O-28: Less Toxic, More Efficient Nanostructured Complexes Based on High Molecular Weight PEI for Cancer Gene Therapy


Hoseini-Alhashemi S *, Dehshahri A , Oskuee R ,

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Objective: Polyethyleneimine (PEI) is a cationic macromolecule and is able to condense DNA into toroidal and globular nanostructures, which are then amenable to cellular internalization via endocytosis. . In the present investigation synthesis of alkylcarboxylate derivatives of PEI were followed by attachment of a hy<font><font>drop</font></font>hilic targeting ligand, lactose bearing galactose group. These modifications were hypothesized to improve plasmid DNA delivery to HepG2 cell line which contains asialoglycoprotein receptors and reduce the cytotoxicity of the nanoplexes with appropriate specifications such as particle size, zeta potential, gene delivery efficiency and DNA binding affinity. Materials and Methods: First, PEI was reacted with a series of ω-bromoalkylcarboxylic acids with 2, 6 and 10 carbon chain lengths. The reaction was allowed to proceed for 24 h at room temperature followed by dialysis against water to remove the unreacted materials. By these modifications, negative charges were introduced into the polymer backbone and the polymer hy<font><font>drop</font></font>hobic-hy<font><font>drop</font></font>hilic balance was improved. In the second step, simple galactosyl residues coupled to hy<font><font>drop</font></font>hobic polyethylenimine by reductive amination reaction. The products were purified by dialysis against water. The binding strength of the modified PEIs to plasmid DNA and the size of the nanoparticles were measured with ethidium bromide (EtBr) exclusion assay and dynamic light scattering studies, respectively. Gene transfer ability and cytotoxicity of the modified PEI were evaluated using Neuro2A and HepG2 cells. Results: Ethidium bromide dye exclusion was used to show the DNA binding ability of the polymers and the cytotoxicity of the free polymers and polymer/plasmid nanoparticles were evaluated in different cell lines such as N2a and HepG2. The results showed the ability of the modified polymer to condense plasmid DNA and form nanoparticles in the range of 70-150 nm. The modifications reduced cytotoxicity and demonstrated that cell viability was strongly dependent on the degree of substitution of primary amines on PEI. Also galactose conjugation significantly increased the ability of nanostructured complexes to transfer plasmid DNA into hepatic cells Conclusion: The most probable reasons for the improved transfection efficiency are a more favorable hy<font><font>drop</font></font>hobic-hy<font><font>drop</font></font>hilic balance in addition to targeting through galactose. A hy<font><font>drop</font></font>hobic environment may also contribute to endosomal release by making the endosomal membrane fragile as a result of acting as a sink for lipids spontaneously released from the endosomal membrane. In conclusion, the hy<font><font>drop</font></font>hobic galactosylated PEI obtained by this safe and inexpensive method can be considered as good nanocarriers both for in vitro and in vivo gene therapy