Shell Nanoparticles for ... - ACS Publications

Subscriber access provided by UNIV OF CALIFORNIA SAN DIEGO LIBRARIES

Article

Au@MnS@ZnS core/shell/shell nanoparticles for magnetic resonance imaging and enhanced cancer radiation therapy Meifang Li, Qi Zhao, Xuan Yi, Xiaoyan Zhong, Guosheng Song, Zhifang Chai, Zhuang Liu, and Kai Yang ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.5b11588 • Publication Date (Web): 04 Apr 2016 Downloaded from http://pubs.acs.org on April 5, 2016

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

ACS Applied Materials & Interfaces is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

ACS Applied Materials & Interfaces

Au@MnS@ZnS Core/Shell/Shell Nanoparticles for Magnetic Resonance Imaging and Enhanced Cancer Radiation Therapy Meifang Li1, Qi Zhao1, Xuan Yi1, Xiaoyan Zhong1, Guosheng Song2, Zhifang Chai1, Zhuang Liu2*, Kai Yang1* 1. School of Radiation Medicine and Protection & School for Radiological and Interdisciplinary Sciences (RAD-X), Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, Medical College of Soochow University, Suzhou, Jiangsu 215123, China 2. Jiangsu Key Laboratory for Carbon-Based Functional Materials & Devices, Institute of Functional Nano & Soft Materials Laboratory (FUNSOM) Soochow University, Suzhou, Jiangsu 215123, China

KEYWORDS: Multifunctional nano-platforms, radiotherapy, MR imaging, Cancer treatment

Core-shell-shell

structure,

Enhanced

ABSTRACT: Although conventional radiotherapy (RT) has been widely used in the clinic to treat cancer, it often has limited therapeutic outcomes and severe toxic effects. There is still a need to develop theranostic agents with both imaging and RT enhancing functions to improve the accuracy and efficiency of RT. Herein, we synthesize Au@MnS@ZnS core/shell/shell nanoparticles with polyethylene glycol (PEG) functionalization, yielding Au@MnS@ZnS-PEG nanoparticles with great stability in different physiological solutions and no significant

ACS Paragon Plus Environment

1


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 2 of 25

cytotoxicity. It is found that Au@MnS@ZnS-PEG nanoparticles can enhance the cancer cell killing efficiency induced by RT, as evidenced by multiple in vitro assays. Owing to the existence of paramagnetic Mn2+ in the nanoparticle shell, our Au@MnS@ZnS-PEG can be used as a contrast agent for T1-weighted magnetic resonance (MR) imaging, which reveals the efficient accumulation and retention of nanoparticles in the tumors of mice after intravenous injection. Importantly, by exposing tumor-bearing mice that were injected with Au@MnS@ZnSPEG to X-ray irradiation, the tumor growth can be significantly inhibited. This result shows clearly improved therapeutic efficacy compared to RT alone. Furthermore, no obvious side effect of Au@MnS@ZnS-PEG is observed in the injected mice. Therefore, our work presents a new type of radiosensitizing agent, which is promising for the imaging-guided enhanced RT treatment of cancer.

1. Introduction Radiotherapy (RT), as the main therapeutics for cancer treatment, has been widely used in the clinic. Almost 50% of cancer patients undergo RT at least once during their treatment course1-4. Relying on the ionization irradiation, RT induces intracellular DNA damage by producing a series of free electrons and ions5-9. However, an important challenge of RT is that ionizing radiation damages both healthy tissues and tumors with little specificity. In recent years, a number of different radio-sensitizers, many of which are nanoparticles containing high Zelements capable of absorbing ionizing radiation, have been widely explored to enhance the therapeutic efficiency and specificity of RT10-22. Once such agents are accumulated in the tumor, they help concentrate the ionizing radiation energy into the tumor. This enhances the RT-induced damage to tumor cells and reduces unnecessary injury to healthy tissues.


2

Page 3 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


To optimize the efficacy of enhanced RT using radiosensitizing agents, imaging is important for locating tumors and providing their size/shape information. In addition, imaging helps determine the best time for applying ionizing radiation, when the radiosensitizing agent reaches its peak level in the tumor. Magnetic resonance (MR) imaging with high spatial and temporal resolution has been widely applied in clinic without inducing harmful effect to patients. Therefore, it may be interesting to develop multifunctional theranostic agents that have the capability of enhancing RT and that offer strong contrast in MR imaging to realize the imagingguided enhanced RT. In this work, we designed a novel theranostic agent that is based on gold / manganese sulfide (MnS) / zinc sulfide (ZnS) (Au@MnS@ZnS) core / shell / shell nanoparticles for MR imagingguided enhanced RT treatment of cancer (Figure 1a). In this core/shell/shell nano-structure, the Au core is able to absorb ionizing radiation to enhance RT, the MnS inner shell offers T1contrast in MR imaging, while the ZnS outer shell is able to stabilize the MnS shell by protecting it from oxidization in aqueous solutions. After being noncovalently modified with polyethylene glycol (PEG), the obtained Au@MnS@ZnS-PEG nanoparticles exhibited high solubility in physiological solutions and did not induce any obvious toxicity to cells. As evidenced by clonogenic assay and γ-H2AX immunofluorescence staining, we found that Au@MnS@ZnSPEG nanoparticles significantly enhanced the in vitro cancer cell killing efficacy induced by Xray irradiation. Under the guidance of in vivo MR imaging, which illustrated the efficient tumor accumulation of our synthesized Au@MnS@ZnS-PEG nanoparticles after intravenous (i.v.) injection, we further designed an in vivo RT study on tumor-bearing mice. It was observed that tumor growth in the mice that were injected with Au@MnS@ZnS-PEG was remarkably inhibited by X-rays radiation compared with the mice treated with RT alone. Importantly, no


3


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 4 of 25

appreciable toxicity to mice was detected using blood chemistry analysis and careful histological examination after i.v. injection of Au@MnS@ZnS-PEG at a rather high dose of 150 mg/kg body weight. Therefore, our newly developed Au@MnS@ZnS-PEG nanoparticles could serve as a novel theranostic agent for the imaging-guided RT treatment with enhanced efficacy.

