Gene Therapy in the Cornea

描述(申请人提供)：角膜疾病是全球失明的主要原因，每年影响150万美国人。组织靶向和局部的角膜基因疗法有望治愈、减少和/或预防由创伤、损伤、感染或遗传性营养不良引起的角膜失明。我们研究的长期目标是确定临床适用的、简单的、微创的、基于基因的治疗角膜失明的方法。最近，我们证明了靶向腺相关病毒(AAV)-Decorin基因治疗有效地抑制了兔体内的角膜纤维化和新生血管。但它并没有 是否有任何短期毒性。我们的中心假设是核心蛋白聚糖是角膜愈合的主要调节因子，作为信号分子，它与许多细胞因子和/或它们的受体相互作用，防止角膜过度愈合和疾病。我们发现角膜中的核心蛋白与转化生长因子相互作用？(？)、表皮生长因子受体(EGFR)和血管内皮细胞生长因子(VEGF)。我们的RNA敲除研究显示，转化生长因子？？Smad7通过Smad信号介导角膜纤维化，Smad7是一个有吸引力的阻断转化生长因子？？的靶点。病理活动。(2)验证靶向AAV-Smad7基因治疗是一种干扰转化生长因子的新方法的假设？在体内传递和防止角膜中肌成纤维细胞的形成和纤维化。(3)体内组织选择性AAV-PEDF基因治疗对兔角膜新生血管的抑制作用，且无明显不良反应。他们还将提供许多关于角膜愈合的基本问题的答案，以及角膜细胞和细胞因子在角膜疾病和疾病中的作用。 公共卫生相关性：创伤、损伤或感染后的角膜疤痕和新生血管会导致失明。通过一种简单的局部技术将靶向基因疗法输送到角膜，通过调节导致这些致盲角膜疾病的过度愈合机制，有望治愈、治疗和/或防止视力丧失。

DESCRIPTION (provided by applicant): Corneal disorders are the leading cause of global blindness and affect 1.5 million Americans each year. Tissue-targeted and localized gene therapy for the cornea offers promise to cure, reduce and/or prevent corneal blindness caused by trauma, injury, infection or inherited dystrophies. The long-range goal of our research is to define clinically applicable, simple, minimally invasive targeted gene-based therapies for corneal blindness. Recently, we demonstrated that targeted adeno-associated virus (AAV)-decorin gene therapy efficiently inhibits corneal fibrosis and neovascularization in vivo in rabbits. It did not show any short-term toxicity. Our central hypothesis is that decorin is a master regulator of corneal healing and, acting as a signaling molecule, it interacts with many cytokines and/or their receptors and prevents excessive healing and disease in the cornea. We found decorin in the cornea interacts with transforming growth factor ? (TGF??), epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF). Our RNA knockdown studies revealed that TGF?? mediates corneal fibrosis via Smad signaling, and Smad7 is an attractive target to block TGF?? pathologic activity. The goals of this project are: i) test whether efficacious AAV-decorin gene therapy has any long-term toxicity for the cornea, ii) collect cornea-specific data and an understanding of the molecular mechanism of decorin functions in the cornea, and iii) identify additional gene-based therapy for corneal blindness. One treatment does not work for all patients due to variations in medical conditions, susceptibility to medication, tolerance level etc. The Specific Aims are: (1) Test the hypothesis that targeted decorin delivery into rabbit keratocytes in vivo does not alter fibril structure or characteristic organization in the stroma, modulates keratocyte differentiation, migration and proliferation, and regulates corneal healing. (2) Test the hypothesis that targeted AAV-Smad7 gene therapy is a novel approach to disrupt TGF? signaling and prevent myofibroblast formation and fibrosis in the cornea in vivo. (3) Test the hypothesis that tissue-selective AAV-PEDF gene therapy delivered into rabbit keratocytes in vivo inhibits corneal neovascularization in rabbits without significant adverse effects. These aims will be tested using well-defined rabbit disease models, recently defined targeted AAV therapy approaches, established protocols and techniques, including slit-lamp, stereomicroscope and optical coherence tomography, immunohistochemistry, immunoblotting, quantitative PCR, southern blot, transmission electron and confocal microscopy, etc. The proposed experiments will provide important answers to how AAV- mediated decorin, Smad7 and PEDF gene therapy may be used, alone and in combination, to restore vision or prevent blindness from corneal scarring or neovascularization. They also will provide answers to many fundamental questions about corneal healing, and the role of keratocyte and cytokines in corneal disorders and diseases. PUBLIC HEALTH RELEVANCE: Corneal scarring and neovascularization following trauma, injury or infection cause blindness. Targeted gene therapy delivered to the cornea with a simple topical technique offers promise to cure, treat and/or prevent vision loss by modulating excessive healing mechanisms that cause these blinding corneal disorders.