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Glycol Chitosan/Heparin Immobilized Iron Oxide Nanoparticles with a Tumor-Targeting Characteristic for Magnetic Resonance Imaging Soon Hong Yuk,*,† Keun Sang Oh,‡ Sun Hang Cho,§ Beom Suk Lee,|| Sang Yoon Kim,|| Byung-Kook Kwak,^ Kwangmeyung Kim,‡ and Ick Chan Kwon‡ †
College of Pharmacy, Korea University, Jochiwon, Yeongi, Chungnam, 339-700, Republic of Korea Biomedical Research Center, Korea Institute of Science and Technology, 39-1 Hawolgok-dong, Seongbuk-gu, Seoul 136-791, Republic of Korea § Nanobiomaterials Laboratories, Korea Research Institute of Chemical Technology, P.O. Box 107, Yusung, Daejeon 305-600, Republic of Korea Department of Otolaryngology, Asan Medical Center, University of Ulsan, College of Medicine, 388-1 Pungnap-dong, Songpa-gu, Seoul 138-736, Republic of Korea ^ Department of Radiology, Chung-Ang University Hospital, 224-1 Heukseok-dong, Dongjak-gu, Seoul 156-755, Republic of Korea
)
‡
bS Supporting Information ABSTRACT: We described the preparation of the glycol chitosan/ heparin immobilized iron oxide nanoparticles (composite NPs) as a magnetic resonance imaging agent with a tumor-targeting characteristic. The iron oxide nanoseeds used clinically as a magnetic resonance imaging agent were immobilized into the glycol chitosan/ heparin network to form the composite NPs. To induce the ionic interaction between the iron oxide nanoseeds and glycol chitosan, gold was deposited on the surface of iron oxide nanoseeds. After the immobilization of gold-deposited iron oxide NPs into the glycol chitosan network, the NPs were stabilized with heparin based on the ionic interaction between cationic glycol chitosan and anionic heparin. FE-SEM (field emission-scanning electron microscopy) and a particle size analyzer were used to observe the formation of the stabilized composite NPs, and a Jobin-Yvon Ultima-C inductively coupled plasma-atomic emission spectrometer (ICP-AES) was used to measure the contents (%) of formed iron oxide nanoseeds as a function of reaction temperature and formed gold deposited on the iron oxide nanoparticles. We also evaluated the time-dependent excretion profile, in vivo biodistribution, circulation time, and tumor-targeting ability of the composite NPs using a noninvasive NIR fluorescence imaging technology. To observe the MRI contrast characteristic, the composite NPs were injected into the tail veins of tumor-bearing mice to demonstrate their selective tumoral distribution. The MR images were collected with conventional T2-weighted spin echo acquisition parameters.
1. INTRODUCTION Early successes in clinical trials have been made with iron oxide nanoparticles (NPs) as a magnetic resonance imaging (MRI) agent.1 However, the in vivo half-life of iron oxide NPs is usually rather short due to the biofouling in the blood plasma and the formation of aggregates which are selectively captured by the reticuloendothelial system and efficiently cleared out of the bloodstream.2,3 To improve the in vivo stability of the iron oxide NPs as an efficient MRI agent, there have been a number of attempts to achieve an iron oxide NPspolymer composite.4 These polymers include polysaccharice,5,6 poly(ethylene glycol),79 polypeptide,10 and poly(vinylpyrrolidone),11 all of which are known to be biocompatible and to result in a long-circulating iron oxide NPs. Passive targeting of the iron oxide NPspolymer composites to solid tumors is expected based on the enhanced permeability and r 2011 American Chemical Society
retention (EPR) effect.12 However, this EPR effect is not a constant feature of tumor vessels,13 and an attractive alternative would be to target NPs to specific molecular receptors in the blood vessels because they are readily available for binding from the bloodstream and because tumor vessels express a wealth of molecules that are not significantly expressed in the vessels of normal tissues.14 For the specific targeting to tumor cells through the molecular recognition of unique cancer-specific markers, targeting ligands were chemically conjugated to the iron oxide NPspolymer composites.1519 However, the targeting efficiency to tumor cells is generally low. Received: March 26, 2011 Revised: April 19, 2011 Published: April 20, 2011 2335
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Scheme 1. Schematic Diagram for the Preparation of (a) Iron Oxide Nanoseeds, (b) Gold-Deposited Iron Oxide NPs, and (c) Gold-Deposited Iron Oxide/Glycol Chitosan NPs with Heparin (Composite NPs)
