Controllable Synthesis of a Smart Multifunctional Nanoscale Metal

Research Article www.acsami.org

Controllable Synthesis of a Smart Multifunctional Nanoscale Metal−Organic Framework for Magnetic Resonance/Optical Imaging and Targeted Drug Delivery Xuechuan Gao,† Manjue Zhai,‡ Weihua Guan,† Jingjuan Liu,† Zhiliang Liu,*,† and Alatangaole Damirin*,‡ †

College of Chemistry and Chemical Engineering, Inner Mongolia University, Hohhot 010021, P. R. China College of Life Sciences, Inner Mongolia University, Hohhot 010021, P. R. China

Downloaded via KAOHSIUNG MEDICAL UNIV on June 19, 2018 at 13:26:40 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

‡

S Supporting Information *

ABSTRACT: As a result of their extraordinarily large surfaces and welldefined pores, the design of a multifunctional metal−organic framework (MOF) is crucial for drug delivery but has rarely been reported. In this paper, a novel drug delivery system (DDS) based on nanoscale MOF was developed for use in cancer diagnosis and therapy. This MOF-based tumor targeting DDS was fabricated by a simple postsynthetic surface modification process. First, magnetic mesoporous nanomaterial Fe-MIL-53-NH2 was used for encapsulating the drug and served as a magnetic resonance contrast agent. Moreover, the Fe-MIL-53-NH2 nanomaterial exhibited a high loading capacity for the model anticancer drug 5-fluorouracil (5-FU). Subsequently, the fluorescence imaging agent 5-carboxyfluorescein (5-FAM) and the targeting reagent folic acid (FA) were conjugated to the 5-FU-loaded Fe-MIL-53-NH2, resulting in the advanced DDS Fe-MIL-53-NH2-FA5-FAM/5-FU. Owing to the multifunctional surface modification, the obtained DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU shows good biocompatibility, tumor enhanced cellular uptake, strong cancer cell growth inhibitory effect, excellent fluorescence imaging, and outstanding magnetic resonance imaging capability. Taken together, this study integrates diagnostic and treatment aspects into a single platform by a simple and efficient strategy, aiming for facilitating new possibilities for MOF use for multifunctional drug delivery. KEYWORDS: nanoscale MOFs, fluorescence imaging, magnetic resonance imaging, targeted drug delivery, multifunctional MOFs been reported often,12−16 the preparation methods are usually complex, and few attempts have been made to incorporate three functionalities: magnetic/fluorescence imaging, cell-targeting, and drug storage delivery in one single platform based on MOF. Considering the above-mentioned disadvantages, designing a targeted drug delivery system (DDS) based on MOF to reduce the secondary action of drugs during clinical treatment and integrating multimodal imaging capabilities is a long-standing challenge in drug development. It is generally known that iron is not only a widespread element in nature but also an essential element in the human body, which makes it extremely appealing for the application of Fe-MOFs-NH2 in diverse biomedical applications.17,18 With fascinating properties such as well-defined pores and low toxicity, nanoscale material Fe-MIL53-NH2 is an ideal candidate for drug carriers.19,20 Further, the Fe-MIL-53-NH2 materials already reported show intense magnetism, which enables them to be effective magnetic resonance imaging (MRI) contrast agents.21 Furthermore, the abundant

1. INTRODUCTION Different from most of the existing pure organic and inorganic carrier materials, metal−organic frameworks (MOFs) as a new class of porous crystalline materials have high stability, high surface area, large pore volume, regular porosity, and an intriguing variety of architectures1−4 and have already emerged as promising drug delivery platforms.5,6 In addition, the variable pore size of MOFs can be achieved by altering the metal or ligand, and the appeared active sites within the framework permit an easier transmission of guest molecules,7,8 offering unexpected advantages in the areas of drug delivery. Starting from Fe-based MOFs as drug delivery carriers demonstrated by Férey and co-workers,9 inspiring efforts have been committed to exploring these fields, and several studies have been carried out. Anionic Zn-based MOF10 was being used as a carrier of cationic drugs, and a biocompatible porous Mg-based MOF11 was being used as an antioxidant carrier. While these MOFs exhibit ideal drug loading and excellent drug delivery ability, the lack of tumor-targeting capability results in strong toxicity to normal cells, which is adverse to the drug efficacy and limits their biomedical applications. Although nanomaterials integrating magnetic or fluorescence imaging and drug delivery have © 2017 American Chemical Society

