244 The Polymer Society of Korea Macromolecular Research, Vol. 20, No. 3, pp 244-249 (2012) www.springer.com/13233 pISSN 1598-5032 eISSN 2092-7673 Enhanced Gene Delivery by Palmitic Acid-Conjugated Low Molecular Weight Polyethylenimine Duhwan Lee 1 , Kaushik Singha 1 , Juhee Park 1 , Seongbong Jo 2 , and Won Jong Kim * ,1 1 Department of Chemistry, BK21 Program, Polymer Research Institute, Pohang University of Science and Technology, Gyeongbuk 790-784, Korea 2 Department of Pharmaceutics, School of Pharmacy, The University of Mississippi, MS 38677, USA Received August 29, 2011; Revised October 17, 2011; Accepted October 20, 2011 Abstract: Palmitic acid-conjugated low molecular weight polyethylenimine (PEI-g-PEG-PA) was successfully synthesized to develop an efficient non-viral gene carrier. The judicious integration of hydrophobic palmitic acid and polyethylenimine via hydrophilic polyethylene glycol (PEG) facilitated the formation of nano-sized complex (nanoplex) with plasmid DNA (pDNA) which provided the protection of pDNA against the serumic enzymatic degradation. Furthermore, the delivery system demonstrated enhanced gene transfection efficiency in comparison to unmodified low molecular weight PEI without inducing any significant cytotoxicity. Keywords: gene delivery, polyethylenimine, nanoplex, non-viral, palmitic acid. Introduction Viral gene carriers such as retroviruses, adenoviruses, and adeno-associated viruses have been known to have high transfection efficiency as compared to non-viral vectors. However, their applicability in clinical practice suffers a great setback due to several impediments associated with them. The demerits including their restricted targeting ability only to dividing cells, random DNA insertion, low capacity for carrying large therapeutic genes in size, risk of replica- tion, and possible host immune reaction impaired their potential as gene delivery tool. 1,2 Therefore, non-viral gene delivery systems such as cationic polymers or lipids have been investigated intensively to overcome the disadvan- tages associated with viral vectors. Among the non-viral gene carriers, polyethylenimine (PEI) is one of the most potent carriers and has been extensively studied in vitro and in vivo since their initial use as gene carrier. 3 Transfection efficiency of PEI, along with its cytotoxicity, is heavily dependent on its molecular weights. 4-6 PEI with a molecular weight higher than 25 kDa exhibits high transfection efficiency and cytotoxicity, whereas low molecular weight PEI (<1.8 kDa) despite its low toxicity profile displays low transfection. The toxicity of PEI primarily arises from the strong interaction potential of positive charged PEI with negatively charged cell surfaces and associated membrane damage. 7 Several strategies have been adopted so far to address the toxicity concern. Polyethylene glycol (PEG) was conjugated to PEI as a hydrophilic segment and its efficacy has been investi- gated by several groups. 8,9 The non-viral gene delivery copoly- mers based on the high molecular weight PEI showed considerable transfection efficiency, however toxicity remained as a problem. In previous reports, biodegradable PEI derived from the low molecular weight PEI and PEG exhib- ited improved transfection efficiency and low cytotoxic- ity. 10-12 Also, water soluble lipopolymer (WSLP) obtained by integrating the cationic headgroup of branched PEI (BPEI, MW=1.8 kDa) with a hydrophobic lipid anchor, cholesterol chloroformate, showed low cytotoxicity, and enhanced trans- fection efficacy in vitro and in vivo was obtained. 13-15 These amphiphilic PEI has also been reported by other research groups. 16,17 The high molecular weight PEI (MW=25 kDa) conjugated with pendant palmitic acids chains improved the biocompatibility markedly when compared to the starting material, but also reduced transfection efficiency. 18 We hypothesized that a similar strategy with low molecular weight PEI, but different structure may generate the differ- ent complex formulation with DNA, thus improve transfec- tion efficiency and biocompatibility. In this study, we successfully synthesized palmitic acid- conjugated PEI (PEI-g-PEG-PA) and studied their physico- chemical characteristics. Furthermore, the potential of PEI- g-PEG-PA as a novel gene carrier was evaluated through luciferase gene transfection assay. Experimental Materials. Glycerol ethoxylate (Glyceryl PEG, M n =1,000 DOI 10.1007/s13233-012-0050-1 *Corresponding Author. E-mail: wjkim@postech.ac.kr