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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

基本信息

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
项目类别：
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

项目摘要

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.

ASXL基因是果蝇Asx基因的同源基因，基因表达的重要调节因子。ASXL 1突变以高频率发生在多种形式的髓系恶性肿瘤患者预后差。ASXL 1突变大多是无义/移码，导致缺乏C-末端PHD指的蛋白质截短，这在ASXL 1患者样本中可检测到突变。我们最近发现，小鼠中ASXL 1截短体（Asxl 1 Y 588 XTg）的转基因表达导致HSC/HPC池增加和骨髓恶性肿瘤发病机制（Blood 2017）。BAP 1是在polycomb蛋白介导的基因抑制过程中被ASXL 1激活去泛素化H2 AK 119。BAP1 突变也发生在MDS患者中。尽管ASXL 1截短突变对细胞增殖有显著影响，骨髓恶性肿瘤的发病机制，基本机制仍在很大程度上未知，阻碍了开发有效的靶向治疗方法。我们的蛋白质组学研究发现ASXL 1aa 1 -587表现出与ASXL 1FL相比，与BAP 1的结合亲和力增加，并获得与BRD 4的相互作用，BRD 4是 BET家族BRD 4参与多种生物过程，包括转录，DNA复制，表观遗传调节和肿瘤发生。我们假设截短的ASXL 1使HSC/HPC失调，通过改变BAP 1去泛素化酶复合物的功能，与BRD 4的互动解决这一关键问题具有重大的翻译影响，也是这个应用程序。在目标1中，我们将使用我们最近产生的Tet-on/off Asxl 1 Y 588 XTg小鼠来评估证明- 通过撤回强力霉素沉默转基因表达是否可以根除ASXL 1aa 1 -587- 介导的异常造血表型和骨髓恶性肿瘤。我们将应用bPPI-seq，一种新的在体内生理条件下确认/鉴定真正ASXL 1aa 1 -587相互作用物的灵敏试验。在目标2中，我们将确定BRD 4-ASXL 1aa 1 -587相互作用在截短的ASXL 1介导的HSC/HPC中的作用，失调和骨髓恶性肿瘤的发展。我们将检查BRD 4抑制剂（EP 11313）治疗能够预防和/或挽救异常的造血表型和骨髓恶性肿瘤。在目标3中，我们将确定BAP 1在截短的ASXL 1介导的细胞凋亡中的作用。异常HSC/HPC行为和体内髓系恶性肿瘤的发病机制。我们将破解 ASXL 1aa 1 -587在体内改变HSC/HPC中BAP 1的功能。ASXL 1aa 1 -587对全基因组的影响 LK细胞中的BAP 1和H2 AK 119 Ub占有率将通过ChIP-seq测定，并与基因表达数据。这些研究是及时的，对于推进我们对 ASXL 1截短突变介导的HSC/HPC失调和髓系恶性肿瘤的发展的该项目的成功将有可能在ASXL 1的获得性相互作用物中确定新的治疗靶点在一些实施方案中，本发明涉及用于治疗具有ASXL 1截短突变的骨髓恶性肿瘤的BRD 4和BAP 1。