Received: November 17, 2016 Accepted: January 12, 2017 Published: January 12, 2017 3455

DOI: 10.1021/acsami.6b14795 ACS Appl. Mater. Interfaces 2017, 9, 3455−3462

Research Article

ACS Applied Materials & Interfaces amino groups on the external surface of Fe-MOFs-NH2 nanocrystals also offer the possibility to functionalize the Fe-based MOF for targeted imaging or therapy, which would have a great significance in further developments. Because the external carboxyl on 5-carboxylfluorescein (5-FAM, Figure S1a) can be covalently connected to the NH2 groups on the surface of Fe-MOFs-NH2 nanocrystals, 5-FAM is appropriate as a fluorescent probe for observing the drug delivery process and realtime imaging in cells.22 To reduce an anticancer drug’s toxicity to normal cells and enhance anticancer effects to cancer cells, targeted molecules should be combined with the drug carrier. As we know, folic acid (FA, Figure S1b) receptors are usually overexpressed on the outside surface of a number of cancer cells, and FA has excellent affinity with folate receptors (FR), leading to a selective uptake within FR-positive cancer cells and avoiding uptake by normal tissues that do not express FR.23,24 Thus, FA is widely chosen as a model targeting reagent to study the efficiency of delivering drugs into tumor cells.25,26 Additionally, the stability and compatibility in physiological conditions facilitate the employment of FA as an ideal targeting molecule for delivery systems. 5-Fluorouracil (5-FU), a pyrimidine analogue (Figure S1c), has been one of the major clinically applied anticancer agents for the treatment of a wide variety of tumors because it can be incorporated into DNA and RNA, leading to cytotoxicity and cancer cell death.27,28 Additionally, the small molecular weight of 5-FU boosts the burgeoning biomedical applications of 5-FU as a model anticancer drug. On the basis of the above background, a multifunctional MOF-based tumor targeting DDS (Fe-MIL-53-NH2-FA-5FAM/5-FU), shown in Figure 1, was developed in the present study. In this composite, Fe-MIL-53-NH2 is used for encapsulating the drug and magnetic resonance imaging, whereas FA serves as the targeted reagent, and 5-FAM is employed for fluorescent imaging. Fe-MIL-53-NH2-FA-5-FAM/5-FU, obtained via a simple postmodification method, displays satisfying magnetic performance and excellent green fluorescence emission. Meanwhile, Fe-MIL-53-NH2-FA-5-FAM/5-FU can target MGC-803 cells and exhibits significant cell growth inhibitory effects on MGC-803 cells due to the sustained release of 5-FU. All results demonstrate that the developed DDS Fe-MIL53-NH2-FA-5-FAM/5-FU possesses great potential as a novel theranostic agent for simultaneous fluorescence/magnetic resonance imaging and drug delivery in biomedicine. We envision that this new hybrid Fe-MIL-53-NH2-FA-5-FAM/ 5-FU nanocomposite will be proven to be a novel drug delivery platform for cancer disease therapy in the near future.

