p21-Activated Kinases as New Therapeutic Targets in Neurofibromatosis Type 1

描述（申请人提供）：1型神经纤维瘤病（NF 1）是一种相对常见的遗传性疾病综合征，由NF 1基因的种系突变引起。大约三分之一的NF 1患者发展为弥漫性丛状神经纤维瘤，可以转化为恶性外周神经鞘瘤-一种经常致命的癌症。值得注意的是，在人类肿瘤和NF 1小鼠模型中，神经纤维瘤几乎总是含有NF 1-null雪旺细胞和NF 1杂合肥大细胞。用野生型骨髓移植这种NF 1易感小鼠可防止肿瘤发生，这意味着骨髓来源的细胞如肥大细胞是发病机制中所需的组分，并且靶向施旺细胞或肥大细胞中的信号传导途径可能具有治疗益处. NF 1基因编码一种对Ras具有GTP酶激活蛋白（GAP）活性的大蛋白。NF 1基因的完全或半合子缺失导致雪旺细胞和肥大细胞中Ras活性增加，伴随着下游效应物的激活，其促进增殖和细胞形状和运动的变化。最近，我们已经表明，p21激活的激酶在激活ERK介导的增殖和p38介导的运动途径下游的Ras在Nf 1缺陷的肥大细胞中发挥重要作用。我们假设Pak功能的丧失也会减少NF 1缺陷的雪旺细胞中关键Ras效应通路的激活，从而使NF 1患者受益。我们提出三个目标：1）我们将鉴定肥大细胞中影响细胞运动性的A组Paks的关键底物：2）使用药理学和遗传学方法使Nf 1缺陷小鼠来源的雪旺细胞中的Paks失活，我们将确定Pak功能的丧失是否逆转体外MAPK的激活和细胞骨架的改变;以及3）我们将使发展恶性外周神经鞘瘤的Krox 20-cre/Nf 1flox/-小鼠与Pak 1敲除小鼠杂交，或将这种NF 1小鼠与Pak 1-/-骨髓细胞移植，以确定Pak 1的缺失，总体上或在骨髓来源的细胞中，影响疾病进展。拟议的研究不仅将增加我们对主要癌症相关信号通路的理解，而且可以将Paks作为这种无法治愈的疾病的治疗干预的合适靶点。 公共卫生相关性：p21激活激酶（Paks）是Ras下游信号传导的关键调节因子，Ras是一种因NF 1基因缺失而激活的蛋白质。目前，对于NF 1相关的恶性肿瘤没有有效的治疗方法。我们已经开发了用于研究Pak在动物中的功能的遗传模型，以及这些酶的第一个特异性化学抑制剂;由于这些原因，我们处于独特的位置，以探索Paks在NF 1中的生物学作用，并确定这些酶是否代表合适的治疗靶点。

DESCRIPTION (provided by applicant): Neurofibromatosis type 1 (NF1) is a relatively common inherited disease syndrome caused by germline mutations in the NF1 gene. About one third of NF1 patients develop diffuse, plexiform neurofibromas that can transform to a malignant peripheral nerve sheath tumor - a cancer that is frequently fatal. Remarkably, in human tumors and in mouse models of NF1, neurofibromas almost invariably contain Nf1-null Schwann cells and Nf1 heterozygous mast cells. Transplanting such NF1-prone mice with wild- type bone marrow prevents tumorigenesis, implying that bone marrow derived cells such as mast cells are a required component in pathogenesis, and that targeting signaling pathways in either Schwann cells or mast cells might be of therapeutic benefit. The NF1 gene encodes a large protein with GTPase Activating Protein (GAP) activity towards Ras. Complete or hemizygous loss of the NF1 gene leads to increased Ras activity in both Schwann cells and mast cells, with concomitant activation of downstream effectors that promote proliferation and changes in cell shape and movement. Recently, we have shown that p21-activated kinases play an important role in activating an Erk-mediated proliferation and a p38- mediated motility pathway downstream of Ras in Nf1-deficient mast cells. We postulate that loss of Pak function will diminish activation of key Ras effector pathways in Nf1-deficient Schwann cells as well, and thus could benefit patients with NF1. We propose three aims: 1) We will identify the key substrates of group A Paks in mast cells that affect cell motility; 2) Using pharmacologic and genetic means to disable Paks in Schwann cells derived from Nf1-deficient mice, we will determine if loss of Pak function reverses the activation of MAPK and cytoskeletal alterations in vitro; and 3) We will cross Krox20-cre/Nf1flox/- mice, which develop malignant peripheral nerve sheath tumors, with Pak1 knock out mice, or transplant such NF1 mice with Pak1-/- bone marrow cells, to determine if loss of Pak1, globally or in bone marrow derived cells, affects disease progression. The proposed studies will not only increase our understanding of cardinal cancer-relevant signaling pathways, but could establish the Paks as suitable targets for therapeutic intervention in this otherwise untreatable disease. PUBLIC HEALTH RELEVANCE: p21-activated kinases (Paks) are key regulators of signaling downstream of Ras, a protein that is activated as a result of loss of the NF1 gene. Currently, there are no effective therapies for NF1-related malignancies. We have developed genetic models for studying Pak function in animals, as well as the first specific chemical inhibitor of these enzymes; for these reasons, we are in a unique position to explore the biological role of Paks in NF1 and to determine if these enzymes represent suitable targets for therapy.