SURFACE FUNCTIONALIZATION OF SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLES FOR POTENTIAL CELL TARGETING, IMAGING, AND CANCER THERAPY APPLICATIONS           HUANG CHAO                     NATIONAL UNIVERSITY OF SINGAPORE 2012         SURFACE FUNCTIONALIZATION OF SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLES FOR POTENTIAL CELL TARGETING, IMAGING, AND CANCER THERAPY APPLICATIONS HUANG CHAO (B.ENG., TIANJIN UNIVERSITY) A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF PHILOSOPHY DEPARTMENT OF CHEMICAL AND BIOMOLECULAR ENGINEERING NATIONAL UNIVERSITY OF SINGAPORE 2012               ACKNOWLEDGEMENTS My sincere and deep gratitude goes first and foremost to my supervisor, Professor Neoh Koon Gee, for her inspired guidance, valuable suggestions, insightful criticism, great patience, and constant encouragement and support throughout the entire period of my research study. Her enthusiasm, rigorous attitude and dedication to scientific research are strongly impressed on my memory. Her expert advices greatly help me improve the depth of my research. The profound and invaluable knowledge that I gained from her will benefit me in my future life and career. I am also very grateful to Professor Kang En-Tang for his kindly permission to access the equipments in his research lab. Further sincere thanks go to all my friends and colleagues for their assistance and support. In particular, big thanks go to Dr. Wang Liang for sharing with me his invaluable experience in the research field. I also owe a debt of gratitude to the lab officers and technical staff of Department of Chemical and Biomolecular Engineering, especially Dr. Yuan Zeliang, Mr. Chia Phai Ann, Dr. Yang Liming, Ms Xu Yanfang and Mr. Chan Chuin Mun for their kindly help during my research. The research scholarship for Ph.D study provided by National University of Singapore is also greatly appreciated. Finally, I would like to express my deepest gratitude to my beloved parents, my husband, and other relatives for their unconditional love and support.     I   TABLE OF CONTENTS ACKNOWLEDGEMENTS I TABLE OF CONTENTS . II SUMMARY VII LIST OF ABBREVIATIONS . VIII LIST OF FIGURES X LIST OF TABLES . XV CHAPTER INTRODUCTION 1.1 Background 1.2 Research Objectives and Scopes . CHAPTER LITERATURE REVIEW 2.1 SPIONs . 2.1.1 Basic Properties of SPIONs . 2.1.2 Synthesis of SPIONs 2.1.2.1 Co-precipitation . 2.1.2.2 Thermal Decomposition . 10 2.1.3 Challenges of SPIONs for Biomedical Applications . 11 2.2 Surface Functionalization of SPIONs 14 2.2.1 Materials for Surface Modification of SPIONs 15 2.2.1.1 Monomer Stabilizers 15 2.2.1.2 Polymer Stabilizers 17 2.2.1.3 Inorganic Stabilizers 18 2.2.2 Methods for Surface Modification of SPIONs . 20 2.2.2.1 Self-assembly . 20 2.2.2.2 Surface-initiated Controlled Polymerization . 22 II   2.3 Biomedical Applications of SPIONs 24 2.3.1 MRI 24 2.3.2 Drug/Gene Delivery . 28 2.3.3 Hyperthermia 31 CHAPTER SURFACE FUNCTIONALIZATION OF SUPERPARAMAGNETIC NANOPARTICLES FOR MODULATION OF MACROPHAGE UPTAKE 33 3.1 Introduction 34 3.2 Materials and Methods . 36 3.2.1 Materials . 36 3.2.2 Preparation of SPIONs . 36 3.2.3 Synthesis of PLMA Copolymers 37 3.2.4 Synthesis of PLMA-PEG Copolymers . 37 3.2.5 Preparation of PLMA-SPIONs and PLMA-PEG-SPIONs 38 3.2.6 In Vitro Quantification of Nanoparticles Uptake by Macrophages 38 3.2.7 Cytotoxicity Assay of Nanoparticles 40 3.2.8 MRI Experiments . 40 3.2.9 Characterization . 42 3.3 Results and Discussion . 45 3.3.1 Characterization of PLMA . 45 3.3.2 Characterization of PLMA-PEG 46 3.3.3 Characterization of PLMA-SPIONs . 48 3.3.4 Characterization of PLMA-PEG-SPIONs 52 3.3.5 Uptake of PLMA-SPIONs by Macrophages 55 3.3.6 Uptake of PLMA-PEG-SPIONs by Macrophages . 59 3.3.7 Cytotoxicity of Nanoparticles 61 3.3.8 Magnetic Properties of Nanoparticles and MR Relaxometry 62 III   3.4 Conclusion 70 CHAPTER SURFACE MODIFIED SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLES FOR HIGH EFFICIENCY FOLATE-RECEPTOR TARGETING WITH LOW UPTAKE BY MACROPHAGES . 71 4.1 Introduction 72 4.2 Materials and Methods . 74 4.2.1 Materials . 74 4.2.2 Preparation of SPIONs . 74 4.2.3 Synthesis of Initiator 75 4.2.4 Synthesis of Initiator Coated SPIONs 76 4.2.5 Surface Initiated ATRP on SPIONs . 76 4.2.6 Chemical Modification of Epoxy Groups with EDA . 77 4.2.7 Folic Acid Conjugation 77 4.2.8 Cell Culture 78 4.2.9 In Vitro Evaluation of Uptake of Nanoparticles . 78 4.2.10 Cytotoxicity Assay . 79 4.2.11 MRI Experiments . 79 4.2.12 Characterization . 79 4.2.13 Statistical Analysis . 80 4.3 Results and Discussion . 81 4.3.1 Synthesis of SPIONs-PGMA-FA . 81 4.3.2 Size and Zeta Potential of Nanoparticles . 85 4.3.3 Magnetic Properties 87 4.3.4 Cellular Uptake of Nanoparticles . 90 4.3.5 Cytotoxicity Assay . 94 4.4 Conclusion 96 IV   CHAPTER COMBINED ATRP AND ‘CLICK’ CHEMISTRY FOR DESIGNING LONG-CIRCULATING TUMOR-TARGETING SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLES 97 5.1 Introduction 98 5.2 Materials and Methods . 100 5.2.1 Materials . 100 5.2.2 Preparation of SPIONs . 100 5.2.3 Preparation of Initiator-coated SPIONs. 100 5.2.4 Preparation of SPIONs-P(GMA-co-PEGMA) via ATRP 100 5.2.5 Preparation of SPIONs-P(GMA-co-PEGMA)-N3 . 102 5.2.6 Preparation of Alkyne-functionalized FA 102 5.2.7 Preparation of SPIONs-P(GMA-co-PEGMA)-FA . 103 5.2.8 Cell Culture 104 5.2.9 Cytotoxicity Assay . 104 5.2.10 In Vitro Evaluation of Folate Receptor Targeting 104 5.2.11 Characterization . 105 5.2.12 Statistical Analysis . 106 5.3 Results and Discussion . 107 5.3.1 Surface Characterization of SPIONs-P(GMA-co-PEGMA)-FA . 107 5.3.2 Nanoparticle Size and Stability 113 5.3.3 Magnetic Property 116 5.3.4 Cytotoxicity Assay . 117 5.3.5 In Vitro Cellular Uptake . 118 5.4 Conclusion 122 CHAPTER CISPLATIN-CONJUGATED MAGNETIC NANOPARTICLES FOR POTENTIAL BLADDER CANCER THERAPY . 