The "Epigenetic Code Replication Machinery", ECREM: a promising drugable target of the epigenetic cell memory

Discrete chemical modifications of the chromatin (DNA and primarily histones) can regulate gene expression or repression and can be transmitted to the descent (cells or organisms) thanks to an epigenetic memory. These modifications involve histone post-translational modifications, DNA methylation at CpG islands and small nuclear RNAs processes. They play fundamental roles in cell proliferation and differentiation. These two processes are crucial in particular during embryonic development, X chromosome inactivation in females, genomic imprinting, gene bookmarking, cell reprogramming, position effect and silencing of retroviral elements. While, only one major DNA modification is known, more than 150 post-translation modifications of histones have been reported, including methylations, acetylations, ubiquitinations, SUMOylations and phosphorylations. How these modifications are inherited from mother cells to daughter cells or from an organism to its descent remains a major scientific challenge. We propose here a macro-molecular complex, called ECREM for "Epigenetic Code REplication Machinery", as being involved in the inheritance of the epigenetic code. The composition of ECREM may vary in a spatio-temporal manner according to the chromatin state, the cell phenotype and the development stage. We describe the members of ECREM, responsible for the epigenetic code inheritance, i.e., enzymes involved in DNA methylation and histone post-translational modifications. Some of them, such as DNA methyltransferases (DNMTs), histone acetyltransferases (HATs) and histone deacetylases (HDACS including sirtuins), have been found to be deregulated in several types of pathologies and are already targeted by inhibitors. ECREM, thus, appears to be an interesting complex to be investigated in order to find new drugs for cancer, metabolic, neuro-degenerative and inflammatory diseases therapy.     

冬凌草甲素上调人前列腺癌PC-3细胞PTEN基因的表达及其诱导凋亡作用

目的：研究冬凌草甲素诱导人前列腺癌PC-3细胞凋亡及与PTEN基因表达的关系。方法：MTT比色法测定冬凌草甲素诱导PC-3细胞的增长抑制;流式细胞术分析DNA倍体测定细胞凋亡;RT-PCR半定量分析冬凌草甲素诱导人前列腺癌PC-3细胞后PTENmRNA的表达情况。结果：MTT比色法测得冬凌草甲素对PC-3细胞具有明显的增殖抑制作用,并随冬凌草甲素浓度的升高及作用时间的延长而增强;流式细胞术分析PC-3细胞经冬凌草甲素诱导后,细胞凋亡比率随作用时间延长而增加。RT-PCR测得PTEN基因mRNA的表达随细胞凋亡的增加而逐渐增高。结论：冬凌草甲素能够明显抑制PC-3细胞的增殖,并诱导前列腺癌细胞凋亡,其作用途径可能与上调PTENmRNA表达有关。     

Ursolic acid-induced apoptosis in K562 cells involving upregulation of PTEN gene expression and inactivation of the PI3K/Akt pathway

Ursolic acid (UA), a pentacyclic triterpenoid derived from a variety of medicinal plants, exhibits potent anticancer activity against many types of cancer cells. However, the anticancer mechanism of UA is not clearly understood. Suppression of phosphatase and a tensin homolog deleted on chromosome 10 (PTEN) gene expression leading to activation of the phosphatidylinositol-3-OH kinase (PI3K)/Akt pathway has been observed in many cancers including leukemia, making the PTEN gene and PI3K/Akt pathway a central target for cancer therapy. Here, we demonstrated that UA was able to inhibit growth, induce apoptosis in a human chronic myelogenous leukemia cell line (K562 cells) via upregulation of PTEN gene expression, inhibit Akt kinase activity, change mitochondrial transmembrane potential and reduce the release of cytochrome c and the activity of caspases. These results suggest that UA may elicit its strong antitumor effects via upregulation of the PTEN gene and inhibition of the PI3K/Akt pathway.     

Targeting epigenetic modifiers in cancer

The chromatin structure of a gene plays an important role in regulating its expression. This structure is established through the action of various protein complexes that remodel nucleosomes, catalyse post-translational modifications, deposit histone variants and methylate DNA. Together these complexes establish epigenetic marks that influence expression of the gene. Some of these epigenetic marks are transient while others, such as those involved in silencing genes are more stable and can require several cell divisions to be fully implemented or reversed. Deregulated gene expression programs are a feature of cancer biology and it is now apparent that epigenetic changes, as well as genetic changes, are important in establishing these aberrant expression patterns. However, unlike genetic alterations, epigenetic changes are reversible. The complexes that catalyse these modifications therefore represent valuable targets for therapeutic intervention. Here we will review the most recent literature describing the protein complexes that catalyse epigenetic modifications and the inhibitors of these complexes that are being pursued as cancer drugs. In addition we will highlight those epigenetic modifiers that provide promise as therapeutic targets but for which inhibitors are not currently available.     

