CpG Island Methylation Research Articles

Protective Effects of Dendrobium nobile Lindl. Alkaloids on Alzheimer's Disease-like Symptoms Induced by High-methionine Diet.

High methionine-diet (HMD) causes Alzheimer's disease (AD)-like symptoms. Previous studies have shown that Dendrobium nobile Lindle. alkaloids (DNLA) have potential benefits for AD Object: The objective of this study has been to explore whether DNLA can improve AD-like symptoms induced by HMD. Mice were fed with 2% HMD diet for 11 weeks; the DNLA20 control group (20 mg/kg), DNLA10 group (10 mg/kg), and DNLA20 group (20 mg/kg) were administered DNLA for 3 months. Morris water maze test was used to detect learning and memory ability. Neuron damage was evaluated by HE and Nissl staining. Levels of homocysteine (Hcy), beta-amyloid 1-42 (Aβ1-42), S-adenosine methionine (SAM) and S-adenosine homocysteine (SAH) were detected by ELISA. Immunofluorescence and western blotting (WB) were used to determine the expression of proteins. CPG island methylation levels were accessed by Methylation-specific PCR (MSP) and MethylTarget methylation detection. Morris water maze test revealed that DNLA improved learning and memory dysfunction. HE, Nissl, and immunofluorescence staining showed that DNLA alleviated neuron damage and reduced the 5-methylcytosine (5-mC), Aβ1-40) and Aβ1-42) levels. DNLA also decreased the levels of Hcy and Aβ1-42) in the serum, along with decreasing SAM/SAH level in the liver tissue. WB results showed that DNLA down-regulated the expression of amyloid-precursor protein (APP), presenilin-1 (PS1), beta-secretase-1 (BACE1), DNA methyltransferase1 (DNMT1), Aβ1-40) and Aβ1-42) proteins. DNLA also up-regulated the proteins expression of insulin-degrading enzyme (IDE), neprilysin (NEP), DNMT3a and DNMT3b. Meanwhile, DNLA increased CPG island methylation levels of APP and BACE1 genes. DNLA alleviated AD-like symptoms induced by HMD via the DNA methylation pathway.

Altered DNA Methylation and Gene Expression Profiles in Radiation-Induced Heart Fibrosis of Sprague-Dawley Rats.

Radiation-induced heart disease (RIHD) is a serious side effect of radiotherapy for thoracic tumors. Advanced myocardial fibrosis in the late phase of RIHD can lead to myocardial remodeling, heart function impairing and heart failure, resulting in serious clinical consequences, and its pathogenesis remains vague. DNA methylation is one of the important epigenetic mechanisms which often occurs in response to environmental stimuli and is crucial in regulating gene expression. We hypothesized DNA methylation may contribute to pathogenesis in radiation-induced heart fibrosis (RIHF) and altered DNA methylation patterns probably influenced the genes expression in RIHF. In present study, we found genome-wide differences in DNA methylation status and RNA expression were demonstrated and we screened out 44 genes whose altered expression maybe were regulated by CpG island methylation within the gene promoter in RIHF of Sprague-Dawley rat by employing gene expression arrays and human CpG island microarrays. Gene expression and CpG island methylation levels of several candidate genes were further validated. Our investigation provided a new dimension to reveal the specific mechanisms of RIHF and explore the potential therapeutic targets for it.

Circular RNAs, a molecule with past and future

Circular RNAs (circRNAs) are a class of recently identified RNAs that are highly expressed in eukaryotes and conserved between species. Their biogenesis is described as a product of a process called back-splicing that occurs during pre-mRNA maturation, in which the binding 3' donor and 5' acceptor sites produce a circular molecule. Their closed-loop structure gives them greater stability and makes them resistant to degradation by some RNAsas in contrast to linear RNAs. Different functions of circRNAs have been postulated such as splicing competition against the gene from which they derive, miRNA sponges, regulators of CpG island methylation, or mediators in pAKT migration to the nucleus. Therefore, circRNAs are considered important post-transcriptional regulators for the regulation of cell survival and have been used as biomarkers in cancer diseases. Their abundance in neural tissue also relates them to the development of different pathologies of the nervous system that are shown in older people. The aim of this review is to summarize current concepts about circRNAs in eukaryotes, their possible functions and to highlight their importance in different systems such as cardiac, nervous, endocrine and digestive. Moreover, their impact on cardiogenesis, neurogenesis and cancer is discussed to be taken into account in molecular biology future research.

