Epigenetic Homeostasis Research Articles

Evaluating the connection between diet quality, EpiNutrient intake and epigenetic age: an observational study

BackgroundDNA methylation (DNAm) has unique properties which makes it a potential biomarker for lifestyle-related exposures. Epigenetic clocks, particularly DNAm-based biological age predictors [epigenetic age (EA)], represent an exciting new area of clinical research and deviations of EA from chronological age [epigenetic age acceleration (EAA)] have been linked to overall health, age-related diseases, and environmental exposures. ObjectivesThis observational study investigates the relationships between biological aging and various dietary factors within the LifeLines-DEEP Cohort. These factors include diet quality, processed food consumption, dietary glycemic load, and intake of vitamins involved in maintaining the epigenetic homeostasis (vitamins B-9, B-12, B-6, B-2, and C). MethodsDietary records collected using food-frequency questionnaires were used to estimate diet quality [LifeLines Diet Score (LLDS)], measure the intake of unprocessed/ultraprocessed food according to the NOVA food classification system, and the adequacy of the dietary intake of vitamins B-9, B-12, B-2, B-6, and C. EA using Horvath, Hannum, Levine, and Horvath2 epigenetic clock models and DNAm-predicted telomere length (DNAm-TL) were calculated from DNAm data in 760 subjects. Associations between dietary factors and EAA were tested, adjusting for sex, energy intake, and body composition. ResultsLLDS was associated with EAA (EAA_Horvath: β: −0.148; P = 1 × 10−4; EAA_Hannum: β: −0.148; P = 9 × 10−5; EAA_Levine: β: −0.174; P = 1 × 10−5; and EAA_Horvath2: β: −0.176; P = 4 × 10−6) and DNAm-TL (β: 0.116; P = 0.003). Particularly, EAA was associated with dietary glycemic load (EAA_Horvath: β: 0.476; P = 9 × 10−10; EAA_Hannum: β: 0.565; P = 1 × 10−13; EAA_Levine: β: 0.469; P = 5 × 10−9; EAA_Horvath2: β: 0.569; P = 1 × 10−13; and DNAmTL adjusted for age: β: −0.340; P = 2 × 10−5) and different measures of food processing (NOVA classes 1 and 4). Positive EAA was also associated with inadequate intake of vitamin B-12 (EAA_Horvath: β: −0.167; P = 0.002; EAA_Hannum: β: −0.144; P = 0.007; and EAA_Horvath2: β: −0.126; P = 0.019) and C (EAA_Hannum: β: −0.136; P = 0.010 and EAA_Horvath2: β: −0.151; P = 0.005). ConclusionsOur findings corroborate the hypothesis that nutrition plays a pivotal role in influencing epigenetic homeostasis, especially DNAm, thereby contributing to individual health trajectories and the pace of aging.

RBMS1 reflects a distinct microenvironment and promotes tumor progression in ocular melanoma

Ocular melanoma, including uveal melanoma (UM) and conjunctival melanoma (CM), is the most common ocular cancer among adults with a high rate of recurrence and poor prognosis. Loss of epigenetic homeostasis disturbed gene expression patterns, resulting in oncogenesis. Herein, we comprehensively analyzed the DNA methylation, transcriptome profiles, and corresponding clinical information of UM patients through multiple machine-learning algorithms, finding that a methylation-driven gene RBMS1 was correlated with poor clinical outcomes of UM patients. RNA-seq and single-cell RNA-seq analyses revealed that RBMS1 reflected diverse tumor microenvironments, where high RBMS1 expression marked an immune active TME. Furthermore, we found that tumor cells were identified to have the higher communication probability in RBMS1+ state. The functional enrichment analysis revealed that RBMS1 was associated with pigment granule and melanosome, participating in cell proliferation as well as apoptotic signaling pathway. Biological experiments were performed and demonstrated that the silencing of RBMS1 inhibited ocular melanoma proliferation and promoted apoptosis. Our study highlighted that RBMS1 reflects a distinct microenvironment and promotes tumor progression in ocular melanoma, contributing to the therapeutic customization and clinical decision-making.

The Loss of an Orphan Nuclear Receptor NR2E3 Augments Wnt/β-catenin Signaling via Epigenetic Dysregulation that Enhances Sp1-β catenin-p300 Interactions in Hepatocellular Carcinoma.

