Epigenetic Events Research Articles

17th International Conference on Malignant Lymphoma, Palazzo dei Congressi, Lugano, Switzerland, 13 - 17 June, 2023.

Background: Diffuse large B-cell lymphoma (DLBCL) is a biologically and clinically heterogeneous disease that requires personalized clinical treatment. To assign patients into different risk categories, cytogenetic abnormalities and genetic mutations have been widely applied to the prognostic stratification of DLBCL. Increasing evidence has demonstrated that deregulated epigenetic modifications and long noncoding RNAs (lncRNAs) contribute to the initiation and progression of DLBCL. However, specific lncRNAs that affect epigenetic regulation and their value in predicting prognosis and therapy response remain uncertain. Methods: LncRNA expression profiles and clinical features of DLBCL samples were collected from the GEO database and 188 DLBCL patients in our center was as a validation cohort. A linear regression based on the modified LASSO algorithm was performed to construct a predictive model named lncRNA-regulating epigenetic event signature (ELncSig). Results: A list of 2,025 epigenetic regulatory genes was generated from GeneCards and 9 lncRNAs (PRKCQ-AS1, C22orf34, HCP5, AC007389.3, APTR, SNHG19, ELFN1-AS1, LINC00487, and LINC00877) were tested and validated to establish the ELncSig which could distinguish different survival outcomes. Functional characterization of ELncSig showed that it could be an indicator of the immune microenvironment and is correlated with distinctive mutational characteristics. Univariate and multivariate analyses showed that ELncSig was independent of traditional prognostic factors. Conclusions: We constructed a lncRNA signature based on epigenetic-related genes to predict the prognosis of DLBCL and also proved that this new signature could affect other coding proteins in addition to epigenetic genes. Importantly, ELncSig might be associated with immune infiltration levels and even the efficacy of tumor immunotherapy. Keywords: aggressive B-cell non-Hodgkin lymphoma, diagnostic and prognostic biomarkers, risk models No conflicts of interests pertinent to the abstract.

Mitochondria in Human Fertility and Infertility.

In human spermatozoa and oocytes (and their surrounding granulosa cells), mitochondria carry out important functions relating to human fertility and infertility. Sperm mitochondria are not transmitted to the future embryo, but are closely related to the generation of energy needed for sperm movement, capacitation, and acrosome reactions, as well as for sperm-oocyte fusion. On the other hand, oocyte mitochondria produce energy required for oocyte meiotic division and their abnormalities can thus cause oocyte and embryo aneuploidy. In addition, they play a role in oocyte calcium metabolism and in essential epigenetic events during the oocyte-to-embryo transition. They are transmitted to the future embryos and may thus cause hereditary diseases in the offspring. Due to the long life span of the female germ cells, the accumulation of mitochondrial DNA abnormalities often causes ovarian aging. Mitochondrial substitution therapy is the only way of dealing with these issues nowadays. New therapies based on mitochondrial DNA editing are under investigation.

A novel cell-based screen identifies chemical entities that reverse the immune-escape phenotype of metastatic tumours.

Genetic and epigenetic events have been implicated in the downregulation of the cellular antigen processing and presentation machinery (APM), which in turn, has been associated with cancer evasion of the immune system. When these essential components are lacking, cancers develop the ability to subvert host immune surveillance allowing cancer cells to become invisible to the immune system and, in turn, promote cancer metastasis. Here we describe and validate the first high-throughput cell-based screening assay to identify chemical extracts and unique chemical entities that reverse the downregulation of APM components in cell lines derived from metastatic tumours. Through the screening of a library of 480 marine invertebrate extracts followed by bioassay-guided fractionation, curcuphenol, a common sesquiterpene phenol derived from turmeric, was identified as the active compound of one of the extracts. We demonstrate that curcuphenol induces the expression of the APM components, TAP-1 and MHC-I molecules, in cell lines derived from both metastatic prostate and lung carcinomas. Turmeric and curcumins that contain curcuphenol have long been utilized not only as a spice in the preparation of food, but also in traditional medicines for treating cancers. The remarkable discovery that a common component of spices can increase the expression of APM components in metastatic tumour cells and, therefore reverse immune-escape mechanisms, provides a rationale for the development of foods and advanced nutraceuticals as therapeutic candidates for harnessing the power of the immune system to recognize and destroy metastatic cancers.