123 6.1 Introduction 124 6.2 Materials and Methods . 127 V   6.2.1 Materials . 127 6.2.2 Preparation of SPIONs . 127 6.2.3 Synthesis of PCL 127 6.2.4 Synthesis of PCL-b-P(PMA-co-PEGMA) . 128 6.2.5 Synthesis of PCL-b-P(PMA-click-MSA-co-PEGMA) 128 6.2.6 Preparation of SPIONs-loaded PNs . 129 6.2.7 Preparation of Cisplatin-conjugated PNs (Pt-Fe-PNs) . 129 6.2.8 In Vitro Cisplatin Release 130 6.2.9 Cell Culture 131 6.2.10 Cytotoxicity Evaluation 131 6.2.11 Cellular Uptake 132 6.2.12 Characterization . 133 6.3 Results and Discussion . 134 6.3.1 Synthesis of PCL-b-P(PMA-click-MSA-co-PEGMA) 135 6.3.2 Preparation of Fe-PNs and Pt-Fe-PNs 139 6.3.3 In Vitro Drug Release . 142 6.3.4 In Vitro Cytotoxicity Evaluation 144 6.4 Conclusion 148 CHAPTER CONCLUSIONS AND RECOMMENDATIONS FOR FUTURE WORK 149 7.1 Conclusions 150 7.2 Recommendations for Future Work . 153 REFERENCES . 155 LIST OF PUBLICATIONS ARISING FROM PHD WORK . 173     VI   SUMMARY Superparamagnetic iron oxide nanoparticles (SPIONs) are very useful for biomedical applications, such as magnetic resonance imaging (MRI), hyperthermia for cancer therapy, cell targeting, drugs or gene delivery. However, once introduced into blood, SPIONs will be captured by the macrophages and then rapidly cleared out from circulation which can drastically reduce the efficiency of SPIONs-based diagnosis and therapy. Therefore, the bio-interfaces of SPIONs are crucial for their biomedical applications. The overall aim of this thesis is to modify SPIONs with different polymers for potential cell targeting, MRI and cancer therapy applications. In the first project, SPIONs were coated with either poly(DL-lactic acid-co-malic acid) (PLMA) or poly(ethylene glycol)-conjugated PLMA (PLMA-PEG) to modulate uptake by macrophages. PLMA-SPIONs are readily taken up by macrophages but the extent of uptake can be reduced by increasing the PEG content of the PLMA-PEG coating. In the second project, SPIONs were coated with either poly(glycidyl methacrylate) or poly(glycidyl methacrylate-co-poly(ethylene glycol) methyl ether methacrylate) combined with a targeting ligand, folic acid, to attain high selectivity in targeting cancers with minimal uptake by macrophages. All these nanoparticles have low cytotoxicity and exhibit higher MRI contrast effects than commercial agents. Finally, SPIONs-loaded, cisplatin-conjugated polymeric nanoparticles were synthesized for potential application against bladder cancer. These nanoparticles show mucoadhesiveness, a sustained release of cisplatin over days and can effectively induce cytotoxicity against the bladder cancer cells.    VII   References Dresco, P. A., V. S. Zaitsev, R. J. Gambino and B. Chu. Preparation and properties of magnetite and polymer magnetite nanoparticles, Langmuir, 15, pp.1945-1951. 1999. Dube, D., M. Francis, J. C. Leroux and F. M. Winnik. Preparation and tumor cell uptake of poly(N-isopropylacrylamide) folate conjugates, Bioconjugate Chem., 13, pp.685-692. 2002. Emerit, J., C. Beaumont and F. Trivin. Iron metabolism, free radicals, and oxidative injury, Biomed. Pharmacother., 55, pp.333-339. 2001. Eroglu, M., S. Irmak, A. Acar and E. B. Denkbas. Design and evaluation of a mucoadhesive therapeutic agent delivery system for postoperative chemotherapy in superficial bladder cancer, Int. J. Pharm., 235, pp.51-59. 2002. Fan, Q. L., K. G. Neoh, E. T. Kang, B. Shuter and S. C. Wang. Solvent-free atom transfer radical polymerization for the preparation of poly(poly(ethyleneglycol) monomethacrylate)-grafted Fe3O4 nanoparticles: Synthesis, characterization and cellular uptake, Biomaterials, 28, pp.5426-5436. 2007. Fortin, J. P., C. Wilhelm, J. Servais, C. Menager, J. C. Bacri and F. Gazeau. Sizesorted anionic iron oxide nanomagnets as colloidal mediators for magnetic hyperthermia, J. Am. Chem. Soc., 129, pp.2628-2635. 2007. Frey, N. A., S. Peng, K. Cheng and S. H. Sun. Magnetic nanoparticles: Synthesis, functionalization, and applications in bioimaging and magnetic energy storage, Chem. Soc. Rev., 38, pp.2532-2542. 2009. Gao, J. M., H. Ai, C. Flask, B. Weinberg, X. Shuai, M. D. Pagel, D. Farrell and J. Duerk. Magnetite-loaded polymeric micelles as ultrasensitive magnetic-resonance probes, Adv. Mater., 17, pp.1949-1952. 2005. Gao, M. Y., F. Q. Hu, L. Wei, Z. Zhou, Y. L. Ran and Z. Li. Preparation of biocompatible magnetite nanocrystals for in vivo magnetic resonance detection of cancer, Adv. Mater., 18, pp.2553-2556. 2006. Gendelman, H. E., H. Dou, C. J. Destache, J. R. Morehead, R. L. Mosley, M. D. Boska, J. Kingsley, S. Gorantla, L. Poluektova, J. A. Nelson, M. Chaubal, J. Werling, J. Kipp and B. E. Rabinow. Development of a macrophage-based nanoparticle platform for antiretroviral drug delivery, Blood, 108, pp.2827-2835. 2006. Gilchrist, R. K., R. Medal, W. D. Shorey, R. C. Hanselman, J. C. Parrott and C. B. Taylor. Selective inductive heating of lymph nodes, Ann. Surg., 146, pp.596-606. 1957. 158 References Gillis, P., F. Moiny and R. A. Brooks. On T2-shortening by strongly magnetized spheres: A partial refocusing model, Magn. Reson. Med., 47, pp.257-263. 2002. Goya, G. F., T. S. Berquo, F. C. Fonseca and M. P. Morales. Static and dynamic magnetic properties of spherical magnetite nanoparticles, J. Appl. Phys., 94, pp.35203528. 2003. Griffith, D. P., D. M. Musher and C. Itin. Urease - primary cause of infection-induced urinary stones, Invest. Urol., 13, pp.346-350. 1976. GuhaSarkar, S. and R. Banerjee. Intravesical drug delivery: Challenges, current status, opportunities and novel strategies, J. Control. Release, 148, pp.147-159. 2010. Gupta, A. K. and M. Gupta. Synthesis and surface engineering of iron oxide nanoparticles for biomedical applications, Biomaterials, 26, pp.3995-4021. 2005. Haam, S., J. Yang, J. Lee, J. Kang, K. Lee, J. S. Suh, H. G. Yoon and Y. M. Huh. Hollow silica nanocontainers as drug delivery vehicles, Langmuir, 24, pp.3417-3421. 