2. Experimental Section 2.1 Synthesis of Au@MnS@ZnS nanoparticles. The Au@MnS@ZnS nanoparticles were prepared via a modified high-temperature chemical synthesis method. Briefly, 100 mg of manganese(II) acetylacetonate (Mn(acac)2) was dissolved in 5 mL of oleylamine in 50 ml threenecked flask that was degassed at 120 °C for 30 min. Meanwhile, 8 mg of HAuCl4•3H2O was dissolved in 5 mL of oleylamine and stirred for 10 min at 120 °C to prepare the Au precursors. Next, the manganese precursor was quickly injected into the solution of Au precursors. Meanwhile, the mixture of 1-dodecanethiol (1-DDT, 55 µL) and tert-Dodecanethiol (t-DDT, 375 µL) was immediately injected into the reacted solution. The reaction temperature was maintained at 280 °C for 1 h to form the Au@MnS core-shell nanoparticles, and then cooled down to 150 °C. On the other hand, 150 mg of Zinc stearate was dissolved in 5 mL oleylamine, heated to 150 °C for 30 min and, then, swiftly injected into the Au@MnS solution for 10 min. Next, the sulfur precursor (50 µL of hexamethyldisilathiane in 5 mL 1-octadecene) was injected into the reaction solution for 10 min. The reaction temperature was maintained at 220°C for 30 min to form ZnScoated Au@MnS nanoparticles. Finally, the reaction was cooled to room temperature. The assynthesized Au@MnS@ZnS nanoparticles were washed three times with a mixture of ethanol and chloroform using centrifugation for 10 min at 8000 rpm. The product was re-dissolved in chloroform and stored at 4°C for next experiments.


4

Page 5 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


For surface coating of Au@MnS@ZnS nanoparticles, PEG-grafted poly (maleic anhydridealt-1-octadecene) (C18PMH-PEG) was synthesized according to our previous protocol23-24. In total, 20 mg of C18PMH-PEG was added to 1 mL of Au@MnS@ZnS nanoparticle solution (1 mg/mL in chloroform) under ultrasonication for 30 min. Then, the solution was stirred for 6 h at room temperature. We dried the solvent by nitrogen and re-dispersed the PEGylated Au@MnS@ZnS nanoparticles (Au@MnS@ZnS-PEG) in water.

2.2. Cell experiments. 4T1 murine breast cancer cells were cultured in a standard cell media recommended by the American type culture collection (ATCC). The standard cell count kit (CCK-8) assay was used to measure the cytotoxicity of Au@MnS@ZnS-PEG nanoparticles. For in vitro-enhanced radiotherapy based on Au@MnS@ZnS-PEG nanoparticles, clonogenic assays and γ-H2AX immunofluorescence were performed. For clonogenic assays, different numbers of 4T1 cancer cells (100, 200, 300, 600 and 1800) were incubated with or without Au@MnS@ZnSPEG nanoparticles (34 µg/mL) for 24 h and, then, irradiated with X-rays at the dose of 0, 2, 4, 6 and 8 Gray (Gy) for clonogenic assays. After that, the cells were cultured for 7-10 days to form colonies. During the colony growth, the cell culture medium was replaced every 3 days. The colonies were stained with Giemsa and were counted only if they contained more than 50 cells. The colony formation rate was calculated using the equation: colony formation rate = (number of colonies / number of seed cells) ×100%. Each treatment was performed in triplicate. For γ-H2AX immunofluorescence analysis, cells seeded in 6 plates were incubated with Au@MnS@ZnS-PEG nanoparticles at a concentration of 34 µg/mL for 6 h. The cells were irradiated with X-rays at a dose of 4 Gy. After additional culturing for 24 h, the cells were collected and fixed by 4% paraformaldehyde for 30 min. Then, the cells were rinsed twice in a


5


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 6 of 25

pure PBS buffer and permeabilized with absolute methanol for 15 min at -20 °C. Then, cells were rinsed in PBS buffer and blocked with a blocking buffer (1% BSA in PBS solution) for 1 h at room temperature. Next, the cells were incubated with mouse monoclonal anti-phosphohistone γ-H2AX primary antibody (1:500 in PBS containing 1% BSA) overnight at 4 °C. After being washed with PBS again, the cells were incubated with sheep anti-mouse Cy633 secondary antibody (1:500 in PBS containing 1% BSA) for 1 h at room temperature. The excess of dyelabeled antibody was removed by rinsing cover slips in PBS buffer. Cell nuclei were stained with DAPI for 5 min at room temperature. Cells were observed and imaged using confocal microscopy (PerkinElmer). 2.3. Blood circulation and biodistribution study. Healthy BALB/c mice were intravenously injected with Au@MnS@ZnS-PEG nanoparticles at a dose of 50 mg/kg per mouse. For blood circulation measurement, blood samples (5-10 µL) were collected from tail veins of mice at different time points (5 min, 30 min, 1 h, 2 h, 6 h, 10 h and 24 h). Then, the level of Au3+ in blood samples was measured using Inductively Coupled Plasma-Atomic Emission Spectrometry (ICP-AES). To evaluate the biodistribution and tumor uptake of Au@MnS@ZnS-PEG nanoparticles, mice bearing 4T1 tumors were intravenously injected with Au@MnS@ZnS-PEG nanoparticles at a dose of 50 mg/kg per mouse and sacrificed at 24 h post injection. The main organs (i.e., liver, spleen, kidney, lung, heart, skin, muscle, bone, brain and tumor) were collected, weighted and solubilized using aqua regia for the ICP-AES measurement.

