantly, in independent studies, CD146 was reported to be a trigger of epithelial to mesenchymal transition in breast cancer cells. The goal of our current study was to verify the potential involvement of epigenetic mechanism behind the regulation of CD146 expression in breast cancer cells, as it has been previously reported in prostate cancer. First, we analysed the response of breast cancer cell lines, differing in the initial CD146 mRNA and protein content, to epigenetic modifier, 5-aza-2-deoxycytidine, and subsequently the methylation status of CD146 gene promoter was investigated, using direct bisulfite sequencing. We observed that treatment with demethylating agent led to induction of CD146 expression in all analysed breast cancer cell lines, both at mRNA and protein level, what was accompanied by increased expression of selected mesenchymal markers. Importantly, CD146 gene promoter analysis showed aberrant CpG island methylation in 2 out of 3 studied breast cancer cells lines, indicating epigenetic regulation of CD146 gene expression. In conclusion, our study revealed, for the first time, that aberrant methylation maybe involved in expression control of CD146, a very potent EMT inducer in breast cancer cells. Altogether, the data obtained may provide the basis for novel therapies as well as diagnostic approaches enabling sensitive and very accurate detection of breast cancer cells.

The transcriptome of CMML monocytes is highly inflammatory and reflects leukemia-specific and age-related alterations

AbstractChronic myelomonocytic leukemia (CMML) is an aggressive myeloid neoplasm of older individuals characterized by persistent monocytosis. Somatic mutations in CMML are heterogeneous and only partially explain the variability in clinical outcomes. Recent data suggest that cardiovascular morbidity is increased in CMML and contributes to reduced survival. Clonal hematopoiesis of indeterminate potential (CHIP), the presence of mutated blood cells in hematologically normal individuals, is a precursor of age-related myeloid neoplasms and associated with increased cardiovascular risk. To isolate CMML-specific alterations from those related to aging, we performed RNA sequencing and DNA methylation profiling on purified monocytes from CMML patients and from age-matched (old) and young healthy controls. We found that the transcriptional signature of CMML monocytes is highly proinflammatory, with upregulation of multiple inflammatory pathways, including tumor necrosis factor and interleukin (IL)-6 and -17 signaling, whereas age per se does not significantly contribute to this pattern. We observed no consistent correlations between aberrant gene expression and CpG island methylation, suggesting that proinflammatory signaling in CMML monocytes is governed by multiple and complex regulatory mechanisms. We propose that proinflammatory monocytes contribute to cardiovascular morbidity in CMML patients and promote progression by selection of mutated cell clones. Our data raise questions of whether asymptomatic patients with CMML benefit from monocyte-depleting or anti-inflammatory therapies.

Abstract 4180: Association between genetic polymorphisms of DNMT genes and breast cancer in a nested case-control study of African Americans