2008. Hao, R., R. J. Xing, Z. C. Xu, Y. L. Hou, S. Gao and S. H. Sun. Synthesis, functionalization, and biomedical applications of multifunctional magnetic nanoparticles, Adv. Mater., 22, pp.2729-2742. 2010. Hardy, P. A. and R. M. Henkelman. Transverse relaxation rate enhancement caused by magnetic particulates, Magn. Reson. Imaging, 7, pp.265-275. 1989. Hatton, T. A. and M. Lattuada. Functionalization of monodisperse magnetic nanoparticles, Langmuir, 23, pp.2158-2168. 2007. Hayashi, K., M. Moriya, W. Sakamoto and T. Yogo. Chemoselective synthesis of folic acid-functionalized magnetite nanoparticles via click chemistry for magnetic hyperthermia, Chem. Mater., 21, pp.1318-1325. 2009. He, B., J. Z. Bei and S. G. Wang. Morphology and degradation of biodegradable poly(L-lactide-co-beta-malic acid), Polym. Advan. Technol., 14, pp.645-652. 2003. He, B., Y. Q. Wan, J. Z. Bei and S. G. Wang. Synthesis and cell affinity of functionalized poly(L-lactide-co-beta-malic acid) with high molecular weight, Biomaterials, 25, pp.5239-5247. 2004. 159 References Hernandezcaselles, T., J. Villalain and J. C. Gomezfernandez. Influence of liposome charge and composition on their interaction with human blood-serum proteins, Mol. Cell. Biochem., 120, pp.119-126. 1993. Horak, D. and P. Shapoval. Reactive poly(glycidyl methacrylate) microspheres prepared by dispersion polymerization, J. Polym. Sci. Polym. Chem. Ed., 38, pp.3855-3863. 2000. Horak, D., M. Trchova, M. J. Benes, M. Veverka and E. Pollert. Monodisperse magnetic composite poly(glycidyl methacrylate)/La0.75Sr0.25MnO3 microspheres by the dispersion polymerization, Polymer, 51, pp.3116-3122. 2010. Hu, F. X., K. G. Neoh, L. Cen and E. T. Kang. Cellular response to magnetic nanoparticles "PEGylated" via surface-initiated atom transfer radical polymerization, Biomacromolecules, 7, pp.809-816. 2006. Hu, F. X., K. G. Neoh and E. T. Kang. Synthesis of folic acid functionalized PLLA-bPPEGMA nanoparticles for cancer cell targeting, Macromol. Rapid Commun., 30, pp.609-614. 2009. Huang, C., K. G. Neoh, L. Wang, E. T. Kang and B. Shuter. Magnetic nanoparticles for magnetic resonance imaging: modulation of macrophage uptake by controlled PEGylation of the surface coating, J. Mater. Chem., 20, pp.8512-8520. 2010. Huh, Y. M., Y. W. Jun, H. T. Song, S. Kim, J. S. Choi, J. H. Lee, S. Yoon, K. S. Kim, J. S. Shin, J. S. Suh and J. Cheon. In vivo magnetic resonance detection of cancer by using multifunctional magnetic nanocrystals, J. Am. Chem. Soc., 127, pp.1238712391. 2005. Huh, Y. M., J. Yang, C. H. Lee, H. J. Ko, J. S. Suh, H. G. Yoon, K. Lee and S. Haam. Multifunctional magneto-polymeric nanohybrids for targeted detection and synergistic therapeutic effects on breast cancer, Angew. Chem. Int. Ed., 46, pp.88368839. 2007. Huynh, V. T., G. J. Chen, P. de Souza and M. H. Stenzel. Thiol-yne and Thiol-ene "click" chemistry as a tool for a variety of platinum drug delivery carriers, from statistical copolymers to crosslinked micelles, Biomacromolecules, 12, pp.1738-1751. 2011a. Huynh, V. T., P. de Souza and M. H. Stenzel. Polymeric micelles with pendant dicarboxylato chelating ligands prepared via a Michael Addition for cis-platinum drug delivery, Macromolecules, 44, pp.7888-7900. 2011b. 160 References Hyeon, T., S. S. Lee, J. Park, Y. Chung and H. Bin Na. Synthesis of highly crystalline and monodisperse maghemite nanocrystallites without a size-selection process, J. Am. Chem. Soc., 123, pp.12798-12801. 2001. Hyeon, T., J. Kim, J. E. Lee, J. Lee, J. H. Yu, B. C. Kim, K. An, Y. Hwang, C. H. Shin, J. G. Park and J. Kim. Magnetic fluorescent delivery vehicle using uniform mesoporous silica spheres embedded with monodisperse magnetic and semiconductor nanocrystals, J. Am. Chem. Soc., 128, pp.688-689. 2006. Ishikawa, T., S. Kataoka and K. Kandori. The influence of carboxylate ions on the growth of beta-Feooh particles, J. Mater. Sci., 28, pp.2693-2698. 1993. Jang, J. T., H. Nah, J. H. Lee, S. H. Moon, M. G. Kim and J. Cheon. Critical enhancements of MRI contrast and hyperthermic effects by dopant-controlled magnetic nanoparticles, Angew. Chem. Int. Ed., 48, pp.1234-1238. 2009. Jun, Y. W., J. H. Lee and J. Cheon. Chemical design of nanoparticle probes for highperformance magnetic resonance imaging, Angew. Chem. Int. Ed., 47, pp.5122-5135. 2008. Kajiyama, T., H. Kobayashi, T. Taguchi, K. Kataoka and J. Tanaka. Improved synthesis with high yield and increased molecular weight of poly(alpha,beta-malic acid) by direct polycondensation, Biomacromolecules, 5, pp.169-174. 2004. Kamat, M., K. El-Boubbou, D. C. Zhu, T. Lansdell, X. Lu, W. Li and X. Huang. Hyaluronic acid immobilized magnetic nanoparticles for active targeting and imaging of macrophages, Bioconjugate Chem., 21, pp.2128-2135. 2010. Ke, C. Y., C. J. Mathias and M. A. Green. Folate-receptor-targeted radionuclide imaging agents, Adv. Drug Deliver Rev., 56, pp.1143-1160. 2004. Ke, J. H., J. J. Lin, J. R. Carey, J. S. Chen, C. Y. Chen and L. F. Wang. A specific tumor-targeting magnetofluorescent nanoprobe for dual-modality molecular imaging, Biomaterials, 31, pp.1707-1715. 2010. Kim, I.-B., H. Shin, A. J. Garcia and U. H. F. Bunz. Use of a folate−PPE conjugate to image cancer cells in vitro, Bioconjugate Chem., 18, pp.815-820. 2007. Kim, J., J. E. Lee, S. H. Lee, J. H. Yu, J. H. Lee, T. G. Park and T. Hyeon. Designed fabrication of a multifunctional polymer nanomedical platform for simultaneous cancer-targeted imaging and magnetically guided drug delivery, Adv. Mater., 20, pp.478-483. 2008. 161 References Komane, L. L., E. H. Mukaya, E. W. Neuse and C. E. J. van Rensburg. Macromolecular antiproliferative agents featuring dicarboxylato-chelated platinum, J. Inorg. Organomet. Polym. Mater., 18, pp.111-123. 