2'-O-[2-[(N,N-dimethylamino)oxy]ethyl]-modified oligonucleotides inhibit expression of mRNA in vitro and in vivo.
Synthesis and antisense activity of oligonucleotides modified with 2'-O-[2-[(N,N-dimethylamino)oxy] ethyl] (2'-O-DMAOE) are described. The 2'-O-DMAOE-modified oligonucleotides showed superior metabolic stability in mice. The phosphorothioate oligonucleotide 'gapmers', with 2'-O-DMAOE- modified nucleoside residues at the ends and 2'-deoxy nucleosides residues in the central region, showed dose-dependent inhibition of mRNA expression in cell culture for two targets. 'Gapmer' oligonucleotides have one or two 2'-O-modified regions and a 2'-deoxyoligonucleotide phosphorothioate region that allows RNase H digestion of target mRNA. To determine the in vivo potency and efficacy, BalbC mice were treated with 2'-O-DMAOE gapmers and a dose-dependent reduction in the targeted C-raf mRNA expression was observed. Oligonucleotides with 2'-O-DMAOE modifications throughout the sequences reduced the intercellular adhesion molecule-1 (ICAM-1) protein expression very efficiently in HUVEC cells with an IC(50) of 1.8 nM. The inhibition of ICAM-1 protein expression by these uniformly modified 2'-O-DMAOE oligonucleotides may be due to selective interference with the formation of the translational initiation complex. These results demonstrate that 2'-O-DMAOE- modified oligonucleotides are useful for antisense-based therapeutics when either RNase H-dependent or RNase H-independent target reduction mechanisms are employed.
Prakash
Thazha P
TP
Isis Pharmaceuticals Inc., 2292 Faraday Avenue, Carlsbad, CA 92008, USA.
Johnston
Joseph F
JF
Graham
Mark J
MJ
Condon
Thomas P
TP
Manoharan
Muthiah
M
D000818:Animals; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D015536:Down-Regulation / Q000187:drug effects*; D005260:Female; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000096:biosynthesis / Q000235:genetics; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D013058:Mass Spectrometry; D051379:Mice; D008807:Mice, Inbred BALB C; D015394:Molecular Structure; D016376:Oligonucleotides, Antisense / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology*; D010442:Peptide Chain Initiation, Translational / Q000187:drug effects; D014176:Protein Biosynthesis / Q000187:drug effects* / Q000235:genetics; D011493:Protein Kinase C / Q000235:genetics; D051571:Protein Kinase C-alpha; D019908:Proto-Oncogene Proteins c-raf / Q000235:genetics; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016914:Ribonuclease H / Q000378:metabolism
D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D018799:Intercellular Adhesion Molecule-1; D019908:Proto-Oncogene Proteins c-raf; C496628:PRKCA protein, human; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D016914:Ribonuclease H