SCGE Disease Models Studies Supplement: Development of LNP-mediated based editing to treat Leber Congenital Amaurosis (LCA) for vision restoration in mouse model

摘要 遗传性失明很普遍，治疗选择有限。尽管取得了巨大的进步， 编辑技术的发展作为潜在的治疗失明，提供编辑的能力， 将复杂的细胞植入视网膜以进行有效编辑仍然是一个重大挑战。目前流行的方法， 编辑机制的递送是通过病毒载体，特别是AAV。对于AAV载体，诸如 免疫原性、基因组整合和长期作用仍然是安全性问题。此外，更多 复杂的编辑策略，包括碱基编辑（BE）和引物编辑（PE），需要包装 将多个组件放入不同的AAV中，这进一步限制了它们的效率。这一提议是为了利用 脂质纳米颗粒（LNP），我们已经在父母SCGE计划中开发了用于mRNA递送的脂质纳米颗粒。 碱基编辑器（ABE）来修复Leber先天性黑蒙（LCA）小鼠模型rd 12中的突变， 有一种人类的突变基因可以恢复视力我们之前已经证明， 介导靶向视网膜色素上皮细胞的Cas9 mRNA-gRNA的递送， 编辑.我们通过以下步骤进一步证明了碱基编辑复合物的LNP递送的可行性： 在小鼠模型中的mRNA。结合，我们将评估新的LNP用于mRNA递送的应用， 通过局部视网膜下注射进行碱基编辑和突变校正，并与功能相关。 恢复RD 12小鼠的视力。该项目的成功将使基于LNP的瞬时交付成为可能 编辑机器，以有效和安全地治疗因RPE突变引起的遗传性失明。

Abstract Genetic blindness is prevalent with limited treatment options. Despite tremendous progress in the development of editing technology as potential therapy for blindness, the ability to deliver the editing complex into the retina for efficient editing remains a major challenge. The current prevailing method of delivery of editing machinery is by viral vectors especially AAV. For AAV vectors, issues such as immunogenicity, genome integration and long-term effect remain a safety concern. Further, more complexed editing strategies including base editing (BE) and prime editing (PE), require the packaging of multiple components into different AAVs, which further limits their efficiency. This proposal is to use lipid nanoparticles (LNP) we have developed in the parental SCGE program for the mRNA delivery of a base editor (ABE) to repair the mutation in a Leber Congenital Amaurosis (LCA) mouse model rd12 that harbors a human mutation for the recovery of vision. We have previously demonstrated that the LNP mediates the delivery of Cas9 mRNA-gRNA that targets the retinal pigment epithelium for efficient editing. We have further demonstrated the feasibility of the LNP delivery of base editing complex by mRNA in a mouse model. Combined, we will evaluate the application of new LNP for mRNA delivery of the base editor and mutation correction by local subretinal injection, and correlate with the functional restoration of vision in the rd12 mice. The success of the project will enable LNP-based transient delivery of editing machinery to treat genetic blindness due to RPE mutations efficiently and safely.