Nucleoside Optimization for RNAi: A High-Throughput Platform

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• 作者：Gabor Butora ; Denise M. Kenski ; Abby J. Cooper ; Wenlang Fu ; Ning Qi ; Jenny J. Li ; W. Michael Flanagan ; Ian W. Davies
• 刊名：Journal of the American Chemical Society
• 出版年：2011
• 出版时间：October 26, 2011
• 年：2011
• 卷：133
• 期：42
• 页码：16766-16769
• 全文大小：892K
• 年卷期：v.133,no.42(October 26, 2011)
• ISSN：1520-5126

文摘

The RNA induced silencing complex (RISC) contains at its core the endonuclease Argonaute (Ago) that allows for guide strand (GS)-mediated sequence-specific cleavage of the target mRNA. Functionalization of the sugar/phosphodiester backbone of the GS, which is in direct contact with Ago, presents a logical opportunity to affect RISC鈥檚 activity. A systematic evaluation of modified nucleosides requires the synthesis of phosphoramidites corresponding to all four canonical bases (A, U, C, and G) and their sequential evaluation at each position along the 21-nucleotide-long GS. With the use of a platform approach, the sequential replacement of canonical bases with inosine greatly simplifies the problem and defines a new activity baseline toward which the corresponding sugar-modified inosines are compared. This approach was validated using 2鈥?O-benzyl modification, which demonstrated that positions 5, 8, 15, and 19 can accommodate this large group. Application of this high-throughput methodology now allows for hypothesis-driven rational design of highly potent, immunologically silent and stable siRNAs suitable for therapeutic applications.