2.4. In vivo MR imaging. Mice bearing 4T1 tumors were intravenously injected with PEGylated Au@MnS@ZnS nanoparticles at a manganese concentration of 4 mg/mL (200 µl)


6

Page 7 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


and imaged using a 3.0 T-animal MRI scanner (Bruker Biospin Corporation, Billerica, MA, USA) at different time points including 0, 0.5, 2, 24 h post injection. 2.5. In vivo enhanced radiotherapy. In total, 30 BALB/C mice bearing 4T1 tumors were randomly divided into 6 groups (n=5 per group), including the untreated group, only X-ray irradiated group, Au@MnS@ZnS-PEG injected but without X-ray irradiation group, and Au@MnS@ZnS-PEG injected and X-ray irradiated group. The dose of Au@MnS@ZnS-PEG nanoparticles was 150 mg/kg, and the X-ray doses were 4 Gy and 6 Gy. The tumor volume was measured every other day. 3. Results and Discussion In our study, Au@MnS@ZnS nanoparticles were synthesized using the high-temperature chemical reaction. Briefly, monodisperse gold nanoparticles (Au) were first prepared using a solvent thermal method and then coated with manganese sulfide (MnS) nanoshells using the high-temperature reaction. Au@MnS nanoparticles were prepared and characterized. Transmission electron microscopy (TEM) showed the core-shell structure of Au@MnS nanoparticles (Supporting information Figure S1a). The powder X-ray diffraction (XRD) of Au@MnS nanoparticles showed that the diffraction peaks appeared at 25.75, 27.55, 29.19, 38.12, 45.38, 49.94, and 53.39 degree, which were attributed to the (100), (002), (101), (102), (110), (103), and (112) planes of the MnS shell (Supporting information Figure S1b). The obtained Au@MnS core-shell nanoparticles, although stable in organic solvent, appeared to be unstable in the presence of water and changed their color from blue to red after being transferred to aqueous solution. This is likely attributed to the dissolving and oxidization of the MnS nanoshell that induced the leakage of gold nanoparticles in the aqueous environment (Supporting information Figure S2, S3 and S9). Therefore, to improve the chemical stability of nanoparticles


7


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 8 of 25

and protect the MnS shell from dissolving and oxidizing, an outer shell of zinc sulfide (ZnS) was induced to coat the Au@MnS nanoparticles and to form the Au@MnS@ZnS core/shell/shell nanoparticles. Although Au@MnS@ZnS nanoparticles contain a multi-layered structure, they can be prepared in one pot by subsequently adding different reagents at desired time points without the need for tedious purification at each growth step. The obtained Au@MnS@ZnS nanoparticles were then carefully characterized. Transmission electron microscopy (TEM) images clearly revealed that the Au core was coated with a MnS@ZnS shell. From the magnified image, we determined that the Au core, MnS shell and ZnS shell were clearly distinguishable, which further confirmed the core/shell/shell structure for our Au@MnS@ZnS nanoparticles (Figure 1b, Supporting information Figure S4 and S5). Energy-dispersive X-ray spectroscopy (EDX) elemental mapping of Au@MnS@ZnS nanoparticles was conducted and showed that three elements (Au, Mn and Zn) were identified in all nanoparticles. This further confirmed that the Au@MnS@ZnS core/shell/shell nanostructure existed in the final product (Figure 1c). The powder X-ray diffraction (XRD) pattern of Au@MnS@ZnS (Au/MZS) core/shell/shell nanoparticles was shown in Figure 2a. The peak at 38.20 degrees can be assigned to the (111) lattice plane of Au nanoparticle (JCPDS 04–0784). Additionally, we observed clear MnS peaks in the XRD spectrum of our final sample. However, the ZnS peaks were not clearly observed, which likely was attributed to the fact that the ZnS shell on those nanoparticles was too thin to be detected using XRD (Supporting information Figure S6). No secondary phases or impurity peaks were detected. To transfer the as-made nanoparticles into aqueous solutions, a PEG-grafted poly(maleic anhydride-alt-1-octadecene) (C18PMH-PEG) amphiphilic polymer was synthesized according to our previously reported procedure23,

