Noncovalent Attachment of Chemical Moieties to siRNAs Using Peptide Nucleic Acid as a Complementary Linker

Bioconjugation of siRNAs with chemical moieties is an effective strategy to improve the stability and cellular uptake of siRNAs. However, chemical conjugations of siRNAs are always challenging because of siRNAs' extremely poor stability. Therefore, a new strategy to attach a chemical moiety to siRNA without chemical reaction is highly needed. Peptide nucleic acids (PNAs) are DNA analogues in which the phosphate ribose ring in the backbone is replaced with a polyamide. Compared to DNA, PNA has a higher affinity for complementary DNA and better chemical stability. We, therefore, employed PNAs as a complementary linker to attach chemical moieties to siRNAs by annealing. The objective of this study is to develop an easy but efficient strategy to noncovalently attach chemical moieties to siRNAs without chemical modification of the siRNAs. We identified a PNA complementary sequence for hybridizing with siRNAs. Also, we compared the stability and silencing effects of different siRNA-PNA chimeras, which were annealed at different termini of the siRNA. siRNAs with a PNA annealed to the 3' end of the sense strand exhibited enhanced stability in the serum and maintained a good silencing effect. The siRNA-PNA chimera was then employed in two delivery systems to deliver the PCBP2 siRNA, a potential antifibrotic siRNA, to hepatic stellate cells. In both systems, the chimera demonstrated high cellular uptake and silencing activity. The results suggested that the siRNA-PNA chimera is an easy and efficient approach to attach targeting ligands or chemical moieties to siRNAs without chemical modification of the siRNA. This new technology will greatly reduce the difficulty and cost in conjugating chemical moieties to siRNAs.