RP1 Dual-AAV Gene Therapy Preserves Retinal Structure and Ameliorates Photoreceptor Degeneration in a Murine Model of Retinitis Pigmentosa.

IF 4.7 2区医学 Q1 OPHTHALMOLOGY

Investigative ophthalmology & visual science Pub Date : 2025-09-02 DOI:10.1167/iovs.66.12.29

Fanfei Liu, Qiqi Li, Jing Su, Jiamei Fu, Xiaoyi Wu, Qiuxia Xu, Chengda Ren, Xiu Jin, Li Song, Fang Lu, Yang Yang

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引用次数: 0

Abstract

Purpose: This study aimed to develop and compare dual-adeno-associated virus (AAV) gene therapy utilizing protein trans-splicing versus mRNA trans-splicing to restore RP1 expression, and to evaluate their safety and efficacy in rescuing photoreceptor degeneration in a mouse model of RP1-associated retinitis pigmentosa (RP).

Methods: By screening various split sites within the RP1 sequence in combination with different inteins or adopting mRNA trans-splicing binding sequences, we generated several plasmid pairs capable of reconstituting RP1 in vitro. Next, we conducted dual-AAV gene therapy via subretinal injection in RP1-/- mice. We validated the RP1 protein reconstitution in vivo by Western blotting. To assess the safety and efficacy of dual-AAV systems in RP1-/- mice, retinal structure was evaluated by histological analysis and transmission electron microscopy (TEM), and visual function was assessed by electroretinography (ERG) and visually guided behavioral tests at 1 and 6 months post-treatment.

Results: The protein trans-splicing dual-AAV system achieved 72.35% of wild-type RP1 expression levels, significantly outperforming the mRNA trans-splicing approach (37.84%) at 1 month post-treatment. Both dual-AAV gene therapies preserved photoreceptor morphology and restored retinal function. The protein trans-splicing dual-AAV system exhibited higher ERG amplitudes and better performance in the water maze than the mRNA trans-splicing system.

Conclusions: Our study demonstrates protein trans-splicing mediated dual-AAV gene therapy as a promising strategy for RP1-associated RP, offering potential therapeutic avenue for patients with RP.

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RP1双aav基因治疗在色素性视网膜炎小鼠模型中保留视网膜结构并改善光感受器变性。

目的：研究利用蛋白反式剪接和mRNA反式剪接双腺相关病毒（AAV）基因疗法恢复RP1的表达，并评价其在RP1相关色素性视网膜炎（RP）小鼠模型中挽救光感受器变性的安全性和有效性。方法：通过筛选RP1序列内不同的分裂位点，结合不同的内部蛋白，或采用mRNA反式剪接结合序列，生成了几种能够在体外重组RP1的质粒对。接下来，我们通过RP1-/-小鼠视网膜下注射进行双aav基因治疗。我们通过Western blotting在体内验证了RP1蛋白的重组。为了评估双aav系统对RP1-/-小鼠的安全性和有效性，在治疗后1个月和6个月，通过组织学分析和透射电子显微镜（TEM）评估视网膜结构，通过视网膜电图（ERG）和视觉引导行为测试评估视觉功能。结果：蛋白反式剪接双aav系统在治疗后1个月达到野生型RP1表达水平的72.35%，显著优于mRNA反式剪接方法（37.84%）。两种双aav基因疗法均能保存感光体形态并恢复视网膜功能。在水迷宫中，蛋白质反式剪接双aav系统比mRNA反式剪接系统表现出更高的ERG振幅和更好的表现。结论：我们的研究表明蛋白质反式剪接介导的双aav基因治疗是一种很有前景的策略，为rp1相关的RP患者提供了潜在的治疗途径。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Investigative ophthalmology & visual science 医学-眼科学

CiteScore

6.90

自引率

4.50%

发文量

339

审稿时长

1 months

期刊介绍： Investigative Ophthalmology & Visual Science (IOVS), published as ready online, is a peer-reviewed academic journal of the Association for Research in Vision and Ophthalmology (ARVO). IOVS features original research, mostly pertaining to clinical and laboratory ophthalmology and vision research in general.