Herein, we report the preparation of the glycol chitosan/ heparin immobilized iron oxide NPs (composite NPs) and their characterization as a MRI agent with a tumor-targeting characteristic. Iron oxide nanoseeds were immobilized in the glycol chitosan/heparin network to form the composite NPs. To induce the ionic interaction between the iron oxide nanoseeds and glycol chitosan, gold was deposited on the iron oxide nanoseeds. After the immobilization of gold-deposited iron oxide NPs into the glycol chitosan network, the NPs were stabilized with heparin via the ionic interaction between cationic glycol chitosan and anionic heparin. The iron oxide nanoseeds and glycol chitosan showed a cationic characteristic in the aqueous solution. To induce the interaction between the iron oxide nanoseeds and glycol chitosan in the aqueous media, gold was deposited on the iron oxide nanoseeds to induce the anionic charge. The glycol chitosan is emerging as a novel carrier of drugs because of its solubility and biocompatibility in vivo.20 It also showed targeting characteristics based on the EPR effect.21 The tumors contain a meshwork of clotted plasma proteins (fibrinogenderived product) in the tumor stroma and the walls of vessels, but no such meshwork is detectable in normal tissues.22,23 The interactions between a fibrinogen-derived product in the solid tumor and heparin
can be expected, and this prompted us to use heparin to enhance the targeting of the composite NPs for efficient MR imaging of tumors with the stabilization of composite NPs.
2. EXPERIMENTAL SECTION Materials. The iron(II) chloride tetrahydrate (FeCl2 3 4H2O), iron(III) chloride hexahydrate (FeCl3 3 6H2O), and glycol chitosan (molecular weight: 250000) were purchased from Sigma Co. (U.S.A.). HAuCl4 3 6H2O and trisodium citrate dihydrate were purchased from Aldrich (U.S.A.). Heparin sodium (189 IU mg1, Mw 12500) was purchased from Celsus Laboratories (Cincinnati, OH, U.S.A.). Resovist (40 mol Fe kg1) was purchased from Schering AG, (Germany). Lowmelting agarose was purchased from BioRad (Hercules, CA). Pluronic F-68 (poly(ethylene oxide)poly(propylene oxide)poly(ethylene oxide) triblock copolymer) was obtained as a gift from BASF Corp., Republic of Korea, and used as received. Pluronic F-68 can be represented by the formula (EO)79(PO)26(EO)79 on the basis of its nominal molecular weight of 8400 and 80% PEO. The monoreactive hydroxysuccinimide ester of Cy5.5 (Cy5.5-NHS) was obtained from Amersham Bioscience (Piscataway, NJ). Preparation of the Gold-Deposited Iron Oxide NPs. Golddeposited iron oxide NPs were formed by the reduction of Au3þ on the iron oxide nanoseeds’ surface. First, the iron oxide nanoseeds were 2336
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In Vitro Characterization of the Formed Iron Oxide Nanoseeds, Gold-Deposited Iron Oxide NPs, and Composite NPs.
Figure 1. (a) Content (%) of the formed iron oxide nanoseeds as a function of reaction temperature. (b) Content (%) of the formed gold on iron oxide nanoseeds and (c) zeta potential of the gold-deposited NPs as a function of the added volume of HAuCl4 aqueous solution. prepared by alkaline coprecipitation of ferrous (Fe2þ) and ferric (Fe3þ) chlorides following the procedure described elsewhere.24 To deposit the iron oxide nanoseeds with the gold, 50 mL of aqueous solution containing the prepared iron oxide nanoseeds were refluxed in a threenecked round-bottom flask at 80 C for 30 min. Then, 3 mL of 1 mM tetrachloroauric acid (HAuCl4) and 3 mL of 1 wt % trisodium citrate dihydrate were added stepwise. Vigorous stirring was continued for another 60 min. The solution was cooled to room temperature and was filtered with a 0.2 μm filter.25 Preparation of the Composite NPs. A total of 0.05 g Tween 80 was mixed with 5 mL of aqueous solution containing gold-deposited iron oxide NPs (0.19 mg/mL), in which 0.5 mL of 0.00125 wt % glycol chitosan solution was subsequently added to form gold-deposited iron oxide/glycol chitosan NPs via ionic interaction. To avoid the formation of the agglomerate composed of gold-deposited iron oxide and glycol chitosan, Tween 80 was used as a surfactant. For the stabilization of the gold-deposited iron oxide NPs/glycol chitosan NPs, 0.5 mL of 0.0625 wt % heparin aqueous solution was added to the colloidal solution. Finally, this colloidal solution was freeze-dried in the 5 mL of F-68 (5 wt %) aqueous solution to form lyophilized composite NPs.