2008. Korgel, B. A., A. T. Heitsch, D. K. Smith, R. N. Patel and D. Ress. Multifunctional particles: Magnetic nanocrystals and gold nanorods coated with fluorescent dyedoped silica shells, J. Solid State Chem., 181, pp.1590-1599. 2008. Kovalenko, M. V., M. I. Bodnarchuk, R. T. Lechner, G. Hesser, F. Schaffler and W. Heiss. Fatty acid salts as stabilizers in size- and shape-controlled nanocrystal synthesis: The case of inverse spinel iron oxide, J. Am. Chem. Soc., 129, pp.63526353. 2007. Krimmer, S. G., H. Z. Pan, J. H. Liu, J. Y. Yang and J. Kopecek. Synthesis and Characterization of Poly(epsilon-caprolactone)-block-poly[N-(2hydroxypropyl)methacrylamide] micelles for drug delivery, Macromol. Biosci., 11, pp.1041-1051. 2011. Kumar, C. S. S. R., J. Hormes and C. Leuschner. Nanofabrication towards biomedical applications : Techniques, tools, applications, and impact. Weinheim, WileyVCH.2005. Kumar, C. S. S. R. and F. Mohammad. Magnetic nanomaterials for hyperthermiabased therapy and controlled drug delivery, Adv. Drug Deliver Rev., 63, pp.789-808. 2011. Labhasetwar, V., M. M. Yallapu, S. P. Foy and T. K. Jain. PEG-functionalized magnetic nanoparticles for drug delivery and magnetic resonance imaging applications, Pharm. Res., 27, pp.2283-2295. 2010. Lai, P. S., J. R. Lai, Y. W. Chang, H. C. Yen, N. Y. Yuan, M. Y. Liao, C. Y. Hsu and J. L. Tsai. Multifunctional doxorubicin/superparamagnetic iron oxide-encapsulated Pluronic F127 micelles used for chemotherapy/magnetic resonance imaging, J. Appl. Phys., 107, pp.2010. Landmark, K. J., S. DiMaggio, J. Ward, C. V. Kelly, S. Vogt, S. Hong, A. Kotlyar, A. Myc, T. P. Thomas, J. E. Penner-Hahn, J. R. Baker, M. M. B. Holl and B. G. Orr. Synthesis, characterization, and in vitro testing of superparamagnetic iron oxide nanoparticles targeted using folic acid-conjugated dendrimers, ACS Nano, 2, pp.773783. 2008. Lattuada, M. and T. A. Hatton. Functionalization of monodisperse magnetic nanoparticles, Langmuir, 23, pp.2158-2168. 2007. 162 References Leakakos, T., C. Ji, G. Lawson, C. Peterson and S. Goodwin. Intravesical administration of doxorubicin to swine bladder using magnetically targeted carriers, Cancer Chemother. Pharmacol., 51, pp.445-450. 2003. Lee, J. H., Y. M. Huh, Y. Jun, J. Seo, J. Jang, H. T. Song, S. Kim, E. J. Cho, H. G. Yoon, J. S. Suh and J. Cheon. Artificially engineered magnetic nanoparticles for ultrasensitive molecular imaging, Nat. Med., 13, pp.95-99. 2007. Lee, K. M., H. L. Liu, C. H. Sonn, J. H. Wu and Y. K. Kim. Synthesis of streptavidinFITC-conjugated core-shell Fe3O4-Au nanocrystals and their application for the purification of CD4(+) lymphocytes, Biomaterials, 29, pp.4003-4011. 2008. Lee, S. M., T. V. O'Halloran and S. T. Nguyen. Polymer-caged nanobins for synergistic cisplatin-doxorubicin combination chemotherapy, J. Am. Chem. Soc., 132, pp.17130-17138. 2010. Lele, B. S. and A. S. Hoffman. Mucoadhesive drug carriers based on complexes of poly(acrylic acid) and PEGylated drugs having hydrolysable PEG-anhydride-drug linkages, J. Control. Release, 69, pp.237-248. 2000. Levchenko, T. S., R. Rammohan, A. N. Lukyanov, K. R. Whiteman and V. P. Torchilin. Liposome clearance in mice: the effect of a separate and combined presence of surface charge and polymer coating, Int. J. Pharm., 240, pp.95-102. 2002. Li, J. H., R. Y. Hong, H. Z. Li, J. Ding, Y. Zheng and D. G. Wei. Simple synthesis and magnetic properties of Fe3O4/BaSO4 multi-core/shell particles, Mater. Chem. Phys., 113, pp.140-144. 2009. Liu, H. M., J. K. Hsiao, H. H. Chu, Y. H. Wang, C. W. Lai, P. T. Choii, S. T. Hsieh and J. L. Wang. Macrophage physiological function after superparamagnetic iron oxide labeling, NMR Biomed., 21, pp.820-829. 2008a. Liu, P., B. Mu, T. M. Wang, Z. H. Wu, H. G. Shi and D. S. Xue. Fabrication of functional block copolymer grafted superparamagnetic nanoparticles for targeted and controlled drug delivery, Colloid. Surface. A, 375, pp.163-168. 2011a. Liu, P., B. M. Mu, B., P. C. Du, Y. Dong and C. Y. Lu. Magnetic-targeted pHresponsive drug delivery system via Layer-by-Layer self-assembly of polyelectrolytes onto drug-containing emulsion droplets and its controlled release, J. Polym. Sci., Part A: Polym. Chem., 49, pp.1969-1976. 2011b. 163 References Liu, X., Y. Wang, K. Nakamura, A. Kubo and D. J. Hnatowich. Cell studies of a three-component antisense MORF/tat/Herceptin nanoparticle designed for improved tumor delivery, Cancer Gene Ther., 15, pp.126-132. 2008b. Liu, Y. T., K. Li, J. Pan, B. Liu and S. S. Feng. Folic acid conjugated nanoparticles of mixed lipid monolayer shell and biodegradable polymer core for targeted delivery of Docetaxel, Biomaterials, 31, pp.330-338. 2010. Lu, A. H., E. L. Salabas and F. Schuth. Magnetic nanoparticles: Synthesis, protection, functionalization, and application, Angew. Chem., Int. Ed., 46, pp.1222-1244. 2007. Lu, J., Z. P. Xiao, K. M. Yang and H. Liang. Synthesis of magnetic, reactive, and thermoresponsive Fe3O4 nanoparticles via surface-initiated RAFT copolymerization of N-isopropylacrylamide and acrolein, J. Polym. Sci. Pol. Chem., 48, pp.542-550. 2010. Luciani, N., F. Gazeau and C. Wilhelm. Reactivity of the monocyte/macrophage system to superparamagnetic anionic nanoparticles, J. Mater. Chem., 19, pp.63736380. 2009. Ma, Z. Y., Y. P. Guan and H. Z. Liu. Synthesis and characterization of micron-sized monodisperse superparamagnetic polymer particles with amino groups, J. Polym. Sci. Polym. Chem. Ed., 43, pp.3433-3439. 2005. Maeda, H. The enhanced permeability and retention (EPR) effect in tumor vasculature: The key role of tumor-selective macromolecular drug targeting, Adv. Enzyme. Regul., 41, pp.189-207. 2001. Maeng, J. H., D. H. Lee, K. H. Jung, Y. H. Bae, I. S. Park, S. Jeong, Y. S. Jeon, C. K. Shim, W. Kim, J. Kim, J. Lee, Y. M. Lee, J. H. Kim, W. H. Kim and S. S. Hong. Multifunctional doxorubicin loaded superparamagnetic iron oxide nanoparticles for chemotherapy and magnetic resonance imaging in liver cancer, Biomaterials, 31, pp.4995-5006. 