Using VEGF expression in inflammatory arthritis to induce targeted apoptosis

描述（由申请人提供）：类风湿性关节炎（RA）涉及滑膜的慢性炎症和增殖，部分原因是致病性效应细胞凋亡缺陷。不适当的低细胞凋亡可能有助于类风湿性关节炎滑膜细胞的持续增殖。在炎性关节炎的动物模型中，Fas受体的激动剂激活可改善疾病，然而，仅在致病细胞中特异性诱导Fas介导的细胞凋亡是一个挑战。RA患者有高水平的滑膜细胞产生的血管内皮生长因子（VEGF），正常细胞中没有发现。我们的应用将利用炎症滑膜分泌的高浓度VEGF直接靶向Fas激动剂诱导这些部位的细胞凋亡。我们开发了一种新的与VEGF结合域（R1FasL）连接的Fas配体融合蛋白。我们发现R1FasL触发细胞凋亡，仅在VEGF聚集的情况下具有细胞毒性。我们的申请将检查VEGF对炎症性关节炎的炎症和细胞凋亡的影响，并利用新型R1FasL融合蛋白作为炎症性关节炎的治疗方法，以确定我们是否可以选择性地消除VEGF产生区域的致病细胞。我们的具体目标是：1)测定小鼠炎性关节炎模型SKG和胶原诱导关节炎（CIA）中VEGF和VEGF受体的表达。2)测定VEGF伴或不伴Fas配体刺激小鼠炎性关节炎滑膜细胞、巨噬细胞和破骨细胞诱导的细胞因子。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)通过炎症性关节炎模型SKG和CIA确定R1FasL治疗的效果。

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.