Preparation, characterization, and transfection efficiency of low molecular weight polyethylenimine-based nanoparticles for delivery of the plasmid encoding CD200 gene
Abstract:Various strategies have been utilized to improve both gene transfer efficiency and cell-induced toxicity of polyethylenimine (PEI), the most extensively investigated cationic polymeric vector. In this study, we sought to enhance transfection efficiency of low molecular weight PEI (LMW PEI) while maintaining its low toxicity by cross-linking LMW PEI via succinic acid linker. These modifications were designed to improve the hydrophilic–hydrophobic balance of the polymer, by enhancing the buffering capacity and m… Show more
“…In this measurement, polymers with a higher buffering capacity need larger amounts of HCl for their pH alteration. A reduction of buffering capacity is frequently associated with hydrophobic modifications . The decrease or increase of buffering capacity is highly associated with conjugation strategies used in the grafting of PEI, since conjugation methods, in that case, lead to a conversion of surface primary amines to secondary amines, thus reducing the buffering capacity .…”
Section: Resultssupporting
confidence: 89%
“…A reduction of buffering capacity is frequently associated with hydrophobic modifications . The decrease or increase of buffering capacity is highly associated with conjugation strategies used in the grafting of PEI, since conjugation methods, in that case, lead to a conversion of surface primary amines to secondary amines, thus reducing the buffering capacity . Here, the hydrophobization of PSI backbone led to substantial increase in the buffering capacity of polymers.…”
The development of polymers with low toxicity and efficient gene delivery remains a significant barrier of nonviral gene therapy. Modification and tuning of chemical structures of carriers is an attractive strategy for efficient nucleic acid delivery. Here, polyplexes consisting of plasmid DNA (pDNA) and dodecylated or non‐dodecylated polysuccinimide (PSI)‐based polycations are designed, and their transfection ability into HeLa cells is investigated by green fluorescent protein (GFP) expressing cells quantification. All cationic polymers show lower cytotoxicity than those of branched polyethyleneimine (bPEI). PSI and bPEI‐based polyplexes have comparable physicochemical properties such as size and charge. Interestingly, a strong interaction between dodecylated polycations and pDNA caused by the hydrophobic moiety is observed in dodecylated PSI derivatives. Moreover, the decrease of GFP expression is associated with lower dissociation of pDNA from polyplexes according to the heparin displacement assay. Besides, a hydrophobization of PSI cationic derivatives with dodecyl side chains can modulate the integrity of polyplexes by hydrophobic interactions, increasing the binding between the polymer and the DNA. These results provide useful information for designing polyplexes with lower toxicity and greater stability and transfection performance.
“…In this measurement, polymers with a higher buffering capacity need larger amounts of HCl for their pH alteration. A reduction of buffering capacity is frequently associated with hydrophobic modifications . The decrease or increase of buffering capacity is highly associated with conjugation strategies used in the grafting of PEI, since conjugation methods, in that case, lead to a conversion of surface primary amines to secondary amines, thus reducing the buffering capacity .…”
Section: Resultssupporting
confidence: 89%
“…A reduction of buffering capacity is frequently associated with hydrophobic modifications . The decrease or increase of buffering capacity is highly associated with conjugation strategies used in the grafting of PEI, since conjugation methods, in that case, lead to a conversion of surface primary amines to secondary amines, thus reducing the buffering capacity . Here, the hydrophobization of PSI backbone led to substantial increase in the buffering capacity of polymers.…”
The development of polymers with low toxicity and efficient gene delivery remains a significant barrier of nonviral gene therapy. Modification and tuning of chemical structures of carriers is an attractive strategy for efficient nucleic acid delivery. Here, polyplexes consisting of plasmid DNA (pDNA) and dodecylated or non‐dodecylated polysuccinimide (PSI)‐based polycations are designed, and their transfection ability into HeLa cells is investigated by green fluorescent protein (GFP) expressing cells quantification. All cationic polymers show lower cytotoxicity than those of branched polyethyleneimine (bPEI). PSI and bPEI‐based polyplexes have comparable physicochemical properties such as size and charge. Interestingly, a strong interaction between dodecylated polycations and pDNA caused by the hydrophobic moiety is observed in dodecylated PSI derivatives. Moreover, the decrease of GFP expression is associated with lower dissociation of pDNA from polyplexes according to the heparin displacement assay. Besides, a hydrophobization of PSI cationic derivatives with dodecyl side chains can modulate the integrity of polyplexes by hydrophobic interactions, increasing the binding between the polymer and the DNA. These results provide useful information for designing polyplexes with lower toxicity and greater stability and transfection performance.
