No-nonsense approach to treat myeloid malignancies with ASXL1 nonsense mutations

治疗具有 ASXL1 无义突变的骨髓恶性肿瘤的严肃方法

• 批准号: 8830950
• 负责人: Mingjiang Xu
• 金额: $ 20.03万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-03 至 2016-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8830950
• 关键词: Acute Myelocytic Leukemia; Alleles; Amino Acids; Aminoglycosides; Animal Model; Attenuated; C-terminal; Cell physiology; Cells; Cessation of life; Chromosomes; Clinical Trials; Comb animal structure; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Data; Development; Disease; Dysmyelopoietic Syndromes; Dystrophin; Frequencies; Generations; Genes; Gentamicins; Goals; Health; Hematologic Neoplasms; Hematopoietic; Histone H3; Human; In Vitro; Intellectual functioning disability; Length; Location; Maps; Mediating; Methylation; Monitor; Mus; Muscular Dystrophies; Mutate; Mutation; Myelofibrosis; Myelogenous; Myeloproliferative disease; N-terminal; Nonsense Codon; Nonsense Mutation; Opitz syndrome; Pathogenesis; Patients; Peptides; Phenotype; Placebos; Plants; Population; Protein Truncation; Proteins; Reading; Role; Sampling; Specimen; Stem cells; Terminator Codon; Testing; Therapeutic; Translations; base; homeodomain; in vivo; innovation; leukemia; loss of function; malformation; mouse model; novel; outcome forecast; overexpression; premature; prevent; protein purification; reading ability; sex; stem; success; transgene expression; treatment strategy

DESCRIPTION (provided by applicant): Read-through compounds (RTCs) are capable of promoting ribosomal read-through of premature termination codons (PTCs), resulting in substitution of the PTC with an amino acid and generation of full-length protein product. The RTCs include aminoglycosides (such as gentamicin) and non-aminoglycoside (such as ataluren and RTC13) compounds, which are specific for PTCs and do not interfere with normal stop codons. These RTCs have been shown to have read-through activity on nonsense mutations in the CFTR and the dystrophin genes in both cultures and animal models. Intriguingly, clinical trials in patients with nonsense mutation muscular dystrophy and cystic fibrosis showed that RTCs induced some expression of functional, previously missing, proteins in both diseases and had some therapeutic benefits. Additional sex comb-like 1 (ASXL1) is mutated at high frequencies in multiple forms of myeloid malignancies, including MDS, MPN, CMML, and AML. De novo ASXL1 mutations cause Bohring-Opitz syndrome. Mutations in ASXL1 are nonsense/frameshift, leading to loss-of-function by truncating the critical PHD domain. Mutations in ASXL1 are associated with poor prognosis. Approximately 1/3 of the myeloid malignancy cases with ASXL1 mutations are nonsense, which are appealing targets for PTC-based read- through. In this application, we will explore the hypothesis that RTCs are capable of reversing ASXL1-PTCs and induce adequate amount of full-length, functional ASXL1 expression in hematopoietic stem/progenitor cells (HSCs/HSPCs), therefore attenuating the ASXL1-deficiency/haploinsufficiency mediated abnormal HSC/HPC function and hematological phenotype. Three specific aims are proposed to test this hypothesis. Aim 1: To determine the ability of RTCs (gentamicin, ataluren and RTC13) to reverse Asxl1-PTCs and yield adequate amount of full-length, functional Asxl1 expression in various hematopoietic cell populations from Asxl1-PTCTg;MxCre mice. Aim 2: To assess the therapeutic potential of RTCs by examining if RTC treatment is capable of preventing or curing the Asxl1-deficiency mediated pathogenesis of myeloid malignancies in Asxl1f/f;MxCre;Asxl1-PTCTg mice (inducible Asxl1-inactivation with simultaneous Asxl1-PTC transgene expression). Aim 3: To validate the therapeutic potential of RTCs using primary cells from myeloid malignancy patients with ASXL1 nonsense mutations. We will determine the ability of RTCs in promoting ribosomal read- through of different ASXL1-PTCs (TGA, TAA, TAG) in various hematopoietic cell populations from myeloid malignancy patients with ASXL1 nonsense mutations. We will examine if RTC treatment is capable of correcting/attenuating the abnormal cellular function of myeloid malignancy HSC/HPCs with ASXL1 nonsense mutations in NSG mice. Success of these studies will advance this novel treatment strategy into not only ASXL1-PTC diseases, but also many hematological malignancies due to nonsense mutation-mediated early termination of translation.

描述（由申请人提供）：通读化合物（RTC）能够促进核糖体通读过早终止密码子（PTC），导致PTC被氨基酸取代并产生全长蛋白质产物。 RTC 包括氨基糖苷类（如庆大霉素）和非氨基糖苷类（如 ataluren 和 RTC13）化合物，它们对 PTC 具有特异性，不会干扰正常终止密码子。这些 RTC 已被证明对培养物和动物模型中 CFTR 和肌营养不良蛋白基因中的无义突变具有通读活性。有趣的是，对无义突变型肌营养不良症和囊性纤维化患者进行的临床试验表明，RTC 可以诱导两种疾病中先前缺失的功能性蛋白质的表达，并具有一定的治疗效果。 额外性梳状蛋白 1 (ASXL1) 在多种形式的骨髓恶性肿瘤中发生高频率突变，包括 MDS、MPN、CMML 和 AML。 ASXL1 新突变会导致 Bohring-Opitz 综合征。 ASXL1 中的突变是无义/移码，通过截断关键的 PHD 结构域而导致功能丧失。 ASXL1 突变与不良预后相关。大约 1/3 的 ASXL1 突变骨髓恶性肿瘤病例是无意义的，这是基于 PTC 的通读的有吸引力的目标。在此应用中，我们将探讨 RTC 能够逆转 ASXL1-PTC 并在造血干细胞/祖细胞 (HSC/HSPC) 中诱导足够量的全长功能性 ASXL1 表达的假设，从而减弱 ASXL1 缺陷/单倍体不足介导的异常 HSC/HPC 功能和血液学表型。提出了三个具体目标来检验这一假设。 目标 1：确定 RTC（庆大霉素、ataluren 和 RTC13）逆转 Asxl1-PTC 并在 Asxl1-PTCTg;MxCre 小鼠的各种造血细胞群中产生足够量的全长、功能性 Asxl1 表达的能力。目标 2：通过检查 RTC 治疗是否能够预防或治愈 Asxl1f/f;MxCre;Asxl1-PTCTg 小鼠（诱导 Asxl1 失活并同时 Asxl1-PTC 转基因表达）中 Asxl1 缺陷介导的骨髓恶性肿瘤发病机制来评估 RTC 的治疗潜力。目标 3：使用来自携带 ASXL1 无义突变的骨髓恶性肿瘤患者的原代细胞来验证 RTC 的治疗潜力。我们将确定 RTC 在促进来自具有 ASXL1 无义突变的骨髓恶性肿瘤患者的各种造血细胞群中不同 ASXL1-PTC（TGA、TAA、TAG）核糖体读取的能力。我们将检查 RTC 治疗是否能够纠正/减弱 NSG 小鼠中带有 ASXL1 无义突变的骨髓恶性肿瘤 HSC/HPC 的异常细胞功能。 这些研究的成功将推动这种新的治疗策略不仅适用于 ASXL1-PTC 疾病，而且适用于许多由于无义突变介导的翻译早期终止而导致的血液恶性肿瘤。