Facilitated Reduction of β-Amyloid Peptide Precursor by Synthetic Oligonucleotides in COS-7 Cells Expressing a Hammerhead Ribozyme

Synthetic deoxyoligonucleotides and phosphorothioate-capped oligonucleotides targeted to bases 112–128 of β-amyloid peptide precursor (βAPP) mRNA were analyzed for their ability to reduce steady-state βAPP in COS-7 cells and in pMEP4-Rz1 cells that express a hammerhead ribozyme targeted to bases βAPP mRNA 133–148. Cells, incubated in the presence of 10 or 25 μM oligonucleotide, remained viable and morphologically identical to untreated control cells for up to 5 days. Antisense deoxyoligonucleotides β112C, β114C, and β116C specifically lowered βAPP in pMEP4-Rz1 cells compared to noncognate and scrambled oligonucleotide controls. The extent of the βAPP reduction did not depend on oligonucleotide length, although it did depend on the presence and proximity of the ribozyme to the oligonucleotides. β117N, a phosphorothioate-capped antisense oligonucleotide, also reduced βAPP levels in pMEP4-Rz1 cells; however, in this case the sense control, β117S, affected βAPP similarly, indicating that the observed reduction may be nonspecific. These data imply that deoxyoligonucleotides targeted immediately upstream of a ribozyme binding site can work cooperativelyin vivo.Localizing the oligonucleotides and ribozyme and substrate targets to the same cellular pools further confirmed this possibility.