Epigenetic dependence of diffuse midline glioma with H3K27M mutation

具有 H3K27M 突变的弥漫性中线胶质瘤的表观遗传依赖性

• 批准号: 10736036
• 负责人: Zhiguo Zhang
• 金额: $ 40.63万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2028-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10736036
• 关键词: ACVR1 gene; ATP phosphohydrolase; Amino Acids; Area; Brain; Brain region; CRISPR screen; Catalytic Domain; Cells; Childhood Glioma; Chromatin; Chromatin Remodeling Factor; Clinical; Complex; DNA Sequence Alteration; Data; Dependence; Diagnosis; Diffuse intrinsic pontine glioma; Disease; Drug Targeting; Epigenetic Process; Extracellular Matrix; Gene Expression; Gene Expression Profile; Gene Silencing; Genes; Genetic Transcription; Glioma; Growth; Heterozygote; Histone H3; In Vitro; Invaded; Lysine; Malignant Neoplasms; Mediating; Methionine; Methods; Methylation; Molecular; Mutation; Neurons; Normal Cell; Paracrine Communication; Pathogenesis; Pathogenicity; Patients; Play; Pontine structure; Proliferating; Proteins; Regulator Genes; Regulatory Element; Research; Role; SMARCA4 gene; SWI/SNF Family Complex; Somatic Mutation; Synapses; Testing; Tissues; Tumor Suppressor Genes; Variant; Work; Writing; brahma; cell growth; chemotherapy; diffuse midline glioma; driver mutation; effective therapy; epigenome; epigenomics; fitness; glioma cell line; in vivo; inhibitor; insight; mouse model; mutant; neoplastic cell; neurodevelopment; new therapeutic target; paralogous gene; patient derived xenograft model; pre-clinical; programs; protein H(3); recruit; therapeutic target; transcription factor; transcriptome sequencing; tumor; tumor growth; tumor initiation; tumorigenesis

Diffuse intrinsic pontine glioma (DIPG) is a deadly disease with the median survival of DIPG patients less than one year after diagnosis. DIPG tumors initiate from the pons and midline of brain, and spread to other brain regions where they mingle with normal cells such as neurons. Recently, it has been shown that neurons promote proliferation and invasion of DIPG cells. However, it is largely unknown how the intrinsic gene expression program of DIPG cells is regulated for their interactions with neurons. Furthermore, currently there are no effective treatments for DIPG patients. Therefore, it is imperative to understand the molecular basis of pathogenesis of DIPG and to identify novel drug targets for this deadly disease. About 80% DIPG tumors contain somatic mutations at genes encoding canonical histone H3 (H3.1) or its variant (H3.3), resulting in replacement of histone H3 lysine 27 with methionine (H3K27M). We found that expression of either H3.1K27M or H3.3K27M proteins leads to a global reduction of di- and tri-methylation (H3K27me2/me3) on wild type histone H3. H3K27me2/me3 marks are catalyzed by the PRC2 complex with Ezh2 as the catalytic subunit, and play important roles in gene silencing. However, H3.1K27M and H3.3K27M DIPG tumors show distinct gene expression signatures and are associated with distinct driver genetic mutations. We hypothesize that epigenome reprograming by H3.1K27M and H3.3K27M creates a dependence of DIPG tumor cells on other chromatin regulators. To test this hypothesis, we performed CRISPR/Cas9 screens and found that Brg1(brahma-related gene 1 or called SMACAR4), the catalytic subunit of mammalian SWI/SNF (mSWI/SNF) chromatin remodeling complexes, and Ezh2, are among top hits. The identification of Ezh2 is expected as we and others have shown that Ezh2 and H3K27me2/me3 are needed to silence tumor suppressor genes in DIPG cells. However, it was not known whether Brg1 has any roles in DIPG. Our results support the hypothesis that Brg1 functions as the catalytic subunit of DIPG-specific mSWI/SNF complexes to control the gene expression and fitness of DIPG cells. Furthermore, in H3.3K27M DIPG cells, transcription factor SOX10 recruits Brg1 to regulate the expression of genes involved in cell growth, extracellular matrix and neural development. Based on these exciting observations, we will 1) identify genes whose expression is regulated by Brg1 directly H3.1K27M DIPG cells; 2) evaluate genetic and epigenetic changes in H3.1K27M DIPG cells that render these cells depends on Brg1; 3) test the hypothesis that Brg1 and its target genes involved in neural development contribute to the neuron-glioma interactions of H3.3K27M DIPG cells; 4) identify other subunits of mSWI/SNF complexes that work with Brg1 to control gene expression and fitness of DIPG cells; and 5) test the hypothesis that inhibition of Brg1 alone and in combination with Ezh2 inhibition impede the growth of DIPG tumors using patient derived xenograft mouse models. Together, these studies will not only provide molecular insight into how epigenomic re-writing by H3K27M mutant proteins promotes tumorigenesis, but also identify drug targets for this deadly disease.

