Surface modification of RNA nanoparticles by ionic interaction for efficient cellular uptake

Small interference RNA (siRNA) delivery has gained much attention for specific regulation of target protein via RNA interference. However, it has been challenged by inherent instability of naked RNA. To address this issue, a variety of siRNA delivery methods have been developed. The synthesis of RNA nanoparticles (RNPs) by complementary rolling circle transcription (cRCT) has been introduced as a novel route for siRNA delivery, because of the enhanced stability in serum condition and the suitable size for enhanced permeation and retention (EPR) effect. However, RNPs are negatively charged due to the innate structure of RNA, resulting in an inefficient cellular uptake without using transfection agents. In this paper, we report a simple surface modification method of RNP with calcium ion. Calcium-layered RNA nanoparticles (CaRNPs) were generated and used to demonstrate enhanced efficiency of siRNA delivery to HeLa cells in vitro. The results revealed that this simple approach could enhance the cellular uptake efficiency of RNPs in contrast to the naked RNP. Moreover, this method is also applicable for other types of RNAs such as messenger RNA (mRNA), microRNA (miRNA), or single guide RNA (sgRNA), which will greatly enhance therapeutic potential of RNPs.