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 数百万人的生命。FDA批准的针对这些疾病的唯一治疗方法是针对特定疾病的基因疗法 导致Leber先天性黑色素和视网膜色素变性的RPE65突变。一大专业 有效疗法开发的限制是使用复制效果不佳的动物模型 人类的状况。特别是对于视锥细胞紊乱，研究使用了视网膜杆状占优势的动物 而且没有真正的斑疹有实质性的局限性。相比之下，非人的富含视锥的黄斑 灵长类动物(NHP)与人类的视网膜非常相似。因此，定义良好的NHP模型 更能预测人类状况的遗传性视网膜疾病，特别是视锥细胞紊乱，是 对于更有效地推进新疗法是必要的。我们计划开发一系列新颖的和 人类遗传性视网膜疾病的自发NHP模型。恒河猴的行为观察 加州国家灵长类研究中心的猕猴(Macaca Mulatta)发现了一系列 表现出明显视力障碍的猕猴。基因检测显示，有四种动物是 PDE6C基因中破坏性突变的纯合子，此前已被认为与 人类的视锥细胞营养不良。对猕猴进行暗视和明视全场视网膜电流图检查 PDE6C突变纯合子显示出相对正常的视杆反应，但没有锥体 不管是什么反应。眼底检查发现了一种轻微但具有特征性的牛眼黄斑病变。 同时进行黄斑中心凹变薄的摄影、蓝色自发荧光和荧光素血管造影 光谱域光学相干层析成像。我们对这一群体的基因调查也发现 其他7个人类视网膜疾病基因突变的猕猴被预测会严重损害 基因或蛋白质的功能，表明可能有其他新的模型。开发PDE6C灵长类动物 视锥细胞营养不良的模型，并使该模型和其他新的NHP模型可用于视觉研究 社区，我们提出了四个具体目标：1)识别新的NHP模型并从基因上描述其特征 通过DNA测序诊断人类视网膜疾病，2)对NHP进行完整的眼部表型鉴定 视网膜疾病的模型，3)培养出患有PDE6C视锥细胞营养不良的NHP群体，4)比较 PDE6C锥体营养不良突变猕猴的细胞和基因替代治疗。 这项工作的成功完成将产生一个具有良好特征的新的遗传动物模型 与现有模型相比，锥体营养不良与人类疾病的相似性明显更大，因此 为后续的人体试验提供了更好的翻译。此外，受影响的动物将 将提供给更广泛的视觉研究社区，以及其他具有类似潜力的新模型 将会被确认。

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 animal models that poorly replicate the human condition. Particularly for cone disorders, studies that use animals with a rod-dominant retina and no true macula have substantive limitations. By contrast, the cone-rich macula of nonhuman primates (NHP) closely mirrors that of the human retina. Consequently, well-defined NHP models of heritable retinal diseases, particularly cone disorders, that are more predictive of human conditions are necessary to more efficiently advance new therapies. We propose to develop a series of novel and spontaneous NHP models of human inherited retinal diseases. Behavioral observations of rhesus macaques (Macaca mulatta) at the California National Primate Research Center identified a series of macaques that displayed apparent visual impairment. Genetic testing showed that four animals are homozygous for a damaging mutation in the PDE6C gene, which has previously been associated with cone dystrophy in humans. Scotopic and photopic full-field electroretinograms performed on macaques homozygous for the PDE6C mutation demonstrated a relatively normal rod response but no cone response whatsoever. A subtle but characteristic bullseye maculopathy was identified using fundus photography, blue autofluorescence, and fluorescein angiography with concurrent foveal thinning using spectral-domain optical coherence tomography. Our genetic survey of this population also identified macaques 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 PDE6C primate model of cone dystrophy, and make this and other new NHP models available to the vision research community, we propose four Specific Aims: 1) to identify and genetically characterize new NHP models of human retinal disease via DNA sequencing, 2) to perform complete ophthalmic phenotyping of NHP models of retinal disease, 3) to breed a colony of NHPs with PDE6C cone dystrophy and 4) to compare cell-based and gene replacement therapies in macaques 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.