Figure 1. 2D plane view of Fe-MIL-53-NH2 nanocrystalline (A and B), schematic of DDS Fe-MIL-53-NH2−FA-5-FAM/5-FU (C), and schematic illustrations of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU for targeting drug delivery (D). of the samples was obtained under a N2 atmosphere and carried out using a NETZSCH STA449F3 thermal analyzer in the temperature range from 40 to 600 °C with a heating rate was 10 °C/min. A laser scanning confocal microscope (OLYMPUS, IX81) was used to confirm the cell targeting process and cell imaging capacity. UV−vis spectra of the samples were recorded on a TU-1901 diode UV−visible spectrophotometer. 2.2. Synthesis of Fe-MIL-53-NH2. Nanocrystalline Fe-MIL-53NH2 was prepared by a facile low-temperature synthesis route via the reaction of FeCl3·6H2O and 2-amino terephthalic acid (NH2-H2BDC) in the ethanol solution.29 Typically, 0.1 mmol FeCl3·6H2O, 0.1 mmol NH2-H2BDC, and 9 mL of ethanol solution were mixed into a threeneck round-bottom flask (25 mL), transferred into a 40 °C oil bath, and stirred for 2 h. The obtained reaction product was purified three times with ethanol and dried at 30 °C in a vacuum oven. To investigate the effect of reactant concentration on the crystal size, the amounts of FeCl3·6H2O and NH2-H2BDC were tuned from 0.1 to 0.4 mmol. 2.3. Preparation of 5-FU-Loaded Fe-MIL-53-NH2. A pure and dried sample of Fe-MIL-53-NH2 (0.1 g) was immersed in 50 mL of a water solution containing 5-FU (0.1 g) for 48 h. After being centrifuged, the obtained product was washed with water several times. The 5-FU loading efficiency on Fe-MIL-53-NH2 can be inferred from the formula as follows: loading efficiency (%) = (m1 − m2)/m. In this equation, m1 represents the initial weight of 5-FU; m2 refers to the weight of 5-FU present in the excess of solvent, and m refers to the total weight of the solid obtained. After 2 days of soaking, the loading efficiency of 5-FU on Fe-MIL-53-NH2 was 28% according to the standard 5-FU concentration curve shown in Figure S2. 2.4. Synthesis of Fe-MIL-53-NH2-FA-5-FAM/5-FU. At room temperature, 0.1 g of 5-FU-loaded Fe-MIL-53-NH2, 0.2 g of FA, and 0.2 g of 5-FAM were added to 50 mL of saturated aqueous solution of 5-FU. After the solution was mixed uniformly, 0.1 g of N-(3-(dimethylamino)propyl)-N-ethylcarbodiimide hydrochloride (EDC) as a

2. EXPERIMENTAL SECTION 2.1. Material and Methods. In this work, all of the starting analytical grade reagents and solvents were acquired from commercial sources and utilized without further purification. A PANalytical empyrean sharp shadow system X-ray diffractometer was used to obtain X-ray diffraction (XRD) patterns of the samples over the 2θ range of 5−80°, and Cu Kα radiation (λ = 1.540598 Å) was used during the testing procedure. Scanning electron microscope (SEM) images with high quality of all samples were acquired using a HITACHI S-4800 scanning electron microscope, and the accelerating voltage in the process of testing is 200 kV. A NEXUS-670 Fourier transform infrared spectrophotometer was utilized to assess the functional groups in all samples, and photoluminescence spectra of fluorescent reagents and the obtained sample were characterized using a Hitachi F-7000 fluorescence spectrophotometer. Thermogravimetric analysis (TGA) 3456


