Targeting an RNA Binding Protein Network in Acute Myeloid Leukemia

靶向急性髓系白血病中的 RNA 结合蛋白网络

• 批准号: 10171812
• 负责人: Omar Abdel-Wahab
• 金额: $ 55.59万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10171812
• 关键词: Ablation; Acute Myelocytic Leukemia; Adaptor Signaling Protein; Address; Adult; Binding; Biological; CD34 gene; CRISPR/Cas technology; Cancer cell line; Cell Line; Cell physiology; Cells; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Data; Dependence; Disease; Drug Targeting; Evaluation; Excision; Exons; Genetic; Genetic Engineering; Genetic Models; Genetic Transcription; Growth; HOXA9 gene; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; Histologic; Human; Impairment; In Vitro; Introns; Link; Liquid substance; Maintenance; Malignant - descriptor; Malignant Neoplasms; Mediating; Messenger RNA; Methods; Modeling; Mutation; Non-Malignant; Nucleosomes; Oncogenic; Patients; Pharmaceutical Preparations; Pharmacology; Physiological; Play; Population; Protein Splicing; Proteins; Proteomics; RNA; RNA Binding; RNA Recognition Motif; RNA Splicing; RNA analysis; RNA-Binding Proteins; Recurrence; Role; Sampling; Solid; Sulfonamides; Time; Tissues; Transcription Elongation; Translations; Up-Regulation; acute myeloid leukemia cell; anti-cancer; base; cancer cell; cancer survival; cell type; conditional knockout; improved; in vivo; insight; leukemia; leukemia initiating cell; loss of function; member; mouse model; mutant; novel; novel strategies; overexpression; patient derived xenograft model; progenitor; stem; treatment strategy; tumorigenesis; ubiquitin ligase

RNA-binding proteins (RBPs) are essential modulators of transcription and translation frequently dysregulated in cancer. We recently systematically interrogated RBP dependencies in a number of human cancers using a comprehensive CRISPR/Cas9 domain-focused screen targeting RNA-binding domains (RBDs) of 490 classical RBPs. This uncovered a network of physically interacting RBPs upregulated in acute myeloid leukemia (AML) and crucial for maintaining physiological RNA splicing and AML survival. Genetic targeting of one key member of this network, RBM39, repressed cassette exon inclusion and promoted intron retention within mRNAs encoding HOXA9 targets as well as in other RBPs preferentially required in AML. Fortuitously, it has recently been described that a class of clinically validated anti-cancer sulfonamide compounds (including the drugs indisulam and E7820) mediate RBM39 degradation as their dominant cellular mechanism of action. This occurs via novel interactions between these compounds and the DCAF15 adapter protein of the CUL4/Ddb1 ubiquitin ligase complex with RBM39 as a neo-substrate. Treatment of AML cells with such compounds in vitro and in vivo resulted in similar lethal cellular effects due to perturbations in RNA splicing. The effects of RBM39 loss on splicing resulted in preferential lethality of spliceosomal mutant AML, providing a novel strategy for treatment of AML bearing recurrent mutations in RBPs that regulated splicing. Overall these data identify RBM39 as central to a network of functionally and physically interacting RBPs upregulated in AML over normal hematopoietic precursors and required for AML maintenance. Despite these insights we do not yet understand the basis for the cell- and context-specific roles of RBM39 in malignant versus normal hematopoietic cells. We also do not understand the exact mechanisms by which RBM39 loss results in eradication of AML. We hypothesize that RBM39 is differentially required in malignant versus normal hematopoietic cells, may be differentially required depending on the precise stage of hematopoiesis, and will be required for leukemia initiation as well as maintenance. These hypotheses will be addressed in two Specific Aims. Aim 1 will determine the biological role of RBM39 in normal and malignant hematopoiesis. In this Aim, we will utilize a novel genetic model for genetic deletion of RBM39 in vivo in a cell- and time-specific manner to rigorously dissect the roles of RBM39 in malignant versus normal hematopoietic stem and progenitor cell populations. Aim 2 will identify the mechanistic basis for cell-type and disease-specific roles for RBM39 in normal and malignant hematopoietic cells. In this Aim we will compare the direct RNA binding targets and effects of RBM39 loss on splicing across normal and malignant hematopoietic cells. In addition, we will evaluate a potential novel for RBM39 in transcriptional elongation by the FACT complex, identified by our preliminary studies as an interactor of RBM39.

RNA 结合蛋白 (RBP) 是经常失调的转录和翻译的重要调节剂 在癌症中。我们最近使用一种方法系统地研究了多种人类癌症中 RBP 的依赖性。 针对 490 个经典的 RNA 结合域 (RBD) 的全面 CRISPR/Cas9 域聚焦筛选 RBP。这揭示了急性髓系白血病 (AML) 中上调的物理相互作用 RBP 网络 对于维持生理性 RNA 剪接和 AML 存活至关重要。一名关键成员的基因靶向 该网络的 RBM39 抑制盒式外显子包含并促进 mRNA 内的内含子保留 编码 HOXA9 目标以及 AML 优先需要的其他 RBP。巧合的是，最近 据描述，一类经过临床验证的抗癌磺酰胺化合物（包括药物 indisulam 和 E7820）介导 RBM39 降解作为其主要细胞作用机制。这 通过这些化合物与 CUL4/Ddb1 的 DCAF15 接头蛋白之间的新型相互作用发生 以 RBM39 作为新底物的泛素连接酶复合物。用此类化合物治疗 AML 细胞 由于 RNA 剪接的扰动，体外和体内导致了类似的致死细胞效应。的影响 RBM39 剪接缺失导致剪接体突变 AML 优先致死，提供了一种新策略 用于治疗带有调节剪接的 RBP 反复突变的 AML。 总的来说，这些数据表明 RBM39 是功能和物理相互作用的 RBP 网络的核心 与正常造血前体细胞相比，在 AML 中表达上调，并且是 AML 维持所需的。尽管有这些 我们尚不了解 RBM39 在恶性细胞中的细胞和环境特异性作用的基础 与正常造血细胞相比。我们也不了解 RBM39 丢失的确切机制 结果是根除 AML。我们假设 RBM39 在恶性和恶性疾病中的需求是不同的 正常造血细胞，根据具体的阶段可能有不同的需要 造血作用，并且是白血病发生和维持所必需的。这些假设 将在两个具体目标中得到解决。目标 1 将确定 RBM39 在正常和 恶性造血。在这个目标中，我们将利用一种新的遗传模型来对 RBM39 进行遗传删除。 以细胞和时间特异性方式在体内严格剖析 RBM39 在恶性与正常中的作用 造血干细胞和祖细胞群。目标 2 将确定细胞类型和 RBM39 在正常和恶性造血细胞中的疾病特异性作用。在这个目标中，我们将比较 直接RNA结合靶标以及RBM39缺失对正常和恶性造血系统剪接的影响 细胞。此外，我们将通过 FACT 评估 RBM39 在转录延伸方面的潜在新颖性 复合物，经我们的初步研究确定为 RBM39 的相互作用因子。