The power of sulfhydryl groups: Thiolated lipid-based nanoparticles enhance cellular uptake of nucleic acids

Daniel Stengel; Arne M Jörgensen; Ilaria Polidori; Paul Kapitza; Fabrizio Ricci; Andreas Bernkop-Schnürch

doi:10.1016/j.jcis.2023.10.039

The power of sulfhydryl groups: Thiolated lipid-based nanoparticles enhance cellular uptake of nucleic acids

J Colloid Interface Sci. 2023 Oct 18;654(Pt B):1136-1145. doi: 10.1016/j.jcis.2023.10.039. Online ahead of print.

Authors

Daniel Stengel¹, Arne M Jörgensen¹, Ilaria Polidori¹, Paul Kapitza², Fabrizio Ricci³, Andreas Bernkop-Schnürch⁴

Affiliations

¹ Department of Pharmaceutical Technology, University of Innsbruck, Institute of Pharmacy, Center for Chemistry and Biomedicine, 6020 Innsbruck, Austria.
² Department of Pharmaceutical Chemistry, University of Innsbruck, Institute of Pharmacy, Center for Chemistry and Biomedicine, 6020 Innsbruck, Austria.
³ Thiomatrix Forschungs- und Beratungs GmbH, Trientlgasse 65, 6020 Innsbruck, Austria.
⁴ Department of Pharmaceutical Technology, University of Innsbruck, Institute of Pharmacy, Center for Chemistry and Biomedicine, 6020 Innsbruck, Austria. Electronic address: Andreas.Bernkop@uibk.ac.at.

PMID: 39491903
DOI: 10.1016/j.jcis.2023.10.039

Abstract

Aim: The aim of the study was to evaluate the effect of thiolation of lipid-based nanoparticles (LNPs) on cellular uptake of nucleic acids.

Methods: A thiolated surfactant was synthesized by binding palmitic acid covalently to cysteine. Green fluorescent protein (GFP) encoding plasmid DNA (pDNA) was used as model nucleic acid and incorporated via hydrophobic ion-pairing with a cationic cholesterol derivate (DcCholesterol) in LNPs that were prepared by solvent injection method using the thiolated surfactant for surface decoration. LNPs were characterized regarding size, polydispersity index, zeta potential and stability in biorelevant media. The endosomal escape properties of LNPs were evaluated by erythrocytes interaction studies. Cell viability and transfection efficiency on HEK293 cells were investigated.

Results: The structure of the thiolated surfactant was confirmed by ¹H NMR and FT-IR. LNPs containing the nucleic acid-DcCholesterol complex with and without the thiolated surfactant were developed and displayed sizes in the range from 173 nm to 233 nm with a narrow size distribution (PDI < 0.3) and a negative zeta potential. LNPs showed no significant increase in size after 4 h of incubation in artificial body fluids. Erythrocytes interaction studies revealed enhanced endosomal escape properties for thiolated LNPs compared to non-thiolated LNPs. LNPs showed in concentrations lower than 0.74 mg/mL cell viability ≥ 80 %. Transfection studies on HEK cells with thiolated LNPs compared to non-thiolated LNP showed a 4.6-fold higher expression of GFP.

Conclusion: Surface thiolation of LNPs represents a promising tool for enhancing intracellular nucleic acid delivery of LNPs.

Keywords: Endosomal escape; Gene delivery; Lipid-based nanoparticles; Thiolated surfactant; Transfection.