弥漫性内在脑桥胶质瘤（DIPG）是一种致命的疾病，DIPG患者的中位生存期小于 诊断后一年。DIPG肿瘤起源于脑桥和中线，并向其他脑组织扩散 它们与正常细胞如神经元混合的区域。最近，研究表明， DIPG细胞的增殖和侵袭。然而，很大程度上不知道内在基因表达是如何 DIPG细胞的程序因其与神经元的相互作用而受到调节。此外，目前没有 DIPG患者的有效治疗。因此，必须了解 DIPG的发病机制，并确定这种致命疾病的新药物靶点。约80%的DIPG肿瘤含有 编码典型组蛋白H3（H3.1）或其变体（H3.3）的基因发生体细胞突变，导致替换 组蛋白H3赖氨酸27与甲硫氨酸（H3 K27 M）的结合。我们发现H3.1K27M或H3.3K27M的表达与H3.1K27M的表达相关。 蛋白导致野生型组蛋白H3上的二甲基化和三甲基化（H3 K27 me 2/me 3）的整体减少。 H3 K27 me 2/me 3标记由PRC 2复合物催化，Ezh 2作为催化亚基， 基因沉默的作用。然而，H3.1K27M和H3.3K27M DIPG肿瘤显示不同的基因表达， 这些特征与不同的驱动基因突变有关。我们假设， 通过H3.1K27M和H3.3K27M的重编程产生了DIPG肿瘤细胞对其他染色质的依赖性 监管部门为了验证这一假设，我们进行了CRISPR/Cas9筛选，发现Brg 1（brahma相关） 基因1或称为SMACAR 4），哺乳动物SWI/SNF（mSWI/SNF）染色质重塑的催化亚基 复合物和EZH 2都是最热门的。正如我们和其他人已经证明的那样，Ezh 2的鉴定是预期的 Ezh 2和H3 K27 me 2/me 3是沉默DIPG细胞中抑癌基因所必需的。但有与会者 不知道Brg 1是否在DIPG中发挥任何作用。我们的研究结果支持Brg 1作为 DIPG特异性mSWI/SNF复合物的催化亚基，以控制DIPG的基因表达和适应性 细胞此外，在H3.3K27M DIPG细胞中，转录因子SOX 10招募Brg 1来调节Brg 1的表达， 细胞生长、细胞外基质和神经发育的基因。基于这些令人兴奋的 通过观察，我们将1）鉴定其表达直接受Brg 1调节的基因H3.1K27M DIPG细胞; 2） 评估H3.1K27 M DIPG细胞中使这些细胞依赖于Brg 1的遗传和表观遗传变化; 3） 验证Brg 1及其靶基因参与神经发育与神经胶质瘤的关系 H3.3K27M DIPG细胞的相互作用; 4）鉴定mSWI/SNF复合物的其他亚基，其与Brg 1起作用， 控制DIPG细胞的基因表达和适应性;和5）检验单独和联合抑制Brg 1的假设。 与Ezh 2抑制的组合使用患者来源的异种移植小鼠阻止DIPG肿瘤的生长 模型总之，这些研究将不仅提供分子洞察如何表观基因组重写， H3 K27 M突变蛋白促进肿瘤发生，但也确定了这种致命疾病的药物靶点。