Abstract Introduction/Purpose. DNA methylation plays a significant role in both the development and progression of various cancers. Although aberrant DNA methylation of CpG islands is a common epigenetic alternation found in cancers, the interpretation of observed DNA methylation and cancer risk remains a challenge. DNA methylation is typically mediated by DNA Methyltransferases (DNMTs). This study took advantage of the infrastructure and existing data and samples of Arkansas Rural Community Health (ARCH) Study cohort to conduct a pilot nested case-control study of breast cancer among African Americans examining the relationship between breast cancer and DNMT single nucleotide polymorphisms (SNPs). Methods: ARCH Study cohort consists of 26,387 women aged 18 to 85 recruited from all 75 counties in Arkansas between 2007 and 2014. All cohort participants donated a saliva sample for DNA and completed a short questionnaire at baseline. 309 African American breast cancer cases and 530 age-group, race, and recruitment year frequency-matched controls were selected for this analysis. Genotyping of DNMT1, DNMT3A, and DNMT3B were conducted using Tagman method. Multivariable logistic regression (odds ratio (OR), 95% confidence interval (CI) was used to examine the association between breast cancer and each SNP while considering the factors of age, education, and status of obesity, child birth, breast feeding, and menstrual-stop as potential confounding variables. Hardy-Weinberg equilibrium for the loci examined were evaluated. Haplotype analyses were conducted. Results: Among 16 SNPs evaluated, heterozygous genotypes rs7575625 in DNMT1 and rs7605753 in DNMT3A were positively associated with breast cancer risk; rs12991495 in DNMT3A and rs17123590 and rs2424910 in DNMT3B were inversely associated with breast cancer. Haplotype AGA of rs8101626, rs2290684, and rs11880388 in DNMT1, which accounts for 43.9% population, differed significantly as a risk factor between cases and controls after adjusting for confounders (OR = 2.05; 95%CI=1.34, 3.12). Haplotype AGGGGT of rs2304429, rs12991495, rs7605753, rs11892646, rs7575625, and rs10196635 in DNMT3A differed significantly as a risk factor between cases and controls (OR = 4.06; 95%CI=1.93, 8.52); while haplotype rs2424905, rs2424910, rs17123590, rs6088008, and rs6058896 in DNMT3B was inversely associated with breast cancer (OR = 0.02; 95%CI=0.001, 0.29). Conclusions: Despite the knowledge of DNMTs on DNA methylation, very few studies examined its role in breast cancer, in particular among underserved and minority population. This study took advantage of the existing sample and data of African Americans in a cohort population and demonstrated DNMT genetic polymorphisms are associated with breast cancer. Further evaluation of methylation patterns could provide knowledge on the mechanistic pathways for the association observed. Citation Format: L Joseph Su, Hui-Yi Lin, Tung-Chin Chiang, Gail Runnells, Lora Rogers, Ping-Ching Hsu, Ronda Henry-Tillman, Susan Kadlubar. Association between genetic polymorphisms of DNMT genes and breast cancer in a nested case-control study of African Americans [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4180.

DNA Methylation Status of SHOX-Flanking CpG Islands in Healthy Individuals and Short Stature Patients with Pseudoautosomal Copy Number Variations

SHOX resides in the short arm pseudoautosomal region (PAR1) of the sex chromosomes and escapes X inactivation. SHOX haploinsufficiency underlies idiopathic short stature (ISS) and Leri-Weill dyschondrosteosis (LWD). A substantial percentage of cases with SHOX haploinsufficiency arise from pseudoautosomal copy number variations (CNVs) involving putative enhancer regions of SHOX. Our previous study using peripheral blood samples showed that some CpG dinucleotides adjacent to SHOX exon 1 were hypomethylated in a healthy woman and methylated in a woman with gross X chromosomal rearrangements. However, it remains unknown whether submicroscopic pseudoautosomal CNVs cause aberrant DNA methylation of SHOX-flanking CpG islands. In this study, we examined the DNA methylation status of SHOX-flanking CpG islands in 50 healthy individuals and 10 ISS/LWD patients with pseudoautosomal CNVs. In silico analysis detected 3 CpG islands within the 20-kb region from the translation start site of SHOX. Pyrosequencing and bisulfite sequencing of genomic DNA samples revealed that these CpG islands were barely methylated in peripheral blood cells and cultured chondrocytes of healthy individuals, as well as in peripheral blood cells of ISS/LWD patients with pseudoautosomal CNVs. These results, in conjunction with our previous findings, indicate that the DNA methylation status of SHOX-flanking CpG islands can be affected by gross X-chromosomal abnormalities, but not by submicroscopic CNVs in PAR1. Such CNVs likely disturb SHOX expression through DNA methylation-independent mechanisms, which need to be determined in future studies.

Predicting cervical intraepithelial neoplasia recurrence in HIV-infected and -noninfected women by detecting aberrant promoter methylation in the CDH1, TIMP3, and MGMT genes.

