Expression Of DNA Methyltransferase Research Articles

Correlation between promoter methylation of the LDH-C4 gene and DNMT expression in breast cancer and their prognostic significance.

DNA methylation plays an important role in tumorigenesis and development. The potential of aberrant DNA methylation to act as a biomarker for tumor diagnosis and prognostic evaluation is currently being explored. Lactate dehydrogenase C4 (LDH-C4) is a member of the cancer/testis antigen family that is expressed in a broad range of human tumor types, with particularly high expression patterns observed in lung cancer, melanoma and breast cancer. However, whether the methylation of the promoter region of LDH-C4 can be used as a tumor marker and its association with prognosis remain poorly understood. The present study aimed to determine the potential of the methylation status of LDH-C4 and DNA methyltransferase (DNMT) expression to be biological markers for the prognostic evaluation of breast cancer. Methylation-specific PCR was conducted to evaluate alterations in the methylation levels of LDH-C4. Immunohistochemical analysis was performed to assess the expression levels of DNMTs in breast cancer tissues. The association between the methylation status of LDH-C4 or the expression of DNMTs, and clinical pathological parameters was also evaluated. The results of the present study revealed that the demethylation rate of LDH-C4 in breast cancer tissues was significantly increased compared with that of adjacent tissues, and associations were identified between the demethylation of LDH-C4 and the histological grade, estrogen receptor, progesterone receptor and HER-2 status, and lymph node metastasis. The level of LDH-C4 demethylation was negatively correlated with the expression of DNMTs. Demethylation of the LDH-C4 promoter and DNMT expression predicted an unfavorable prognosis of patients with breast cancer. In addition, demethylation of the LDH-C4 promoter, high expression of DNMT3a and DNMT3b, histological grade and lymph node metastasis were all discovered to be independent prognostic factors in patients with breast cancer. In conclusion, results of the present study indicated that the demethylation of LDH-C4 and DNMT expression levels may be closely associated with the occurrence and development of breast cancer, in addition to lymph node metastasis. Thus, both may be used to assist the clinical evaluation of prognosis.

DNA Methylome Mapping Identifies Epigenetic Abnormalities in Intestinal Lymphocyte Regulation in Human Necrotizing Enterocolitis.

Epigenetic changes occur in response to environmental factors during the pathogenesis of necrotizing enterocolitis (NEC) in animal models, but the DNA methylation signature in human patients with NEC has not been examined. To illustrate the signature and function of DNA methylation in the intestine of human NEC. DNA methyltransferases (DNMTs) were compared between intestinal tissue with NEC and control. Genome-wide DNA methylation was analyzed by reduced representation bisulfite sequencing (RRBS). The biological functions of the potential methylation regulated genes were analyzed by Gene Ontology. Gene methylation and expression were confirmed by bisulfite genomic sequencing (BGS) and RT-qPCR. By screening the expression of DNMTs, we identified a marked reduction in DNMT3A at both the mRNA and protein levels in NEC. Genome-wide variation of DNA methylation was detected in NEC lesions. The CG methylation level in almost all unique regions except CpG islands (CGIs) was lower in NEC compared with control. A total of 287 differentially methylated regions (DMRs) were identified across the whole genome in NEC, 123 of them are located on the CGI in the promoter. The DMR-associated genes were linked to intestinal epithelial permeability, platelet aggregation, and lymphocyte proliferation. Four genes (ZNF335, MPL, RASAL3, and KDM6A) with roles in the regulation of lymphocytes that may predispose the intestine to imbalanced immune processes were further confirmed to be hypermethylated and transcriptionally downregulated. These findings underscore the novel relationship between epigenetic changes and lymphocyte regulation in human NEC, which may have potential diagnostic and therapeutic relevance for NEC.

Methylation and Expression of Mutant FUS in Motor Neurons Differentiated From Induced Pluripotent Stem Cells From ALS Patients.

