Huntington's disease (HD) is one of the most common, dominantly inherited neurodegenerative disorders. It affects the striatum, cerebral cortex, and other subcortical structures leading to involuntary movement abnormalities, emotional disturbances, and cognitive impairments. HD is caused by a CAG•CTG trinucleotide-repeat expansion in exon 1 of the huntingtin (HTT) gene leading to the formation of mutant HTT (mtHTT) protein aggregates. Besides the toxicity of the mutated protein, there is also evidence that mtHTT transcripts contribute to the disease. Thus, the reduction of both mutated mRNA and protein would be most beneficial as a treatment. Previously, we designed a novel anti-gene oligonucleotide (AGO)-based strategy directly targeting the HTT trinucleotide-repeats in DNA and reported downregulation of mRNA and protein in HD patient fibroblasts. In this study, we differentiate HD patient-derived induced pluripotent stem cells to investigate the efficacy of the AGO, a DNA/Locked Nucleic Acid mixmer with phosphorothioate backbone, to modulate HTT transcription during neural in vitro development. For the first time, we demonstrate downregulation of HTT mRNA following both naked and magnetofected delivery into neural stem cells (NSCs) and show that neither emergence of neural rosette structures nor self-renewal of NSCs is compromised. Furthermore, the inhibition potency of both HTT mRNA and protein without off-target effects is confirmed in neurons. These results further validate an anti-gene approach for the treatment of HD.

中文翻译：

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靶向 DNA 重复序列的寡核苷酸下调亨廷顿患者衍生神经模型系统中的亨廷顿基因表达

亨廷顿病 (HD) 是最常见的显性遗传神经退行性疾病之一。它影响纹状体、大脑皮层和其他皮层下结构，导致不自主运动异常、情绪障碍和认知障碍。HD 是由亨廷顿( HTT ) 基因的外显子 1 中的 CAG•CTG 三核苷酸重复扩增导致突变 HTT (mtHTT) 蛋白聚集体的形成引起的。除了突变蛋白的毒性外，还有证据表明 mt HTT转录物会导致该疾病。因此，减少突变的 mRNA 和蛋白质将是最有益的治疗。此前，我们设计了一种新的基于抗基因寡核苷酸 (AGO) 的策略，直接针对HTTDNA 中的三核苷酸重复并报道了 HD 患者成纤维细胞中 mRNA 和蛋白质的下调。在这项研究中，我们分化了源自 HD 患者的诱导多能干细胞，以研究 AGO（一种具有硫代磷酸酯骨架的 DNA/锁定核酸混合物）在神经体外发育过程中调节HTT转录的功效。我们首次证明了在裸露和磁转染递送到神经干细胞 (NSCs) 后HTT mRNA 的下调，并表明神经花环结构的出现和 NSCs 的自我更新都没有受到损害。此外，两种HTT的抑制效力在神经元中证实了没有脱靶效应的 mRNA 和蛋白质。这些结果进一步验证了用于治疗 HD 的抗基因方法。