Aberrant DNA methylation is present in virtually all types of human cancer. There is no clear evidence that methylation status can predict bad prognosis in patients with CIN recurrence in HIV infected. This study evaluates the relationship between aberrant methylation of CpG islands of CDH1, TIMP3 and MGMT genes and CIN recurrence in HIV-infected and -noninfected women. This is a nested case-control study involving 33 cases with CIN recurrence and 114 controls without recurrence, HIV infected and noninfected, treated with LEEP, between 1999 and 2004. Recurrence diagnosis was established after biopsy. Genes methylation profile was assessed by MSP-PCR technique in formalin-fixed, paraffin-embedded cone specimens. Statistical analysis was performed to compare categorical variables, using χ2 test with Yates correction and Fisher's exact test. Multivariate analysis was carried out using logistic regression. CIN recurrence was more frequent in women with glandular involvement (OR 11.6; 95% CI 2.93-45.89) and compromised surgical margins (OR 2.5; 95% CI 0.87-7.27) in the cervical cone and in HIV-infected women (OR 2.47; 95% CI 0.87-7.05). One methylated allele of CDH1, TIMP3 and MGMT genes was present in 87.9% women with CIN recurrence. Promoter hypermethylation of TIMP3 and MGMT was detected in women with CIN recurrence and without CIN recurrence independent of HIV infection with significant difference between groups (p = 0.04 and p = 0.02, respectively). CIN recurrence was associated with glandular involvement and compromised margins in cone biopsy and HIV infection. The presence of CpG islands hemimethylation in TIMP3 and MGMT genes is a promising triage method in CIN recurrence.

Promoter epigenetics of APC gene and its implication in sporadic breast cancer patients from South Indian population

BackgroundThe adenomatous polyposis coli (APC) gene is a tumor suppressor gene associated with both familial and sporadic cancer, whose protein plays an important role in the WNT signal transduction pathway. WNT signaling is a key developmental pathway involved in embryonic development, cell differentiation, cell proliferation and tissue maintenance in adults. APC protein acts as an antagonist of the WNT signaling pathway by binding and regulating the β-catenin protein. Impaired APC gene function usually leads to a lack of degradation of β-catenin, which could cause uncontrolled cell growth and lead to tumorigenic transformation. APC gene may be inactivated by aberrant DNA methylation of CpG islands in their promoter regions. ObjectiveHence the present study was designed to determine the role of promoter methylation of APC genes in sporadic breast cancer patients from South Indian population. MethodsDNA methylation analyses of APC gene was performed by methylation-specific polymerase chain reaction (MSP).·Fifty biopsy samples of breast tumor and their corresponding non-malignant portions as controls were studied comparatively. APC mRNA expression analysis was also done using real time PCR. ResultsThirty eight percent (38%) cancerous tissue samples (19/50) showed promoter hypermethylation in APC gene; however, none of normal tissues showed promoter hypermethylation. The difference in methylation frequency between cancerous and normal tissue was statistically significant (p = 0.0001). APC promoter methylation was positively associated with lymph node involvement (p = 0.048), APC Promote hyper methylation and decreased mRNA expression was significantly associated (p = 0.0034). Lower APC-mRNA expression was associated with patient's age above 50 years (p = 0.044). ConclusionIn conclusion, our data showed that promoter hypermethylation of APC gene was a prognostic factor in breast cancer patients from South Indian population. The loss of APC gene function usually leads to an accumulation of β-catenin, which may promote tumorigenesis.

O6-Methyguanine-DNA Methyl Transferase (MGMT) Promoter Methylation in Serum DNA of Iranian Patients with Colorectal Cancer

Introduction:Colorectal cancer (CRC) is a leading cause of cancer deaths worldwide but current molecular targeted therapy is not providing major success in CRC treatment, so early detection by non-invasive methods continues to be vital. Aberrant methylation of CpG islands in promoter regions is associated with inactivation of various tumor suppressor genes. O6-methyguanine-DNA methyltransferase (MGMT) is a DNA repair enzyme that removes mutagenic and cytotoxic adducts from O6-guanine in DNA. Aberrant hypermethylation of the MGMT promoter has been associated with lack of mRNA expression, with concomitant loss of protein content and enzyme activity.AIM:Our aim was to determine whether MGMT promoter methylation might be detectable in circulating free DNA in the serum of CRC patients and normal individuals using a methylation specific (MSP) polymerase chain reaction (PCR) method.Methods:A total of 70 subjects were enrolled in the study. Of these, 30 patients who were diagnosed previously as untreated colon adenocarcinoma by a gastroenterologist and the other 40 were nearly age-matched individuals who had a normal colonoscopic evaluation (except for hemorrhoids or fissures) and normal pathologic reports. After bisulphite modification of DNA, serum samples were examined for MGMT promoter methylation using MSP.Results:Ninety percent of CRC patients had MGMT promoter hypermethylation as compared to no methylation in normal subjects’ serum. Most of the cancers were stage П and moderately differentiated adenocarcinomas; nearly 60% were found in the left colon. No statistically significant correlation was found between the promoter methylation status and gender and age.Discussion and Conclusions:MGMT hypermethylation can be detected in free circulating DNA in serum of CRC patients and can be used “as a clinical biomarker” for early diagnosis and prognostic assessment of the disease. Our data confirm previous studies indicating utility for free circulating DNA as a serum biomarker for early detection, diagnosis and monitoring of CRC patients.