Functional characterization of age-dependent p16 epimutation reveals biological drivers and therapeutic targets for colorectal cancer

BackgroundMethylation of the p16 promoter resulting in epigenetic gene silencing—known as p16 epimutation—is frequently found in human colorectal cancer and is also common in normal-appearing colonic mucosa of aging individuals. Thus, to improve clinical care of colorectal cancer (CRC) patients, we explored the role of age-related p16 epimutation in intestinal tumorigenesis.MethodsWe established a mouse model that replicates two common genetic and epigenetic events observed in human CRCs: Apc mutation and p16 epimutation. We conducted long-term survival and histological analysis of tumor development and progression. Colonic epithelial cells and tumors were collected from mice and analyzed by RNA sequencing (RNA-seq), quantitative PCR, and flow cytometry. We performed single-cell RNA sequencing (scRNA-seq) to characterize tumor-infiltrating immune cells throughout tumor progression. We tested whether anti-PD-L1 immunotherapy affects overall survival of tumor-bearing mice and whether inhibition of both epigenetic regulation and immune checkpoint is more efficacious.ResultsMice carrying combined Apc mutation and p16 epimutation had significantly shortened survival and increased tumor growth compared to those with Apc mutation only. Intriguingly, colon tumors with p16 epimutation exhibited an activated interferon pathway, increased expression of programmed death-ligand 1 (Pdl1), and enhanced infiltration of immune cells. scRNA-seq further revealed the presence of Foxp3+ Tregs and γδT17 cells, which contribute to an immunosuppressive tumor microenvironment (TME). Furthermore, we showed that a combined therapy using an inhibitor of DNA methylation and a PD-L1 immune checkpoint inhibitor is more effective for improving survival in tumor-bearing mice than blockade of either pathway alone.ConclusionsOur study demonstrated that age-dependent p16 epimutation creates a permissive microenvironment for malignant transformation of polyps to colon cancer. Our findings provide a mechanistic rationale for future targeted therapy in patients with p16 epimutation.

Abstract #1406834: Study of Methylation Status and Expression of Vitamin D Receptor Gene in Polycystic Ovarian Syndrome Patients of Kashmiri Population

Downregulation of Vitamin D Receptor (VDR) gene expression resulting from epigenetics events, like methylation in promoter of VDR gene play an important role in pathogenesis, clinical and bio-chemical parameters of polycystic ovary syndrome, but the results from different studies are inconsistent. We aim to determine the methylation status of promoter of VDR gene and VDR gene expression analysis in addition to the co-relation between promoter methylation and expression profile of VDR genes in PCOS patients in Kashmiri Population. This study was a case control, comprising of 100 women with PCOS and 100 normal healthy control women aged between 15-45 years. PCOS patients were selected after proper diagnosis using Rotterdam criteria (2004) and obtaining written consent. DNA extraction was carried out by the non-enzymatic Salting out method and extracted DNA samples were subjected to bisulphite treatment using EZ DNA Methylation-Gold Kit (Zymo Research, Irvine, California) to produces a chemical conversion of unmethylated cytosine to uracil, which is detected as a thymine after PCR. Methylation specific PCR (MS-PCR) was performed following the bisulfite conversion, using pre-designed sets of methylated and unmethylated primers to confirm the prominence of methylation in the promoter VDR region and to specially acknowledge methylated and unmethylated VDR fragments. The VDR expression analysis was carried out following RNA extraction from the blood sample and processed ahead for cDNA synthesis using reverse transcriptase reaction and then relative expression analysis by RT-PCR using SYBR Green fluorescent dye. We found that vitamin D level among test group was significantly lower compared to controls (13.95±9.60 VS. 16.63±7.46; p=0.026). Methylation was significantly higher in test group compared to controls (84% vs 62%; p<0.0001) with OR of 3.21. Furthermore, there exists a significant difference between the average cyclic threshold (CT) value of VDR gene among test and control groups (24.64 vs. 26.14;p <0.0001). The average CT value corresponding to GAPDH (glyceraldehyde phosphate dehydrogenase) is 29.76 in test group and 32.11 in control group. The expression of VDR gene was 0.55 times down regulated in test group in comparison to controls. In the present study, we have comprehensively shown that VDR hypermethylation is significantly associated with reduced vitamin D level and decreased expression level of VDR gene among PCOS women. Increased VDR methylation in response to decreased vitamin D levels could be involved in the impairment of VDR mediated effects resulting in the peculiar clinical and bio-chemical derangements in patients of polycystic ovary syndrome. However, the enhanced ligand availability results in decreased receptor expression to maintain homeostasis of vitamin D signalling. Hence, this, inverse association could have a significant role in the altered gene expression of VDR and thereby effecting the cell response in PCOS patients.

