HYBRIDIZATION SPECIFICITY, ENZYMATIC-ACTIVITY AND BIOLOGICAL (HA-RAS)ACTIVITY OF OLIGONUCLEOTIDES CONTAINING 2,4-DIDEOXY-BETA-D-ERYTHRO-HEXOPYRANOSYL NUCLEOSIDES

Antisense oligonucleotides with a 2,4-dideoxyhexopyranosyl nucleoside incorporated at the 3'-end and at a mutation site of the Ha-ras oncoge ne mRNA were synthesized. Melting temperature studies revealed that an A-G mismatch is more stable than an A*-T mismatch with these hexopyr anosyl nucleosides incorporated at the mutation site. The oligonucleot ides are stable against enzymatic degradation. RNase H mediated cleava ge studies revealed selective cleavage of mutated Ha-ras mRNA. The oli gonucleotide containing two pyranose nucleosides at the penultimate po sition activates RNase H more strongly than natural oligonucleotides. No correlation, however, was found between DNA-DNA or RNA-DNA melting temperatures and RNase H mediated cleavage capacity. Although the A-G mismatch gives more stable hybridization than the A-T base pairing, only the oligonucleotides containing an A-T base pair are recognized by RNase H. This modification is situated 3 base pairs upstream to the cleavage site. Finally, the double pyranose modified oligonucleotide was able to reduce the growth of T24 cells (bladder carcinoma) while t he unmodified antisense oligonucleotide was not.