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.

项目总结 急性髓系白血病(AML)是一种以造血成熟受阻为特征的癌症。 和扩散优势。它几乎是无法治愈的，5年存活率只有25%，中位存活率 65岁以上患者1年。目前的化疗耐受性很差，大多数患者会复发。 克服分化障碍是一条很有希望的治疗途径，然而对于急性髓细胞白血病来说，主要缺乏这种方法。 没有已知致癌基因的亚型。我们的研究考察了针对这两个主要方面的新途径 对白血病细胞的存活、分化的阻断和增殖能力的增强，这可以被证明是有效的 跨多个子类型。 可逆的蛋白质磷酸化控制着这种细胞命运决定的许多方面，而 急性髓系白血病的发病机制之一是蛋白水解酶和磷酸酶活性正常。然而，虽然反常的 在急性髓细胞白血病中常见的激活的激酶被很好地研究和开发用于治疗目的，失活的 磷酸酶在急性髓细胞白血病中也很普遍，但经常被忽视。这项提案的重点是丝氨酸苏氨酸 磷酸酶肿瘤抑制蛋白磷酸酶2A(PP2A)及其在影响细胞增殖和凋亡中的作用 急性髓系白血病的差异化决策。对于F99期，我们确定了PP2A在驱动髓系中的一个新角色 急性髓系白血病的分化和细胞周期停滞。根据我们的初步数据，我们的总体假设是PP2A 通过细胞周期调节因子p21促进分化和细胞周期停滞。我们特别寻求建立 PP2A介导的p21诱导机制及潜在下游因子在视网膜母细胞瘤中的作用 蛋白质和CEBP蛋白在AML基因过度表达和基因敲除促进分化中的作用 系统，芯片序列和使用质量细胞术的单细胞分析。 K00阶段着重于鉴定DNA损伤诱导分化的因素。而DNA 损伤已被证明可以诱导AML和造血祖细胞分化，分子 机制还不是很清楚。有趣的是，DNA修复机制包括ATM和ATR丝氨酸苏氨酸 激酶保护白血病细胞免受损伤诱导分化，并在急性髓细胞白血病中被过度激活。这就提高了 抑制DNA修复可以促进基因组胁迫诱导分化的可能性。我的假设是 K00期是PP2A拮抗DNA修复激酶，促进DNA反应分化的阶段 AML中的损坏。我将确定PP2A和从初步数据中确定的其他潜在候选人的角色 和文献，如GADD45蛋白和p21在DNA损伤诱导AML分化中使用基因 和PP2A的药理激活、GADD45的过表达、敲除和敲除以及全球 体外和体内RNAi和全基因组CRISER Screen分析。成功完成本建议书 将对能够诱导细胞生长停滞和终末分化的细胞通路产生新的见解 成熟可以阻止AML等癌症的发生。