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: 8666589
• 负责人: Jing Zhang
• 金额: $ 36.48万
• 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/8666589
• 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(申请人提供)：MEK/ERK是RAS下游的一个主要信号通路。最近，我们描述了人类骨髓增生性疾病(MPD)的条件小鼠模型，这些模型分别在造血室表达一个或两个致癌NRAS(NRAS G12D/+和NRAS G12D/G12D)。在两种MPD模型中，在细胞因子刺激下，MEK/ERK通路在造血干/祖细胞(HSPC)中被过度激活。此外，基因改变的HSCs需要启动和维持MPD表型，从而作为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信号以剂量依赖的方式结构性地过度激活HSC中的ERK1/2。这种异常信号与NRAS G12D/+HSCs的自我更新能力增强有关，而与NRAS G12D/G12D HSCs的自我更新和耗竭能力降低有关。微阵列分析一致地将MEK/ERK通路确定为两组HSC中受影响最大的功能基因本体论类别。总之，这些研究提出，MEK/ERK通路的过度激活以剂量依赖的方式调节HSCs的自我更新和分化，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 HSCs的耗竭，有效地控制了MPD表型，并显著延长了患病小鼠的生存时间。为了在体内进一步验证我们的假设，我们产生了一个新的条件性MEK1敲击等位基因，该等位基因携带模拟丝氨酸218和丝氨酸222处的磷酸化的点突变(LSL MEK1 S218；222E)。MEK1 S218；222E是MEK1的一种结构性活性形式。作为我们阐明控制造血的新的信号机制的长期目标的一部分，本建议的具体目标是：1)确定MEK/ERK通路的过度激活对于NRAS G12D介导的MPD表型是否必要且充分；2)确定NRAS G12D信号是否通过剂量依赖的MEK/ERK通路激活来调节HSC的自我更新和分化；3)确定抑制MEK1/2和JAK2是否阻止NRAS G12D/G12D介导的MPD及其向爆炸性危机的转化。总之，在这两个特定目标中提出的实验将为深入了解MEK/ERK信号在造血中的核心作用提供见解。这些见解不仅将使我们更接近于了解基因改变的HSCs在肿瘤发生中的作用，而且还将增进对由Ras/Raf/MEK信号异常引起的发育性和骨髓增生性疾病的基本理解。