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疾病（1500 - 2500例活产婴儿中有1例），由PTPN11（50%）、SOS1（20%）或KRAS（<5%）突变引起，以面部畸形、先天性心脏畸形、骨骼异常和各种血液学异常为特征，包括易患幼年髓细胞白血病（JMML）。虽然在NS中观察到的异常是多种多样的，但一些并发症可归因于巨噬细胞或单核细胞来源细胞的功能或数量增加。NS患者巨噬细胞过度产生的最极端和致命的例子是儿童白血病（JMML）。JMML的临床特征是骨髓单核细胞的过量产生和造血祖细胞对粒细胞-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phospho-Erk水平升高外，我们发现激活Shp2突变（Shp2E76K和Shp2D61Y）促进了以牺牲其他髓系谱系为代价的向单核细胞分化的转变，这与在JMML患者中观察到的疾病表型一致。这种向单核细胞谱系倾斜的分化与c-Jun的增加和GATA2表达的减少有关。基于这些发现，我们假设激活shp2诱导的Ras过度激活会改变转录谱，从而以牺牲替代髓系谱系为代价增强单核细胞分化。在机制上，我们提出Ras过度激活产生组成型c-Jun表达，与PU.1合作，允许过度的单核细胞分化和降低GATA2表达。该应用的目的是：1)描述JMML相关的Shp2功能获得突变是否诱导c-Jun表达，并以jnk依赖或jnk独立的方式促进异常单核细胞分化；2)研究组蛋白去乙酰化酶抑制剂曲古斯汀A降低C - jun和PU.1以及重新激活GATA2和C/EBP的潜力。表达shp2突变体造血祖细胞中的表达；3)检测人JMML样本中与正常对照相比的造血特异性转录因子水平。我们期望这些研究的结果将为JMML（一种致命的儿童白血病和努南综合征并发症）的治疗带来新的策略。公共卫生相关性：这项名为“Shp2功能获得突变体诱导的异常单核细胞分化”的提案关注的是一种特别致命的儿童髓性白血病，即幼年髓性单核细胞白血病（JMML）。JMML是一种5岁以下儿童的致命白血病，其特点是单核细胞大量过量产生。不幸的是，JMML对化疗具有耐药性，大多数患病儿童由于恶性单核细胞和巨噬细胞的器官浸润而死于疾病，最终以出血和感染而告终。本提案中概述的实验旨在确定增强单核细胞过度生产的分子机制，以描绘JMML治疗干预的新靶点。

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.