title: Design of antisense oligonucleotides stabilized by locked nucleic acids.
abstract: The design of antisense oligonucleotides containing locked nucleic acids (LNA) was optimized and compared to intensively studied DNA oligonucleotides, phosphorothioates and 2'-O-methyl gapmers. In contradiction to the literature, a stretch of seven or eight DNA monomers in the center of a chimeric DNA/LNA oligonucleotide is necessary for full activation of RNase H to cleave the target RNA. For 2'-O-methyl gapmers a stretch of six DNA monomers is sufficient to recruit RNase H. Compared to the 18mer DNA the oligonucleotides containing LNA have an increased melting temperature of 1.5-4 degrees C per LNA depending on the positions of the modified residues. 2'-O-methyl nucleotides increase the T(m) by only <1 degree C per modification and the T(m) of the phosphorothioate is reduced. The efficiency of an oligonucleotide in supporting RNase H cleavage correlates with its affinity for the target RNA, i.e. LNA > 2'-O-methyl > DNA > phosphorothioate. Three LNAs at each end of the oligonucleotide are sufficient to stabilize the oligonucleotide in human serum 10-fold compared to an unmodified oligodeoxynucleotide (from t(1/2) = approximately 1.5 h to t(1/2) = approximately 15 h). These chimeric LNA/DNA oligonucleotides are more stable than isosequential phosphorothioates and 2'-O-methyl gapmers, which have half-lives of 10 and 12 h, respectively.
authors.affiliation: Freie Universität Berlin, Institut für Chemie/Biochemie, Thielallee 63, 14195 Berlin, Germany.
authors.forename: Jens
authors.initials: J
authors.lastname: Kurreck
authors.forename: Eliza
authors.initials: E
authors.lastname: Wyszko
authors.forename: Clemens
authors.initials: C
authors.lastname: Gillen
authors.forename: Volker A
authors.initials: VA
authors.lastname: Erdmann
mesh_terms: D015195:Drug Design; D004355:Drug Stability; D006207:Half-Life; D006801:Humans; D007700:Kinetics; D009691:Nucleic Acid Denaturation; D020319:Oligodeoxyribonucleotides, Antisense / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000097:blood / Q000737:chemistry* / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D011955:Receptors, Drug / Q000235:genetics; D016914:Ribonuclease H / Q000737:chemistry; D012265:Ribonucleotides / Q000737:chemistry; D050916:TRPV Cation Channels; D013696:Temperature; D013873:Thionucleotides / Q000378:metabolism
chemical_list: D020319:Oligodeoxyribonucleotides, Antisense; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D011955:Receptors, Drug; D012265:Ribonucleotides; D050916:TRPV Cation Channels; C494484:TRPV1 receptor; D013873:Thionucleotides; D016914:Ribonuclease H