PT  - JOURNAL ARTICLE
AU  - Marco Herrera
AU  - Jeonghwan Kim
AU  - Yulia Eygeris
AU  - Antony Jozic
AU  - Gaurav Sahay
TI  - Illuminating endosomal escape of polymorphic lipid nanoparticles that boost mRNA delivery
AID  - 10.1101/2020.12.02.407601
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.12.02.407601
4099  - http://biorxiv.org/content/early/2020/12/03/2020.12.02.407601.short
4100  - http://biorxiv.org/content/early/2020/12/03/2020.12.02.407601.full
AB  - Lipid-based nanoparticles (LNPs) for the delivery of mRNA have jumped to the forefront of non-viral gene delivery. Despite this exciting development, poor endosomal escape after LNP cell entry remains an unsolved, rate-limiting bottleneck. Here we report the use of galectin 8-GFP (Gal8-GFP) cell reporter system to visualize the endosomal escaping capabilities of LNPs encapsulating mRNA. LNPs substituted with phytosterols in place of cholesterol exhibited various levels of Gal8 recruitment in the Gal8-GFP reporter system. In live-cell imaging, the results of the LNPs containing β-sitosterol (LNP-Sito) showed a 10-fold increase in detectable endosomal perturbation events when compared to the standard cholesterol LNPs (LNP-Chol), suggesting the superior capability of LNP-Sito to escape from endosomal entrapment. Trafficking studies of these LNPs showed strong localization with late endosomes. This highly sensitive and robust Gal8-GFP reporter system can be a valuable tool to elucidate intricacies of LNP trafficking and ephemeral endosomal escape events, enabling the advancement in gene delivery.Competing Interest StatementG.S. is a co-inventor in patent application US20200129445A1 that details LNP formulations containing cholesterol derivatives.