Protein Phosphatase PP2A and DNA damage in cell fate decisions of acute myeloid leukemic cells

蛋白磷酸酶 PP2A 和 DNA 损伤在急性髓系白血病细胞命运决定中的作用

• 批准号: 10019487
• 负责人: Swagata Goswami
• 金额: $ 3.8万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-17 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10019487
• 关键词: ATR gene; Acute Myelocytic Leukemia; Acute Promyelocytic Leukemia; Apoptosis; Automobile Driving; CD34 gene; Cell Cycle; Cell Cycle Arrest; Cell Death; Cell Differentiation process; Cell Fate Control; Cell Line; Cell Survival; Cells; Cellular biology; Cessation of life; ChIP-seq; Cytometry; DNA Damage; DNA Repair; Data; Differentiation Therapy; Differentiation and Growth; Flow Cytometry; GADD45 protein; GADD45A gene; Genetic; Genomics; Goals; Growth; Hematologic Neoplasms; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic System; Hematopoietic stem cells; Homeostasis; In Vitro; Knock-out; Knowledge; Lead; Leukemic Cell; Link; Literature; Malignant Neoplasms; Mediating; Mind; Molecular; Mutation; Myelogenous; Oncogenes; Oncogenic; Pathogenesis; Pathway interactions; Patients; Pharmaceutical Preparations; Pharmacology; Phase; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Play; Protein Serine/Threonine Phosphatase; Protein phosphatase; Proteins; RNA Interference; RUNX3 gene; Research; Research Project Grants; Research Proposals; Resistance; Retinoblastoma; Retinoblastoma Protein; Role; Safety; Signal Transduction; Small Interfering RNA; Stress; Survival Rate; System; Therapeutic; Therapeutic Agents; Tretinoin; Tumor Suppressor Proteins; Work; acute myeloid leukemia cell; base; c-myc Genes; cell injury; chemotherapy; clinical development; design; effective therapy; genetic manipulation; genome-wide; in vivo; inhibitor/antagonist; insight; interest; kinase inhibitor; knock-down; leukemia; leukemic stem cell; mouse model; novel; novel strategies; novel therapeutics; oncoprotein p21; overexpression; pre-clinical; protein activation; relapse patients; repaired; research clinical testing; response; single cell analysis; skills; stem cell population; therapeutic candidate; tool; transcription factor; transcriptome sequencing

PROJECT SUMMARY Acute Myeloid Leukemia (AML) is a cancer characterized by a co-operative block in hematopoietic maturation and proliferative advantage. It is near incurable, with a 5-year survival rate of only 25% and median survival <1year in patients over 65 years. Current chemotherapy is very poorly tolerated and majority of patients relapse. Overcoming the differentiation block is a promising therapeutic avenue, however is majorly lacking for AML subtypes without a known driver oncogene. Our study examines new pathways to target the two major aspects of leukemic cell survival, the differentiation block and increased proliferative ability, that can prove effective across multiple subtypes. Reversible protein phosphorylation controls many aspects of such cell fate decisions, and an imbalance in the normal activities of kinases and phosphatases contribute to pathogenesis of AML. However, while the aberrant activation of kinases, frequent in AML, is well studied and exploited for therapeutic purposes, inactivation of phosphatases, also prevalent in AML, is often overlooked. This proposal focuses on the serine threonine phosphatase tumor suppressor Protein Phosphatase 2A (PP2A) and its role in influencing proliferation vs differentiation decisions in AML. For the F99 phase, we identified a novel role for PP2A in driving myeloid differentiation and cell cycle arrest in AML. Based on our preliminary data, our overall hypothesis is that PP2A drives differentiation and cell cycle arrest through the cell cycle regulator p21. We specifically seek to establish the mechanism of PP2A mediated p21 induction, and the role of potential downstream factors Retinoblastoma protein and CEBP proteins in driving differentiation in AML using genetic overexpression and knockdown systems, CHIP-Seq and single cell analysis using Mass Cytometry. The K00 phase focuses on identification of factors mediating DNA damage induced differentiation. While DNA damage has been shown to induce differentiation in AML and hematopoietic progenitors, the molecular mechanism is poorly understood. Interestingly, DNA repair machinery including ATM and ATR serine threonine kinases protect leukemic cells from damage induced differentiation and are hyperactivated in AML. This raises the possibility that inhibiting DNA repair can promote genomic stress induced differentiation. My hypothesis for the K00 phase is that PP2A can antagonize DNA repair kinases and promote differentiation in response to DNA damage in AML. I will establish the role of PP2A and other potential candidates identified from preliminary data and literature such as GADD45 proteins and p21 in DNA damage induced differentiation in AML using genetic and pharmacological activation of PP2A, overexpression, knockdown and knockout of GADD45 as well as global in-vitro and in-vivo RNAi and genome wide CRISPER screen analyses. Successful completion of this proposal will yield new insight into cellular pathways capable of inducing growth arrest and terminal differentiation of maturation blocked cancers such as AML.

项目摘要 急性髓样白血病（AML）是一种癌症，其特征是造血成熟中的合作障碍 和增殖优势。它几乎无法治愈，5年生存率仅为25％，中位存活率 在65年以上的患者中<1年。当前的化学疗法的耐受性较差，大多数患者复发。 克服差异化是一个有前途的治疗大道，但是AML主要缺乏 亚型没有已知的驱动器癌基因。我们的研究研究了针对两个主要方面的新途径 白血病细胞存活，分化阻滞和增强能力的提高，这可能证明有效 跨多个子类型。 可逆蛋白质磷酸化控制了这种细胞命运决定的许多方面，并且在 激酶和磷酸酶的正常活性有助于AML的发病机理。但是，虽然异常 激活AML中经常的激活是为了治疗目的，对AML的经常进行了充分的研究和利用 通常在AML中盛行的磷酸酶通常被忽略。该提议重点介绍丝氨酸苏氨酸 磷酸酶肿瘤抑制蛋白磷酸酶2a（PP2A）及其在影响增殖中的作用 AML的分化决策。对于F99阶段，我们确定了PP2A在驱动髓样中的新作用 AML中的分化和细胞周期停滞。基于我们的初步数据，我们的总体假设是PP2A 通过细胞周期调节剂p21驱动分化和细胞周期停滞。我们特别寻求建立 PP2A介导的P21诱导的机制以及潜在下游因子的作用。 使用遗传过表达和敲低的蛋白质和CEBP蛋白在驱动AML的分化方面进行分化 使用质量细胞仪的系统，芯片序列和单细胞分析。 K00期的重点是识别介导DNA损伤引起的分化的因素。而DNA 损害已显示可诱导AML和造血祖细胞的分化，分子 机制知之甚少。有趣的是，DNA维修机械包括ATM和ATR丝氨酸苏氨酸 激酶可保护白血病细胞免受损伤诱导的分化，并在AML中过度激活。这加剧了 抑制DNA修复可以促进基因组应激引起的分化的可能性。我的假设 K00相是PP2A可以拮抗DNA修复激酶并促进对DNA的分化 AML损坏。我将确定从初步数据确定的PP2A和其他潜在候选者的作用 和诸如GADD45蛋白和DNA损伤中P21之类的文献使用遗传诱导AML诱导分化 PP2A的药理激活，GADD45的过表达，敲低和敲除以及全球 体内和体内RNAi和基因组宽阔的筛网分析。成功完成此建议 将对能够诱导生长停滞和终末分化的细胞途径产生新的见解 成熟阻塞了癌症，例如AML。