Using VEGF expression in inflammatory arthritis to induce targeted apoptosis

描述（由申请人提供）：风湿性关节炎（RA）涉及滑膜的慢性炎症和增殖，其部分原因是致病效应细胞的凋亡缺陷。不适当的低细胞凋亡可能有助于类风湿关节炎中持续的滑膜细胞增殖。在炎性关节炎的动物模型中，Fas受体的激动剂活化改善疾病，然而挑战是仅在致病细胞中特异性诱导Fas介导的凋亡。RA患者具有高水平的由滑膜细胞产生的血管内皮生长因子（VEGF），而正常细胞中未发现。我们的应用将利用炎症滑膜分泌的高局部浓度的VEGF直接靶向Fas激动剂，以诱导这些部位的细胞凋亡。我们开发了一种新的融合蛋白与Fas配体连接到VEGF结合域（R1 FasL）。我们发现，R1FasL触发细胞凋亡和细胞毒性，只有与聚集的VEGF。我们的申请将检查VEGF对炎症和炎症性关节炎中细胞凋亡的影响，并利用新型R1FasL融合蛋白治疗炎症性关节炎，以确定我们是否可以选择性地消除产生VEGF的区域中的致病细胞。我们的具体目标是：1）测定VEGF和VEGF受体在鼠炎性关节炎模型SKG和胶原诱导的关节炎（CIA）中的表达。2)测定VEGF诱导的细胞因子，有或没有Fas配体刺激炎症性关节炎小鼠的滑膜细胞、巨噬细胞和破骨细胞。3)测定VEGF对Fas介导的滑膜细胞、巨噬细胞和破骨细胞凋亡的影响。4)使用炎性关节炎模型SKG和CIA确定R1 FasL治疗的效果。 公共卫生相关性：风湿性关节炎是一种慢性和使人衰弱的炎症性疾病，尽管治疗取得了重大进展，但由于缺乏可用疗法的反应或副作用，患者继续患有活动性疾病。我们的应用程序调查VEGF或血管内皮生长因子在这种疾病的发病机制中的作用，并利用滑膜细胞VEGF的异常产生来选择性地去除这些致病细胞。我们的研究将进一步了解炎症性关节炎的发病机制，并在这种疾病的动物模型中研究一种新型的治疗方法。

DESCRIPTION (provided by applicant): Rheumatoid arthritis (RA) involves chronic inflammation and proliferation of the synovium, which is in part due to defective apoptosis of pathogenic effector cells. Inappropriately low apoptosis likely contributes to persistent synovial cell proliferation in rheumatoid arthritis. In animal models of inflammatory arthritis, agonist activation of the Fas Receptor improves disease, however a challenge is to specifically induce Fas-mediated apoptosis only in pathogenic cells. RA patients have high levels of Vascular Endothelial Growth Factor (VEGF) produced by synoviocytes, not found with normal cells. Our application will exploit the high local concentrations of VEGF secreted by inflamed synovium to directly target a Fas agonist to induce apoptosis at those sites. We developed a novel fusion protein with Fas ligand linked to a VEGF binding domain (R1FasL). We show that R1FasL triggers apoptosis and is cytotoxic only with clustering by VEGF. Our application will examine VEGF effects on inflammation and apoptosis in inflammatory arthritis and utilize the novel R1FasL fusion protein as a treatment for inflammatory arthritis to determine if we can selectively eliminate pathogenic cells in areas where VEGF is produced. Our specific aims are: 1) Determine VEGF and VEGF receptor expression in the murine inflammatory arthritis models SKG and collagen induced arthritis (CIA). 2) Determine cytokines induced by VEGF with or without Fas ligand stimulation of synoviocytes, macrophages and osteoclasts from mice with inflammatory arthritis. 3) Determine the effect of VEGF on Fas-mediated apoptosis of synoviocytes, macrophages and osteoclasts. 4) Determine the effect of R1FasL treatment using the inflammatory arthritis models SKG and CIA. PUBLIC HEALTH RELEVANCE: Rheumatoid Arthritis is a chronic and debilitating inflammatory disease and despite significant advances in treatment, patients continue with active disease due to a lack of response or side effects with available therapies. Our application investigates the role of VEGF or vascular endothelial growth factor in pathogenesis of this disease and utilizes the abnormal production of VEGF by synovial cells to selectively remove these pathogenic cells. Our studies will further our understanding of disease pathogenesis in inflammatory arthritis and examine a novel type of therapeutic in animal models for this disease.