Amyotrophic lateral sclerosis (ALS) is a rapidly progressive disease leading to degeneration of motor neurons (MNs). Epigenetic modification of gene expression is increasingly recognized as potential disease mechanism. In the present study we generated motor neurons from induced pluripotent stem cells from ALS patients carrying a mutation in the fused in sarcoma gene (FUS) and analyzed expression and promoter methylation of the FUS gene and expression of DNA methyltransferases (DNMTs) compared to healthy control cell lines. While mutant FUS neural progenitor cells (NPCs) did not show a difference in FUS and DNMT expression compared to healthy controls, differentiated mutant FUS motor neurons showed significantly lower FUS expression, higher DNMT expression and higher methylation of the proximal FUS gene promoter. Immunofluorescence revealed perceived proximity of cytoplasmic FUS aggregates in ALS MNs together with 5-methylcytosin (5-mC). Targeting disturbed methylation in ALS may therefore restore transcriptional alterations and represent a novel therapeutic strategy.

Olaparib Is a Mitochondrial Complex I Inhibitor That Kills Temozolomide-Resistant Human Glioblastoma Cells.

Glioblastoma represents the highest grade of brain tumors. Despite maximal resection surgery associated with radiotherapy and concomitant followed by adjuvant chemotherapy with temozolomide (TMZ), patients have a very poor prognosis due to the rapid recurrence and the acquisition of resistance to TMZ. Here, initially considering that TMZ is a prodrug whose activation is pH-dependent, we explored the contribution of glioblastoma cell metabolism to TMZ resistance. Using isogenic TMZ-sensitive and TMZ-resistant human glioblastoma cells, we report that the expression of O6-methylguanine DNA methyltransferase (MGMT), which is known to repair TMZ-induced DNA methylation, does not primarily account for TMZ resistance. Rather, fitter mitochondria in TMZ-resistant glioblastoma cells are a direct cause of chemoresistance that can be targeted by inhibiting oxidative phosphorylation and/or autophagy/mitophagy. Unexpectedly, we found that PARP inhibitor olaparib, but not talazoparib, is also a mitochondrial Complex I inhibitor. Hence, we propose that the anticancer activities of olaparib in glioblastoma and other cancer types combine DNA repair inhibition and impairment of cancer cell respiration.

Blood-Based Epigenetic Markers of FKBP5 Gene Methylation in Patients With Dilated Cardiomyopathy.

BackgroundBlood‐based DNA methylation patterns are linked to types of diseases. FKBP prolyl isomerase 5 (FKBP5), a protein cochaperone, is known to be associated with the inflammatory response, but the regulatory mechanisms by leukocyte FKBP5 DNA methylation in patients with dilated cardiomyopathy (DCM) remain unclear.Methods and ResultsThe present study enrolled patients with DCM (n=31) and age‐matched and sex‐matched control participants (n=43). We assessed FKBP5 CpG (cytosine‐phosphate‐guanine) methylation of CpG islands at the 5′ side as well as putative promoter regions by methylation‐specific quantitative polymerase chain reaction using leukocyte DNA isolated from the peripheral blood. FKBP5 CpG methylation levels at the CpG island of the gene body and the promoter regions were significantly decreased in patients with DCM. Leukocyte FKBP5 and IL‐1β (interleukin 1β) mRNA expression levels were significantly higher in patients with DCM than in controls. The protein expressions of DNMT1 (DNA methyltransferase 1) and DNMT3A (DNA methyltransferase 3A) in leukocytes were significantly reduced in patients with DCM. In vitro methylation assay revealed that FKBP5 promoter activity was inhibited at the methylated conditions in response to immune stimulation, suggesting that the decreased FKBP5 CpG methylation was functionally associated with elevation of FKBP5 mRNA expressions. Histological analysis using a mouse model with pressure overload showed that FKBP5‐expressing cells were substantially infiltrated in the myocardial interstitium in the failing hearts, indicating a possible role of FKBP5 expressions of immune cells in the cardiac remodeling.ConclusionsOur findings demonstrate a link between specific CpG hypomethylation of leukocyte FKBP5 and DCM. Blood‐based epigenetic modification in FKBP5 may be a novel molecular mechanism that contributes to the pathogenesis of DCM.