Prominent epigenetic and transcriptomic changes in CD4+ and CD8+ T cells during and after pregnancy in women with multiple sclerosis and controls

BackgroundMultiple sclerosis (MS) is a neuroinflammatory disease in which pregnancy leads to a temporary amelioration in disease activity as indicated by the profound decrease in relapses rate during the 3rd trimester of pregnancy. CD4+ and CD8+ T cells are implicated in MS pathogenesis as being key regulators of inflammation and brain lesion formation. Although Tcells are prime candidates for the pregnancy-associated improvement of MS, the precise mechanisms are yet unclear, and in particular, a deep characterization of the epigenetic and transcriptomic events that occur in peripheral T cells during pregnancy in MS is lacking.MethodsWomen with MS and healthy controls were longitudinally sampled before, during (1st, 2nd and 3rd trimesters) and after pregnancy. DNA methylation array and RNA sequencing were performed on paired CD4+ and CD8+ T cells samples. Differential analysis and network-based approaches were used to analyze the global dynamics of epigenetic and transcriptomic changes.ResultsBoth DNA methylation and RNA sequencing revealed a prominent regulation, mostly peaking in the 3rd trimester and reversing post-partum, thus mirroring the clinical course with improvement followed by a worsening in disease activity. This rebound pattern was found to represent a general adaptation of the maternal immune system, with only minor differences between MS and controls. By using a network-based approach, we highlighted several genes at the core of this pregnancy-induced regulation, which were found to be enriched for genes and pathways previously reported to be involved in MS. Moreover, these pathways were enriched for in vitro stimulated genes and pregnancy hormones targets.ConclusionThis study represents, to our knowledge, the first in-depth investigation of the methylation and expression changes in peripheral CD4+ and CD8+ T cells during pregnancy in MS. Our findings indicate that pregnancy induces profound changes in peripheral T cells, in both MS and healthy controls, which are associated with the modulation of inflammation and MS activity.

Role of histone deacetylase 3 (HDAC3) in adipose tissue metabolism and immunophenotype: Selected Abstract - SITeCS Congress 2022

Introduction: Obesity is associated with comorbidities such as cardiovascular disease and type 2 diabetes. HDAC3 regulates adipose tissue physiology (WAT), and its genetic inactivation causes metabolic reprogramming of white adipocytes toward browning. The aim of this work is to evaluate the effect of HDAC3 silencing at different stages of differentiation and investigate the influence of adipocyte metabolism on the immunophenotype of WAT. Materials and Methods: Following HDAC3 silencing in mesenchymal stem cells and mature adipocytes, adipocyte function, RNA, DNA and protein levels, and proliferation at the end of differentiation were analyzed. Visceral WAT immunophenotype (vWAT) of Hdac3 KO mice in WAT (Hdac3fatKO) and controls (FL) was performed by FACS. Results: Silencing HDAC3 in precursors amplifies the expression of genes and proteins that regulate differentiation, oxidative metabolism, browning and mitochondrial activity. Following silencing, we found increased 1)phosphorylation of AKT (1.64 fold change, P&lt;0.0001), indicative of increased insulin signaling, and 2)proliferation, characteristic of the early phase of differentiation. Mitochondrial content was unchanged, but increased mitochondrial activity was observed in terms of maximal respiration (1.42 fold change, P=0.0151) and uncoupling of the electron transport chain (+11.6%, P&lt;0.0001). No difference was observed following HDAC3 silencing in mature adipocytes. We hypothesized that the enhancement of oxidative metabolism may cause cellular damage or senescence and, consequently, the immunophenotype of vWAT might be affected by HDAC3 ablation. Analysis reveals an increase of macrophages (2.48 fold change, P=0.0311) in the vWAT of Hdac3fatKO mice polarizing toward the M2 population. Coculture of adipocytes with macrophages from bone marrow indicates that HDAC3 silencing in adipocytes stimulates macrophage activation. Conclusions: HDAC3 is a key factor in the WAT phenotype, and its inactivation triggers mechanisms that support browning. Early epigenetic events mediated by HDAC3 silencing are crucial in directing adipocyte precursors toward the oxidative phenotype. Finally, results obtained from ex vivo and in vitro studies suggest that specific factors produced by KO adipocytes may be involved in determining the observed immunophenotype. [FONDAZIONE CARIPLO 2015-0641]

Early Life Exposure to C16-Ceramide Improves Learning and Short-Term Memory Behavior during Adulthood in Mice.