Analysis of DNA Methylation in Tissues Exposed to Inflammation.

Induction of aberrant DNA methylation is one of the most important mechanisms mediating the effect of inflammation on cancer development. Aberrant methylation of promoter CpG islands of tumor suppressor genes can silence their downstream genes, and that in cancer tissues is associated with prognosis or therapeutic effects. In addition, aberrant methylation can occur in tissues exposed to specific types of inflammation, producing a so-called "epigenetic field for cancerization," and its accumulation is correlated with cancer risk. Thus, aberrant methylation at specific loci is an important biomarker and mediator of the carcinogenic effect of inflammation. DNA methylation at specific genomic regions can be analyzed by various methods based upon bisulfite-mediated DNA conversion, which specifically converts unmethylated cytosines into uracils under appropriate conditions. Methylation-specific PCR (MSP), quantitative MSP, and bisulfite sequencing are widely used, and this chapter provides protocols for bisulfite-mediated conversion, quantitative MSP, and bisulfite sequencing.

Epigenetic modifications of gene expression by lifestyle and environment

Epigenetics oftenly described as the heritable changes in gene expression independent of changes in DNA sequence. Various environmental factors such as nutrition-dietary components, lifestyle, exercise, physical activity, toxins, and other contributing factors remodel the genome either in a constructive or detrimental way. Since epigenetic changes are reversible and nutrition is one of the many epigenetic regulators that modify gene expression without changing the DNA sequence, dietary nutrients and bioactive food components contribute to epigenetic phenomena either by directly suppressing DNA methylation or histone catalyzing enzymes or by changing the availability of substrates required for enzymatic reactions. Diets that contain catechol-dominant polyphenols are reported to suppress enzyme activity and activate epigenetically silenced genes. Furthermore, several dietary nutrients play a crucial role in one-carbon metabolism including folate, cobalamin, riboflavin, pyridoxine, and methionine by directly affecting S-adenosyl-L-methionine. Soy polyphenols block DNA methyltransferases and histone deacetylases to reverse aberrant CpG island methylation. Organosulfur rich compounds such as the sulforaphane found in broccoli appear to normalize DNA methylation and activate miR-140 expression, which represses SOX9 and ALDH1 and decreases tumor growth. The purpose of this short communication is to overview the epigenetic regulatory mechanisms of diet and other environmental factors. We discuss the epigenetic contributions of dietary components with a particular focus on nutritional polyphenols and flavonoids as epigenetic mediators that modify epigenetic tags and control gene expression. These mechanisms provide new insights to better understand the influence of dietary nutrients on epigenetic modifications and gene expression.