Regulation of the cellular redox state and the expression of DNA methyltransferase-1 in peripheral blood mononuclear cells from patients with Graves’ disease

BackgroundGraves’ disease is an autoimmune disorder characterised by excessive production of thyroid hormones, which induces increased cellular metabolism in most tissues and increased production of reactive oxygen species (ROS). The aim of this work was to analyse the effect of ROS on cell viability and the expression of catalase (CAT), glutathione peroxidase-1 (GPx-1), superoxide dismutase (SOD-1) and DNA methyltransferase-1 (DNMT-1) in peripheral blood mononuclear cells (PBMC) from patients with newly diagnosed Graves’ disease or treated with methimazole. Patients and methodsFor this study, women patients with newly diagnosed Graves’ disease (n=18), treated with methimazole (n=6) and healthy subjects (n=15) were recruited. ROS were evaluated by flow cytometry, and the viability/apoptosis of PBMC was analysed by flow cytometry and fluorescence microscopy. Genomic expression of CAT, GPx-1, SOD-1 and DNMT-1 was quantified by real-time PCR. ResultsWe found high levels of ROS and increased expression of CAT, GPx-1, SOD-1 and DNMT-1 in PBMC from patients with newly diagnosed Graves’ disease. Methimazole treatment reversed these parameters. Cell viability was similar in all study groups. ConclusionsROS induces the expression of CAT, GPx-1, and SOD-1. The activity of these enzymes may contribute to the protection of PBMC from the harmful effect of free radicals on cell viability. Increased expression of DNMT-1 may be associated with aberrant methylation patterns in immunoregulatory genes contributing to autoimmunity in Graves’ disease.

Expression of DNMTs and H3K9ac in Ameloblastoma and Ameloblastic Carcinoma

Objectives: DNA methyltransferases (DNMTs) and the histone modification H3K9ac are epigenetic markers. This study aimed to describe the immunohistochemical expression of DNMT1, DNMT3A, DNMT3B, and H3K9ac in the dental follicle (DF), ameloblastoma (AME), and ameloblastic carcinoma (AC), correlating these expressions with the recurrence and aggressive behavior in ameloblastoma.Study Design: Immunohistochemical reactions were performed in 10 human DFs, 38 ameloblastomas, and 6 AC samples. Another 59 ameloblastomas assembled in a tissue microarray were used to compare the immunoexpression with the clinical, radiographic, and histopathological characteristics and the presence of BRAFv600e mutation. Each slide was digitized as a high-resolution image and quantified by Aperio ScanScope Nuclear V9 software. All statistical analyzes were performed using GraphPad Prism statistical software.Results: DNMT3B expression was higher in ameloblastomas than in the DFs, while the AC overexpressed all proteins. The ameloblastomas with BRAFv600e mutation, vestibular/lingual, or vestibular/palatine bone cortical disruption and maxilla involvement showed DNMT1 overexpression, while recurrent cases had high DNMT3B levels.Conclusions: DNA methylation and histone modification might play a role in the development, clinical aggressiveness, and recurrence rates of ameloblastoma, such as the progression to AC. Further investigation about gene methylations in ameloblastomas is needed to better understand its relationship with aggressiveness and recurrence.

Resveratrol ameliorates toxic effects of cadmium on placental development in mouse placenta and human trophoblast cells.