异黄酮对前列腺癌细胞及PTEN基因的影响

目的 通过大豆异黄酮抑制前列腺癌PC-3和LNCaP细胞生长和相关基因表达的影响，探讨大豆异黄酮类抑制前列腺癌的机制。方法 对染料木黄酮和大豆甙元作用后的雄激素非依赖型前列腺癌(PC-3)、雄激素依赖型前列腺癌LNCaP细胞的存活率、细胞毒作用、细胞周期和PTEN基因的mRNA表达等进行了研究。结果 染料木黄酮和大豆甙元对PC-3、LNCaP细胞抑制效应具有时间和剂量依赖性，具有诱导凋亡和引起坏死效应；染料木黄酮能诱导PC-3和LNCaP细胞的(PTEN)表达，而大豆甙元只能诱导LNCaP细胞PTEN表达。结论 PTEN基因表达的变化可能在染料木黄酮和大豆甙元抑制PC-3和LNCaP细胞中具有重要的意义。染料木黄酮和大豆甙元抑制PC-3和LNCaP细胞可能存在多种作用途径。     

The complexity of PTEN: mutation, marker and potential target for therapeutic intervention

Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is a phosphatase that removes phosphates primarily from lipids. It has also been called mutated in multiple advanced cancers 1 and transforming growth factor-beta regulated epithelial cell-enriched phosphatase 1. The best described substrate of PTEN is phosphatidyliniositol (3,4,5)-tris-phosphate [PtdIns(3,4,5)P3]. PTEN removes the phosphate in PtdIns(3,4,5)P(3) to generate PtdIns(4,5)P(2). PTEN serves to counter-balance the effects of phosphoinositide 3' kinase, which normally adds a phosphate to PtdIns(4,5)P(2) to generate PtdIns(3,4,5)P(3). PtdIns(3,4,5)P(3) recruits kinases such as phosphoinositide-dependent kinase 1, which in turn phosphorylate Akt, which phosphorylates other downstream proteins involved in regulation of apoptosis and cell-cycle progression. PTEN removal of the phosphate from PtdIns(3,4,5)P(3) inhibits this pathway by preventing localisation of proteins with pleckstrin homology domains to the cell membrane. Alterations of the PTEN gene are associated with cancer and other diseases. Novel therapeutic approaches have been developed to counteract the deletion/mutation of PTEN in human cancer. This review will discuss the role of PTEN in signal transduction and cancer as well as pharmacological approaches to combat PTEN loss in human cancer.     

抑癌基因PTEN在急性白血病中的表达及意义

目的 研究在急性白血病（AL）中抑癌基因PTEN的表达情况,并对急性白血病发生与预后的关系进行探究分析.方法 使用聚合酶链式反应法（PCR）对2010年2月至2011年8月安徽医科大学第一附属医院血液科就诊的住院及门诊收治的111例急性白血病（AL）患者抑癌基因的表达情况进行检测,并分析PTEN表达情况与临床指标之间的相关性.结果 111例AL患者中检测到34例患者抑癌基因PTEN表达阴性缺失,AL患者中PTEN（-）患者的完全缓解率低于PTEN（＋）的患者（P＜0.05）;PTEN基因阴性缺失与骨髓原始细胞百分比、外周血白细胞计数存在正相关.结论 急性白血病的发生可能有抑癌基因PTEN的参与,其对急性白血病患者的转归和复发具有重要意义.     

