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

基本信息

批准号：
10443596
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
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-07-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10443596
关键词：
Acute Myelocytic Leukemia Address Affect Biochemical 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 Secondary acute myeloid leukemia 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

项目摘要

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 组蛋白赖氨酸去甲基酶作为潜在的新型骨髓肿瘤抑制剂。超过 20% 的新发正常病例中存在异柠檬酸脱氢酶（IDH1 和 IDH2）突变核型 AML 以及 10-20% 的继发性 AML 病例由白血病转化引起骨髓增生异常综合征（MDS）或骨髓增生性肿瘤（MPN）。突变的 IDH 通过以下方式改变细胞产生 R-2-羟基戊二酸 (R-2HG)，这是一种致癌代谢物，可以抑制多种细胞的活性酶，包括 TET2，一种调节 DNA 甲基化状态的骨髓肿瘤抑制因子。它不是已知 R-2HG 在 AML 中是否还有除 TET2 之外的其他致病靶点。然而，IDH突变体的表型和TET2突变的骨髓疾病有很大不同。这一观察构成了我们的前提的基础提议，即 R-2HG 对其他途径的抑制有助于突变 IDH 介导转变。我们进行了公正的正选 CRISPR-Cas9 筛选，以鉴定新型肿瘤抑制因子 AML，我们确定了两种组蛋白赖氨酸去甲基酶 KDM5A 和 KDM5C 作为潜在的致病靶点 IDH 突变 AML 中的 R-2HG。我们的中心假设是 KDM5A 和 KDM5C 调节造血干 R-2HG 对 KDM5A 和 KDM5C 活性的破坏有助于突变 IDH 介导转变。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 缺失促进 TF-1 细胞的细胞因子非依赖性增殖的机制建立因子依赖性人类 AML 细胞系？ cDNA拯救实验和基因组分析组蛋白赖氨酸甲基化和转录将与 KDM5 的遗传操作结合使用酶，阐明 KDM5 亚型在 AML 中的独特和共享功能。二、什么是证据在 IDH 突变型 AML 中，R-2HG 会抑制 KDM5 酶吗？我们将分析组蛋白原代 IDH 突变体和 IDH 野生型 AML 患者样本的甲基化状态，并将表征表达野生型和突变型 IDH 的同基因细胞系的表观遗传和转录状态，以确定是否 R-2HG 对 KDM5 酶的抑制有助于突变 IDH 介导的转化。最后， Kdm5a 缺失如何影响正常小鼠造血和由 Kdm5a 诱导的克隆造血经典骨髓肿瘤抑制因子的丧失？我们将使用条件性 Kdm5a 敲除小鼠对单独缺乏 Kdm5a 和联合缺乏 Kdm5a 的小鼠的造血功能进行详细分析与 Dnmt3a、Nf1 或 Tet2 一起使用。这些问题的答案将使我们更好地了解 KDM5 去甲基酶在正常和恶性造血，特别是 IDH 突变 AML。这项工作在概念上创新之处在于它将建立一种有助于 AML 的新型表观遗传机制，并且在它有可能带来治疗白血病患者的新治疗方法。