The role of ATRX mutation in the epigenetic dysregulation of cell cycle in pediatric high-grade glioma

ATRX 突变在儿童高级别胶质瘤细胞周期表观遗传失调中的作用

• 批准号: 10432082
• 负责人: Carl J Koschmann
• 金额: $ 39万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10432082
• 关键词: ATRX gene; Automobile Driving; Binding; Brain; Brain Stem; CHEK1 gene; CHEK2 gene; Cell Cycle; Cell Cycle Checkpoint; Cell Cycle Progression; Cell Cycle Regulation; Cells; Childhood; Childhood Glioma; DNA; DNA Damage; DNA Repair; DNA replication fork; Data; Dependence; Deposition; Development; Diffuse intrinsic pontine glioma; Enhancers; Epigenetic Process; Functional disorder; Genetic; Genetically Engineered Mouse; Glioma; H3 K27M mutation; Histone H3; Histones; Human; Impairment; Knowledge; Maintenance; Malignant Childhood Neoplasm; Modeling; Mus; Mutate; Mutation; Outcome; Pathway interactions; Patients; Pharmacology; Phase; Phenotype; Pre-Clinical Model; Proteins; Psychological reinforcement; Radiation Tolerance; Radiation therapy; Radiation-Sensitizing Agents; Radiosensitization; Regulator Genes; Research; Role; Science; Site; Subgroup; Survival Rate; Tissues; Transcriptional Regulation; Translating; Untranslated RNA; Variant; base; chromatin remodeling; embryonic stem cell; experimental study; gain of function mutation; genomic locus; inhibitor; irradiation; knock-down; loss of function mutation; mouse model; mutant; new therapeutic target; novel; overexpression; precursor cell; promoter; targeted treatment

PROJECT SUMMARY / ABSTRACT Background and long-term objectives: Pediatric high-grade glioma (pHGG) is among the most lethal pediatric cancers, and new targeted therapies are desperately needed. Approved therapies for pHGG remain non- targeted and 2-year survival rates are less than 20%. Loss of function mutations in the chromatin remodeling protein ATRX are found in 30% of pHGG and DIPG, usually with concurrent mutation in the histone variant H3F3A (H3.3). We previously developed a mouse model of ATRX-deficient GBM and showed that loss of ATRX results in increased sensitivity to radiation treatment. We recently discovered that HGG cells with isogenic ATRX loss demonstrate inappropriate release of G1/S and G2/M checkpoint after irradiation and radio-sensitization with inhibitors of the master cell cycle regulator ATM. However, the mechanism driving this phenotype has not been established, and no models utilizing a background of pHGG mutations (e.g. H3.3) have been employed to study ATRX loss. Thus, there is a critical need to determine how ATRX loss deregulates cell cycle checkpoints, and to clarify the impact of concurrent H3F3A mutation on cell cycle regulation and radiation sensitizing therapy. In the absence of such knowledge, the ability to translate therapies targeted to the cell cycle checkpoint deficit in ATRX-deficient pHGG will remain unlikely. Our overall objective in this proposal is to determine the epigenetic mechanism of cell cycle dysfunction in ATRX mutated-pHGG and the impact/targetability of concurrent H3F3A mutation. Our central hypothesis is that ATRX mutation in pHGG results in reduced H3.3-promotor binding and expression of the cell cycle checkpoint regulator Checkpoint Kinase 1 (CHK1), leading to permissive cell cycle checkpoints after DNA damage. We propose that co-occurrence of H3K27M mutation will enhance this deficit and increase radio-sensitization with ATM inhibition. This is based on our preliminary data demonstrating (i) ATRX/H3.3 deposition at CHEK1 promoter sites, (ii) reduction in Chk1 expression and checkpoint maintenance after irradiation in ATRX deficient models, and (iii) increased cell cycle release with ATM inhibition in H3K27M cells compared to controls. Specific Aim 1: Determine the mechanism of cell-cycle phase dysfunction in ATRX-deficient pHGG. We will accomplish this by integrating complementary experimental approaches of multiple human and mouse pre- clinical models of ATRX loss in pHGG, including epigenetic, cell cycle and DNA-damage repair experiments. Specific Aim 2: Determine the impact of co-occurring H3F3A mutation on the targetability of ATRX- deficient pHGG. We will accomplish this Aim by integrating multiple human and mouse pre-clinical models of ATRX loss in pHGG, including a novel genetically engineered mouse model with isogenic control of H3F3A and ATRX, to isolate contribution of each driver on cell-cycle deficit and targetability. Our integrative experimental approach will establish the mechanism behind the phenotypes we have recently discovered and open new windows for therapies targeted to the unique features of ATRX-deficient pHGG.

项目摘要/摘要 背景和长期目标：儿童高级别胶质瘤(PHGG)是儿童最致命的肿瘤之一。 癌症和新的靶向治疗是迫切需要的。已批准的PHGG治疗方法仍然没有 靶向存活率和2年存活率均低于20%。染色质重塑中的功能突变丢失 在30%的PHGG和DIPG中发现ATRX蛋白，通常在组蛋白变体中同时发生突变 H3F3A(H3.3)。我们以前建立了ATRX缺陷的GBM的小鼠模型，并表明ATRX的丢失 结果增加了对放射治疗的敏感性。我们最近发现具有等基因ATRX的HGG细胞 丢失显示照射和放射增敏后G1/S和G2/M检查点的不当释放 使用主细胞周期调节器ATM的抑制剂。然而，驱动这种表型的机制并没有 已经建立，并且没有使用PHGG突变背景(例如H3.3)的模型来 研究ATRX的损失。因此，迫切需要确定ATRX丢失如何解除对细胞周期检查点的调控， 并阐明并发H3F3A突变对细胞周期调节和放射增敏治疗的影响。 在缺乏这样的知识的情况下，将针对细胞周期检查点缺陷的治疗转化为 在ATRX缺乏的情况下，PHGG仍然不太可能。 我们在这个方案中的总体目标是确定ATRX细胞周期功能障碍的表观遗传学机制 突变-PHGG和并发H3F3A突变的影响/靶向性。我们的中心假设是ATRX PHGG突变导致H3.3启动子结合减少和细胞周期检查点调节因子表达减少 Checkpoint Kinase 1(CHK1)，导致DNA损伤后允许的细胞周期检查点。我们建议 H3K27M突变的并存将加强这一缺陷，并增加ATM抑制的放射增敏作用。 这是基于我们的初步数据显示：(I)ATRX/H3.3沉积在CHEK1启动子位置，(Ii) 在ATRX缺陷模型中，照射后Chk1表达减少和检查点维护，以及(Iii) 与对照组相比，在ATM抑制下，H3K27M细胞的细胞周期释放增加。 具体目的1：确定ATRX缺陷型PHGG细胞周期时相功能障碍的机制。我们 将通过整合多个人和小鼠Pre-Pre的互补实验方法来实现这一点 PHGG中ATRX丢失的临床模型，包括表观遗传学、细胞周期和DNA损伤修复实验。 特定目标2：确定共发生的H3F3A突变对ATRX靶向性的影响 缺乏性PHGG。我们将通过整合多种人和小鼠的临床前模型来实现这一目标 PHGG中ATRX缺失，包括具有H3F3A等基因控制的新的基因工程小鼠模型 和ATRX，以分离每个驱动器对细胞周期缺陷和靶向性的贡献。 我们的综合实验方法将建立我们最近获得的表型背后的机制 发现并打开了针对ATRX缺乏PHGG的独特特征的治疗的新窗口。