Preparation of the Cy5.5-Labeled Composite NPs with Heparin. The composite NPs were labeled with NIR dye cyanine 5.5 (Cy5.5) to visualize their tumor-targeting ability in vivo using a NIR fluorescence imaging system. In brief, 50 mg composite NPs dispersed in 10 mL of distilleddeionized water were modified with 1 mg/mL Cy5.5 dissolved in DMSO overnight. Cy5.5 was immobilized into the composite NPs by ionic interaction. To remove free Cy5.5, the mixed solution was dialyzed (MWCO = 60008000, Spectrum, U.S.A.) for 3 days and lyophilized. The content of Cy5.5 molecules was fixed to 1 wt %, as determined by using a UV/vis spectrophotometer at 680 nm.
The contents (%) of the formed iron oxide nanoseeds as a function of reaction temperature and formed gold deposited on the iron oxide NPs were measured using a Jobin-Yvon Ultima-C inductively coupled plasma-atomic emission spectrometer (ICP-AES). The content of metal (gold and iron oxide) in the composite NPs was also measured using ICP-AES. To evaluate the magnetic property of composite NPs, the magnetization measurements were performed using a MPMS-XL superconducting quantum interference device (SQUID) magnetometer (Quantum Design Inc., San Diego, CA). The particle size distribution, zeta potential, and stability of composite NPs (3 mg/mL of composite NPs dispersed in PBS) were measured using electrophoretic light scattering (ELS-8000, Otsuka Electronics, Japan) at 25 þ 0.1 C. When the difference between the measured and the calculated baselines was less than 0.1%, the correlation function was accepted. A nonlinear regularized inverse Laplacian transformation technique was used to obtain the distribution of the decay constant. The mean diameter was evaluated by the StokesEinstein equation. Also, the FE-SEM pictures were taken to observe the morphology of composite NPs. To prepare a sample for FE-SEM, 0.1 wt % of aqueous solution of the composite NPs was prepared in distilleddeionized water. Each solution was dropped on a carbon mount and then dried at 25 C in a vacuum oven for 24 h. FE-SEM measurement was performed on a JSM-6700F operating at 5 kV. Cellular Cytotoxicity of the Composite NPs. The cytotoxicity of the iron oxide nanoseeds, gold-deposited iron oxide NPs, and golddeposited iron oxide/glycol chitosan NPs with or without heparin was evaluated using MTT assays. Murine SCC-7 (squamous cell carcinoma) cells were cultured in RPMI 1640 (Gibco, Grand Island, NY, U.S.A.) containing 10% (v/v) FBS (Gibco) and 1% (w/v) penicillinstreptomycin at 37 C in a humidified 5% CO295% air atmosphere. The cells were seeded at a density of 5 103 cells/well in 96-well flat-bottomed plates and allowed to adhere overnight. The cells were washed twice with PBS and incubated for 1 day with various concentrations of each samples. The cells were then washed twice with PBS to eliminate the remaining NPs. A total of 25 μL of MTT solution (5 mg/mL in PBS) was added to each well and the cells were incubated further for 2 h at 37 C. The cells were then added to and dissolved in 200 μL of DMSO. Absorbance at 570 nm was measured with a microplate reader (VERSAmax, Molecular Devices Corp., Sunnyvale, CA, U.S.A.).
