TY - JOUR
T1 - Investigating the mechanism of action of DNA-loaded PEGylated lipid nanoparticles
AU - Digiacomo, Luca
AU - Renzi, Serena
AU - Quagliarini, Erica
AU - Pozzi, Daniela
AU - Amenitsch, Heinz
AU - Ferri, Gianmarco
AU - Pesce, Luca
AU - De Lorenzi, Valentina
AU - Matteoli, Giulia
AU - Cardarelli, Francesco
AU - Caracciolo, Giulio
N1 - Publisher Copyright:
© 2023 The Authors
PY - 2023/9
Y1 - 2023/9
N2 - PEGylated lipid nanoparticles (LNPs) are commonly used to deliver bioactive molecules, but the role of PEGylation in DNA-loaded LNP interactions at the cellular and subcellular levels remains poorly understood. In this study, we investigated the mechanism of action of DNA-loaded PEGylated LNPs using gene reporter technologies, dynamic light scattering (DLS), synchrotron small angle X-ray scattering (SAXS), and fluorescence confocal microscopy (FCS). We found that PEG has no significant impact on the size or nanostructure of DNA LNPs but reduces their zeta potential and interaction with anionic cell membranes. PEGylation increases the structural stability of LNPs and results in lower DNA unloading. FCS experiments revealed that PEGylated LNPs are internalized intact inside cells and largely shuttled to lysosomes, while unPEGylated LNPs undergo massive destabilization on the plasma membrane. These findings can inform the design, optimization, and validation of DNA-loaded LNPs for gene delivery and vaccine development.
AB - PEGylated lipid nanoparticles (LNPs) are commonly used to deliver bioactive molecules, but the role of PEGylation in DNA-loaded LNP interactions at the cellular and subcellular levels remains poorly understood. In this study, we investigated the mechanism of action of DNA-loaded PEGylated LNPs using gene reporter technologies, dynamic light scattering (DLS), synchrotron small angle X-ray scattering (SAXS), and fluorescence confocal microscopy (FCS). We found that PEG has no significant impact on the size or nanostructure of DNA LNPs but reduces their zeta potential and interaction with anionic cell membranes. PEGylation increases the structural stability of LNPs and results in lower DNA unloading. FCS experiments revealed that PEGylated LNPs are internalized intact inside cells and largely shuttled to lysosomes, while unPEGylated LNPs undergo massive destabilization on the plasma membrane. These findings can inform the design, optimization, and validation of DNA-loaded LNPs for gene delivery and vaccine development.
KW - DNA delivery
KW - Nanoparticle-membrane interactions
KW - PEGylation, lipid nanoparticles
UR - http://www.scopus.com/inward/record.url?scp=85167834467&partnerID=8YFLogxK
U2 - 10.1016/j.nano.2023.102697
DO - 10.1016/j.nano.2023.102697
M3 - Article
C2 - 37507061
AN - SCOPUS:85167834467
SN - 1549-9634
VL - 53
JO - Nanomedicine: Nanotechnology, Biology, and Medicine
JF - Nanomedicine: Nanotechnology, Biology, and Medicine
M1 - 102697
ER -