Expanding the chemical diversity of therapeutic oligonucleotides to treat neurodegenerative disorders

摘要： 治疗性寡核苷酸(例如，小干扰RNA(SiRNA)和反义)有望成为 治疗遗传性神经退行性疾病的变革性药物，包括 亨廷顿病(HD)和肌萎缩侧索硬化症(ALS)。SiRNA通过以下方式沉默致病基因 靶向其同源的mRNAs进行降解，从而防止有毒基因产物的表达。 它们固有的序列特异性和持久的活性提供了一个强大的治疗平台，只要 这种疾病是从基因上定义的，并且可以将其输送到相关的靶组织。然而，siRNA可以 不能通过血脑屏障和局部注射输送中枢神经系统往往会导致滞留、分布不良 或者毒性。因此，高效和无毒的传递是基于RNAi的发展的主要障碍 治疗神经退行性疾病的药物。 这项提议的目标是开发和表征新型化学支架， 高效、无毒的寡核苷酸递送和治疗靶点的有效沉默 神经系统。我们描述了一类完全化学稳定的疏水siRNAs(HsiRNAs)，它可以诱导 脑脊液(CSF)注射后，整个大脑持久而有效的沉默。 修饰包括寡核苷酸结构、核糖、主干和添加亲脂结合物- 例如，神经活性类固醇、内源性大麻素样脂、神经节苷脂。广泛的结构-活性关系 研究表明，结合物的类型定义了分布、保留、有效性、作用持续时间和 Hsirna结合物对中枢神经系统的毒性。 这项提案的完成将(I)确定和表征两种新型的化学支架 有效、特异和持久地沉默中枢神经系统中的靶基因，以及(Ii)验证这一点 HD和ALS动物模型的新平台，为两种疾病的新治疗开辟了一条道路 神经退行性疾病。这项提议建立了一个平台，允许直接靶向任何基因 在啮齿动物的中枢神经系统的任何区域都有表达。这项工作的圆满完成将 使中枢神经系统中的基因功能研究成为可能，并为开发新的 基因定义的神经退行性疾病的基于寡核苷酸的治疗。

ABSTRACT: Therapeutic oligonucleotides (e.g., small interfering RNAs (siRNAs) and antisense) hold promise as transformative drugs for the treatment of genetically defined neurodegenerative disorders, including Huntington's disease (HD) and amyotrophic lateral sclerosis (ALS). siRNAs silence disease-causing genes by targeting their cognate mRNAs for degradation, thereby preventing the expression of toxic gene products. Their inherent sequence specificity and prolonged activity provide a powerful therapeutic platform, as long as the disease is genetically defined and delivery to the relevant target tissue is achievable. However, siRNAs do not cross the blood–brain barrier and local CNS delivery by injection often results in poor retention, distribution or toxicity. Thus, efficient and non-toxic delivery represents a major hurdle in the development of RNAi-based drugs to treat neurodegenerative disorders. The goal of this proposal is to develop and characterize novel chemical scaffolds that promote simple, efficient, and non-toxic delivery of oligonucleotides and potent silencing of therapeutic targets in the central nervous system. We describe a class of fully chemically stabilized hydrophobic siRNAs (hsiRNAs) that elicit durable and potent silencing throughout the brain following bolus cerebrospinal fluid (CSF) infusion. Modifications include oligonucleotides structure, ribose, backbone and the addition of lipophilic conjugates— e.g., neuroactive steroids, endocannabinoid-like lipids, gangliosides. Extensive structure-activity relationship studies reveal that the type of conjugate defines the distribution, retention, efficacy, duration of effect, and toxicity of hsiRNA-conjugates in the central nervous system. Completion of this proposal will (i) define and characterize two novel chemical scaffolds that support potent, specific, and long-lasting silencing of target genes in the central nervous system, and (ii) validate this new platform in animal models of HD and ALS, establishing a path towards novel treatments for two neurodegenerative diseases. This proposal establishes a platform that allows direct targeting of any gene expressed in any region of the central nervous system in a rodent. Successful completion of this work will enable studies of gene function in the central nervous system and pave the way towards development of novel oligonucleotide-based therapies for genetically defined neurodegenerative diseases.