25

. Then, it was used to functionalize the


8

Page 9 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


excellent stability in physiological solutions (Supporting information Figure S7). To measure the stability and dispersibility after PEGylation, the hydrodynamic diameter of Au@MnS@ZnSPEG in water and phosphate buffer solution (PBS) was recorded for one week. No obvious change was observed following the dynamic light scattering (DLS) measurement, which revealed significant stability and uniform size distribution of Au@MnS@ZnS-PEG (Figure 2b). The hydrodynamic size of Au@MnS@ZnS-PEG measured using DLS was ~110 nm, which was bigger than that observed using TEM imaging (approximately 30-40 nm). We thought that it could have been the condensed surface polymer coating that resulted in the increased hydrodynamic size of Au@MnS@ZnS-PEG nanoparticles. To test the serum stability of Au@MnS@ZnS-PEG, we incubated the Au@MnS@ZnS nanoparticles in mouse plasma at 37 °C for 24 h, and found that hardly any Au or Mn leaked from the nanoparticles (Supporting information, Figure S8). UV-vis-NIR spectra of Au@MnS@ZnS-PEG nanoparticles exhibited a typical absorbance peak at 618 nm due gold nanoparticles as their cores (Figure 2c). The red shifting of plasmon band of gold nanoparticles after shell coating was attributed to the high refractive index of the MnS shell. In recent years, manganese-based nanomaterials as T1-weighted contrast agents have been extensively studied for in vivo MR imaging26-31. In this work, T1-weighted magnetic resonance (MR) imaging of Au@MnS@ZnS-PEG solution with different concentrations acquired using a 3.0 T MR scanner showed the concentration-dependent whitening effect. The T1 relativity of Au@MnS@ZnS-PEG was determined to be 7.86 mM-1S-1 (Figure 2d), which suggested that Au@MnS@ZnS-PEG was expected to be an excellent MR imaging contrast agent. Although the paramagnetic MnS is encapsulated inside the ZnS shell, it still can induce the effective shortening of T1 relaxation of the surrounding water protons, which is similar to many other


9


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 10 of 25

magnetic nanoparticles shelled by the SiO2 or Bi2Se3 coatings22, 32. The ZnS shell was very thin and could be somewhat porous. Therefore, the T1-contrast of the nanoparticles remained strong. To measure the cytotoxicity of Au@MnS@ZnS-PEG nanoparticles, the standard cell count kit (CCK-8) assay was used to determine relative cell viabilities after being incubated with different concentrations of Au@MnS@ZnS-PEG nanoparticles for 24 h. It was found that Au@MnS@ZnS-PEG exhibited no obvious toxicity to cells even at a high concentration (Figure 3a). To determine the efficacy of radio-sensitization of Au@MnS@ZnS-PEG nanoparticles, 4T1 cells that were incubated with Au@MnS@ZnS-PEG at a concentration of 34 µg/ml for 12 h were irradiated with X-rays at different doses (0, 2, 4, 6 and 8 Grey). A clonogenic assay was used to determine the resulting impact on the cancer cell growth by measuring the capacity of single tumor cells to grow into cell colonies after various treatments. It was found that 4T1 cells that were incubated with Au@MnS@ZnS-PEG nanoparticles exhibited a clear decrease of survival fractions under X-ray irradiation compared with the control group after being treated by the same doses of X-rays. The sensitizing enhancement ratio of Au@MnS@ZnS-PEG nanoparticles was estimated to be 1.22 (Figure 3 b &c), which was consistent with previous studies using gold nanoparticles as radiosensitizers12, 33-35 To understand the radio-sensitizing mechanism, the double-strand breaks (DSBs) of DNA inside tumor cells were recorded using γ-H2AX staining to evaluate the effect of enhanced DNA damage during radiotherapy7-8,

36-38

. Immunofluorescent imaging of 4T1 cells showed that

minimal amount of DSBs of DNA (red fluorescent spots) was observed in cells without X-ray irradiation, regardless of the presence of Au@MnS@ZnS-PEG nanoparticles (Figure 3d). Additionally, the DSBs of DNA induced by RT alone was not evident at our tested condition (4 Gy). In contrast, high levels of DSBs of DNA were clearly observed within the nuclei of cells


10

Page 11 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


that received X-ray irradiation in the presence of Au@MnS@ZnS-PEG nanoparticles, which suggested severe DNA damage in those cells (Figure 3d). Therefore, our synthesized Au@MnS@ZnS-PEG nanoparticles could act as a promising radio-sensitizing agent for enhanced radiotherapy. We then studied in vivo MR imaging guided enhanced radiotherapy using Au@MnS@ZnSPEG as a therapeutic agent. Mice bearing 4T1 tumors were intravenously (i.v.) injected with 200 µL of Au@MnS@ZnS-PEG (with a manganese concentration of 4 mg/mL) and then imaged using a 3.0-T animal MRI scanner at different time points post injection (p.i.). The MR imaging of mice uncovered high tumor uptake and a clear whitening effects in tumors. Additionally, the T1 MR signals increased by 280% in tumor sites at 24 h p.i. (Figure 4a&b). In order to further investigate

the

in

vivo

behavior

including

blood

circulation

and

biodistribution,

Au@MnS@ZnS-PEG nanoparticles were i.v. injected into healthy BALB/c mice (50 mg/kg). Blood samples were drawn from tail veins of mice at different time points p.i. and examined by measuring the Au3+ levels using ICP-AES. The Au3+ levels in blood decayed in accordance with a two-compartment model, with first ( t1/2 ) and second ( t'1/2 ) phases of circulation half-lives calculated to be 0.142 ± 0.058 h and 21.715 ± 3.53 h, respectively (Figure 4c). For biodistribution analysis, various organs collected from mice treated with Au@MnS@ZnS-PEG at 24 h p.i were tested using ICP-AES based on the Au3+ levels. It was found that Au@MnS@ZnS-PEG could passively accumulate in the tumor (~12 %ID/g), likely owing to long blood circulation half-lives of Au@MnS@ZnS-PEG and the enhanced penetration and retention (EPR) effect of cancerous tumors (Figure 4d). Besides, the reticuloendothelial system (RES) including liver and spleen showed high accumulation of Au@MnS@ZnS-PEG due to the macrophage uptake of nanoparticles (Figure 4d).


