骨髓增生异常综合征去甲基化药物治疗的研究进展

骨髓增生异常综合征(myelodysplastic syndrome,MDS)的发病机制涉及多阶段、多因素,基因改变与表观遗传修饰可能共同参与了这一过程.DNA甲基化是表观遗传学中一种最为重要的修饰,MDS患者常表现为总体DNA高甲基化.使用DNA甲基转移酶(DNA methyltransferase,DNMT)抑制剂降低总体甲基化水平,在MDS患者中取得了富有成效的临床反应及血液学改善.DNMT抑制剂可分为两类:5-氮杂胞苷(5-azacytidine,5-Aza-CdR)、地西他滨(5-Aza-2-deoxycytidine,decitabine)等核苷和核苷衍生物类抑制剂,它们可提高MDS患者的临床完全反应率、部分反应率及血液学改善,但缓解率、疗效尚不够令人满意;肼苯哒嗪等非核苷类抑制剂.非核苷类抑制剂与丙戊酸镁联合应用治疗MDS获得成功,为MDS去甲基化治疗药物的研究开启了一种新思路.     

DNA甲基化靶向药物在急性髓系白血病中的应用

DNA甲基化和组蛋白甲基化是急性髓系白血病（acute myeloid leukemia,AML）表观遗传学调控的常见模式,针对甲基化过程的靶向治疗包括DNA甲基转移酶抑制剂、甲基化调节蛋白抑制剂及组蛋白甲基化调控蛋白抑制剂,其中DNA甲基转移酶（DNA methyltransferase,DNMT）抑制剂阿扎胞苷、地西他滨已上市进入临床,针对甲基化调节蛋白IDH1/2抑制剂也已进入Ⅱ期临床研究。此外,针对组蛋白甲基化调控蛋白EZH2、LSD1抑制剂也显示出良好体外抗白血病活性,部分已进入Ⅰ期临床研究,为AML的治疗提供新的选择。      

骨髓增生异常综合征去甲基化治疗的研究新进展*

表观遗传学异常是骨髓增生异常综合征（myelodysplastic syndromes，MDS ）发病的重要机制之一，DNA 甲基化异常是MDS 最常见的表观遗传学改变。抑制DNA 异常甲基化可以改善部分MDS 患者的病情，延长其生存期。目前被批准用于MDS 临床治疗的去甲基化药物主要有阿扎胞苷和地西他滨，这两种药物在MDS 患者中均显示出一定的疗效。本文对该药的应用及进展研究进行综述。      

骨髓增生异常综合征患者DNMT3A基因突变的情况及临床意义

目的:了解DNMT3A基因突变在骨髓增生异常综合征（MDS）患者中的发生率、分布情况及临床意义。方法:选取2012年 4月至2018年6月在湘雅二医院和湘潭市中心医院住院的85例MDS患者为研究对象。提取患者外周血基因组DNA,针对DNMT3A基因突变热点R882位点设计合成引物,采用聚合酶链式反应法扩增DNMT3A基因23号外显子整个编码区基因片段,再将扩增产物纯化后测序,分析DNMT3A基因突变在本组患者中的发生率、分布情况及临床意义。结果:85例MDS患者中检测到5例DNMT3A基因突变,突变阳性率5.9%,其中R882H和R882C基因突变各2例,R882P基因突变1例,未见R882S基因突变。常规化疗联合去甲基化药物地西他滨治疗后,1例患者一度获得血液学完全缓解,复杂染色体核型恢复正常,但是1月后转为急性白血病并发严重感染死亡,1例治疗后稳定,2例治疗后疾病进展,1例死于肺部感染。结论:MDS中DNMT3A基因突变率低,其突变多预示预后较差,并可能更快地向急性髓性白血病转化,选择去甲基化药物治疗可能获益。     