Research Article

ACS Applied Materials & Interfaces dehydrating agent was added to the above mixture, which was subsequently stirred for 16 h in the dark. At the end of the reaction, the obtained Fe-MIL-53-NH2-FA-5-FAM/5-FU nanocomposite was isolated from the solution through centrifugation, followed by washing with water and drying under vacuum at 25 °C. Through use of a similar method to calculate loading efficiency of 5-FU on Fe-MIL53-NH2, the loading efficiency of 5-FU on Fe-MIL-53-NH2-FA5-FAM/5-FU nanocomposite was calculated to be 23%, and the loading amounts of FA and 5-FAM on Fe-MIL-53-NH2-FA-5-FAM/ 5-FU nanocomposite were 2.6 and 1.5%, respectively. Additionally, the standard concentration curves of FA and 5-FAM are shown in Figures S3 and S4. The aforementioned synthetic procedure was subsequently used to synthesize Fe-MIL-53-NH2-FA-5-FAM and Fe-MIL-53-NH2-5-FAM/ 5-FU. Details of the procedure are provided in the Supporting Information. 2.5. Cytotoxicity Study. MGC-803 and HASMC cells were used to evaluate the in vitro cytotoxicities and antitumor efficiencies of Fe-MIL-53-NH2-FA-5-FAM, Fe-MIL-53-NH2-FA-5-FAM/5-FU, and 5-FU via the MTT method. In short, MGC-803 and HASMC cells were seeded onto 96-well plates with a culture density of 104 cells per well. After MGC-803 and HASMC cells were incubated for 24 h, the initial culture medium was removed and replaced by the dispersions of Fe-MIL-53-NH2-FA-5-FAM, Fe-MIL-53-NH2-FA-5-FAM/5-FU, and 5-FU in various concentrations (0, 6.25, 12.5, 25, 50, 100, and 200 μg/mL). After being incubated for another 24 h, 20 μL of MTT solution was added to each well in which the previous culture medium has been removed and incubated for another 4 h. At last, all media were removed from each well before 100 μL of DMSO was added, and the cell viability was obtained by monitoring the absorbance value of each sample at 570 nm using a microplate reader. The cell viability was expressed as a percentage of the absorbance value of the sample well compared to that of the control cell. All experiments were executed in triplicate, and the results were averaged. 2.6. Cellular Uptake Study. The in vitro targeting ability of the FA to FR was evaluated by luminescence imaging in MGC-803 and HASMC cells. The cells were attached onto a 6-well plate for 24 h. Afterward, the MGC-803 cells were treated with Fe-MIL-53-NH2-FA5-FAM/5-FU and Fe-MIL-53-NH2-5-FAM/5-FU for 4 h, and HASMC cells were incubated with Fe-MIL-53-NH2-FA-5-FAM/ 5-FU for 4 h (concentration was 0.1 μg/mL). Before being observed using a laser confocal microscope (OLYMPUS, IX81) under an excitation wavelength of 473 nm, the MGC-803 and HASMC cells were washed several times with phosphate buffered saline (PBS). 2.7. In Vivo Magnetic Resonance Imaging. All animal experiments were conducted according to the guidelines approved by the Institutional Animal Care and Use Committee of Peking University. To carry out the in vivo imaging study, athymic nude mice bearing glioblastoma were injected with Fe-MIL-53-NH2-FA-5-FAM/ 5-FU aqueous solution (5 mg/kg). At 1 h after intratumoral injection, magnetic resonance imaging was taken by a Siemens Prisma 3.0 T MR scanner (Erlangen, Germany) with a gradient strength up to 80 mT/m. 2.8. In Vivo Biodistribution in Different Organs. To investigate the biodistribution of Fe-MIL-53-NH2-FA-5-FAM/5-FU in different organs, athymic nude mice were administered the solutions via intraperitoneal injection (10 mg/kg) and were sacrificed at the 24 h time point postinjection. The major organs were collected, and the fluorescence signal intensity was recorded on an IVIS imaging system (Iumina II) at an excitation wavelength of 495 nm. After autofluorescence was deducted, the biodistribution of different organs was concluded as the ratio of average fluorescence intensity of the individual organ to the average fluorescence intensity of all measured organs. 2.9. In Vitro Drug Release Study. The sustained 5-FU release assays were carried out by soaking the samples in PBS (pH 7.4 and 5). At 37 °C, a dialysis containing 0.05 g of Fe-MIL-53-NH2-FA-5-FAM/ 5-FU was put into a 10 mL PBS solution in a 50 mL centrifuge tube. For real-time monitoring of the release behavior of 5-FU, 2 mL of solution was withdrawn from the centrifuge tube at appropriate time intervals, and the UV−vis absorption values of the solutions were

recorded at 265 nm. The cumulative release profiles were expressed by cumulative release percentages of 5-FU from Fe-MIL-53-NH2-FA5-FAM/5-FU as a function of time, in which the cumulative release percentages of 5-FU from Fe-MIL-53-NH2-FA-5-FAM/5-FU were calculated as follows: cumulative 5-FU release = amount of released 5-FU/amount of total 5-FU × 100%.

