KDM5 histone lysine demethylases as potential novel myeloid tumor suppressors

KDM5 组蛋白赖氨酸去甲基酶作为潜在的新型骨髓肿瘤抑制剂

• 批准号: 10225299
• 负责人: Julie Aurore Losman
• 金额: $ 38.27万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10225299
• 关键词: Acute Myelocytic Leukemia; Address; Affect; Biochemical; Biological; Biological Assay; Biology; Blast Cell; CRISPR screen; CRISPR/Cas technology; Cell Line; Cells; Chromatin; Clinical; Combined Modality Therapy; Complementary DNA; DNA; DNMT3a; Dependence; Development; Disease; Dysmyelopoietic Syndromes; Enzymes; Epigenetic Process; Family; Gene Expression Profiling; Genes; Genetic; Genetic Transcription; Genomics; Goals; Hematopoiesis; Hematopoietic stem cells; Histones; Human; In Vitro; Isocitrate Dehydrogenase; Karyotype; Knockout Mice; Lead; Learning; Libraries; Lysine; Malignant - descriptor; Malignant Neoplasms; Maps; Mediating; Methylation; Mixed Function Oxygenases; Mus; Mutation; Myelogenous; Myeloproliferative disease; NF1 gene; Pathogenicity; Pathway interactions; Patients; Phenocopy; Phenotype; Play; Protein Isoforms; Proteins; Recombinants; Role; Sampling; Testing; Tumor Suppressor Proteins; Work; alpha ketoglutarate; base; cell transformation; cytokine; demethylation; epigenetic profiling; epigenomics; experimental study; functional genomics; genetic manipulation; histone demethylase; histone methylation; in vivo; innovation; insight; leukemia; leukemic transformation; metaplastic cell transformation; methylation pattern; mortality; mouse model; mutant; novel; novel therapeutic intervention; programs; stem cell function; targeted treatment; therapeutic target

ABSTRACT: KDM5 histone lysine demethylases as potential novel myeloid tumor suppressors. Mutations in Isocitrate Dehydrogenase (IDH1 and IDH2) are present in over 20% of cases of de novo normal karyotype AML and in 10-20% of cases of secondary AML that result from leukemic transformation of myelodysplastic syndrome (MDS) or myeloproliferative neoplasm (MPN). Mutant IDH transforms cells by producing R-2-hydroxyglutarate (R-2HG), an oncometabolite that can inhibit the activity of a number of cellular enzymes, including TET2, a myeloid tumor suppressor that regulates the methylation state of DNA. It is not known if R-2HG has other pathogenic targets besides TET2 in AML. However, the phenotypes of IDH mutant and TET2 mutant myeloid diseases are quite different. This observation forms the basis of the premise of our proposal, which is that inhibition of other pathways by R-2HG contributes to mutant IDH-mediated transformation. We performed an unbiased positive-selection CRISPR-Cas9 screen to identify novel tumor suppressors in AML, and we identified two histone lysine demethylases, KDM5A and KDM5C, as potential pathogenic targets of R-2HG in IDH mutant AML. Our central hypothesis is that KDM5A and KDM5C regulate hematopoietic stem cell function, and that disruption of KDM5A and KDM5C activity by R-2HG contributes to mutant IDH-mediated transformation. Our proposed studies address this hypothesis by asking three key questions. First, what is the mechanism by which KDM5 loss promotes cytokine-independent proliferation of TF-1 cells, an established factor-dependent human AML cell line? cDNA rescue experiments and genomic profiling of histone lysine methylation and transcription will be used, in conjunction with genetic manipulation of KDM5 enzymes, to elucidate the unique and shared functions of KDM5 isoforms in AML. Second, what evidence is there that KDM5 enzymes are inhibited by R-2HG in IDH mutant AML? We will profile the histone methylation state of primary IDH mutant and IDH wild-type AML patient samples, and will characterize the epigenetic and transcriptional states of isogenic cell lines that express wild-type and mutant IDH to determine if inhibition of KDM5 enzymes by R-2HG contributes to the mutant IDH-mediated transformation. And finally, how does loss of Kdm5a affect normal murine hematopoiesis and clonal hematopoiesis induced by loss of canonical myeloid tumor suppressors? We will employ conditional Kdm5a knock-out mice to perform detailed analyses of hematopoiesis in mice that lack Kdm5a alone and that lack Kdm5a in combination with Dnmt3a, Nf1 or Tet2. The answers to these questions will give us a greater understanding of the role of KDM5 demethylases in normal and malignant hematopoiesis, and in IDH mutant AML in particular. This work is conceptually innovative in that it will establish a novel epigenetic mechanism that contributes to AML, and is significant in that it has the potential to lead to novel therapeutic approaches to treat patients with leukemia.

摘要：KDM5组蛋白赖氨酸脱甲基酶是潜在的新型髓样肿瘤抑制剂。 在20％的从头正常病例中，异位酸脱氢酶（IDH1和IDH2）的突变存在 核型AML和10-20％的继发性AML病例是由白血病转化导致的 骨髓增生综合征（MDS）或骨髓增生性肿瘤（MPN）。突变IDH通过 产生R-2-羟基戊二酸（R-2HG），这是一种可以抑制许多细胞活性的oncometabolite 酶，包括TET2，一种调节DNA甲基化状态的髓样肿瘤抑制剂。它不是 知道除了AML中的TET2以外，R-2HG是否具有其他致病靶标。但是，IDH突变体的表型 和TET2突变髓样疾病完全不同。这种观察构成了我们前提的基础 提案，即通过R-2HG抑制其他途径会导致突变体IDH介导 转型。 我们进行了公正的正选择CRISPR-CAS9屏幕，以识别新的肿瘤抑制剂 AML，我们确定了两个组蛋白赖氨酸脱甲基酶KDM5A和KDM5C，为潜在的致病靶标 IDH突变体AML中的R-2HG。我们的中心假设是KDM5A和KDM5C调节造血茎 细胞功能，R-2HG对KDM5A和KDM5C活性的破坏有助于突变体IDH介导 转型。我们提出的研究通过提出三个关键问题来解决这一假设。首先，什么是 KDM5损失促进TF-1细胞的细胞因子非依赖性增殖的机制 建立的因子依赖性人AML细胞系？ cDNA救援实验和基因组分析 组蛋白赖氨酸甲基化和转录将与KDM5的遗传操纵一起使用 酶，以阐明AML中KDM5同工型的唯一和共享功能。第二，什么证据是 在IDH突变体AML中，R-2HG抑制KDM5酶？我们将介绍组蛋白 主要IDH突变体和IDH野生型AML患者样品的甲基化状态，并将表征 表达野生型和突变体IDH的等生细胞系的表观遗传和转录状态，以确定是否是否 通过R-2HG抑制KDM5酶有助于突变体IDH介导的转化。最后， KDM5A的丧失如何影响正常的鼠造血和克隆造血作用。 失去了骨髓肿瘤的损失？我们将使用有条件的KDM5A敲除小鼠 对仅缺乏KDM5A的小鼠进行造血的详细分析，并且缺乏KDM5A组合 使用DNMT3A，NF1或TET2。 这些问题的答案将使我们对KDM5脱甲基酶在 正常和恶性造血，尤其是在IDH突变体中。这项工作在概念上是 创新的是，它将建立一种新型的表观遗传机制，从而有助于AML，并且在 它有可能导致新型治疗方法治疗白血病患者。