2010. Maitra, A. and S. Bisht. Dextran-doxorubicin/chitosan nanoparticles for solid tumor therapy, Wiley Interdiscip. Rev. Nanomed. Nanobiotechnol., 1, pp.415-425. 2009. Majewski, P. and B. Thierry. Functionalized magnetite nanoparticles - Synthesis, properties, and bio-applications, Crit. Rev. Solid State Mater. Sci., 32, pp.203-215. 2007. Malik, N., E. G. Evagorou and R. Duncan. Dendrimer-platinate: A novel approach to cancer chemotherapy, Anti-Cancer Drugs, 10, pp.767-776. 1999. 164 References McBain, S. C., H. H. P. Yiu and J. Dobson. Magnetic nanoparticles for gene and drug delivery, Int. J. Nanomed., 3, pp.169-180. 2008. Mindt, T. L., C. Muller, M. Melis, M. de Jong and R. Schibli. "Click-to-Chelate": In vitro and in vivo comparison of a Tc-99m(CO)(3)-labeled N(tau)-histidine folate derivative with its isostructural, clicked 1,2,3-triazole analogue, Bioconjugate Chem., 19, pp.1689-1695. 2008. Minotti, G. Sources and role of iron in lipid-peroxidation, Chem. Res. Toxicol., 6, pp.134-146. 1993. Moghimi, S. M., A. C. Hunter and J. C. Murray. Long-circulating and target-specific nanoparticles: Theory to practice, Pharmacol. Rev., 53, pp.283-318. 2001. Moghimi, S. M. and J. Szebeni. Stealth liposomes and long circulating nanoparticles: critical issues in pharmacokinetics, opsonization and protein-binding properties, Prog. Lipid Res., 42, pp.463-478. 2003. Morales, M. P., S. Veintemillas-Verdaguer, M. I. Montero, C. J. Serna, A. Roig, L. Casas, B. Martinez and F. Sandiumenge. Surface and internal spin canting in gammaFe2O3 nanoparticles, Chem. Mater., 11, pp.3058-3064. 1999. Moulder, J. F. and J. Chastain. Handbook of X-ray photoelectron spectroscopy : a referen ce book of standard spectra for identification and interp retation of XPS data. Eden Prairie, Minn., : Perkin-Elmer Corporation.1992. Mugabe, C., B. A. Hadaschik, R. K. Kainthan, D. E. Brooks, A. I. So, M. E. Gleave and H. M. Burt. Paclitaxel incorporated in hydrophobically derivatized hyperbranched polyglycerols for intravesical bladder cancer therapy, BJU Int., 103, pp.978-986. 2009. Muller, R. N., S. Laurent, D. Forge, M. Port, A. Roch, C. Robic and L. V. Elst. Magnetic iron oxide nanoparticles: Synthesis, stabilization, vectorization, physicochemical characterizations, and biological applications, Chem. Rev., 108, pp.2064-2110. 2008. Munteanu, M., S. Choi and H. Ritter. Cyclodextrin methacrylate via microwaveassisted click reaction, Macromolecules, 41, pp.9619-9623. 2008. Nel, A. E., M. Liong, J. Lu, M. Kovochich, T. Xia, S. G. Ruehm, F. Tamanoi and J. I. Zink. Multifunctional inorganic nanoparticles for imaging, targeting, and drug delivery, ACS Nano, 2, pp.889-896. 2008. 165 References Neugebauer, D., K. Bury and T. Biela. Methacrylate copolymers with hydroxyl terminated caprolactone chains via ATRP. A route to grafted copolymers, React. Funct. Polym., 71, pp.616-624. 2011. Neuwelt, E. A., P. Varallyay, G. Nesbit, L. L. Muldoon, R. R. Nixon, J. Delashaw, J. I. Cohen, A. Petrillo and D. Rink. Comparison of two superparamagnetic viral-sized iron oxide particles ferumoxides and ferumoxtran-10 with a gadolinium chelate in imaging intracranial tumors, Am. J. Neuroradiol., 23, pp.510-519. 2002. Nishiyama, N., M. Yokoyama, T. Aoyagi, T. Okano, Y. Sakurai and K. Kataoka. Preparation and characterization of self-assembled polymer-metal complex micelle from cis-dichlorodiammineplatinum(II) and poly(ethylene glycol)-poly(alpha,betaaspartic acid) block copolymer in an aqueous medium, Langmuir, 15, pp.377-383. 1999. Nishiyama, N. and K. Kataoka. Preparation and characterization of size-controlled polymeric micelle containing cis-dichlorodiammineplatinum(II) in the core, J. Control. Release, 74, pp.83-94. 2001. Nishiyama, N., S. Okazaki, H. Cabral, M. Miyamoto, Y. Kato, Y. Sugiyama, K. Nishio, Y. Matsumura and K. Kataoka. Novel cisplatin-incorporated polymeric micelles can eradicate solid tumors in mice, Cancer Res., 63, pp.8977-8983. 2003. Noth, U., A. Heymer, D. Haddad, M. Weber, U. Gbureck, P. M. Jakob and J. Eulert. Iron oxide labelling of human mesenchymal stem cells in collagen hydrogels for articular cartilage repair, Biomaterials, 29, pp.1473-1483. 2008. Oh, J. K. and J. M. Park. Iron oxide-based superparamagnetic polymeric nanomaterials: Design, preparation, and biomedical application, Prog. Polym. Sci., 36, pp.168-189. 2011. Onoa, G. B., V. Moreno, M. Font-Bardia, X. Solans, J. M. Perez and C. Alonso. Structural and cytotoxic study of new Pt(II) and Pd(II) complexes with the biheterocyclic ligand mepirizole, J. Inorg. Biochem., 75, pp.205-212. 1999. Pan, J. and S. S. Feng. Targeting and imaging cancer cells by Folate-decorated, quantum dots (QDs)-loaded nanoparticles of biodegradable polymers, Biomaterials, 30, pp.1176-1183. 2009. Pankhurst, Q. A., J. Connolly, S. K. Jones and J. Dobson. Applications of magnetic nanoparticles in biomedicine, J. Phys. D: Appl. Phys., 36, pp.167-181. 2003. 166 References Penczek, S., A. Duda and R. Szymanski. Intra- and intermolecular chain transfer to macromolecules with chain scission. The case of cyclic esters, Macromol. Symp., 132, pp.441-449. 1998. Photos, P. J., L. Bacakova, B. Discher, F. S. Bates and D. E. Discher. Polymer vesicles in vivo: Correlations with PEG molecular weight, Journal of Control. Release, 90, pp.323-334. 2003. Ploeg, M., K. K. H. Aben and L. A. Kiemeney. The present and future burden of urinary bladder cancer in the world, World J. Urol., 27, pp.289-293. 2009. Qiao, R. R., C. H. Yang and M. Y. Gao. Superparamagnetic iron oxide nanoparticles: from preparations to in vivo MRI applications, J. Mater. Chem., 19, pp.6274-6293. 