3. RESULTS AND DISCUSSION 3.1. Fabrication and Characterization of Fe-MIL-53NH2-FA-5-FAM/5-FU. Different reactant concentrations were applied to control the size of Fe-MIL-53-NH2, and the results are shown in Figure 2. Figures 2A−D show the SEM images of

Figure 2. SEM images of nanocrystalline Fe-MIL-53-NH2 with different reactant concentrations (A−D).

Figure 3. XRD patterns of simulated Fe-MIL-53-NH2 (A) and Fe-MIL53-NH2 nanocrystallines prepared with different reactant concentrations (B−E).

the nanocrystalline Fe-MIL-53-NH2 prepared by employing decreasing amounts of FeCl3·6H2O from 0.4 to 0.1 mmol and BDC-NH2 from 0.4 to 0.1 mmol. Interestingly, as the reactant concentrations decrease, the sizes of the nanoparticles change from 120 nm to 1 μm. Thus, tuning the amount of the reactant is a facile and robust way to control the particle size. To check the crystallinity, the materials were further investigated by XRD, which is depicted in Figure 3. The XRD reflections of all 3457


Research Article

ACS Applied Materials & Interfaces

Figure 4. (a) FTIR spectra of nanocrystalline Fe-MIL-53-NH2 (A), FA (B), 5-FAM (C), and nanocrystalline Fe-MIL-53-NH2-FA-5-FAM (D); and (b) FTIR spectra of 5-FU (A), Fe-MIL-53-NH2-FA-5-FAM/5-FU nanocomposites (B), and Fe-MIL-53-NH2-FA-5-FAM nanocomposites (C).

Fe-MIL-53-NH2 size of about 120 nm was chosen to form the multifunctional drug carriers by conjugation with the targeted agent (FA) an fluorescent reagent (5-FAM). To identify the successful synthesis of Fe-MIL-53-NH2-FA5-FAM nanocomposites, FTIR spectra of Fe-MIL-53-NH2 (Figure 4aA), FA (Figure 4aB), 5-FAM (Figure 4aC), and Fe-MIL-53-NH2-FA-5-FAM (Figure 4aD) were recorded. Fe-MIL-53-NH2 exhibits the N−H symmetrical deformation mode at 1573 cm−1, while the stretching vibrations of the CO of carboxyl groups in FA and 5-FAM appear at 1696 and 1693 cm−1, respectively. For comparison, the FTIR spectrum of Fe-MIL-53-NH2-FA-5-FAM shows typical bands of Fe-MIL-53NH2, FA, and 5-FAM, as revealed by Figure 4aD. However, no band around 1696 cm−1 that is representative of CO of carboxyl groups is observed in Figure 4aD, and the characteristic peak of CO of amide groups appears at 1596 cm−1, suggesting the successful conjugation of FA and 5-FAM to NH2 on the surface of Fe-MIL-53-NH2. Further, confocal laser scanning microscopy images of Fe-MIL-53-NH2-FA-5-FAM (Figure S5) show a strong green luminescent signal, originated from the representative fluorescence of 5-FAM, confirming the successful conjugation of 5-FAM. After being loaded with 5-FU, the FTIR spectrum of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU was collected. The FTIR spectra further indicate the presence of 5-FU on the DDS (Figure 4b). The peak at 1655 cm−1 belongs to the vibrations of CO in 5-FU (Figure 4bA). The broad band at 1596 cm−1 is attributed to CO of amide groups in the Fe-MIL-53-NH2FA-5-FAM nanomaterial (Figure 4bC). Generally, Fe-MIL-53NH2-FA-5-FAM/5-FU exhibits the typical absorption of 5-FU and Fe-MIL-53-NH2-FA-5-FAM, which indicates that the loading of 5-FU with Fe-MIL-53-NH2-FA-5-FAM was successful. Due to the formation of DDS Fe-MIL-53-NH2-FA5-FAM/5-FU, the TGA curve of Fe-MIL-53-NH2-FA-5-FAM/ 5-FU is extremely different from the TGA curve of Fe-MIL53-NH2, as shown in Figure S6. The DDS Fe-MIL-53-NH2FA-5-FAM/5-FU exhibits larger weight loss because of decomposition of the loaded 5-FU, FA, and 5-FAM. Additionally, postmodification and 5-FU loading cause the XRD peaks to broaden, while the dominating peaks of the XRD spectrum of Fe-MIL-53-NH2-FA-5-FAM/5-FU correlates with the FeMIL-53-NH2 pattern, as shown in Figure S7, suggesting the crystalline integrity of Fe-MIL-53-NH2-FA-5-FAM/5-FU. 3.2. Properties and Applications of Fe-MIL-53-NH2FA-5-FAM/5-FU. The transverse relaxation times (T2) of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU with various concentrations were measured, and the relaxivity values (r2) of DDS

