Institute for Research in Biomedicine (IRB Barcelona) and Institute for Advanced Chemistry of Catalonia (IQAC), Spanish Research Council (CSIC), Cluster Building, Baldiri i Reixac 10, 08028 Barcelona, Spain.
Chembiochem. 2013 Mar 4;14(4):510-20. doi: 10.1002/cbic.201200611. Epub 2013 Jan 29.
The development of nucleic acid derivatives to generate novel medical treatments has become increasingly popular, but the high vulnerability of oligonucleotides to nucleases limits their practical use. We explored the possibility of increasing the stability against 3'-exonucleases by replacing the two 3'-terminal nucleotides by N-ethyl-N-coupled nucleosides. Molecular dynamics simulations of 3'-N-ethyl-N-modified DNA:Klenow fragment complexes suggested that this kind of alteration has negative effects on the correct positioning of the adjacent scissile phosphodiester bond at the active site of the enzyme, and accordingly was expected to protect the oligonucleotide from degradation. We verified that these modifications conferred complete resistance to 3'-exonucleases. Furthermore, cellular RNAi experiments with 3'-N-ethyl-N-modified siRNAs showed that these modifications were compatible with the RNAi machinery. Overall, our experimental and theoretical studies strongly suggest that these modified oligonucleotides could be valuable for therapeutic applications.
核酸衍生物的开发已成为生成新型医疗手段的热点，但寡核苷酸对核酸酶的高度脆弱性限制了其实际应用。我们探索了通过用 N-乙基-N-连接核苷取代两个 3'-末端核苷酸来增加对 3'-外切核酸酶的稳定性的可能性。3'-N-乙基-N-修饰 DNA：Klenow 片段复合物的分子动力学模拟表明，这种改变对酶活性部位相邻可切割磷酸二酯键的正确定位有负面影响，因此有望保护寡核苷酸免受降解。我们验证了这些修饰赋予了完全的抗 3'-外切核酸酶性。此外，用 3'-N-乙基-N-修饰的 siRNA 进行的细胞 RNAi 实验表明，这些修饰与 RNAi 机制兼容。总的来说，我们的实验和理论研究强烈表明，这些修饰的寡核苷酸可能对治疗应用具有重要价值。
