Small Oligonucleotides As Therapeutic Agents Of Spinal Muscular Atrophy

小寡核苷酸作为脊髓性肌萎缩症的治疗剂

• 批准号: 8296504
• 负责人: RAVINDRA N SINGH
• 金额: $ 18.26万
• 依托单位: IOWA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-15 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8296504
• 关键词: Address; Alleles; Antisense Oligonucleotides; Applications Grants; Belief; Biological; Brain; Cell Line; Cells; Characteristics; Chemicals; Chemistry; Collaborations; Consensus; Coupled; Custom; Dose; Drug Delivery Systems; Drug Kinetics; Exons; GC Rich Sequence; Generations; Genetic; Goals; Growth and Development function; Human; Immune response; Infant Mortality; Lead; Length; Life; Lipids; Longevity; Modeling; Modification; Motor Neurons; Mus; Neuromuscular Diseases; Nucleotides; Oligonucleotides; Outcome Measure; Patients; Pharmaceutical Preparations; Phenotype; Production; Property; Proteins; RNA Splicing; Rana; Reporting; Research Personnel; Running; SMN1 gene; SMN2 gene; Series; Site; Specificity; Spinal; Spinal Cord; Spinal Muscular Atrophy; Suggestion; Testing; Therapeutic; Therapeutic Agents; TimeLine; Tissues; Transgenic Mice; Treatment Efficacy; Variant; Vertebral column; Werdnig-Hoffmann Disease; base; cost; efficacy testing; functional loss; improved; in vivo; mRNA Precursor; mouse model; nanoparticle; novel; offspring; phosphorothioate; preclinical study; pregnant; pup; research study; response; success; survival motor neuron gene; therapy development

DESCRIPTION (provided by applicant): Spinal Muscular Atrophy (SMA) is a leading genetic cause of infant mortality. Most commonly, SMA results from the reduced levels of full-length SMN protein (SMN) in motor neurons and spinal chord due to the loss of functional Survival Motor Neuron (SMN1) alleles. A nearly identical copy of this gene, SMN2, fails to provide protection from SMA due to production of a truncated SMN because of skipping of SMN2 exon 7 during pre-mRNA splicing. There is a near consensus among researchers that strategies aimed at promotion of SMN2 exon 7 inclusion resulting into the increased levels of full- length SMN would cure SMA. Towards this goal we have recently reported an eight-nucleotide GC- rich intronic target, sequestering of which by an antisense oligonucleotide (ASO) fully restored SMN2 exon 7 inclusion in SMA patient cells. Specificity and efficiency of antisense response by our 8-mer lead ASO (3UP8) targeting GC-rich sequence constitute the first such report of splicing correction by a short ASO in a patient cell line. The unmatched benefits of a short ASO as a therapeutic agent include but not limited to the expected high specificity, low cost of synthesis, ease of modifications and increased chances of delivery across biological barriers. Currently SMA has no cure. As one of the best hopes of SMA therapy, here we propose to develop an optimized variant of our lead ASO that efficiently corrects SMN2 exon 7 splicing and raise the levels of full-length SMN in all tissues including brain and spinal cord of mice models of SMA. In Aim 1, we will optimize our short ASO for application in vivo. These will be accomplished by a series of custom modifications including different combinations of the terminal and backbone chemistries. We will test the efficacy of modified short ASOs in transgenic mice containing human SMN2. Our initial results validate the proof-of-principle that custom modifications improve the efficacy of a short ASO in splicing correction in vivo. We will run a series of tests to confirm the in vivo efficacy of the custom-modified ASOs. These include but not limited to stability, dose response, off-target effects, immune response and pharmacokinetic properties. We believe that a combination of chemical modifications will allow us to obtain an optimized lead ASO that could be effectively delivered in all tissues including brain and spinal cord. Will also test an alternatively lipid-nanoparticle-based approach to efficiently deliver our optimized lead ASO in different tissues including across BBB. In Aim 2, we will perform experiments in mild as well as severe SMA mouse models. We will determine the effect of different doses of our optimized lead ASO on the phenotype, growth and development of SMA mice. In particular, we seek to improve the longevity of SMA mice. We will also conduct experiments in pregnant mice to see the effect of pre-natal drug delivery on the phenotype and longevity of SMA offspring. The success of this proposal will provide a mechanism-based and target- specific drug for the treatment of SMA. )

描述（由申请人提供）：脊髓性肌萎缩症（SMA）是婴儿死亡的主要原因。最常见的是，SMA 是由于功能性运动神经元存活 (SMN1) 等位基因的丧失导致运动神经元和脊髓中全长 SMN 蛋白 (SMN) 水平降低所致。该基因的几乎相同的副本 SMN2 无法提供针对 SMA 的保护，因为在前 mRNA 剪接过程中跳过了 SMN2 外显子 7，从而产生了截短的 SMN。研究人员几乎达成共识，旨在促进 SMN2 外显子 7 包含并导致全长 SMN 水平增加的策略将治愈 SMA。为了实现这一目标，我们最近报道了一种富含 8 核苷酸 GC 的内含子靶标，通过反义寡核苷酸 (ASO) 隔离该靶标，完全恢复了 SMA 患者细胞中的 SMN2 外显子 7 包含。我们的 8 聚体先导 ASO (3UP8) 靶向富含 GC 的序列的反义反应的特异性和效率构成了第一个在患者细胞系中通过短 ASO 进行剪接校正的此类报告。短 ASO 作为治疗剂具有无与伦比的优势，包括但不限于预期的高特异性、合成成本低、易于修饰以及增加跨越生物屏障的递送机会。目前 SMA 尚无治愈方法。作为 SMA 治疗的最大希望之一，我们在这里建议开发我们的先导 ASO 的优化变体，它可以有效纠正 SMN2 外显子 7 剪接，并提高 SMA 小鼠模型的所有组织（包括大脑和脊髓）中全长 SMN 的水平。 SMA。 在目标 1 中，我们将优化我们的短 ASO 以便在体内应用。这些将通过一系列定制修改来完成，包括末端和主链化学物质的不同组合。我们将在含有人 SMN2 的转基因小鼠中测试改良的短 ASO 的功效。我们的初步结果验证了原理证明，即定制修饰可提高短 ASO 在体内剪接校正中的功效。我们将进行一系列测试来确认定制修饰的 ASO 的体内功效。这些包括但不限于稳定性、剂量反应、脱靶效应、免疫反应和药代动力学特性。我们相信，化学修饰的组合将使我们能够获得优化的先导 ASO，它可以有效地输送到包括大脑和脊髓在内的所有组织中。还将测试另一种基于脂质纳米颗粒的方法，以在不同组织（包括整个血脑屏障）中有效地传递我们优化的先导 ASO。在目标 2 中，我们将在轻度和重度 SMA 小鼠模型中进行实验。我们将确定不同剂量的优化先导 ASO 对 SMA 小鼠表型、生长和发育的影响。特别是，我们寻求提高 SMA 小鼠的寿命。我们还将在怀孕小鼠身上进行实验，观察产前给药对 SMA 后代表型和寿命的影响。该提案的成功将为SMA的治疗提供基于机制、靶向特异性的药物。 ）