A novel exon-specific U1 snRNA strategy to correct splicing in Familial Dysautonomia

家族性自主神经功能障碍（FD）是一种隐性遗传性神经退行性疾病， IKBKAP基因，导致外显子20的组织特异性可变跳读。这反过来又导致了 IKAP/ELP 1蛋白在中枢和外周神经系统中的表达。虽然FD患者患有多种 神经系统症状，进行性失明大大降低了生活质量，是一个主要关注的问题， 患者及其家属。在这个提议中，我们概述了一种新的策略，靶向视网膜中的mRNA剪接， FD视网膜变性的潜在治疗方法。Franco帕加尼博士的实验室开发了一种新型的改良 剪接体U1 snRNA（ExSpeU 1 s）的版本，可以特异性靶向改善包括 跳过的外显子这一策略已被证明在其他几种剪接疾病中是成功的，包括脊髓肌肉 萎缩和囊性纤维化。最近，他的实验室已经创建了IKBKAP特定的U1，具有精确的靶向碱基- 我们已经证明，它们在FD细胞和正常人中都能正确剪接并增加IKAP蛋白。 使用AAV 9通过腹膜内递送在体内进行。在过去的几年里，我们一直在努力创造一个新的FD 小鼠模型，其既再现人类疾病表型又再现组织特异性剪接缺陷。我们 新的小鼠是唯一可用于评估IKBKAP剪接调节的治疗效果的模型 疾病表型。改变mRNA剪接的突变占所有人类遗传基因的很大一部分。 疾病，以及纠正剪接的方法，包括小分子和寡核苷酸，已经在临床试验中。 mRNA剪接是一个复杂的过程，因此无法预测这些方法的有效性 在不同的突变中，事实上，联合治疗可能最终是必要的。在这 我们将研究一种新的治疗策略，即递送外显子靶向的U1 snRNAs，我们 将测试他们的疗效，特别是在视网膜的速度与AAV定向基因治疗视网膜病变。 疾病正在进入诊所。我们已经组建了一个专家小组，我们的研究不仅涉及 关键未满足的医疗需求，但也将使我们能够揭示这些外显子的确切机制， 靶向分子纠正剪接。成功地实现所述目标肯定会对以下方面产生影响： 治疗FD以外的遗传剪接疾病，进一步增加了这一建议的重要性。

Familial dysautonomia (FD) is a recessive neurodegenerative disease caused by a splice mutation in the IKBKAP gene that leads to tissue-specific variable skipping of exon 20. This, in turn, leads to a drastic reduction of IKAP/ELP1 protein in the central and peripheral nervous system. Although FD patients suffer from multiple neurological symptoms, progressive blindness drastically reduces quality of life and is a major concern of patients and their families. In this proposal, we outline a novel strategy to target mRNA splicing in the retina as a potential therapy for retinal degeneration in FD. Dr. Franco Pagani's laboratory has developed novel modified versions of the spliceosomal U1snRNAs (ExSpeU1s) that can be specifically targeted to improve inclusion of skipped exons. This strategy has proven successful in several other splicing disorders, including spinal muscular atrophy and cystic fibrosis. Recently, his lab has created IKBKAP specific U1s with precisely targeted base- pairing and together we have shown that they correct splicing and increase IKAP protein both in FD cells and in vivo by intraperitoneal delivery using AAV9. For the past several years, we have worked to generate a new FD mouse model that both recapitulates the human disease phenotype and the tissue-specific splice defect. Our new mouse is the only model that can be used to evaluate the therapeutic effect of IKBKAP splicing modulation on disease phenotype. Mutations that alter mRNA splicing account for a large fraction of all human genetic disease, and methods to correct splicing, including small molecules and oligos, are already in clinical trials. mRNA splicing is a complex process, and therefore it is impossible to predict the efficacy of these methods across various mutations, and in fact it is likely that combination therapies will ultimately be necessary. In this proposal we will study a novel therapeutic strategy, namely the delivery of exon-targeted U1 snRNAs, and we will test their efficacy specifically in the retina given the speed with which AAV directed gene therapy for retinal disease is moving into the clinic. We have assembled an expert team and our studies will not only address a critical unmet medical need in FD but will also allow us to uncover the precise mechanism by which these exon targeted molecules correct splicing. Successful completion of the stated Aims will certainly have implications for treating genetic splicing disease beyond FD, further increasing the significance of this proposal.