Aberrant Monocytic Differentiation Induced by Gain-of-Function Shp2 Mutants

描述(由申请人提供)：努南综合征(NS)是一种常见的常染色体显性遗传病(每1500至2500名活产儿中就有一名)，由PTPN11(50%)、SOS1(20%)或KRAS(5%)突变引起，其特征是面部畸形、先天性心脏畸形、骨骼异常和各种血液异常，包括青少年粒单核细胞白血病(JMML)的易感性。虽然在NS中观察到的异常多种多样，但一些并发症可归因于巨噬细胞或单核细胞来源的细胞功能或数量的增加。NS患者中巨噬细胞过度产生的最极端和最致命的例子是儿童白血病，JMML。JMML的临床特征是粒单核细胞过度生成和造血祖细胞对粒-巨噬细胞集落刺激因子(GM-CSF)的体外表型超敏。除了GM-CSF刺激的过度增殖和磷酸化ERK水平升高外，我们发现激活Shp2突变(Shp2E76K和Shp2D61Y)促进了向单核细胞分化的转变，损害了其他髓系，这与在JMML患者中观察到的疾病表型一致。这种偏向于单核细胞系的分化与c-jun的增加和GATA2的表达减少有关。基于这些发现，我们假设激活Shp2诱导的RAS过度激活改变了转录谱，导致单核细胞分化增强，代价是不同的髓系。从机制上讲，我们认为RAS的过度激活产生了结构性的c-jun表达，与PU.1协同，允许过度的单核细胞分化和减少GATA2的表达。本申请的目的是1)阐明JMML相关Shp2功能获得突变是否以JNK依赖或非JNK依赖的方式诱导c-jun表达并促进异常单核细胞分化；2)研究组蛋白去乙酰酶抑制剂trichostatin A降低c-jun和PU.1并重新激活GATA2和C/EBP的可能性。在表达突变Shp2的造血祖细胞中的表达；以及3)与正常对照相比，检测JMML患者样本中的造血特异性转录因子水平。我们预计，这些研究的结果将导致JMML的新治疗策略，JMML是一种致命的儿童白血病，也是Noonan综合征的并发症。公共卫生相关性：这项名为“功能获得Shp2突变诱导的异常单核细胞分化”的提案关注于一种特别致命的儿童髓系白血病，称为幼年粒单核细胞白血病(JMML)。JMML是一种5岁以下儿童的致命性白血病，其特征是单核细胞大量过剩。不幸的是，JMML对化疗具有抵抗力，大多数儿童因器官恶性单核细胞和巨噬细胞浸润而死于疾病，最终导致出血和感染。本提案中概述的实验旨在确定促进单核细胞过度生产的分子机制，以便为JMML的治疗干预勾勒出新的靶点。

DESCRIPTION (provided by applicant): Noonan syndrome (NS) is a common (1 in 1500 to 2500 live births) autosomal-dominant disorder caused by mutations in PTPN11 (50%), SOS1 (20%), or KRAS (<5%) and is characterized by dysmorphic facial features, congenital heart malformations, skeletal anomalies, and a variety of hematologic abnormalities including a predisposition to juvenile myelomonocytic leukemia (JMML). Although the anomalies observed in NS are diverse, several of the complications can be attributed to the increased function or number of macrophages or monocyte-derived cells. The most extreme and lethal example of macrophage overproduction in NS patients is the childhood leukemia, JMML. JMML is characterized clinically by overproduction of myelomonocytic cells and by the in vitro phenotype of hematopoietic progenitor hypersensitivity to granulocyte-macrophage colony- stimulating factor (GM-CSF). In addition to GM-CSF-stimulated hyperproliferation and elevated levels of phospho-Erk, we found that activating Shp2 mutations (Shp2E76K and Shp2D61Y) promote a shift toward monocytic differentiation at the expense of other myeloid lineages, consistent with the disease phenotype observed in JMML patients. This skewed differentiation toward the monocytic lineage is associated with increased c-Jun and decreased GATA2 expression. Based on these findings, we hypothesize that activating Shp2-induced Ras hyperactivation alters the transcriptional profile leading to enhanced monocytic differentiation at the expense of alternative myeloid lineages. Mechanistically, we propose that Ras hyperactivation produces constitutive c-Jun expression permitting, in collaboration with PU.1, excessive monocytic differentiation and reduced GATA2 expression. The objectives of this application are to 1) delineate if JMML associated Shp2 gain-of-function mutations induce c-Jun expression and promote aberrant monocytic differentiation in a JNK-dependent or a JNK-independent manner; 2) investigate the potential of trichostatin A, a histone deacetylase inhibitor, to reduce c-Jun and PU.1 and to re-activate GATA2 and C/EBP? expression in mutant Shp2-expressing hematopoietic progenitors; and 3) examine hematopoietic- specific transcription factor levels in human samples of JMML compared to normal controls. We anticipate that results of these studies will lead to novel therapeutic strategies in JMML, a lethal childhood leukemia and complication of Noonan syndrome. PUBLIC HEALTH RELEVANCE: This proposal, "Aberrant Monocytic Differentiation Induced by Gain-of-Function Shp2 Mutants" focuses on a particularly lethal form of childhood myeloid leukemia called juvenile myelomonocytic leukemia (JMML). JMML is a lethal leukemia of children less than 5 years of age characterized by massive overproduction of monocytic cells. Unfortunately, JMML is resistant to chemotherapy and most afflicted children succumb to disease due to organ infiltration with malignant monocytes and macrophages, ending terminally in bleeding and infection. Experiments outlined in this proposal are directed at identifying the molecular mechanisms that enhance monocyte overproduction in order to delineate novel targets for therapeutic intervention in JMML.