Diabetes mellitus in relation to colorectal tumor molecular subtypes: A pooled analysis of more than 9000 cases.

Diabetes is an established risk factor for colorectal cancer. However, colorectal cancer is a heterogeneous disease and it is not well understood whether diabetes is more strongly associated with some tumor molecular subtypes than others. A better understanding of the association between diabetes and colorectal cancer according to molecular subtypes could provide important insights into the biology of this association. We used data on lifestyle and clinical characteristics from the Colorectal Cancer Family Registry (CCFR) and the Genetics and Epidemiology of Colorectal Cancer Consortium (GECCO), including 9756 colorectal cancer cases (with tumor marker data) and 9985 controls, to evaluate associations between reported diabetes and risk of colorectal cancer according to molecular subtypes. Tumor markers included BRAF and KRAS mutations, microsatellite instability and CpG island methylator phenotype. In the multinomial logistic regression model, comparing colorectal cancer cases to cancer‐free controls, diabetes was positively associated with colorectal cancer regardless of subtype. The highest OR estimate was found for BRAF‐mutated colorectal cancer, n = 1086 (ORfully adj: 1.67, 95% confidence intervals [CI]: 1.36‐2.05), with an attenuated association observed between diabetes and colorectal cancer without BRAF‐mutations, n = 7959 (ORfully adj: 1.33, 95% CI: 1.19‐1.48). In the case only analysis, BRAF‐mutation was differentially associated with diabetes (P difference = .03). For the other markers, associations with diabetes were similar across tumor subtypes. In conclusion, our study confirms the established association between diabetes and colorectal cancer risk, and suggests that it particularly increases the risk of BRAF‐mutated tumors.

Aberrant promoter methylation contributes to LRIG1 silencing in basal/triple-negative breast cancer

BackgroundLRIG1, the founding member of the LRIG (leucine-rich repeat and immunoglobulin-like domain) family of transmembrane proteins, is a negative regulator of receptor tyrosine kinases and a tumour suppressor. Decreased LRIG1 expression is consistently observed in cancer, across diverse tumour types, and is linked to poor patient prognosis. However, mechanisms by which LRIG1 is repressed are not fully understood. Silencing of LRIG1 through promoter CpG island methylation has been reported in colorectal and cervical cancer but studies in breast cancer remain limited.MethodsIn silico analysis of human breast cancer patient data were used to demonstrate a correlation between DNA methylation and LRIG1 silencing in basal/triple-negative breast cancer, and its impact on patient survival. LRIG1 gene expression, protein abundance, and methylation enrichment were examined by quantitative reverse-transcription PCR, immunoblotting, and methylation immunoprecipitation, respectively, in breast cancer cell lines in vitro. We examined the impact of global demethylation on LRIG1 expression and methylation enrichment using 5-aza-2’-deoxycytidine. We also examined the effects of targeted demethylation of the LRIG1 CpG island, and transcriptional activation of LRIG1 expression, using the RNA guided deadCas9 transactivation system.ResultsAcross breast cancer subtypes, LRIG1 expression is lowest in the basal/triple-negative subtype so we investigated whether differential methylation may contribute to this. Indeed, we find that LRIG1 CpG island methylation is most prominent in basal/triple-negative cell lines and patient samples. Use of the global demethylating agent 5-aza-2’-deoxycytidine decreases methylation leading to increased LRIG1 transcript expression in basal/triple-negative cell lines, while having no effect on LRIG1 expression in luminal/ER-positive cell lines. Using a CRISPR/deadCas9 (dCas9)-based targeting approach, we demonstrate that TET1-mediated demethylation (Tet1-dCas9) along with VP64-mediated transcriptional activation (VP64-dCas9) at the CpG island, increased endogenous LRIG1 expression in basal/triple-negative breast cancer cells, without transcriptional upregulation at predicted off-target sites. Activation of LRIG1 by the dCas9 transactivation system significantly increased LRIG1 protein abundance, reduced site-specific methylation, and reduced cancer cell viability. Our findings suggest that CRISPR-mediated targeted activation may be a feasible way to restore LRIG1 expression in cancer.ConclusionsOur study contributes novel insight into mechanisms which repress LRIG1 in triple-negative breast cancer and demonstrates for the first time that targeted de-repression of LRIG1 in cancer cells is possible. Understanding the epigenetic mechanisms associated with repression of tumour suppressor genes holds potential for the advancement of therapeutic approaches.

