Huntington's disease is a neurodegenerative disorder caused by a CAG repeat expansion in the first exon of huntingtin gene (HTT) encoding for a toxic polyglutamine protein. This disease is characterized by motor, psychiatric, and cognitive impairments. Currently, there is no disease modifying treatment. However, reducing the expression of the huntingtin protein (HTT) using antisense oligonucleotides (ASOs) has been shown as a promising therapeutic strategy. In this study, we explore the therapeutic potential of ASO made of tricyclo-DNA (tcDNA), a conformationally constrained DNA analog, to silence HTT. We used a gapmer ASO, containing central DNA nucleotides flanked by tcDNA modifications on 5' and 3' ends, allowing the recruitment of RNAse H and subsequent degradation of the messenger RNA. After transfection of tcDNA-ASO in patient-derived fibroblast cell lines, we show a strong decrease of HTT mRNA and protein levels. As a control, 2'O-methyl-RNA targeting the same region of HTT was also tested and did not induce a significant effect. tcDNA-ASO were also evaluated in vivo in the YAC128 mice, containing the full-length human HTT gene with 128 CAG repeat expansion. Single intracerebroventricular (ICV) injections of tcDNA induce a significant decrease of HTT messenger and protein levels in the cortex, hippocampus, striatum, and cerebellum of treated mice. tcDNA-ASO were found well distributed in the central nervous system (CNS) and show long lasting effect with protein levels still low, 12 weeks after a single ICV injection. This proof of concept study suggests the therapeutic potential of gapmer tcDNA ASO to downregulate huntingtin in vitro and in vivo.

中文翻译：

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使用三环DNA反义寡核苷酸降低亨廷顿病突变体作为亨廷顿氏病的治疗方法。

亨廷顿舞蹈病是一种神经退行性疾病，由亨廷顿基因（HTT）编码有毒的聚谷氨酰胺蛋白的第一个外显子中的CAG重复扩增引起。这种疾病的特征是运动，精神病和认知障碍。当前，没有疾病改良治疗。然而，使用反义寡核苷酸（ASO）降低亨廷顿蛋白（HTT）的表达已被证明是一种有前途的治疗策略。在这项研究中，我们探索由三环DNA（tcDNA）（一种构象受限的DNA类似物）制成的ASO的治疗潜力，以使HTT沉默。我们使用了gapmer ASO，其包含侧翼为5'和3'末端的tcDNA修饰的中央DNA核苷酸，从而允许募集RNAse H和随后降解信使RNA。在患者来源的成纤维细胞系中转染tcDNA-ASO后，我们显示HTT mRNA和蛋白水平大大降低。作为对照，还测试了靶向HTT相同区域的2'O-甲基-RNA，并且没有引起明显的作用。还在YAC128小鼠体内评估了tcDNA-ASO，该小鼠包含具有128 CAG重复扩增的全长人HTT基因。单次脑室内（ICV）注射tcDNA会在治疗小鼠的皮质，海马，纹状体和小脑中显着降低HTT信使和蛋白质水平。一次ICV注射12周后，发现tcDNA-ASO在中枢神经系统（CNS）中分布良好，并显示出长效作用，蛋白质水平仍然很低。