Chitosan-Graft-Branched Polyethylenimine Copolymers: Influence of Degree of Grafting on Transfection Behavior

Abstract

BackgroundSuccessful non-viral gene delivery currently requires compromises to achieve useful transfection levels while minimizing toxicity. Despite high molecular weight (MW) branched polyethylenimine (bPEI) is considered the gold standard polymeric transfectant, it suffers from high cytotoxicity. Inversely, its low MW counterpart is less toxic and effective in transfection. Moreover, chitosan is a highly biocompatible and biodegradable polymer but characterized by very low transfection efficiency. In this scenario, a straightforward approach widely exploited to develop effective transfectants relies on the synthesis of chitosan-graft-low MW bPEIs (Chi-g-bPEIx) but, despite the vast amount of work that has been done in developing promising polymeric assemblies, the possible influence of the degree of grafting on the overall behavior of copolymers for gene delivery has been largely overlooked.Methodology/Principal FindingsWith the aim of providing a comprehensive evaluation of the pivotal role of the degree of grafting in modulating the overall transfection effectiveness of copolymeric vectors, we have synthesized seven Chi-g-bPEIx derivatives with a variable amount of bPEI grafts (minimum: 0.6%; maximum: 8.8%). Along the Chi-g-bPEIx series, the higher the degree of grafting, the greater the ζ-potential and the cytotoxicity of the resulting polyplexes. Most important, in all cell lines tested the intermediate degree of grafting of 2.7% conferred low cytotoxicity and higher transfection efficiency compared to other Chi-g-bPEIx copolymers. We emphasize that, in transfection experiments carried out in primary articular chondrocytes, Chi-g-bPEI2.7% was as effective as and less cytotoxic than the gold standard 25 kDa bPEI.Conclusions/SignificanceThis work underlines for the first time the pivotal role of the degree of grafting in modulating the overall transfection effectiveness of Chi-g-bPEIx copolymers. Crucially, we have demonstrated that, along the copolymer series, the fine tuning of the degree of grafting directly affected the overall charge of polyplexes and, altogether, had a direct effect on cytotoxicity.

Size matters for in vitro gene delivery: investigating the relationships among complexation protocol, transfection medium, size and sedimentation

“…Indeed, it is noteworthy that serum-enriched medium is required for mid-to-long-term survival of cells in culture and is normally used to evaluate the serum-resistance of gene delivery vectors prior to animal studies72223. However, serum-free media are also often employed in transfection, such as for the production of recombinant proteins for biopharmaceutical applications2425.…”

Preparation and Characterization of Shape Memory Polymer Scaffolds via Solvent Casting/Particulate Leaching

“…The study of new technologies to process advanced materials has been actively promoted (26,27), from fundamental to applied materials science, in different fields (28)(29)(30)(31). Namely, the study on SMP foams has been originally proposed by Hayashi and Sokolowski (10,(32)(33)(34), and their applications in medical devices by the latter in cooperation with Prof. Yahia (8)(9)(10)35).…”

Hydrophobe-substituted bPEI derivatives: boosting transfection on primary vascular cells

“…It is worth to note that high molecular weight (MW) PEIs display not only higher transfection efficiency but also greater cytotoxicity with respect to their low MW counterparts. To improve its efficacy and cytocompatibility, PEI has been chemically tailored with a number of different substituents [9,11,12], among which, hydrophobic modification has been proven as a valuable approach. In fact, the grafting of hydrophobic chains on PEI results in increased cellular uptake via endocytosis and improved interactions between polyplexes and cell membranes [13].…”