A large-scale chemical modification screen identifies design rules to generate siRNAs with high activity, high stability and low toxicity.
The use of chemically synthesized short interfering RNAs (siRNAs) is currently the method of choice to manipulate gene expression in mammalian cell culture, yet improvements of siRNA design is expectably required for successful application in vivo. Several studies have aimed at improving siRNA performance through the introduction of chemical modifications but a direct comparison of these results is difficult. We have directly compared the effect of 21 types of chemical modifications on siRNA activity and toxicity in a total of 2160 siRNA duplexes. We demonstrate that siRNA activity is primarily enhanced by favouring the incorporation of the intended antisense strand during RNA-induced silencing complex (RISC) loading by modulation of siRNA thermodynamic asymmetry and engineering of siRNA 3'-overhangs. Collectively, our results provide unique insights into the tolerance for chemical modifications and provide a simple guide to successful chemical modification of siRNAs with improved activity, stability and low toxicity.
Bramsen
Jesper B
JB
Department of Molecular Biology, University of Aarhus, Arhus, Nucleic Acid Center, University of Southern Denmark, Odense, Denmark. jebb@mb.au.dk
Laursen
Maria B
MB
Nielsen
Anne F
AF
Hansen
Thomas B
TB
Bus
Claus
C
Langkjaer
Niels
N
Babu
B Ravindra
BR
Højland
Torben
T
Abramov
Mikhail
M
Van Aerschot
Arthur
A
Odadzic
Dalibor
D
Smicius
Romualdas
R
Haas
Jens
J
Andree
Cordula
C
Barman
Jharna
J
Wenska
Malgorzata
M
Srivastava
Puneet
P
Zhou
Chuanzheng
C
Honcharenko
Dmytro
D
Hess
Simone
S
Müller
Elke
E
Bobkov
Georgii V
GV
Mikhailov
Sergey N
SN
Fava
Eugenio
E
Meyer
Thomas F
TF
Chattopadhyaya
Jyoti
J
Zerial
Marino
M
Engels
Joachim W
JW
Herdewijn
Piet
P
Wengel
Jesper
J
Kjems
Jørgen
J
D045744:Cell Line, Tumor; D002470:Cell Survival; D006801:Humans; D034622:RNA Interference; D020871:RNA Stability; D034741:RNA, Small Interfering / Q000097:blood / Q000737:chemistry* / Q000633:toxicity; D034743:RNA-Induced Silencing Complex / Q000378:metabolism
D034741:RNA, Small Interfering; D034743:RNA-Induced Silencing Complex