Ref-1 in Retinal Neovascularization

项目总结 视网膜新生血管损害视觉功能，是几种新生血管性眼病的标志， 包括早产儿视网膜病变(ROP)和增殖性糖尿病视网膜病变(PDR)。糖尿病视网膜病变 (DR)是劳动年龄成年人视力丧失的主要原因，ROP是可预防的主要原因 儿童失明。目前的治疗方法包括玻璃体内(IVT)注射抗血管内皮生长药物 因子(血管内皮生长因子)生物制品。然而，这些疗法往往伴随着较高的治疗负担和 对治疗的抗拒。先前的研究表明，仅有血管内皮生长因子不足以诱导 新生血管，表明多条与疾病相关的途径可能被靶向增加 治疗反应。因此，迫切需要开发新的疗法来调节多种疾病- 相关途径和绕过静脉注射的问题。先前的研究表明，APE1/Ref-1，a 同时具有核酸内切酶(APE1)和氧化还原活性(Ref-1)的多功能蛋白，调节 与视网膜新生血管有关的转录因子。激光诱导脉络膜的初步数据 新生血管模型显示Ref-1在新生血管形成过程中上调， 给予小分子Ref-1氧化还原抑制剂可降低疾病严重程度。人视网膜的RNA序列 内皮细胞显示，Ref-1氧化还原抑制剂下调Wnt信号基因，这意味着Ref-1 氧化还原功能通过Wnt信号调节新生血管形成。但是，潜在的分子 新的REF-1靶点在视网膜新生血管形成中的作用机制尚未阐明。识别 REF-1在视网膜新生血管中的分子机制是开发新的治疗方法的关键 减少视力损失。这一提议的假设是Ref-1氧化还原活性促进视网膜的诱导 通过激活Wnt信号通路和降低Ref-1氧化还原活性的新方法促进新生血管形成 REF-1氧化还原抑制剂将通过抑制Wnt信号通路发挥有效的治疗作用。目标1将 氧诱导视网膜病变视网膜新生血管中Ref-1的时空表达分析 (OIR)小鼠模型、Vldlr-/-小鼠模型和人PDR组织中。目标2将分析其作用机制 REF-1对视网膜内皮细胞Wnt通路的调节作用目标3将 口服Ref-1抑制剂在OIR小鼠体内的抗血管生成作用 建立新生血管模型并评估其变化。对Wnt信号通路的影响也将是 用免疫荧光法进行评估。这项研究的完成将加强对Ref-1的了解 并提供了一种新的治疗策略来对抗因以下原因造成的视力丧失和失明 这些疾病。

PROJECT SUMMARY Retinal neovascularization impairs visual function and is a hallmark of several neovascular eye diseases, including retinopathy of prematurity (ROP) and proliferative diabetic retinopathy (PDR). Diabetic retinopathy (DR) is the leading cause of vision-loss in working age adults, and ROP is the leading cause of preventable blindness in children. Current treatments include intravitreal (IVT) injections of anti-vascular endothelial growth factor (VEGF) biologics. However, these therapeutics are often accompanied by high treatment burden and resistance to therapy. Previous research indicates that VEGF alone is not sufficient for induction of neovascularization, suggesting that multiple disease-relevant pathways may be targeted to increase therapeutic response. Thus, there is a critical need to develop novel therapies that modulate multiple disease- relevant pathways and circumvent the issues with IVT injections. Prior studies indicate that APE1/Ref-1, a multifunctional protein with both endonuclease (APE1) and redox activity (Ref-1), regulates multiple transcription factors that are linked to retinal neovascularization. Preliminary data in the laser-induced choroidal neovascularization model demonstrated upregulation of Ref-1 during neovascularization, and oral administration of a small molecule Ref-1 redox inhibitor decreased disease severity. RNA-seq of human retinal endothelial cells revealed that a Ref-1 redox inhibitor downregulated Wnt signaling genes, implying that Ref-1 redox function modulates neovascularization through the Wnt signaling. But, the underlying molecular mechanisms of the novel Ref-1 target in retinal neovascularization have yet to be elucidated. Identifying the molecular mechanism underlying Ref-1 in retinal neovascularization is critical in developing novel therapies to reduce vision loss. The hypothesis of this proposal is that Ref-1 redox activity promotes induction of retinal neovascularization via activation of the Wnt signaling pathway, and reducing Ref-1 redox activity with a novel Ref-1 redox inhibitor will exhibit potent therapeutic effects via inhibition of the Wnt signaling pathway. Aim 1 will analyze the spatiotemporal expression of Ref-1 in retinal neovascularization in the oxygen-induced retinopathy (OIR) mouse model, the Vldlr-/- mouse model, and in human PDR tissue. Aim 2 will analyze the mechanism of Ref-1 regulation of the Wnt pathway in retinal endothelial cells through functional in vitro assays. Aim 3 will assess anti-angiogenic effects of a Ref-1 inhibitor in vivo by administering oral Ref-1 inhibitor in the OIR mouse model and evaluating changes in neovascularization. Effects on the Wnt signaling pathway will also be assessed using immunofluorescence. Completion of this study will bolster understanding of Ref-1 in neovascular eye diseases and provide a novel therapeutic strategy to combat vision loss and blindness due to these diseases.