Modified poly(lactic-co-glycolic acid) nanoparticles for enhanced cellular uptake and gene editing in the lung

Abstract

Surface-modified poly(lactic-co-glycolic acid) (PLGA)/poly(β-aminoester)(PBAE)nanoparticles (NPs) have shown great promise in gene delivery. In this work, the pulmonary cellular uptake of these NPs is evaluated and surface-modified PLGA/PBAE NPs are shown to achieve higher cellular association and gene editing than traditional NPs composed of PLGA or PLGA/PBAE blends alone.

Keywords: cell-penetrating peptide; cellular uptake; gene editing; nanoparticles; pulmonary delivery.

Figure 1

A) Schematic representation of NPs. NPs were formulated with a polymer blend of PLGA and PBAE. Particles encapsulated plasmid DNA (not shown in schematic) or PNA/DNA cargo and were surface modified with a DSPE-PEG-MPG moiety. B) SEM image of NPs. C) Size characterization of NPs when suspended in aqueous solution (hydrodynamic diameter) or after lyophilization (dry powder diameter).

Figure 2

Lung tissue from mice treated with, NP-0, NP-15, and NP-15-MPG particles. Lung tissue was harvested 4 hours post administration. The nucleus (Dapi) is shown in blue. Macrophages and CECs are shown in red, while NPs are shown in green. Insets depict co-localization of NPs with macrophages or CECs. All images were taken at 20X magnification. White scale bar is equal to 58 µm.

Figure 3

FACS analysis of mice lungs treated with C6 loaded NPs four hours post IN administration. A) FL1/FSC channel of all lung cells. B) Percentage of C6+ lung cells. C) Percentage of C6+ Macrophages. D) Percentage of C6+ Alveolar Epithelial Cells (Type 1). *Indicates statistical significance in particle cellular association of NP-15-MPG relative to untreated and unmodified particle formulations; p<0.05.

Figure 4

Expression of EGFP after correction of EGFP gene splice site with PNA/DNA oligos. A) Schematic of PNA mediated homologous site-specific recombination of donor DNA to correct EGFP gene and restore EGFP expression. B) Percentage of GFP+ lung cells after treatment with different NP formulations. C) Percentage of GFP+ macrophage cells. D) Percentage of GFP+ type I alveolar epithelial cells. Experiments were repeated for NP-15-MPG and NP-15 Blank treated groups to increase the number of replicates (red symbols). *Indicates statistical significance in gene correction for the of NP-15-MPG treated groups above control (NP-15 Blank); p<0.05.