Improved Methods of Antisense Therapy

Antisense oligonucleotides, either externally administered or generated by transcription ofinserted genes, show promise as agents for controlling the expression of specific genes. However, theduplexes formed between antisense oligonucleotides and a target RNA can be dissociated by ribosomesduring translation. Our objective research is to develop a technique for attaching an antisense moleculeto its target after paring. Such a complex should be resistant to dissociation and thus increase both theeffectiveness of the antisense procedure and the number of sequences that can be targeted. Theapproach can also be adapted to antigene strategies. In this case, a triplex is formed in which the addedthird stand is attached to the target duplex DNA. The antigene approach has the advantage ofinactivating gene expression at the DNA source, which is generally present in only one of a few copies,whereas RNA can be present in thousands of copies which are constantly being replaced. The antisenseversion of this proposal will be developed using tumor necrosis factor alpha mRNA as a model target.Phase II will involve applications of this procedure to the inhibition of gene expression in cultured cellsand its extension to DNA targets.