title: 2'-O-methyl-modified phosphorothioate antisense oligonucleotides have reduced non-specific effects in vitro.
abstract: Antisense oligodeoxynucleotides (ODNs) have biological activity in treating various forms of cancer. The antisense effects of two types of 20mer ODNs, phosphorothioate-modified ODNs (S-ODNs) and S-ODNs with 12 2'-O-methyl groups (Me-S-ODNs), targeted to sites 109 and 277 of bcl-2 mRNA, were compared. Both types were at least as effective as G3139 (Genta, Inc.) in reducing the level of Bcl-2 protein in T24 cells following a 4 h transfection at a dose of 0.1 micro M. Circular dichroism spectra showed that both types formed A-form duplexes with the complementary RNA, and the melting temperatures were in the order of Me-S-ODN.RNA > normal DNA.RNA > S-ODN.RNA. In comparison with the S-ODN, the Me-S-ODN had reduced toxic growth inhibitory effects, was less prone to bind the DNA-binding domain A of human replication protein A, and was as resistant to serum nucleases. Neither type of oligomer induced apoptosis, according to a PARP-cleavage assay. Hybrids formed with Me-S-ODN sequences were less sensitive to RNase H degradation than those formed with S-ODN sequences. Despite this latter disadvantage, the addition of 2'-O-methyl groups to a phosphorothioate-modified ODN is advantageous because of increased stability of binding and reduced non-specific effects.
authors.affiliation: Department of Molecular and Cell Biology, Mail Stop FO31, The University of Texas at Dallas, PO Box 830688, Richardson, TX 75083-0688, USA.
authors.forename: Byong Hoon
authors.initials: BH
authors.lastname: Yoo
authors.forename: Elena
authors.initials: E
authors.lastname: Bochkareva
authors.forename: Alexey
authors.initials: A
authors.lastname: Bochkarev
authors.forename: Tung-Chung
authors.initials: TC
authors.lastname: Mou
authors.forename: Donald M
authors.initials: DM
authors.lastname: Gray
mesh_terms: D017209:Apoptosis; D002455:Cell Division; D045744:Cell Line, Tumor; D002942:Circular Dichroism; D004247:DNA / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation; D006801:Humans; D008745:Methylation; D016376:Oligonucleotides, Antisense / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D012313:RNA / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D051716:Replication Protein A; D016914:Ribonuclease H / Q000378:metabolism; D013696:Temperature; D013873:Thionucleotides / Q000737:chemistry*
chemical_list: D004268:DNA-Binding Proteins; D016376:Oligonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; C497167:RPA1 protein, human; D051716:Replication Protein A; D013873:Thionucleotides; D012313:RNA; D004247:DNA; D016914:Ribonuclease H