胃癌组织中PTEN和MMP-9的表达与腹腔游离癌细胞的相关性

目的 观察PTEN和基质金属蛋白酶9（MMP-9）在胃癌组织中的表达,探讨其在癌变机制中的作用。方法 采用SP法检测65例胃癌组织中PTEN、MMP-9的表达,同时应用实时荧光RT-PCR方法检测胃癌术中腹腔灌洗液中游离癌细胞。结果 （1）PTEN、MMP-9阳性表达率在有无腹腔游离癌细胞组之间的差异均有显著性（P〈0．05）;（2）PTEN与MMP-9的表达呈负相关。结论 PTEN、MMP-9可以反映腹腔游离癌细胞是否存在,为评估预后及指导化疗提供参考。     

Enzymatic and non-enzymatic activities of SHIP-1 in signal transduction and cancer

PI3K cascade is a central signaling pathway regulating cell proliferation, growth, differentiation, and survival. Tight regulation of the PI3K signaling pathway is necessary to avoid aberrant cell proliferation and cancer development. Together with SHIP-1, the inositol phosphatases PTEN and SHIP-2 are the gatekeepers of this pathway. In this review, we will focus on SHIP-1 functions. Negative regulation of immune cell activation by SHIP-1 is well characterized. Besides its catalytic activity, SHIP-1 also displays non-enzymatic activity playing role in several immune pathways. Indeed, SHIP-1 exhibits several domains that mediate protein-protein interaction. This review emphasizes the negative regulation of immune cell activation by SHIP-1 that is mediated by its protein-protein interaction.     

DNMT1-mediated PTEN hypermethylation confers hepatic stellate cell activation and liver fibrogenesis in rats

Hepatic stellate cell (HSC) activation is an essential event during liver fibrogenesis. Phosphatase and tension homolog deleted on chromosome 10 (PTEN), a tumor suppressor, is a negative regulator of this process. PTEN promoter hypermethylation is a major epigenetic silencing mechanism in tumors. The present study aimed to investigate whether PTEN promoter methylation was involved in HSC activation and liver fibrosis. Treatment of activated HSCs with the DNA methylation inhibitor 5-aza-2′-deoxycytidine (5-azadC) decreased aberrant hypermethylation of the PTEN gene promoter and prevented the loss of PTEN expression that occurred during HSC activation. Silencing DNA methyltransferase 1 (DNMT1) gene also decreased the PTEN gene promoter methylation and upregulated the PTEN gene expression in activated HSC-T6 cells. In addition, knockdown of DNMT1 inhibited the activation of both ERK and AKT pathways in HSC-T6 cells. These results suggest that DNMT1-mediated PTEN hypermethylation caused the loss of PTEN expression, followed by the activation of the PI3K/AKT and ERK pathways, resulting in HSC activation.     

微小 RNA-425-5p靶向调控 PTEN表达促进宫颈癌海拉细胞增殖、侵袭和迁移的实验研究

目的 探讨微小RNA(miR)-425-5p对宫颈癌海拉细胞增殖、侵袭和迁移的影响及其机制.方法 2018年10月至2019年10月,利用Lipofectamine TM 2000将miR-425-5p模拟物(mimics)、抑制剂(inhibitor)及其阴性对照转染至海拉细胞中,采用实时荧光定量PCR检测细胞中miR-425-5p的表达情况,MTT法检测细胞增殖活力,平板克隆形成实验检测细胞克隆形成能力,Transwell小室实验检测细胞侵袭和迁移,蛋白质印迹法(Western blotting)检测细胞中磷酸酶及张力蛋白同源基因(PTEN)蛋白的表达.采用生物信息学软件预测、双荧光素酶报告基因实验验证miR-425-5p和PTEN的靶向关系.将PTEN过表达质粒pcDNA3.1-PTEN和PTEN干扰序列siRNA-PTEN转染至海拉细胞后,观察PTEN对海拉细胞增殖活力、克隆形成能力、侵袭能力和迁移能力的影响.结果 与各自阴性对照比较,miR-425-5p过表达可促进海拉细胞增殖[(142.25±11.32)％比(100.00±6.67)％]、克隆形成、侵袭[(133.28±9.86)个比(77.46±5.32)个]和迁移[(187.56±15.12)个比(115.35±10.26)个]并下调PTEN蛋白表达,而miR-425-5p低表达则抑制海拉细胞增殖[(58.38±3.45)比(100.00±5.74)]、克隆形成、侵袭[(43.32±3.62)个比(75.65±6.15)个]和迁移[(62.28±4.05)个比(109.72±9.84)个]并上调PTEN蛋白表达(P<0.05).双荧光素酶报告基因实验证实,PTEN是miR-425-5p的靶基因;PTEN过表达可促进海拉细胞增殖[(66.52±4.36)％比(100.00±6.18)％]、克隆形成、侵袭[(46.23±3.13)个比(71.65±5.24)个]和迁移[(62.65±4.26)个比(108.42±8.57)个],而PTEN低表达则抑制海拉细胞增殖[(136.52±9.85)个比(100.00±7.05)个]、克隆形成、侵袭[(110.27±9.85)个比(70.52±6.18)个]和迁移[(168.78±16.96)个比(113.26±10.13)个].结论 miR-425-5p可通过靶向调控PTEN表达促进宫颈癌海拉细胞增殖、侵袭和迁移.     

