Tailoring of Chitosans for Gene Delivery: Novel Self-Branched Glycosylated Chitosan Oligomers with Improved Functional Properties

Abstract

Chitosan is a promising biomaterial with an attractive safety profile; however, its application potential for gene delivery is hampered by poor compatibility at physiological pH values. Here we have tailored the molecular architecture of chitosan to improve the functional properties and gene transfer efficacy of chitosan oligomers and have developed self-branched glycosylated chitosan oligomer (SB-TCO) substituted with a trisaccharide containing N-acetylglucosamine, AAM. SB-TCO was prepared by controlled depolymerization of chitosan, followed by simultaneous branching and AAM substitution. The product was fully soluble at physiological pH and complexed plasmid DNA into polyplexes of high colloidal and physical stability. SB-TCO displayed high transfection efficacy in HEK293 cells, reaching transfection efficiencies of up to 70%, and large amounts of transgene were produced. Gene transfer efficacy was confirmed in HepG2 cells, where gene expression levels mediated by SB-TCO were up to 10 and 4 times higher than those obtained with unsubstituted and substituted linear oligomers, respectively. The rapid onset of transgene expression in both cell lines indicates efficient DNA release and transcription from SB-TCO polyplexes. In comparison with 22 kDa linear PEI-based transfection reagent used as the control, SB-TCO possessed higher gene transfer efficacy, significantly lower cytotoxicity, and improved serum compatibility.

Low Molecular Weight Chitosan (LMWC)-based Polyplexes for pDNA Delivery: From Bench to Bedside

“…One of the main drawbacks related to LMWC-pDNA polyplexes is their poor colloidal stability in physiological conditions [46]. Thus, the presence of polyanions such as blood proteins, heparin or glycosaminoglycans (GAG) in the body may cause the dissociation of the polyplex and the early release of the pDNA before it reaches the desired target tissue [47,48].…”

“…It has been reported that the size of LMWC-pDNA polyplexes increased from 100 nm to more than 1 micron after 1 h of incubation in phosphate buffered saline (PBS) [46]. In addition, bovine serum albumin (BSA)-induced aggregation of polyplexes has been assessed by measuring changes in the turbidity of the solution after their incubation with BSA.…”

“…Moreover, grafting Ch with arginine has been proven to increase cell uptake and PEG-succinimidyl ester or Folate-PEG-COOH molecules could enhance pharmacokinetics [76] Finally, the reductive N-alkylation method serves to graft an aldehyde derivative on the amine of Ch. Grafted species include fluorescent probes as 9-anthraldehyde [77], phosphorylcholine [78,79], oligosaccharides [18,46,52,80] and dextran like polymers [81]. Normally, reductive N-alkylation is directed to reduce cytotoxicity and increase transfection efficiency.…”

Quantitative 3-D colocalization analysis as a tool to study the intracellular trafficking and dissociation of pDNA-chitosan polyplexes

“…19 The superior biocompatibility of chitosan compared with other polycations makes it attractive for biomedical applications. 20,21 The ability of chitosan to condense pDNA and improve the gene transfection efficacy strongly depends on its structural variables. [22][23][24][25] We have synthesized a new generation of self-branched glycosylated (trisaccharide-substituted) chitosan oligomers (SBTCOs) that demonstrate higher gene transfection efficacy than linear chitosan oligomers (LCOs) of the same chain length.…”

Augmentation of adenovirus 5 vector-mediated gene transduction under physiological pH conditions by a chitosan/NaHCO3 solution

“…26,27 However, such complex modification of chitosan can interfere with its chemical and biological properties [27][28][29] and the procedures of preparing chitosan derivatives, for example, self-branched glycosylated chitosan oligomers, are complex and not easily adapted for most laboratories. 30 An alternative to these approaches is to develop a new preparation procedure, which can dissolve chitosan under physiological pH conditions. This strategy can circumvent the complicated chemical modification and synthesis, as well as avoid the interference to its biological activities.…”