Serrated Colorectal Lesions: An Up-to-Date Review from Histological Pattern to Molecular Pathogenesis

Until 2010, colorectal serrated lesions were generally considered as harmless lesions and reported as hyperplastic polyps (HPs) by pathologists and gastroenterologists. However, recent evidence showed that they may bear the potential to develop into colorectal carcinoma (CRC). Therefore, the World Health Organization (WHO) classification has identified four categories of serrated lesions: hyperplastic polyps (HPs), sessile serrated lesions (SSLs), traditional serrated adenoma (TSAs) and unclassified serrated adenomas. SSLs with dysplasia and TSAs are the most common precursors of CRC. CRCs arising from serrated lesions originate via two different molecular pathways, namely sporadic microsatellite instability (MSI) and the CpG island methylator phenotype (CIMP), the latter being considered as the major mechanism that drives the serrated pathway towards CRC. Unlike CRCs arising through the adenoma–carcinoma pathway, APC-inactivating mutations are rarely shown in the serrated neoplasia pathway.

Evaluation of methylated DCR1 as a biomarker for response to adjuvant irinotecan-based therapy in stage III colon cancer: cancer and leukaemia Group B 89803 (Alliance)

ABSTRACT Aberrantly methylated genes contribute to the landscape of epigenetic alterations in colorectal adenocarcinoma. The global CpG Island methylator phenotype (CIMP) and individually methylated genes are potential prognostic/predictive biomarkers. Research suggests an association between methylated DCR1 (mDCR1) and lack of benefit with irinotecan (IFL) treatment. We assessed the association between DCR1 methylation status and survival in patients receiving adjuvant fluorouracil/ leucovorin (5-FU/LV) or IFL. We analysed data from patients with stage III colon adenocarcinoma randomly assigned to adjuvant 5-FU/LV or IFL in CALGB 89803 (Alliance). The primary endpoint was overall survival (OS), and the secondary endpoint was disease-free survival (DFS). Using tumour sample DNA, we evaluated the association between survival, DCR1 methylation status, and molecular subgroups (BRAF, KRAS, mismatch repair status, CIMP status) using Kaplan–Meier estimator and Cox proportional hazard model. mDCR1 was observed in 221/400 (55%) colon cancers. Histopathologic features were similar between mDCR1 and unmethylated DCR1 (unDCR1) colon cancers. There was no difference in OS (p = 0.83) or DFS (p = 0.85) based on DCR1 methylation status. There was no association between methylation status and response to IFL . In patients with unDCR1 and KRAS-wildtype tumours, those who received IFL had a nearly two-fold worse DFS compared to patients who received 5-FU/LV (HR = 1.85, 95% CI (0.97–3.53, p = 0.06). This relationship was not notable among other subgroups. In stage III colon cancer patients, mDCR1 status did not associate with response to irinotecan. Larger studies may suggest an association between the iridocene response and molecular subgroups.

Sessile serrated lesions: Clinicopathological characteristics, endoscopic diagnosis, and management.

The 2019 World Health Organization (WHO) Classification of Tumours of the Digestive System (5th edition) introduced the term "sessile serrated lesion" (SSL) to replace the term "sessile serrated adenoma/polyp" (SSA/P). SSLs are early precursor lesions in the serrated neoplasia pathway that result in colorectal carcinomas with BRAF mutations, methylation for DNA repair genes, a CpG island methylator phenotype, and high levels of microsatellite instability. Some of these lesions can rapidly become dysplastic or invasive carcinomas that exhibit high lymphatic invasion and lymph node metastasis potential. The 2019 WHO classification noted that dysplasia arising in an SSL most likely is an advanced polyp, regardless of the morphologic grade of the dysplasia. Detecting SSLs with or without dysplasia is critical; however, detection of SSLs is challenging, and theiridentification by endoscopists and pathologists is inconsistent. Furthermore, indications for their endoscopic treatment have not been established. Moreover, SSLs are considered to contribute to the development of post-colonoscopy colorectal cancers. Herein, the clinicopathological and endoscopic characteristics of SSLs, including features determined using white light and image-enhanced endoscopy, therapeutic indications, therapeutic methods, and surveillance are reviewed based on the literature. This information may lead to more intensive research to improve detection, diagnosis, and rates of complete resection of these lesions and reduce post-colonoscopy colorectal cancer rates.

