Smart Nanoparticles Based on Hyaluronic Acid for Redox-Responsive and CD44 Receptor-Mediated Targeting of Tumor

Park et al. Nanoscale Research Letters (2015) 10:288 DOI 10.1186/s11671-015-0981-5 NANO EXPRESS Open Access Smart Nanoparticles Based on Hyaluronic Acid for Redox-Responsive and CD44 Receptor-Mediated Targeting of Tumor Hyung-Kyu Park1, Sang Joon Lee2, Jong-Suk Oh1, Sam-Gyu Lee3, Young-IL Jeong4 and Hyun Chul Lee1* Abstract Background: Since aggressive cancer cells highly express the CD44 receptor compared to normal cells, hyaluronic acid (HA) can be used for CD44 targeting molecule. Since glutathione (GSH) level is normally elevated in the intracellular compartment and in the tumor cell, the fact that disulfide bond can be cleaved by GSH is widely used for intracellular drug delivery. Methods: HA was connected with poly(DL-lactide-co-glycolide) (PLGA) using disulfide linkage, and then a diblock copolymer (HAssLG) was prepared. Doxorubicin (DOX)-loaded HAssLG nanoparticles were prepared by dialysis procedures. Results and Discussion: DOX-loaded HAssLG nanoparticles have spherical shapes with small particle size of less than 300 nm. In fluorescence measurement, DOX was dose-dependently liberated from nanoparticles by the addition of GSH. DOX release rate from HAssLG nanoparticles was increased by the addition of GSH. To confirm CD44 receptor-mediated endocytosis of nanoparticles, CD44-positive MDA-MB231 cells were employed and fluorescence intensity was strong when nanoparticles were treated to tumor cells. However, fluorescence intensity was significantly decreased through blocking of the CD44 receptor by pretreatment of cells with free HA. Fluorescence intensity of cells was increased again when GSH was added, indicating that HAssLG nanoparticles have CD44 receptor targetability and potential of redox-responsive drug delivery. For animal imaging study, CD44-positive MDA-MB231 cells and CD44-negative NIH3T3 cells were simultaneously implanted into the right flank and left flank of mice, respectively. Fluorescence intensity was significantly stronger at tumor mass of MDA-MB231 cells than solid mass of NIH3T3 cells, indicating that HAssLG nanoparticles were specifically delivered to tumor cells. Conclusions: The results indicated that HAssLG nanoparticles have specificity against the CD44 receptor and can be used for anticancer drug targeting. We recommend HAssLG nanoparticles as a promising vehicle for cancer drug targeting. Keywords: Hyaluronic acid; CD44 receptor-mediated endocytosis; Redox-responsive; Block copolymer; Nanoparticles * Correspondence: 1 Department of Microbiology, Chonnam National University Medical School, 160, Baekseo-ro, Dong-gu, Gwangju 501-746, Republic of Korea Full list of author information is available at the end of the article © 2015 Park et al. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. Park et al. Nanoscale Research Letters (2015) 10:288 Background Nanomedicines based on nanotechnology have been extensively investigated for drug delivery issues, and they can be considered as appropriate vehicles for tumor targeting due to their small sizes [1–4]. Especially, nanosized vehicles such as nanoparticles or polymeric micelles have superior potentials for solid tumor targeting because the enhanced permeation and retention (EPR) effect against tumor blood vessels is regarded as a preferential advantage of nanovehicles for solid tumor targeting [5, 6]. Furthermore, nanoparticles can be decorated with various targeting moieties for specific targeting of cancer cells since they have a big surface area [7–10]. Due to these intrinsic properties, nanoparticles can concentrate cytotoxic anticancer drug to the solid tumor with minimal unwanted side effects for normal tissues/cells. From these points of view, nanoparticles decorated with targeting moieties have been also extensively investigated [7–10]. Aggressive tumor cells frequently overexpress the CD44 receptor in the cell membrane [11–18]. This is a specific receptor for hyaluronic acid (HA), and HA is known to stimulate spreading, migration, invasion, and metastasis of cancer cells [11–14, 17]. From these reasons, hyaluronic acid has been extensively used as a vehicle for specific targeting of the CD44 receptor of tumor cells [8, 18–22]. Especially, Prestwich’s group extensively investigated the potential of CD44 receptor delivery of HA-anticancer agent conjugates against cancer cells [19–22]. Furthermore, HA was also investigated for tissue engineering, gene delivery, and peptide delivery due to the biodegradability and biocompatibility [23–25]. Nanoparticles based on redox-responsive drug targeting for cancer cells also have been extensively investigated in the last decades [26–29]. Redox-responsive drug delivery is based on the fact that disulfide bond can be reduced by glutathione and intracellular glutathione (GSH) level is significantly higher than the extracellular fluid level [30]. This peculiar property between GSH and disulfide bond stimulated the redox-responsive drug delivery concept. For example, Sun et al. synthesized a poly(ethylene glycol)/poly (ε-caprolactone) block copolymer using disulfide bond [27]. They showed that drug release rate was increased by the addition of dithiothreitol (DTT) and enhanced tumor cell uptake of anticancer drugs was observed. Furthermore, redox-responsive delivery of DNA drugs using disulfidelinked polymer nanovehicles was also investigated by several authors [26–29]. In spite of successful development of the redox-responsive drug delivery system, GSH levels in normal tissues and low specificity of nanoparticles are still obstacles for efficient drug targeting. For example, GSH levels can be increased in the tissues of a tumor-bearing mice compared to a normal mice [30]. Therefore, multiple targeting strategies are required to attain efficient targeting of tumors. Page 2 of 10 In this study, we synthesized a HA-b-PLGA (HAssLG) copolymer having disulfide linkage between HA and PLGA for targeting of tumor cells through the CD44 receptor and redox-responsive process. Since PLGA is a biodegradable and lipophilic polymer, PLGA must form the core of the nanoparticles while HA would form the outer shell of the nanoparticles. We studied the potential of CD44 receptor-mediated and redox-responsive targeting of tumor cells by HAssLG nanoparticles using tumor cells and fibroblast cells in vitro and in vivo. Methods Materials HA (molecular weight from the manufacturer’s data, 7460 g/mol) was purchased from Lifecore Biomedical (Chaska, MN, USA). Triethylamine (TEA), L-glutathione (GSH), sodium cyanoborohydride, N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride, and cystamine hydrochloride were purchased from Sigma Chem. Co. (St. Louis, USA). N,N′-dicyclohexyl carbodiimide (DCC) and N-hydroxysuccimide (NHS) were purc (...truncated)