Search Results (65)

Bladder cancer pathogenesis is closely linked to epigenetic alterations, particularly DNA methylation and demethylation processes. Environmental carcinogens and persistent inflammatory stimuli—such as recurrent urinary tract infections—can induce aberrant DNA methylation, altering gene expression profiles and contributing to malignant transformation. This review synthesizes current evidence on the role of DNA methyltransferases (DNMT1, DNMT3a, DNMT3b) and the hypermethylation of key tumour suppressor genes, including A2BP1, NPTX2, SOX11, PENK, NKX6-2, DBC1, MYO3A, and CA10, in bladder cancer. It also evaluates the therapeutic application of DNA-demethylating agents such as 5-azacytidine and highlights the impact of chronic inflammation on epigenetic regulation. Promoter hypermethylation of tumour suppressor genes leads to transcriptional silencing and unchecked cell proliferation. Urine-based DNA methylation assays provide a sensitive and specific method for non-invasive early detection, with single-target approaches offering high diagnostic precision. Animal models are increasingly employed to validate these findings, allowing the study of methylation dynamics and gene–environment interactions in vivo. DNA methylation represents a key epigenetic mechanism in bladder cancer, with significant diagnostic, prognostic, and therapeutic implications. Integration of human and experimental data supports the use of methylation-based biomarkers for early detection and targeted treatment, paving the way for personalized approaches in bladder cancer management. Full article

Background/Objectives: Prostate cancer (PCa) is diagnosed at an earlier median age, more advanced stage, and has worse clinical outcomes in African American (AA) men compared to European Americans (EA). Methods: To investigate the role of aberrant DNA methylation in tumor-adjacent stroma (TAS), methyl binding domain sequencing (MBD-seq) was performed on AA (n = 17) and EA (n = 15) PCa patients. This was independently confirmed using the long interspersed nuclear element-1 (LINE-1) assay. Pathway analysis was performed on statistically significantly differentially methylated genes for AA and EA TAS. DNA methylation profiles of primary cultured AA and EA carcinoma-associated fibroblasts (CAFs) were compared with AA and EA TAS. AA and EA CAFs were treated with demethylating agent 5-Azacytidine (5-AzaC). Results: AA TAS exhibited higher global DNA methylation than EA TAS (p-value < 0.001). Of the 3268 differentially methylated regions identified (DMRs, p-value < 0.05), 85% (2787 DMRs) showed increased DNA methylation in AA TAS, comprising 1648 genes, of which 1379 were protein-coding genes. Based on DNA methylation levels, two AA subgroups were identified. Notably, AA patients with higher DNA methylation were predominantly those with higher Gleason scores. Pathway analysis linked methylated genes in AA TAS to several key signaling pathways (p-value < 0.05), including immune response (e.g., IL-1, IL-15, IL-7, IL-8, IL-3, and chemokine), Wnt/β-catenin, androgen, PTEN, p53, TGF-β, and circadian clock regulation. A total of 168 concordantly methylated genes were identified, with 109 genes (65%) showing increased methylation in AA CAFs and TAS (p-value < 0.05). Treatment with 5-AzaC significantly reduced DNA methylation of concordant genes in AA CAFs (p-value < 0.001). Conclusions: These findings suggest a distinct stromal methylome in AA, providing a foundation for integrating demethylating agents into standard therapies. This approach targets the tumor microenvironment, potentially addressing PCa disparities in AA men. Full article