DNA甲基转移酶基因在MDS患者中的表达及其临床意义

研究DNA甲基转移酶（DNA methyltransferase,DNMT）基因在骨髓增生异常综合征（MDS）患者中的表达及其与抑癌基因p15INK4B甲基化状态的相关性,并进一步探讨其与临床预后的关系。方法:采用SYBR Green I实时定量逆转录聚合酶链反应（Real-time RT-PCR）方法对48例初治MDS患者和20例正常人骨髓进行DNMT1、DNMT3A、DNMT3B mRNA水平检测;采用甲基化特异性PCR（MSP-PCR）方法检测48例初治MDS患者和20例正常人p15INK4B基因的甲基化状态。结果:低危组MDS患者3种DNMTs mRNA与正常对照组相比,表达水平差异均无统计学意义（P>0.05）;高危组MDS患者3种DNMTs mRNA表达显著高于低危组和正常对照组（P均<0.01）;MDS患者DNMTs mRNA表达水平与p15INK4B基因甲基化程度呈正相关。12例高危MDS患者接受了地西他滨治疗,另外15例高危MDS患者接受了IA/DA联合化疗,地西他滨组疗效与联合化疗组比较差异无统计学意义（P>0.05）。结论:DNMTs基因的异常高表达,导致细胞周期调控相关的p15INK4B等抑癌基因启动子CpG岛过甲基化失活,在MDS患者由低危向高危转变乃至进展为急性髓系白血病（AML）过程中,起着至关重要的作用,DNMTs mRNA表达水平可以作为一种判断MDS预后的指标。      

开展以遗传学类型分类骨髓增生异常综合征的针对性治疗研究

 骨髓增生异常综合征（MDS）是一组起源于造血干（祖）细胞的克隆性异质性疾患,其异质性不仅表现在外周血和骨髓象上,也表现在细胞遗传学、病程、转归和对治疗的反应上。MDS遗传学分型应是一种能反映疾病本质的分型方法。国内应开展MDS遗传学分型的生物学特征研究,以MDS遗传学分型指导MDS的临床分型治疗,提高其临床疗效。     

伴SF3B1及JAK2基因突变的骨髓增生异常/骨髓增殖性肿瘤伴环形铁粒幼细胞和血小板增多合并骨髓纤维化一例并文献复习

目的:探讨骨髓增生异常/骨髓增殖性肿瘤伴环形铁粒幼细胞和血小板增多（MDS/MPN-RS-T）合并骨髓纤维化患者的临床特点及治疗。方法:回顾性分析南方医科大学深圳医院2018年5月收治的1例伴SF3B1及JAK2基因突变的MDS/MPN-RS-T合并骨髓纤维化患者的临床资料,并进行相关文献复习。结果:患者以头晕、乏力为主要症状,诊断为伴SF3B1及JAK2基因突变的MDS/MPN-RS-T合并骨髓纤维化,予羟基脲降细胞治疗及芦可替尼靶向治疗,效果良好。结论:MDS/MPN-RS-T是血液肿瘤系统新定义的克隆性造血干细胞疾病,具有两种疾病的双重特点,不易鉴别,易漏诊,其有效的治疗选择仍有待探索。     

雷利度胺治疗骨髓增生异常综合征的研究进展

 【摘要】　可用于治疗骨髓增生异常综合征（MDS）的药物十分有限，而且很多类型的MDS预后较差，近年来，雷利度胺开始得到大家的关注。目前，雷利度胺已被美国食品与药物管理局（FDA）批准用于治疗国际预后评分系统（IPSS）评分中低危／中危-1且核型为5q31-伴或不伴其他细胞遗传学异常的MDS。但其作用机制仍不清楚。现有研究发现，雷利度胺对其他类型的MDS也有较好的疗效，或可推广应用于其他类型MDS的治疗。     

骨髓增生异常综合征诊断和治疗进展

骨髓增生异常综合征（myelodysplastic syndromes,MDS）是一组起源于造血干细胞向白血病转化的克隆性疾病,其异质性和临床预后差异性极大。随着二代测序技术的出现,在MDS中已经确定了涉及表观遗传学调控、染色质修饰、剪接体、信号传导和DNA修复通路的体细胞基因突变。最新的WHO分型更强调了血细胞多系异常造血及基因突变,这些突变对MDS的分型及预后发挥着越来越重要的作用,可用于指导治疗及评价预后。目前MDS的批准用药主要有去甲基化药物及免疫抑制剂,但均无法治愈MDS,异基因造血干细胞移植（autologous stem cell transplantation,ASCT）仍然是唯一有效的治愈方法。本文就MDS诊断、治疗进展和预后作一综述。     

骨髓增生异常综合征的分子生物学研究：现况与启示

应用第2代测序技术对大系列骨髓增生异常综合征（MDS）患者的研究初步揭示了MDS发病的分子基础.约90％的MDS患者至少存在≥1个基因突变,最常受累基因有SF3B1、TET2、SRSF2、ASXL1、DNMT3A和RUNX1.随着MDS克隆演变和受累基因功能研究的深入,有望进一步完善MDS患者的预后判断和治疗方案的制定.     