Ceramides, structural components of the cell, are known to play a range of roles in glucose metabolism and apoptosis. C16-ceramide, an abundant molecular species of endogenous ceramide, has not had its influence on learning and memory explored. We administered C16-ceramide to mice immediately after weaning and examined the learning and memory behavior of these mice during adulthood. Mice given C16-ceramide early in life showed improved adult learning/short-term memory behavior without affecting their glucose metabolism. Looking for a plausible mechanism for this, we found that calcium influx, CaMKII/CREB, and the Erk-relevant signaling transduction are increased after C16-ceramide stimulation in primary neurons in vitro. Possible downstream epigenetic molecular events, such as H3K4 methylation and Egr-1 abundance, were also found to be upregulated. Utilizing J20 mice, an Alzheimer disease mice model in which mice were injected after weaning with C16-ceramide, we found that these mice also show improved learning and short-term memory behavior when assessed by the Morris water maze test. Taken together, giving C16-ceramide early in life would seem to benefit learning and short-term memory behavior during adulthood.

Abstract 6021: Genetic and epigenetic features OF KMT2D, CREBBP, ATM, TSC2 and GNAS in uterine leyomiosarcomas

Abstract Background: Uterine leiomyosarcoma (LMS) comprises 60% to 70% of the uterine sarcoma (US). These tumors present high rates of recurrence and metastasis, and the patients’ survival rates are very low (20% in five years). It is known that somatic mutations and modifications in the DNA methylation levels are associated with several types of neoplasms. Previously, our group identified missense and loss of function mutations in KMT2D, CREBBP, ATM, TSC2, and GNAS, using a genetic screening method in LMS samples. Based on these results, we decided to investigate whether a genetic and epigenetic crosstalk could help to understand the biological processes involved in the LMS origin, development, and pathogenesis. The present study aimed to evaluate the KMT2D, CREBBP, ATM, TSC2 and GNAS gene expression profile in LMS cell line, and their methylation levels in formalin-fixed and paraffin-embedded (FFPE) patients´ samples. Methods: We selected 15 LMS - FFPE samples obtained via surgical procedures performed between 2012 and 2019 at the Instituto do Cancer do Estado de São Paulo (ICESP). Genomic DNA was extracted using the QIAamp DNA FFPE Tissue Kit and we evaluated methylation levels using the Illumina Infinium Methylation EPIC BeadChip system 850k, including samples treated with bisulfite-treated DNA. Leiomyoma (LM) (THESCs CRL-4003) and LMS (SK-UT-1) cell lines were cultivated for gene expression analysis. Cells were growth for 24, 48, 72, and 96 hours, and the total RNA was extracted by TRIzol. High-Capacity cDNA Reverse Transcription Kit was used for cDNA synthesis and for gene expression evaluation, real-time PCR reactions were performed using TaqMan Universal PCR Master Mix and inventoried TaqMan probes. Results: KMT2D and TSC2 showed increased expression in LMS compared to LM (cut-off ≤ -2 and ≥ 2 for down and upregulation, respectively), with higher expression in 24 hours of KMT2D [Fold Regulation (FR): 2.77] and TSC2 (FR: 2.58). In contrast, CREBBP was downregulated, with lowest expression in 72 hours (FR: -6.57). ATM was upregulated at 24, 48 with higher expression in 72 hours (FR: 2.32), and GNAS was downregulated, with lowest expression in 96 hours (FR: -4.31). Moreover, the methylation analyzes showed an expressive hypomethylation in LMS samples, compared to myometrium, for KMT2D (β value: 0.45; p= 0.004); CREBBP (β value: 0.51 p &amp;lt; 0.0001); ATM (β value: 0.47 p &amp;lt; 0.0001); TSC2 (β value: 0.53 p &amp;lt; 0.0001) and GNAS (β value: 0.41 p &amp;lt; 0.0001). In conclusion, our study showed that potentially pathogenic mutations may be associated with an aberrant hypomethylation profile, as well as increased expression of KMT2D, TSC2, ATM and loss expression of CREBBP and GNAS in LMS. The interconnection of these genetic and epigenetic events is essential for understanding the complex biology of these tumors, in addition to enabling identification of biomarkers. Citation Format: Laura G. dos Anjos, Daniela Bizinelli, Katia C. Carvalho. Genetic and epigenetic features OF KMT2D, CREBBP, ATM, TSC2 and GNAS in uterine leyomiosarcomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6021.

