Hexanoic acid and polyethylene glycol double grafted amphiphilic chitosan for enhanced gene delivery: influence of hydrophobic and hydrophilic substitution degree.
Gene therapy holds immense potential as a future therapeutic strategy for the treatment of numerous genetic diseases which are incurable to date. Nevertheless, safe and efficient gene delivery remains the most challenging aspects of gene therapy. To overcome this difficulty a series of hexanoic acid (HA) and monomethoxy poly(ethylene glycol) (mPEG) double grafted chitosan-based (HPC) nanomicelles were developed as nonviral gene carrier. HPC polymers with various HA and mPEG substitution degrees were synthesized, and their chemical structures were confirmed by (1)H NMR spectroscopy. HPC nanomicelles exhibited excellent blood compatibility and cell viability, as demonstrated by in vitro hemolysis and MTT assay, respectively. The cationic HPC nanomicelles retained the plasmid DNA (pDNA) binding capacity of chitosan and formed stable HPC/pDNA polyplexes with diameters below 200 nm. Both hydrophobic and hydrophilic substitution resulted in suppressed nonspecific protein adsorption on HPC/pDNA polyplexes and increased pDNA dissociation. However, resistance against DNase I degradation was enhanced by HA conjugation while being inhibited by mPEG substitution. Amphiphilic modification resulted in 3-4.5-fold higher cellular uptake in human embryonic kidney 293 cells (HEK 293) mainly through clathrin-mediated pathway. The optimal HPC/pDNA polyplexes displayed 50-fold and 1.2-fold higher gene transfection compared to unmodified chitosan and Fugene, respectively, in HEK 293 cells. Moreover, both the cellular uptake and in vitro transfection study suggested a clear dependence of gene expression on the extent of HA and mPEG substitution. These findings demonstrate that amphiphilic HPC nanomicelles with the proper combination of HA and mPEG substitution could be used as a promising gene carrier for efficient gene therapy.
Department of Pharmaceutical Sciences, College of Pharmacy, Nursing, and Allied Sciences, North Dakota State University , Fargo, North Dakota 58105, United States.
Buddhadev
B
Layek
Manas K
MK
Haldar
Gitanjali
G
Sharma
Lindsey
L
Lipp
Sanku
S
Mallik
Jagdish
J
Singh
D000818:Animals; D002208:Caproates / Q000737:chemistry* / Q000378:metabolism; D048271:Chitosan / Q000737:chemistry* / Q000378:metabolism; D004247:DNA / Q000008:administration & dosage; D024202:Electrophoretic Mobility Shift Assay; D004912:Erythrocytes / Q000378:metabolism; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000378:metabolism; D057809:HEK293 Cells; D006367:HeLa Cells; D006461:Hemolysis / Q000235:genetics; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D009682:Magnetic Resonance Spectroscopy; D053758:Nanoparticles / Q000737:chemistry* / Q000378:metabolism; D010957:Plasmids / Q000008:administration & dosage*; D011092:Polyethylene Glycols / Q000737:chemistry* / Q000378:metabolism; D011108:Polymers / Q000737:chemistry* / Q000378:metabolism; D051381:Rats; D017550:Spectroscopy, Fourier Transform Infrared
D002208:Caproates; D011108:Polymers; D049452:Green Fluorescent Proteins; C037652:hexanoic acid; D011092:Polyethylene Glycols; C028210:monomethoxypolyethylene glycol; D004247:DNA; D048271:Chitosan
chitosan; gene delivery; hexanoic acid; poly(ethylene glycol); transfection