Targeting CNG-Ca2+ channels: Evaluation of pharmacological and antisense oligonukleotide approaches for the treatment of retinitis pigmentosa.

本研究的目的是发现和评价药理学或反义寡核苷酸(AON)介导的、以环核苷酸门控钙通道(CNG通道)为靶点的神经保护剂，以防止视网膜色素变性(RP)中的光感受器退化。虽然许多经过临床测试的CNG通道的药理抑制剂很容易获得，可能允许快速临床翻译，但AON具有提高特异性和减少副作用的潜力。这项研究将在最初的18-24个月内分为体外筛选和验证，然后在相关的RP动物模型(RD1、RD2和RD10小鼠)上进行体内试验。药物化合物和AON对CNG通道的影响将在体外细胞培养和器官型视网膜外植体培养中进行测试，以快速建立原理证明并获得剂量-反应曲线。此外，还将对视网膜切片进行体外钙离子成像，以衡量潜在的脱靶效应。在三种不同的RP动物模型中，最有希望的化合物或AON将通过全身(腹膜内)或局部(玻璃体内)应用范例被带到体内水平。在这里，我们将使用扫描激光眼底镜(SLO)和光学相干断层扫描(OCT)活体成像技术在纵向研究中评估治疗效果，并结合视网膜电图(ERG)进行功能测试。这项研究计划旨在明确CNG通道靶向方法是否构成治疗RP的可行治疗策略，并提供第一批体内数据，以便于以后对此类方法进行临床前和临床测试。

The aim of this study is to identify and evaluate pharmacological or antisense oligonucleotide (AON) mediated, knock-down neuroprotectors that target cyclic nucleotide gated Ca2+ channels (CNG channels) to prevent photoreceptor degeneration in retinitis pigmentosa (RP). While numerous clinically-tested pharmacological inhibitors of CNG channels are readily available, possibly allowing for a rapid clinical translation, AONs bear the potential for improved specificity and reduced side-effects. The study will be split up into in vitro screening and validation in the first 18-24 project months, to then be carried forward to in vivo testing in relevant RP animal models (rd1, rd2, and rd10 mouse). The effect of pharmacological compounds and AONs on CNG channels will be tested in cell cultures and organotypic retinal explant cultures in vitro, to quickly establish proof-of-principle and to obtain dose-response curves. This will be supplemented by in vitro Ca2+ imaging on retinal slice preparations to weigh up potential off-target effects. The most promising compound or AON will be taken to the in vivo level using either systemic (intraperitoneal) or local (intravitreal) application paradigms, in three different RP animal models. Here, we will use scanning laser ophthalmoscopy (SLO) and optic coherence tomography (OCT) in vivo imaging techniques to assess treatment effects in longitudinal studies, combined with electroretinography (ERG) for functional testing. The study program is designed to clearly establish whether CNG channel targeting approaches constitute a viable therapeutic strategy for the treatment of RP and to yield first in vivo data to facilitate later pre-clinical and clinical testing of such approaches.