Abstract 6007: p16 epimutation modulates the tumor microenvironment and promotes malignancy of Apc-mutant colon cancer

Abstract Methylation of the p16 promoter resulting in mitotically stable gene silencing—known as p16 epimutation—is the most common epigenetic event in human colorectal cancer (CRC) and is also common in normal-appearing colonic mucosa of aging individuals. However, the functional role of p16 epimutation in CRC carcinogenesis is not completely understood. Here, using a mouse model of engineered p16 promoter hypermethylation, we show that p16 epimutation cooperates with mutant Apc to promote colon cancer development. Mice carrying combined Apc mutation and p16 epimutation display significantly shortened survival and increased tumor growth compared to those with Apc mutation only. Moreover, colon tumors from these animals express increased levels of programmed death-ligand 1 (Pdl1). Single-cell RNA sequencing (scRNA-seq) analysis further revealed the presence of dysfunctional cytotoxic CD8+ T cells together with Foxp3+ Tregs and γδT17 cells, which promote an immunosuppressive tumor microenvironment (TME). Lastly, we show that a combined therapy using an inhibitor of DNA methylation and a PD-L1 immune checkpoint inhibitor is more effective for improving survival in tumor-bearing mice than anti-PD-L1 treatment alone. These findings provide a mechanistic rationale for targeted epigenetic and immunotherapy in CRC patients, with the potential to improve colon cancer treatment. Citation Format: Li Yang, Xiaomin Chen, Christy Lee, Jiejun Shi, Emily B. Lawrence, Lanjing Zhang, Yumei Li, Nan Gao, Sung Yun Jung, Chad J. Creighton, Jingyi Jessica Li, Ya Cui, Sumimasa Arimura, Wei Li, Lanlan Shen. p16 epimutation modulates the tumor microenvironment and promotes malignancy of Apc-mutant colon cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6007.

The reduced expression of trimethylation of histone H3 at lysine 27 in primary cutaneous mucinous carcinoma

Primary cutaneous mucinous carcinoma (PCMC) is rare, and its carcinogenesis is unclear. Trimethylation of histone H3 at lysine 27 (H3K27me3) is a key regulator in chromatin remodeling-controlled transcription. Focusing on the epigenetic mechanism, we aimed to investigate the expression of H3K27me3 in PCMC by immunohistochemistry. A retrospective cohort of PCMC patients from a tertiary hospital in Taiwan was enrolled to evaluate the clinicopathologic features, treatment outcome, and protein expression. Immunohistochemistry for H3K27me3 was performed on all PCMCs and a comparison group of colonic mucinous adenocarcinoma and pure mucinous carcinoma of the breast. The percentage of H3K27me3-negative tumor cells was calculated and analyzed. Three patients with PCMC were recruited. All PCMCs were solitary and slow growing, arising from the head-and-neck region. All PCMCs had tumor excision without local recurrence or metastasis. The loss of H3K27me3 expression was significant in PCMCs (mean ± standard deviation [SD]: 21.0% ± 6.6%) compared to other mucinous carcinomas (mean ± SD: 3.8% ± 1.7%) (P = 0.019). In conclusion, we report a reduction in H3K27me3 expression in PCMC. In contrast, H3K27me3 expression is retained or mildly reduced in other mucinous carcinomas. This is the first study to indicate a possible role of epigenetic events in the pathobiology of PCMC.

PHF6 recruits BPTF to promote HIF-dependent pathway and progression in YAP-high breast cancer

BackgroundAberrant epigenetic remodeling events contribute to progression and metastasis of breast cancer (Bca). The specific mechanims that epigenetic factors rely on to mediate tumor aggressiveness remain unclear. We aimed to elucidate the roles of epigenetic protein PHF6 in breast tumorigenesis.MethodsPublished datasets and tissue samples with PHF6 staining were used to investigate the clinical relevance of PHF6 in Bca. CCK-8, clony formation assays were used to assess cell growth capacity. Cell migration and invasion abilities were measured by Transwell assay. The gene mRNA and protein levels were measured by quantitative real-time PCR and western blot. Chromatin immunoprecipitation (ChIP)-qPCR assays were used to investigate transcriptional relationships among genes. The Co-immunoprecipitation (Co-IP) assay was used to validate interactions between proteins. The CRISPR/Cas9 editing technology was used to construct double HIF knockout (HIF-DKO) cells. The subcutaneous xenograft model and orthotopic implantation tumor model were used to asess in vivo tumor growth.ResultsIn this study, we utilized MTT assay to screen that PHF6 is required for Bca growth. PHF6 promotes Bca proliferation and migration. By analyzing The Cancer Genome Atlas breast cancer (TCGA-Bca) cohort, we found that PHF6 was significantly higher in tumor versus normal tissues. Mechanistically, PHF6 physically interacts with HIF-1α and HIF-2α to potentiate HIF-driven transcriptional events to initiate breast tumorigenesis. HIF-DKO abolished PHF6-mediated breast tumor growth, and PHF6 deficiency in turn impaired HIF transcriptional effects. Besides, hypoxia could also rely on YAP activation, but not HIF, to sustain PHF6 expressions in Bca cells. In addition, PHF6 recuits BPTF to mediate epigenetic remodeling to augment HIF transcriptional activity. Targeting PHF6 or BPTF inhibitor (AU1) is effective in mice models. Lastly, PHF6 correlated with HIF target gene expression in human breast tumors, which is an independent prognostic regulator.ConclusionsCollectively, this study identified PHF6 as a prognostic epigenetic regulator for Bca, functioning as a HIF coactivator. The fundamental mechanisms underlying YAP/PHF6/HIF axis in breast tumors endowed novel epigenegtic targets for Bca treatment.