Figure 5. Relaxation rate 1/T2 versus concentrations of DDS Fe-MIL53-NH2-FA-5-FAM/5-FU using a 3T MRI scanner. T2-weighted MRI of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU with diverse Fe concentrations in vitro (the inset).

Figure 6. Excitation (A) and emission (B) spectra of 5-FAM, and the excitation (C) and emission (D) spectra of DDS Fe-MIL-53-NH2-FA5-FAM/5-FU.

the obtained Fe-MIL-53-NH2 nanocrystallines correspond well with the simulated pattern, readily indexing high crystallinity of the obtained Fe-MIL-53-NH2 nanomaterials and the same crystal structures of all the obtained nanocrystalline Fe-MIL53-NH2. Due to the abundant NH2 on the surface of nanocrystalline Fe-MIL-53-NH2 and its small particle size, the 3458


Research Article


Figure 7. Fluorescence imaging of live MGC-803 cells cultured with Fe-MIL-53-NH2-FA-5-FAM/5-FU (A) and Fe-MIL-53-NH2-5-FAM/5-FU (B) and HASMC cells cultured with Fe-MIL-53-NH2-FA-5-FAM/5-FU (C) for 4 h. Left, middle, and right panels are dark-field images, bright-field images, and overlays, respectively. Scale bar: 100 μm.

band with a maximum at 530 nm (excited at 480 nm), DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU generates a broad band with a maximum at 560 nm (excited at 495 nm), as shown in Figure 6, which belongs to green emission and fulfills the requirements of fluorescence imaging. To gain more insight into the targeting ability of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU, we carried out fluorescence imaging tests using FA-positive MGC-803 cells and FA-negative HASMC cells, and the results are shown in Figure 7. When MGC-803 cells are incubated with Fe-MIL-53-NH2-FA-5-FAM/ 5-FU, bright green fluorescence is observed in the cell. However, no obvious fluorescence contrast is obtained in MGC-803 cells incubated with Fe-MIL-53-NH2-5-FAM/5-FU or HASMC cells incubated with Fe-MIL-53-NH2-FA-5-FAM/5-FU. This observation shows that only the FA-conjugated Fe-MIL-53NH2-FA-5-FAM/5-FU nanocomposite shows a high affinity to the cancer cells but has little interaction with normal cells, confirming the targeted fluorescence imaging ability of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU. We also investigated the magnetic resonance imaging capability of Fe-MIL-53-NH2-FA5-FAM/5-FU in vivo (Figure 8). Clearly, at 1 h postinjection, the tumor became darker, and the T2-weighted MR image exhibits intense contrast enhancement. In brief, Fe-MIL-53NH2-FA-5-FAM/5-FU displays good imaging capability similar to that of the fluorescence and magnetic resonance imaging