The orphan nuclear receptor NR2E3 (Nuclear receptor subfamily 2 group E, Member 3) is an epigenetic player that modulates chromatin accessibility to activate p53 during liver injury. Nonetheless, a precise tumor suppressive and epigenetic role of NR2E3 in hepatocellular carcinoma (HCC) development remains unclear. HCC patients expressing low NR2E3 exhibit unfavorable clinical outcomes, aligning with heightened activation of the Wnt/β-catenin signaling pathway. The murine HCC models utilizing NR2E3 knockout mice consistently exhibits accelerated liver tumor formation accompanied by enhanced activation of Wnt/β-catenin signaling pathway and inactivation of p53 signaling. At cellular level, the loss of NR2E3 increases the acquisition of aggressive cancer cell phenotype and tumorigenicity and upregulates key genes in the WNT/β-catenin pathway with increased chromatin accessibility. This event is mediated through increased formation of active transcription complex involving Sp1, β-catenin, and p300, a histone acetyltransferase, on the promoters of target genes. These findings demonstrate that the loss of NR2E3 activates Wnt/β-catenin signaling at cellular and organism levels and this dysregulation is associated with aggressive HCC development and poor clinical outcomes. In summary, NR2E3 is a novel tumor suppressor with a significant prognostic value, maintaining epigenetic homeostasis to suppress the Wnt/β-catenin signaling pathway that promotes HCC development.

Abstract 3011: The loss of an orphan nuclear receptor NR2E3 augments Wnt/β-Catenin signaling via epigenetic dysregulation that links to the Sp1-β catenin-p300 interactions in hepatocellular carcinoma

Abstract Hepatocellular carcinoma (HCC) stands as the fourth leading cause of global cancer-related mortality due to late-stage detection, tumor heterogeneity, and drug resistance. The pivotal role of epigenetic factors in HCC development and progression necessitates the identification of critical epigenetic regulators. The orphan nuclear receptor NR2E3 (Nuclear receptor subfamily 2 group E, Member 3), primarily associated with retinal development, has emerged as a potential tumor suppressor. Earlier studies showed that the loss of NR2E3 leads to epigenetic repression of the estrogen receptor α (ER) through the involvement of Lysine Specific histone Demethylase 1 (LSD1). NR2E3 also forms a transcriptional complex with Sp1, governing the aryl hydrocarbon receptor (AHR) expression, a crucial factor in liver xenobiotic metabolism. In vivo, NR2E3 ablation results in p53 inactivation and severe liver injuries when exposed to liver toxicants, highlighting its role in activating p53. Nonetheless, the precise tumor suppressive and epigenetic role of NR2E3 in HCC remains unclear. HCC patients expressing low NR2E3 exhibit unfavorable clinical outcomes, aligning with heightened activation of the Wnt/β-catenin signaling pathway. The murine HCC models utilizing NR2E3 knockout mice show accelerated liver tumor formation and progression accompanied by enhanced activation of the Wnt/β-catenin signaling pathway and inactivation of the p53 signaling pathway. At a cellular level, losing NR2E3 increases the acquisition of aggressive cancer cell phenotype and tumorigenicity and upregulates vital genes in the Wnt/β-catenin pathway with enhanced chromatin accessibility. This event is mediated through the increased formation of active transcription complex involving Sp1, β-catenin, and p300, a histone acetyltransferase, on the promoters of target genes. Correspondingly, treatment of p300 inhibitor C646 decreased the expression of critical genes in the Wnt/β-catenin signaling pathway. These findings demonstrate that the loss of NR2E3 promotes Wnt/β-catenin signaling activation at cellular, organismal, and clinical levels. In summary, NR2E3 is a novel tumor suppressor that maintains epigenetic homeostasis, thereby preventing activation of Wnt/β-catenin signaling that promotes HCC formation. Citation Format: Yuet-Kin Leung, Sung-Gwon Lee, Jiang Wang, Shuk-Mei Ho, Nancy Rusch, Chungoo Park, Kyounghyun Kim. The loss of an orphan nuclear receptor NR2E3 augments Wnt/β-Catenin signaling via epigenetic dysregulation that links to the Sp1-β catenin-p300 interactions in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3011.

