Comprehensive analysis of sequence-specific stability of siRNA.
Double-stranded small interfering RNAs (siRNAs) are important modulators of biological processes and hold great promise for therapeutic applications. However, serum processing of synthetic siRNAs is still largely unknown. To address this issue, serum degradation assays of 125 siRNAs were first performed in this study. Four siRNA categories of distinct serum stability were identified, including a group of siRNAs that were stable in their native form for both in vitro and in vivo assays. Fine mapping of the cleavage events occurring in serum treatment demonstrated that most occurred at two vulnerable sites, leading to a speculation that rational modification of these sites might protect most siRNAs from serum degradation. For proof of concept, an exhaustive siRNA modification study was performed. In addition to the consistent stabilization pattern revealed at these sites, our study further showed that a single modification made at the cleavage site stabilized the siRNAs to a large extent, highlighting the importance of these sites in siRNA degradation. In summary, the present study provided a comprehensive picture of serum processing of siRNA as well as a starting point for a rational siRNA modification strategy, both of which are of great importance to in vivo and therapeutic applications of siRNA.
Hong
Junmei
J
Institute of Molecular Medicine, Peking University, Beijing, China.
Huang
Yuanyu
Y
Li
Jun
J
Yi
Fan
F
Zheng
Jie
J
Huang
Huang
H
Wei
Na
N
Shan
Yongqiang
Y
An
Mingrui
M
Zhang
Hongyan
H
Ji
Jianguo
J
Zhang
Peizhuo
P
Xi
Zhen
Z
Du
Quan
Q
Liang
Zicai
Z
D000818:Animals; D002460:Cell Line; D006801:Humans; D008297:Male; D051379:Mice; D034622:RNA Interference / Q000502:physiology; D020871:RNA Stability / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D034741:RNA, Small Interfering