A highly effective and long-lasting inhibition of miRNAs with PNA-based antisense oligonucleotides

Su Young Oh, YeongSoon Ju, Heekyung Park
2009 Molecules and Cells  
MiRNAs are non-coding RNAs that play a role in the regulation of major processes. The inhibition of miRNAs using antisense oligonucleotides (ASOs) is a unique and effective technique for the characterization and subsequent therapeutic targeting of miRNA function. Recent advances in ASO chemistry have been used to increase both the resistance to nucleases and the target affinity and specificity of these ASOs. Peptide nucleic acids (PNAs) are artificial oligonucleotides constructed on a
more » ... ke backbone. PNAs have a stronger affinity and greater specificity to DNA or RNA than natural nucleic acids and are resistant to nucleases, which is an essential characteristic for a miRNA inhibitor that will be exposed to serum and cellular nucleases. For increasing cell penetration, PNAs were conjugated with cell penetrating peptides (CPPs) at N-terminal. Among the tested CPPs, Tat-modified peptide-conjugated PNAs have most effective function for miRNA inhibition. PNAbased ASO was more effective miRNA inhibitor than other DNA-based ASOs and did not show cytotoxicity at concentration up to 1,000 nM. The effects of PNA-based ASOs were shown to persist for 9 days. Also, PNA-based ASOs showed considerable stability at storage temperature. These results suggest that PNA-based ASOs are more effective ASOs of miRNA than DNA-based ASOs and PNAbased ASO technology, compared with other technologies used to inhibit miRNA activity can be an effective tool for investigating miRNA functions. MATERIALS AND METHODS Construction of reporter plasmid The plasmids used were the pGL3-control vectors (which encodes for the firefly luciferase) and the pRL-TK vectors which Molecules and Cells
doi:10.1007/s10059-009-0134-8 pmid:19812898 fatcat:3iiiyixmyvhl5phudqp4xvhnne