Fe-MIL-53-NH2-FA-5-FAM/5-FU can be calculated by the slope of a plot of the relaxation rate 1/T2 versus Fe concentration. As we know, dipolar interaction between the magnetic moments of Fe-MIL-53-NH2-FA-5-FAM/5-FU and the protons in the water can cut down T2. Thus, 1/T2 increases linearly within the tested range of Fe concentration, as revealed by Figure 5, and DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU exhibits high relaxivity values (r2 = 18.8 mM−1 s−1) for potential clinical use as contrast agents for T2-weighted MRI. The T2-weighted MRI images of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU are shown in the inset of Figure 5, and the iron concentrations in images 2 to 7 are 0.115, 0.2308, 0.4617, 0.9235, 1.847, and 3.694 mM, respectively. It is evident that the signal intensity decreases (darkening) with the increase in the iron concentration, suggesting that Fe-MIL-53-NH2-FA-5-FAM/ 5-FU is a better candidate for T2-weighted MRI. Furthermore, the fluorescence testing of DDS Fe-MIL-53-NH2-FA-5-FAM/ 5-FU was also performed. Clearly, compared to the fluorescence spectrum of 5-FAM, the excitation and emission spectra of Fe-MIL-53-NH2-FA-5-FAM/5-FU are similar, suggesting that the luminescent property of Fe-MIL-53-NH2FA-5-FAM/5-FU is mostly derived from the 5-FAM. It is noted that the slight discrepancies in the position of excitation and emission wavelengths are due to the combination of 5-FAM and Fe-MIL-53-NH2. While 5-FAM exhibits a broad emission 3459


Research Article


the liver, and the XRD of Fe-MIL-53-NH2-FA-5-FAM/5-FU after being soaked for 7 days in PBS agrees well with the original Fe-MIL-53-NH2-FA-5-FAM/5-FU (Figure S8), suggesting that this DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU is stable under physiological conditions. It is necessary to investigate the cytotoxicity of Fe-MIL-53NH2-FA-5-FAM/5-FU for its potential application in bioimaging. Figure 10 shows the effects of Fe-MIL-53-NH2-FA-5-FAM, DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU, and 5-FU on the viabilities of MGC-803 and HASMC cells using the MTT method. After MGC-803 and HASMC cells were incubated with Fe-MIL-53-NH2-FA-5-FAM at various concentrations, little toxicity for the nanocomposite was observed at concentrations up to 200 μg/mL, and the cell viability decreases by 80%, as shown in Figures 10aA and bA, indicating that the Fe-MIL-53-NH2-FA-5-FAM nanocomposite displays excellent biocompatibility. Similarly, DDS Fe-MIL-53-NH2-FA-5-FAM/ 5-FU and 5-FU show little toxicity to HASMC cells, and more than 80% of cells survive at a concentration of 200 μg/mL (Figures 10aB and aC). In contrast, incubation of DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU with MGC-803 cells exhibits a dose-dependent toxicity similar to that of 5-FU and induces 45% death of MGC-803 cells when the concentration of Fe-MIL-53-NH2-FA-5-FAM/5-FU is 200 μg/mL (Figures 10bB and bC). These data demonstrate that DDS Fe-MIL-53-NH2FA-5-FAM/5-FU gives highly efficient and selective treatment of tumor cells due to the targeting ability of FA and the release of 5-FU. Due to the stable and large pore size, the Fe-MIL-53-NH2 nanocomposite displays BET surface area up to 198 m2/g (Figure S9), and the drug loading efficiency of Fe-MIL-53NH2-FA-5-FAM/5-FU is 23%. To validate the sustained release property of 5-FU from the Fe-MIL-53-NH2-FA-5-FAM/5-FU system, release experiments were carried out in PBS buffer solutions (pH 7.4 and 5) at 37 °C, and Figure 11 shows the drug delivery profile. Apparently, the release rates of 5-FU both in pH 7.4 and 5 gradually decrease with time. During the first 5 h, a distinctly rapid 5-FU release from DDS Fe-MIL-53-NH2FA-5-FAM/5-FU was detected, which is mostly due to the fast release of 5-FU molecules on the Fe-MIL-53-NH2-FA-5-FAM/ 5-FU surface. Subsequently, drug release slows, and the gentle release lasts for 25 h in pH 7.4, while the gentle release lasts for 20 h in pH 5. This prolonged 5-FU release is ascribed to the extended diffusion route of 5-FU from pores and cavities of Fe-MIL-53-NH2-FA-5-FAM/5-FU to the surrounding environment. The faster and more stable drug release in pH 5 may be due to the slight dissolution of Fe-MIL-53-NH2 in the acidic environment. Clearly, DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU exhibits remarkable time-release properties, making its presence known in the area of targeted drug delivery.

