Novel model systems for the study of cone disorders and other heritable retinal diseases

项目摘要/摘要 视网膜变性疾病是全世界无法治愈的失明的常见原因，影响着数百万人的生活。FDA批准的治疗这些疾病的唯一方法是针对导致Leber先天性黑色素和视网膜色素变性的特定RPE65突变进行基因治疗。开发有效疗法的一大限制是使用模型系统，这些系统很难复制人类的情况。特别是对于视锥细胞障碍，使用视网膜杆状占优势且没有真正的黄斑的模型系统进行的研究有实质性的局限性。我们建议建立一系列新颖的、自发的人类遗传性视网膜疾病的动物模型。在初步分析中，我们已经确定了一种与人类视网膜疾病直接相关的新的自发模型。基因测试确定了四个人是PDE6C基因自然发生的破坏性突变的纯合个体，该基因以前曾与人类锥体营养不良有关。在PDE6C突变纯合子动物身上进行的暗视和明视全场视网膜电图显示相对正常的视杆反应，但没有任何视锥反应。使用光谱域光学相干断层扫描，通过眼底照相、蓝色自发荧光和荧光素血管造影术，同时进行黄斑中心凹变薄，确定了一种微小但具有特征性的视网膜病变。我们的基因调查还发现了其他7个人类视网膜疾病基因突变的个体，这些基因预计会严重损害基因或蛋白质功能，并指出可能会有其他新的模型。为了开发PDE6C视锥细胞营养不良的新模型，并将该模型和其他新模型提供给视觉研究团体，我们提出了四个具体目标：1)通过DNA测序识别并从遗传学上表征人类视网膜疾病的新动物模型，2)对新的视网膜疾病模型进行完整的眼部表型鉴定，3)培育PDE6C视锥细胞营养不良的动物群体，4)比较基于细胞的治疗和基因替代疗法在这些具有PDE6C视锥细胞营养不良突变的受试者中的作用。这项工作的成功完成将产生一个具有良好特征的遗传性视锥细胞营养不良的新动物模型，与现有模型相比，它与人类疾病的相似性要大得多，从而为后续的人体试验提供更好的翻译。此外，受影响的动物将提供给更广泛的视力研究社区，并将确定其他具有类似潜力的新模型。

Project Summary/Abstract Retinal degeneration diseases are a common cause of untreatable blindness worldwide, affecting the lives of millions. The only FDA-approved treatment for these disorders is gene therapy for specific RPE65 mutations that cause Leber’s congenital amaurosis and retinitis pigmentosa. One major limitation to the development of effective therapies is the use of model systems that poorly replicate the human condition. Particularly for cone disorders, studies that use model systems with a rod-dominant retina and no true macula have substantive limitations. We propose to develop a series of novel and spontaneous animal models of human inherited retinal diseases. In preliminary analyses, we have identified a new spontaneous model directly relevant to human retinal disease. Genetic testing identified four individuals homozygous for a naturally occurring damaging mutation in the PDE6C gene, which has previously been associated with cone dystrophy in humans. Scotopic and photopic full-field electroretinograms performed on animals homozygous for the PDE6C mutation demonstrated a relatively normal rod response but no cone response whatsoever. A subtle but characteristic retinopathy was identified using fundus photography, blue autofluorescence, and fluorescein angiography with concurrent foveal thinning using spectral-domain optical coherence tomography. Our genetic survey also identified individuals with mutations in 7 other human retinal disease genes that are predicted to severely damage gene or protein function, pointing to possible additional new models. To develop the new model of PDE6C cone dystrophy, and make this and other new models available to the vision research community, we propose four Specific Aims: 1) to identify and genetically characterize new animal models of human retinal disease via DNA sequencing, 2) to perform complete ophthalmic phenotyping of the new models of retinal disease, 3) to breed a colony of animals with PDE6C cone dystrophy and 4) to compare cell-based and gene replacement therapies in these subjects with PDE6C cone dystrophy mutations. Successful completion of this work will produce a well-characterized new animal model of inherited cone dystrophy with significantly greater similarity to human disease than existing models, thus providing substantially better translation to subsequent human trials. In addition, affected animals will be made available to the wider vision research community, and other new models with similar potential will be identified.