Targeting the histone orthography of cancer: drugs for writers,erasers and readers

Gene expression is dynamically controlled by epigenetics through post-translational modifications of histones, chromatin-associated proteins and DNA itself. All these elements are required for the maintenance of chromatin structure and cell identity in the context of a normal cellular phenotype. Disruption of epigenetic regulation is a common event in human cancer. Here, we review the key protein families that control epigenetic signalling through writing, erasing or reading specific post-translational modifications. By exploiting the leading role of epigenetics in tumour development and the reversibility of epigenetic modifications, promising novel epigenetic-based therapies are being developed. In this article, we highlight the emerging low MW inhibitors targeting each class of chromatin-associated protein, their current use in preclinical and clinical trials and the likelihood of their being approved in the near future.     

非小细胞肺癌中抑癌基因PTEN和FHIT蛋白的表达及其与临床病理的关系

目的 探讨抑癌基因PTEN和FHIT与非小细胞肺癌发生、发展的关系.方法 应用免疫组化Elivision法检测78例非小细胞肺癌组织和40例非肿瘤肺组织中PTEN和FHIT在基因蛋白的表达.结果 非肿瘤肺组织FHIT和PTEN蛋白阳性表达率明显高于肺癌组织(P＜0.05);肺癌组织中FHIT和PTEN蛋白阳性表达率与肿瘤分化程度和临床分期有明显相关性(P＜0.05);而FHIT和PTEN蛋白的阳性表达率在不同类型肺癌组织以及患者年龄、肿瘤大小、有无淋巴结转移等情况的比较中差异无显著性(P＞0.05).结论 PTEN和FHIT基因蛋白表达缺失在不同类型和不同分化程度的非小细胞肺癌发生、发展中可能存在着普通意义,可作为判断肺癌生物学行为和患者预后的参考指标.     

Post-translational modification of heat-shock protein 90: impact on chaperone function

Heat-shock protein 90 (Hsp90) is a molecular chaperone required for the stability and function of many signaling proteins that are often activated, mutated or overexpressed in cancer cells and that underly cancer cell proliferation and survival. Hsp90 is a conformationally flexible protein that associates with a distinct set of cochaperones depending on ATP or ADP occupancy of an N-terminal binding pocket. Nucleotide exchange and ATP hydrolysis by Hsp90 itself, with the assistance of cochaperones, drive the Hsp90 chaperone machine to bind, chaperone and release client proteins. Cycling of the Hsp90 chaperone machine is critical to its function. Although ATP binding and hydrolysis have been convincingly implicated in regulating the Hsp90 cycle, growing evidence suggests that various post-translational modifications of Hsp90, including phosphorylation, acetylation and other modifications, provide an additional overlapping or parallel level of regulation. A more complete understanding of how these various protein modifications are regulated and interact with each other at the cellular level to modulate Hsp90 chaperone activity is critical to the design of novel approaches to inhibit this medically important molecular target.     

PTEN在胃癌中的研究进展

胃癌是我国最常见的恶性肿瘤之一,其发生及发展是一个多阶段、多步骤、有序的过程。第十号染色体同源缺失性磷酸酶-张力蛋白基因(PTEN)作为继P53基因之后发现的最重要的肿瘤抑制基因,调控着细胞周期和多种信号途径,与人类多种恶性肿瘤的发生发展相关,其表达减少或缺失在胃癌的发生发展过程中亦有重要作用。深入研究PTEN在胃癌发生中的相关机制,将为胃癌的早期诊断、治疗以及预后评价提供新的方法和途径。     

The complexity of PTEN: mutation,marker and potential target for therapeutic intervention

Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is a phosphatase that removes phosphates primarily from lipids. It has also been called mutated in multiple advanced cancers 1 and transforming growth factor-β regulated epithelial cell-enriched phosphatase 1. The best described substrate of PTEN is phosphatidyliniositol (3,4,5)-tris-phosphate [PtdIns(3,4,5)P3]. PTEN removes the phosphate in PtdIns(3,4,5)P₃ to generate PtdIns(4,5)P₂. PTEN serves to counter-balance the effects of phosphoinositide 3′ kinase, which normally adds a phosphate to PtdIns(4,5)P₂ to generate PtdIns(3,4,5)P₃. PtdIns(3,4,5)P₃ recruits kinases such as phosphoinositide-dependent kinase 1, which in turn phosphorylate Akt, which phosphorylates other downstream proteins involved in regulation of apoptosis and cell-cycle progression. PTEN removal of the phosphate from PtdIns(3,4,5)P₃ inhibits this pathway by preventing localisation of proteins with pleckstrin homology domains to the cell membrane. Alterations of the PTEN gene are associated with cancer and other diseases. Novel therapeutic approaches have been developed to counteract the deletion/mutation of PTEN in human cancer. This review will discuss the role of PTEN in signal transduction and cancer as well as pharmacological approaches to combat PTEN loss in human cancer.     

The miR-21/PTEN/Akt signaling pathway is involved in the anti-tumoral effects of zoledronic acid in human breast cancer cell lines

Preclinical data indicate a direct anti-tumor effect of zoledronic acid (ZA) outside the skeleton, but its molecular mechanism is still not completely clarified. The aim of this study was to investigate the anti-cancer effects of ZA in human breast cancer cell lines, suggesting that they may in part be mediated via the miR-21/PTEN/Akt signaling pathway. The effect of ZA on cell viability was measured by MTT assay, and cell death induction was analyzed using either a double AO/EtBr staining and M30 ELISA assay. A Proteome Profiler Human Apoptosis Array was executed to evaluate the molecular basis of ZA-induced apoptosis. Cell cycle analysis was executed by flow cytometry. The effect of ZA on miR-21 expression was quantified by qRT-PCR, and the amount of PTEN protein and its targets were analyzed by Western blot. ZA inhibited cell growth in a concentration- and time-dependent manner, through the activation of cell death pathways and arrest of cell cycle progression. ZA downregulated the expression of miR-21, resulting in dephosphorilation of Akt and Bad and in a significant increase of p21 and p27 proteins expression. These results were observed also in MDA-MB-231 cells, commonly used as an experimental model of bone metastasis of breast cancer. This study revealed, for the first time, an involvement of the miR-21/PTEN/Akt signaling pathway in the mechanism of ZA anti-cancer actions in breast cancer cells. We would like to underline that this pathway is present both in the hormone responsive BC cell line (MCF-7) as well as in a triple negative cell line (MDA-MB-231). Taken together these results reinforce the use of ZA in clinical practice, suggesting the role of miR-21 as a possible mediator of its therapeutic efficacy.     

Treatment of PTEN-Null Breast Cancer by a Synthetic Lethal Approach Involving PARP1 Gene Silencing

The loss of the phosphatase and tensin homolog (PTEN) deleted from chromosome 10 is frequently observed in a variety of human cancers and appears to be an ideal target in synthetic lethality–based treatment. In this study, the synthetic lethal interaction between PTEN loss and the gene silencing of poly [ADP-ribose] polymerase 1 (PARP1) was examined in human triple-negative breast cancer cells (PTEN-null MDA-MB-468 and PTEN-positive MDA-MB-231 cells). Polycation liposomes previously developed by us were employed to deliver the small interfering ribonucleic acid (siRNA) targeted toward PARP1 (siPARP1) into the cancer cells. The silencing of the PARP1 gene exerted a cytocidal effect on the MDA-MB-468 cells but had no effect on the MDA-MB-231 cells and the human umbilical vein endothelial cells employed as normal cells. The simultaneous knockdown of PARP1 and PTEN in the MDA-MB-231 cells resulted in the significant inhibition of cell growth. The data suggest that the effects of the PARP1 knockdown on the cells were dependent on the PTEN status. A significant increase in the DNA breaks and the extent of apoptosis, possibly due to the failure of DNA repair, was observed upon PARP1 knockdown in the MDA-MB-468 cells compared with the case in the MDA-MB-231 cells. Our findings suggest that the synthetic lethal approach via PARP1 gene silencing holds promise for the treatment of patients with PTEN-null breast cancer.