IDH mutations in cancer and progress toward development of targeted therapeutics

Isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) are key metabolic enzymes, converting isocitrate to &agr;-ketoglutarate (&agr;KG). IDH1 and IDH2 mutations have been identified in multiple tumor types, including gliomas and myeloid malignancies such as acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS). Here we provide an overview of the function of normal and mutated IDH, discuss the role of IDH mutations in tumorigenesis and progression and review the key clinical considerations when treating IDH-mutated tumors based on emerging clinical data from mutant IDH1/2 inhibitor trials. IDH1 and IDH2 mutations confer neomorphic activity in the mutant protein, resulting in the conversion of &agr;KG to the oncometabolite, D-2-hydroxyglutarate (2-HG). The subsequent accumulation of 2-HG results in epigenetic dysregulation via inhibition of &agr;KG-dependent histone and DNA demethylases, and a block in cellular differentiation. There is growing preclinical and clinical evidence suggesting that IDH mutations are involved in neoplasia. Furthermore, preclinical studies assessing small molecule inhibitors of mutant IDH1/2 enzymes have provided proof of concept that this approach decreases intracellular 2-HG levels, reverses epigenetic dysregulation and induces cellular differentiation. Phase I studies of mutant IDH inhibitors are currently ongoing in patients with IDH-mutant hematologic and solid tumors, with early data in hematologic tumors suggesting a manageable safety profile as well as clinical benefit, with a mechanism of action based on differentiation of malignant cells. Inhibition of mutant IDH shows promise as a treatment approach in hematologic malignancies, with further development ongoing in solid tumors and glioma. The mutant IDH inhibitors may have clinical utility both as single agents and in combination strategies that target additional oncogenic pathways.     

Inhibitors of DNA methylation: beyond myelodysplastic syndromes

DNA methyltransferase (DNMT) inhibitors, azacitidine (Vidaza, Pharmion, Boulder, CO, USA) and decitabine (Dacogen; SuperGen Inc, Dublin, CA, USA, and MGI Pharma Inc, Bloomington, MN, USA), have had a significant impact on the treatment paradigm of myelodysplastic syndromes (MDSs), previously managed mainly by supportive care and hematopoietic-stem-cell transplantation. The positive clinical experience seen in MDS to date coupled with the persistent challenges faced in the treatment of other hematologic malignancies has served as the impetus for further exploration of the therapeutic value of DNMT inhibitors beyond MDS. In that respect, the majority of data for these agents are in the setting of acute myelogenous leukemia (AML). Experience with these agents in patients with refractory anemia with excess blasts in transformation (reclassified by the World Health Organization as AML) was also reported in the clinical trials submitted to the FDA for approval of azacitidine for MDS. Some use has also been described in chronic myelogenous leukemia and acute lymphocytic leukemia. Further studies are needed to clarify the appropriate dose and the number and duration of cycles in the treatment of leukemias, and to identify ideal candidates for therapy, explore the role of DNMT inhibitors in combination with other agents, especially histone deacetylase inhibitors, delineate differences between the commercially available agents, and establish the long-term safety of these agents. To this end, experience with DNMT inhibitors in hematologic malignancies other than MDS is reviewed in an effort to better understand the therapeutic potential of these agents and to define areas of future exploration in these settings.     

非髓性白血病的其他恶性血液病中DNMT3A基因突变的检测及临床意义

目的:了解DNMT3A基因突变在非髓性白血病的其他恶性血液肿瘤患者中的发生率、分布情况及临床意义.方法:选取196例非髓性白血病的其他血液肿瘤患者为研究对象,提取患者外周血基因组DNA,针对DNMT3A基因突变热点R882位点设计引物,采用聚合酶链式反应(PCR)法扩增DNMT3A基因23号外显子整个编码区基因片段,再将扩增产物纯化后测序,分析DNMT3A基因突变在本组恶性血液病患者中的发生率、分布情况及临床意义.结果:在57例非霍奇金淋巴瘤及34例骨髓增生异常综合征患者中各检出1例伴DNMT3A基因突变,在25例急性淋巴细胞白血病、45例多发性骨髓瘤和35例骨髓增殖性肿瘤患者中均未检测到DNMT'3A基因突变.结论:非髓性白血病的其他血液肿瘤患者中DNMT3A基因突变少见,伴DNMT3A基因突变的1例T淋巴母细胞白血病/淋巴瘤患者预后不良,联合去甲基化药物的化疗方案使伴该基因突变的1例骨髓增生异常综合征患者一度获得血液学完全缓解,但短期内转为急性白血病并发严重感染死亡.     