Cadmium (Cd) is a common heavy metal pollutant. Prenatal exposure to Cd results in adverse effects on fetal development. Placental apoptosis, inflammation, and epigenetic disruption have been implicated in Cd-induced placental toxicity. Resveratrol (Res) is a naturally occurring polyphenol with anti-apoptotic, anti-inflammatory, and epigenetic regulatory activities. In present study, the effects of Res on placental toxicity induced by Cd were evaluated. Pregnant CD-1 mice were fed with base diet containing 0.2% Res started on gestational day 0 (GD0), and intraperitoneally injected with 4.5mg/kg CdCl2 or saline once on GD9. JEG-3 cells were treated with 20 μM Res for 24 hr in the absence or presence of 20 μM CdCl2 for the second 12 hr. The fetal outcomes, the apoptosis in placenta and JEG-3 cells, the expression of inflammatory cytokines and chemokines including tumor necrosis factor-α (TNF-α), interferon-gamma (IFN-γ), monocyte chemoattractant protein 1 (MCP-1), macrophage inflammatory protein-2 (MIP-2) and chemokine (C-X-C motif) ligand 1 (KC), and expression of endoplasmic reticulum (ER) stress markers were evaluated. The expression and activities of DNA methyltransferase (DNMT), and the activation of Akt signaling pathway were detected. Cd exposure resulted in decreased fetal weight and crown-rump length while Res ameliorated these outcomes. Res suppressed Cd-induced apoptosis in placenta and JEG-3 cells, and decreased Cd-induced expression of TNF-α, IFN-γ, MCP-1, MIP-2, and KC in placenta. Cd greatly increased ER stress in placenta in mice, which was partially ameliorated by Res treatment. Res decreased Cd-induced upregulation of DNMT activity and suppressed Cd-induced expression of DNMT3B. Res restored estradiol secretion, enhances activity and protein levels of SIRT1 and inhibited Cd-induced activation of Akt signaling pathway. Res ameliorated Cd-induced placental toxicity and regulated DNMT3 expression and PI3K/Akt pathway activation.

Cyclophosphamide inhibits Pax5 methylation to regulate the growth of retinoblastoma via the Notch1 pathway.

Retinoblastoma (Rb) is the most common intraocular malignant tumor in infants. Here, we investigated the function and mechanism of cyclophosphamide (CTX) in the development of Rb. Real-time quantitative polymerase chain reaction (RT-qPCR) results showed that paired box protein 5 (Pax5) expression was down-regulated in Rb tissues and cell lines. Methylation-specific PCR (MSP) results showed that the methylation level of Pax5 was up-regulated in Rb. After treatment with CTX, the Pax5 expression in Rb cell lines was increased significantly. The methylation of Pax5 and the expression of DNA methyltransferases (DNMTs) were down-regulated in the CTX group. Cyclophosphamide inhibited cell proliferation, migration, and invasion, promoted cell apoptosis via the Notch1 pathway. DNA methyltransferase inhibitor SGI-1027 had synergistic effects with CTX. Paired box protein 5 siRNA was transfected into Y79 cells treated with CTX. The expression of DNMTs, Pax5, the Notch1 pathway and apoptosis marker protein was detected by Western blotting, and changes in cell behavior were detected, respectively. Results showed that knockdown of Pax5 reversed the effects of CTX. Moreover, the Notch1 activator Valproic acid (VPA) abolished the inhibitory effects of CTX on Rb development. Moreover, CTX inhibited tumor growth in nude mice. These findings demonstrated that CTX up-regulated Pax5 expression by down-regulating DNMTs expression, and then inhibited the Notch1 signaling pathway activation and Rb growth.

Global histone modification analysis reveals hypoacetylated H3 and H4 histones in B Cells from systemic lupus erythematosus patients

ObjectiveHistone modification is an epigenetic alteration which either activates or suppresses gene transcription. Studies revealed the association of altered global histone modification in T cells and monocytes with the pathogenesis of Systemic lupus erythematosus (SLE). Herein, we investigated the level of global histone 3 (H3) and histone 4 (H4) acetylation in B cells of SLE patients. MethodsTotal 20 SLE patients and 10 healthy donors were recruited. Global H3 and H4 acetylation in B cells was assessed by Epigentek assay kits. Expression of DNA methyltransferase 1 (DNMT1) in B cells was analyzed by staining cells with anti-CD19/20 and anti-DNMT1 antibody. The concentration of BAFF and APRIL was measured using LegendPlex Human B cells panel and circulating ANAs were determined using indirect immunofluorescence. ResultsCompared to healthy donors, B cells from SLE patients were found to be hypoacetylated on both H3 and H4 histones together with a decrease in the expression of DNMT1. Indeed, stratification of SLE patients on the basis of disease activity did not show any variation, as the amount of H3 and H4 acetylation in both inactive and active SLE patients was almost uniform. ConclusionThese findings suggest that SLE-B cells were manifested with aberrant histone acetylation levels.

Misregulation of the expression and activity of DNA methyltransferases in cancer.

