The Role of SPARC in Neuroblastoma Angiogenesis

描述(申请人提供)：儿童癌症神经母细胞瘤(NB)具有生物学和临床异质性。神经母细胞瘤包括神经母细胞/神经节细胞和雪旺氏间质。丰富的Schwannian间质的存在有利于影响预后，与肿瘤血管呈负相关。雪旺细胞和分化的NB肿瘤细胞产生一系列血管生成抑制物，包括金属蛋白酶组织抑制物-2(TIMP-2)和色素上皮衍生因子(PEDF)。最近，层析研究发现了另一种雪旺细胞来源的分泌型血管生成抑制因子，分泌型酸性蛋白和富含半胱氨酸的蛋白(SPARC)。体内外研究表明，SPARC能有效抑制血管生成，诱导内皮细胞凋亡。我们推测，雪旺细胞分泌的一系列血管生成抑制物导致神经母细胞瘤的低血管水平和更良性的临床行为，而SPARC是雪旺细胞分泌的抗血管生成活性的关键贡献者。在目前的提案中，将研究SPARC在调控NB血管生成中所起的作用。第一个具体目的是检测SPARC对肿瘤血管和体内生长的影响。我们计划扩大和延长我们的初步临床前研究，这些研究表明SPARC抑制NB血管生成。SPARC将用于皮下移植NB的动物以及使用ALZET渗透泵的原位NB裸鼠模型。还将进行其他实验，以测试外源表达的SPARC对肿瘤生长的影响。第二个具体目标是生产和鉴定重组SPARC，以及具有血管生成抑制活性的SPARC结构域和多肽。被发现具有抗血管生成活性的重组蛋白和多肽将在临床前模型中进一步评估。这项资助的第三个具体目标是描述SPARC诱导细胞凋亡的机制，并分析其对内皮细胞功能的影响。这项资助中概述的研究将有助于进一步了解NB中血管生成的复杂调控，并为SPARC在确定NB表型中所起的作用提供洞察力。这项研究拨款的长期目标是产生有效的抗血管生成SPARC衍生分子，这些分子将被证明对患有高度血管性、临床侵袭性NB的儿童有效。

DESCRIPTION (provided by applicant): The pediatric cancer neuroblastoma (NB) is biologically and clinically heterogeneous. NB tumors contain both neuroblastic/ganglionic cells and Schwannian stroma. The presence of abundant Schwannian stroma favorably impacts prognosis and is inversely correlated with tumor vascularity. Schwann cells as well as differentiated NB tumor cells produce a spectrum of angiogenesis inhibitors including tissue inhibitor of metalloproteinase-2 (TIMP-2) and pigment epithelium-derived factor (PEDF). Chromatography studies have recently led to the identification of another Schwann cell-derived secreted inhibitor of angiogenesis, Secreted Protein Acidic and Rich in Cysteine (SPARC). In vitro and in vivo studies show that SPARC potently inhibits angiogenesis, and that SPARC induces endothelial cell apoptosis. We hypothesize that the low level of vascularity and more benign clinical behavior of NB tumors with abundant Schwannian stroma results from the Schwann cell production of a spectrum of angiogenesis inhibitors, and that SPARC is a key contributor to the anti-angiogenic activity of factors secreted by the Schwann cells. In the current proposal, the role SPARC plays in the regulation of NB angiogenesis will be investigated. The first specific aim is to examine the effects of SPARC on NB tumor vascularity and growth in vivo. We plan to expand and extend our preliminary preclinical studies that suggest that SPARC inhibits NB angiogenesis. SPARC will be administered to animals with subcutaneous NB xenografts as well as an orthotopic NB nude mouse model using ALZET osmotic pumps. Additional experiments will be performed testing the effects of exogenously expressed SPARC on tumor growth. The second specific aim is to produce and characterize recombinant SPARC, and SPARC domains and peptides that confer angiogenesis inhibitory activity. Recombinant proteins and peptides found to have anti-angiogenesis activity will be further evaluated in the preclinical models. The third specific aim of this grant is to delineate the mechanisms by which SPARC induces apoptosis and analyze its effects on endothelial cell function. The studies outlined in this grant will lead to a further understanding of the complex regulation of angiogenesis in NB, and provide insight into the role that SPARC plays in determining NB phenotype. The long-term goal of this research grant is to generate potent anti-angiogenic SPARC-derived molecules that will prove to be effective in the treatment of children with highly vascular, clinically aggressive NB.