The role of mutations in epigenetic regulators in myeloid malignancies

Recent genomic studies have identified novel recurrent somatic mutations in patients with myeloid malignancies, including myeloproliferative neoplasms (MPNs), myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML). In some cases these mutations occur in genes with known roles in regulating chromatin and/or methylation states in haematopoietic progenitors, and in other cases genetic and functional studies have elucidated a role for specific mutations in altering epigenetic patterning in myeloid malignancies. In this Review we discuss recent genetic and functional data implicating mutations in epigenetic modifiers, including tet methylcytosine dioxygenase 2 (TET2), isocitrate dehydrogenase 1 (IDH1), IDH2, additional sex combs-like 1 (ASXL1), enhancer of zeste homologue 2 (EZH2) and DNA methyltransferase 3A (DNMT3A), in the pathogenesis of MPN, MDS and AML, and discuss how this knowledge is leading to novel clinical, biological and therapeutic insights.     

1号染色体克隆性异常在恶性血液病中的发生及其临床意义

 【摘要】　目的　分析1号染色体克隆性异常在恶性血液病中的发生频率、类型,并了解其临床和生物学意义。方法　对256例恶性血液病患者细胞遗传学R显带核型分析结果进行回顾性分析,将累及1号染色体的异常核型及临床资料进行总结,并结合文献复习。结果　256例中涉及1号染色体异常的恶性血液病25例（9.8 %）,分别见于急性淋巴细胞白血病（ALL）-L2、急性非淋巴细胞白血病（ANLL）-M2、骨髓增生异常综全征（MDS）、多发性骨髓瘤（MM）、淋巴瘤骨髓浸润、慢性粒细胞加速急变期、浆细胞白血病、慢性粒单核细胞白血病。1号染色体与其他染色体之间易位11例,其中t（1;19）3例,t（1;14） 2例,1号染色体部分增加或缺失7例,增加1条完整或部分缺失的1号染色体7例,1q等臂染色体3例。t（1;19）见于B淋巴细胞增殖性疾病,t（1;14）见于急性T淋巴细胞白血病。随访涉及1号染色体异常的20例患者,临床疗效差,生存期短。结论　发生1号染色体克隆性异常的恶性血液病主要见于急性白血病、MDS、MM,非特异性的异常多见,特异性的异常主要是与其他染色体之间的易位和1q三体。具有重现性的异常有t（1;19）、t（1;14）,均与白血病免疫表型相关;而1q三体和1q21扩增分别对MDS和MM的治疗、预后有指导意义。     

Updates in Cytogenetics and Molecular Markers in MDS

Myelodysplastic syndromes (MDS) are clonal hematologic neoplasms that can result in cytopenias and increase the risk of leukemic transformation. The disease is characterized by several recurrent cytogenetic defects, which can affect diagnosis, prognosis, and treatment. Metaphase cytogenetics (MC) is the gold standard in karyotypic analysis in hematology. Progress in molecular analysis, including additional karyotypic tools exemplified by fluorescence in situ hybridization, comparative genomic hybridization, and more importantly, single nucleotide polymorphism array (SNP-A) analysis, has led to increased detection of chromosomal abnormalities in myeloid malignancies and improved prognostic risk stratification. SNP-A, together with MC, has also been instrumental in the discovery of genes that have improved our understanding of the biology of MDS. Newly elucidated molecular abnormalities in MDS include mutations in CBL, TET2, ASXL1, IDH1/IDH2, EZH2, DNMT3A, and UTX. This review provides an update on the changing landscape of molecular and cytogenetic characterization in MDS and its significance in disease biology and clinical practice.     