In mammals, DNA methyltransferases DNMT1 and DNMT3’s (A, B and L) deposit and maintain DNA methylation in dividing and nondividing cells. Although these enzymes have an unremarkable DNA sequence specificity (CpG), their regional specificity is regulated by interactions with various protein factors, chromatin modifiers, and post-translational modifications of histones. Changes in the DNMT expression or interacting partners affect DNA methylation patterns. Consequently, the acquired gene expression may increase the proliferative potential of cells, often concomitant with loss of cell identity as found in cancer. Aberrant DNA methylation, including hypermethylation and hypomethylation at various genomic regions, therefore, is a hallmark of most cancers. Additionally, somatic mutations in DNMTs that affect catalytic activity were mapped in Acute Myeloid Leukemia cancer cells. Despite being very effective in some cancers, the clinically approved DNMT inhibitors lack specificity, which could result in a wide range of deleterious effects. Elucidating distinct molecular mechanisms of DNMTs will facilitate the discovery of alternative cancer therapeutic targets. This review is focused on: (i) the structure and characteristics of DNMTs, (ii) the prevalence of mutations and abnormal expression of DNMTs in cancer, (iii) factors that mediate their abnormal expression and (iv) the effect of anomalous DNMT-complexes in cancer.

Exposure to carbon black nanoparticles during pregnancy aggravates lipopolysaccharide-induced lung injury in offspring: an intergenerational effect

Carbon black nanoparticles (CBNPs) are one of the most frequently used nanoparticles. Exposure to CBNPs during pregnancy (PrE to CBNPs) can directly induce inflammation, lung injury, and genotoxicity in dams and results in abnormalities in offspring. However, whether exposure to CBNPs during pregnancy enhances the susceptibility of offspring to environmental stimuli remains unknown. To address this issue, in this study, we intranasally treated pregnant mice with mock or CBNPs from gestational day (GD) 9 to GD18, and F1 and F2 offspring were normally obtained. By intratracheal instillation of mice with lipopolysaccharide (LPS) to trigger a classic animal model for acute lung injury, we intriguingly found that after LPS treatment, F1 and F2 offspring after exposure during pregnancy to CBNPs both exhibited more pronounced lung injury symptoms, including more degenerative histopathological changes, vascular leakage, elevated MPO activity, and activation of inflammation-related signaling transduction, compared with F1 and F2 offspring in the mock group, suggesting PrE to CBNPs would aggravate LPS-induced lung injury in offspring, and this effect was intergenerational. We also observed that PrE to CBNPs upregulated the mRNA expression of DNA methyltransferases (Dnmt) 1/3a/3b and DNA hypermethylation in both F1 and F2 offspring, which might partially account for the intergenerational effect. Together, our study demonstrates for the first time that PrE to CBNPs can enhance sensitivity to LPS in both F1 and F2 offspring, and this intergenerational effect may be related to DNA hypermethylation caused by CBNPs.

An Exploratory Pilot Study of Changes in Global DNA Methylation in Patients Undergoing Major Breast Surgery Under Opioid-Based General Anesthesia.

This study aimed to investigate DNA methylation levels in patients undergoing major breast surgery under opioid-based general anesthesia. Blood samples were collected from eleven enrolled patients, before, during and after anesthesia. PBMC were isolated and global DNA methylation levels as well as DNA methyltransferase (DNMT) and cytokine gene expression were assessed. DNA methylation levels significantly declined by 26%, reversing the direction after the end of surgery. Likewise, DNMT1a mRNA expression was significantly reduced at all time points, with lowest level of −68%. DNMT3a and DNMT3b decreased by 65 and 71%, respectively. Inflammatory cytokines IL6 and TNFα mRNA levels showed a trend for increased expression at early time-points to end with a significant decrease at 48 h after surgery. This exploratory study revealed for the first time intraoperative global DNA hypomethylation in patients undergoing major breast surgery under general anesthesia with fentanyl. The alterations of global DNA methylation here observed seem to be in agreement with DNMTs gene expression changes. Furthermore, based on perioperative variations of IL6 and TNFα gene expression, we hypothesize that DNA hypomethylation may occur as a response to surgical stress rather than to opiate exposure.

