Dose-dependent Activation of the MEK/ERK Pathway in Hematopoiesis

造血过程中 MEK/ERK 通路的剂量依赖性激活

• 批准号: 8504631
• 负责人: Jing Zhang
• 金额: $ 35.43万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8504631
• 关键词: Acute; Affect; Alleles; Biological Assay; Blast Phase; Bone Marrow Transplantation; Categories; Cell Survival; Cells; Chronic; Clinic; Clinical; Cytokine Signaling; Data; Development; Disease; Disease model; Dose; Engineering; Equilibrium; Exposure to; Genes; Genetically Engineered Mouse; Goals; Hematopoiesis; Hematopoietic; Hematopoietic Neoplasms; Human; In Vitro; Knockout Mice; Knowledge; Lead; Leukemic Cell; Lymphoid Cell; MAP2K1 gene; MAPK3 gene; MEK inhibition; MEKs; Malignant Neoplasms; Measures; Mediating; Methods; Microarray Analysis; Mus; Myelogenous; Myeloid Cells; Myeloproliferative disease; Oncogenic; Ontology; Pathway interactions; Patients; Phase I Clinical Trials; Phosphorylation; Play; Point Mutation; Ras/Raf; Recruitment Activity; Regimen; Research; Role; Sampling; Serine; Signal Pathway; Signal Transduction; Stem cells; Testing; Transplant Recipients; base; cytokine; disease phenotype; exhaustion; gain of function mutation; human disease; improved; in vivo; inhibitor/antagonist; insight; mouse model; novel; novel therapeutics; progenitor; public health relevance; research study; self-renewal; stem; tumorigenesis

DESCRIPTION (provided by applicant): MEK/ERK is a major signaling pathway downstream of Ras. Recently, we characterized conditional mouse models of human myeloproliferative diseases (MPDs) that express one or two copies of oncogenic Nras in the hematopoietic compartment (Nras G12D/+ and Nras G12D/G12D), respectively. In both MPD models, the MEK/ERK pathway is hyperactivated in hematopoietic stem/progenitor cells (HSPCs) upon cytokine stimulation. Moreover, genetically altered HSCs are required to initiate and maintain MPD phenotypes and thus serve as MPD initiating cells. Interestingly, unlike in HSPCs and other progenitors, we found that Nras G12D signaling constitutively hyperactivates ERK1/2 in HSCs in a dose-dependent manner. This aberrant signaling is associated with increased self-renewal capability of Nras G12D/+ HSCs but reduced self-renewal and exhaustion of Nras G12D/G12D HSCs. Consistently, microarray analysis identifies the MEK/ERK pathway as the most strongly affected functional gene ontology category in both groups of HSCs. Collectively, these studies raise the possibility that hyperactivation of the MEK/ERK pathway regulates the self-renewal and differentiation of HSCs in a dose-dependent manner and is a critical step leading to MPD. In support of this hypothesis, AZD6244, a MEK inhibitor, rescues exhaustion of Nras G12D/G12D HSCs, effectively controls MPD phenotypes, and significantly prolongs the survival of diseased mice. To further test our hypothesis in vivo, we have generated a novel conditional MEK1 knockin allele carrying point mutations that mimic phosphorylation at serine 218 and serine 222 (LSL MEK1 S218;222E). MEK1 S218;222E is a constitutively active form of MEK1. As a part of our long-term goal to elucidate novel signaling mechanisms that control hematopoiesis, the specific aims of this proposal are to: 1) determine whether hyperactivation of the MEK/ERK pathway is necessary and sufficient for Nras G12D-mediated MPD phenotypes; 2) determine whether Nras G12D signaling regulates HSC self-renewal and differentiation through a dose-dependent activation of the MEK/ERK pathway; 3) determine whether inhibition of MEK1/2 and Jak2 blocks Nras G12D/G12D-mediated MPD and its transformation to a blast crisis. Together, the experiments proposed in these two specific aims will provide insights into the central role of MEK/ERK signaling in hematopoiesis. These insights will not only bring us closer to understanding the role of genetically altered HSCs in tumorigenesis, but also improve fundamental understanding of developmental and myeloproliferative diseases caused by dysfunctional Ras/Raf/MEK signaling.

描述（由申请人提供）：MEK/ERK是RAS下游的主要信号通路。最近，我们表征了人脊髓增生性疾病（MPD）的条件小鼠模型，该模型分别表达造血室（NRAS G12D/+和NRAS G12D/G12D）中表达一两个致癌NRA的副本NRA。在两个MPD模型中，在细胞因子刺激后，MEK/ERK途径在造血干/祖细胞（HSPC）中过度激活。此外，需要遗传改变的HSC来启动和维持MPD表型，从而充当MPD启动细胞。有趣的是，与HSPC和其他祖细胞不同，我们发现NRAS G12D信号传导以剂量依赖性方式在HSC中组成性过度激活ERK1/2。这种异常信号传导与NRAS G12D/+ HSC的自我更新能力的提高有关，但减少了NRAS G12D/G12D HSC的自我更新和精疲力尽。一致地，微阵列分析将MEK/ERK途径确定为两组HSC中最强烈影响的功能基因本体学类别。总的来说，这些研究提出了MEK/ERK途径的过度激活以剂量依赖性的方式调节HSC的自我更新和分化的可能性，并且是导致MPD的关键步骤。为了支持这一假设，MEK抑制剂AZD6244挽救了NRAS G12D/G12D HSC的精疲力尽，有效地控制了MPD表型，并显着延长了患病小鼠的存活。为了进一步检验我们在体内的假设，我们产生了一种新型的有条件的MEK1敲击等位基因携带点突变，该突变模仿丝氨酸218和丝氨酸222（LSL MEK1 S218; 222E）。 MEK1 S218; 222E是MEK1的组成型活跃形式。作为控制造血的新型信号传导机制的长期目标的一部分，该提案的具体目的是：1）确定MEK/ERK途径的过度激活是否需要NRAS G12D介导的MPD MPD表型； 2）确定NRAS G12D信号是否通过MEK/ERK途径的剂量依赖性激活来调节HSC自我更新和分化； 3）确定抑制MEK1/2和JAK2是否阻止NRAS G12D/G12D介导的MPD及其转化为爆炸危机。总之，这两个特定目标中提出的实验将提供有关MEK/ERK信号传导在造血中的核心作用的见解。这些见解不仅会使我们更加了解遗传改变的HSC在肿瘤发生中的作用，而且还提高了对由功能失调的RAS/RAF/MEK信号引起的对发育和骨髓增生性疾病的基本理解。