Combinatorial selection and delivery of thioaptamers.
Oligonucleotide-based agents are emerging as potential therapeutic agents that can be attractive alternatives for the small-molecule chemical drugs. Monothiophosphate-backbone-modified DNA aptamers (thioaptamers) that specifically and tightly bind to the RNase H domain of the HIV RT (reverse transcriptase) have been isolated from nucleic acid libraries using combinatorial selection methods. The selected thioaptamer inhibited RNase H activity of the HIV RT in in vitro studies. In cell cultures, the transfected thioaptamer markedly reduced HIV production in a dose-dependent manner. Gel electrophoretic mobility-shift assays and NMR spectroscopy showed that the selected thioaptamer binds to the isolated RNase H domain, but did not bind to a structurally similar RNase H from Escherichia coli. In cell cultures, the transfected thioaptamer showed a dose-dependent inhibition of HIV replication, with a maximal inhibition of 83%. Using various liposome-delivery agents, the DNA thioaptamer was transfected into HIV-infected astrocytoma adherent cells with greater than 70% efficiency.
Thiviyanathan
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Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555-1157, USA. thivi@nmr.utmb.edu
Somasunderam
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AD
Gorenstein
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D052157:Aptamers, Nucleotide / Q000008:administration & dosage* / Q000737:chemistry*; D020650:Combinatorial Chemistry Techniques; D004926:Escherichia coli / Q000201:enzymology; D018014:Gene Transfer Techniques; D005821:Genetic Techniques; D006678:HIV / Q000378:metabolism; D006801:Humans; D008081:Liposomes / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008954:Models, Biological; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D016914:Ribonuclease H / Q000737:chemistry / Q000378:metabolism; D014162:Transfection
D052157:Aptamers, Nucleotide; D008081:Liposomes; D016914:Ribonuclease H