A novel aerosolised mRNA-based vaccine delivery platform

The SARS-CoV-2 pandemic vastly accelerated development of lipid nanoparticle (LNP) complexed mRNA-based vaccines. However, LNP vaccines present significant challenges due to lack of stability and requirement for ultra-cold storage (Echaide, M. et al., 2023). mRNA encoding GFP was complexed with ToRNAdo™ Nucleic-Acid Delivery System (Factor Bioscience Ltd. MA, USA) or other commercially available reagents. Complexes were applied to lung epithelial cells either fresh, snap frozen (stored at -80C), or lyophilised (stored at RT for 28d). Complexes were also passed through a vibrating mesh nebuliser, collected and added to A549 lung epithelial cells and protein expression assessed by flow cytometry after 48h. Lipofectamine™ 3000 performed best (P < 0.001) for lyophilised transfection complexes (65.6% GFP positive cells), while ToRNAdo™ outperformed In Vivo-jetPEI® across all preparations with a range of 50.9 - 90.5% GFP positive cells (P < 0.0001). ToRNAdo™ was used to transfect A549 cells with mRNA encoding SARS-CoV-2 full-length spike glycoprotein. Supernatant harvest after 24h was added to HEK 293T-hACE2 cells. After 1h cells were incubated with SARS-CoV-2 Spike pseudotyped lentivirus (Luc-eGFP dual reporter) for 24h and transduction measured. Incubation with supernatant containing Spike protein significantly reduced viral transduction (P < 0.05), possibly due to Spike binding of HEK ACE2 receptors. Taken together we have shown <italic>in vitro</italic> that we can produce mRNA lipoplexes that are robust in terms of handling and storage and can be used to produce functional SARS-CoV-2 proteins that can be delivered by nebulisation. A preclinical vaccine mouse study is underway.