DNMT family induces down-regulation of NDRG1 via DNA methylation and clinicopathological significance in gastric cancer.

BackgroundAberrant DNA methylation of tumor suppressor genes is a common event in the development and progression of gastric cancer (GC). Our previous study showed NDRG1, which could suppress cell invasion and migration, was frequently down-regulated by DNA methylation of its promoter in GC.Purpose and MethodsTo analyze the relationship between the expression and DNA methylation of NDRG1 and DNA methyltransferase (DNMT) family. We performed a comprehensive comparison analysis using 407 patients including sequencing analysis data of GC from TCGA.ResultsNDRG1 was down-regulated in GC, and was negatively correlative to DNMT1 (r = −0.11, p = 0.03), DNMT3A (r = −0.10, p = 0.01), DNMT3B (r = −0.01, p = 0.88), respectively, whereas the DNA methylation of NDRG1 was positively correlative to DNMT family (DNMT1 r = 0.20, p < 0.01; DNMT3A r = 0.26, p < 0.001; DNMT3B r = 0.03, p = 0.57, respectively). NDRG1 expression was significantly inverse correlated with invasion depth (p = 0.023), but DNMT1 was significantly positive correlated with invasion depth (p = 0.049). DNMT3B was significantly correlated with the degree of tumor cell differentiation (p = 0.030). However, there was no association between the expression of DNMT3A and clinicopathological features. The KM plotter showed that NDRG1 (HR = 0.95, 95% CI [0.8–1.12], p = 0.53) and DNMT1 (HR = 1.04, 95% CI [0.88–1.23], p = 0.67) had no association with prognosis of GC patients, while, DNMT3A (p = 0.0064) and DNMT3B (p = 0.00025) displayed significantly association. But the overall survival of high expression of NDRG1 tended to be prolonged.ConclusionThese data suggest that down-regulation of NDRG1expression in GC may be due to its promoter DNA methylation via DNMT family. The demethylating agent maybe a potential target drug for GC patients.

Fetal e-cigarette exposure programs a neonatal brain hypoxic-ischemic sensitive phenotype via altering DNA methylation patterns and autophagy signaling pathway.

Maternal e-cigarette (e-cig) exposure is a pressing perinatal health concern. Emerging evidence reveals its potential adverse impacts on brain development in offspring, yet the underlying mechanisms are poorly understood. The present study tested the hypothesis that fetal e-cig exposure induces an aberrant DNA methylation profile in the developing brain, leading to alteration of autophagic flux signaling and programming of a sensitive phenotype to neonatal hypoxic-ischemic encephalopathy (HIE). Pregnant rats were exposed to chronic intermittent e-cig aerosol. Neonates were examined at the age of 9 days old. Maternal e-cig exposure decreased the body weight and brain weight but enhanced the brain-to-body weight ratio in the neonates. E-cig exposure induced a gender-dependent increase in hypoxic-ischemia-induced brain injury in male neonates associated with enhanced reactive oxygen species (ROS) activity. It differentially altered DNA methyltransferase expression and enhanced both global DNA methylation levels and specific CpG methylation at the autophagy-related gene 5 (ATG5) promoter. In addition, maternal e-cig exposure caused downregulations of ATG5, microtubule-associated protein 1 light chain 3β, and sirtuin 1 expression in neonatal brains. Of importance, knockdown of ATG5 in neonatal pups exaggerated neonatal HIE. In conclusion, the present study reveals that maternal e-cig exposure downregulates autophagy-related gene expression via DNA hypermethylation, leading to programming of a hypoxic-ischemic sensitive phenotype in the neonatal brain.