11


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 12 of 25

Considering the good MR imaging and biodistribution results, we performed an in vivo radiotherapy study using Au@MnS@ZnS-PEG as a radiosensitizer. BALB/c mice bearing 4T1 tumors were randomly divided into six groups with five mice per group and then differently treated with: 1) saline, 2) 4 Gy irradiation, 3) 6 Gy irradiation, 4) Au@MnS@ZnS-PEG only, 5) Au@MnS@ZnS-PEG + 4 Gy irradiation, and 6) Au@MnS@ZnS-PEG + 6 Gy irradiation. The tumor volumes measured with caliper were used to evaluate the treatment effect. It was found that the tumors on mice treated with Au@MnS@ZnS-PEG (150 mg/kg by i.v. injection) under irradiation at the doses of 4 Gy and 6 Gy were significantly inhibited with clearly observed decreased growth rates (Figure 5 a&c). However, under RT alone with 4 Gy or 6 Gy of X-ray irradiation, the tumor growth was only partially delayed at the early stage post treatment and regained the rapid growth 8 days after the treatment (Figure 5a). Furthermore, the tumors in all control groups without X-ray irradiation showed similar growth rates, suggesting that the Au@MnS@ZnS-PEG injection by itself would not affect tumor development (Figure 5 a&c). In addition, we monitored body weight changes of mice with different treatments and found no significant body weight change during this period of observation (Figure 5b). Therefore, our Au@MnS@ZnS-PEG nanoparticles could be expected to be an excellent radio-sensitizer for in vivo radiotherapy treatment of cancer. The potential in vivo toxicity is still a significant problem for the application of nanomaterials in biomedicine. In our work, the potential in vivo toxicity of Au@MnS@ZnS-PEG was systematically investigated via blood analysis and histological examination. Healthy BALB/c mice that were i.v. injected with Au@MnS@ZnS-PEG at a dose of 150 mg/kg were sacrificed at 1, 7 and 20 days post injection for blood collection. Blood from the age matched control untreated mice was collected 20 days p.i. (5 mice per group). Serum biochemical parameters


12

Page 13 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


including liver function markers (alanine aminotransferase (ALT), alkaline phosphatase (ALP) and aspartate aminotransferase (AST)), kidney function marker urea nitrogen (BUN) and albumin/globin ration were investigated and found that all tested markers were within normal ranges (Figure 6 a-d). We also investigated the blood routine including white blood cells, red blood cells, hemoglobin, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelet count and mean corpuscular hemoglobin. Compared with the control group, all parameters were normal and were within the normal range (Figure 6 e–l). Meanwhile, the major organs from the abovementioned treated mice were collected for Hematoxylin & Eosin (H&E) staining. Compared to the control group (Figure 6 m), neither visible organ damage nor inflammation from the treated mice was observed. Therefore, our Au@MnS@ZnS-PEG nanoparticles induced no significant side effect on the treated mice. 4. Conclusion In summary, we designed a novel radio-sensitizer for the in vivo MR imaging guided enhanced radiotherapy of tumors. With a biocompatible PEG coating, the obtained Au@MnS@ZnS-PEG nanoparticles exhibited significant stability in physiological solutions and long blood circulation time. Utilizing the magnetic property of the MnS nano-shell, T1 weighed MR imaging was performed. As a result of the EPR effect, a high passive tumor accumulation of our nanoparticles was shown, and the desired MR imaging enhanced effect was achieved. Furthermore, using the high atomic number of gold, we used Au@MnS@ZnS-PEG nanoparticles as a radio-sensitizer for enhanced RT treatment and achieved significant tumor inhibition using X-ray irradiation at 6 Gy. No obvious toxicity or side effects of our Au@MnS@ZnS-PEG nanoparticles was observed at our tested doses. Therefore, the combination of RT with other treatments, such as chemotherapy or photothermal therapy, based


13


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 14 of 25

on Au@MnS@ZnS-PEG may offer further opportunities in future studies. Our work highlights the potential of multifunctional nanomaterials for external RT cancer treatment under imaging guidance.


14

Page 15 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 1. Preparation and characterization of Au@MnS@ZnS core@shell@shell nanoparticles. (a) Schematic illustration of the design of Au@MnS@ZnS nanoparticles. (b) TEM image of Au@MnS@ZnS nanoparticles. (c) Elemental mapping (Au, Mn, and Zn) of a Au@MnS@ZnS nanoparticle.


15


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 16 of 25

Figure 2. (a) X-ray powder diffraction pattern of Au@MnS@ZnS nanoparticles. (b) Dynamic light scattering (DLS) data of Au@MnS@ZnS-PEG in water and PBS solution. (c) UV-vis-NIR spectra of Au@MnS@ZnS-PEG nanoparticles. Inset: photo of a Au@MnS@ZnS-PEG solution. (d) T1 relaxation rates (R1) of Au@MnS@ZnS-PEG solutions at different manganese concentrations. Inset: T1-weighted MR images of Au@MnS@ZnS-PEG solutions.


