title: Selective binding of trisamine-modified phosphorothioate antisense DNA to target mRNA improves antisense activity and reduces toxicity.
abstract: Antisense activity in living cells has been thought to occur via a mechanism involving both DNA-mediated hybridization arrest of target mRNA and RNase H-mediated mRNA digestion. Therefore an ideal antisense agent should be permeable to the cell and possess capacities (1) to form a thermally stable duplex in vivo with its target, (2) to discriminate between mRNAs with different degrees of complementarity, and (3) to form antisense/RNA complexes that are susceptible to RNase H hydrolysis. A trisamine-modified deoxyuridine derivative of a novel phosphorothioate DNA 15-mer that meets all these criteria is described here. Compared with the unmodified phosphorothioate oligomer, the phosphorothioate derivative exhibits a higher antisense activity as well as reduced cytotoxicity in cells infected with HIV-1. Our data suggest that the melting temperature (T(m)) between antisense DNA and the target mRNA is not only one of the factors contributing to this derivative's improved antisense activity. Also important are an enhanced ability to discriminate between sequences and an increased susceptibility of the DNA/mRNA complex to RNase H hydrolysis. These results will be useful in designing more active, clinically useful antisense drugs.
authors.affiliation: Department of Child Development, Kumamoto University School of Medicine, Kumamoto City 860-0811, Japan. mtmk@kaiju.medic.kumamoto-u.ac.jp
authors.forename: Makoto
authors.initials: M
authors.lastname: Matsukura
authors.forename: Tsuyosi
authors.initials: T
authors.lastname: Okamoto
authors.forename: Teruhisa
authors.initials: T
authors.lastname: Miike
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Sawai
authors.forename: Kazuo
authors.initials: K
authors.lastname: Shinozuka
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology; D001483:Base Sequence; D002264:Carboxylic Acids / Q000138:chemical synthesis; D004247:DNA / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D006678:HIV / Q000187:drug effects; D006868:Hydrolysis; D008956:Models, Chemical; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism*; D016914:Ribonuclease H / Q000378:metabolism*; D013696:Temperature; D013816:Thermodynamics; D013997:Time Factors
chemical_list: D019380:Anti-HIV Agents; D002264:Carboxylic Acids; D018076:DNA, Complementary; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D004247:DNA; D016914:Ribonuclease H