2009. Qin, J., S. Laurent, Y. S. Jo, A. Roch, M. Mikhaylova, Z. M. Bhujwalla, R. N. Muller and M. Muhammed. A high-performance magnetic resonance imaging T2 contrast agent, Adv. Mater., 19, pp.1874-1878. 2007. Reimhult, E., E. Amstad, T. Gillich, I. Bilecka and M. Textor. Ultrastable iron oxide nanoparticle colloidal suspensions using dispersants with catechol-derived anchor groups, Nano Lett., 9, pp.4042-4048. 2009. Rockenberger, J., E. C. Scher and A. P. Alivisatos. A new nonhydrolytic singleprecursor approach to surfactant-capped nanocrystals of transition metal oxides, J. Am. Chem. Soc., 121, pp.11595-11596. 1999. Rogers, W. J. and P. Basu. Factors regulating macrophage endocytosis of nanoparticles: Implications for targeted magnetic resonance plaque imaging, Atherosclerosis, 178, pp.67-73. 2005. Rohrer, M., H. Bauer, J. Mintorovitch, M. Requardt and H. J. Weinmann. Comparison of magnetic properties of MRI contrast media solutions at different magnetic field strengths, Invest. Radiol., 40, pp.715-724. 2005. Ross, J. F., P. K. Chaudhuri and M. Ratnam. Differential regulation of folate receptor isoforms in normal and malignant tissues in vivo and in established cell-lines. Physiological and clinical implications, Cancer, 73, pp.2432-2443. 1994. Rutt, B. K., C. V. Bowen, X. W. Zhang, G. Saab and P. J. Gareau. Application of the static dephasing regime theory to superparamagnetic iron-oxide loaded cells, Magn. Reson. Med., 48, pp.52-61. 2002. 167 References Saeed, A. O., J. P. Magnusson, E. Moradi, M. Soliman, W. X. Wang, S. Stolnik, K. J. Thurecht, S. M. Howdle and C. Alexander. Modular construction of multifunctional bioresponsive cell-targeted nanoparticles for gene delivery, Bioconjugate Chem., 22, pp.156-168. 2011. Satarkar, N. S. and J. Z. Hilt. Magnetic hydrogel nanocomposites for remote controlled pulsatile drug release, J. Control. Release, 130, pp.246-251. 2008. Schladt, T. D., K. Schneider, H. Schild and W. Tremel. Synthesis and biofunctionalization of magnetic nanoparticles for medical diagnosis and treatment, Dalton T., 40, pp.6315-6343. 2011. Schmidt, A. M., A. Kaiser and S. Dutz. Kinetic studies of surface-initiated atom transfer radical polymerization in the synthesis of magnetic fluids, J. Polym. Sci., Part A: Polym. Chem., 47, pp.7012-7020. 2009. Sengupta, S., P. Tyagi, T. Velpandian, Y. K. Gupta and S. K. Gupta. Etoposide encapsulated in positively charged liposomes: Pharmacokinetic studies in mice and formulation stability studies, Pharmacol. Res., 42, pp.459-464. 2000. Seo, W. S., J. H. Lee, X. M. Sun, Y. Suzuki, D. Mann, Z. Liu, M. Terashima, P. C. Yang, M. V. McConnell, D. G. Nishimura and H. J. Dai. FeCo/graphitic-shell nanocrystals as advanced magnetic-resonance-imaging and near-infrared agents, Nat. Mater., 5, pp.971-976. 2006. Setua, S., D. Menon, A. Asok, S. Nair and M. Koyakutty. Folate receptor targeted, rare-earth oxide nanocrystals for bi-modal fluorescence and magnetic imaging of cancer cells, Biomaterials, 31, pp.714-729. 2010. Sha, K., D. S. Li, Y. P. Li, X. T. Liu, S. W. Wang, J. Q. Guan and J. Y. Wang. Synthesis, characterization, and micellization of an epoxy-based amphiphilic diblock copolymer of epsilon-caprolactone and glycidyl methacrylate by enzymatic ringopening polymerization and atom transfer radical polymerization, J. Polym. Sci. Polym. Chem. Ed., 45, pp.5037-5049. 2007. Shen, Z. C., T. Shen, M. G. Wientjes, M. A. O'Donnell and J. L. S. Au. Intravesical treatments of bladder cancer: Review, Pharm. Res., 25, pp.1500-1510. 2008. Shuai, X. T., X. Q. Yang, Y. H. Chen, R. X. Yuan, G. H. Chen, E. Blanco and J. M. Gao. Folate-encoded and Fe3O4-loaded polymeric micelles for dual targeting of cancer cells, Polymer, 49, pp.3477-3485. 2008. 168 References Shubayev, V. I., T. R. Pisanic and S. H. Jin. Magnetic nanoparticles for theragnostics, Adv. Drug Deliver Rev., 61, pp.467-477. 2009. Smart, J. D. The basics and underlying mechanisms of mucoadhesion, Adv. Drug Deliver Rev., 57, pp.1556-1568. 2005. Stickler, D. J. and S. D. Morgan. Modulation of crystalline Proteus mirabilis biofilm development on urinary catheters, J. Med. Microbiol., 55, pp.489-494. 2006. Stoll, G., C. Wesemeier, R. Gold, L. Solymosi, K. V. Toyka and M. Bendszus. In vivo monitoring of macrophage infiltration in experimental autoimmune neuritis by magnetic resonance imaging, J. Neuroimmunol., 149, pp.142-146. 2004. Sun, C., R. Sze and M. Q. Zhang. Folic acid-PEG conjugated superparamagnetic nanoparticles for targeted cellular uptake and detection by MRI, J. Biomed. Mater. Res. A, 78A, pp.550-557. 2006. Sun, C., J. S. H. Lee and M. Q. Zhang. Magnetic nanoparticles in MR imaging and drug delivery, Adv. Drug Del. Rev., 60, pp.1252-1265. 2008. Sun, S., J. Xie, C. Xu, N. Kohler and Y. Hou. Controlled PEGylation of monodisperse Fe3O4 nanoparticles for reduced non-specific uptake by macrophage cells, Adv. Mater., 19, pp.3163-3166. 2007a. Sun, S. H. and H. Zeng. Size-controlled synthesis of magnetite nanoparticles, J. Am. Chem. Soc., 124, pp.8204-8205. 2002. Sun, S. H., H. Zeng, D. B. Robinson, S. Raoux, P. M. Rice, S. X. Wang and G. X. Li. Monodisperse MFe2O4 (M = Fe, Co, Mn) nanoparticles, J. Am. Chem. Soc., 126, pp.273-279. 2004. Sun, Y. B., X. B. Ding, Z. H. Zheng, X. Cheng, X. H. Hu and Y. X. Peng. Surface initiated ATRP in the synthesis of iron oxide/polystyrene core/shell nanoparticles, Eur. Polym. J., 43, pp.762-772. 2007b. Tan, K. L., L. L. Woon, H. K. Wong, E. T. Kang and K. G. Neoh. Surface modification of plasma-pretreated poly(tetrafluoroethylene) films by graftcopolymerization, Macromolecules, 26, pp.2832-2836. 1993. Tartaj, P., T. Gonzalez-Carreno and C. J. Serna. Synthesis of nanomagnets dispersed in colloidal silica cages with applications in chemical separation, Langmuir, 18, pp.4556-4558. 2002. 169 References Teja, A. S. and P. Y. Koh. Synthesis, properties, and applications of magnetic iron oxide nanoparticles, Prog. Cryst. Growth Charact. Mater., 55, pp.22-45. 