Pre-birth acquisition of personhood: Incremental accrual of attributes as the framework for individualization by serial and concurrently acting developmental factors.

Discrete events and processes influence development of individual humans. Attribution of personhood to any individual human being cannot be disconnected from the underlying biological events and processes of early human development. Nonetheless, the philosophical, sociological and legal components that are integral to the meaning of the term as commonly used cannot be deduced from biology alone. The challenge for biomedical scientists to inform discussion in this arena then rests on profiling the key biological events and processes that must be assessed when considering how one might objectively reason about the task of superimposing the concept of personhood onto the developing biological entity of a potential human being. Endogenous genetic and epigenetic events and exogenous developmental milieu processes diversify developmental trajectories of potential individual humans prior to livebirth. First, fertilization and epigenetic resetting of each individual's organismic clock to time zero (t = 0) at the gastrulation/primitive streak stage (day 15 of embryogenesis), are two discrete unseen biological events that impact a potential individual human's attributes. Second, those two discrete unseen biological events are immersed in the continuous developmental process spanning pre-fertilization and gestation, further driving individualization of diverse attributes of each future human before the third discrete and blatant biological event of parturition and livebirth. Exposures of the gravida to multiple diverse exogenous exposures means that morphogenesis and physiogenesis of every embryo/fetus has individualized attributes for its future human lifespan. Our proposed framework based on the biological discrete events and processes spanning pre-fertilization and prenatal development, implies that personhood should be incrementally attributed, and societal protections should be graduated and applied progressively across the pre-birth timespan.

Critical role of epigenetic modification in the pathogenesis of atopic dermatitis.

Atopic dermatitis is a chronic inflammatory skin disease characterised by recurrent eczema-like lesions and severe pruritus, along with drying and decrustation of skin. Current research relates the pathogenesis of atopic dermatitis mainly to genetic susceptibility, abnormal skin barrier function, immune disorders, Staphylococcus aureus colonisation, microbiological dysfunction and vitamin D insufficiency. Epigenetic modifications are distinct genetic phenotypes resulting from environment-driven changes in chromosome functions in the absence of nuclear DNA sequence variation. Classic epigenetic events include DNA methylation, histone protein modifications and non-coding RNA regulation. Increasing evidence has indicated that epigenetic events are involved in the pathogenesis of atopic dermatitis by their effects on multiple signalling pathways which in turn influence the above factors. This review primarily analyses the function of epigenetic regulation in the pathogenesis of atopic dermatitis. In addition, it tries to make recommendations for personalised epigenetic treatment strategies for atopic dermatitis in the future.

A genomic enhancer signature associates with hepatocellular carcinoma prognosis

Background & AimsLifestyle and environmental-related exposures are important risk factors for hepatocellular carcinoma (HCC), suggesting that epigenetic dysregulation significantly underpins HCC. We profiled 30 surgically resected tumours and the matched adjacent normal tissues to understand the aberrant epigenetic events associated with HCC. MethodsWe identified tumour differential enhancers and the associated genes by analysing H3K27 acetylation (H3K27ac) chromatin immunoprecipitation sequencing (ChIP-seq) and Hi-C/HiChIP data from the resected tumour samples of 30 patients with early-stage HCC. This epigenome dataset was analysed with previously reported genome and transcriptome data of the overlapping group of patients from the same cohort. We performed patient-specific differential expression testing using multiregion sequencing data to identify genes that undergo both enhancer and gene expression changes. Based on the genes selected, we identified two patient groups and performed a recurrence-free survival analysis. ResultsWe observed large-scale changes in the enhancer distribution between HCC tumours and the adjacent normal samples. Many of the gain-in-tumour enhancers showed corresponding upregulation of the associated genes and vice versa, but much of the enhancer and gene expression changes were patient-specific. A subset of the upregulated genes was activated in a subgroup of patients’ tumours. Recurrence-free survival analysis revealed that the patients with a more robust upregulation of those genes showed a worse prognosis. ConclusionsWe report the genomic enhancer signature associated with differential prognosis in HCC. Findings that cohere with oncofoetal reprogramming in HCC were underpinned by genome-wide enhancer rewiring. Our results present the epigenetic changes in HCC that offer the rational selection of epigenetic-driven gene targets for therapeutic intervention or disease prognostication in HCC. Impact and ImplicationsLifestyle and environmental-related exposures are the important risk factors of hepatocellular carcinoma (HCC), suggesting that tumour-associated epigenetic dysregulations may significantly underpin HCC. We profiled tumour tissues and their matched normal from 30 patients with early-stage HCC to study the dysregulated epigenetic changes associated with HCC. By also analysing the patients’ RNA-seq and clinical data, we found the signature genes – with epigenetic and transcriptomic dysregulation – associated with worse prognosis. Our findings suggest that systemic approaches are needed to consider the surrounding cellular environmental and epigenetic changes in HCC tumours.