DNA methylation plays a pivotal role in the epigenetic regulation of gene expression and holds promise for enhancing livestock meat production. In this study, we analyzed the DNA methylome and transcriptome of the longissimus dorsi muscle (LDM) in Duroc pigs with varying growth rates. Our results reveal that DNA methylation suppressed the expression of key muscle development markers (MYOD, MYOG, MHC1) and proliferation markers (PI67, PCNA), as well as the protein expression and phosphorylation of PI3K and AKT (p < 0.05). Dual-luciferase reporter assays and EMSA showed that SP1 overexpression enhanced the luciferase activity of the wild-type LPAR1 promoter, an effect amplified by the demethylating agent 5-AZA (p < 0.05). The EMSA further demonstrates the relationship between SP1 and the LPAR1 promoter region. Overexpression of SP1 upregulated LPAR1 expression at both the mRNA and protein levels (p < 0.05). Knockdown of LPAR1 reduced muscle marker gene expression and delayed myotube formation, while silencing SP1 disrupted the expression of LPAR1, MEF2C, and MHC1 (p < 0.05), and the demethylation induced by 5-AZA partially reversed these effects. These findings suggest that the DNA methylation/SP1/LPAR1 axis is critical for skeletal muscle growth and development, underscoring the regulatory role of DNA methylation in muscle formation. Full article

Ethylene-insensitive 3/ethylene-insensitive 3-like (EIN3/EIL) transcription factors are central regulators of ethylene signaling and stress adaptation in plants. However, their roles in Osmanthus fragrans, a globally cherished ornamental and aromatic plant with significant economic value, remain poorly characterized. Here, we identified nine OfEIL genes across eight chromosomes in the O. fragrans “Liuye Jingui” genome. Conserved motif analysis revealed core domains (Motif1/2/4/7), and promoter cis-elements highlighting hormone-related, stress-related, and growth-related regulatory potential. During late flowering stages, six OfEILs (3/4/5/6/7/9) were significantly upregulated. Under 5-azacytidine (AZA, a DNA demethylation agent), OfEIL2 and OfEIL7 were downregulated, whereas the ETH treatment activated OfEIL3/7/8/9. Strikingly, OfEIL7 exhibited dual regulatory roles, correlating strongly with natural flowering progression, AZA-induced demethylation, and ETH responses. Functional divergence was observed in petal senescence, with OfEIL2–5 and OfEIL7–9 showing stage-specific and tissue-specific expression patterns. These results position OfEIL7 as a key hub integrating epigenetic and hormonal signals to modulate floral longevity and stress adaptation. Our study provides the first genome-wide characterization of the EIL family in O. fragrans, offering critical insights for molecular breeding aimed at enhancing ornamental traits and environmental resilience in this economically significant species. Full article

Skin cancer prevalence has increased during the last decades, with the last years serving as a pivotal moment for comprehending its epidemiological patterns and its impact on public health. Melanoma is one of the most frequently occurring malignancies, arising from a complex interplay of genetic factors, environmental factors, lifestyle and socio-economic conditions. Epigenetic changes play a critical role in tumor development, influencing progression and aggressiveness. Epigenetic therapies could represent novel therapeutic options, while drug repositioning may serve as a viable strategy for cancer treatment. Demethylating agents, commonly used in hematological malignancies, show promising results on solid tumors, including melanoma. Methylation patterns are responsible for tumor development by modulating gene expression, while histone acetylation influences DNA processes such as transcription, replication, repair, and recombination. This review aims to identify existing potential therapeutical approaches using therapeutic agents that can modulate DNA methylation and histone modification, which can lead to tumor inhibition, cell death initiation and reactivation of tumor suppressor genes. Full article

