Down-regulation of VEGF mRNA expression by 2'-O,4'-C-ethylene-bridged nucleic acid (ENA) antisense oligonucleotides and investigation of non-target gene expression.
We studied the properties of 2'-O,4'-C-ethylene-bridged nucleic acid (ENA) oligonucleotides as antisense molecules. Compared to a phosphorothioate (PS) DNA and RNA heteroduplex, a duplex of an ENA/PS/ENA gapmer with RNA was a more effective substrate for RNase H-mediated cleavage. We designed ENA antisense oligonucleotides (AON) targeting human vascular endothelial growth factor (VEGF) mRNA. ENA AON with a cationic polymer considerably down-regulated VEGF mRNA expression, while no down-regulation by PS AON was observed. We also investigated the influence on the gene expression of genes other than VEGF by a gene database homology search and RT-PCR analysis. This method turned out to be an efficient approach to search for non-target genes sequence-specifically down-regulated by AON. In rat plasma, an ENA/PS/ENA gapmer was very stable after 24 hr, while a bridged nucleic acid (BNA) or locked nucleic acid (LNA) oligonucleotide and a PS oligonucleotide were half degraded in 4 hr. The high stability of ENA oligonucleotides in plasma would make ENA oligonucleotides ideal for in vivo studies.
Morita
Koji
K
Exploratory Chemistry Research Laboratories, Sankyo Co., Ltd., Tokyo 140-8710, Japan.
Yamate
Koji
K
Kurakata
Shin-ichi
S
Abe
Koji
K
Imanishi
Takeshi
T
Koizumi
Makoto
M
D015536:Down-Regulation / Q000187:drug effects*; D016228:Endothelial Growth Factors / Q000235:genetics*; D004789:Enzyme Activation; D015870:Gene Expression / Q000187:drug effects; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics*; D008222:Lymphokines / Q000235:genetics*; D009696:Nucleic Acids / Q000235:genetics*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics*; D016914:Ribonuclease H / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D036341:Intercellular Signaling Peptides and Proteins; D008222:Lymphokines; D009696:Nucleic Acids; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D016914:Ribonuclease H