Properties and synergistic effects of a nonionic backbone and aminoalkyl modified nucleosides in RNAs.

In this study, we report the synthesis of two formacetal (FA)-linked dimer building blocks, namely 2'-O-methyluridyl-2'-O-methyluridine and 2'-O-methyluridyl-2'-O-aminoethyluridine. We utilize the former dimer in combination with (S)-5'-C-aminopropyl-2'-O-methylnucleosides (5'-APs) as a neutral trimer unit, and the latter dimer as a cationic unit. Double-stranded RNA containing the neutral trimer unit exhibits greater stability compared to the cationic unit and maintains nuclease stability in a serum-containing buffer. Furthermore, this unit appears to establish additional hydrogen bonds with complementary bases, as supported by modeling simulations and mismatch melting temperature assays. Importantly, siRNAs modified with this unit enhance RNA interference activity in cultured cells. These findings suggest that the trimer unit holds promise for therapeutic siRNAs.

Synthesis of siRNAs incorporated with cationic peptides R8G7 and R8A7 and the effect of the modifications on siRNA properties.

Small interfering RNA (siRNA) can be used as an innovative next-generation drug. However, there are several challenges in the therapeutic application of siRNAs, including their low cell membrane permeability. In this study, we designed and synthesized siRNAs, incorporating the cationic peptides R8G7 and R8A7 to improve cell membrane permeability of siRNAs. Thermal denaturation studies revealed that R8G7 and R8A7 modifications increased the thermal stability of the siRNA duplexes. Incorporating these peptides at the 3'-ends of the siRNA passenger strands increased the stability of the siRNAs in a buffer containing bovine serum. Further, we found that the peptide-siRNA conjugates did not show sufficient RNA interference (RNAi) activity in the absence of the transfection reagent; however, when the transfection reagent was used, the peptide-siRNA conjugates preserved their RNAi activity.