“…Our results were consistent with the other investigations reporting the lower toxicity of bPEI and its derivatives following the attachment of ligands for receptor mediated delivery 9 , 50 . Since the non-targeted PEI derivatives enter the cells through adsorptive endocytosis, their accumulation in the medium lead to higher toxic effects 9 , 15 , 18 , 50 , 51 . Figure 6 Viability of bPEI and its conjugate complexed with pUMVC3-hIL12 plasmid at C/P ratios of 0.25, 4 and 8 evaluated by MTT assay in ( a ) OVCAR-3 and ( b ) HepG2 cell lines.…”
Section: Biophysical Characterization Of Polymer and Polyplexesmentioning
The objective of the present study is to conjugate L-thyroxine PEI derivative onto another PEI to compensate the amine content of the whole structure which has been utilized for the ligand conjugation. Since αvβ3 integrin receptors are over-expressed on cancer cells and there is binding site for L-thyroxine on these receptors, PEI conjugation by L-thyroxine along with restoring the PEI amine content might be an efficient strategy for targeted delivery using polymeric nanoparticles. The results demonstrated the ability of the PEI conjugate in the formation of nanoparticles with the size of around 210 nm with higher buffering capacity. The conjugated PEI derivative increased the transfection efficiency in the cell lines over-expressing integrin by up to two folds higher than unmodified PEI, whereas in the cell lines lacking the integrin receptors there was no ligand conjugation-associated difference in gene transfer ability. The specificity of transfection demonstrated the delivery of plasmid DNA through integrin receptors. Also, the results of in vivo imaging of the polyplexes revealed that 99mTc-labeled PEI/plasmid DNA complexes accumulated in kidney and bladder 4 h post injection. Therefore, this PEI derivative could be considered as an efficient targeted delivery system for plasmid DNA.
“…This reaction provided the free carboxylate moieties on PEI.The initial feed of the succinic anhydride was adjusted to result in one carboxylic group per molecule, which corresponds to 0.17 mol% of amines. The succinylated PEI was confirmed by 1 H-NMR and FT-IR as described previously21,22,25 and labeled as PEI-SUC derivative. On the other hand, the coupling of lactose-bearing galactose to the amines of LMW PEI molecule was performed between the surface amines of PEI and the aldehyde group of the glucose residue in lactose as described prevously 23.…”
The objective of the present investigation was to design a targeted polyethylenimine (PEI)-based polyplex by conjugating lactose bearing galactose groups on low molecular weight PEI (LMW PEI) grafted to a high molecular weight PEI (HMW PEI) via a succinic acid linker in order to restore the amine content of the whole conjugate used for ligand conjugation. The PEI conjugate was synthesized and characterized in terms of buffering capacity, particle size, zeta potential, plasmid condensation ability, and protection of DNA against degrading enzymes. Also, the transfection efficiency and cytotoxicity were evaluated in the cell line over-expressing asialoglycoprotein receptors (ASGPRs) and compared with the cells lacking the receptors. The results demonstrated the ability of PEI conjugate in condensation of plasmid DNA and protection against enzyme degradation. The PEI conjugate formed nanoparticles of around 75 nm with higher buffering capacity compared with unmodified PEI. The polyplexes prepared by the modified PEI could increase the level of transgene up to four folds in the cells over-expressing the receptor. The results demonstrated the separation of targeting and delivery domains could be considered as a strategy to restore the amine content of the PEI molecule utilized for targeting ligand conjugation.
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