Evaluation of Bruton’s Tyrosine Kinase and p110 delta in Mutant Shp2-Induced JMML

突变型 Shp2 诱导的 JMML 中 Bruton 酪氨酸激酶和 p110 delta 的评估

• 批准号: 9326811
• 负责人: Lisa Deng Yuen
• 金额: $ 3.65万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-01 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9326811
• 关键词: AKT Signaling Pathway; Accounting; Address; Agammaglobulinaemia tyrosine kinase; Allogeneic Bone Marrow Transplantation; B-Cell Leukemia; B-Lymphocytes; Binding; Biochemical; Blood Cells; Cancerous; Catalytic Domain; Cells; Childhood; Chronic Lymphocytic Leukemia; Data; Development; Disease; Dominant-Negative Mutation; Evaluation; Feedback; GTP-Binding Protein alpha Subunits, Gs; Genetic; Granulocyte-Macrophage Colony-Stimulating Factor; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic stem cells; Hypersensitivity; In Vitro; Juvenile Myelomonocytic Leukemia; Knock-out; Laboratories; Lead; Link; Malignant Childhood Neoplasm; Malignant lymphoid neoplasm; Mantle Cell Lymphoma; Mediating; Modeling; Molecular Models; Mus; Mutation; Myeloid Cells; Myeloproliferative disease; PTPN11 gene; Pathway interactions; Patients; Pharmacology; Phospholipase C; Phospholipids; Protein Kinase C; Protein Tyrosine Kinase; Protein Tyrosine Phosphatase; Proteins; Receptor Signaling; Receptors, Antigen, B-Cell; Regulation; Relapse; Role; Safety; Series; Signal Transduction; Site; Structure; Testing; Time; Waldenstrom Macroglobulinemia; Work; base; curative treatments; efficacy testing; experimental study; gain of function; gain of function mutation; hyperactive Ras; improved; in vivo; inhibitor/antagonist; kinase inhibitor; leukemia; molecular modeling; mutant; novel; novel strategies; phosphoinositide-3,4,5-triphosphate

PROJECT SUMMARY/ABSTRACT Juvenile myelomonocytic leukemia (JMML) is a fatal childhood myeloproliferative neoplasm in which myeloid cells are overproduced and hematopoietic progenitors are hypersensitive to granulocyte-macrophage colony stimulating factor (GM-CSF). The only curative treatment is allogeneic bone marrow transplant, but even this rigorous therapy yields only a 50% relapse-free 5-year survival. A majority of patients have hyperactive Ras signaling, with the most common mutation being somatic gain-of-function (GOF) mutations in PTPN11, which encodes the protein tyrosine phosphatase Shp2. Previous work in the Chan lab has shown that the PI3K catalytic subunit p110δ is needed for both Akt and Erk hyperactivation, and promotes GOF Shp2-induced GM- CSF hypersensitivity and hyperproliferation, contributing to the progression of JMML. Based on the significant role of p110δ in promoting GOF Shp2-induced leukemia, we investigated potential tyrosine kinases that can cooperate with p110δ to promote Akt and Erk activation and lead to hyperproliferation of myeloid cells. Bruton's Tyrosine Kinase (BTK) has been identified as a critical molecule in lymphoid malignancies and the BTK inhibitor, ibrutinib, was recently approved for use in patients with B cell leukemias. As BTK has been well- studied in the context of B cell receptor signaling, it is known that BTK is activated downstream of p110δ and that active BTK phosphorylates B cell adaptor for PI3K (BCAP), allowing phospho-BCAP to bind to the regulatory p85α subunit and promote activation of PI3K. Based on what is known in B cells, we hypothesize that BTK and BCAP are critical players in the GOF Shp2-mediated hyperactivation of p110δ signaling in JMML. To address this hypothesis, we propose three aims: (1) we will use murine cells expressing the GOF Shp2 mutation, E76K, with genetic knockout of BTK to perform proliferation and biochemical analysis in vitro, as well as examine the progression of myeloproliferative disease in p110δ inhibitor-treated mice in vivo; (2) we will use cells from mice lacking expression of p85α along with p85α site-specific mutant constructs to explore the physical interaction between BCAP and p85α; and (3) we will test the efficacy of BTK (ACP-196) and p110δ (ACP-319) inhibitors provided by AcertaPharma on Shp2E76K-expressing mice in vivo, as well as on primary JMML patient cells in vitro. These studies are timely and significant, as the AcertaPharma inhibitors are currently being tested in combination for safety and efficacy in chronic lymphocytic leukemia patients, and therefore have the potential to be a novel treatment in patients with JMML.

项目总结/摘要 青少年粒单核细胞白血病（JMML）是一种致命的儿童骨髓增生性肿瘤，其中髓系 细胞过度产生，造血祖细胞对粒细胞-巨噬细胞集落高度敏感 刺激因子（GM-CSF）。唯一的治疗方法是异基因骨髓移植，但即使这样， 严格的治疗只能使5年无复发生存率达到50%。大多数患者有过度活跃的Ras 最常见的突变是PTPN 11中的体细胞功能获得性（GOF）突变， 编码蛋白酪氨酸磷酸酶Shp 2。Chan实验室以前的工作表明，PI 3 K 催化亚基p110δ是Akt和Erk过度活化所必需的，并促进GOF Shp 2诱导的GM-1。 CSF超敏反应和过度增殖，促进JMML的进展。基于重大 p110δ在促进GOF Shp 2诱导的白血病中的作用，我们研究了潜在的酪氨酸激酶， 与p110δ协同作用，促进Akt和Erk活化，导致髓系细胞过度增殖。布鲁顿氏 酪氨酸激酶（BTK）已被确定为淋巴恶性肿瘤的关键分子， 抑制剂伊曲替尼最近被批准用于B细胞白血病患者。因为BTK一直很好- 在B细胞受体信号传导的背景下研究，已知BTK在p110δ的下游被激活， 活性BTK磷酸化B细胞PI 3 K衔接子（BCAP），使磷酸化BCAP结合到 调节p85α亚基并促进PI 3 K的活化。基于对B细胞的了解，我们假设 BTK和BCAP是JMML中GOF Shp 2介导的p110δ信号过度激活的关键参与者。 为了解决这一假设，我们提出了三个目标：（1）我们将使用表达GOF Shp 2的鼠细胞 突变，E76 K与BTK的基因敲除一起进行体外增殖和生化分析，以及 作为检查骨髓增生性疾病在p110δ受体治疗的小鼠体内的进展;（2）我们将使用 来自缺乏p85α表达的小鼠的细胞与p85α位点特异性突变构建体一起沿着，以探索 BCAP和p85α之间的物理相互作用;（3）我们将测试BTK（ACP-196）和p110δ的功效 AcertaPharma提供的（ACP-319）抑制剂对Shp 2 E76 K表达小鼠的体内试验以及对原代 体外JMML患者细胞。这些研究是及时和重要的，因为AcertaPharma抑制剂是 目前正在慢性淋巴细胞白血病患者中测试联合用药的安全性和有效性， 因此有可能成为JMML患者的新治疗方法。