Increased Potency of an Aptameric G-Rich Oligonucleotide Is Associated with Novel Functional Properties of Phosphorothioate Linkages

We previously showed that inhibition of the expression of CD28 (an essential immune receptor on T cells) mediated by a phosphorothioate (PS)-modified aptameric oligodeoxynucleotide (ODN) sequence, GR1, resulted in reduced T cell responses in vitro and in vivo. Using GR1 sequences differing only in the amount of terminal PS linkages (chimeric SO-ODN), the present study demonstrated that even after a substantial reduction in PS linkages, this 18-mer ODN sequence could still confer functionality in the ODN-mediated inhibition of CD28 expression. We showed that secondary structure and full retention of the ability to form a specific protein-ODN complex and to increase cellular uptake in activated Jurkat T cells were critical parameters in the determination of the magnitude of bioactivity of chimeric SO-ODN. We report that a chimeric SO-ODN with terminal PS linkages that total 9 (ICN 17221) or 12 (ICN 17263) was sufficient to inhibit CD28 expression and suppress in vivo inflammatory ear responses to contact allergen in mice with similar potency to the 17-thioate S-ODN (ICN 16064). Interestingly, all chimeric SO-ODN showed similar in vitro nuclease resistance. These data suggest alternate functional properties for PS linkages, unrelated to nuclease resistance, in enhancing the bioactivity of a G-rich aptamer.