Dual centrifugation as fast and novel screening approach for optimal RNA loaded lipid-based nanoparticles

The last decade has shown increased benefits for non-viral gene delivery. To overcome the challenges of nucleic acid administration, appropriate drug delivery systems (DDS) are required. The recently approved RNA formulations have demonstrated that lipid nanoparticles (LNPs) are suitable DDS for delivering RNAs. LNPs are commonly composed of cationic and/or ionizable lipids, helper lipids and PEGylated lipids. Conventional manufacturing procedures for LNPs are mixing systems, such as microfluidics, with drawbacks in terms of time and resource consumption. The LNPs produced also pose problems with storage stability. Based on a microRNA (miRNA) model, we present dual centrifugation (DC) as a novel and reproducible way for preparing RNA loaded LNP formulations via in-vial homogenization. Our formulations show promising results in size characteristics, as well as in their cell performance. Depending on the lipid composition of the LNPs, a remarkable knockdown efficiency is achieved. With a net formulation time of 7 min, an enormously fast approach can be presented. DC offers the capability for fast LNP screenings, with a loading capacity of up to 40 vials per run. The simplicity of the method could take advantage of bedside preparation, overcoming the hurdles of storage stability for LNP formulations.