Epigenetic regulation of autophagy: A key modification in cancer cells and cancer stem cells

Aberrant epigenetic alterations play a decisive role in cancer initiation and propagation via the regulation of key tumor suppressor genes and oncogenes or by modulation of essential signaling pathways. Autophagy is a highly regulated mechanism required for the recycling and degradation of surplus and damaged cytoplasmic constituents in a lysosome dependent manner. In cancer, autophagy has a divergent role. For instance, autophagy elicits tumor promoting functions by facilitating metabolic adaption and plasticity in cancer stem cells (CSCs) and cancer cells. Moreover, autophagy exerts pro-survival mechanisms to these cancerous cells by influencing survival, dormancy, immunosurveillance, invasion, metastasis, and resistance to anti-cancer therapies. In addition, recent studies have demonstrated that various tumor suppressor genes and oncogenes involved in autophagy, are tightly regulated via different epigenetic modifications, such as DNA methylation, histone modifications and non-coding RNAs. The impact of epigenetic regulation of autophagy in cancer cells and CSCs is not well-understood. Therefore, uncovering the complex mechanism of epigenetic regulation of autophagy provides an opportunity to improve and discover novel cancer therapeutics. Subsequently, this would aid in improving clinical outcome for cancer patients. In this review, we provide a comprehensive overview of the existing knowledge available on epigenetic regulation of autophagy and its importance in the maintenance and homeostasis of CSCs and cancer cells.     

多能性胚胎干细胞与DNA甲基化的关系

胚胎干细胞是一种能够维持自我更新、具有无限扩增能力的多能性干细胞。灵长类多能干细胞（iPSCs）根据其发育能力、细胞形态、基因表达谱以及表观遗传学的差异分为初始态多能干细胞（pPSCs）和原始态多能干细胞（nPSCs）。nPSCs因其容易进行基因工程处理以及体内外再生出功能组织器官等优势而在临床潜在应用上备受关注,因而有效维持ESCs的原始状态对其用于基础及临床研究具有重要意义。nPSCs的线粒体活性和自我更新能力高于pPSCs,且这两种多能性干细胞在DNA甲基化等方面都存在明显差别,DNA甲基化在nPSCs的转化及代谢中起到重要的作用。本文综述了DNA甲基化对ESCs的作用,特别是维持原始态的作用。     

DNA methylation and hydroxymethylation in stem cells

In mammals, DNA methylation and hydroxymethylation are specific epigenetic mechanisms that can contribute to the regulation of gene expression and cellular functions. DNA methylation is important for the function of embryonic stem cells and adult stem cells (such as haematopoietic stem cells, neural stem cells and germline stem cells), and changes in DNA methylation patterns are essential for successful nuclear reprogramming. In the past several years, the rediscovery of hydroxymethylation and the TET enzymes expanded our insights tremendously and uncovered more dynamic aspects of cytosine methylation regulation. Here, we review the current knowledge and highlight the most recent advances in DNA methylation and hydroxymethylation in embryonic stem cells, induced pluripotent stem cells and several well‐studied adult stems cells. Our current understanding of stem cell epigenetics and new advances in the field will undoubtedly stimulate further clinical applications of regenerative medicine in the future. Copyright © 2015 John Wiley & Sons, Ltd.     

Epigenetic gene regulation in stem cells and correlation to cancer

Through the classic study of genetics, much has been learned about the regulation and progression of human disease. Specifically, cancer has been defined as a disease driven by genetic alterations, including mutations in tumor-suppressor genes and oncogenes, as well as chromosomal abnormalities. However, the study of normal human development has identified that in addition to classical genetics, regulation of gene expression is also modified by ‘epigenetic’ alterations including chromatin remodeling and histone variants, DNA methylation, the regulation of polycomb group proteins, and the epigenetic function of non-coding RNA. These changes are modifications inherited during both meiosis and mitosis, yet they do not result in alterations of the actual DNA sequence. A number of biological questions are directly influenced by epigenetics, such as how does a cell know when to divide, differentiate or remain quiescent, and more importantly, what happens when these pathways become altered? Do these alterations lead to the development and/or progression of cancer? This review will focus on summarizing the limited current literature involving epigenetic alterations in the context of human cancer stems cells (CSCs). The extent to which epigenetic changes define cell fate, identity, and phenotype are still under intense investigation, and many questions remain largely unanswered. Before discussing epigenetic gene silencing in CSCs, the different classifications of stem cells and their properties will be introduced. This will be followed by an introduction to the different epigenetic mechanisms. Finally, there will be a discussion of the current knowledge of epigenetic modifications in stem cells, specifically what is known from rodent systems and established cancer cell lines, and how they are leading us to understand human stem cells.     

