Overcoming technological and biological barriers to pulmonary mRNA delivery with lipid-polymer hybrid nanoparticles

Delivering mRNA to the lungs promises treatment of genetic, infectious and non-communicable lung diseases. Yet, conventional lipid nanoparticle formulations lack the stability needed to endure the harsh mechanical stress during nebulization. Additionally, once deposited in the respiratory tract, gene delivery systems may first have to overcome the mucus barrier, before actually transfecting the target cells. To overcome these challenges, we utilize lipid-polymer hybrid nanoparticles (LPH), merging the stability of pharmaceutical polymers with the transfection efficiency of cationic and ionizable lipids. LPH are produced in a robust microfluidic process, accomplish near-complete encapsulation efficiency, even after nebulization, and confer high transfection efficiency in A549 cells. To provide colloidal stability after aerosolization, we developed a coating strategy with hyaluronic acid (HA) and poly(vinyl alcohol) (PVA), ensuring ideal nebulization stability by steric shielding. This formulation showed good transfection efficiency of air-liquid interface grown Calu-3 cells, producing and covered by mucus. The potential of LPH to deliver mRNA to mammalian cells was first shown in primary human lung fibroblasts in vitro and could also be confirmed after intratracheal administration to mice in vivo. Together, these findings validate LPH as a versatile and aerosolizable mRNA delivery platform, overcoming the key technological and biological barriers in pulmonary gene delivery.