International Journal of Pharmaceutics 376 (2009) 141–152 Contents lists available at ScienceDirect International Journal of Pharmaceutics journal homepage: www.elsevier.com/locate/ijpharm Pharmaceutical Nanotechnology Hyperbranched polysiloxysilane nanoparticles: Surface charge control of nonviral gene delivery vectors and nanoprobes Won Jin Kim a,b , Adela C. Bonoiu a , Teruaki Hayakawa c , Cheng Xia c , Masa-aki Kakimoto c , Haridas E. Pudavar a , Kwang-Sup Lee a,b,∗ , Paras N. Prasad a,∗∗ a Institute for Lasers, Photonics, and Biophotonics, University at Buffalo, The State University of New York, Buffalo, NY 14260-3000, USA b Department of Advanced Materials, Hannam University, Daejeon 305-811, Republic of Korea c Department of Organic and Polymeric Materials, Tokyo Institute of Technology, Tokyo 152-8552, Japan article info Article history: Received 14 November 2008 Received in revised form 1 April 2009 Accepted 20 April 2009 Available online 3 May 2009 Keywords: Polymer nanoparticle Surface charge controllable nanoparticle Nonviral gene delivery vector Nanoprobe Hyperbranched polysiloxysilane Multi-photon cellular imaging abstract New hyperbranched polysiloxysilane (HBPS) materials containing terminal carboxylic acid and qua- ternary ammonium groups were designed and synthesized to obtain fluorescent-dye-encapsulated nanoparticles. These polymers exhibited desirable characteristics, including amphiphilicity for nanopar- ticle formation, and contained various terminal groups for surface-charge control on the nanoparticles or for further bioconjugation for targeted imaging. Nanoprobes composed of polysiloxysilane nanoparticles encapsulating two-photon dyes were also prepared for optical bioimaging with controlled surface charge density (zeta potential) for modulation of cellular uptake. Intracellular delivery of these structurally sim- ilar polysiloxysilane nanoparticles, with substantially different surface charges, was investigated using confocal and two-photon fluorescence microscopy as well as flow cytometry. Finally, the use of these nanoparticles as efficient gene delivery vectors was demonstrated by means of in vitro transfection study using -galactosidase plasmid and pEGFP-N1 plasmid and the most efficient combination was obtained using HBPS-CN30:70. © 2009 Elsevier B.V. All rights reserved. 1. Introduction The use of nanoparticles as diagnostic probes and effective car- rier vehicles in targeted therapy is gaining considerable interest. Recently, various organic, inorganic, and hybrid nanoparticle sys- tems have been used for the nanocarriers and gene delivery vectors (Prasad, 2003). There have been several recent papers that have demonstrated the capability of tailored synthetic polymers such as dendrimers, block copolymers, hyperbranched polymers, and star polymers (Ramzi et al., 1997; Grohn et al., 2001; Tian et al., 2005; Pistel et al., 1999), consequently, to form nanoparticle as a nanocarrier. These polymeric approaches show enhanced ther- apeutic efficacy. Particularly, polymeric nanocarriers/vectors are typically prepared in the form of micellar nanoparticle because the molecular self-assembling architectures lead to the formation of the stable nanostructures which has two main components such as hydrophobic core and hydrophilic corona. The hydrophilic corona surrounding the core is known to play an important role in the ∗ Corresponding author at: Hannam University, Daejeon 305-811, Republic of Korea. Tel.: +82 42 629 8857; fax: +82 420629 8854. ∗∗ Corresponding author at: University at Buffalo, SUNY, Buffalo, NY 14260-3000, USA. Tel.: +1 716 645 6800x2098; fax: +1 716 645 6945. E-mail addresses: kslee@hnu.kr (K.-S. Lee), pnprasad@buffalo.edu (P.N. Prasad). cellular uptake, and hydrophobic core of micelles can act as the reservoir of drugs. It can be used in biological applications such as drug delivery vehicles, targeted cellular imaging probes, and gene delivery vectors (Benns et al., 2000; Midoux and Monsigny, 1999; Liu and Yao, 2002; Thanou et al., 2002; Wetering et al., 1999; Fischer et al., 1999; Marschall et al., 1999; Campeau et al., 2001). However, most of these nanocarrier systems exhibit only limited capability of controlling the surface charge of the nanoparticles to optimize cellular uptake and gene delivery efficiency. Further- more, many of these carriers are often cytotoxic thus eliminating them from in vivo applications. To address some of these issues, we synthesized hyperbranched polysiloxysilane (HBPS) polymers containing amphiphilic components with different end groups. While the amphiphilicity of these polymers helps in the formation of aqueous nanoparticle dispersions, the combination of different functional end groups makes it possible to tailor the surface-charge distribution of the formed nanoparticles. The novelty of these amphiphilic HBPS polymers lie in the fact that nanoparticles with different zeta () potential (surface charge density) can be easily tailored and functionalized. Indeed, the resulting hydrophobic cores found within the nanoparticles can be loaded with hydrophobic dye molecules or other hydrophobic drugs, which can be used as fluorescent probes in gene delivery or in drug delivery applications (Kim et al., 2002a,b, Midoux and Monsigny, 1999). Furthermore, HBPS polymers are composed of 0378-5173/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.ijpharm.2009.04.023