Global DNA methylation profiles of\xa0buffalo (Bubalus bubalis)\xa0preimplantation embryos produced by handmade cloning and in vitro fertilization

Somatic cell nuclear transfer technique (SCNT) has proved to be an outstanding method of multiplication of elite animals but accompanied with low efficiency and live birth rate of cloned animals. Epigenetic alterations of DNA has been one of the culprits behind this issue. Cloned embryos are found to deviate slightly from regular pattern of demethylation and re-methylation at the time of nuclear reprogramming and embryonic development when compared with embryos produced by in vitro fertilization (IVF). Thus, the present study was aimed at evaluating global DNA methylation profiles of cloned embryos at 2-cell, 8-cell and blastocyst stages and compare it with corresponding stages of embryos produced by IVF by using MeDIP-Sequencing on Illumina-based platform. We found out that cloned embryos exhibited significantly different DNA methylation pattern as compared to IVF embryos with respect to distribution of differentially methylated regions in different components of genome, CpG islands distribution and methylation status, gene ontological profiles and pathways affected throughout the developmental stages. The data generated from MeDIP-Seq was validated at blastocyst stage cloned and IVF embryos by bisulfite-sequencing PCR on five randomly selected gene regions.

High‐fat, sucrose and salt‐rich diet during rat spermatogenesis lead to the development of chronic kidney disease in the female offspring of the F2 generation

Effects of feeding male rats during spermatogenesis a high-fat, high-sucrose and high-saltdiet (HFSSD) over two generations (F0 and F1) on renal outcomes are unknown. Male F0 and F1 rats were fed either control diet (F0CD+F1CD) or HFSSD (F0HD+F1HD). The outcomes were glomerular filtration rate and urinary albumin excretion in F1 and F2 offspring. If both outcomes were altered a morphological and molecular assessment was done. F2 offspring of both sexes had a decreased GFR. However, increased urinary albumin excretion was only observed in female F2 F0HD+F1HD offspring compared with controls. F0HD+F1HD female F2 offspring developed glomerulosclerosis (+31%; p<.01) and increased renal interstitial fibrosis (+52%; p<.05). RNA sequencing followed by qRT-PCR validation showed that four genes (Enpp6, Tmem144, Cd300lf, and Actr3b) were differentially regulated in the kidneys of female F2 offspring. lncRNA XR-146683.1 expression decreased in female F0HD+F1HD F2 offspring and its expression was (r=0.44, p=.027) correlated with the expression of Tmem144. Methylation of CpG islands in the promoter region of the Cd300lf gene was increased (p=.001) in female F2 F0HD+F1HD offspring compared to controls. Promoter CpG island methylation rate of Cd300lf was inversely correlated with Cd300lf mRNA expression in F2 female offspring (r=-0.483, p=.012). Cd300lf mRNA expression was inversely correlated with the urinary albumin-to-creatinine ratio in female F2 offspring (r=-0.588, p=.005). Paternal pre-conceptional unhealthy diet given for two generations predispose female F2 offspring to chronic kidney disease due to epigenetic alterations of renal gene expression. Particularly, Cd300lf gene promotor methylation was inversely associated with Cd300lf mRNA expression and Cd300lf mRNA expression itself was inversely associated with urinary albumin excretion in F2 female offspring whose fathers and grandfathers got a pre-conceptional unhealthy diet.

Targeting DNA Methylation as an Epigenetic Leukocyte Counting Tool.

The methylation of CpG islands in gene promoters is a well-stablished epigenetic mechanism essential to mammalian development. During development, a dynamic process involving both de novo DNA methylation and demethylation in the genome affects the pattern of DNA methylation. This process results in differentiated cells with distinctive and stable DNA methylation signatures that regulate tissue-specific transcriptional programming of genes (1). Specific patterns of DNA methylation can therefore be explored as valuable biomarkers to stratify cell lineage and cell subsets in complex samples such as whole blood. DNA methylation markers that can be used to distinguish and quantify T cells, B cells, monocytes, eosinophils, basophils, neutrophils, and natural killer cells may have a broad clinical application. Leukocyte counts are usually determined based on morphological analysis, flow cytometric measurements, and immunophenotypic classification of lymphocyte subsets. New techniques based on DNA methylation signature may complement and help to overcome several limitations from conventional methods.