DNA methylation in embryonic stem cells

Embryonic stem cells (ESCs) are pluripotent, self‐renewing cells. These cells can be used in applications such as cell therapy, drug development, disease modeling, and the study of cellular differentiation. Investigating the interplay of epigenetics, genetics, and gene expression in control of pluripotence and differentiation could give important insights on how these cells function. One of the best known epigenetic factors is DNA methylation, which is a major mechanism for regulation of gene expression. This phenomenon is mostly seen in imprinted genes and X‐chromosome inactivation where DNA methylation of promoter regions leads to repression of gene expression. Differential DNA methylation of pluripotence‐associated genes such as Nanog and Oct4/Pou5f1 has been observed between pluripotent and differentiated cells. It is clear that tight regulation of DNA methylation is necessary for normal development. As more associations between aberrant DNA methylation and disease are reported, the demand for high‐throughput approaches for DNA methylation analysis has increased. In this article, we highlight these methods and discuss recent DNA methylation studies on ESCs. J. Cell. Biochem. 109: 1–6, 2010. © 2009 Wiley‐Liss, Inc.     

The epigenome of AML stem and progenitor cells

Acute myeloid leukemia (AML) is sustained by a population of cancer stem cells (CSCs or cancer-initiating cell). The mechanisms underlying switches from CSCs to non-CSCs in vivo remain to be understood. We address this issue in AML from the aspect of epigenetics using genome-wide screening for DNA methylation and selected histone modifications. We found no major differences in DNA methylation, especially in promoter CpG islands, between CSCs and non-CSCs. By contrast, we found thousands of genes that change H3K4me3 and/or H3K27me3 status between stem and progenitor cells as well as between progenitor and mature cells. Stem cell related pathways and proliferation or metabolism related pathways characterize genes differentially enriched for H3K4me3/H3K27me3 in stem and progenitor populations. Bivalent genes in stem cells are more plastic during differentiation and are more likely to lose H3K4me3 than to lose H3K27me3, consistent with increasingly closed chromatin state with differentiation. Our data indicates that histone modifications but not promoter DNA methylation are involved in switches from CSCs to non-CSCs in AML.     

Epigenetic modulators for brain cancer stem cells: Implications for anticancer treatment

Primary malignant brain tumors are a major cause of morbidity and mortality in both adults and children, with a dismal prognosis despite multimodal therapeutic approaches. In the last years, a specific subpopulation of cells within the tumor bulk, named cancer stem cells(CSCs) or tumor-initiating cells, have been identified in brain tumors as responsible for cancer growth and disease progression. Stemness features of tumor cells strongly affect treatment response, leading to the escape from conventional therapeutic approaches and subsequently causing tumor relapse. Recent research efforts have focused at identifying new therapeutic strategies capable of specifically targeting CSCs in cancers by taking into consideration their complex nature. Aberrant epigenetic machinery plays a key role in the genesis and progression of brain tumors as well as inducing CSC reprogramming and preserving CSC characteristics. Thus, reverting the cancer epigenome can be considered a promising therapeutic strategy. Three main epigenetic mechanisms have been described: DNA methylation, histone modifications, and non-coding RNA, particularly micro RNAs. Each of these mechanisms has been proven to be targetable by chemical compounds, known as epigeneticbased drugs or epidrugs, that specifically target epigenetic marks. We review here recent advances in the study of epigenetic modulators promoting and sustaining brain tumor stem-like cells. We focus on their potential role in cancer therapy.     

DNA methylation in mouse embryonic stem cells and development

Mammalian development is associated with considerable changes in global DNA methylation levels at times of genomic reprogramming. Normal DNA methylation is essential for development but, despite considerable advances in our understanding of the DNA methyltransferases, the reason that development fails when DNA methylation is deficient remains unclear. Furthermore, although much is known about the enzymes that cause DNA methylation, comparatively little is known about the mechanisms or significance of active demethylation in early development. In this review, we discuss the roles of the various DNA methyltransferases and their likely functions in development.     

Regulation beyond genome sequences: DNA and histone methylation in embryonic stem cells

Embryonic stem (ES) cells distinct themselves from other cell type populations by their pluripotent ability. The unique features of ES cells are controlled by both genetic and epigenetic factors. Studies have shown that the methylation status of DNA and histones in ES cells is quite different from that of differentiated cells and somatic stem cells. Herein, we summarized recent advances in DNA and histone methylation studies of mammalian ES cells. The methylation status of several key pluripotent regulatory genes is also discussed.     

