Suppression of platelet-type 12-lipoxygenase activity in human erythroleukemia cells by an RNA-cleaving DNAzyme.
Human platelet-type 12-lipoxygenase (12-LOX) and its metabolites play a crucial role in tumor angiogenesis. A "10-23" deoxyribozyme (DNAzyme) and its phosphorothioate-modified version were designed and synthesized against the 12-LOX mRNA. Both DNAzymes were able to cleave their substrate efficiently in a time- and concentration-dependent manner in vitro. Under a multiple turnover condition, both performed well at 37 degrees C, showing the k(cat) of 1 and 0.26 min(-1), respectively. The phosphorothioate modification of the DNAzyme significantly increased its stability in cells without a substantial loss of kinetic efficiency in vitro. In a cell culture system, transfection of the DNAzymes into HEL cells resulted in a significant down-regulation of the 12-LOX mRNA. Furthermore, the cell extracts from the DNAzyme-transfected cells exhibited a marked reduction in the 12-LOX enzyme activity. The present results indicated the potential use of DNAzyme technology for gene function study and cancer therapy.
Liu
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Department of Molecular Virology and Bioengineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100080, People's Republic of China.
Cheng
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R
Sun
L Q
LQ
Tien
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P
D001092:Arachidonate 12-Lipoxygenase / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D001792:Blood Platelets / Q000201:enzymology*; D021881:DNA, Catalytic / Q000737:chemistry / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D004915:Leukemia, Erythroblastic, Acute; D009843:Oligoribonucleotides / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D021881:DNA, Catalytic; D009843:Oligoribonucleotides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D001092:Arachidonate 12-Lipoxygenase