Tet methylcytosine dioxygenase 2 (TET2) deficiency elicits EGFR-TKI (tyrosine kinase inhibitors) resistance in non-small cell lung cancer

Despite epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI) have shown remarkable efficacy in patients with EGFR-mutant non-small cell lung cancer (NSCLC), acquired resistance inevitably develops, limiting clinical efficacy. We found that TET2 was poly-ubiquitinated by E3 ligase CUL7FBXW11 and degraded in EGFR-TKI resistant NSCLC cells. Genetic perturbation of TET2 rendered parental cells more tolerant to TKI treatment. TET2 was stabilized by MEK1 phosphorylation at Ser 1107, while MEK1 inactivation promoted its proteasome degradation by enhancing the recruitment of CUL7FBXW11. Loss of TET2 resulted in the upregulation of TNF/NF-κB signaling that confers the EGFR-TKI resistance. Genetic or pharmacological inhibition of NF-κB attenuate the TKI resistance both in vitro and in vivo. Our findings exemplified how a cell growth controlling kinase MEK1 leveraged the epigenetic homeostasis by regulating TET2, and demonstrated an alternative path of non-mutational acquired EGFR-TKI resistance modulated by TET2 deficiency. Therefore, combined strategy exploiting EGFR-TKI and inhibitors of TET2/NF-κB axis holds therapeutic potential for treating NSCLC patients who suffered from this resistance.

Oncogenic Viruses and the Epigenome: How Viruses Hijack Epigenetic Mechanisms to Drive Cancer.

Globally, viral infections substantially contribute to cancer development. Oncogenic viruses are taxonomically heterogeneous and drive cancers using diverse strategies, including epigenomic dysregulation. Here, we discuss how oncogenic viruses disrupt epigenetic homeostasis to drive cancer and focus on how virally mediated dysregulation of host and viral epigenomes impacts the hallmarks of cancer. To illustrate the relationship between epigenetics and viral life cycles, we describe how epigenetic changes facilitate the human papillomavirus (HPV) life cycle and how changes to this process can spur malignancy. We also highlight the clinical impact of virally mediated epigenetic changes on cancer diagnosis, prognosis, and treatment.

Methyl Donor Micronutrients: A Potential Dietary Epigenetic Target in Systemic Lupus Erythematosus Patients.

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by an aberrant immune response and persistent inflammation. Its pathogenesis remains unknown; however, a complex interaction between environmental, genetic, and epigenetic factors has been suggested to cause disease onset. Several studies have demonstrated that epigenetic alterations, such as DNA hypomethylation, miRNA overexpression, and altered histone acetylation, may contribute to SLE onset and the disease's clinical manifestations. Epigenetic changes, especially methylation patterns, are modifiable and susceptible to environmental factors such as diet. It is well known that methyl donor nutrients, such as folate, methionine, choline, and some B vitamins, play a relevant role in DNA methylation by participating as methyl donors or coenzymes in one-carbon metabolism. Based on this knowledge, this critical literature review aimed to integrate the evidence in animal models and humans regarding the role of nutrients in epigenetic homeostasis and their impact on immune system regulation to suggest a potential epigenetic diet that could serve as adjuvant therapy in SLE.

High nuclear expression of DNMT1 in correlation with inactivation of TET1 portray worst prognosis among the cervical carcinoma patients: clinical implications.

In this study, we aimed to understand the interplay of the epigenetic modifier genes DNMT1 and TET1 along with HPV infection in the cervical epithelium and how it changes during tumorigenesis. For this purpose, initially the bioinformatical analysis (methylation and expression profile) of DNMT1 and TET1 was analyzed in the TCGA dataset. Next genetic (deletion) and epigenetic profiling (promoter methylation) of DNMT1 and TET1 were done in our sample pool and also validated in CACX cell lines as well. The results were further correlated with different clinicopathological parameters. Our data revealed that HPV infection in basal/parabasal layers of cervical epithelium actually disrupts the epigenetic homeostasis of DNMT1 and TET1 proteins which ultimately leads to the high expression of DNMT1 along with further reduction in TET1 protein during the development of carcinoma. Further, in-depth look into the results revealed that comparatively low methylation frequency of DNMT1 coupled with high promoter methylation and deletion frequency [22-46%] of TET1 were the plausible reasons of their antagonistic expression profile during the progression of the disease. Interestingly, the prevalence of DNMT1 [9.1%] and TET1 promoter methylation [22.7%] found in both the plasma DNA of the respective CACX patients implicated its diagnostic importance in this study. Lastly, molecular alteration of TET1 alone or in combination with DNMT1 showed the worst overall survival among the patients. Hence, it may be concluded that an inverse molecular profile of DNMT1 and TET1 genes seen in the proliferative basal-parabasal layers of the cervical epithelium was aggravated during the development of CACX along with genetic and epigenetic changes due to HPV infection.