2009. Theil, E. C. Ferritin: At the crossroads of iron and oxygen metabolism, J. Nutr., 133, pp.1549-1553. 2003. Tsourkas, A., D. L. J. Thorek, A. Chen and J. Czupryna. Superparamagnetic iron oxide nanoparticle probes for molecular imaging, Ann. Biomed. Eng., 34, pp.23-38. 2006. Tyagi, P., P. C. Wu, M. Chancellor, N. Yoshimura and L. Huang. Recent advances in intravesical drug/gene delivery, Mol. Pharm., 3, pp.369-379. 2006. Uchino, H., Y. Matsumura, T. Negishi, F. Koizumi, T. Hayashi, T. Honda, N. Nishiyama, K. Kataoka, S. Naito and T. Kakizoe. Cisplatin-incorporating polymeric micelles (NC-6004) can reduce nephrotoxicity and neurotoxicity of cisplatin in rats, Br. J. Cancer, 93, pp.678-687. 2005. Ulman, A. Formation and structure of self-assembled monolayers, Chem. Rev., 96, pp.1533-1554. 1996. Veiseh, O., F. M. Kievit, R. G. Ellenbogen and M. Q. Zhang. Cancer cell invasion: Treatment and monitoring opportunities in nanomedicine, Adv. Drug Deliver Rev., 63, pp.582-596. 2011. Vyas, S. P., S. Jain, V. Mishra, P. Singh, P. K. Dubey and D. K. Saraf. RGD-anchored magnetic liposomes for monocytes/neutrophils-mediated brain targeting, Int. J. Pharm., 261, pp.43-55. 2003. Waite, J. H. and M. L. Tanzer. Polyphenolic substance of Mytilus-Edulis - Novel adhesive containing L-Dopa and hydroxyproline, Science, 212, pp.1038-1040. 1981. Wang, L., K. G. Neoh, E. T. Kang and B. Shuter. Multifunctional polyglycerolgrafted Fe3O4@SiO2 nanoparticles for targeting ovarian cancer cells, Biomaterials, 32, pp.2166-2173. 2011. Wang, S., R. J. Lee, C. J. Mathias, M. A. Green and P. S. Low. Synthesis, purification, and tumor cell uptake of Ga-67-deferoxamine-folate, a potential radiopharmaceutical for tumor imaging, Bioconjugate Chem., 7, pp.56-62. 1996. 170 References Wang, S. H., X. Y. Shi, M. Van Antwerp, Z. Y. Cao, S. D. Swanson, X. D. Bi and J. R. Baker. Dendrimer-functionalized iron oxide nanoparticles for specific targeting and imaging of cancer cells, Adv. Funct. Mater., 17, pp.3043-3050. 2007. Wang, W. C., Q. Zhang, B. B. Zhang, D. N. Li, X. Q. Dong, L. Zhang and J. Chang. Preparation of monodisperse, superparamagnetic, luminescent., and multifunctional PGMA microspheres with amino-groups, Chin. Sci. Bull., 53, pp.1165-1170. 2008. Weissleder, R., A. S. Lee, A. J. Fischman, P. Reimer, T. Shen, R. Wilkinson, R. J. Callahan and T. J. Brady. Polyclonal human immunoglobulin-G labeled with polymeric iron-oxide - antibody MR imaging, Radiology, 181, pp.245-249. 1991. Wilhelm, C., N. Luciani and F. Gazeau. Reactivity of the monocyte/macrophage system to superparamagnetic anionic nanoparticles, J. Mater. Chem., 19, pp.63736380. 2009. Williams, M. E. and A. H. Latham. Versatile routes toward functional, water-soluble nanoparticles via trifluoroethylester-PEG-thiol ligands, Langmuir, 22, pp.4319-4326. 2006. Woo, K., J. Hong, S. Choi, H. W. Lee, J. P. Ahn, C. S. Kim and S. W. Lee. Easy synthesis and magnetic properties of iron oxide nanoparticles, Chem. Mater., 16, pp.2814-2818. 2004. Wuang, S. C., K. G. Neoh, E. T. Kang, D. W. Pack and D. E. Leckband. Heparinized magnetic nanoparticles: In vitro assessment for biomedical applications, Adv. Funct. Mater., 16, pp.1723-1730. 2006. Xia, W. and P. S. Low. Folate-Targeted Therapies for Cancer, J. Med. Chem., 53, pp.6811-6824. 2010. Xie, J., C. Xu, N. Kohler, Y. Hou and S. Sun. Controlled PEGylation of monodisperse Fe3O4 nanoparticles for reduced non-specific uptake by macrophage cells, Adv. Mater., 19, pp.3163-3166. 2007. Xu, B., C. J. Xu, K. M. Xu, H. W. Gu, R. K. Zheng, H. Liu, X. X. Zhang and Z. H. Guo. Dopamine as a robust anchor to immobilize functional molecules on the iron oxide shell of magnetic nanoparticles, J. Am. Chem. Soc., 126, pp.9938-9939. 2004. Yang, J., E. K. Lim, H. J. Lee, J. Park, S. C. Lee, K. Lee, H. G. Yoon, J. S. Suh, Y. M. Huh and S. Haam. Fluorescent magnetic nanohybrids as multimodal imaging agents for human epithelial cancer detection, Biomaterials, 29, pp.2548-2555. 2008. 171 References Ye, L., K. Letchford, M. Heller, R. Liggins, D. Guan, J. N. Kizhakkedathu, D. E. Brooks, J. K. Jackson and H. M. Burt. Synthesis and characterization of carboxylic acid conjugated, hydrophobically derivatized, hyperbranched polyglycerols as nanoparticulate drug carriers for cisplatin, Biomacromolecules, 12, pp.145-155. 2011. Zhang, M. Q., N. Kohler, C. Sun, A. Fichtenholtz, J. Gunn and C. Fang. Methotrexate-immobilized poly(ethylene glycol) magnetic nanoparticles for MR imaging and drug delivery, Small, 2, pp.785-792. 2006a. Zhang, M. Q., C. Sun and R. Sze. Folic acid-PEG conjugated superparamagnetic nanoparticles for targeted cellular uptake and detection by MRI, J. Biomed. Mater. Res. A, 78A, pp.550-557. 2006b. Zhang, Q., C. H. Wang, L. Qiao, H. S. Yan and K. L. Liu. Superparamagnetic iron oxide nanoparticles coated with a folate-conjugated polymer, J. Mater. Chem., 19, pp.8393-8402. 2009. Zhang, X. Q., S. W. Gong, Y. Zhang, T. Yang, C. Y. Wang and N. Gu. Prussian blue modified iron oxide magnetic nanoparticles and their high peroxidase-like activity, J. Mater. Chem., 20, pp.5110-5116. 2010. Zhang, Z. J. and C. R. Vestal. Atom transfer radical polymerization synthesis and magnetic characterization of MnFe2O4/polystyrene core/shell nanoparticles, J. Am. Chem. Soc., 124, pp.14312-14313. 2002. Zhou, P., Z. Y. Li and Y. Chau. Synthesis, characterization, and in vivo evaluation of poly(ethylene oxide-co-glycidol)-platinate conjugate, Eur. J. Pharm. Sci., 41, pp.464472. 2010. Zhu, W., Y. L. Li, L. X. Liu, W. L. Zhang, Y. M. Chen and F. Xi. Biamphiphilic triblock copolymer micelles as a multifunctional platform for anticancer drug delivery, J. Biomed. Mater. Res. A, 96A, pp.330-340. 2011. 