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

基本信息

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
项目类别：
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

项目摘要

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 下游的主要信号通路。最近，我们表征了人类骨髓增殖性疾病（MPD）的条件性小鼠模型，这些模型在造血室中分别表达一个或两个致癌Nras拷贝（Nras G12D / +和Nras G12D / G12D）。在两种 MPD 模型中，造血干细胞/祖细胞 (HSPC) 中的 MEK/ERK 通路在细胞因子刺激后过度激活。此外，基因改变的 HSC 是启动和维持 MPD 表型所必需的，因此可作为 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 信号以剂量依赖性方式持续过度激活 HSC 中的 ERK1/2。这种异常信号传导与 Nras G12D/+ HSC 的自我更新能力增强有关，但与 Nras G12D/G12D HSC 的自我更新能力降低和耗尽有关。一致地，微阵列分析将 MEK/ERK 通路确定为两组 HSC 中受影响最强烈的功能基因本体类别。总的来说，这些研究提出了这样的可能性：MEK/ERK 通路的过度激活以剂量依赖性方式调节 HSC 的自我更新和分化，并且是导致 MPD 的关键步骤。为了支持这一假设，MEK 抑制剂 AZD6244 可以挽救 Nras G12D/G12D HSC 的耗竭，有效控制 MPD 表型，并显着延长患病小鼠的生存期。为了进一步在体内验证我们的假设，我们生成了一种新型条件性 MEK1 敲入等位基因，其携带模拟丝氨酸 218 和丝氨酸 222 磷酸化的点突变 (LSL MEK1 S218;222E)。 MEK1 S218;222E 是 MEK1 的组成型活性形式。作为我们阐明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 表型是否是必要和充分的； 2) 确定Nras G12D信号是否通过MEK/ERK通路的剂量依赖性激活来调节HSC的自我更新和分化； 3)确定MEK1/2和Jak2的抑制是否阻断Nras G12D/G12D介导的MPD及其向急变危机的转变。总之，在这两个具体目标中提出的实验将深入了解 MEK/ERK 信号在造血中的核心作用。这些见解不仅将使我们更深入地了解基因改变的 HSC 在肿瘤发生中的作用，而且还能提高对 Ras/Raf/MEK 信号传导功能障碍引起的发育和骨髓增殖性疾病的基本了解。