Published online 2 April 2004
Nucleic Acids Research, 2004, Vol. 32, No. 6 2008-2016
© 2004 Oxford University Press
2'-O-methyl-modified phosphorothioate antisense oligonucleotides have reduced non-specific effects in vitro
Department of Molecular and Cell Biology, Mail Stop FO31, The University of Texas at Dallas, PO Box 830688, Richardson, TX 75083-0688, USA, 1 Department of Biochemistry and Molecular Biology, The University of Oklahoma Health Sciences Center, Oklahoma City, OK 73190, USA and 2 Howard Hughes Medical Institute, The University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Boulevard, Dallas, TX 75390, USA
*To whom correspondence should be addressed at Department of Molecular and Cell Biology, Mail Stop FO31, The University of Texas at Dallas, PO Box 830688, Richardson, TX 75083-0688, USA. Tel: +1 972 883 2513; Fax: +1 972-883-2409; Email: dongray{at}utdallas.edu
Received January 19, 2004; Revised March 1, 2004; Accepted March 9, 2004
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 µM. 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.
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