172   LIST OF PUBLICATIONS ARISING FROM PHD WORK (1) Huang C., K. G. Neoh, L. Wang, E. T. Kang, and B. Shuter. Magnetic nanoparticles for magnetic resonance imaging: modulation of macrophage uptake by controlled PEGylation of the surface coating. J. Mater. Chem., 20, pp.8512-8520. 2010. (2) Huang C., K. G. Neoh, L. Wang, E. T. Kang, B. Shuter. Surface functionalization of superparamagnetic nanoparticles for the development of highly efficient magnetic resonance probe for macrophages. Contrast Media Mol. Imaging, 6, pp.298-307.2011. (3) Huang C., K. G. Neoh, E. T. Kang, and B. Shuter. Surface modified superparamagnetic iron oxide nanoparticles (SPIONs) for high efficiency folatereceptor targeting with low uptake by macrophages. J. Mater. Chem., 21, pp.16094-16102. 2011. (4) Huang C., K. G. Neoh, and E. T. Kang. Combined ATRP and ‘Click’ chemistry for designing stable tumor-targeting superparamagnetic iron oxide nanoparticles. Langmuir, 28, pp.563-571. 2012. (5) Huang C., K. G. Neoh, L. Q. Xu, E. T. Kang and E. Chiong. Polymeric nanoparticles with encapsulated superparamagnetic iron oxide and conjugated cisplatin for potential bladder cancer therapy. Biomacromolecules, 13, pp.2513-2520. 2012. 173 [...]... technologies of the 21st century, and it has greatly enabled the design of advanced functional nanomaterials of dimensions of 11000 nm in the biomedical field (Gupta and Gupta 2005) Among the different types of nanomaterials, magnetic iron oxide nanoparticles are of intense current interest and have been successfully used for clinical applications, for example, molecular imaging Magnetic iron oxide nanoparticles. .. organs is crucial for SPIONs-based biomedical applications 1.2 Research Objectives and Scopes The overall aim of this thesis is to develop various functional polymers as surface coatings of SPIONs for potential cell targeting, MRI, and cancer therapy applications This thesis consists of seven chapters In Chapter 1, a general introduction of the current problems for SPIONs-based biomedical applications, ... (s-1) and 1/T2 (s-1) in water as a function of the iron concentration of (a) PLMA-2-SPIONs and (b) PLMA-1PEG-3-SPIONs (for all plots, correlation coefficient R2 > 0.97) Relaxometric measurements were performed by MRI.      Figure 3-14  MR images of phantoms containing (a) macrophages without nanoparticles at a cell density of 200×103 cells/mL, and PLMA-2SPIONs-labeled macrophages at a cell density of. .. enhancing the selectivity in targeting of cancer cells ATRP of GMA and poly(ethylene glycol) methyl ether methacrylate (PEGMA) from the surface of SPIONs was first carried out, followed by ‘click’ chemistry to conjugate FA with controlled surface densities In Chapter 6, the preparation of nanoparticles incorporating SPIONs and drug for potential bladder cancer therapy is described Amphiphilic poly(ε-caprolactone)-b-poly(propargyl... (blue), and (c) combined with FITC and DAPI channels Scale bar=100 μm.  Figure 6-11 In vitro cytotoxicity profile of (a) free cisplatin, (b) Fe-PNs, and PtFe-PNs against UMUC3 bladder cancer cells Cells were exposed to the drug or nanoparticles for 2 h and further cultured with fresh medium for 72 h                                             XIV   LIST OF TABLES Table 3-1 Molecular weight and PEG... PLMA-2SPIONs, and (b) with PLMA-2-SPIONs at an iron concentration of 0.5 mM, after staining with Prussian Blue.      Figure 3-9  Uptake of PLMA-2-SPIONs by macrophages (a) as a function of incubation time at incubated iron concentration of 0.5 mM and (b) as a function of incubated iron concentration for an incubation period of 4 h Inset is the image of macrophages incubated with PLMA-2-SPIONs at an iron concentration... growth of nanotechnology over the past decade provides exciting possibilities for synthesis, characterization, and functionalization of nanoscale materials for biomedical applications and diagnostics (Schladt et al 2011) Among the variety of promising nanoscale materials, SPIONs have gained significant attention due to their great potential for various biomedical applications, including MRI for cell. .. nanoparticles by the addition of a base to an aqueous mixture of Fe3+ and Fe2+ salts under an inert atmosphere The size, shape, and composition of the iron oxide nanoparticles depends on the type of salts (e.g nitrates, chlorides, sulphates, etc.), the ratio of Fe3+ /Fe2+, the reaction temperature, the pH value and ionic strength of the medium 8  Chapter 2 The formation of magnetite (Fe3O4) is expected... kidney, and thyroid (Moghimi et al 2001) Figure 2-3 In vivo behavior of nanoparticles in blood vessels The EPR effect of nanoparticles is greatest at tumors (Jun et al 2008) 2.2 Surface Functionalization of SPIONs Although superparamagnetic iron oxide nanoparticles synthesized by hightemperature decomposition method are monodisperse, they are typically coated with hydrophobic ligands, such as oleic acid and. .. acidstabilized SPIONs and (b) Pt-Fe-PNs Figure 6-8 Release profiles of cisplatin from Pt-Fe-PNs in DI water, PBS, and artificial urine at 37 oC Inset shows the release profile in the first 10 h Figure 6-9 Size increase of a mixture of Pt-Fe-PNs and mucin after incubation at 37oC for 2 h Pure mucin and nanoparticle suspensions were used as controls Figure 6-10 Fluorescence images of UMUC3 bladder cancer cells after .           NATIONAL UNIVERSITY OF SINGAPORE 2012     SURFACE FUNCTIONALIZATION OF SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLES FOR POTENTIAL CELL TARGETING, IMAGING, AND CANCER THERAPY APPLICATIONS .     SURFACE FUNCTIONALIZATION OF SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLES FOR POTENTIAL CELL TARGETING, IMAGING, AND CANCER THERAPY APPLICATIONS      HUANG. Superparamagnetic iron oxide nanoparticles (SPIONs) are very useful for biomedical applications, such as magnetic resonance imaging (MRI), hyperthermia for cancer therapy, cell targeting, drugs