Role of Asxl1 in normal hematopoiesis and pathogenesis of myeloid malignancies

Asxl1在正常造血和骨髓恶性肿瘤发病机制中的作用

• 批准号: 8628269
• 负责人: Mingjiang Xu
• 金额: $ 32.37万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-01-01 至 2014-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8628269
• 关键词: Acute Myelocytic Leukemia; Age; Apoptosis; Behavior; Biological Assay; Biology; Bone Marrow Cells; C-terminal; CD34 gene; Cell Cycle; Cell Differentiation process; Cell physiology; Cells; ChIP-seq; Characteristics; Chromatin; Chromatin Structure; Chromosome Mapping; Chromosomes; Comb animal structure; Complex; Data; Defect; Development; Dysmyelopoietic Syndromes; Enzymes; Exhibits; Frequencies; Gene Expression; Gene Expression Profiling; Gene Targeting; Genes; Genomics; Goals; Hematopoiesis; Hematopoietic; Histones; Homeostasis; Human; In Vitro; Knock-in Mouse; Knockout Mice; Lead; Maintenance; Maps; Mediating; Methylation; Methyltransferase; Mitosis; Modeling; Molecular; Molecular Target; Monoubiquitination; Mus; Mutant Strains Mice; Mutate; Mutation; Myelofibrosis; Myelogenous; Myeloid Cells; Myelopoiesis; Myeloproliferative disease; N-terminal; NF1 gene; Other Genetics; Pathogenesis; Pathologic; Patients; Phenotype; Physiological; Plants; Protein Truncation; Regulation; Reporting; Role; Stem cells; Testing; Transplantation; Tumor Suppressor Genes; Tumor Suppressor Proteins; genome-wide; histone modification; homeodomain; leukemia; leukemogenesis; loss of function; mouse model; mutant; novel; outcome forecast; public health relevance; reconstitution; self-renewal; sex; small hairpin RNA; stem

DESCRIPTION (provided by applicant): Asxl1 gene is mutated and/or deleted with high frequencies in multiple forms of myeloid malignancies including CMML, MDS, MPN and AML. The majority of the Asxl1 mutations in these patients are heterozygous, leading to nonsense/frameshift, suggesting loss of function. Mutations in ASXL1 are associated with poor prognosis. Therefore, ASXL1 has been speculated to be a putative tumor suppressor gene that is strongly implicated in the pathogenesis of myeloid malignancies. The objective of this project is to define the physiological function of ASXL1 and its role in the pathogenesis of myeloid malignancies. We generated several Asxl1-targeted murine models. Haploinsufficiency of Asxl1 (+/-) leads to the development of MDS in mice, which can progress to MDS/MPN and leukemia as they age. The Asxl1-targeted mouse model, therefore, recapitulates the pathologic situation of patients with myeloid malignancy, thus allows us to gain the needed information about Asxl1 biology in a timely fashion. Deleting Asxl1 leads to increased apoptosis and mitosis of bone marrow cells, characteristic cellular feature of MDS. A competitive reconstitution assay showed that Asxl1-/- hematopoietic stem/progenitor cells (HSC/HPCs) had a decreased hematopoietic repopulating capacity. We, therefore, hypothesize that Asxl1 acts as a tumor suppressor in myelopoiesis by altering the behavior of HSC/HPCs. We will test this hypothesis in 3 specific aims: Aim 1: To determine if Asxl1 acts as a tumor suppressor in myelopoiesis by characterizing the phenotype of various Asxl1-deficient mice, including MxCre or Vav1Cre mediated Asxl1 conditional knock-out mice. In addition, we will evaluate the role of Asxl1 haploinsufficiency in cooperation with other genetic alterations, such as Nf1 haploinsufficiency, for triggering myeloid malignancies. Aim 2: To define the cellular mechanisms by which loss of Asxl1 function in mice leads to myeloid malignancies. Specifically, we will examine the effects of Asxl1 deletion/haploinsufficiency on the proliferation, differentiation, apoptosis and cell cycle o HSC/HPCs. In addition, we will determine the effect of Asxl1 deletion/haploinsufficiency on self-renewal and differentiation potential of HSC/HPCs by serial transplantation. Aim 3: To define the molecular mechanisms by which Asxl1 regulates normal hematopoiesis and exerts its tumor suppressor function in myelopoiesis. We will identify Asxl1-target genes by mapping the genomic distribution of Asxl1 and its interacting histone modifying enzymes by ChIP-Seq in Asxl1:Tag and WT or Asxl1-/- HSC/HPCs, respectively. Furthermore, we will dissect Asxl1- deletion induced misregulation of H3 methylation and H2A monoubiquitination in HSC/HPCs with ChIP-Seq and correlate with the gene expression profiling. Accomplishment of these studies allows us to uncover the role of Asxl1 in normal hematopoiesis and in the multiple-step pathogenesis of myeloid malignancies, which may lead to the identification of novel molecular targets for the treatment of patients with myeloid malignancies.

