The research team led by Daniel Anderson at the Massachusetts Institute of Technology has published a research paper titled “Combinatorial development of nebulized mRNA delivery formulations for the lungs” in the journal Nature Nanotechnology.

The study developed the most efficient aerosolized lipid nanoparticles (LNPs) formulation to date for delivering mRNA to the lungs, offering a more promising delivery vehicle for gene therapy in lung diseases. The delivery of mRNA in vivo holds potential for treating and preventing various genetic and infectious diseases, as well as other conditions. Notably, intramuscular injection of mRNA vaccines based on LNPs has demonstrated excellent protective efficacy against the novel coronavirus driving the COVID-19 pandemic. Beyond infectious disease vaccines, mRNA technology has been employed in cancer therapeutic vaccines, protein replacement therapies, and gene editing.

The challenges in aerosolized mRNA delivery based on LNPs are associated with several aerosolization and lung-specific barriers.

Firstly, aerosolization induces strong shear forces that can disrupt nanoparticle structures and induce aggregation.

Secondly, transfection of the bronchial epithelium requires penetration through the mucous layer, posing spatial and chemical barriers to diffusion.

Additionally, although nanoparticle uptake is most effective on the basal side of polarized lung epithelial cells, the tight junctions on the basal side make it challenging for delivered LNPs to enter.

To address aerosolization challenges, the research team first optimized the LNP component ratios to enhance stability and further improved stability through rational design of buffer conditions and additives. Subsequently, the team screened a lipid library for mRNA delivery to lung epithelium using an in vitro air-liquid interface (ALI) culture. ALI culture was proven to have good predictive capability and yielded two candidate lipids—IR-117-17 and IR-19-Py—with excellent in vivo performance.

When nebulized under optimized conditions, both lipids significantly outperformed state-of-the-art mRNA delivery vehicle formulations for the lungs and nasal cavity. Nebulized IR-117-17 LNP demonstrated a 300-fold improvement over LNPs in delivering mRNA to the lungs and a twofold improvement over previously reported hyperbranched poly(β-amino ester) (PBAE), including a 45-fold improvement in delivering to the large airways.

Inhalable mRNA holds potential for treating various diseases. However, aerosolized LNPs face unique challenges, including stability during aerosolization and penetration of cellular and extracellular barriers. The study developed a combination approach to address these obstacles. Firstly, by altering the aerosolization buffer to increase the charge of LNPs during aerosolization and adding branched polymer additives, the LNP formulation could be stabilized to resist aerosolization-induced aggregation. Subsequently, the research team synthesized ionizable and degradable lipid combinations and evaluated their delivery potential using fully differentiated air-liquid interface (ALI) cultures of primary lung epithelial cells. The final development of a combination with ionizable lipids, charge-stabilizing formulations, and stability-enhancing additives showed significant improvement in the efficiency of delivering mRNA to the lungs compared to the current state-of-the-art LNPs and polymer nanoparticles.