Full deacylation of polyethylenimine dramatically boosts its gene delivery efficiency and specificity to mouse lung

Abstract

High-molecular-mass polyethylenimines (PEIs) are widely used vectors for nucleic acid delivery. We found that removal of the residual N-acyl moieties from commercial linear 25-kDa PEI enhanced its plasmid DNA delivery efficiency 21 times in vitro, as well as 10,000 times in mice with a concomitant 1,500-fold enhancement in lung specificity. Several additional linear PEIs were synthesized by acid-catalyzed hydrolysis of poly(2-ethyl-2-oxazoline), yielding the pure polycations. PEI87 and PEI217 exhibited the highest efficiency in vitro: 115-fold and 6-fold above those of the commercial and deacylated PEI25s, respectively; moreover, PEI87 delivered DNA to mouse lung as efficiently as the pure PEI25 but at a lower concentration and with a 200-fold lung specificity. These improvements stem from an increase in the number of protonatable nitrogens, which presumably results in a tighter condensation of plasmid DNA and a better endosomal escape of the PEI͞DNA complexes. As a validation of the potential of such linear, fully deacylated PEIs in gene therapy for lung diseases, systemic delivery in mice of the complexes of a short interfering RNA (siRNA) against a model gene, firefly luciferase, and PEI25 or PEI87 afforded a 77% and 93% suppression of the gene expression in the lungs, respectively. Furthermore, a polyplex of a siRNA against the influenza viral nucleocapsid protein gene and PEI87 resulted in a 94% drop of virus titers in the lungs of influenza-infected animals.influenza ͉ linear polyethylenimine ͉ short interfering RNA ͉ flu therapy ͉ transfection

Simple Modifications of Branched PEI Lead to Highly Efficient siRNA Carriers with Low Toxicity

“…These data fit very well with literature, where several modifications of branched PEI led to decreased charge and lower toxicity of the polymer (17,25). The reduction of positive charge, however, might also reduce activity of polycation in nucleic acid transfer as previously reported (13,14,19). Within our series of modifications, only two products with the highest degrees of modification showed inactivation: PEI-PROP-31 for both DNA and siRNA transfer, and PEI-SUC-19 for siRNA transfer.…”

“…Backbone modification of linear PEI 22 kDa, which on its own is one of the most potent transfection agents (6), have reduced the high gene transfer activity (13). In fact, it was demonstrated that the high activity depends on complete removal of propionamide residues from the polyisoxazoline precursor polymer (14). Linear PEI22 on its own has limited efficiency in siRNA delivery (7); the activity however can be enhanced with concatemerizing "sticky" siRNA (ssiRNA) applied for polyplex formation (15).…”

Cancer-associated Mutations Activate the Nonreceptor Tyrosine Kinase Ack1

“…Cos7 cells (from American Type Culture Collection, Manassas, VA) were maintained in Dulbecco's modified Eagle's medium with 10% iron-supplemented calf serum (JRH, Lenexa, KS) and penicillin/streptomycin. Transient transfection of cells seeded on acid-washed glass coverslips was performed using fully deacylated polyethylenimine reagent, prepared from 200-kDa poly(2-ethyl-2-oxazoline) (Sigma-Aldrich) as described (41). Transfection mixtures contained 2 g of DNA and 12 l of polyethylenimine reagent/35-mm dish.…”

Cholesterol improves the transfection efficiency of polyallylamine as a non-viral gene delivery vector

“…Zintchenko et al (2008) reported similar results for PEI modified with propionic acid or succinic acid. Thomas et al (2005) argue that transfection ability is reduced if polycation positive charge is highly reduced while in previous studies, Oskuee et al (2015b) concluded that polyplex transfection activity was not influenced by hydrophobic-modified PAA65 kDa. Additionally, according to the results of the present study regarding transfection efficiency, there is no significant difference between cholesterol modified PAA 65 kDa and the unmodified one.…”