In Vivo Tumor-Targeting Ability of the Composite NPs in Tumor-Bearing Mice Using a NIR Fluorescence Imaging System. To observe the in vivo tumor-targeting ability of composite NPs with or without heparin in tumor-bearing mice, SCC-7 (squamous cell carcinoma) cells were induced in male C3H/HeN mice (5.5 weeks old, ORIENT BIO Inc., Republic of Korea) by subcutaneous injection of 1.0 106 cells suspended in cell culture media (RPMI 1640, 10% fetal bovine serum, 1% antibiotic agent). When the tumor volume reached approximately 250300 mm3, mice received an IV injection of the Cy5.5/ composite NPs with 10 mg metal NPs/kg. As a control, free Cy5.5 was injected. The tumor- targeting ability was measured by positioning the animal on the eXplore Optix system (Advanced Research Technologies Inc., Montreal, Canada). The laser power and count time settings were optimized at 5 W and 0.3 s per point, respectively. Excitation and emission spots were raster-scanned in 1 mm steps over the selected region of interest to generate emission wavelength scans. A 670 nm pulsed laser diode was used to excite Cy5.5 molecules. The NIR fluorescence emission at 700 nm was collected and detected through a fast photomultiplier tube (Hamamatsu, Japan) and a time-correlated single photon counting system (Becker and Hickl GmbH, Berlin, Germany). All animal experiments were carried out in accordance with the guidelines for animal experiments at the Korea Institute of Science and Technology, Republic of Korea. In Vivo MR Characteristics of the Composite NPs. Prior to in vivo MR imaging, phantom imaging was measured with composite 2337
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Figure 2. (a) Size distribution of the composited NPs in the distilleddeionized water; (b) FE-SEM images of the composite NPs after freeze-drying in the Pluronic F-68 aqueous solution.
Figure 3. Size distribution of the composite NPs in the PBS (pH 7.4, temperature: 36.5 C) for (a) 7 days and (b) 10 days. NPs. Samples for phantom imaging were prepared by suspending weighed amounts of the composite NPs in 50 mL of 1 wt % low-melting agarose. Suspensions were loaded into a prefabricated 24-well agarose sample holder and allowed to solidify at 48 C. In vivo MR images were collected to observe the accumulation of composite NPs at the tumor sites. Accumulation of the composite NPs containing iron oxide at the tumor site shortens the spinspin relaxation time (T2*) by dephasing the spin of neighboring water protons and results in the darkening of T2*-weighted images. SCC-7 cells (ATCC, Rockville, MD, U.S.A.) were cultured in RPMI medium 1640 supplemented with 10% FBS at 37 C in a humidified 5% CO2 incubator. A suspension of 1 106 cells in a physiological saline was inoculated in the subcutaneous right calf of C3H/HeN mice (7 weeks old, 2025 g). When the tumor size was 1 cm in diameter, 0.75 mg of the composite NPs mixed with 0.1 mL of saline was administered through the tail veins of the tumor bearing mice. MR images were acquired at 1, 3, and 6 h post-injection. MR images were acquired using a 4.7 T Bruker Biospin imager (Bruker Medical Systems, Karlsruhe, Germany) with a conventional T2*-weighted gradient (GRE) sequence (repetition time msec/echo time msec, 400/10.4; flip angle, 30). Specific parameters of the images are as follows: spatial resolution, 256 256 matrix; field of view, 30 30 mm; section thickness, 0.7 mm; section gap, 0.7 mm; and number of sections, 12.
Ex Vivo Histological Analysis of the Composite NPs. To validate the targeting specificity, the mice were sacrificed after the postinjection scan at 12 h, and the tumors were sectioned and subjected to a histological study. The tumor tissues were excised and stored in 10 wt % paraformaldehyde. Paraffin-embedded histological slices (10 mm thickness) were stained with hematoxylin-eosin (H&E). To identify the cluster in the histological sections of the tumor, mounting was also demonstrated by using a Prussian blue staining kit and silver enhancer kit (Sigma-Aldrich, U.S.A.). Slides were rinsed with distilleddeionized water and images were obtained on an OLYMPUS BX 51 microscope (OLYMPUS Microsystems, Japan). Statistical Analysis. Data are expressed as the mean ( SEM of at least three experiments. Data for in vivo characteristics using a NIR fluorescence system were calculated by using the region of interest (ROI) function of the Analysis Workstation software (ART Advanced Research Technologies Inc., Montreal, Canada). All data processing was performed using the ORIGIN 7.0 statistical software program (OriginLab Corp., U.S.A.).
3. RESULTS AND DISCUSSION The strategy for the preparation of composite NPs comprised four principal steps: (1) the preparation of gold-deposited iron 2338
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Figure 4. (a) Cytotoxicity of the iron oxide nanoseeds, gold-deposited iron oxide NPs, gold-deposited iron oxide/glycol chitosan NPs, and composite NPs. (b) In vivo noninvasive NIR images and quantification analysis of the Cy5.5/composite NPs with or without heparin. An average normalized counter was obtained from the defined areas.