Aberrant epigenetic regulation of estrogen and progesterone signaling at the level of endometrial/endometriotic tissue in the pathomechanism of endometriosis.

Endometriosis is a term referring to a condition whereby the endometrial tissue is found outside the uterine cavity. This progressive and debilitating condition affects up to 15% of women of reproductive age. Due to the fact that endometriosis cells may express estrogen receptors (ERα, Erβ, GPER) and progesterone (P4) receptors (PR-A, PR-B), their growth, cyclic proliferation, and breakdown are similar to the processes occurring in the endometrium. The underlying etiology and pathogenesis of endometriosis are still not fully explained. The retrograde transport of viable menstrual endometrial cells with the retained ability to attach within the pelvic cavity, proliferate, differentiate and invade into the surrounding tissue explains the most widely accepted implantation theory. Endometrial stromal cells (EnSCs) with clonogenic potential constitute the most abundant population of cells within endometrium that resemble the properties of mesenchymal stem cells (MSCs). Accordingly, formation of the endometriotic foci in endometriosis may be due to a kind of EnSCs dysfunction. Increasing evidence indicates the underestimated role of epigenetic mechanisms in the pathogenesis of endometriosis. Hormone-mediated epigenetic modifications of the genome in EnSCs or even MSCs were attributed an important role in the etiopathogenesis of endometriosis. The roles of excess estrogen exposure and P4 resistance were also found to be crucial in the development of epigenetic homeostasis failure. Therefore, the aim of this review was to consolidate the current knowledge regarding the epigenetic background of EnSCs and MSCs and the changed properties due to estrogen/P4 imbalances in the context of the etiopathogenesis of endometriosis.

Crosstalk between metabolic reprogramming and epigenetics in cancer: updates on mechanisms and therapeutic opportunities

Reversible, spatial, and temporal regulation of metabolic reprogramming and epigenetic homeostasis are prominent hallmarks of carcinogenesis. Cancer cells reprogram their metabolism to meet the high bioenergetic and biosynthetic demands for vigorous proliferation. Epigenetic dysregulation is a common feature of human cancers, which contributes to tumorigenesis and maintenance of the malignant phenotypes by regulating gene expression. The epigenome is sensitive to metabolic changes. Metabolism produces various metabolites that are substrates, cofactors, or inhibitors of epigenetic enzymes. Alterations in metabolic pathways and fluctuations in intermediate metabolites convey information regarding the intracellular metabolic status into the nucleus by modulating the activity of epigenetic enzymes and thus remodeling the epigenetic landscape, inducing transcriptional responses to heterogeneous metabolic requirements. Cancer metabolism is regulated by epigenetic machinery at both transcriptional and post-transcriptional levels. Epigenetic modifiers, chromatin remodelers and non-coding RNAs are integral contributors to the regulatory networks involved in cancer metabolism, facilitating malignant transformation. However, the significance of the close connection between metabolism and epigenetics in the context of cancer has not been fully deciphered. Thus, it will be constructive to summarize and update the emerging new evidence supporting this bidirectional crosstalk and deeply assess how the crosstalk between metabolic reprogramming and epigenetic abnormalities could be exploited to optimize treatment paradigms and establish new therapeutic options. In this review, we summarize the central mechanisms by which epigenetics and metabolism reciprocally modulate each other in cancer and elaborate upon and update the major contributions of the interplays between epigenetic aberrations and metabolic rewiring to cancer initiation and development. Finally, we highlight the potential therapeutic opportunities for hematological malignancies and solid tumors by targeting this epigenetic-metabolic circuit. In summary, we endeavored to depict the current understanding of the coordination between these fundamental abnormalities more comprehensively and provide new perspectives for utilizing metabolic and epigenetic targets for cancer treatment.

Regulation of epigenetic homeostasis in uveal melanoma and retinoblastoma.

Uveal melanoma (UM) and retinoblastoma (RB), which cause blindness and even death, are the most frequently observed primary intraocular malignancies in adults and children, respectively. Epigenetic studies have shown that changes in the epigenome contribute to the rapid progression of both UM and RB following classic genetic changes. The loss of epigenetic homeostasis plays an important role in oncogenesis by disrupting the normal patterns of gene expression. The targetable nature of epigenetic modifications provides a unique opportunity to optimize treatment paradigms and establish new therapeutic options for both UM and RB with these aberrant epigenetic modifications. We aimed to review the research findings regarding relevant epigenetic changes in UM and RB. Herein, we 1) summarize the literature, with an emphasis on epigenetic alterations, including DNA methylation, histone modifications, RNA modifications, noncoding RNAs and an abnormal chromosomal architecture; 2) elaborate on the regulatory role of epigenetic modifications in biological processes during tumorigenesis; and 3) propose promising therapeutic candidates for epigenetic targets and update the list of epigenetic drugs for the treatment of UM and RB. In summary, we endeavour to depict the epigenetic landscape of primary intraocular malignancy tumorigenesis and provide potential epigenetic targets in the treatment of these tumours.