Clinical applications of epigenetic markers and epigenetic profiling in myeloid malignancies

Aberrant DNA methylation is frequent in the myeloid malignancies, particularly myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML). Promoter CpG methylation is correlated with silencing of tumor-suppressor genes (TSGs) in specific pathways that are also targets of mutation or other mechanisms of inactivation, and is thought to contribute to disease progression and poor prognosis. Epigenetic contributions to myeloid pathogenesis are more complex. Examples include TSG inactivation and oncogenic activation associated with formation of altered chromatin separate from CpG methylation. Epigenetic dysregulation occurs at multiple disease stages and at non-CpG island genomic sites, and also includes genomic hypomethylation and small RNA mechanisms of epigenetic regulation. Identification of recurrent mutations in potential epigenetic regulators, including TET2, IDH1, IDH2, DNMT3A, UTX, and ASXL1, were recently described. Accordingly, therapeutics directed towards epigenetic mechanisms including methylation inhibitors and histone deacetylase (HDAC) inhibitors have had some clinical success when applied to MDS and AML. However, identification of the underlying mechanisms associated with clinical responses and drug resistance remain enigmatic. Remarkably, in spite of significant molecular and translational progress, there are currently no epigenetic biomarkers in widespread clinical use. In this review, we explore the potential applications of epigenetic biomarker discovery, including epigenetic profiling for myeloid malignancy pathogenesis understanding, diagnostic classification, and development of effective treatment paradigms for these generally considered poor prognosis disorders.     

血液肿瘤患者IDH1/IDH2基因突变发生率及其血液病分布特点研究

目的研究血液肿瘤患者IDH1/IDH2基因突变发生率和其血液病的分布特点。方法选取300例不同病情的血液患者骨髓的样品,其中245例是血液肿瘤患者,55例是非血液肿瘤患者,对比检查IDH2和IDH1基因是否在血液肿瘤患者中有突变行为。结果实验共发现了7个错义的突变基因,其中2个为IDH2基因突变,4个为IDH1基因突变;还有2个IDH1同义突变。4个IDH1的突变类型是:199M、R132C、R131D及R132H,前3个突变患者都出现在3例急性髓系的白血病患者中,最后1例出现在急性淋巴细胞的白血病患者中。2个IDH2的错义突变类型是:P167R和R172K,这2例也都出现在急性髓系的白血病患者中。在发生IDH1错义基因突变的4例患者中,其体内白细胞的计数明显升高,且患者年龄均大于60岁。在发生IDH2基因突变的2例患者中,其体内白细胞的计数明显降低,且患者的年龄均小于60岁。结论 96例的急性白血病患者的检测中,共发现了9例突变患者,其IDH的突变率是9.3%,且7例发生于急性髓系的白血病患者中,说明急性髓系的白血病的发病和IDH的突变有紧密的联系。     

Efficient induction of differentiation and growth inhibition in IDH1 mutant glioma cells by the DNMT Inhibitor Decitabine

Mutation in the IDH1 or IDH2 genes occurs frequently in gliomas and other human malignancies. In intermediate grade gliomas, IDH1 mutation is found in over 70% of tumors. These mutations impart the mutant IDH enzyme with a neomorphic activity – the ability to synthesize 2-hydroxyglutarate (2-HG). This ability leads to a reprogramming of chromatin state, a block in differentiation, and the establishment of the glioma hypermethylator phenotype (G-CIMP). It has been hypothesized but not proven that the extensive DNA methylation that occurs in G-CIMP tumors helps maintain and “lock in” glioma cancer cells in a dedifferentiated state. Here, we tested this hypothesis by treating patient derived IDH1 mutant glioma initiating cells (GIC) with non-cytotoxic, epigenetically targeted doses of the DNMT inhibitor decitabine. Global methylome analysis of treated IDH1 mutant GICs showed that DAC treatment resulted in reversal of DNA methylation marks induced by IDH and the re-expression of genes associated with differentiation. Accordingly, treatment of IDH1 mutant glioma cells resulted in a dramatic loss of stem-like properties and efficient adoption of markers of differentiation, effects not seen in decitabine treated IDH wild-type GICs. Induction of differentiation was much more efficient than that seen following treatment with a specific inhibitor of mutant IDH enzyme (Agios). Decitabine also decreased replicative potential and tumor growth in vivo. Reexpression of polycomb regulated genes accompanied these DAC-induced phenotypes. In total, our data indicates that targeting the pathologic DNA methylation in IDH mutant cells can reverse mutant IDH induced hypermethylation and block in differentiation and promote tumor control. These findings have substantial impact for exploring new treatment strategies for patients with IDH mutant gliomas.