Properties of PEI-based Polyplex Nanoparticles That Correlate With Their Transfection Efficacy

Abstract

We have evaluated the key properties of the polyethylenimine (PEI)-polyethylene glycol (PEG)-TAT peptide polyplex nanoparticles including their behavior in cells and compared them with the transfection efficacy (TE) using 11 different cell lines. We found statistically significant positive correlation between TE and the share of 50-75 nm fraction in the whole mixture of nanoparticles estimated with atomic force microscopy. Variations in PEG/PEI and N/P ratios (PEI nitrogen to DNA phosphate ratio) enabled us to find their optimal combinations, which resulted in up to 100% TE for several cell lines. Surfaces of the TE dependence of both PEG/PEI and N/P turned out to be similar in appearance for all investigated cell lines, while maximum TEs were different. We investigated subcellular transport kinetics and unpacking of the polyplex nanoparticles labeled with quantum dots (plasmid DNA) and AlexaFluor647 (block-copolymer part) using Förster Resonance Energy Transfer approach. The results demonstrated clear and statistically significant positive correlation of TE with the cellular uptake rate of the nanoparticles and negative correlation with the rate constant of their unpacking within endo/lysosomal compartments in the living cells.

Specific effects of PEGylation on gene delivery efficacy of polyethylenimine: Interplay between PEG substitution and N/P ratio

“…While this makes direct comparisons between studies difficult, it also suggests that the screening of any PEI-PEG library requires an indepth study using a wide range of substitutions and N/P ratios. While many studies have noted the importance of these variables, the relevance of the two variables acting in concert has only been considered via a transfection surface recently: Ulasov et al [33] reported the transfection surfaces (PEG chains/PEI vs N/P vs %transfection) of numerous cell types, all having similar peaks, but with the elevation dependent on cell type. The present study investigated a broader range of substitutions and N/P ratios in comparison, using 0-37.2 PEG/PEI with a maximum N/P = $70, whereas Ulasov et al used 0-8 PEG/PEI with a maximum N/P = $40.…”

“…Previous studies suggest that siRNA delivery is less inhibited by PEI PEGylation than plasmid delivery [25,28], and lower stability of PEGylated complexes was attributed to greater cargo release intracellularly. By extension, it is possible that higher cytosolic unpacking may be the same reason for a high degree of PEGylation decreasing plasmid transfection; premature unpacking of polyplexes in the cytosol decreases the likelihood of the plasmid bypassing the nuclear envelope, owing to its larger size and negative charge [33]. The question remains then, how does a low degree of PEG substitution increase the transfection efficiency of PEI, compared with unmodified PEI?…”

Evaluation of genotoxicity and cytotoxicity induced by different molecular weights of polyethylenimine/DNA nanoparticles

“…However, toxicity remains one of the challenges of this vector. There are many targets and assays that can be used to give better insight to different aspects of toxicity (Ulasov, 2011;Fischer et al, 2003;), while little is known about genotoxicity and intracellular ROS generation effects of PEI.…”

“…Cationic polymers and liposomes are the most studied nonviral vectors, with cationic polymers reported with significant differences because of their unique characteristics and potential to generate nanoparticles with DNA for gene delivery (Nakayama et al, 2005;Kafil and Omidi, 2011). Polyethylenimine (PEI) is one of the best-known cationic polymers, which has been widely used for nonviral transfection in vitro and in vivo (Ulasov et al, 2011). PEI is a cationic synthetic polymer available in linear and branched forms, and in a vast range of molecular weights, from <1000 Da up to 1.6 × 10 6 Da (Godbey and Mikos, 2001;Neu et al, 2005;Oskuee et al, 2009).…”

Fast Characterization of Polyplexes by Taylor Dispersion Analysis

“…10.1021/acs.macromol.5b01824 Macromolecules XXXX, XXX, XXX−XXX D the quantity of free (unbound) PLKC20 was ∼82% of the initial concentration (18% are complexed with DNA).…”