Breast cancer (BC) is a widespread malignancy that affects the lives of millions of women each year, and its resulting financial and healthcare hardships cannot be overstated. These issues, in combination with side effects and obstacles associated with the current standard of care, generate considerable interest in new potential targets for treatment as well as means for BC prevention. One potential preventive compound is Withaferin A (WFA), a traditional medicinal compound found in winter cherries. WFA has shown promise as an anticancer agent and is thought to act primarily through its effects on the epigenome, including, in particular, the methylome. However, the relative importance of specific genes’ methylation states to WFA function remains unclear. To address this, we utilized human BC cell lines in combination with CRISPR-dCas9 fused to DNA methylation modifiers (i.e., epigenetic editors) to elucidate the importance of specific genes’ promoter methylation states to WFA function and cancer cell viability. We found that targeted demethylation of promoters of the tumor suppressors p21 and p53 within MDA-MB-231/MCF7 cells resulted in around 1.7×/1.5× and 1.2×/1.3× increases in expression, respectively. Targeted methylation of the promoter of the oncogene CCND1 within MDA-MB-231/MCF7 cells resulted in 0.5×/0.8× decreases in gene expression. These changes to p21, p53, and CCND1 were also associated with decreases in cell viability of around 25%/50%, 5%/35%, and 12%/16%, respectively, for MDA-MB-231/MCF7 cells. When given in combination with WFA in both p53 mutant and wild type cells, we discovered that targeted methylation of the p21 promoter was able to modulate the anticancer effects of WFA, while targeted methylation or demethylation of the promoters of p53 and CCND1 had no significant effect on viability decreases from WFA treatment. Taken together, these results indicate that p21, p53, and CCND1 may be important targets for future in vivo studies that may lead to epigenetic editing therapies and that WFA may have utility in the prevention of BC through its effect on p21 promoter methylation independent of p53 function. Full article

Background: Epigenetic dysregulation is a common feature of cancer. Promoter demethylation of tumor-promoting genes and global DNA hypomethylation may trigger tumor progression. Epigenetic changes are unstable; thus, research has focused on detecting remedies that target epigenetic regulators. Previous studies have suggested that concordantly targeting hypomethylation and hypermethylation is beneficial for suppressing both the oncogenic and pro-metastatic functions of cancer cells. Therefore, we aimed to investigate the effect of a combination of S-adenosylmethionine (SAM) and the demethylating agent decitabine on prostate cancer cells. Materials and Methods: Prostate cancer cells (PC-3) were treated with SAM, decitabine, or a combination of both. Proliferation, migration, invasion, and methylation assays were also performed. A transcriptome study was conducted to detect different gene clusters between the treatment groups, followed by analyses using the Kyoto Encyclopedia of Genes and Genomes pathway and ingenuity pathway analysis. Finally, to gain information on differential gene expression, promoter methylation studies were performed. Results: Groups treated with decitabine, SAM, or their combination showed reduced proliferative capacity. The decitabine-treated group showed a marginal increase in cell migration and invasion, whereas the SAM-treated and combination treatment groups showed reduced invasion and migration potential. Methylation assays demonstrated the restoration of decitabine-induced demethylation in prostate cancer samples, whereas the transcriptome study revealed the upregulation of different gene clusters between the treatment groups. Methylation studies confirmed that SAM could restore the decitabine-induced demethylation of proto-oncogenes, but it did not induce the re-methylation of tumor-suppressor genes. Conclusions: Combination treatment with SAM and decitabine had an additive effect and did not nullify each other. Full article

Objective: Bisphenol A and phthalate are known endocrine disruptors and capable of inducing epigenetic changes in the human population. However, their impact on the placenta is less well studied. Our objective was to measure the effect of exposure to bisphenol A and benzyl butyl phthalate in first-trimester HTR8-SVneo and third-trimester 3A-sub E trophoblast cells by profiling the DNA methylation pattern of the imprinting control region of the IGF2 (insulin-like growth factor) and H19 genes. Methods: Human placental HTR8-SVneo and 3A-sub E cell lines were treated with two sub-lethal concentrations of bisphenol A and benzyl butyl phthalate. Demethylating agent, 5-azacytidine, was used as a positive control. Cells were harvested on post-treatment days 1 and 4. The methylation profile of six CpG dinucleotide sites, part of the CTCF 6 binding site of the IGF2/H19 imprinting control region, was determined by pyrosequencing. Results: In the first-trimester HTR8-SVneo cell line, we observed a significant increased methylation of the CpG sites 3, 4 when treated with a high concentration of bisphenol A or benzyl butyl phthalate while increased methylation at site 6 for both high and low dose treatment on day 4. Demethylation of the CpG sites 1, 4, and 6 was observed when treated with 5-azacytidine on day 4. In the third-trimester 3A-sub E cell line, no significant changes in the methylation profile were observed under any treatment conditions. Conclusions: The results of this study demonstrate the capability of epigenetic changes in human placenta cells induced by bisphenol A and benzyl butyl phthalate. The observed methylation changes only in the first-trimester HTR8-SVneo cells phthalate may reflect a window of epigenetic susceptibility related to these environmental toxicants. Full article

