Morphological Characterization of Self-Amplifying mRNA Lipid Nanoparticles

作者：Jacob L. Thelen, Wellington Leite, Volker S. Urban, Hugh O’Neill, Alexander Grishaev, Joseph E. Curtis, Susan Krueger, Maria Monica Castellanos · 发表于：ACS Nano · 年份：2024 · DOI：10.1021/acsnano.3c08014 · 被引用次数：44 · 研究领域：RNA Interference and Gene Delivery、Bacteriophages and microbial interactions、Advanced biosensing and bioanalysis techniques

The mRNA technology has emerged as a rapid modality to develop vaccines during pandemic situations with the potential to protect against endemic diseases. The success of mRNA in producing an antigen is dependent on the ability to deliver mRNA to the cells using a vehicle, which typically consists of a lipid nanoparticle (LNP). Self-amplifying mRNA (SAM) is a synthetic mRNA platform that, besides encoding for the antigen of interest, includes the replication machinery for mRNA amplification in the cells. Thus, SAM can generate many antigen encoding mRNA copies and prolong expression of the antigen with lower doses than those required for conventional mRNA. This work describes the morphology of LNPs containing encapsulated SAM (SAM LNPs), with SAM being three to four times larger than conventional mRNA. We show evidence that SAM changes its conformational structure when encapsulated in LNPs, becoming more compact than the free SAM form. A characteristic "bleb" structure is observed in SAM LNPs, which consists of a lipid-rich core and an aqueous RNA-rich core, both surrounded by a DSPC-rich lipid shell. We used SANS and SAXS data to confirm that the prevalent morphology of the LNP consists of two-core compartments where components are heterogeneously distributed between the two cores and the shell. A capped cylinder core-shell model with two interior compartments was built to capture the overall morphology of the LNP. These findings provide evidence that bleb two-compartment str...