Epigenetic Pathways Offer Targets for Ovarian Cancer Treatment

Introduction Ovarian cancer is the most severe disease of the female reproductive tract as it is associated with the highest mortality rates. There are 3 categories of ovarian cancer including epithelial, stromal, and germ cell tumors. From these categories, epithelial and in particular serous epithelial ovarian cancer is the most common type. Treatment of the cancer depends a lot on its stage. When it is organ confined, it can be surgically removed. However, in the case where it has progressed beyond the ovaries, a combination of surgery and chemotherapy is needed. An important prognostic factor that could predict survival is whether the tumor size is reduced after surgery and to what extent. In chemotherapy, the 2 most important drugs used in the treatment of ovarian cancer are carboplatin and paclitaxel. Nowadays, one of the greatest challenges associated with chemotherapy is the fact that cancers have started developing resistance against such drugs, and consequently, their effectiveness in eradicating the tumor has been significantly reduced. This has raised the need for the development of a new treatment that would remain unaffected by these mechanisms of resistance. Epigenetics is a field which focuses on the study of heritable changes which do not result from changes in the DNA sequence. Epigenetic changes include DNA methylation, which is the addition of a methyl group to a cytosine (C) residue with the aid of an enzyme known as DNA methyltransferase (DNMT), and histone modifications. These are divided into histone acetylation, deacetylation, and methylation events carried out by histone acetyltransferases, histone deacetylases (HDACs), and histone methyltransferases (HMTs), respectively. Recently, research has focused on epigenetic changes and their relation to cancer. In fact, aberrant methylation of CpG islands which are found in close proximity to gene transcription initiation sites and are normally methylated has been linked to tumor initiation and progression. Because of their involvement in tumorigenesis, the biology of these changes is now being investigated so that its potential in the treatment of ovarian cancer is fully understood. This mini-review will describe the common molecular mechanisms of ovarian cancer and discuss the prospective of epigenetic remedies in the treatment of ovarian cancer.

Clinical and prognosis value of the CIMP status combined with MLH1 or p16 INK4a methylation in colorectal cancer.

Aberrant DNA methylation of CpG islands occurred frequently in CRC and associated with transcriptional silencing of key genes. In this study, the CIMP combined with MLH1 or p16 INK4a methylation status was determined in CRC patients and correlated with clinicopathological parameters and overall survival. Our data showed that CIMP+ CRCs were identified in 32.9% of cases and that CACNAG1 is the most frequently methylated promoter. When we combined the CIMP with the MLH1 or the p16 INK4a methylation status, we found that CIMP-/MLH1-U (37.8%) and CIMP-/p16 INK4a -U (35.4%) tumors were the most frequent among the four subtypes. Statistical analysis showed that tumor location, lymphovascular invasion, TNM stage, and MSI differed among the group of patients. Kaplan-Meier analyses revealed differences in overall survival according to the CIMP combined with MLH1 or p16 INK4a methylation status. In a multivariate analysis, CIMP/MLH1 and CIMP/p16 INK4a methylation statuses were predictive of prognosis, and the OS was longer for patients with tumors CIMP-/MLH1-M, as well as CIMP-/p16 INK4a -M. Furthermore, DNMT1 is significantly overexpressed in tumors than in normal tissues as well as in CIMP+ than CIMP- tumors. Our results suggest that tumor classification based on the CIMP status combined with MLH1 or p16 INK4a methylation is useful to predict prognosis in CRC patients.

Genome-wide screening of DNA methylation associated with lymph node metastasis in esophageal squamous cell carcinoma.

Lymph node metastasis (LNM) of esophageal squamous cell carcinoma (ESCC) is well-known to be an early event associated with poor prognosis in patients with ESCC. Recently, tumor-specific aberrant DNA methylation of CpG islands around the promoter regions of tumor-related genes has been investigated as a possible biomarker for use in early diagnosis and prediction of prognosis. However, there are few DNA methylation markers able to predict the presence of LNM in ESCC. To identify DNA methylation markers associated with LNM of ESCC, we performed a genome-wide screening of DNA methylation status in a discovery cohort of 67 primary ESCC tissues and their paired normal esophageal tissues using the Illumina Infinium HumanMethylation450 BeadChip. In this screening, we focused on differentially methylated regions (DMRs) that were associated with LNM of ESCC, as prime candidates for DNA methylation markers. We extracted three genes, HOXB2, SLC15A3, and SEPT9, as candidates predicting LNM of ESCC, using pyrosequencing and several statistical analyses in the discovery cohort. We confirmed that HOXB2 and SEPT9 were highly methylated in LNM-positive tumors in 59 ESCC validation samples. These results suggested that HOXB2 and SEPT9 may be useful epigenetic biomarkers for the prediction of the presence of LNM in ESCC.

