Gene delivery of PEI incorporating with functional block copolymer via non-covalent assembly strategy Yuling Hu a , Dezhong Zhou a , Congxin Li a , Hao Zhou b , Jiatong Chen b , Zhengpu Zhang a , Tianying Guo a,⇑ a Key Laboratory of Functional Polymer Materials, Ministry of Education, Institute of Polymer Chemistry, College of Chemistry, Nankai University, Weijin Road, No. 94, Tianjin 300071, China b Department of Biochemistry and Molecular Biology, College of Life Science, Nankai University, Weijin Road, No. 94, Tianjin 300071, China article info Article history: Received 21 May 2012 Received in revised form 19 September 2012 Accepted 25 September 2012 Available online 2 October 2012 Keywords: Poly(ethyleneimine) Functional diblock copolymer Ternary complexes Gene vector abstract A novel functional diblock polymer P(PEGMA-b-MAH) is prepared and incorporated to improve the gene delivery efficiency of poly(ethyleneimine) PEI via non-covalent assembly strategy. First, P(PEG- MA-b-MAH) is prepared from L-methacrylamidohistidine methyl ester (MAH) by reversible addition fragmentation chain transfer polymerization, with poly[poly(ethylene glycol) methyl ether methacrylate] (P(PEGMA)) as the macroinitiator. Then P(PEGMA-b-MAH) is assembled with plasmid DNA (pDNA) and PEI (M w = 10 kDa) to form PEI/P(PEGMA-b-MAH)/pDNA ternary complexes. The agarose gel retardation assay shows that the presence of P(PEGMA-b-MAH) does not interfere with DNA condensation by the PEI. Dynamic light scattering tests show that PEI/P(PEGMA-b-MAH)/pDNA ternary complexes have excel- lent serum stability. In vitro transfection indicates that, compared to the P(PEGMA-b-MAH) free PEI-25k/ pDNA binary complexes, PEI-10k/P(PEGMA-b-MAH)/pDNA ternary complexes have lower cytotoxicity and higher gene transfection efficiency, especially under serum conditions. The ternary complexes pro- posed here can inspire a new strategy for the development of gene and drug delivery vectors. Ó 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved. 1. Introduction Gene delivery systems have attracted a great deal of attention in recent years, and can be generally categorized into viral vectors and nonviral vectors [1–4]. Due to the limitations of viral vectors, such as immunogenicity, oncogenicity, high cost and restrictions in the size of the DNA, nonviral vectors are considered to be pref- erable for gene delivery [5–7]. One of numerous nonviral vectors, poly(ethyleneimine) (PEI), a polycation with a high positive charge density and strong buffering capacity, can protect DNA from nucle- ase degradation and conduce the release of the complex from endosomes via the ‘‘proton sponge effect’’ [8–11]. The gene trans- fection efficiency and cytotoxicity of PEI are heavily correlated with its molecular weight (MW) [12–14]. PEI with high molecular weight (HMW) (e.g. M w = 25 kDa) has high transfection activity; however, its ultrahigh cytotoxicity and aggregation in the blood- stream severely restrict its clinical application [15]. By contrast, PEI with low molecular weight (LMW) is much less cytotoxic, though, due to its low DNA condensation capacity, the gene trans- fection efficiency of LMW PEI is very low [16]. To achieve high gene transfection efficiency as well as low tox- icity, many strategies have been proposed to modify PEI, such as covalent grafting [17], cross linking [18] and electrostatic coating [19]. Poly(ethylene glycol) (PEG) is a biocompatible hydrophilic polymer with good serum protein shielding ability via the ‘‘stealth’’ effect [20–22], and grafting PEI with PEG is one of the most popular strategies to reduce the cytotoxicity and complex aggregation in the bloodstream [23,24]. However, the vast majority of grafting modifications on PEI with PEG are covalent strategies. One of the problems with covalent modifications is that, in coupling the amines with a functional group, the positive charge density of PEI is reduced, and this can affect DNA condensation by the mod- ified PEI; therefore, higher N/P ratios for compact DNA condensa- tion are needed. In order to circumvent the limitations involved in the covalent strategy, a non-covalent strategy is proposed. By this strategy, the cationic property of polycations can be main- tained without change, while the gene delivery performance can be improved by introducing additional functional polymers. Hamada et al. [19,25] devised a DNA/PEI/chondroitin sulfate ter- nary complex and studied its transfection efficiency in ovarian can- cer cells compared with DNA/PEI/hyaluronic acid complex. Zeng et al. [26] combined a positive-targeted peptide with a plasmid DNA (pDNA)/PEI system to form ternary complexes which were capable of mediating gene delivery efficiently and specifically into cells expressing the NGF receptor TrkA. Kurosaki et al. [27] used a polyanion, r-polyglutamic acid, as the coating in preparing the pDNA/PEI/r-polyglutamic acid complex, which showed high 1742-7061/$ - see front matter Ó 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.actbio.2012.09.033 ⇑ Corresponding author. Tel./fax: +86 22 235 015 97. E-mail address: tyguo@nankai.edu.cn (T. Guo). Acta Biomaterialia 9 (2013) 5003–5012 Contents lists available at SciVerse ScienceDirect Acta Biomaterialia journal homepage: www.elsevier.com/locate/actabiomat