16

Page 17 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 3. In vitro experiments. (a) Relative viabilities of 4T1 cells incubated with different concentrations of Au@MnS@ZnS-PEG for 24 h. (b) Surviving fractions of 4T1 cells treated with (black) or without (red) Au@MnS@ZnS-PEG at radiation doses of 0, 2, 4, 6 and 8 Gy. (c) Growth of 4T1 cancer cell colonies following 7 days after 4 Gy of X-ray irradiation. (d) Immunofluorescent imaging of γ-H2AX foci in 4T1 cells incubated with or without Au@MnS@ZnS-PEG under X-ray irradiation. The results showed that Au@MnS@ZnS-PEG could act as a powerful theranostic agent for enhanced radiotherapy of cancer cells.


17


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 18 of 25

Figure 4. In vivo experiments. (a) MR images of mice i.v. injected with Au@MnS@ZnS-PEG recorded at different time points post-injection (Red arrows point to the tumor sites). (b) Relative T1 signal intensities of the tumor on mice recorded at different time points post Au@MnS@ZnS-PEG injection. (c) Blood circulation of Au@MnS@ZnS-PEG by measuring Au3+ levels at different time points p.i. (d) Biodistribution of Au@MnS@ZnS-PEG by measuring the Au 3+ levels in tumors and other major organs at 24 h p.i. All data showed that Au@MnS@ZnS-PEG could passively accumulate in the tumor after intravenous injection.


18

Page 19 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 5. In vivo radiotherapy. (a&b) Tumor growth curves (a) and body weight curves (b) of mice after different treatments indicated for 4T1 tumor-bearing mice. c) Representative photos of tumor-bearing mice after different treatment taken at 0, 10 and 20 days post RT treatment. Six groups (5 mice for each group) were used in this experiment including the untreated mice (Control), X-ray treated mice without Au@MnS@ZnS-PEG injection (4 Gy and 6 Gy), Au@MnS@ZnS-PEG injected mice without X-ray irradiation, and Au@MnS@ZnS-PEG injected mice with X-ray irradiation at 4 Gy and 6 Gy.


19


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 20 of 25

Figure 6. In vivo toxicity investigation. (a-l) Blood biochemistry (a-d) and routine blood analysis (e-l) from mice treated with or without Au@MnS@ZnS-PEG (dose = 150 mg/kg) at 0, 7 and 20 day after i.v. injection. (m) H&E-stained tissue sections of major organs including heart, liver spleen, lung and kidney from the untreated healthy mice and the mice injected with Au@MnS@ZnS-PEG at 20 days p.i. All blood chemistry and hematological data were within the normal range. The statistics was based on 5 mice per data-point.


20

Page 21 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


ASSOCIATED CONTENT Supporting Information The photos of Au@MnS@ZnS nanoparticles before and after PEGylation in different physiological solution. This material is available free of charge via the Internet at http://pubs.acs.org.

AUTHOR INFORMATION Corresponding Authors *Tel.: 86-512-65882925. E-mail: [email protected]. *Tel.: 86-512-65882036. E-mail: [email protected].

Notes The authors declare no competing financial interests. ACKNOWLEDGMENTS This work was partially supported by the National Natural Science Foundation of China (81471716, 81302383, 31400861), a Juangsu Natural Science Fund for Distinguished Young Scholars (BK20140320), and a Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).


