Biodegradable poly(ester amine) based on glycerol dimethacrylate and polyethylenimine as a gene carrier.
BACKGROUND
Polyethylenimine (PEI) vectors are widely used in gene delivery because of their high transfection efficiency owing to a unique proton sponge effect. An increase in molecular weight increases transfection efficiency, but simultaneously results in increased toxicity. Therefore, the design and synthesis of new degradable gene delivery carriers having high transfection efficiencies and reduced cytotoxicity are necessary.


METHODS
In the present study degradable poly(ester amines) (PEAs) based on glycerol dimethacrylate (GDM) and low molecular weight branched polyethylenimine (LMW-PEI) were synthesized in anhydrous methanol at 60 degrees C following Michael addition reaction. The transfection efficiencies of the synthesized PEA/DNA complexes were evaluated using three different cell lines (HeLa, HepG2 and 293T cells) in vitro.


RESULTS
PEAs with zeta potential in the range of 30-55 mV (at physiological pH) condensed plasmid DNA into nanosized particles (<150 nm) suitable for intracellular delivery. The PEAs degraded in a controlled fashion (t(1/2) of approximately 9-10 days). Compared with PEI 25K, the PEAs showed significantly lower cytotoxicity in three different cells. The PEAs demonstrated much higher transfection efficiency compared to conventional PEI 25K and Lipofectamine. The PEA synthesized using a 1 : 4 mole ratio of GDM to PEI [GDM/PEI-1.2 (1:4)] showed the highest transfection efficiency in HepG2 cells. Significantly higher pEGFP-N(2) reporter gene expression in 293T cells was achieved using these PEAs. The hyperosmotic effect of PEAs was demonstrated by the reduction in packed cell volume (PCV). The GDM/PEI-1.2 (1:4) showed comparable reduction in PCV with respect to glycerol in 293T cells. The effect of bafilomycin A(1) on transfection efficiency of PEAs on 293T cells indicated its endosomal buffering capacity.


CONCLUSIONS
We hypothesized that the higher transfection efficiency of PEAs was the synergistic effect arising from hyperosmotic glycerol and endosomal buffering capacity of PEAs resulting from the presence of a glycerol backbone and PEI amine groups, respectively.
Arote
Rohidas B
RB
Department of Agricultural Biotechnology, Seoul National University, Seoul, South Korea.
Hwang
Soon-Kyung
SK
Yoo
Mi-Kyong
MK
Jere
Dhananjay
D
Jiang
Hu-Lin
HL
Kim
You-Kyoung
YK
Choi
Yun-Jai
YJ
Nah
Jae-Woon
JW
Cho
Myung-Haing
MH
Cho
Chong-Su
CS
D001672:Biocompatible Materials / Q000737:chemistry / Q000378:metabolism; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004337:Drug Carriers / Q000737:chemistry / Q000378:metabolism; D018014:Gene Transfer Techniques; D005990:Glycerol / Q000737:chemistry*; D006367:HeLa Cells; D006801:Humans; D018942:Macrolides / Q000494:pharmacology; D008689:Methacrylates / Q000737:chemistry*; D018613:Microscopy, Confocal; D008970:Molecular Weight; D011073:Polyamines / Q000737:chemistry / Q000378:metabolism; D011091:Polyesters / Q000737:chemistry / Q000378:metabolism; D011094:Polyethyleneimine / Q000737:chemistry*; D014162:Transfection
D001672:Biocompatible Materials; D004337:Drug Carriers; D018942:Macrolides; D008689:Methacrylates; D011073:Polyamines; D011091:Polyesters; C516446:poly(caprolactone-block-ethyleneimine); C040929:bafilomycin A1; D011094:Polyethyleneimine; D004247:DNA; D005990:Glycerol