Epigenetic changes are common in cancer and include aberrant DNA methylation and histone modifications, including both acetylation or methylation. DNA methylation in the promoter regions and histone deacetylation are usually accompanied by gene silencing, and may lead to the suppression of tumor suppressors in cancer cells. An interaction between epigenetic pathways has been reported that could be exploited to more efficiently target aggressive cancer cells, particularly those against which current treatments usually fail, such as pancreatic cancer. In this study, we explored the possibility to combine the DNA demethylating agent 5-AZA with HDAC inhibitor SAHA to treat pancreatic cancer cell lines, focusing on the acetylation of mutp53 and the consequences on its stability, as well as on the interaction of this protein with c-myc and BRCA-1, key molecules in cancer survival. The results obtained suggest that SAHA/5-AZA combination was more effective than single treatments to promote the degradation of mutp53, to upregulate p21 and downregulate c-Myc and BRCA-1, thus increasing DNA damage and cytotoxicity in pancreatic cancer cells. Full article

Epigenetic modifications, including aberrant DNA methylation occurring at the promoters of oncogenes and oncosuppressor genes and histone modifications, can contribute to carcinogenesis. Aberrant methylation mediated by histone methylatransferases, alongside histones, can affect methylation of proteins involved in the regulation of pro-survival pathways such as JAK/STAT and contribute to their activation. In this study, we used DNA or histone demethylating agents, 5-Azacytidine (5-AZA) or DS-3201 (valemetostat), respectively, to treat primary effusion lymphoma (PEL) cells, alone or in combination with AG490, a Signal transducer and activator of transcription 3 (STAT3) inhibitor. Cell viability was investigated by trypan blue assay and FACS analysis. The molecular changes induced by 5-AZA and/or AG490 treatments were investigated by Western blot analysis, while cytokine release by PEL cells treated by these drugs was evaluated by Luminex. Statistical analyses were performed with Graphpad Prism^® software (version 9) and analyzed by Student’s t test or a nonparametric one-way ANOVA test. The results obtained in this study suggest that 5-AZA upregulated molecules that inhibit STAT3 tyrosine phosphorylation, namely Suppressor of Cytokine Signaling 3 (SOCS3) and tyrosine–protein phosphatase non-receptor type (PTPN) 6/Src homology region 2 domain-containing phosphatase-1 (SHP-1), reducing STAT3 activation and downregulating several STAT3 pro-survival targets in PEL cells. As this lymphoma is highly dependent on the constitutive activation of STAT3, 5-AZA impaired PEL cell survival, and when used in combination with AG490 JAK2/STAT3 inhibitor, it potentiated its cytotoxic effect. Differently from 5-AZA, the inhibition of the EZH1/2 histone methyltransferase by DS-3201, reported to contribute to STAT3 activation in other cancers, slightly affected STAT3 phosphorylation or survival in PEL cells, either alone or in combination with AG490. This study suggests that 5-AZA, by upregulating the expression level of SOCS3 and PTPN6/SHP1, reduced STAT3 activation and improved the outcome of treatment targeting this transcription factor in PEL cells. Full article