Abstract 1614: Creation of malignant peripheral nerve sheath tumor models deficient for polycomb repressive complex 2 and identification of therapeutic vulnerabilities

Abstract We constructed models of nerve sheath tumors that arise in Neurofibromatosis Type 1 (NF1) patients and conducted therapeutics discovery using synthetic lethal pharmacogenomic screens. Given both plexiform neurofibromas and malignant peripheral nerve sheath tumors (MPNSTs) arise within the Schwann cell lineage, we developed a drug discovery pipeline to identify targeted therapeutics for treating NF1-related neoplasia. Using CRISPR/Cas9, immortalized human Schwann cell lines were created lacking the NF1 gene or NF1 and components of Polycomb Repressive Complex 2 (PRC2), such as SUZ12. ~80% of all MPNST harbor loss of function mutations in PRC2 genes, which is highly suggestive that perturbation of epigenetic homeostasis plays a role in malignant transformation of neurofibromas. Our models mimic the genetics of MPNSTs and allowed us to identify selectively lethal drugs which exploit vulnerabilities specific to these genetic drivers. We describe the identification of these novel drugs, extensive in vivo testing in models of MPNST, and mechanistic studies to explain why they act specifically on MPNSTs. We identified compounds showing selective lethality towards NF1/SUZ12 double mutant cells. These include drugs affecting epigenetic homeostasis, such as HDAC inhibitors (HDACi). Moreover, many of these drugs showed strong synergy when tested in combination with a MEK inhibitor (MEKi) against NF1/SUZ12 deficient human Schwann and MPNST cell lines. We investigated the mechanism of this synergy using genetics, transcriptome, and proteome analysis. Interestingly, these mechanistic studies revealed possible emerging avenues of resistance the cells are utilizing to compensate for and survive therapeutic intervention. For instance, proteomics analysis of PRC2 deficient MPNST cell lines treated with combination therapy of MEKi and HDACi indicates activation of specific survival and stress response pathways. Targeting these pathways in conjunction with the therapeutics we have identified could prevent emergence of resistance and tumor escape. Clinically interesting drug candidates were advanced and tested as single agents and in combination in multiple in vivo models of MPNST (including patient derived and cell line xenografts). The FDA approved drugs vorinostat (HDACi) and selumetinib (MEKi) have shown dramatic efficacy, with combination therapy exhibiting strong synergy in vivo. This includes durable responses in survival studies and the ability to dramatically shrink established tumors, with treated tumors showing markers of apoptosis. The discovery of novel agents effective against several models of MPNST is exciting as there are currently no approved targeted therapies for MPNSTs. Our results implicate targeting of epigenetic homeostasis, in combination with MEKi, as a major vulnerability of MPNSTs deficient for PRC2 activity. Citation Format: Kyle B. Williams, Alex Larsson, Justin Tibbits, Christopher L. Moertel, David A. Largaespada. Creation of malignant peripheral nerve sheath tumor models deficient for polycomb repressive complex 2 and identification of therapeutic vulnerabilities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1614.

Therapeutic Targeting of Cancer: Epigenetic Homeostasis.

A large number of studies have revealed that epigenetics plays an important role in cancer development. However, the currently-developed epigenetic drugs cannot achieve a stable curative effect. Thus, it may be necessary to redefine the role of epigenetics in cancer development. It has been shown that embryonic development and tumor development share significant similarities in terms of biological behavior and molecular expression patterns, and epigenetics may be the link between them. Cell differentiation is likely a manifestation of epigenetic homeostasis at the cellular level. In this article, we introduced the importance of epigenetic homeostasis in cancer development and analyzed the shortcomings of current epigenetic treatment regimens. Understanding the dynamic process of epigenetic homeostasis in organ development can help us characterize cancer according to its differentiation stages, explore new targets for cancer treatment, and improve the clinical prognosis of patients with cancer.

The effects of vitamins and dietary pattern on epigenetic modification of non-communicable diseases.

