Tissue-targeted and localized AAV5-DCN and AAV5-PEDF combination gene therapy abrogates corneal fibrosis and concurrent neovascularization in rabbit eyes in vivo

IF 5.9 1区医学 Q1 OPHTHALMOLOGY Ocular Surface Pub Date : 2024-01-06 DOI:10.1016/j.jtos.2024.01.001

Rajiv R. Mohan , Suneel Gupta , Rajnish Kumar , Nishant R. Sinha , James Landreneau , Prashant R. Sinha , Ashish Tandon , Shyam S. Chaurasia , Nathan P. Hesemann

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Abstract

Purpose

Corneal fibrosis and neovascularization (CNV) after ocular trauma impairs vision. This study tested therapeutic potential of tissue-targeted adeno-associated virus5 (AAV5) mediated decorin (DCN) and pigment epithelium-derived factor (PEDF) combination genes in vivo.

Methods

Corneal fibrosis and CNV were induced in New Zealand White rabbits via chemical trauma. Gene therapy in stroma was delivered 30-min after chemical-trauma via topical AAV5-DCN and AAV5-PEDF application using a cloning cylinder. Clinical eye examinations and multimodal imaging in live rabbits were performed periodically and corneal tissues were collected 9-day and 15-day post euthanasia. Histological, cellular, and molecular and apoptosis assays were used for efficacy, tolerability, and mechanistic studies.

Results

The AAV5-DCN and AAV5-PEDF combination gene therapy significantly reduced corneal fibrosis (p < 0.01 or p < 0.001) and CNV (p < 0.001) in therapy-given (chemical-trauma and AAV5-DCN + AAV5-PEDF) rabbit eyes compared to the no-therapy given eyes (chemical-trauma and AAV5-naked vector). Histopathological analyses demonstrated significantly reduced fibrotic α-smooth muscle actin and endothelial lectin expression in therapy-given corneas compared to no-therapy corneas on day-9 (p < 0.001) and day-15 (p < 0.001). Further, therapy-given corneas showed significantly increased Fas-ligand mRNA levels (p < 0.001) and apoptotic cell death in neovessels (p < 0.001) compared to no-therapy corneas. AAV5 delivered 2.69 × 10⁷ copies of DCN and 2.31 × 10⁷ copies of PEDF genes per μg of DNA. AAV5 vector and delivered DCN and PEDF genes found tolerable to the rabbit eyes and caused no significant toxicity to the cornea.

Conclusion

The combination AAV5-DCN and AAV5-PEDF topical gene therapy effectively reduces corneal fibrosis and CNV with high tolerability in vivo in rabbits. Additional studies are warranted.

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组织靶向和定位 AAV5-DCN 和 AAV5-PEDF 组合基因疗法可在体内减轻兔眼的角膜纤维化和并发新生血管形成

目的眼外伤后角膜纤维化和新生血管形成（CNV）会损害视力。本研究测试了组织靶向腺相关病毒 5（AAV5）介导的装饰素（DCN）和色素上皮衍生因子（PEDF）组合基因在体内的治疗潜力。化学创伤后 30 分钟，使用克隆圆筒通过局部应用 AAV5-DCN 和 AAV5-PEDF 在基质中进行基因治疗。定期对活兔进行临床眼部检查和多模态成像，并在安乐死后 9 天和 15 天收集角膜组织。结果 AAV5-DCN 和 AAV5-PEDF 联合基因疗法显著减少了角膜纤维化（p < 0.01 或 p < 0.001）和 CNV（p < 0.001）。组织病理学分析表明，在第 9 天（p < 0.001）和第 15 天（p < 0.001），接受治疗的角膜与未接受治疗的角膜相比，纤维化的 α-平滑肌肌动蛋白和内皮凝集素表达明显减少。此外，与未接受治疗的角膜相比，接受治疗的角膜的 Fas-配体 mRNA 水平（p <0.001）和新生血管细胞凋亡（p <0.001）明显增加。AAV5 每微克 DNA 可提供 2.69×107 个 DCN 和 2.31×107 个 PEDF 基因拷贝。结论 AAV5-DCN 和 AAV5-PEDF 联合局部基因疗法能有效减少兔子体内的角膜纤维化和 CNV，且具有很高的耐受性。有必要进行更多研究。

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来源期刊

Ocular Surface 医学-眼科学

CiteScore

11.60

自引率

14.10%

发文量

审稿时长

39 days

期刊介绍： The Ocular Surface, a quarterly, a peer-reviewed journal, is an authoritative resource that integrates and interprets major findings in diverse fields related to the ocular surface, including ophthalmology, optometry, genetics, molecular biology, pharmacology, immunology, infectious disease, and epidemiology. Its critical review articles cover the most current knowledge on medical and surgical management of ocular surface pathology, new understandings of ocular surface physiology, the meaning of recent discoveries on how the ocular surface responds to injury and disease, and updates on drug and device development. The journal also publishes select original research reports and articles describing cutting-edge techniques and technology in the field. Benefits to authors We also provide many author benefits, such as free PDFs, a liberal copyright policy, special discounts on Elsevier publications and much more. Please click here for more information on our author services. Please see our Guide for Authors for information on article submission. If you require any further information or help, please visit our Support Center