Effect of 5'-fluoro-2'-deoxycytidine and sodium butyrate on the genes of the intrinsic apoptotic pathway, p21, p53, cell viability, and apoptosis in human hepatocellular carcinoma cell lines

Backgrounds: Epigenetic regulation such as DNA methylation plays a major role in chromatin organization and gene transcription. Additionally, histone modification is an epigenetic regulator of chromatin structure and influences chromatin organization and gene expression. The relationship between DNA methyltransferase (DNMTs) expression and promoter methylation of the tumor suppressor genes (TSGs) has been reported in various cancers. Previously, the effect of 5-aza-2'-deoxycytidine (5-AZA-CdR), trichostatin A (TSA), and valproic acid (VPA) was shown on various cancers. This study aimed to investigate the effect of 5'-fluoro-2'-deoxycytidine (FdCyd) and sodium butyrate on the genes of the intrinsic apoptotic pathway, p21, p53, cell viability, and apoptosis in human hepatocellular carcinoma SNU449, SNU475, and SNU368 cell lines. Materials and Methods: In this lab trial study, the SNU449, SNU475, and SNU368 cells were cultured and treated with 5'-fluoro-2'-deoxycytidine and sodium butyrate. To determine cell viability, cell apoptosis, and the relative gene expression level, MTT assay, flow cytometry assay, and qRT-PCR were done respectively. Results: 5'-fluoro-2'-deoxycytidine and sodium butyrate changed the expression level of the BAX, BAK, APAF1, Bcl-2, Bcl-xL, p21, and p53 gene (P&lt;0.0001) by which induced cell apoptosis and inhibit cell growth in all three cell lines, SNU449, SNU475, and SNU368. Conclusion: Both compounds played their roles through the intrinsic apoptotic pathway to induce cell apoptosis.

Hsa_circ_0040809 regulates colorectal cancer development by upregulating methyltransferase DNMT1 via targeting miR-515-5p.

Circular RNAs (circRNAs) are key regulators in the progression of various cancers. Abnormal DNA methylation patterns feature prominently in the regulation of the expression of tumor-related genes. This study is aimed at investigating the molecular mechanism of circ_0040809 affecting colorectal cancer (CRC) progression by regulating DNA methyltransferase 1 (DNMT1). circ_0040809 was selected from the circRNA microarray datasets (GSE142837 and GSE138589). Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to examine the expression of circ_0040809, miR-515-5p, and DNMT1 mRNA in paired cancerous and paracancerous tissues of 40 CRC patients, as well as in cell lines. Western blotting was conducted for detecting DNMT1 protein expression in CRC cells. Cell proliferation, migration, and apoptosis were assessed through CCK-8, Transwell, and flow cytometry assays. Bioinformatics and dual-luciferase gene assay were conducted to predict and verify, respectively, the targeted relationships between circ_0040809 and miR-515-5p, as well as between miR-515-5p and DNMT1 mRNA. In CRC tissues and cells, circ_0040809 and DNMT1 expression are markedly increased, whereas miR-515-5p expression is decreased. Also, high circ_0040809 expression is significantly linked to shorter overall survival. Cell function compensation experiments reveal that circ_0040809 silencing inhibits CRC cell proliferation and migration and promotes apoptosis, while circ_0040809 overexpression has the opposite effects. Mechanistically, circ_0040809 competitively binds to miR-515-5p to elevate DNMT1 expression. Rescue assay reveals that overexpressed miR-515-5p partly counteracts the tumor-facilitating impact of circ_0040809. circ_0040809 facilitates CRC cell proliferation and migration, and inhibits apoptosis, through modulating miR-515-5p/DNMT1 axis. Our study implies that targeting circ_0040809 may be a therapy strategy for CRC treatment.

A novel small-molecule antagonist enhances the sensitivity of osteosarcoma to cabozantinib in vitro and in vivo by targeting DNMT-1 correlated with disease severity in human patients