DNA damage responses in cancer stem cells: Implications for cancer therapeutic strategies

The identification of cancer stem cells(CSCs) that are responsible for tumor initiation, growth, metastasis, and therapeutic resistance might lead to a new thinking on cancer treatments. Similar to stem cells,CSCs also display high resistance to radiotherapy and chemotherapy with genotoxic agents. Thus, conventional therapy may shrink the tumor volume but cannot eliminate cancer. Eradiation of CSCs represents a novel therapeutic strategy. CSCs possess a highly efficient DNA damage response(DDR) system, which is considered as a contributor to the resistance of these cells from exposures to DNA damaging agents. Targeting of enhanced DDR in CSCs is thus proposed to facilitate the eradication of CSCs by conventional therapeutics. To achieve this aim, a better understanding of the cellular responses to DNA damage in CSCs is needed. In addition to the protein kinases and enzymes that are involved in DDR, other processes that affect the DDR including chromatin remodeling should also be explored.     

CAR-T therapy: Prospects in targeting cancer stem cells

Cancer stem cells (CSCs), a group of tumour cells with stem cell characteristics, have the ability of self-renewal, multi-lineage differentiation and tumour formation. Since CSCs are resistant to conventional radiotherapy and chemotherapy, their existence may be one of the root causes of cancer treatment failure and tumour progression. The elimination of CSCs may be effective for eventual tumour eradication. Because of the good therapeutic effects without major histocompatibility complex (MHC) restriction and the unique characteristics of CSCs, chimeric antigen receptor T-cell (CAR-T) therapy is expected to be an important method to eliminate CSCs. In this review, we have discussed the feasibility of CSCs-targeted CAR-T therapy for cancer treatment, summarized current research and clinical trials of targeting CSCs with CAR-T cells and forecasted the challenges and future direction from the perspectives of toxicity, persistence and potency, trafficking, infiltration, immunosuppressive tumour microenvironment, and tumour heterogeneity.     

DNA methylation and demethylation link the properties of mesenchymal stem cells: Regeneration and immunomodulation

Mesenchymal stem cells (MSCs) are a heterogeneous population that can be isolated from various tissues, including bone marrow, adipose tissue, umbilical cord blood, and craniofacial tissue. MSCs have attracted increasingly more attention over the years due to their regenerative capacity and function in immunomodulation. The foundation of tissue regeneration is the potential of cells to differentiate into multiple cell lineages and give rise to multiple tissue types. In addition,the immunoregulatory function of MSCs has provided insights into therapeutic treatments for immune-mediated diseases. DNA methylation and demethylation are important epigenetic mechanisms that have been shown to modulate embryonic stem cell maintenance, proliferation, differentiation and apoptosis by activating or suppressing a number of genes. In most studies, DNA hypermethylation is associated with gene suppression, while hypomethylation or demethylation is associated with gene activation. The dynamic balance of DNA methylation and demethylation is required for normal mammalian development and inhibits the onset of abnormal phenotypes. However, the exact role of DNA methylation and demethylation in MSC-based tissue regeneration and immunomodulation requires further investigation. In this review, we discuss how DNA methylation and demethylation function in multi-lineage cell differentiation and immunomodulation of MSCs based on previously published work. Furthermore, we discuss the implications of the role of DNA methylation and demethylation in MSCs for the treatment of metabolic or immune-related diseases.     

癌症中DNA甲基化基因模块筛选

肿瘤的发生受遗传学和表观遗传学修饰的共同影响。DNA甲基化是一种重要的表观遗传修饰,在癌症的发生与发展中起着重大的作用。因此找到癌症的甲基化标记物在癌症的诊断和治疗中具有重大意义。本文利用权重基因共表达网络分析的方法（WGCNA）筛选出甲基化基因模块,并分析模块向量基因,进行功能注释,最后对基因模块进行功能分析,得到DNA甲基化与肿瘤间的关系。结果显示,这些甲基化异常的基因模块与癌症的发生有着显著的关联。同时还发现某些甲基化异常的基因模块与多种癌症的发生都有着显著的关联。     

DNA methylation and epigenetic defects in carcinogenesis

It is frequently assumed that DNA-damaging agents are carcinogenic because they induce mutations. However, another strong possibility is that the damage leads to heritable changes in the methylation of cytosine in DNA. Considerable evidence exists that gene expression in mammalian cells is in part controlled by methylation of specific DNA sequences. Carcinogens may act by altering the normal epigenetic controls of gene activity in specialised cells, and thereby produce aberrant heritable phenotypes. It is known that agents which inhibit DNA methylation can be carcinogenic and that tumour cells are altered in DNA methylation.     

