Synthetic siRNA molecules without chemical modifications are easily degraded in the body, and 2'--modifications are frequently introduced to enhance stability. However, such chemical modifications tend to impact the gene knockdown potency of siRNA negatively. To circumvent this problem, we previously developed a prodrug-type siRNA bearing 2'--methyldithiomethyl (MDTM) groups, which can be converted into unmodified siRNA under the reductive environment in cells. In this study, we developed a nuclease-resistant prodrug-type 2'--MDTM siRNA for deployment in future animal experiments. To rationally design siRNA modified with a minimal number of 2'--MDTM nucleotide residues, we identified the sites susceptible to nuclease digestion and tolerant to 2'--methyl (2'-OMe) modification in the antisense strand of apolipoprotein B-targeted siRNA. Subsequently, we optimized the positions where the 2'-OMe and 2'--MDTM groups should be incorporated. siRNA bearing the 2'--MDTM and 2'-OMe groups at their respective optimized positions exhibited efficient knockdown potency and enhanced stability in serum.

中文翻译：

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合理设计前药型 apoB 靶向 siRNA，在不影响基因沉默效力的情况下改善核酸酶抗性

未经化学修饰的合成siRNA分子很容易在体内降解，并且经常引入2'-修饰以增强稳定性。然而，此类化学修饰往往会对 siRNA 的基因敲除效力产生负面影响。为了解决这个问题，我们之前开发了一种带有2'--甲基二硫甲基（MDTM）基团的前药型siRNA，它可以在细胞的还原环境下转化为未修饰的siRNA。在这项研究中，我们开发了一种核酸酶抗性前药类型2'--MDTM siRNA，用于未来的动物实验。为了合理设计用最少数量的2'-MDTM核苷酸残基修饰的siRNA，我们在载脂蛋白B靶向的反义链中鉴定了易受核酸酶消化和耐受2'-甲基（2'-OMe）修饰的位点siRNA。随后，我们优化了 2'-OMe 和 2'-MDTM 基团应合并的位置。在各自优化位置带有2'-MDTM和2'-OMe基团的siRNA表现出有效的敲低效力和增强的血清稳定性。