Identifying genetic vulnerabilities in KIAA1549-BRAF mutant pediatric low-grade gliomas

识别 KIAA1549-BRAF 突变儿童低级别神经胶质瘤的遗传脆弱性

• 批准号: 10752212
• 负责人: Sean Alexander Misek
• 金额: $ 7.18万
• 依托单位: BROAD INSTITUTE, INC.
• 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 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10752212
• 关键词: Affect; BRAF gene; Binding; Brain; CRISPR screen; Cell membrane; Cells; Childhood; Chronic; Clinical; Complex; DNA Sequence Alteration; Dependence; Development; Dose; Dose Limiting; Essential Genes; Event; GIT2 gene; Genes; Genetic Predisposition to Disease; Genome; Glioma; Goals; Human; Integral Membrane Protein; Knowledge; Length; MAP Kinase Gene; Mediating; Membrane; Modeling; Mutation; N-terminal; Oncogenes; Oncogenic; Pathway interactions; Patients; Plasma Enhancement; Post-Translational Protein Processing; Proteins; Proteome; Resistance; Signal Transduction; Site; Testing; Therapeutic; Toxic effect; Transmembrane Domain; Withholding Treatment; cell growth; cofactor; extracellular; genome-wide; inhibitor; loss of function; member; mutant; new therapeutic target; novel therapeutics; prevent; protein-O-mannosyltransferase 1; protein-O-mannosyltransferase 2; relapse prevention; therapeutic target; tumor

Project Summary: Pediatric low-grade gliomas (pLGGs) typically harbor only a single oncogenic driver event which almost universally result in MAPK pathway activation. The most frequent of these alterations is a rearrangement that fuses the C-terminus of BRAF to the large transmembrane protein KIAA1549. Many patients whose tumors harbor this fusion respond to BRAF inhibitors, but challenges remain in extending clinical benefit to all patients. A key challenge is that patients require chronic treatment with these inhibitors throughout childhood because tumors often rebound upon cessation of treatment. These inhibitors are associated with significant dose-limiting toxicity issues, likely because the MAPK pathway is essential for brain development. These observations suggest that we need to identify new therapeutic targets for pLGG. The two core members of the POMT complex and two complex cofactors emerged as the top four dependencies in a genome-scale loss of function CRISPR/Cas9 screen. This dependency was specific for cells harboring the KIAA1549-BRAF fusion and were not dependencies in cells expressing BRAFV600E. The POMT complex is necessary for O-mannosylation of secreted and transmembrane proteins. This is striking because across all proteins in the human proteome, KIAA1549 one of the proteins most affected by this post-translational modification. It was originally hypothesized that the KIAA1549-BRAF fusion activates BRAF by truncating its negative regulatory domain, but this observation suggests that the KIAA1549 portion of the fusion may also be necessary for activating BRAF. The primary goals of this proposal are to understand why the POMT complex is a vulnerability in KIAA1549- BRAF mutant cells, and to understand how the KIAA1549 portion of KIAA1549-BRAF regulates the activity of the fusion. In Aim 1 I will test the hypothesis that POMT complex activity is necessary for the survival of KIAA1549-BRAF-dependent cells. In Aim 2 I will test the hypothesis that the POMT complex is required for O- mannosylation of KIAA1549-BRAF, and that O-mannosylation of KIAA1549-BRAF is necessary for oncogenic signaling. In Aim 3 I will test the hypothesis that the KIAA1549 portion of the KIAA1549-BRAF fusion tethers BRAF to the plasma membrane and enhances its ability to activate the MAPK pathway.

项目概要： 儿童低级别胶质瘤（pLGG）通常只有一个致癌驱动事件， 普遍导致MAPK通路激活。这些改变中最常见的是重排， 将BRAF的C末端融合到大跨膜蛋白KIAA 1549。许多患者的肿瘤 尽管这些患者对BRAF抑制剂有反应，但在将临床获益扩展至所有患者方面仍存在挑战。 一个关键的挑战是，患者需要在整个儿童期长期使用这些抑制剂治疗， 肿瘤经常在停止治疗后反弹。这些抑制剂与显著的剂量限制性 毒性问题，可能是因为MAPK途径对大脑发育至关重要。这些观察结果表明 我们需要为pLGG找到新的治疗靶点。 POMT复合体的两个核心成员和两个复合辅因子成为前四个依赖项 在基因组规模的功能丧失CRISPR/Cas9筛选中。这种依赖性是特定的细胞， KIAA 1549-BRAF融合蛋白在表达BRAFV 600 E的细胞中不具有依赖性。POMT复合体是 分泌和跨膜蛋白的O-甘露糖基化所必需的。这是惊人的，因为在所有 KIAA 1549是人类蛋白质组中最受这种翻译后表达影响的蛋白质之一， 改性最初假设KIAA 1549-BRAF融合物通过截短其 负调控结构域，但这一观察结果表明，融合的KIAA 1549部分也可能是 激活BRAF所必需的。 本提案的主要目标是了解为什么POMT复合体是KIAA 1549中的一个漏洞- BRAF突变细胞，并了解KIAA 1549-BRAF的KIAA 1549部分如何调节BRAF突变细胞的活性。 融合。在目标1中，我将检验POMT复合物活性对于细胞存活是必要的这一假设。 KIAA 1549-BRAF依赖性细胞。在目标2中，我将检验POMT复合物是O-所需的假设。 KIAA 1549-BRAF的O-甘露糖基化，并且KIAA 1549-BRAF的O-甘露糖基化是致癌性所必需的。 信号在目标3中，我将检验KIAA 1549-BRAF融合系链的KIAA 1549部分 BRAF可以促进细胞膜的粘附，并增强其激活MAPK通路的能力.