Improving RNA Interference in Mammalian Cells by 4‘-Thio-Modified Small Interfering RNA (siRNA):  Effect on siRNA Activity and Nuclease Stability When Used in Combination with 2‘-O-Alkyl Modifications

• Published in: Journal of medicinal chemistry 2006-03, Vol.49 (5), p.1624-1634
• Main Authors: Dande, Prasad, Prakash, Thazha P, Sioufi, Namir, Gaus, Hans, Jarres, Russell, Berdeja, Andreas, Swayze, Eric E, Griffey, Richard H, Bhat, Balkrishen
• Format: Article
• Language: English
• Subjects: Animals; Biological and medical sciences; Drug Stability; Heating; HeLa Cells; Humans; In Vitro Techniques; Medical sciences; Mice; Miscellaneous; Organophosphorus Compounds - chemical synthesis; Organophosphorus Compounds - chemistry; Pharmacology. Drug treatments; Plasma; PTEN Phosphohydrolase - biosynthesis; PTEN Phosphohydrolase - genetics; Ribonucleases - blood; Ribose - chemistry; RNA Interference; RNA, Messenger - biosynthesis; RNA, Messenger - genetics; RNA, Small Interfering - chemistry; RNA, Small Interfering - metabolism; RNA, Small Interfering - pharmacology; Stereoisomerism; Structure-Activity Relationship; Sulfoxides - chemistry; Thionucleosides - chemical synthesis; Thionucleosides - chemistry; Thionucleosides - pharmacology
• ISSN: 0022-2623; 1520-4804
• DOI: 10.1021/jm050822c

A systematic structure−activity relationship study of 4‘-thioribose containing small interfering RNAs (siRNAs) has led to the identification of highly potent and stable antisense constructs. To enable this optimization effort for both in vitro and in vivo applications, we have significantly improved the yields of 4‘-thioribonucleosides by using a chirally pure (R)-sulfoxide precursor. siRNA duplexes containing strategically placed regions of 4‘-thio-RNA were synthesized and evaluated for RNA interference activity and plasma stability. Stretches of 4‘-thio-RNA were well tolerated in both the antisense and sense strands. However, optimization of both the number and placement of 4‘-thioribonucleosides was necessary for maximal potency. These optimized siRNAs were generally equipotent or superior to native siRNAs and exhibited increased thermal and plasma stability. Furthermore, significant improvements in siRNA activity and plasma stability were achieved by judicious combination of 4‘-thioribose with 2‘-O-methyl and 2‘-O-methoxyethyl modifications. These optimized 4‘-thio-siRNAs may be valuable for developing stable siRNAs for therapeutic applications.