Poly[Lys-(AEDTP)]:  A Cationic Polymer That Allows Dissociation of pDNA/Cationic Polymer Complexes in a Reductive Medium and Enhances Polyfection

Abstract

Polyplexes of high stability resulting from the condensation of a plasmid DNA by a cationic polymer are widely used to develop polymer-based gene delivery systems. However, the plasmid must be released from its vector once inside the cells for an efficient expression of the exogenous gene in the cell nucleus. We have designed a disulfide-containing cationic polymer termed poly[Lys-(AEDTP)] which allowed for the formation of polyplexes and the release of the plasmid in a reductive medium. The amino groups of polylysine were substituted with 3-(2-aminoethyldithio)propionyl residues in order to have each amino group of poly[Lys-(AEDTP)] interacting with a phosphate DNA linked to the polymer backbone via a disulfide bond. As evidenced by agarose gel electrophoresis and ethidium bromide/pDNA fluorescence restoration, poly[Lys-(AEDTP)] polyplexes were decondensed and the plasmid released upon treatment with either dithiothreitol, glutathione in the presence of glutathione reductase, or the thioredoxin reductase. Electron microscopy showed that polyplexes exhibiting spherical particles of a mean size at about 100 nm were decondensed in the presence of glutathione and exhibited filamentous aggregates. Finally, we found that the transfection of 293T7 and HepG2 cells was 10- and 50-fold more efficient with poly[Lys-(AEDTP)] polyplexes, respectively, than with poly[Lys] polyplexes. These results indicate that disulfide-containing cationic polymers must be borne in mind for developing polymer-base gene delivery systems.

An influenza virus-inspired polymer system for the timed release of siRNA

“…The accumulation of cationic species could result in unwanted toxicity especially when administered in multiple doses. To overcome the release problem of siRNA, polymers have been designed to incorporate side chain molecules that trigger a release when stimulated using temperature 23 , pH 24 , redox potential 25,26 , light 27 , electrical pulse 28,29 and enzymatic degradation 30,31 . Such release mechanisms rely on either external (remote) or environmental stimuli.…”

Evaluation of cystamine-modified hyaluronic acid/chitosan polyplex as retinal gene vector

“…These assays were performed as described previously [8,9]. Briefly, a 0.4 M glutathione reduced-form (Sigma) stock solution was prepared in 0.1 M potassium phosphate buffer (pH 7.5), with 2 mM EDTA, pH 7.5 (potassium phosphate buffer with EDTA from here on referred to as PB).…”

“…In this study, HA was modified with cystamine through a coupling reaction with the carboxyl groups of HA. This modification with cystamine not only adds amine groups to the polymer but it also contains a disulfide bond that can be cleaved in the presence of intercellular glutathione, thereby promoting a more rapid release of genetic material [8,9].…”

Bioreducible Cationic Polymers for Gene Transfection

“…A typical synthesis route is the preparation of cationic polymers with pyridyldithio residue, which is then modified with suitable thiol compounds via an exchange reaction (Figure 5a). By this method, the pLL containing disulfide linkages in the polymer side chains (termed as poly[Lys-(AEDTP)]) was prepared through chemical modification of the primary amines in pLL with Nsuccinimidyl-3-(2-pyridyldithio)propionate, followed by an exchange reaction with mercapthoethylamine (Pichon et al, 2002). The polyplexes of poly[Lys-(AEDTP)] can transfect HeLa cells with a level of gene expression 10-fold higher than that of parent pLL.…”