Supplement to Accelerating photoreceptor replacement therapy with in-vivo cellular imaging of retinal function

为了恢复患者的高质量、可用视力，在动物身上开发再生疗法是很重要的。 具有人类视觉系统关键特征的模型。非人灵长类动物实际上有一个完全相同的 视网膜解剖学和生理学与人类的关系，在哺乳动物模型中是独一无二的，共同的中心凹， 视网膜区域专门用于高视力视力的区域。而非人类的灵长类动物是黄金标准动物 用于人类视觉研究的模型，其在临床前测试中的应用由于两者的缺失而受到限制 视力丧失的模型和展示恢复功能的困难。在之前的AGI资助下， 罗切斯特大学的高级视网膜成像联盟最近克服了这些挑战 创建利用自适应光纤技术的临床前测试平台，以：1.创建本地化区域 灵长类动物的光感受器消融是轴向受限的，以及2。光学读出修复的视网膜 神经节细胞通过对活体眼睛进行细胞尺度的钙成像来实现功能。这个系统是 开发以满足光感受器替代疗法的需求，该疗法需要光感受器丢失 保存了宿主视网膜回路。此外，一种高分辨率的活体成像方法非常适合于 再生治疗的临床评价，其中恢复连接的时间尺度未知，并且 功能整合发生在细胞尺度上。在此提案中，我们将使用我们的平台来生成预 临床数据将为未来光感受器替代疗法在患者中的临床试验提供信息。功能性 移植的光感受器与宿主视网膜的整合需要高密度的高密度输送 优质的供体光感受器和具有突触形成能力的宿主视网膜。我们已经组建了一支 可以探索和优化这种互动双方的财团。继续与团队合作 在威斯康星大学，由临床医生和人类发育专家大卫·伽姆领导的 多能干细胞(HPSC)衍生的光感受器替代疗法，我们将评估患者的存活率和 移植的光感受器前体在灵长类中心凹输送到视网膜下的功能整合 空间作为集合体或结合到定制的可生物降解支架。与 加州大学伯克利分校的团队由视网膜重塑专家Teresa Puthussery领导 变性模型和灵长类组织学，我们将检查光感受器信号丢失的影响 在灵长类的内视网膜上。我们将探索去传入的锥体双极细胞是否可以重塑和功能 与供体光感受器整合，以及灵长类动物是否会像啮齿动物那样出现视网膜过度活动。 在恢复视力方面取得有意义的进展，这些患者生活在许多年的视力丧失中 ，我们将研究这些现象是如何随着时间的推移在灵长类中心凹中发展的，以及 宿主的再生能力可以通过维甲酸阻滞剂等治疗干预措施得到改善。 这些研究将使我们能够充分描述我们的灵长类光感受器消融模型，并将其应用于 光感受器替代疗法将该领域推向临床试验。

To restore high quality, usable vision in patients, it is important to develop regenerative therapies in an animal model that shares key features of the human visual system. The non-human primate has a virtually identical retinal anatomy and physiology to the human and, uniquely amongst mammalian models, shares a fovea, the retinal area specialized for high acuity vision. Whilst the non-human primate is the gold standard animal model for research on human vision, its utility for pre-clinical testing has been limited due to both the absence of models of vision loss and the difficulty of demonstrating restored function. Under previous AGI funding, the Advanced Retinal Imaging Alliance at the University of Rochester has recently overcome these challenges to create a pre-clinical testing platform leveraging adaptive optics technology to: 1.) Create localized regions of photoreceptor ablation in primate that are axially confined, and 2.) Optically read out restored retinal ganglion cell function by performing cellular scale calcium imaging in the living eye. This system was developed to meet the needs of photoreceptor replacement therapy which requires photoreceptor loss with preserved host retinal circuitry. Furthermore, a high-resolution in vivo imaging approach is well suited for pre- clinical evaluation of regenerative therapies where the timescales of restored connectivity are unknown and functional integration occurs on the cellular scale. In this proposal, we will use our platform to generate pre- clinical data that will inform future clinical trials of photoreceptor replacement therapy in patients. Functional integration of transplanted photoreceptors with the host retina requires both high density delivery of high- quality donor photoreceptors and a host retina with the capacity for synaptogenesis. We have assembled a consortium that can explore and optimize both sides of this interaction. In continued collaboration with a team at the University of Wisconsin led by David Gamm, a clinician and expert in the development of human pluripotent stem cell (hPSC) derived photoreceptor replacement therapy, we will evaluate survival and functional integration of transplanted photoreceptor precursors in primate fovea delivered to the sub-retinal space as aggregates or following incorporation into custom biodegradable scaffolds. In collaboration with a team at University of California, Berkeley led by Teresa Puthussery, an expert in retinal remodelling in retinal degeneration models and primate histology, we will examine the impact of the loss of photoreceptor signalling on primate inner retina. We will explore whether deafferented cone bipolar cells can remodel and functionally integrate with donor photoreceptors and whether retinal hyperactivity develops in primate as it does in rodent. To make meaningful progress toward restoring vision in patients who have lived with vision loss for many years, we will examine how these phenomena develop in the primate fovea over time and whether the regenerative potential of the host can be improved by therapeutic interventions such as retinoic acid blockers. These studies will allow us to fully characterize our primate photoreceptor ablation model and deploy it with photoreceptor replacement therapies to advance the field toward clinical trials.