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: 10321955
• 负责人: Mingjiang Xu
• 金额: --
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-01-02 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10321955
• 关键词: Affinity; BRD2 gene; Behavior; Binding; Biological; Biological Assay; Biological Process; Blood; Bromodomain; C-terminal; Cells; ChIP-seq; Chromatin; Comb animal structure; Complex; DNA biosynthesis; Data; Development; Doxycycline; Drosophila genus; Dysmyelopoietic Syndromes; Event; Exhibits; Family; Family member; Frequencies; Gene Expression; Genes; Genetic Transcription; Goals; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; Homologous Gene; Human; Impairment; Knowledge; Lead; Lysine; Malignant Neoplasms; Mediating; Molecular; Mus; Mutation; Myeloproliferative disease; Oncogenic; PHD Finger; Pathogenesis; Pathologic; Patients; Phenotype; Physiological; Polycomb; Prognosis; Protein Truncation; Proteins; Proteomics; Regulation; Regulator Genes; Reporting; Role; Sampling; Tetanus Helper Peptide; Therapeutic; Time; Transgenic Organisms; Xenograft procedure; anticancer activity; epigenetic regulation; gene repression; genome-wide; in vivo; inhibitor; member; mouse model; new therapeutic target; novel; operation; prevent; protein complex; sex; success; targeted treatment; therapeutic target; therapeutically effective; transgene expression; translational impact; transplant model; tumorigenesis; ubiquitinated H2A

Additional sex comb-like (ASXL) genes are human homologues of Drosophila-Asx gene which encode important regulators of gene expression. ASXL1 mutations occur at high frequencies in multiple forms of myeloid malignancy patients with poor prognosis. ASXL1 mutations are mostly nonsense/frameshift, causing truncation of the protein lacking the C-terminal PHD finger, which is detectable in patient samples with ASXL1 mutations. We have recently shown that transgenic expression of an ASXL1 truncation in mice (Asxl1Y588XTg) results in increased HSC/HPC pools and pathogenesis of myeloid malignancies (Blood 2017). BAP1 is activated by ASXL1 to deubiquitinate H2AK119 during polycomb protein-mediated gene repression. BAP1 mutations also occur in patients with MDS. Despite the significant impact of ASXL1 truncation mutations on the pathogenesis of myeloid malignancies, the underlying mechanisms remain largely unknown, hindering the development of effective targeted therapeutics. Our proteomics studies discovered that ASXL1aa1-587 exhibits an increased binding affinity to BAP1 compared to ASXL1FL and gains an interaction with BRD4, a member of the BET family. BRD4 is involved in multiple biologic processes, including transcription, DNA replication, epigenetic regulation, and tumorigenesis. We hypothesize that truncated ASXL1 dysregulates HSC/HPCs and causes myeloid malignancies through altering the function of the BAP1 deubiquitinase complex and gaining interaction with BRD4. Challenging this critical question has great translational impact and is the major goal of this application. In Aim 1, we will use our recently generated Tet-on/off Asxl1Y588XTg mice to assess in prove- of-concept whether silencing transgene expression by withdrawing doxycycline can eradicate the ASXL1aa1-587- mediated abnormal hematopoietic phenotype and myeloid malignancies. We will apply bPPI-seq, a novel sensitive assay to confirm/identify the true ASXL1aa1-587 interactors at physiological conditions in vivo. In Aim 2, we will determine the role of BRD4-ASXL1aa1-587 interaction in truncated ASXL1-mediated HSC/HPC dysregulation and myeloid malignancy development. We will examine whether BRD4 inhibitor (EP11313) treatment is capable of preventing and/or rescuing the abnormal hematopoietic phenotype and myeloid malignancies in Asxl1Y588XTg mice. In Aim 3, we will determine the role of BAP1 in truncated ASXL1-mediated abnormal HSC/HPC behavior and pathogenesis of myeloid malignancies in vivo. We will decipher how ASXL1aa1-587 alters the function of BAP1 in HSC/HPCs in vivo. The effects of ASXL1aa1-587 on genome-wide BAP1 and H2AK119Ub occupancy in LK cells will be determined by ChIP-seq and correlated with the gene expression data. These studies are timely and fundamentally important for advancing our knowledge on ASXL1 truncation mutation-mediated HSC/HPC dysregulation and myeloid malignancy development. The success of this project will likely identify novel therapeutic targets in the gained-interactors for ASXL1 truncation, BRD4 and BAP1, for the treatment of myeloid malignancies with ASXL1 truncation mutations.

附加性梳状(ASXL)基因是编码果蝇-ASX基因的人类同源基因 基因表达的重要调节因子。ASXL1突变在多种形式的 髓系恶性肿瘤患者预后较差。ASXL1突变大多是无稽之谈/移码，导致 在ASXL1患者样本中可检测到的缺失C-末端PHD手指的蛋白质的截断 突变。我们最近在小鼠(Asxl1Y588XTg)中发现了ASXL1截断的转基因表达。 结果增加了HSC/HPC池和髓系恶性肿瘤的发病机制(血液2017)。BAP1是 在多梳蛋白介导的基因抑制过程中，被ASXL1激活以去泛素化H_2AK119。BAP1 MDS患者也会发生突变。尽管ASXL1截断突变对 髓系恶性肿瘤的发病机制仍很大程度上尚不清楚，阻碍了 开发有效的靶向疗法。我们的蛋白质组学研究发现ASXL1aa1-587展示了 与ASXL1FL相比，与BAP1的结合亲和力增强，并获得与BRD4的相互作用 贝特家族。BRD4参与多种生物过程，包括转录、DNA复制、 表观遗传调控和肿瘤发生。我们假设截短的ASXL1失调了HSC/HPC和 通过改变BAP1脱泛素酶复合体的功能并获得 与BRD4的交互。挑战这一关键问题具有巨大的翻译影响，也是 这个应用程序。在目标1中，我们将使用我们最近开发的Tet-On/Off Asxl1Y588XTg小鼠来评估在证明- 停用多西环素沉默转基因表达能否根除ASXL1aa1-587 介导的异常造血表型和髓系恶性肿瘤。我们将应用bppi-seq，一部小说 在体内生理条件下确认/鉴定真实ASXL1aa1-587相互作用的灵敏方法。在目标2中， 我们将确定BRD4-ASXL1aa1-587相互作用在截短ASXL1介导的HSC/HPC中的作用 调节失调和髓系恶性肿瘤的发展。我们将检查BRD4抑制剂(EP11313) 治疗能够预防和/或挽救异常的造血表型和髓系 Asxl1Y588XTg小鼠的恶性肿瘤。在目标3中，我们将确定BAP1在截短ASXL1介导的 HSC/HPC异常行为与体内髓系恶性肿瘤的发病机制我们将破译如何 ASXL1aa1-587可在体内改变BAP1在HSC/HPC中的功能。ASXL1aa1-587对全基因组的影响 BAP1和H2AK119Ub在LK细胞中的占有率将由CHIP-SEQ确定并与基因相关 表达式数据。这些研究对于增进我们对……的认识具有及时和根本的重要性。 ASXL1截短突变介导的HSC/HPC失调与髓系恶性肿瘤的发生发展这个 该项目的成功将可能在ASXL1获得的相互作用中确定新的治疗靶点 截断，BRD4和BAP1，用于治疗带有ASXL1截断突变的髓系恶性肿瘤。