Single Stranded Fully Modified-Phosphorothioate Oligonucleotides can Induce Structured Nuclear Inclusions, Alter Nuclear Protein Localization and Disturb the Transcriptome In Vitro.
Single Stranded Fully Modified-Phosphorothioate Oligonucleotides can Induce Structured Nuclear Inclusions, Alter Nuclear Protein Localization and Disturb the Transcriptome In Vitro.
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DOI：
10.3389/fgene.2022.791416
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发表时间：
2022
影响因子：
3.7
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中科院分区：
文献类型：
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可以改变基因表达的寡核苷酸和核酸类似物现在在人类疾病中显示出治疗前景。虽然对合成核酸进行修饰以防止核酸酶降解和影响药物功能是常见的做法，但这种修饰也可能带来意想不到的物理化学和生物学特性。硫代磷酸骨架上的Gapmer混合修饰和DNA寡核苷酸可以非特异性地与细胞内蛋白质结合，形成各种有毒包裹体，受硫代磷酸连接的驱动，也受寡核苷酸序列的影响。最近，人们对2‘O-完全修饰的硫代磷酸连接寡核苷酸的非反义或其他非靶点效应有了更好的了解。在这里，我们报告了由修饰或未修饰碱基组成的、具有硫代键的寡核苷酸对亚核细胞器的化学特异性影响，并显示了核蛋白分布的改变，在原代人成纤维细胞和其他培养细胞中出现了高度稳定和显著结构的核包涵体，以及干扰了RNA的加工。磷酸二酯、磷二酰胺吗啉低聚物和退火硫代硫代低聚物二聚体没有引起这样的后果。亚核结构和蛋白质的破坏会引起严重的表型紊乱，这是通过对表现这种破坏的转基因成纤维细胞的转录分析揭示的。我们的数据增加了越来越多的证据，证明了一些硫代磷酸类药物在原代细胞中的非靶标效应，并提出了缓解这些效应的替代方法。
Oligonucleotides and nucleic acid analogues that alter gene expression are now showing therapeutic promise in human disease. Whilst the modification of synthetic nucleic acids to protect against nuclease degradation and to influence drug function is common practice, such modifications may also confer unexpected physicochemical and biological properties. Gapmer mixed-modified and DNA oligonucleotides on a phosphorothioate backbone can bind non-specifically to intracellular proteins to form a variety of toxic inclusions, driven by the phosphorothioate linkages, but also influenced by the oligonucleotide sequence. Recently, the non-antisense or other off-target effects of 2′ O- fully modified phosphorothioate linkage oligonucleotides are becoming better understood. Here, we report chemistry-specific effects of oligonucleotides composed of modified or unmodified bases, with phosphorothioate linkages, on subnuclear organelles and show altered distribution of nuclear proteins, the appearance of highly stable and strikingly structured nuclear inclusions, and disturbed RNA processing in primary human fibroblasts and other cultured cells. Phosphodiester, phosphorodiamidate morpholino oligomers, and annealed complimentary phosphorothioate oligomer duplexes elicited no such consequences. Disruption of subnuclear structures and proteins elicit severe phenotypic disturbances, revealed by transcriptomic analysis of transfected fibroblasts exhibiting such disruption. Our data add to the growing body of evidence of off-target effects of some phosphorothioate nucleic acid drugs in primary cells and suggest alternative approaches to mitigate these effects.