Investigating the role of dysfunctional histone H3.3 in driving early neuronal development and pediatric high-grade gliomas

摘要/Abstract 小儿脑肿瘤是儿童中最常见的实体肿瘤，大约有5000例新发病例 每年在美国确诊的在0-14岁的儿童中，约17%的脑肿瘤是高级别的 神经胶质瘤（HGG），目前无法治愈。缺乏有效的治疗方法突出了迫切需要 确定基于机制的治疗方法。最近有大量的实验证据表明 携带H3.3-G34 R的儿科HGG（pHGG）表现出高基因组不稳定性和高水平表达， 的神经元标记物，表明这些肿瘤代表了一个独特的亚型pHGG相比，其他 类型，包括H3.3-K27 M突变的类型。超过90%的H3.3-G34 R胶质瘤也含有ATRX 功能丧失突变使用新建立的基因工程小鼠模型（GEMM），我们 证明了H3.3-G34 R突变和ATRX缺失在恶性前神经干细胞（PM-NSC）中， Trp 53-/-背景可强烈促进胶质瘤的发生。这些肿瘤表现出人类的典型特征， H3.3-G34 R的pHGG，因此该GEMM为我们提供了一个可靠的研究分子生物学的工具， H3.3-G34 R突变和ATRX缺失的协同作用的潜在机制以及用于鉴定 新的治疗靶点。我们发现H3.3-G34 R突变改变了组蛋白修饰， 并导致FoxD 1和HoxA 1的高表达，FoxD 1和HoxA 1是早期神经元发育所必需的转录因子。 发展鉴于FoxD 1和HoxA 1的富集与胶质瘤的预后不良相关， 患者，他们为pHGG提供了2个新的治疗靶点。此外，我们发现ATRX损失导致ALT 激活，这使得肿瘤细胞对其线粒体功能的扰动敏感。根据这些 观察，我们假设H3.3-G34 R突变和ATRX丢失诱导的独特的表观遗传特征， 驱动胶质瘤的发生，并导致涉及功能失调的端粒和受损的靶向脆弱性， 线粒体活性为了验证这一假设，我们计划1）确定FoxD 1和HoxA 1在H3.3中的作用- G34 R驱动的胶质瘤发生，2）定义由pHGG中ATRX缺陷诱导的治疗脆弱性，以及 3)阐明H3.3-G34 R突变和ATRX缺失对表观遗传重编程的协同作用， 神经胶质瘤完成拟议的研究不仅将填补我们对H3.3-G34 R如何 和ATRX损失改变表观基因组，导致正常的神经元发育和胶质瘤形成，但也- 更重要的是，有助于开发靶向pHGG的治疗策略， 深入了解表观遗传调控在大脑发育和胶质瘤形成中的作用。

Summary Statement/Abstract Pediatric brain tumors are the most common solid tumors in children, with approximately 5000 new cases diagnosed per year in the United States. Around 17% of brain tumors in children age 0–14 years are high-grade gliomas (HGGs), which are currently incurable. The lack of effective treatments highlights the urgent need to identify mechanism-based therapeutic approaches. Substantial experimental evidence has recently revealed that H3.3-G34R–harboring pediatric HGGs (pHGGs) exhibit high genomic instability and high-level expression of neuronal markers, indicating that these tumors represent a distinct subtype of pHGG compared with other types, including the one with an H3.3-K27M mutation. More than 90% of H3.3-G34R gliomas also harbor ATRX loss-of-function mutations. Using a newly established genetically engineered murine model (GEMM), we demonstrated that H3.3-G34R mutation and ATRX deletion in premalignant neural stem cells (PM-NSCs) with the Trp53-/- background could strongly promote gliomagenesis. These tumors exhibit typical features of human H3.3-G34R–harboring pHGGs, so this GEMM provides us with a faithful tool for studying the molecular mechanisms underlying the synergistic effects of H3.3-G34R mutation and ATRX deletion and for identifying novel therapeutic targets. We have found that H3.3-G34R mutation changes histone modifications both locally and globally and leads to high expression of FoxD1 and HoxA1, transcription factors essential for early neuronal development. Given that enrichments of FoxD1 and HoxA1 are associated with worse prognosis in glioma patients, they provide 2 novel therapeutic targets for pHGGs. In addition, we found that ATRX loss leads to ALT activation, which makes tumor cells sensitive to perturbation of their mitochondrial function. On the basis of these observations, we hypothesize that distinctive epigenetic profiles induced by H3.3-G34R mutation and ATRX loss drive gliomagenesis and lead to targetable vulnerabilities involving dysfunctional telomeres and impaired mitochondrial activity. To test this hypothesis, we plan to 1) determine the roles of FoxD1 and HoxA1 in H3.3- G34R–driven gliomagenesis, 2) define the therapeutic vulnerability induced by ATRX deficiency in pHGGs, and 3) elucidate the synergistic effect of H3.3-G34R mutation and ATRX loss on epigenetic reprogramming in gliomas. The completion of the proposed studies will not only fill the gaps in our knowledge of how H3.3-G34R and ATRX loss change the epigenome to lead to normal neuronal development and gliomagenesis, but also— and more importantly—contribute to the development of therapeutic strategies that target pHGGs and provide insights into the role of epigenetic regulation in brain development and gliomagenesis.