oxide NPs, (2) the incorporation of gold-deposited iron oxide NPs into the glycol chitosan matrix, (3) the induction of ionic interaction between glycol chitosan/heparin to form composite NPs, and (4) the freeze-drying of composite NPs in a Pluronic F-68 aqueous solution to obtain the composite NPs in the powdery state (Scheme 1). The iron oxide nanoseeds and glycol chitosan show a cationic characteristic in the aqueous solution. To induce the interaction
between the iron oxide NPs and glycol chitosan in the aqueous media, gold was deposited on the surface of iron oxide NPs to induce the anionic charge on the surface. As shown in Figure 1a, the yield of iron oxide nanoseeds was increased with an increase in the reaction temperature and iron oxide nanoseeds formed at 80 C were used throughout the study. The formation of gold on the iron oxide nanoseeds was induced as a function of added volume of 1 mM HAuCl4 aqueous 2339
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Figure 5. (a) Phantom images and quantified MR signal intensity of the composite NPs and Resovist in the same concentration gradient in distilleddeionized water. (b) Magnetization curves of the composite NPs and Resovist.
solution. The aggregation or precipitation of NPs was observed with an increase in the volume of HAuCl4 aqueous solution. Based on these results, gold-deposited NPs formed with 3 mL of HAuCl4 aqueous solution were used throughout the experiment (Figure 1b). Figure 1c shows the change of zeta potential with the deposition of gold on the iron oxide nanoseeds. An anionic charge was developed on the iron oxide nanoseeds with the deposition of gold. Figure 2a shows the size distribution of the composite NPs. The diameter of composite NPs was approximately 120 nm. Freezedrying of the composite NPs was performed in a Pluronic F-68 aqueous solution to obtain the powder form of composite NPs. During the freeze-drying, Pluronic F-68 was adsorbed into the composite NPs to avoid aggregation with improving the stability in the aqueous media. Figure 2b shows the FE-SEM images of the Pluronic F-68-adsorbed composite NPs after freeze-drying. Because of the aggregation of the composite NPs caused by their magnetic property, the size distribution shown in Figure 3 does not reflect the real size of the NPs in the aqueous media. However, we could predict the stability change of the composite NPs in the aqueous media. To observe the stability of the composite NPs in the aqueous media, the size distribution was observed in the PBS as a function of equilibrium time at 36.5 C. Because the composite NPs are prepared by ionic interactions between the components utilized
in the formation of composite NPs and because there are various ionic species in the blood, the ion exchange will occur between composite NPs and ionic species in the blood. This may lead to the rapid disintegration of composite NPs. As shown in Figure 3, we may expect the stability of composite NPs in the blood during MR imaging. Although the composite NPs did not show an aggregation during 5 days of equilibrium in the PBS, the presence of NPs with approximately 50 nm diameter was observed after 7 days, and this trend became significant after 10 days. This indicates that the disintegration of the composite NPs occurred in the PBS with the leakage and aggregation of the gold-deposited iron oxide NPs. After MR imaging, the agent should be cleared from blood. From the experimental result, we can expect the disintegration of composite NPs after a certain period of time. All the samples showed cytotoxicity, which increased as the concentration increased. The iron oxide nanoseeds or golddeposited iron oxide NPs caused a significant reduction in cell viability, even at a low concentration (12.5 μg/mL), and induced further reductions at higher concentrations (below 20 μg/mL), resulting in about 50% loss of cell viability. However, the composite NPs showed lower cytotoxicity than the iron oxide nanoseeds and gold-deposited iron oxide NPs (Figure 4a). The Cy5.5/composite NPs with heparin in mice (10 mg/kg) showed a strong NIR fluorescence signal in the tumor area within 2340
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Figure 6. In vivo MR images of (a) the composite NPs, (b) the composite NPs without heparin, and (c) Resovist.
1 h of the injection, indicating rapid nanoparticles circulation, but the Cy5.5/composite NPs without heparin did not show significant enhancement of the signal in the tumor area (Figure 4b). Although free Cy5.5 used as a control showed a signal in the tumor site, the Cy5.5/composite NPs showed the most enhanced signal and we were able to distinguish tumors from the surrounding background tissue at 1 h postinjection, with a maximum NIR signal at 9 h postinjection and the appearance of a fluorescence signal for more than 48 h at the tumor site. This indicates that the EPR effect causes the composite NPs to accumulate preferentially in tumor tissue. Figure 5 shows phantom images of the composite NPs and Resovist in the same concentration gradient in distilleddeionized
water. It also shows the signal intensity values converted by the image analysis tool for quantitative measurement. The composite NPs showed the enhanced T2* negative images comparied with Resovist (Figure 5a). Also, the composite NPs showed higher magnetic properties compared with Resovist. The increased magnetization of composite NPs may be due to the agglomeration of the magnetic NPs (gold-deposited iron oxide NPs) in the glycol chitosan/heparin composite matrix (Figure 5b). After 1 h post-injection, MR intensity at the tumor site increased (Figure 6a). Up to 6 h after injection, the significant enhancement of MR intensity at the tumor site was observed compared with the 2341
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Figure 7. Histochemical analyses of tumor tissue 12 h after IV injection using (a) H&E staining and Prussian blue staining and (b) silver staining.