Cutaneous melanoma is rapidly on the rise globally, surpassing the growth rate of other cancers, with metastasis being the primary cause of death in melanoma patients. Consequently, understanding the mechanisms behind this metastatic process and exploring innovative treatments is of paramount importance. Recent research has shown promise in unravelling the role of epigenetic factors in melanoma progression to metastasis. While DNA hypermethylation at gene promoters typically suppresses gene expression, we have contributed to establishing the newly understood mechanism of paradoxical activation of genes via DNA methylation, where high methylation coincides with increased gene activity. This mechanism challenges the conventional paradigm that promoter methylation solely silences genes, suggesting that, for specific genes, it might actually activate them. Traditionally, altering DNA methylation in vitro has involved using global demethylating agents, which is insufficient for studying the mechanism and testing the direct consequence of gene methylation changes. To investigate promoter hypermethylation and its association with gene activation, we employed a novel approach utilising a CRISPR-SunTag All-in-one system. Here, we focused on editing the DNA methylation of a specific gene promoter segment (EBF3) in melanoma cells using the All-in-one system. Using bisulfite sequencing and qPCR with RNA-Seq, we successfully demonstrated highly effective methylation and demethylation of the EBF3 promoter, with subsequent gene expression changes, to establish and validate the paradoxical role of DNA methylation. Further, our study provides novel insights into the function of the EBF3 gene, which remains largely unknown. Overall, this study challenges the conventional view of methylation as solely a gene-silencing mechanism and demonstrates a potential function of EBF3 in IFN pathway signalling, potentially uncovering new insights into epigenetic drivers of malignancy and metastasis. Full article

Epigenetic modifications, mainly aberrant DNA methylation, have been shown to silence the expression of genes involved in epigenetic diseases, including cancer suppression genes. Almost all conventional cancer therapeutic agents, such as the DNA hypomethylation drug 5-aza-2-deoxycytidine, have insurmountable side effects. To investigate the role of the well-known DNA protectant (ectoine) in skin cell DNA methylation and cancer cell proliferation, comprehensive methylome sequence analysis, 5-methyl cytosine (5mC) analysis, proliferation and tumorigenicity assays, and DNA epigenetic modifications-related gene analysis were performed. The results showed that extended ectoine treatment globally hypomethylated DNA in skin cells, especially in the CpG island (CGIs) element, and 5mC percentage was significantly reduced. Moreover, ectoine mildly inhibited skin cell proliferation and did not induce tumorigenicity in HaCaT cells injected into athymic nude mice. HaCaT cells treated with ectoine for 24 weeks modulated the mRNA expression levels of Dnmt1, Dnmt3a, Dnmt3b, Dnmt3l, Hdac1, Hdac2, Kdm3a, Mettl3, Mettl14, Snrpn, and Mest. Overall, ectoine mildly demethylates DNA in skin cells, modulates the expression of epigenetic modification-related genes, and reduces cell proliferation. This evidence suggests that ectoine is a potential anti-aging agent that prevents DNA hypermethylation and subsequently activates cancer-suppressing genes. Full article

Epigenetic dysregulation, particularly alterations in DNA methylation and hydroxymethylation, plays a pivotal role in cancer initiation and progression. Ten-eleven translocation (TET) proteins catalyze the successive oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) and further oxidized methylcytosines in DNA, thereby serving as central modulators of DNA methylation–demethylation dynamics. TET loss of function is causally related to neoplastic transformation across various cell types while its genetic or pharmacological activation exhibits anti-cancer effects, making TET proteins promising targets for epigenetic cancer therapy. Here, we developed a robust cell-based screening system to identify novel TET activators and evaluated their potential as anti-cancer agents. Using a carefully curated library of 4533 compounds provided by the National Cancer Institute, Bethesda, MD, USA, we identified mitoxantrone as a potent TET agonist. Through rigorous validation employing various assays, including immunohistochemistry and dot blot studies, we demonstrated that mitoxantrone significantly elevated 5hmC levels. Notably, this elevation manifested only in wild-type (WT) but not TET-deficient mouse embryonic fibroblasts, primary bone marrow-derived macrophages, and leukemia cell lines. Furthermore, mitoxantrone-induced cell death in leukemia cell lines occurred in a TET-dependent manner, indicating the critical role of TET proteins in mediating its anti-cancer effects. Our findings highlight mitoxantrone’s potential to induce tumor cell death via a novel mechanism involving the restoration of TET activity, paving the way for targeted epigenetic therapies in cancer treatment. Full article