Background: Non-communicable diseases (NCDs) have received more attention because of high prevalence and mortality rate. Besides genetic and environmental factors, the epigenetic abnormality is also involved in the pathogenesis of NCDs. Methylation of DNA, chromatin remodeling, modification of histone, and long non-coding RNAs are the main components of epigenetic phenomena. Methodology: In this review paper, the mechanistic role of vitamins and dietary patterns on epigenetic modification was discussed. All papers indexed in scientific databases, including PubMed, Scopus, Embase, Google Scholar, and Elsevier were searched during 2000 - 2021 using, vitamins, diet, epigenetic repression, histones, methylation, acetylation, and NCDs as keywords. Results: The components of healthy dietary patterns like Mediterranean and dietary approaches to stop hypertension diets have a beneficial effect on epigenetic hemostasis. Both quality and quantity of dietary components influence epigenetic phenomena. A diet with calorie deficiency in protein content and methyl-donor agents in a long time, with a high level of fat, disrupts epigenetic hemostasis and finally, causes genome instability. Also, soluble and insoluble vitamins have an obvious role in epigenetic modifications. Most vitamins interact directly with methylation, acetylation, and phosphorylation pathways of histone and DNA. However, numerous indirect functions related to the cell cycle stability and genome integrity have been recognized. Conclusion: Considering the crucial role of a healthy diet in epigenetic homeostasis, adherence to a healthy dietary pattern containing enough levels of vitamin and avoiding the western diet seems to be necessary. Having a healthy diet and consuming the recommended dietary level of vitamins can also contribute to epigenetic stability.

P–230 The NAD+ precursor nicotinamide riboside protects against postovulatary aging in vitro

Abstract Study question Can nicotinamide riboside, one of the NAD+ precursor, protect against postovulatary aging in vitro? Summary answer The NAD+ precursor nicotinamide riboside can protect against postovulatary aging in vitro. What is known already Postovulatory aging(POA) has been considered one of the most intractable challenges that limit the successful rate of ART. Multiple cellular and molecular changes have been involved during the process of POA. The NAD+ is a prominent redox cofactor which is indispensable to DNA repair, energy metabolism, autophagy, genomic stability as well as epigenetic homeostasis. Over the last several decades, increasingly studies have eported that NAD+ contents decline with age across multiple tissues and loss of it are implicated in various diseases associated to aging. As one of a precursor of NAD+, nicotinamide riboside play important role in regulating oxidative stress. Study design, size, duration In this study, we take advantage of in vitro aging model to explore the influences of NR administration on the postovulatory aged oocytes in mice. We analyzed the association of NR supplementation with the aging-related deterioration of oocyte quality, such as mitochondrial dysfunction, mislocalization of cortical granules, followed by embryonic development potential and the NAD+/SIRT1 signaling. We used 3582 oocytes totally. Participants/materials, setting, methods CD–1 mice oocytes/ in vitro culture/ UPLC-MS/MS for NAD+ contents measurement ,DCFH-DA staining for ROS detecting, γH2AX staining for DNA damage measurement, BODIPY FL ATP staining for ATP detecting, LCA-FITC staining to assess the distribution and dynamics of cortical granules (CGs), In vitro fertilization, Quantitative real time PCR Main results and the role of chance NR supplementation exerted protective effects on morphological defects of oocytes, and that these protective effects were concentration dependent. We detected a significantly decline in NAD+ levels in aging oocytes, however, NAD+ accumulation was present in aging oocytes after 200μM NR treatment, indicating that NR administration might be a feasible strategy to enhance the quality of aging oocytes. Furthermore, NR indeed elevated the embryonic development potential of POA oocytes after fertilization. Our findings revealed that NR administration effectively ameliorated ROS accumulation in POA oocytes.Then we tried to uncover the effects of NR on the meiotic apparatus in aging oocytes. NR supplementation can partially restores normal spindle assembly and chromosome alignment in postovulatory aging oocytes. Furthermore, we investigated the protective effects of NR on mitochondrial function through following expects: mitochondrial distribution, ATP production and mitochondrial membrane potential. NR treatment could promote mitochondrial function in oocytes during postovulatory aging in vitro. In addition, DNA damage and mislocalized CGs during postovulatory aging might be rescued by the supplementation of NR. Based on these evidences, we identified that NR improved the quality of aging oocytes by NAD+/SIRT1 axis. Limitations, reasons for caution Only in vitro, shown only in mice. Wider implications of the findings: Our work represents a clinically effective pharmacological chemical to improve infertility caused by POA process. Further studies are needed to define the related mechanisms of NR supplementation on oocyte quality as well as reproductive outcomes clinically. Trial registration number N