肿瘤干细胞与表观遗传学调控

关于恶性肿瘤发生、复发与转移机制的研究由来已久,但目前的临床治疗方法依然不能克服肿瘤复发与转移的难题,肿瘤患者的生存率并未得到显著改善。近年来的研究提示肿瘤的起源、复发与转移的真正原因可能是存在于肿瘤内的极少数具有干细胞特性的细胞,即肿瘤干细胞（cancer stem cells,CSC）。与此同时,越来越多的研究表明,对于肿瘤干细胞的发生与功能维持,表观遗传学的调控机制可能发挥着极其重要的作用。该文简要综述目前肿瘤干细胞和表观遗传学相关领域的研究进展,并对肿瘤干细胞形成及发展过程中表观遗传学的调控作用及机制进行重点介绍。     

Therapies targeting cancer stem cells: Current trends and future challenges

Traditional therapies against cancer, chemo- and radiotherapy, have multiple limitations that lead to treatment failure and cancer recurrence. These limitations are related to systemic and local toxicity, while treatment failure and cancer relapse are due to drug resistance and self-renewal, properties of a small population of tumor cells called cancer stem cells (CSCs). These cells are involved in cancer initiation, maintenance, metastasis and recurrence. Therefore, in order to develop efficient treatments that can induce a long-lasting clinical response preventing tumor relapse it is important to develop drugs that can specifically target and eliminate CSCs. Recent identification of surface markers and understanding of molecular feature associated with CSC phenotype helped with the design of effective treatments. In this review we discuss targeting surface biomarkers, signaling pathways that regulate CSCs self-renewal and differentiation, drug-efflux pumps involved in apoptosis resistance, microenvironmental signals that sustain CSCs growth, manipulation of miRNA expression, and induction of CSCs apoptosis and differentiation, with specific aim to hamper CSCs regeneration and cancer relapse. Some of these agents are under evaluation in preclinical and clinical studies, most of them for using in combination with traditional therapies. The combined therapy using conventional anticancer drugs with CSCs-targeting agents, may offer a promising strategy for management and eradication of different types of cancers.     

Identification of DNA methylation changes at cis-regulatory elements during early steps of HSC differentiation using tagmentation-based whole genome bisulfite sequencing

Epigenetic alterations during cellular differentiation are a key molecular mechanism which both instructs and reinforces the process of lineage commitment. Within the haematopoietic system, progressive changes in the DNA methylome of haematopoietic stem cells (HSCs) are essential for the effective production of mature blood cells. Inhibition or loss of function of the cellular DNA methylation machinery has been shown to lead to a severe perturbation in blood production and is also an important driver of malignant transformation. HSCs constitute a very rare cell population in the bone marrow, capable of life-long self-renewal and multi-lineage differentiation. The low abundance of HSCs has been a major technological barrier to the global analysis of the CpG methylation status within both HSCs and their immediate progeny, the multipotent progenitors (MPPs). Within this Extra View article, we review the current understanding of how the DNA methylome regulates normal and malignant hematopoiesis. We also discuss the current methodologies that are available for interrogating the DNA methylation status of HSCs and MPPs and describe a new data set that was generated using tagmentation-based whole genome bisulfite sequencing (TWGBS) in order to comprehensively map methylated cytosines using the limited amount of genomic DNA that can be harvested from rare cell populations. Extended analysis of this data set clearly demonstrates the added value of genome-wide sequencing of methylated cytosines and identifies novel important cis-acting regulatory regions that are dynamically remodeled during the first steps of haematopoietic differentiation.     

表观遗传学与肿瘤干细胞

肿瘤干细胞模型是关于肿瘤形成及生物学特征的一种重要观点。该模型认为肿瘤发生的核心是一群类似于成体干细胞的肿瘤细胞, 具有自我增殖和分化潜能, 称为肿瘤干细胞(Cancer stem cells, CSCs)。目前在多种肿瘤中都发现了CSCs, 其不仅能导致肿瘤发生, 还是引起肿瘤转移、复发、抗药的关键原因。因此, 研究CSC的调控机制具有重要意义。近年来的研究发现, 除了基础的遗传学因素外, 表观遗传学在CSCs的调控中同样具有重要作用。目前主要的表观遗传学机制包括DNA甲基化、组蛋白修饰、染色质重塑及miRNA等, 能有效调节基因表达及细胞表型, 也是肿瘤研究的新热点。文章主要围绕近几年CSCs的特性研究及表观遗传学线索, 阐述表观遗传学机制调控CSCs的最新进展。