Facilitated reduction of beta-amyloid peptide precursor by synthetic oligonucleotides in COS-7 cells expressing a...

Synthetic deoxyoligonucleotides and phosphorothioate-capped oligonucleotides targeted to bases 112-128 of beta-amyloid peptide precursor (beta APP) mRNA were analyzed for their ability to reduce steady-state beta APP in COS-7 cells and in pMEP4-Rz1 cells that express a hammerhead ribozyme targeted to bases beta APP mRNA 133-148. Cells, incubated in the presence of 10 or 25 microM oligonucleotide, remained viable and morphologically identical to untreated control cells for up to 5 days. Antisense deoxyoligonucleotides beta 112C, beta 114C, and beta 116C specifically lowered beta APP in pMEP4-Rz1 cells compared to noncognate and scrambled oligonucleotide controls. The extent of the beta APP reduction did not depend on oligonucleotide length, although it did depend on the presence and proximity of the ribozyme to the oligonucleotides. beta 117N, a phosphorothioate-capped antisense oligonucleotide, also reduced beta APP levels in pMEP4-Rz1 cells; however, in this case the sense control, beta 117S, affected beta APP similarly, indicating that the observed reduction may be nonspecific. These data imply that deoxyoligonucleotides targeted immediately upstream of a ribozyme binding site can work cooperatively in vivo. Localizing the oligonucleotides and ribozyme and substrate targets to the same cellular pools further confirmed this possibility.