21


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 22 of 25

REFERENCES (1) Jemal, A.; Bray, F.; Center, M. M.; Ferlay, J.; Ward, E.; Forman, D., Global Cancer Statistics. CA. Cancer J. Clin. 2011, 61, 69-90. (2) Pearson, J. G., The Present Status and Future Potential of Radiotherapy in the Management of Esophageal Cancer. Cancer 1977, 39, 882-890. (3) Su, X. Y.; Liu, P. D.; Wu, H.; Gu, N., Enhancement of Radiosensitization by Metal-based Nanoparticles in Cancer Radiation Therapy. Cancer Biol. Med. 2014, 11, 86-91. (4) Xing, H.; Zheng, X.; Ren, Q.; Bu, W.; Ge, W.; Xiao, Q.; Zhang, S.; Wei, C.; Qu, H.; Wang, Z.; Hua, Y.; Zhou, L.; Peng, W.; Zhao, K.; Shi, J., Computed Tomography Imaging-Guided Radiotherapy by Targeting Upconversion Nanocubes with Significant Imaging and Radiosensitization Enhancements. Sci. Rep. 2013,3, 0175 (5) Khanna, K. K.; Jackson, S. P., DNA Double-Strand Breaks: Signaling, Repair and the Cancer Connection. Nat. Genet. 2001, 27, 247-254. (6) Wang, H.; Zeng, Z. C.; Bui, T. A.; DiBiase, S. J.; Qin, W.; Xia, F.; Powell, S. N.; Iliakis, G., Nonhomologous End-Joining of Ionizing Radiation-Induced DNA Double-Stranded Breaks in Human Tumor Cells Deficient in BRCA1 or BRCA2. Cancer Res. 2001, 61, 270-277. (7) Dizdaroglu, M.; Jaruga, P.; Birincioglu, M.; Rodriguez, H., Free Radical-Induced Damage to DNA: Mechanisms and Measurement. Free Radical Biol. Med. 2002, 32, 1102-1115. (8) Vignard, J.; Mirey, G.; Salles, B., Ionizing-Radiation Induced DNA Double-Strand Breaks: A Direct and Indirect Lighting Up. Radiother. Oncol. 2013, 108, 362-369. (9) Taleei, R.; Girard, P. M.; Nikjoo, H., DSB Repair Model for Mammalian Cells in Early S and G1 Phases of the Cell Cycle: Application to Damage Induced by Ionizing Radiation of Different Quality. Mutation Res. Genet. Toxicol. Environ. Mutagen. 2015, 779, 5-14. (10) Al Zaki, A.; Joh, D.; Cheng, Z.; De Barros, A. L.; Kao, G.; Dorsey, J.; Tsourkas, A., GoldLoaded Polymeric Micelles for Computed Tomography-Guided Radiation Therapy Treatment and Radiosensitization. ACS Nano 2014, 8, 104-112. (11) Ngwa, W.; Kumar, R.; Sridhar, S.; Korideck, H.; Zygmanski, P.; Cormack, R. A.; Berbeco, R.; Makrigiorgos, G. M., Targeted Radiotherapy with Gold Nanoparticles: Current Status and Future Perspectives. Nanomedicine 2014, 9, 1063-1082. (12) Chithrani, D. B.; Jelveh, S.; Jalali, F.; van Prooijen, M.; Allen, C.; Bristow, R. G.; Hill, R. P.; Jaffray, D. A., Gold Nanoparticles as Radiation Sensitizers in Cancer Therapy. Radiat. Res. 2010, 173, 719-728. (13) McQuade, C.; Al Zaki, A.; Desai, Y.; Vido, M.; Sakhuja, T.; Cheng, Z.; Hickey, R. J.; Joh, D.; Park, S. J.; Kao, G.; Dorsey, J. F.; Tsourkas, A., A Multifunctional Nanoplatform for Imaging, Radiotherapy, and the Prediction of Therapeutic Response. Small 2015, 11, 834843. (14) Yang, Y. S.; Carney, R. P.; Stellacci, F.; Irvine, D. J., Enhancing Radiotherapy by Lipid Nanocapsule-Mediated Delivery of Amphiphilic Gold Nanoparticles to Intracellular Membranes. ACS Nano 2014, 8, 8992-9002. (15) Zhang, X. D.; Luo, Z.; Chen, J.; Shen, X.; Song, S.; Sun, Y.; Fan, S.; Fan, F.; Leong, D. T.; Xie, J., Ultrasmall Au(10-12)(SG)(10-12) Nanomolecules for High Tumor Specificity and Cancer Radiotherapy. Adv. Mater. 2014, 26, 4565–4568 (16) Zhang, X. D.; Wu, D.; Shen, X.; Chen, J.; Sun, Y. M.; Liu, P. X.; Liang, X. J., SizeDependent Radiosensitization of PEG-Coated Gold Nanoparticles for Cancer Radiation Therapy. Biomaterials 2012, 33, 6408-6419.


22

Page 23 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(17) Ma, M.; Huang, Y.; Chen, H.; Jia, X.; Wang, S.; Wang, Z.; Shi, J., Bi2S3-Embedded Mesoporous Silica Nanoparticles for Efficient Drug Delivery and Interstitial Radiotherapy Sensitization. Biomaterials 2015, 37, 447-455. (18) Zhang, X.-D.; Chen, J.; Min, Y.; Park, G. B.; Shen, X.; Song, S.-S.; Sun, Y.-M.; Wang, H.; Long, W.; Xie, J.; Gao, K.; Zhang, L.; Fan, S.; Fan, F.; Jeong, U., Metabolizable Bi2Se3 Nanoplates: Biodistribution, Toxicity, and Uses for Cancer Radiation Therapy and Imaging. Adv. Funct. Mater. 2014, 24, 1718-1729. (19) Yao, M. H.; Ma, M.; Chen, Y.; Jia, X. Q.; Xu, G.; Xu, H. X.; Chen, H. R.; Wu, R., Multifunctional Bi2S3/PLGA Nanocapsule for Combined HIFU/Radiation Therapy. Biomaterials 2014, 35, 8197-8205. (20) Chiu, S. J.; Lee, M. Y.; Chou, W. G.; Lin, L. Y., Germanium Oxide Enhances the Radiosensitivity of Cells. Radiat. Res. 2003, 159, 391-400. (21) Lin, M. H.; Hsu, T. S.; Yang, P. M.; Tsai, M. Y.; Perng, T. P.; Lin, L. Y., Comparison of Organic and Inorganic Germanium Compounds in Cellular Radiosensitivity and Preparation of Germanium Nanoparticles as a Radiosensitizer. Int. J. Radiat. Biol. 2009, 85, 214-226. (22) Song, G.; Liang, C.; Gong, H.; Li, M.; Zheng, X.; Cheng, L.; Yang, K.; Jiang, X.; Liu, Z., Core-Shell MnSe@Bi2Se3 Fabricated via a Cation Exchange Method as Novel Nanotheranostics for Multimodal Imaging and Synergistic Thermoradiotherapy. Adv. Mater. 2015, 27, 6110–6117. (23) Yang, K.; Wan, J.; Zhang, S.; Tian, B.; Zhang, Y.; Liu, Z., The Influence of Surface Chemistry and Size of Nanoscale Graphene Oxide on Photothermal Therapy of Cancer Using Ultra-Low Laser Power. Biomaterials 2012, 33, 2206-2214. (24) Liu, X.; Tao, H.; Yang, K.; Zhang, S.; Lee, S. T.; Liu, Z., Optimization of Surface Chemistry on Single-Walled Carbon Nanotubes for In Vivo Photothermal Ablation of Tumors. Biomaterials 2011, 32, 144-151. (25) Prencipe, G.; Tabakman, S. M.; Welsher, K.; Liu, Z.; Goodwin, A. P.; Zhang, L.; Henry, J.; Dai, H., PEG Branched Polymer for Functionalization of Nanomaterials with Ultralong Blood Circulation. J. Am. Chem. Soc. 2009, 131, 4783-4787. (26) Chen, Y.; Yin, Q.; Ji, X.; Zhang, S.; Chen, H.; Zheng, Y.; Sun, Y.; Qu, H.; Wang, Z.; Li, Y.; Wang, X.; Zhang, K.; Zhang, L.; Shi, J., Manganese Oxide-Based Multifunctionalized Mesoporous Silica Nanoparticles for pH-Responsive MRI, Ultrasonography and Circumvention of MDR in Cancer Cells. Biomaterials 2012, 33, 7126-7137. (27) Kim, T.; Momin, E.; Choi, J.; Yuan, K.; Zaidi, H.; Kim, J.; Park, M.; Lee, N.; McMahon, M. T.; Quinones-Hinojosa, A.; Bulte, J. W.; Hyeon, T.; Gilad, A. A., Mesoporous SilicaCoated Hollow Manganese Oxide Nanoparticles as Positive T1 Contrast Agents for Labeling and MRI Tracking of Adipose-Derived Mesenchymal Stem Cells. J. Am. Chem. Soc. 2011, 133, 2955-2961. (28) Yang, H.; Zhang, C.; Shi, X.; Hu, H.; Du, X.; Fang, Y.; Ma, Y.; Wu, H.; Yang, S., WaterSoluble Superparamagnetic Manganese Ferrite Nanoparticles for Magnetic Resonance Imaging. Biomaterials 2010, 31, 3667-3673. (29) Tromsdorf, U. I.; Bigall, N. C.; Kaul, M. G.; Bruns, O. T.; Nikolic, M. S.; Mollwitz, B.; Sperling, R. A.; Reimer, R.; Hohenberg, H.; Parak, W. J.; Forster, S.; Beisiegel, U.; Adam, G.; Weller, H., Size and Surface Effects on the MRI Relaxivity of Manganese Ferrite Nanoparticle Contrast Agents. Nano Lett. 2007, 7, 2422-2427.


