The role of a histone point mutation in pediatric gliomagenesis

组蛋白点突变在儿童胶质瘤发生中的作用

• 批准号: 10207450
• 负责人: Kasey R Skinner
• 金额: $ 4.04万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-29 至 2023-06-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10207450
• 关键词: ATRX gene; Affect; Aggressive course; Allografting; Amino Acids; Biochemical; Biochemistry; Bioinformatics; Biological; Biological Assay; Brain; Brain Stem; Cancer Biology; Cancer Model; Cell Line; Cells; ChIP-seq; Childhood; Childhood Glioma; Clinic; Clinical; Communication; Critical Thinking; Data; Data Set; Deposition; Development; Diffuse; Disease; Dominant-Negative Mutation; Enterobacteria phage P1 Cre recombinase; Environment; Epigenetic Process; Ethics; Genes; Genetic Transcription; Genetically Engineered Mouse; Genome; Genomics; Glioma; Gliomagenesis; Goals; Harvest; Heterogeneity; Histone H3; Histones; Human; In Vitro; Invaded; Knowledge; Lead; Location; Lysine; Malignant - descriptor; Malignant neoplasm of brain; Methionine; Modeling; Modernization; Modification; Molecular; Mus; Mutate; Mutation; Neurosciences; Oncogenes; Oncogenic; PDGFRA gene; Pathogenesis; Patients; Pediatric Neoplasm; Phenotype; Platelet-Derived Growth Factor; Point Mutation; Primary Brain Neoplasms; Prognosis; Publishing; Recurrence; Research; Research Proposals; Role; Sampling; Signal Transduction; Somatic Mutation; Structure; System; TP53 gene; Technical Expertise; Testing; Training; Translating; Tumor Suppressor Proteins; Variant; Virus; Western Blotting; base; brain cell; chromatin remodeling; druggable target; effective therapy; epigenetic therapy; epigenomics; gene repression; histone modification; human disease; immunocytochemistry; live cell imaging; mouse model; mutant; mutational status; neoplastic cell; nerve stem cell; nestin protein; overexpression; patient derived xenograft model; self-renewal; skills; stem cells; stemness; therapeutic target; transcriptome; transcriptome sequencing; transcriptomics; tumor; tumorigenesis

PROJECT SUMMARY Pediatric high-grade gliomas (pHGG) are the most common and malignant primary brain tumors in children. Mutations in H3F3A, one of the genes that encodes histone variant H3.3, are definitional for pHGG and H3.3 mutation status correlates with clinical variables such as survival and tumor location. pHGG with a lysine 27 to methionine substitution (H3.3K27M) occur in the brain stem and correlate with particularly poor survival. H3.3K27M is insufficient to drive tumorigenesis in mouse models. However, it co-occurs in humans with PDGFRA activation and TP53 and ATRX deletion, suggesting that H3.3K27M cooperates with these oncogenic mutations to induce tumors. Further, the phenotypic differences between H3.3K27M pHGG and other pHGG suggest differing epigenetic alterations and mechanisms of tumorigenesis. This proposal is focused on identifying the epigenetic and transcriptomic changes induced in murine neural stem cells (NSC) by H3.3K27M and its co-occurring mutations and how those mutations lead to glioma cell phenotypes. To model H3.3K27M pHGG as accurately as possible, I will use an RCAS-TVA-based model in which NSC harvested from Nestin tv-a; Trp53fl/fl or Nestin tv- a; Trp53fl/fl; Atrxfl/fl mice are infected with RCAS viruses encoding Cre recombinase, PDGFβ, and H3.3K27M. The resulting NSC will be used to characterize the global and local epigenetic and transcriptomic changes induced by H3.3K27M and its co-occurring mutations in NSC using western blots, ChIP-seq, and RNA-seq. Further, I will characterize the phenotypic changes caused by these alterations using immunocytochemistry, extreme limiting dilution, proliferation assays, and orthotopic allografts. Finally, I will validate the epigenetic and transcriptomic signatures of H3.3K27M mouse NSC in a panel of H3.3WT and H3.3K27M patient derived xenograft (PDX) cell lines via ChIP-seq and RNA-seq, as well as with existing datasets from human tumor samples. These studies will identify mechanisms of pathogenesis for H3.3K27M pHGG and potential epigenetic therapeutic targets that are both tractable in mice and relevant to humans. By completing this research proposal, I will gain technical skills applicable to cancer biology, neuroscience, epigenetics, and biochemistry while also developing my critical thinking, scientific communication, and ethical research skills.

项目概要 儿童高级别胶质瘤（pHGG）是儿童中最常见和恶性的原发性脑肿瘤。 H3F3A（编码组蛋白变体 H3.3 的基因之一）的突变是 pHGG 和 H3.3 的定义 突变状态与生存和肿瘤位置等临床变量相关。 pHGG 与赖氨酸 27 蛋氨酸替代 (H3.3K27M) 发生在脑干中，与特别差的生存率相关。 H3.3K27M 不足以驱动小鼠模型中的肿瘤发生。然而，它在人类中与 PDGFRA 激活同时发生 TP53 和 ATRX 缺失，表明 H3.3K27M 与这些致癌突变协同诱导 肿瘤。此外，H3.3K27M pHGG 和其他 pHGG 之间的表型差异表明不同 表观遗传改变和肿瘤发生机制。该提案的重点是确定表观遗传 H3.3K27M 及其同时发生的小鼠神经干细胞 (NSC) 诱导的转录组变化 突变以及这些突变如何导致神经胶质瘤细胞表型。对 H3.3K27M pHGG 进行精确建模 可能的话，我将使用基于 RCAS-TVA 的模型，其中 NSC 从 Nestin tv-a 中获取； Trp53fl/fl 或 Nestin 电视- 一个; Trp53fl/fl； Atrxfl/fl 小鼠感染编码 Cre 重组酶、PDGFβ 和 H3.3K27M 的 RCAS 病毒。这 由此产生的 NSC 将用于表征引起的全局和局部表观遗传和转录组变化 使用蛋白质印迹、ChIP-seq 和 RNA-seq 检测 H3.3K27M 及其在 NSC 中同时发生的突变。此外，我将 使用免疫细胞化学描述由这些改变引起的表型变化，极端限制 稀释、增殖测定和原位同种异体移植。最后，我将验证表观遗传学和转录组学 H3.3WT 和 H3.3K27M 患者来源的异种移植 (PDX) 细胞系中 H3.3K27M 小鼠 NSC 的特征 通过 ChIP-seq 和 RNA-seq，以及来自人类肿瘤样本的现有数据集。这些研究将 确定 H3.3K27M pHGG 的发病机制和潜在的表观遗传治疗靶点 对小鼠来说很容易处理，并且与人类相关。通过完成这个研究计划，我将获得技术技能 适用于癌症生物学、神经科学、表观遗传学和生物化学，同时也发展我的批判性思维 思维、科学沟通和道德研究技能。