Advanced osteosarcoma (OSA) is highly aggressive and can lead to distant metastasis or recurrence. Here, a novel small-molecule inhibitor/antagonist of DNA methyltransferase 1 (DNMT-1) named DI-1 (inhibitor of DNMT-1) was explored to enhance the antitumor effect of a molecular-targeted agent, cabozantinib, on OSA cell lines. In patients with OSA, expression of DNMT-1 was negatively related with that of microRNA (miR)-34a and associated with a poor prognosis. In OSA cell lines (OSA cell line U2OS and an OSA cell line U2OSR resistance to cabozantinib), DI-1 treatment enhanced miR-34a expression by inhibiting hypermethylation of the promoter region of miR-34a mediated by DNMT-1. DI-1 enhanced the sensitivity of OSA cells (U2OS, 143B and MG63) to cabozantinib and other molecular-targeted agents by enhancing miR-34a expression and repressing activation of the Notch pathway. Mechanistically, DI-1 repressed recruitment of DNMT-1 to the promoter region of miR-34a and, in turn, decreased the methylation rate in the promoter region of miR-34a in OSA cells. These results suggest that repressing DNMT-1 activation by DI-1 enhances miR-34a expression in OSA cells and could be a promising therapeutic strategy for OSA.

A Pan-Cancer Analysis of the Prognostic Value and Expression of Adenylate Cyclase 7 (ADCY7) in Human Tumors.

BackgroundThe role of adenylate cyclase 7 (ADCY7) in cancer is still unclear. This study analyzed the interrelationship between the expression and immune function of ADCY7.MethodsADCY7 expression in multiple human cancers was analyzed using the databases of Genotype-Tissue Expression Project (GTEx), Cancer Cell Line Encyclopedia (CCLE), and The Cancer Genome Atlas (TCGA). Correlations among ADCY7 gene expression, mismatch repair (MMR) gene expression, and DNA methyltransferase (DNMT) expression were assessed using Spearman correlation analysis. Univariate survival analysis and Kaplan–Meier (KM) curve were used to examine the effect of ADCY7 expression on prognosis. The Tumor Immune Estimation Resource (TIMER) database was used to evaluate the relationship between ADCY7 gene expression and tumor immune invasion or immune checkpoint gene (ICG) expression.ResultsADCY7 was abnormally expressed in multiple human cancers and was correlated with MMR genes and DNMT expression. Univariate survival analysis and KM curve showed that ADCY7 expression influences the overall survival (OS) of six types of cancer, disease-specific survival (DSS) of eight, and progression-free interval (PFI) of three. The high expression of ADCY7 in OS, DSS, and PFI was strongly associated with poor outcomes in patients with breast cancer and lung squamous cell carcinoma. ADCY7 expression was strongly associated with immune cell infiltration and ICG expression.ConclusionThe results of this study indicated that ADCY7 may be a prognostic biomarker of tumorigenesis. The study may also provide a new perspective on the role of ADCY7 in human cancers.

High DNMT1 Is Associated With Worse Local Control in Early-Stage Laryngeal Squamous Cell Carcinoma.

Objectives/HypothesisEarly‐stage laryngeal squamous cell carcinoma (LSCC) has yielded local control rates of 75% after radiotherapy. DNA methylation, in which DNA methyltransferases play an important role, has influence on tumorigenesis. In this study, we investigated the association between the expression of DNA methyltransferase 1 (DNMT1) and local control in early‐stage LSCC treated with radiotherapy.Study DesignRetrospective cohort study.MethodsWe analyzed a well‐defined homogeneous series of 125 LSCC patients treated with radiotherapy with curative intent. The association of immunohistochemical expression of DNMT1 with local control was evaluated using Cox proportional hazard regression models.ResultsWith a median follow‐up of 58 months, 29 local recurrences (23%) were observed. On univariate analysis, worse local control was associated with high DNMT1 expression (hazard ratio [HR] 2.57, 95% confidence interval [CI] 1.10–6.01). Also, higher T‐stage (HR 2.48, 95% CI 1.06–5.80) and positive N‐status (HR 2.62, 95% CI 1.06–6.47) were associated with worse local control. Multivariate Cox regression demonstrated that high DNMT1 (HR 2.81; 95% CI 1.20–6.58) was independently associated with worse local control.ConclusionsWe found an association between high DNMT1 expression and worse local control in a homogeneous well‐defined cohort of early‐stage LSCC patients treated with definitive radiotherapy. The association between DNA methylation status as determined by DNMT1 expression and local control suggests that DNMT1 acts as a potential prognostic tumor marker in treatment decision‐making in early‐stage laryngeal carcinoma.Level of evidenceNA Laryngoscope, 132:801–805, 2022