Characterization of lipid nanoparticles using macro mass photometry: Insights into size and mass

Lipid nanoparticles (LNPs) have become an important delivery system for nucleic acids, as applied in the first RNAi drug and two COVID-19 mRNA vaccines approved by the FDA. Despite advantages of their high cargo capacity, low immunogenicity allowing for redosing, scalability and low-cost manufacturing, challenges such as liver accumulation and difficulties in quality control persist for LNPs development. Conjugation of antibodies or antibody fragments onto LNPs holds promise in achieving precise targeting and higher stability of the targeting moieties on LNP surfaces. However, quality control of such multiple-component products poses additional challenges compared to LNP alone. LNP size and mass are critical quality attributes which play important roles in determining LNPs' nucleic acid cargo loading, antibody conjugation level, biodistribution, targeting capability, and overall efficacy.

Macro mass photometry (MMP), a single-molecule technique, enabled orthogonal characterization of LNPs by incorporating contrast analysis as a proxy for particle mass and combining it with size measurement. This approach thus improves the definition of LNP species within heterogeneous systems. Using MMP, this study revealed the effects of PEG-lipid concentration, mRNA encapsulation, and antibody conjugation on LNP size and mass. Additionally, the study demonstrated the MMP's utility in monitoring LNPs under various stress conditions, including high pH, low pH, oxidation, freeze-thaw cycles, and agitation.

This study represents the first use of mass photometry for LNP analysis, simultaneously characterizing the mass and size of LNPs. It highlights the potential of MMP as a valuable orthogonal tool for LNP characterization and supports LNP formulation development.