Therapeutic strategies for microsatellite expansion diseases using RNA-targeting CRISPR/Cas

项目摘要 CRISPR/Cas系统已经广泛应用于临床。 用于DNA识别，使基因组编辑和招募效应蛋白等应用能够 DNA影响转录或使成像。最近在Nelles等人的Cell，2016中，我们表明， 修饰，用含有PAM识别的修饰的寡核苷酸催化死亡的Cas9 序列能够结合活的哺乳动物细胞中的特定mRNA，并能够跟踪它们的运动， 开启了Cas蛋白的许多RNA应用的潜力。在这项建议中，我们的目的是发展一个 基于腺相关病毒（AAV）的强直性肌营养不良症治疗策略，微卫星扩增 以CTG重复序列扩增为特征的疾病，使用RNA靶向CRISPR/Cas9（RCas 9）。我们将 进行我们基于AAV的治疗的体内安全性和有效性研究，并评估以下假设： 替代的RNA加工生物标志物对于测量RCas 9在肌强直性疾病中的治疗功效是可靠的。 营养不良如果成功的话，我们将在开发治疗一类 微卫星扩张病！

Project Summary The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas system has become widely adopted for DNA recognition, enabling applications such as genome editing and recruiting effector proteins to DNA to affect transcription or enable imaging. Recently in Nelles et al, Cell, 2016, we showed that with modifications, catalytically dead Cas9 with a modified oligonucleotide containing the PAM recognition sequence is able to bind specific mRNAs in living mammalian cells and is able to track their movement, opening up the potential for many RNA applications of Cas proteins. In this proposal, we aim at developing a adeno-associated virus (AAV)-based therapeutic strategy for myotonic dystrophy, a microsatellite expansion disease characterized by expanded CTG repeats, using RNA-targeting CRISPR/Cas9 (RCas9). We will perform in vivo safety and efficacy studies of our AAV-based therapy and evaluate the hypothesis that alternative RNA processing biomarkers are reliable for measuring treatment efficacy of RCas9 in myotonic dystrophy. If successful, we will have taken a significant step forward in developing a treatment for a class of microsatellite expansion diseases.!