Polymers, Vol. 17, Pages 2979: PEGylation Enhances Colloidal Stability and Promotes Ligand-Mediated Targeting of LAF–Xenopeptide mRNA Complexes

Polymers doi: 10.3390/polym17222979

Authors:
Paul Folda
Eric Weidinger
Johanna Seidl
Mina Yazdi
Jana Pöhmerer
Melina Grau
David P. Minde
Mayar Ali
Ceren Kimna
Ernst Wagner

For complexation of mRNA into polyplexes, double-pH-responsive lipo-xenopeptides (XP), comprising tetraethylene pentamino succinic acid (Stp) and lipoamino fatty acids (LAFs), were combined with PEGylated lipids, either DMG-PEG 2 kDa (DMG-PEG) or azido-group-containing DSPE-PEG 2 kDa (DSPE-PEG-N3), to increase colloidal stability and to facilitate ligand-mediated targeted mRNA delivery. LAF-XPs mixed with DMG-PEG at low (1.5% and 3%) molar ratios improved colloidal stability and retained transfection efficiency. PEGylation also enabled the formulation of otherwise unstable carrier complexes and prevented aggregation induced by salt, proteins, and serum. PEGylation of more positively charged Stp-LAF2 mRNA polyplexes decreased fibrinogen adsorption. More neutral, LAF-rich Stp-LAF4 polyplexes exhibited low fibrinogen binding without PEGylation. Intravenous administration of these stabilized mRNA complexes demonstrated enhanced biosafety while preserving transfection efficiency. DSPE-PEG-N3 was selected for cell targeting after strain-promoted azide-alkyne cycloaddition (SPAAC)-mediated click-coupling of DBCO-modified ligands. Higher PEG ratios (10% and 20%) provided effective shielding but reduced transfection efficiency, a drawback known as the “PEG dilemma”. Functionalization with an EGFR-targeting ligand restored transfection in EGFR-positive cell lines in a ligand-specific manner. High transfection efficiency is consistent with a lipophilic-to-hydrophilic polarity switch of LAF-XP carriers upon endosomal protonation, triggering dissociation of the PEG lipids and deshielding of the polyplex.

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Paul Folda www.mdpi.com