composite NPs without heparin and Resovist (Figure 6b,c). This indicates that the composite NPs in this study can be utilized as a MR imaging agent with the enhanced targeting characteristic. It was reported that most solid tumors contain considerable amounts of fibrinogen-derived product (a meshwork of clotted plasma proteins). This suggests that fibrin is important in tumor formation.26 Although inhibition of fibrin formation at a tumor site by heparin is not effective in the treatment of solid tumors,27 the localization of heparin-immobilized NPs at the tumor can be expected based on the interactions between fibrinogen-derived product in the solid tumor and heparin immobilized in the composite NPs. The enhanced targeting of composite NPs can be explained in terms of the aforementioned interactions. Also, MR imaging clearly showed
that composite NPs had extravasated out of tumor blood vessels 12 h after injection and were mostly distributed in the perivascular area. To validate the targeting specificity, the mice were sacrificed after 12 h post-injection and the tumors were sectioned and subjected to histological studies. Figure 7a shows the images from an H&E staining and Prussian blue staining. As shown in Figure 7b, a tumor was sectioned into three parts (inside, line, and outside). The brown color labeled by a silver enhancer kit was observed at one part of the tumor tissue classified as a line, indicating the existence of composite NPs in the tumor tissue after 12 h post-injection. All these results further confirmed the tumor-targeting of the composite NPs after administration. 2342
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4. CONCLUSIONS A novel method for the preparation of the gold-deposited iron oxide/glycol chitosan NPs with heparin (the composite NPs) was reported for efficient MR imaging. FE-SEM measurement and size distribution analysis showed that the composite NPs were formed based on the preparation method in this study. Improved T2* negative images of the composite NPs compared with Resovist were verified by phantom imaging. In vivo MR imaging also suggested the composite NPs exhibited tumorspecific targeting. These results indicate that the composite NPs in this study can be utilized in the design of MR imaging agent with a tumor-targeting characteristic. ’ ASSOCIATED CONTENT
bS
Supporting Information. Experimental results including characteristics and the calculated IC50 of the prepared NPs in this study. This material is available free of charge via the Internet at http://pubs.acs.org.
’ AUTHOR INFORMATION Corresponding Author
*Tel.: þ82-41-860-1612. Fax: þ82-41-860-1606. E-mail: shyuk@ korea.ac.kr.
’ ACKNOWLEDGMENT This work was financially supported by the Ministry of Science and Technology (2010K001253, 2010K001249, and 2010K001254) and grant from the fundamental R&D program for core technology of materials funded by the Ministry of Knowledge Economy, Republic of Korea.