Deleterious point mutations in T‐cell acute lymphoblastic leukemia: Mechanistic insights into leukemogenesis

T-cell acute lymphoblastic leukemia (T-ALL) is characterized by the leukemogenic transformation of immature T-cells, which accumulate an array of genetic and epigenetic lesions, leading to a sustained proliferation of abnormal T cells. Genetic alterations in the DNA repair genes, protooncogenes, transcription factors, and epigenetic modifiers have been studied in the past decade using next-generation sequencing and high-resolution copy number arrays. While other genomic lesions like chromosomal rearrangements, inversions, insertions, and gene fusions have been well studied at functional level, the mechanism of generation of driver mutations in T-ALL is the subject of current investigation. Novel oncogenic mutations in the TP53, BRCA2, PTEN, IL7R, RAS, NOTCH1, ETV6, BCL11B, WT1, DNMT3A, PRC2, PHF6, USP7, KDM6A and an array of other genes disrupt the genetic and epigenetic homeostasis in T-ALL. In this review, we have summarized the mechanistic role of deleterious driver mutations in T-ALL initiation and progression. We speculate that the formation of non-B DNA structures could be one of the primary reasons for the occurrence of different genomic lesions seen in T-ALL, which warrants further investigation. Understanding the mechanism behind the genesis of oncogenic mutations will pave the way to develop targeted therapies that can improve the overall survival and treatment outcome.

Extensive Placental Methylation Profiling in Normal Pregnancies.

The placental methylation pattern is crucial for the regulation of genes involved in trophoblast invasion and placental development, both key events for fetal growth. We investigated LINE-1 methylation and methylome profiling using a methylation EPIC array and the targeted methylation sequencing of 154 normal, full-term pregnancies, stratified by birth weight percentiles. LINE-1 methylation showed evidence of a more pronounced hypomethylation in small neonates compared with normal and large for gestational age. Genome-wide methylation, performed in two subsets of pregnancies, showed very similar methylation profiles among cord blood samples while placentae from different pregnancies appeared very variable. A unique methylation profile emerged in each placenta, which could represent the sum of adjustments that the placenta made during the pregnancy to preserve the epigenetic homeostasis of the fetus. Investigations into the 1000 most variable sites between cord blood and the placenta showed that promoters and gene bodies that are hypermethylated in the placenta are associated with blood-specific functions, whereas those that are hypomethylated belong mainly to pathways involved in cancer. These features support the functional analogies between a placenta and cancer. Our results, which provide a comprehensive analysis of DNA methylation profiling in the human placenta, suggest that its peculiar dynamicity can be relevant for understanding placental plasticity in response to the environment.

EPCO-27. GLIOMA SINGLE CELL MULTI-OMIC ANALYSES REVEALS REGULATORS OF PLASTICITY AND ADAPTIVE STRESS RESPONSE

Abstract Extensive intra- and intertumoral heterogeneity in glioma contributes to therapeutic resistance and poor patient outcomes. Alterations to DNA methylation (DNAme) modulate epigenetic homeostasis, allowing tumor cells to sample alternative cell states to promote tumorigenesis. However, the epigenetic mechanisms that promote cellular plasticity and regulate cell states are still poorly understood. To characterize the epigenetic mechanisms underlying glioma heterogeneity we profiled 914 single-cell methylomes, 55,284 single-cell transcriptomes, and bulk whole genomes across 11 patient samples spanning initial and recurrent time points and 3 molecular subtypes delineated by IDH mutation status. Local DNAme disorder, defined as epimutation burden, was increased in tumor cells relative to nontumor cells, higher in IDH wild-type than in IDH mutant glioma and was positively associated with copy number alteration (CNA). Epimutation was positively associated with transcriptional variability and enriched at genes involved in cellular differentiation. Epimutation was also increased in the binding sites of transcription factors (TFs) associated with response to extracellular stimuli, suggesting that stochastic DNAme alterations enable cellular plasticity and diverse responses to microenvironmental stressors. Integrative clustering of DNAme and scRNAseq profiles defined stem-like and differentiated-like cell states which exhibited differences in TF activity. Stem-like cells were enriched for differentially methylated binding sites of TFs associated with hypoxia response. scDNAme and scRNAseq-derived copy number profiles were compared with bulk copy number profiles and inferred tumor phylogenies to assess how the timing of CNAs impact epigenetic instability, with results suggesting that early CNA events propagate both genetic and epigenetic heterogeneity. Bulk longitudinal data was used to validate the relationship of epigenetic instability with CNA burden as well as differentially methylated binding sites of cell stress response TFs. Our work suggests that local DNAme disorder promotes cellular plasticity and enables adaptive response to cellular stress such as hypoxia.

