Osman Erkan Y, Washington Charles W, Kaifer Kevin A, Mazzasette Chiara, Patitucci Teresa N, Florea Kyra M, Simon Madeline E, Ko Chien-Ping, Ebert Allison D, Lorson Christian L
Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Missouri, Columbia, Missouri, USA.
Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, Missouri, USA.
Mol Ther. 2016 Sep;24(9):1592-601. doi: 10.1038/mt.2016.145. Epub 2016 Jul 9.
Loss of Survival Motor Neuron-1 (SMN1) causes Spinal Muscular Atrophy, a devastating neurodegenerative disease. SMN2 is a nearly identical copy gene; however SMN2 cannot prevent disease development in the absence of SMN1 since the majority of SMN2-derived transcripts are alternatively spliced, encoding a truncated, unstable protein lacking exon 7. Nevertheless, SMN2 retains the ability to produce low levels of functional protein. Previously we have described a splice-switching Morpholino antisense oligonucleotide (ASO) sequence that targets a potent intronic repressor, Element1 (E1), located upstream of SMN2 exon 7. In this study, we have assessed a novel panel of Morpholino ASOs with the goal of optimizing E1 ASO activity. Screening for efficacy in the SMNΔ7 mouse model, a single ASO variant was more active in vivo compared with the original E1(MO)-ASO. Sequence variant eleven (E1(MOv11)) consistently showed greater efficacy by increasing the lifespan of severe Spinal Muscular Atrophy mice after a single intracerebroventricular injection in the central nervous system, exhibited a strong dose-response across an order of magnitude, and demonstrated excellent target engagement by partially reversing the pathogenic SMN2 splicing event. We conclude that Morpholino modified ASOs are effective in modifying SMN2 splicing and have the potential for future Spinal Muscular Atrophy clinical applications.
生存运动神经元1（SMN1）的缺失会导致脊髓性肌萎缩症，这是一种毁灭性的神经退行性疾病。SMN2是一个几乎完全相同的拷贝基因；然而，在没有SMN1的情况下，SMN2无法阻止疾病的发展，因为大多数源自SMN2的转录本会发生可变剪接，编码一种缺少外显子7的截短、不稳定的蛋白质。尽管如此，SMN2仍保留产生低水平功能蛋白的能力。此前我们描述了一种剪接转换吗啉代反义寡核苷酸（ASO）序列，其靶向位于SMN2外显子7上游的一个强效内含子阻遏元件，元件1（E1）。在本研究中，我们评估了一组新型的吗啉代ASO，目的是优化E1 ASO的活性。在SMNΔ7小鼠模型中筛选疗效时，与原始的E1（MO）-ASO相比，单个ASO变体在体内表现出更高的活性。序列变体十一（E1（MOv11））在单次脑室内注射至中枢神经系统后，通过延长严重脊髓性肌萎缩症小鼠的寿命，始终显示出更高的疗效，在一个数量级范围内呈现出强烈的剂量反应，并通过部分逆转致病性SMN2剪接事件证明了出色的靶点结合能力。我们得出结论，吗啉代修饰的ASO在改变SMN2剪接方面是有效的，并且具有未来用于脊髓性肌萎缩症临床应用的潜力。