DNA methyltransferase 1 inhibits microRNA-497 and elevates GPRC5A expression to promote chemotherapy resistance and metastasis in breast cancer

BackgroundAbnormal DNA methylation of tumor suppressor gene promoter has been found in breast cancer. Therefore, the current study set out to explore how DNA methyltransferase 1 (DNMT1) affects breast cancer through mediating miR-497/GPRC5A axis.MethodsAfter loss and gain-of-function approaches were conducted in MCF-7/ADR and MCF-7 cells, cell viability, IC50 value, invasion, migration and apoptosis were measured, respectively. In addition, drug resistance, metastasis and apoptosis-related protein expression were examined using immunoblotting. ChIP and dual-luciferase reporter gene assays were carried out to validate relationship among DNMT1, miR-497, and GPRC5RA. Subcutaneous xenograft tumor model in nude mice was established to detect effects of DNMT1 on growth and metastasis of breast cancer in vivo.ResultsIt was found that DNMT1 was notably increased, while miR-497 was poorly-expressed in breast cancer. Highly-expressed DNMT1 could promote chemotherapy resistance and metastasis of breast cancer. Meanwhile, DNMT1 modified methylation of CpG island in miR-497 promoter region, thereby repressing miR-497 level. In addition, miR-497 targeted GPRC5A expression to curb chemotherapy resistance and metastasis of breast cancer cells. Lastly, in vivo experiments showed that knockdown of DNMT1 could suppress breast cancer growth and metastasis.ConclusionsCollectively, our findings indicated that DNMT1 may inhibit miR-497 and boost the expression of GPRC5A through methylation, thus augmenting breast cancer chemotherapy resistance and metastasis, which provides novel mechanistic insight into the unrecognized roles of DNMT1 in breast cancer.

Epigenetic changes in the promoter of the fragile histidine triad (FHIT) gene in human sebocytes under the influence of in vitro culture

Due to the lack of tumor suppressor function of the fragile histidine triad (FHIT) gene product, sebaceous gland carcinomas can develop. The model of the sebocyte cell line SZ95 was used to identify methylated CpG islands at the 5'-end of the FHIT gene and the decrease of gene expression as well as the increase of double-stranded (ds) DNA breaks were examined. Methylation, immunofluorescence analysis, promotor sequencing and treatment of SZ95 cells with 5‑azacytidine/trichostatinA (TSA). The cultivation was accompanied by an increasing methylation of the CpG islands, adecrease of the FHIT gene expression and an accumulation of ds-DNA breaks. Treatment with 5‑azacytidine/TSA showed adecrease in DNA methylation and are-expression of FHIT transcripts. Epigenetic changes in the cellular genome are caused by in vitro cell culture. Consequently, apositive selection of sebocytes with an epigenetically inactivated FHIT locus occurs.

SDHB knockout and succinate accumulation are insufficient for tumorigenesis but dual SDHB/NF1 loss yields SDHx-like pheochromocytomas.

SUMMARYInherited pathogenic succinate dehydrogenase (SDHx) gene mutations cause the hereditary pheochromocytoma and paraganglioma tumor syndrome. Syndromic tumors exhibit elevated succinate, an oncometabolite that is proposed to drive tumorigenesis via DNA and histone hypermethylation, mitochondrial expansion, and pseudohypoxia-related gene expression. To interrogate this prevailing model, we disrupt mouse adrenal medulla SDHB expression, which recapitulates several key molecular features of human SDHx tumors, including succinate accumulation but not 5hmC loss, HIF accumulation, or tumorigenesis. By contrast, concomitant SDHB and the neurofibromin 1 tumor suppressor disruption yields SDHx-like pheochromocytomas. Unexpectedly, in vivo depletion of the 2-oxoglutarate (2-OG) dioxygenase cofactor ascorbate reduces SDHB-deficient cell survival, indicating that SDHx loss may be better tolerated by tissues with high antioxidant capacity. Contrary to the prevailing oncometabolite model, succinate accumulation and 2-OG-dependent dioxygenase inhibition are insufficient for mouse pheochromocytoma tumorigenesis, which requires additional growth-regulatory pathway activation.

1α,25(OH)2D3 reverses exhaustion and enhances antitumor immunity of human cytotoxic T cells

BackgroundEpidemiological surveys have revealed that low serum vitamin D level was correlated with increased risk of tumors. Dysfunctional T cells in patients with tumor are characterized as exhausted with high...