23


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 24 of 25

(30) Huang, Y.; Hu, L.; Zhang, T.; Zhong, H.; Zhou, J.; Liu, Z.; Wang, H.; Guo, Z.; Chen, Q., Mn(3)[Co(CN)(6)](2)@SiO(2) Core-Shell Nanocubes: Novel Bimodal Contrast Agents for MRI and Optical Imaging. Sci. Rep. 2013, 3, 2647-2653. (31) Lu, J.; Ma, S.; Sun, J.; Xia, C.; Liu, C.; Wang, Z.; Zhao, X.; Gao, F.; Gong, Q.; Song, B.; Shuai, X.; Ai, H.; Gu, Z., Manganese Ferrite Nanoparticle Micellar Nanocomposites as MRI Contrast Agent for Liver Imaging. Biomaterials 2009, 30, 2919-2928. (32) Schladt, T. D.; Koll, K.; Prüfer, S.; Bauer, H.; Natalio, F.; Dumele, O.; Raidoo, R.; Weber, S.; Wolfrum, U.; Schreiber, L. M.; Radsak, M. P.; Schild, H.; Tremel, W., Multifunctional Superparamagnetic MnO@SiO2 Core/Shell Nanoparticles and Their Application for Optical and Magnetic Resonance Imaging. J. Mater. Chem. 2012, 22, 9253-9262. (33) Hainfeld, J. F.; Smilowitz, H. M.; O'Connor, M. J.; Dilmanian, F. A.; Slatkin, D. N., Gold Nanoparticle Imaging and Radiotherapy of Brain Tumors in Mice. Nanomedicine 2013, 8, 1601-1609. (34) Hainfeld, J. F.; Dilmanian, F. A.; Slatkin, D. N.; Smilowitz, H. M., Radiotherapy Enhancement with Gold Nanoparticles. J. Pharm. Pharmacol. 2008, 60, 977-985. (35) Hainfeld, J. F.; Slatkin, D. N.; Smilowitz, H. M., The Use of Gold Nanoparticles to Enhance Radiotherapy in Mice. Phys. Med. Biol. 2004, 49, N309-N315. (36) Lomax, M. E.; Folkes, L. K.; O'Neill, P., Biological Consequences of Radiation-Induced DNA Damage: Relevance to Radiotherapy. Clin. Oncol. 2013, 25, 578-585. (37) Hei, T. K.; Zhou, H.; Chai, Y.; Ponnaiya, B.; Ivanov, V. N., Radiation Induced NonTargeted Response: Mechanism and Potential Clinical Implications. Curr. Mol. Pharmacol. 2011, 4, 96-105. (38) Huang, Y. Y.; Luo, Y.; Zheng, W. J.; Chen, T. F., Rational Design of Cancer-Targeted BSA Protein Nanoparticles as Radiosensitizer to Overcome Cancer Radioresistance. ACS Appl. Mater. Interfaces. 2014, 6, 19217-19228.


24

Page 25 of 25

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


For Table of Contents only


25

Shell Nanoparticles for ... - ACS Publications

Recommend Documents