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

描述(申请人提供)：神经纤维瘤病1型(NF1)是一种相对常见的遗传性疾病综合征，由NF1基因的胚系突变引起。大约三分之一的NF1患者发展为弥漫性丛状神经纤维瘤，可能会转变为恶性周围神经鞘瘤--一种经常致命的癌症。值得注意的是，在人类肿瘤和NF1小鼠模型中，神经纤维瘤几乎总是包含NF1缺失的雪旺细胞和NF1杂合肥大细胞。将这种倾向于NF1的小鼠移植到野生型骨髓中可以防止肿瘤的发生，这意味着骨髓来源的细胞，如肥大细胞是发病过程中必需的组成部分，并且靶向雪旺细胞或肥大细胞中的信号通路可能对治疗有益。NF1基因编码一个对RAS具有GTP酶激活蛋白活性的大蛋白。NF1基因的完全或半合子缺失会导致雪旺细胞和肥大细胞中RAS活性增加，伴随而来的是下游效应因子的激活，从而促进细胞增殖和细胞形状和运动的变化。最近，我们发现p21激活的激酶在NF1缺陷的肥大细胞中激活ERK介导的增殖和p38介导的RAS下游的运动通路中发挥重要作用。我们推测，Pak功能的丧失也会减少NF1缺陷的雪旺细胞中关键的RAS效应通路的激活，从而可能使NF1患者受益。我们提出了三个目标：1)我们将确定肥大细胞中影响细胞运动的A组PAK的关键底物；2)使用药理学和遗传学手段禁用来自NF1缺陷小鼠的施万细胞中的PAK，我们将确定Pak功能的丧失是否在体外逆转MAPK的激活和细胞骨架的变化；以及3)我们将Krox20-cre/Nf1flx/-发展为恶性周围神经鞘瘤的小鼠与Ak1基因敲除小鼠进行杂交，或将此类NF1小鼠与Ak1-/骨髓细胞移植，以确定Ak1基因缺失是否会影响疾病的进展。拟议的研究不仅将增加我们对与大癌相关的信号通路的了解，还可能建立PAK作为治疗这种原本无法治愈的疾病的合适靶点。 公共卫生相关性：p21激活的激酶(PAK)是RAS下游信号的关键调节者，RAS是一种因NF1基因丢失而被激活的蛋白质。目前，还没有有效的治疗NF1相关恶性肿瘤的方法。我们已经开发了用于研究动物体内PAK功能的遗传模型，以及第一个这些酶的特定化学抑制剂；出于这些原因，我们处于独特的地位，可以探索PAK在NF1中的生物学作用，并确定这些酶是否代表了合适的治疗靶点。

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.