Researchers Develop Acid-Degradable LNPs to Enhance mRNA Delivery

Niren Murthy from the University of California, Berkeley, Aijun Wang from the University of California, Davis, and others published a research paper titled "Acid-degradable lipid nanoparticles enhance the delivery of mRNA" in Nature Nanotechnology. The study developed an acid-degradable linker called "azido-acetal", which was used to synthesize degradable lipids composed of polyethylene glycol lipids, anionic lipids, and cationic lipids, and based on this, LNPs (RD-LNPs) that hydrolyze rapidly in endosomes were synthesized. In in vitro and in vivo experiments, RD-LNP significantly improved the performance of LNP-mRNA complexes. Compared with traditional LNPs, mRNA is more effectively delivered to the liver, lungs, spleen, and brain of mice, as well as hematopoietic stem/progenitor cells in vitro.

Acid-degradable lipids (ADLs) have great potential for generating RD-LNPs, given the pH gradient between endosomes and blood. For example, the pH of early endosomes is 6.0, the pH of late endosomes is 5.0, and the pH of blood is 7.4. Therefore, developing ADLs and using them to prepare RD-LNPs has attracted great interest among researchers. ADLs have been synthesized through linkages such as vinyl ethers, orthoesters, ketols, acetals, and hydrazines, and have been incorporated into liposomes, cationic liposomes, and LNPs with encouraging results.

However, existing ADLs degrade within days in the pH range of 6.0-6.8 and within hours in the pH range of 5.0-6.0, and are therefore transported to lysosomes for degradation. Developing ADLs that degrade rapidly around pH 6.0 has been a challenge. The hydrolysis rate of traditional acid-degradable linkers is proportional to the hydrogen ion concentration, so linkers that hydrolyze rapidly at pH 6.0-6.8 are also unstable and difficult to use at pH 7.4.

In this latest study, the research team developed an acid-degradable linker called "azido-acetal", which can be hydrolyzed within minutes in the endosome but is stable for 21 days at pH 7.4. The azido-acetal linker consists of a benzaldehyde acetal with an azide group in the para position and is hydrolyzed by a two-step mechanism that requires reduction and acid hydrolysis. This two-step hydrolysis mechanism gives azido-acetal unique stability and rapid hydrolysis ability at mildly acidic pH.

Figure 1. RD-LNPs enhance the delivery of mRNA in multiple organs in vivo. (Zhao S, et al., 2024)

Due to the weak electron-attracting properties of azido, the hydrolysis rate of azido-acetal is slow, which enables the synthesis of ADL in aqueous environments with high yields and its incorporation into LNPs. Prior to administration, azido-acetal is reduced to amines by the addition of thiols. This reduction accelerates the hydrolysis rate of azido-acetal because amines are highly electron-donating.

The research team demonstrated that the azido-acetal linker can be used as a platform to generate RD-LNPs and further demonstrated that RD-LNPs are superior to conventional LNPs in delivering mRNA to the liver, lungs, spleen, and brain of mice and hematopoietic stem/progenitor cells (HSPCs) in vitro. For example, RD-LNPs designed to deliver mRNA to the lungs successfully rescued mice from acute lung injury by delivering IL-22 mRNA, which conventional LNPs could not do. In addition, RD-LNPs containing high levels of PEG modification efficiently delivered Cas9 mRNA and gRNA to brain tissue, and the number of gene-edited brain cells was 5 times that of conventional LNPs.

Overall, this study demonstrates that engineering LNP hydrolysis rates in vivo has great potential to expand the application of LNPs in medicine.

Reference

Zhao S, et al. Acid-degradable lipid nanoparticles enhance the delivery of mRNA. Nature Nanotechnology, 2024: 1-10.