UV Radiation and Its Relation to DNA Methylation in Epidermal Cells: A Review.

DNA methylation is the most studied epigenetic mark, and it can be altered by environmental factors. Among these factors, ultraviolet radiation (UV) is little explored within this context. While the relationship between UV radiation and DNA mutations is clear, little is known about the relationship between UV radiation and epimutations. The present study aimed to perform a literature review to determine the influence of artificial or natural (solar) UV radiation on the global and site-specific methylation profile of epidermal cells. A systematic review of the literature was carried out using the databases PubMed, Scopus, Cochrane, and Web of Science. Observational and intervention studies in cultured cells and animal or human models were included. Most studies showed a relationship between UV radiation and changes in the methylation profile, both global and site-specific. Hypermethylation and hypomethylation changes were detected, which varied according to the studied CpG site. In conclusion, UV radiation can alter the DNA methylation profile in epidermal cells derived from the skin. These data can be used as potential biomarkers for environmental exposure and skin diseases, in addition to being targets for treatments. On the other hand, UV radiation (phototherapy) can also be used as a tool to treat skin diseases. Thus, the data suggest that epigenetic homeostasis can be disrupted or restored by exposure to UV radiation according to the applied wavelength.

Abstract 4106: High throughput chemical profiling of chemoresistant triple negative breast cancer

Abstract Triple-negative breast cancer (TNBC) makes up approximately 10-20% of all breast cancers, is characterized by a lack of targeted therapies, and in approximately 40-50% of cases a significant residual cancer burden remains after neoadjuvant chemotherapy, thus presenting a need to identify actionable targets in both treatment naive and resistant settings. To address the unmet treatment needs of chemo-resistant patients, our multi-institutional collaborative of clinical oncologists and basic researchers are assembling a panel of patient-derived xenograft (PDX) models of TNBC as part of an ongoing IRB-approved clinical trial at MD Anderson Cancer Center (ARTEMIS; NCT02276443). PDX models are generated from serial fine needle aspirates of primary TNBC from patients (1) prior to treatment, and in non-responding tumors (2) after four cycles of Adriamycin plus cyclophosphamide (AC), and (3) after a 3-month course of targeted therapy. We first developed a standardized intake procedure that simultaneously optimized cell seeding densities in 384-well plates, tested multiple cell culturing medias, determined baseline phenotypic characteristics, and established robust assay controls. After optimization, we performed high-throughput drug screens with a panel of 634 mechanistically annotated chemical probes that are FDA approved, late stage investigational or mechanistic probes. A bioluminescent reporter quantifying the amount of ATP (CellTiter Glo, Promega) was used as a surrogate readout of cell viability, which is well suited for the majority of PDX cultures that grew as non-adherent multi-cellular aggregates as well as being integrable into larger datasets run in a similar format. Screening assays are performed with 3-4 technical replicates over a three log concentration range (10.0, 1.0, 0.1 uM). The effectiveness of each drug is summarized as an area under the fitted concentration-response curve, simultaneously capturing alterations in potency and efficacy. In total, we have tested 21 PDX models isolated from 14 unique patients using this strategy. From these data, we show that PDX models generated from individual patients have unique pharmacologic profiles with little correspondence to molecularly defined sub-classifications, which may suggest the need for the development of an actionable sub-classification strategy to better stratify patients. Despite the significant heterogeneity across pharmacologic profiles, we identified a consolidated set of drugs targeting specific epigenetic processes and protein homeostasis that have increased activity in a subset of chemo-resistant samples. We are currently collecting orthogonal data including next-generation sequencing and transcriptomics to further elucidate the mechanism of these targets and identify potential biomarkers. Citation Format: Reid Trenton Powell, Abena Redwood, Lei Guo, Shirong Cai, Yizheng Tu, Yan Jiang, Xuan Liu, Xinhui Zhou, Clifford Stephan, Peter Davies, Jeffrey T. Chang, Stacey Moulder, William F. Symmans, Helen Piwnica-Worms. High throughput chemical profiling of chemoresistant triple negative breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4106.