Complex host cell responses to antisense suppression of ACHE gene expression

N. Galyam, D. Grisaru, M. Grifman, N. Melamed-Book, F. Eckstein, S. Seidman, A. Eldor, H. Soreq*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

3′-End-capped, 20-mer antisense oligodeoxynucleotides (AS-ODN) protected with 2′-O-methyl (Me) or phosphorothioate (PS) substitutions were targeted to acetylcholinesterase (AChE) mRNA and studied in PC12 cells. Me-modified AS-ODN suppressed AChE activity up to 50% at concentrations of 0.62-100 nM. PS-ODN was effective at 1-100 nM. Both AS-ODN displayed progressively decreased efficacy above 10 nM. In situ hybridization and confocal microscopy demonstrated dose-dependent decreases, then increases, in AChE mRNA. Moreover, labeling at nuclear foci suggested facilitated transcription or stabilization of AChE mRNA or both under AS-ODN. Intracellular concentrations of biotinylated oligonucleotide equaled those of target mRNA at extracellular concentrations of 0.02 nM yet increased only 6-fold at 1 μM ODN. Above 50 nM, sequence-independent swelling of cellular, but not nuclear, volume was observed. Our findings demonstrate suppressed AChE expression using extremely low concentrations of AS-ODN and attribute reduced efficacy at higher concentrations to complex host cell feedback responses.

Original languageEnglish
Pages (from-to)51-57
Number of pages7
JournalAntisense and Nucleic Acid Drug Development
Volume11
Issue number1
DOIs
StatePublished - 2001

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