CpG Island Methylation of the Rap1Gap Gene in Medullary Thyroid Cancer

Medullary thyroid cancer (MTC) is a rare type of neuroendocrine tumor. This study aimed to investigate the gene and protein expression of RAP1GAP and DNA methylation patterns of its CpG74a , CpG74b , and CpG24 in an Iranian population with MTC. In this case-control study, we selected 55 individuals who underwent thyroidectomy in Erfan hospital, Tehran, between 2018 and 2020. Samples were divided into normal thyroid tissues (control; n=20), benign nodule (n=20), and MTC (n=15). DNA methylation patterns were investigated using MSP (methylation-specific PCR). The protein level and mRNA expression of RAP1GAP were also evaluated using western blotting and real-time PCR, respectively. The hyper-methylation rates of CpG24 and CpG74a in the MTC samples were considerably higher than the controls (83% versus 15% and 74% versus 17%, respectively; P<0.001). The methylation/unmethylation ratio of CpG74a , and CpG24 was considerably higher than the controls (P<0.001). The methylation/unmethylation ratio of CpG24 in the benign nodules was also considerably greater than the controls (P<0.001). The mRNA expression and the protein level of RAP1GAP in the MTC group were considerably lower than the controls (P=0.005 and P=0.035, respectively). In the MTC group, aberrant methylation of CpG74a and CpG24 was significantly correlated with decreasing expression of the Rap1Gap gene (R2 : 0.23; P=0.032 and R2 : 0.56; P=0.001, respectively). Hyper-methylation in CpG24 and CpG74a and decreasing expression of RAP1GAP can be considered as diagnostic biomarkers for MTC.

Role of Epigenetic alterations in the development of cancers

Many different factors are involved in the progression of cancers. Genes mutations and chromosomal abnormalities are normally considered main cause of cancers but there are some other reason for the development of cancers. Other cancer causing factors are known as epigenetic alterations [1,2]. Epigentic modification of genome is known as epigenetic alterations, lead toward cancer cells production. Epigentic modification does not cause change in sequences of nucleotide. Similar to genetic alteration epigenetic alteration can’t be ignored [3]. Basically mechanisms behind epigenetic modifications are deregulation of DNA proteins, change in CpG island methylation, change in histone, oncogenes activation and deactivation of tumor suppressor [4].&#x0D; Epigenetic alterations is directly linked with functional alterations of genome. Alteration in DNA methylation, histone degeneration and functional and structural abnormalities of chromosomes are the major examples of epigenetic modifications [5]. The main function of all epigenetic alterations is to modulate gene expression with same DNA sequences. Means these changes never effect main basal sequence oF DNA [6], which remain same in cell division [7].&#x0D; Many different types of cancers contains large number of epigenetic alterations, the most important of these are epigenetic alterations that occurs in DNA repair genes. These DNA repair genes drive slow expression of DNA proteins. These abnormalities cause genetic unreliability, which is mainly considered as characteristic of various cancers [8,9].

The methylation of SDC2 and TFPI2 defined three methylator phenotypes of colorectal cancer

BackgroundMethylated SDC2 and TFPI2 are widely used for colorectal cancer (CRC) detection. However, they often miss some CRCs, which directly diminishes the sensitivity. Further investigations of the underlying mechanisms leading to the missed samples will facilitate developing more eligible methylation markers.MethodsCRC samples from TCGA and GEO datasets were divided into three groups, High-methylation/ High-methylation (HH), High-methylation/Low-methylation (HL), and Low-methylation/Low-methylation (LL) according to the methylation status of SDC2 and TFPI2 promoters. Variations in age, tumor location and microsatellite instable were then assessed between the three groups and verified in our custom cohort.ResultsSamples of HL group preferred to derive from left-sided CRCs (P < 0.05). HH samples showed the highest microsatellite instability and mutation load (mean nonsynonymous mutations for HH/HL/LL: 10.55/3.91/7.02, P = 0.0055). Almost all mutations of BRAF, one of the five typical CpG island methylator phenotype (CIMP) related genes, were observed in HH group (HH/HL/LL: 51/0/1, P = 0.018). Besides, older patients were frequently found in HH group. Expression analysis identified 37, 84, and 22 group-specific differentially expressed genes (DEGs) for HH, HL, and LL, respectively. Functional enrichment analysis revealed that HH-specific DEGs were mainly related to transcription regulation, while LL-specific DEGs were enriched in the biological processes of extracellular matrix interaction and cell migration.ConclusionsThe current study revealed that the performance of methylation-based markers might be affected by tumor location, patient age, mutation load and MSI, and these respective sides should be considered when developing new methylation markers for CRC detection.

