Ionically Crosslinked Chitosan Nanoparticles as Gene Delivery Systems: Effect of PEGylation Degree on <I>In Vitro</I> and <I>In Vivo</I> Gene Transfer

Abstract

The purpose of this work was to develop a new type of nanoparticles made of PEG-grafted chitosans (CS-g-PEG) using tripolyphosphate (TPP) as a polyanionic crosslinker and to investigate the potential of these nanostructures as gene carriers. The formation of these nanoparticles was optimised by the evaluation of the combined effects of pH, PEGylation degree and chitosan/crosslinker ratio on the particle formation. The selected CS-g-PEG/TPP nanoparticles were studied with regard to their physico-chemical properties, DNA association efficiency as well as to their toxicity and gene expression in vitro. Furthermore, the best performing nanoparticle prototypes were also evaluated for their potential for in vivo gene delivery. CS-g-PEG/TPP nanoparticles displayed a high DNA association efficiency combined with high stability and low cellular toxicity. The results of the in vitro transfection assays showed positive effects of the PEGylation in the case of particles prepared from high molecular weight chitosan, while the presence of PEG slightly decreased the efficiency of the nanoparticles based on low molecular weight chitosan. Overall, high and long lasting gene expression levels could be observed for all types of nanoparticles. Moreover, CS-g-PEG/TPP nanoparticles also mediated high gene expression levels in vivo, following nasal administration. These results indicate the potential of ionically crosslinked CS-g-PEG/TPP nanoparticles as transmucosal gene delivery systems.

Effects of ionic strength on the size and compactness of chitosan nanoparticles

“…In this work, chitosan nanoparticles were prepared using a well-known preparation method in which the positively charged chitosan is ionically crosslinked with negatively charged TPP-anions. Such chitosan nanoparticles have previously been reported to possess positive zeta potentials [13,[20][21][22][23][24][25][26][27][28][29][30]37], which is in accordance with the results found in this study. But as can be seen in Fig.…”

“…Chitosan nanoparticles crosslinked with TPP (chitosan-TPP nanoparticles) have been employed in drug delivery of both small molecular drugs, like estradiol and venlafaxine, and macromolecules, such as insulin and DNA [4,13,17,[19][20][21][22][23][24][25][26][27], and is thus a promising nanoparticulate drug delivery system. Several factors affecting the characteristics of chitosan-TPP nanoparticles have been studied, such as the ratio between crosslinker and polymer [17,[20][21][22][23][28][29][30], the molecular weight of the chitosan [17,20,21,24,[26][27][28][29], the chitosan concentration [22,23,26,28,31] and the pH [19,22,28,29]. For instance, increasing the molecular weight and the concentration of the chitosan employed in the preparation of chitosan-TPP nanoparticles have resulted in the formation of larger particles [20-24, 26-29, 31].…”

The potential of chitosan for pulmonary drug delivery

“…These nanoparticles presented an adequate size range (100-200 nm, depending on chitosan molecular weight, MW), a positive surface charge (+22 to +35 mV) and very high DNA association efficiency (>90%). The ability of the nanoparticles to entrap pDNA was in agreement with the previous results obtained for other chitosan-based nanometric systems, and it can be easily explained by the high affinity of chitosan for the DNA (86,87). Indeed, it is known that the strong electrostatic interaction exists between the phosphate groups of DNA and the amino groups of CS, as well as hydrophobic and hydrogen bonds (88).…”

Hyaluronic Acid/Chitosan-g-Poly(ethylene glycol) Nanoparticles for Gene Therapy: An Application for pDNA and siRNA Delivery

“…Grafting of PEG to chitosan was carried out by a carbodiimide-mediated reaction, using a carboxylic acid derivative of PEG (MeO-PEG-OCH 2 CO 2 H) as previously reported (10,16,17). MeO-PEG-OCH 2 CO 2 H was synthesized from a commercially available MeO-PEG-OH.…”

“…Among them, chitosan (CS)-based carriers (both polyplexes and nanoparticles) are an appealing approach due to their interesting properties and their efficiency in delivering genetic material, including plasmid DNA (pDNA) (3), oligonucleotides (4) and siRNA (5,6). Within this context, we have recently reported the feasibility and efficacy of nanoparticles made of poly(ethylene glycol)-grafted CS (PEG-g-CS) and also those from hyaluronic acid (HA) and CS as delivery carriers for pDNA (7)(8)(9)(10). The results of this previous work indicated that the presence of either PEG or HA in the polymer nanostructure has a very positive role, not only in terms of enhancing their transfection capacity but also in terms of improving the toxicity profile of classical CS-based nanosystems.…”