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 essentia l immune receptor on T cells) mediated by a phosphorothioate (PS)-modified aptameric oligodeoxynucleotide (ODN) sequence, GR1, resulted in reduced T c ell responses in vitro and in vivo. Using GR1 sequences differing only in t he amount of terminal PS linkages (chimeric SO-ODN), the present study demo nstrated that even after a substantial reduction in PS linkages, this 18-me r ODN sequence could still confer functionality in the ODN-mediated inhibit ion of CD28 expression. We showed that secondary structure and full retenti on of the ability to form a specific protein-ODN complex and to increase ce llular uptake in activated Jurkat T cells were critical parameters in the d etermination 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) o r 12 (ICN 17263) was sufficient to inhibit CD28 expression and suppress in vivo inflammatory ear responses to contact allergen in mice with similar po tency to the 17-thioate S-ODN (ICN 16064). Interestingly, all chimeric SO-O DN showed similar in vitro nuclease resistance. These data suggest alternat e functional properties for PS linkages, unrelated to nuclease resistance, in enhancing the bioactivity of a G-rich aptamer.