Interaction between the HIF-1α gene rs1957757 polymorphism and CpG island methylation in the promoter region is associated with the risk of anti-tuberculosis drug-induced liver injury in humans: A case-control study.

HIF-1α gene polymorphisms, including rs11549465, rs11549467, rs1957757 and rs10873142, cause liver cell damage or pulmonary disease. The aim of this study was to analyse the association between the polymorphisms of the loci of the HIF-1α gene and its CpG island methylation in the promoter region with the risk of anti-tuberculosis drug-induced liver injury (ADLI). 286 patients with tuberculosis (TB) and ADLI (case group) and 286 patients with TB but without liver injury (control group) were matched one-to-one, among the 1728 TB patients recruited from July 2019 to July 2020. Genotyping of the four loci of the HIF-1α gene was confirmed using PCR-RFLP technology. Methylation of the HIF-1α gene was measured using the MSP method. The comparison of risk factors, HIF-1α genotype and methylation status between the case group and the control group was all achieved through univariate and multivariate conditional logistic regression. Univariate analysis showed that the frequency of rs1957757 mutation genotype and CpG island methylation was significantly higher in the case group than in the control group (P<0.001, all). In contrast, there was no statistical difference in the frequency of mutated genotypes at the other three loci between the two groups (p=0.21, p=0.12 and p=0.55, respectively). Further, multivariate analysis showed that CpG islands were methylated, the mutation genotype of the rs1957757 locus was independently associated with the high risk of ADLI, and the adjusted OR (95%CI) reached 1.92 (1.32-2.63) and 2.01 (1.32-2.83), respectively. Furthermore, taking the rs1957757 locus wild genotype and CpG islands without methylation as the reference group, the mutation genotype and CpG island methylation increased the risk of ADLI, and the probability of ADLI could reach 4.73 times that of the reference group. This is the first demonstration of the association of HIF-1α gene polymorphism and CpG island methylation with ADLI risk stratification. The interaction between CpG islands methylated in the promoter region of the HIF-1α gene and its rs1957757 locus mutant genotype was associated with a higher risk of ADLI.

Stage-differentiated ensemble modeling of DNA methylation landscapes uncovers salient biomarkers and prognostic signatures in colorectal cancer progression

Aberrant DNA methylation acts epigenetically to skew the gene transcription rate up or down, contributing to cancer etiology. A gap in our understanding concerns the epigenomics of stagewise cancer progression. In this study, we have developed a comprehensive computational framework for the stage-differentiated modelling of DNA methylation landscapes in colorectal cancer (CRC). The methylation β-matrix was derived from the public-domain TCGA data, converted into M-value matrix, annotated with AJCC stages, and analysed for stage-salient genes using an ensemble of approaches involving stage-differentiated modelling of methylation patterns and/or expression patterns. Differentially methylated genes (DMGs) were identified using a contrast against controls (adjusted p-value <0.001 and |log fold-change of M-value| >2), and then filtered using a series of all possible pairwise stage contrasts (p-value <0.05) to obtain stage-salient DMGs. These were then subjected to a consensus analysis, followed by matching with clinical data and performing Kaplan-Meier survival analysis to evaluate the impact of methylation patterns of consensus stage-salient biomarkers on disease prognosis. We found significant genome-wide changes in methylation patterns in cancer cases relative to controls agnostic of stage. The stage-differentiated models yielded the following consensus salient genes: one stage-I gene (FBN1), one stage-II gene (FOXG1), one stage-III gene (HCN1) and four stage-IV genes (NELL1, ZNF135, FAM123A, LAMA1). All the biomarkers were significantly hypermethylated in the promoter regions, indicating down-regulation of expression and implying a putative CpG island Methylator Phenotype (CIMP) manifestation. A prognostic signature consisting of FBN1 and FOXG1 survived all the analytical filters, and represents a novel early-stage epigenetic biomarker / target. We have designed and executed a workflow for stage-differentiated epigenomic analysis of colorectal cancer progression, and identified several stage-salient diagnostic biomarkers, and an early-stage prognostic biomarker panel. The study has led to the discovery of an alternative CIMP-like signature in colorectal cancer, reinforcing the role of CIMP drivers in tumor pathophysiology.