Altered expression of DNA methyltransferases and methylation status of the TLR4 and TNF-α promoters in COVID-19

Epigenetic modifications play a significant role in the host's immune response to viral infection. Two epigenetic events, DNA methylation and histone acetylation, are crucial for modifying the chromatin architecture and the location of regulatory elements such as promoters and enhancers. In this case-control study, we evaluated the expression of genes involved in epigenetic machinery (DNMT1, DNMT3A, DNMT3B, HDAC2, and HDAC3) and the degree of methylation of promoters of immune response genes (IFITM1/2/3, TLR3/4, TNF-α, NF-κB, and MYD88) as well as global methylation (LINE-1 and global 5–mC) in blood samples from 120 COVID-19 patients (30 mild, 30 moderate, 30 severe, and 30 critical) and 30 healthy subjects without COVID-19. In contrast to previous reports, DNMT3A and DNMT3B expression was found to be significantly downregulated in COVID-19 cases, whereas DNMT1, HDAC2, and HDAC3 expression did not change. DNMT1 and DNMT3A were negatively correlated with COVID-19 severity. Critically ill patients had lower HDAC3 expression levels. TLR4 and TNF-α had increased promoter methylation, whereas IFITM1/2/3, TLR3, NF-κB, MYD88, and LINE-1 did not differ between cases and controls. Methylation of the TNF-α promoter increased as disease severity increased. Significantly less methylation of the TLR3 promoter was observed in patients with a positive outcome (recovery). We also found a correlation between the expression of DNMT3B and the methylation level of the TLR4 promoter. In milder cases, the global 5–mC levels were lower than that in more severe cases. Our findings suggest the exclusion of DNMTs inhibitors previously recommended for COVID-19 treatment and the need for additional research in this area.

Kaposi's Sarcoma-Associated Herpesvirus Immediate Early Proteins Trigger FOXQ1 Expression in Oral Epithelial Cells, Engaging in a Novel Lytic Cycle-Sustaining Positive Feedback Loop.

Kaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic gammaherpesvirus that can replicate in oral epithelial cells to promote viral transmission via saliva. To identify novel regulators of KSHV oral infection, we performed a transcriptome analysis of KSHV-infected primary human gingival epithelial (HGEP) cells, which identified the gene coding for the host transcription factor FOXQ1 as the top induced host gene. FOXQ1 is nearly undetectable in uninfected HGEP and telomerase-immortalized gingival keratinocytes (TIGK) cells but is highly expressed within hours of KSHV infection. We found that while the FOXQ1 promoter lacks activating histone acetylation marks in uninfected oral epithelial cells, these marks accumulate in the FOXQ1 promoter in infected cells, revealing a rapid epigenetic reprogramming event. To evaluate FOXQ1 function, we depleted FOXQ1 in KSHV-infected TIGK cells, which resulted in reduced accumulation of KSHV lytic proteins and viral DNA over the course of 4 days of infection, uncovering a novel lytic cycle-sustaining role of FOXQ1. A screen of KSHV lytic proteins demonstrated that the immediate early proteins ORF45 and replication and transcription activator (RTA) were both sufficient for FOXQ1 induction in oral epithelial cells, indicating active involvement of incoming and rapidly expressed factors in altering host gene expression. ORF45 is known to sustain extracellular signal-regulated kinase (ERK) p90 ribosomal s6 kinase (RSK) pathway activity to promote lytic infection. We found that an ORF45 mutant lacking RSK activation function failed to induce FOXQ1 in TIGK cells, revealing that ORF45 uses a shared mechanism to rapidly induce both host and viral genes to sustain lytic infection in oral epithelial cells. IMPORTANCE The oral cavity is a primary site of initial contact and entry for many viruses. Viral replication in the oral epithelium promotes viral shedding in saliva, allowing interpersonal transmission, as well as spread to other cell types, where chronic infection can be established. Understanding the regulation of KSHV infection in the oral epithelium would allow for the design of universal strategies to target the first stage of viral infection, thereby halting systemic viral pathogenesis. Overall, we uncover a novel positive feedback loop in which immediate early KSHV factors drive rapid host reprogramming of oral epithelial cells to sustain the lytic cycle in the oral cavity.

