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.

弥漫性内在蓬托胶质瘤（DIPG）是一种致命疾病，DIPG患者的中位存活率小于 诊断后一年。 DIPG肿瘤是从大脑的PON和中线引发的，并扩散到其他大脑 它们与正常细胞（如神经元）混合的区域。最近，已经表明神经元促进 DIPG细胞的增殖和侵袭。但是，在很大程度上未知固有基因表达如何 DIPG细胞的程序与神经元的相互作用受到调节。此外，目前没有 DIPG患者的有效治疗方法。因此，必须了解 DIPG的发病机理并确定这种致命疾病的新药物靶标。大约80％DIPG肿瘤包含 编码规范组蛋白H3（H3.1）或其变体（H3.3）的基因的体细胞突变，导致替换 用蛋氨酸（H3K27M）组蛋白H3赖氨酸27的组成。我们发现H3.1K27M或H3.3K27M的表达 蛋白质导致野生型组蛋白H3上的Di-和三甲基化（H3K27ME2/ME3）的全球降低。 H3K27ME2/ME3标记是由EZH2作为催化亚基的PRC2复合物催化的，并且发挥重要作用 基因沉默中的作用。但是，H3.1K27M和H3.3K27M DIPG肿瘤显示出独特的基因表达 签名，与不同的驱动基因突变有关。我们假设表观基因组 通过H3.1K27M和H3.3K27M重新编程可产生DIPG肿瘤细胞对其他染色质的依赖性 监管机构。为了检验该假设，我们进行了CRISPR/CAS9屏幕，发现BRG1（梵天有关 基因1或称为Smacar4），哺乳动物SWI/SNF（MSWI/SNF）染色质重塑的催化亚基 复合物和EZH2是最受欢迎的热门单曲。正如我们和其他人所表明的那样，EZH2的识别是可以的 需要EZH2和H3K27ME2/ME3来使DIPG细胞中的肿瘤抑制基因沉默。但是，是 不知道BRG1在DIPG中是否有任何作用。我们的结果支持BRG1作为 DIPG特异性MSWI/SNF复合物的催化亚基控制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SOX10募集BRG1来调节表达 涉及细胞生长，细胞外基质和神经发育的基因。基于这些令人兴奋的 观察结果，我们将1）识别其表达受BRG1直接调节的基因，直接H3.1K27M DIPG细胞； 2） 评估造成这些细胞的H3.1K27M DIPG细胞中的遗传和表观遗传变化取决于BRG1； 3） 检验BRG1及其与神经发育涉及的靶基因的假设有助于神经元 - 胶质瘤 H3.3K27M DIPG细胞的相互作用； 4）确定与BRG1一起使用的MSWI/SNF复合物的其他亚基 控制DIPG细胞的基因表达和适应性； 5）检验单独抑制BRG1和IN的假设 与EZH2抑制结合使用患者衍生的异种移植小鼠阻碍DIPG肿瘤的生长 型号。总之，这些研究不仅将提供分子的洞察力，了解表观基因组如何通过 H3K27M突变蛋白促进肿瘤发生，但也鉴定出该致命疾病的药物靶标。