Promoter Hypermethylation and Its Impact on Expression of MGMT Gene in the GIT Malignant Patients of Kashmiri Origin

ABSTRACTEpigenetic alterations, in addition to multiple gene abnormalities, are involved in the genesis and progression of human cancers. Gastrointestinal tract (GIT) cancer is a major medical and economic burden worldwide. Aberrant methylation of CpG islands within promoter regions is associated with transcriptional inactivation of various tumor suppressor genes. Although a number of cancer-associated genes have been found to be hypermethylated in GIT cancer, valuable methylation markers for early diagnosis and prognostic evaluation of this cancer remain largely unknown. O6-methyguanine DNA methyltransferase (MGMT) is a DNA-repair gene that removes mutagenic and cytotoxic adducts from the O6 position of guanine induced by alkylating agents. MGMT promoter hypermethylation and reduced expression have been found in some primary human carcinomas. We studied DNA methylation of CpG islands of the MGMT gene and its relation with MGMT protein expression in human GIT carcinomas. A total of 210 GIT tumor samples and 90 adjacent normal tissues were analyzed for MGMT promoter methylation by methylation-specific polymerase chain reaction after bisulfite modification of DNA and same samples were analyzed for MGMT protein expression by Western blotting. The methylation frequencies of MGMT gene promoter were 41.4%, 34.2%, and 44.2% in stomach, esophageal, and colorectal cancer cases while as 16.6, 13.3, and 13.3 in respective controls. MGMT protein was found downregulated in controls of all GIT. The results suggest that methylation at CpG islands of MGMT may be responsible for the downregulation of MGMT protein expression in GIT cancers.

Mapping of DNA Hypermethylation and Hypomethylation induced by Folate Deficiency in Sporadic Colorectal Cancer and Clinical Implication Analysis of Hypermethylation Pattern in CBS Promoter.

Aberrant DNA methylation patterns play a major role in tumorigenesis and the effects of nutrients, especially folate in the diet, on methylation changes is of great importance in colorectal cancer (CRC). Folate deficiency would disrupt methylation patterns; however, its exact effects on DNA methylation patterns in CRC are unclear. This study was performed to gain insight into the methylation changes induced by folate deficiency and the putative role of methylation pattern diversities of related genes in the clinical outcome of CRC. The NimbleGen MeDIP chip (Methylated DNA Immunoprecipitation chip) assay was used in high-resolution mapping of DNA methylation patterns in the normal human colon mucosal epithelial cell line, NCM460 cultured with or without folate. Aberrant CpG island methylation patterns in the promoter of genes were identified by chip assay and then were confirmed in paired colorectal tissues and corresponding non-malignant tissues obtained from patients by bisulfate sequencing PCR (BSP). Of the total, the expression of cystathionine-beta-synthase (CBS) involved in methyl metabolism and its important substrate, homocysteine, were all detected by realtime RT-PCR and immunostaining. We also analyzed the data of its hypermethylation level statistically correlated with pathological parameters and the clinical outcome in malignant tissues. The chip assay showed that there are 17 genes with hyper or hypomethylation in CpG islands of promoter on chromosome 21, and 8 of them seemed to be associated with tumorigenesis. Among the total, a hypermethylation patterns existed in the promoter of CBS in CRC (p < 0.001), and the hypermethylation is related with the down-regulation of CBS and the accumulation of homocysteine in vitro and vivo (p < 0.001). Univariate analysis showed CBS hypermethylation level is correlated with age (p < 0.001), pT stage (p = 0.008), pN stage (p = 0.038), liver metastasis (p = 0.017), pTNM stage (p = 0.032), Dukes' stage (p = 0.022), recurrence (p = 0.041), five-year survival (p = 0.034), recurrence-free probability (p = 0.011), and overall survival (p = 0.018). Multivariate analysis showed that CBS hypermethylation level significantly correlated with recurrence rate (p = 0.039) and overall survival (p = 0.012) independent of pT stage, pN stage, and liver metastasis. Folate deficiency could induce aberrant DNA methylation patterns and gene expressions in CRC. CBS plays a critical role in tumorigenesis and could serve as a prognostic marker for tumor progression.