Modulation of thermal stability can enhance the potency of siRNA

被引：50

作者：

Addepalli, Haripriya ^{[1
]}

Meena ^{[1
]}

Peng, Chang G. ^{[1
]}

Wang, Gang ^{[1
]}

Fan, Yupeng ^{[1
]}

Charisse, Klaus ^{[1
]}

Jayaprakash, K. Narayanannair ^{[1
]}

Rajeev, Kallanthottathil G. ^{[1
]}

Pandey, Rajendra K. ^{[1
]}

Lavine, Gary ^{[1
]}

Zhang, Ligang ^{[1
]}

Jahn-Hofmann, Kerstin ^{[2
]}

Hadwiger, Philipp ^{[2
]}

Manoharan, Muthiah ^{[1
]}

Maier, Martin A. ^{[1
]}

机构：

[1] Alnylam Pharmaceut Inc, Cambridge, MA 02142 USA

[2] Alnylam Europe AG, D-95326 Kulmbach, Germany

来源：

NUCLEIC ACIDS RESEARCH | 2010年 / 38卷 / 20期

关键词：

RNA INTERFERENCE; PASSENGER-STRAND; CLEAVAGE; ARGONAUTE2; INSIGHTS; DUPLEXES;

D O I：

10.1093/nar/gkq568

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

During RNA-induced silencing complex (RISC) assembly the guide (or antisense) strand has to separate from its complementary passenger (or sense) strand to generate the active RISC complex. Although this process was found to be facilitated through sense strand cleavage, there is evidence for an alternate mechanism, in which the strands are dissociated without prior cleavage. Here we show that the potency of siRNA can be improved by modulating the internal thermodynamic stability profile with chemical modifications. Using a model siRNA targeting the firefly luciferase gene with subnanomolar IC50, we found that placement of thermally destabilizing modifications, such as non-canonical bases like 2,4-difluorotoluene or single base pair mismatches in the central region of the sense strand (9-12 nt), significantly improve the potency. For this particular siRNA, the strongest correlation between the decrease in thermal stability and the increase in potency was found at position 10. Controls with stabilized sugar-phosphate backbone indicate that enzymatic cleavage of the sense strand prior to strand dissociation is not required for silencing activity. Similar potency-enhancing effects were observed as this approach was applied to other functional siRNAs targeting a different site on the firefly luciferase transcript or endogenously expressed PTEN.

引用

页码：7320 / 7331

页数：12

共 25 条

[1] Fully 2′-modified oligonucleotide duplexes with improved in vitro potency and stability compared to unmodified small interfering RNA [J].