描述（由申请人提供）：ASXL1基因被突变和/或用高频率以多种形式的髓样恶性肿瘤（包括CMML，MDS，MPN，MPN和AML）删除。这些患者中的大多数ASXL1突变都是杂合的，导致胡说八道/移封，表明功能丧失。 ASXL1中的突变与预后不良有关。因此，据推测，ASXL1是一种推定的肿瘤抑制基因，与髓样恶性肿瘤的发病机理有很大的影响。该项目的目的是定义ASXL1的生理功能及其在髓样恶性肿瘤的发病机理中的作用。我们生成了几种以ASXL1为目标的鼠模型。 ASXL1（+/-）的单倍不足导致小鼠MD的发展，随着年龄的增长，可以发展为MDS/MPN和白血病。因此，以ASXL1为目标的小鼠模型概括了髓样恶性肿瘤患者的病理状况，因此使我们能够及时获得有关ASXL1生物学所需的信息。删除ASXL1导致骨髓细胞的凋亡和有丝分裂增加，MDS的特征性细胞特征。竞争性的重建测定法显示，ASXL1 - / - 造血茎/祖细胞（HSC/HPC）的造血重塑能力降低。因此，我们假设ASXL1通过改变HSC/HPC的行为来充当骨髓病中的肿瘤抑制因子。我们将在3个特定目的中检验这一假设：目标1：确定ASXL1是否通过表征各种ASXL1缺陷小鼠的表型（包括MXCRE或VAV1CRE介导的ASXL1 ASXL1条件敲除小鼠）来确定骨髓病中的肿瘤抑制剂。此外，我们将评估ASXL1单倍不使性在与其他遗传变化（例如NF1单倍氨酸疾病）合作中的作用，以触发髓样恶性肿瘤。目标2：定义细胞机制，通过小鼠中ASXL1功能导致髓样恶性肿瘤的损失。具体而言，我们将检查ASXL1缺失/单倍宽度供应对增殖，分化，凋亡和细胞周期O HSC/HPC的影响。此外，我们将确定ASXL1删除/单倍度不足对HSC/HPC通过串行移植的自我更新和分化潜力的影响。目标3：定义ASXL1调节正常造血的分子机制，并在骨髓中发挥其肿瘤抑制功能。我们将通过映射ASXL1的基因组分布及其相互作用的组蛋白通过chip-seq在ASXL1：TAG和WT或ASXL1或ASXL1 - / - HSC/HPC中的酶修饰酶来鉴定ASXL1-TARGET基因。此外，我们将在HSC/HPC中剖析H3甲基化和H2A单晶体的不正调，并用chip-seq进行了单调性，并与基因表达谱分析相关。 这些研究的完成使我们能够揭示ASXL1在正常造血作用和髓样恶性肿瘤的多步发病机理中的作用，这可能导致鉴定出新的分子靶标，以治疗髓样恶性肿瘤患者。