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(15) Nasongkla, N.; Bey, E.; Ren, J.; Ai, H.; Khemtong, C.; Guthi, J. S.; Chin, S.-F.; Sherry, A. D.; Boothman, D. A.; Gao, J. Nano Lett. 2006, 6, 2427–2430. (16) Sun, C.; Sze, R.; Zhang, M. J. Biomed. Mater. Res., Part A 2006, 78, 550–557. (17) DeNardo, S. J.; DeNardo, G. L.; Miers, L. A.; Natarajan, A.; Foreman, A. R.; Gruettner, C.; Adamson, G. N.; Ivkov, R. Clin. Cancer Res. 2005, 11, 7087s–7092s. (18) Akerman, M. E.; Chan, W. C.; Laakkonen, P.; Bhatia, S. N.; Ruoslahti, E. Proc. Natl. Acad. Sci. U.S.A. 2002, 99, 12617–12621. (19) Simberg, D.; Duza, T.; Park, J. H.; Essler, M.; Pilch, J.; Zhang, L.; Derfus, A. M.; Yang, M.; Hoffman, R. M.; Bhatia, S.; Sailor, M. J.; Ruoslahti, E. Proc. Natl. Acad. Sci. U.S.A. 2007, 104, 932–936. (20) Kim, J. H.; Kim, Y. S.; Kim, S.; Park, J. H.; Kim, K.; Choi, K.; Chung, H.; Jeong, S. Y.; Park, R. W.; Kim, I. S.; Kwon, I. C. J. Controlled Release 2006, 111, 228–234. (21) Park, K.; Kim, J. H.; Nam, Y. S.; Lee, S.; Nam, H. Y.; Kim, K.; Park, J. H.; Kim, I. S.; Choi, K.; Kim, S. Y.; Kwon, I. C. J. Controlled Release 2007, 122, 305–314. (22) Abe, K.; Shoji, M.; Chen, J.; Bierhaus, A.; Danave, I.; Micko, C.; Casper, K.; Dillehay, D. L.; Nawroth, P. P.; Rickles, F. R. Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 8663–8668. (23) Pilch, J.; Brown, D. M.; Komatsu, M.; Jarvinen, T. A.; Yang, M.; Peters, D.; Hoffman, R. M.; Ruoslahti, E. Proc. Natl. Acad. Sci. U.S.A. 2006, 10, 2800–2804. (24) Lim, H.; Lee, E.; Kim, D. K.; Jang, N. K.; Jeong, S. Y.; Jon, S. J. Am. Chem. Soc. 2006, 128, 7383–7389. (25) Lin, J.; Zhou, W.; Kumbhar, H. R.; Fang, W. J.; Carpenter, E. E.; O’Corner, C. J. J. Solid State Chem. 2001, 159, 26–31. (26) Costantini, V.; Zacharski, L. R. Thromb. Haemost. 1993, 69, 406–414. (27) Niers, T. M .H.; Klerk, C. P. W.; DiNisio, M.; Van Noorden, C. J. F.; Buller, H. R.; Reitsma, P. H.; Richel, D. J. Crit. Rev. Oncol./ Hematol. 2007, 61, 195–207.
’ REFERENCES (1) Desai, N.; Trieu, V.; Yao, Z.; Louie, L.; Ci, S.; Yang, A.; Tao, C.; De, T.; Beals, B.; Dykes, D. Clin. Cancer Res. 2006, 12, 1317–1324. (2) Raynal, I.; Prigent, P.; Peyramaure, S.; Najid, A.; Rebuzzi, C.; Corot, C. Invest. Radiol. 2004, 39, 56–63. (3) Rogers, W. J.; Basu, P. Atherosclerosis 2005, 178, 67–73. (4) Oh, J. K.; Park, J. M. Prog. Polym. Sci. 2011, 36, 168–189. (5) Berry, C. C.; Wells, S.; Charles, S.; Curtis, A. S. G. Biomaterials 2003, 24, 551–4557. (6) Lemarchand, C.; Gref, R.; Couvreur, P. Eur. J. Pharm. Biopharm. 2004, 58, 327–341. (7) Kohler, N.; Sun, C.; Fichtenholtz, A.; Gunn, J.; Fang, C.; Zhang, M. Small 2006, 2, 785–792. (8) Lutz, J.; Stiller, S.; Hoth, A.; Kaufner, L.; Pison, U.; Cartier, R. Biomacromolecules 2006, 73, 132–3138. (9) Chanana, M.; Jahn, S.; Georgieva, R.; Lutz, J.; Baumler, H.; Wang, D. Chem. Mater. 2009, 21, 1906–1914. (10) Th€unemann, A. F.; Sch€utt, D.; Kaufner, L.; Pison, U.; M€ohwald, H. Langmuir 2006, 22, 2351–2357. (11) Lee, H. Y.; Lim, N. H.; Seo, J. A.; Yuk, S. H.; Kwak, B. K.; Khang, G.; Lee, H. B.; Cho, S. H. J. Biomed. Mater. Res., Part B 2006, 79, 142–150. (12) Maeda, H.; Wu, J.; Sawa, T.; Matsumura, Y.; Hori, K. J. Controlled Release 2000, 65, 271–284. (13) Sinek, J.; Frieboes, H.; Zheng, X.; Cristini, V. Biomed. Microdev. 2004, 6, 297–309. (14) Hoffman, J. A.; Giraudo, E.; Singh, M.; Zhang, L.; Inoue, M.; Porkka, K.; Hanahan, D.; Ruoslahti, E. Cancer Cell 2003, 4, 383–391. 2343
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