Magnetic resonance imaging-based quantification of endosomal escape using iron oxide nanoparticles-loaded lipid nanoparticles

Lipid nanoparticles (LNPs) have attracted attention as a promising and advanced platform for the delivery of nucleic acid-based therapeutics. The therapeutic efficacy of LNP-based drugs depends heavily on endosomal escape. However, few methods are available for quantifying the efficiency of endosomal escape. In this study, we developed a novel method to quantify the endosomal escape efficiency using magnetic resonance imaging (MRI). We synthesized ultrasmall iron oxide nanoparticles (IONPs) and incorporated them into LNPs to produce IO@LNPs. After cells internalized the IO@LNPs, we observed a decline in the R ₂ relaxation over time, suggesting that free IONPs were dispersed due to endosomal escape. Biological electron microscopy further corroborated this finding, showing a strong correlation between the R ₂ relaxation and the number of intracellular vesicles harboring the intact IO@LNPs. Furthermore, in-vivo MRI experiments in mice demonstrated an initial drop and a gradual increase in the T ₂ signal at the tissue site where IO@LNPs were injected, indicating the potential for in-vivo application of our method. Our findings could lead to advancements in LNP-based nucleic acid delivery by enhancing the understanding of endosomal escape dynamics.