Human health risk assessment of 6:2 Cl-PFESA through quantitative in vitro to in vivo extrapolation by integrating cell-based assays, an epigenetic key event, and physiologically based pharmacokinetic modeling

Human health risk assessment of chemicals is essential but often relies on time-consuming and animal and labor-extensive procedures. Here, we develop a population-based, quantitative in vitro to in vivo extrapolation (QIVIVE) approach which depended on cellular effects monitored by in vitro assays, considered chemical internal concentration determined by LC-MS/MS, extrapolated into in vivo target tissue concentration through physiologically based pharmacokinetic (PBPK) modelling, and assessed populational health risk using in silico modelling. By applying this QIVIVE approach to 6:2 chlorinated polyfluorinated ether sulfonate (6:2 Cl-PFESA), as a representative of the emerging pollutants, we find that 6:2 Cl-PFESA disturbed lipid homeostasis in HepG2 cells through enhancement of lipid accumulation and fatty acid β-oxidation, during which miR-93-5p served as a key event towards toxicity and thus, could serve as an efficient toxicity marker for risk assessment; further, the disruption potency of lipid homeostasis of 6:2 Cl-PFESA for the most of studied populations in China might be of moderate concern. Together, our approach improved the reliability of QIVIVE during human health risk assessment, which can readily be used for other chemicals.

Hydroxyphenylpyruvate Dioxygenase Is a Metabolic Immune Checkpoint for UTX-deficient Colorectal Cancer

Aberrant epigenetic events mediated by histone methyltransferases and demethylases contribute to malignant progression of colorectal cancer (CRC). However, the role of the histone demethylase ubiquitously transcribed tetratricopeptide repeat on chromosome X (UTX) in CRC remains poorly understood. UTX conditional knockout mice and UTX-silenced MC38 cells were used to investigate UTX function in tumorigenesis and development of CRC. We performed time of flight mass cytometry to clarify the functional role of UTX in remodeling immune microenvironment of CRC. To investigate metabolic interaction between myeloid-derived suppressor cells (MDSCs) and CRC, we analyzed metabolomics data to identify metabolites secreted by UTX-deficient cancer cells and taken up by MDSCs. We unraveled a tyrosine-mediated metabolic symbiosis between MDSC and UTX-deficient CRC. Loss of UTX in CRC resulted in methylation of phenylalanine hydroxylase, preventing its degradation and subsequently increasing tyrosine synthesis and secretion. Tyrosine taken up by MDSCs was metabolized to homogentisic acid by hydroxyphenylpyruvate dioxygenase. Homogentisic acid modified protein inhibitor of activated STAT3 via carbonylation of Cys 176, and relieved the inhibitory effect of protein inhibitor of activated STAT3 on signal transducer and activator of transcription 5 transcriptional activity. This in turn, promoted MDSC survival and accumulation, enabling CRC cells to acquire invasive and metastatic traits. Collectively, these findings highlight hydroxyphenylpyruvate dioxygenase as a metabolic checkpoint to restrict immunosuppressive MDSCs and to counteract malignant progression of UTX-deficient CRC.

A Systematic Review of Apicomplexa Looking into Epigenetic Pathways and the Opportunity for Novel Therapies

Interest in host epigenetic changes during apicomplexan infections increased in the last decade, mainly due to the emergence of new therapies directed to these alterations. This review aims to carry out a bibliometric analysis of the publications related to host epigenetic changes during apicomplexan infections and to summarize the main studied pathways in this context, pointing out those that represent putative drug targets. We used four databases for the article search. After screening, 116 studies were included. The bibliometric analysis revealed that the USA and China had the highest number of relevant publications. The evaluation of the selected studies revealed that Toxoplasma gondii was considered in most of the studies, non-coding RNA was the most frequently reported epigenetic event, and host defense was the most explored pathway. These findings were reinforced by an analysis of the co-occurrence of keywords. Even though we present putative targets for repurposing epidrugs and ncRNA-based drugs in apicomplexan infections, we understand that more detailed knowledge of the hosts' epigenetic pathways is still needed before establishing a definitive drug target.