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：
10543761
负责人：
Mingjiang Xu
金额：
$ 23.3万
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/10543761
关键词：
Acetylation Affinity BRD2 gene Behavior Binding Biological Biological Assay Biological Process Blood Bromodomain C-terminal Cells ChIP-seq Chromatin 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 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 Therapeutic Xenograft procedure anticancer activity cohesin epigenetic regulation gene repression genome-wide in vivo inhibitor member mouse model new therapeutic target novel prevent protein complex sex success targeted treatment therapeutic target therapeutically effective transgene expression translational impact transplant model tumorigenesis ubiquitin isopeptidase

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.

其他性梳状（ASXL）基因是果蝇-ASX基因的人类同源物基因表达的重要调节剂。 ASXL1突变以多种形式的高频出现预后不良的髓样恶性肿瘤患者。 ASXL1突变大多是胡说八道/移架，导致缺乏C末端PHD手指的蛋白质的截断，该手指在ASXL1的患者样品中可检测到突变。我们最近表明，小鼠ASXL1截断的转基因表达（ASXL1Y588XTG）导致HSC/HPC池增加和髓样恶性肿瘤的发病机理（血液2017）。 Bap1是在多孔蛋白介导的基因抑制过程中，被ASXL1激活以去泛素化H2AK119。 BAP1 MDS患者也会发生突变。尽管ASXL1截断突变对髓样恶性肿瘤的发病机理，基本机制在很大程度上未知，阻碍了开发有效的靶向治疗学。我们的蛋白质组学研究发现ASXL1AA1-587展示与ASXL1FL相比，与BAP1的结合亲和力增加，并获得了与BRD4的相互作用， BET家族。 BRD4参与了多种生物过程，包括转录，DNA复制，表观遗传调节和肿瘤发生。我们假设截断的ASXL1失调HSC/HPC和通过改变BAP1去泛素酶复合物的功能并获得的功能，从而导致髓样恶性肿瘤与BRD4的相互作用。挑战这个关键问题具有巨大的翻译影响，是此应用程序。在AIM 1中，我们将使用我们最近生成的Tet-On/Off ASXL1Y588XTG小鼠进行评估。概念是否通过撤回强力霉素来使沉默的转基因表达能否消除ASXL1AA1-587- 介导的异常造血表型和髓样恶性肿瘤。我们将应用Bppi-Seq，一部小说敏感测定法确认/识别体内生理条件下的真实ASXL1AA1-587相互作用。在AIM 2中，我们将确定BRD4-ASXL1AA1-587相互作用在截短的ASXL1介导的HSC/HPC中的作用失调和髓样恶性肿瘤的发展。我们将检查BRD4抑制剂（EP11313）是否是否治疗能够预防和/或营救异常的造血表型和髓样 ASXL1Y588XTG小鼠的恶性肿瘤。在AIM 3中，我们将确定BAP1在截短的ASXL1介导的作用体内髓样恶性肿瘤的异常HSC/HPC行为和发病机理。我们将解密如何 ASXL1AA1-587改变了BAP1在体内HSC/HPC中的功能。 ASXL1AA1-587对全基因组的影响 LK细胞中的BAP1和H2AK119UB占用率将由芯片序列确定，并与基因相关表达数据。这些研究对于促进我们的知识至关重要 ASXL1截断突变介导的HSC/HPC失调和髓样恶性肿瘤的发展。这该项目的成功可能会确定ASXL1获得的相互作用的新型治疗靶标截断，BRD4和BAP1，用于用ASXL1截断突变处理髓样恶性肿瘤。