Tyrosine Phosphatases in Endothelial Growth Control

描述(申请人提供)：调节失调的毛细血管和动脉生长对我们的健康有重大影响，并导致许多恶性、缺血性和炎症性疾病。对调节血管新生血管生长的内在分子控制的定义有望为包括心脏病、糖尿病和癌症在内的各种疾病提供新的治疗方法。尽管以前的研究已经确定了诱导血管形成的内皮受体，但对传递“血管抑制”信号的功能性内皮受体知之甚少。我们提出的假设是，受体酪氨酸磷酸酶(RPTPs)通过与对抗性受体的旁分泌接触，在细胞-细胞接触时发出内皮生长停止的信号。我们从人肾微血管内皮细胞中分离到一种RPTP，CD148(DEP-1/PTPq)，结果表明：1)CD148在发育和成熟的各种器官的血管内皮细胞中大量表达，2)CD148积聚在内皮细胞间，其活性随着细胞密度的增加而增加，3)CD148纯合子突变小鼠在孕中期死于血管形成障碍，伴随着内皮血管生长和内皮细胞增殖的紊乱，4)针对胞外结构域序列的CD148单抗具有显著的抗血管生成活性，抑制培养的内皮细胞的增殖，阻断体内血管生成，5)CD148通过胞外结构域(分子间结合)相互作用，强迫CD148二聚化抑制细胞增殖；6)CD148与成纤维细胞生长因子受体结合；7)CD148胞浆结构域与PI3激酶p85亚单位和p120连环蛋白分子相互作用并去磷酸化。这些发现表明CD148在调节细胞-CEU接触时内皮细胞的生长中起着关键作用。为了进一步确定CD148在血管形成中的作用，本文描述了以下实验方法：1)定位负责介导分子间联系的细胞外亚结构域，并阐明其在调节CD148活性和密度介导的内皮细胞生长停滞中的重要性；2)确定CDt48与成纤维细胞生长因子受体、p85和p120连环蛋白之间的分子相互作用在内皮生长控制中的生物学意义；以及3)确定CD148在内皮血管形成中的作用。这些互补的努力将确定一条通过CD148调节内皮细胞生长的新途径，并有望成为抗血管生成治疗的新策略。

DESCRIPTION (provided by applicant): Dysregulated capillary and arterial growth has a major impact on our health and contributes to numerous malignant, ischemic and inflammatory disorders. Definition of the intrinsic molecular controls that regulate angiogenic blood vessel growth promises novel therapeutic approaches for a variety of diseases including heart disease, diabetes and cancer. Although previous works have identified the endothelial receptors that induce vessel formation, little is known about the functional endothelial receptors that transduce "angiostatic" signals. We have advanced the hypothesis that receptor tyrosine phosphatases (RPTPs) signal endothelial growth arrest upon cell-cell contacts, through juxtacrine contact with counter-receptors. We have isolated a RPTP, CD148 (DEP-1/PTPq), from human renal microvascular endothelial cells and shown that: 1) CD148 is abundantly expressed in developing and mature vascular endothelium of various organs, 2) CD148 is accumulated at interendothelial sites and its activity increases with cell density, 3) the CD148 homozygous mutant mice die at mid-gestation due to vascularization failure accompanied by disordered endothelial vessel growth and endothelial cell proliferation, 4) a CD148 monoclonal antibody against the ectodomain sequence has a prominent anti-angiogenic activity to inhibit proliferation of culture endothelial cells and block angiogenesis in vivo, 5) CD148 interacts through the ectodomain (inter-molecular association) and forced CD148 dimerization inhibits cell proliferation, 6) CD148 constitutively associates with FGF receptor, and 7) CD148 cytoplasmic domain interacts with- and dephosphorylates the PI3 kinase p85 subunit and p120 catenin molecules. These findings suggest a pivotal role of CD148 to regulate endothelial cell growth upon cell-ceU contacts. To further define role of CD148 in blood vessel formation, the present proposal describes experimental approaches to: 1) map the extracellular subdomain responsible for mediating inter-molecular associations and elucidate its importance in regulation of CD148 activity and density-mediated endothelial growth arrest, 2) define biological importance of the molecular interaction between CDt48 and FGF receptor, p85, and p120 catenin in endothelial growth control, and 3) determine role of CD148 in endothelial vessel formation. These complementary efforts will identify a novel pathway that regulates endothelial cell growth through CD148 and promise new strategies for anti-angiogenesis therapy.