Dual ABCA4-AAV Vector Treatment Reduces Pathogenic Retinal A2E Accumulation in a Mouse Model of Autosomal Recessive Stargardt Disease
Dual ABCA4-AAV Vector Treatment Reduces Pathogenic Retinal A2E Accumulation in a Mouse Model of Autosomal Recessive Stargardt Disease
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DOI：
10.1089/hum.2019.132
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发表时间：
2019-09-30
影响因子：
4.2
通讯作者：
Hauswirth, William W.
中科院分区：
文献类型：
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作者：
Dyka, Frank M.;Molday, Laurie L.;Hauswirth, William W.
常染色体隐性遗传性Stargardt病是人类最常见的遗传性黄斑变性。它是由视网膜特异性ATP结合盒转运蛋白A4（ABCA 4）突变引起的，ABCA 4对于从感光细胞清除全反式视网膜至关重要。该功能的丧失导致毒性双维甲酸在外节椎间盘膜中的积累，并随后转移到相邻的视网膜色素上皮（RPE）细胞中。这最终导致视网膜自发荧光增加和进行性RPE和光感受器细胞损失的Stargardt病表型。腺相关病毒（AAV）载体已广泛用于基因治疗应用中，但其类似于4.5kb的有限cDNA包装能力阻碍了其用于超过该限制的转基因。开发了AAV双载体以克服这种大小限制。在这项研究中，我们评估了ABCA 4的体外表达使用三种选择：重叠，transplicing，和杂交ABCA 4双载体系统。杂交系统是这些双载体替代方案中最有效的，并且用于在Abca 4（-/-）小鼠中表达全长ABCA 4。全长ABCA 4蛋白正确定位于光感受器外节。此外，用该ABCA 4杂合双载体系统治疗Abca 4（-/-）小鼠导致脂褐质/N-亚视黄基-N-视黄乙醇胺（A2 E）自体荧光在体内的积累减少，并且视网膜A2 E定量支持这些发现。这些结果表明，杂合AAV双载体选项在小鼠中是安全的和治疗性的，并且递送的ABCA 4转基因是功能性的，并且对减少Stargardt病的Abca 4（-/-）小鼠模型中的A2 E积累具有显著作用。
Autosomal recessive Stargardt disease is the most common inherited macular degeneration in humans. It is caused by mutations in the retina-specific ATP binding cassette transporter A4 (ABCA4) that is essential for the clearance of all-trans-retinal from photoreceptor cells. Loss of this function results in the accumulation of toxic bisretinoids in the outer segment disk membranes and their subsequent transfer into adjacent retinal pigment epithelium (RPE) cells. This ultimately leads to the Stargardt disease phenotype of increased retinal autofluorescence and progressive RPE and photoreceptor cell loss. Adeno-associated virus (AAV) vectors have been widely used in gene therapeutic applications, but their limited cDNA packaging capacity of similar to 4.5 kb has impeded their use for transgenes exceeding this limit. AAV dual vectors were developed to overcome this size restriction. In this study, we have evaluated the in vitro expression of ABCA4 using three options: overlap, transplicing, and hybrid ABCA4 dual vector systems. The hybrid system was the most efficient of these dual vector alternatives and used to express the full-length ABCA4 in Abca4(-/-) mice. The full-length ABCA4 protein correctly localized to photoreceptor outer segments. Moreover, treatment of Abca4(-/-) mice with this ABCA4 hybrid dual vector system resulted in a reduced accumulation of the lipofuscin/N-retinylidene-N-retinylethanolamine (A2E) autofluorescence in vivo, and retinal A2E quantification supported these findings. These results show that the hybrid AAV dual vector option is both safe and therapeutic in mice, and the delivered ABCA4 transgene is functional and has a significant effect on reducing A2E accumulation in the Abca4(-/-) mouse model of Stargardt disease.