The recent evolution of molecular targeted therapy has improved clinical outcomes in several human malignancies. The translocation of anaplastic lymphoma kinase (ALK) was originally identified in anaplastic large-cell lymphoma (ALCL) and subsequently in non-small cell lung carcinoma (NSCLC). Since ALK fusion gene products act as a driver of carcinogenesis in both ALCL and NSCLC, several ALK tyrosine kinase inhibitors (TKIs) have been developed. Crizotinib and alectinib are first- and second-generation ALK TKIs, respectively, approved for the treatment of ALK-positive ALCL (ALK⁺ ALCL) and ALK⁺ NSCLC. Although most ALK⁺ NSCLC patients respond to crizotinib and alectinib, they generally relapse after several years of treatment. We previously found that DNA-demethylating agents enhanced the efficacy of ABL TKIs in chronic myeloid leukemia cells. Moreover, aberrant DNA methylation has also been observed in ALCL cells. Thus, to improve the clinical outcomes of ALK⁺ ALCL therapy, we investigated the synergistic efficacy of the combination of alectinib and the DNA-demethylating agent azacytidine, decitabine, or OR-2100 (an orally bioavailable decitabine derivative). As expected, the combination of alectinib and DNA-demethylating agents synergistically suppressed ALK⁺ ALCL cell proliferation, concomitant with DNA hypomethylation and a reduction in STAT3 (a downstream target of ALK fusion proteins) phosphorylation. The combination of alectinib and OR-2100 markedly altered gene expression in ALCL cells, including that of genes implicated in apoptotic signaling, which possibly contributed to the synergistic anti-ALCL effects of this drug combination. Therefore, alectinib and OR-2100 combination therapy has the potential to improve the outcomes of patients with ALK⁺ ALCL. Full article

Temozolomide (TMZ) is an important first-line treatment for glioblastoma (GBM), but there are limitations to TMZ response in terms of durability and dependence on the promoter methylation status of the DNA repair gene O⁶-methylguanine DNA methyltransferase (MGMT). MGMT-promoter-hypermethylated (MGMT-M) GBMs are more sensitive to TMZ than MGMT-promoter-hypomethylated (MGMT-UM) GBMs. Moreover, TMZ resistance is inevitable even in TMZ-sensitive MGMT-M GBMs. Hence, epigenetic reprogramming strategies are desperately needed in order to enhance TMZ response in both MGMT-M and MGMT-UM GBMs. In this study, we present novel evidence that the epigenetic reactivation of Tumor Suppressor Candidate 3 (TUSC3) can reprogram sensitivity of GBM stem cells (GSCs) to TMZ irrespective of MGMT promoter methylation status. Interrogation of TCGA patient GBM datasets confirmed TUSC3 promoter regulation of TUSC3 expression and also revealed a strong positive correlation between TUSC3 expression and GBM patient survival. Using a combination of loss-of-function, gain-of-function and rescue studies, we demonstrate that TUSC3 reactivation is associated with enhanced TMZ response in both MGMT-M and MGMT-UM GSCs. Further, we provide novel evidence that the demethylating agent 5-Azacitidine (5-Aza) reactivates TUSC3 expression in MGMT-M GSCs, whereas the combination of 5-Aza and MGMT inhibitor Lomeguatrib is necessary for TUSC3 reactivation in MGMT-UM GSCs. Lastly, we propose a pharmacological epigenetic reactivation strategy involving TUSC3 that leads to significantly prolonged survival in MGMT-M and MGMT-UM orthotopic GSCs models. Collectively, our findings provide a framework and rationale to further explore TUSC3-mediated epigenetic reprogramming strategies that could enhance TMZ sensitivity and outcomes in GBM. Mechanistic and translational evidence gained from such studies could contribute towards optimal design of impactful trials for MGMT-UM GBMs that currently do not have good treatment options. Full article