Antisense oligonucleotide (ASO) has the potential to induce off-target effects due to complementary binding between the ASO and unintended RNA with a sequence similar to the target RNA. Conventional animal studies cannot be used to assess toxicity induced by off-target effects because of differences in the genome sequence between humans and other animals. Consequently, the assessment of off-target effects with in silico analysis using a human RNA database and/or in vitro expression analysis using human cells has been proposed. Our previous study showed that the number of complementary regions of ASOs with mismatches in the human RNA sequences increases dramatically as the number of tolerated mismatches increases. However, to what extent the expression of genes with mismatches is affected by off-target effects at the cellular level is not clear. In this study, we evaluated off-target effects of gapmer ASOs, which cleave the target RNA in an RNase H-dependent manner, by introducing the ASO into human cells and performing microarray analysis. Our data indicate that gapmer ASOs induce off-target effects depending on the degree of complementarity between the ASO and off-target candidate genes. Based on our results, we also propose a scheme for the assessment of off-target effects of gapmer ASOs.

中文翻译：

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使用人细胞评估gapmer反义寡核苷酸的脱靶效应。

反义寡核苷酸（ASO）由于具有与靶RNA相似的序列的ASO和意外RNA之间的互补结合而具有诱导脱靶效应的潜力。由于人类与其他动物之间基因组序列的差异，传统的动物研究不能用于评估脱靶效应诱导的毒性。因此，已经提出了通过使用人RNA数据库的计算机分析和/或使用人细胞的体外表达分析来评估脱靶效应。我们以前的研究表明，随着人类容许的错配数目的增加，人RNA序列中错配的ASO互补区域的数目急剧增加。然而，尚不清楚在细胞水平上错配基因的表达受到脱靶作用的影响程度。在这项研究中，我们通过将ASO引入人细胞并进行微阵列分析，评估了gapmer ASO的脱靶作用，后者以RNase H依赖性方式切割靶RNA。我们的数据表明，gapmer ASO诱导脱靶效应，具体取决于ASO与脱靶候选基因之间的互补程度。根据我们的结果，我们还提出了一种评估gapmer ASO脱靶效应的方案。