Figure 8. T2-weighted MR images of tumor-bearing athymic nude mice without Fe-MIL-53-NH2-FA-5-FAM/5-FU (A) and with Fe-MIL-53-NH2-FA-5-FAM/5-FU via intratumoral injection (B).

Figure 9. Accumulation of Fe-MIL-53-NH2-FA-5-FAM/5-FU in major organs excised from athymic nude mice 24 h postinjection, and ex vivo fluorescence imaging of different organs at 24 h postinjection (inset).

systems previously reported.30−33 Further, Fe-MIL-53-NH2FA-5-FAM/5-FU shows stable drug loading and release behavior, making it superior for use in practical applications. To explore the dispersibility and stability of this DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU for in vivo application, the biodistribution of Fe-MIL-53-NH2-FA-5-FAM/5-FU was analyzed by ex vivo fluorescence intensity of different organs 24 h after deducting autofluorescence, and the result is shown in Figure 9. It can be found that DDS Fe-MIL-53-NH2-FA-5FAM/5-FU is able to transport to tissues in vivo, and the intensity from the liver was highest among all measured organs. Additionally, the ex vivo fluorescence imaging of different organs 24 h postinjection is shown in the inset of Figure 9. Apparently, strong fluorescence signals can still be detected in

4. CONCLUSION Tumor-targeting and programmable chemotherapeutic delivery are demonstrated by the designed DDS Fe-MIL-53-NH2-FA-5FAM/5-FU. This smart system provides a unique MOF platform for fluorescence/magnetic resonance dual-mode imaging and targeted drug delivery. The Fe-MIL-53-NH2-FA-5-FAM/ 5-FU nanocomposite was prepared using a reflux method at low temperature, followed by a simple postmodification method. In vitro fluorescence imaging verifies that DDS Fe-MIL-53-NH2FA-5-FAM/5-FU exhibits excellent receptor-specific targeting fluorescence imaging for FA-positive MGC-803 cells, and the high transverse relaxivity of the nanocomposites gives it potential 3460


Research Article


Figure 10. (a) Viabilities of HASMC cells cultured with Fe-MIL-53-NH2-FA-5-FAM (A), DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU (B), and 5-FU (C) evaluated by MTT. (b) Viabilities of MGC-803 cells cultured with Fe-MIL-53-NH2-FA-5-FAM (A), DDS Fe-MIL-53-NH2-FA-5-FAM/5-FU (B), and 5-FU (C) evaluated by MTT.

■

ACKNOWLEDGMENTS All authors received funding from the Natural Science Foundation of China (Grant 21361016) and the Inner Mongolia Autonomous Region Fund for Natural Science (Grant 2013ZD09).

■

(1) Della Rocca, J.; Liu, D.; Lin, W. Nanoscale Metal−Organic Frameworks for Biomedical Imaging and Drug Delivery. Acc. Chem. Res. 2011, 44, 957−968. (2) Yoon, M.; Srirambalaji, R.; Kim, K. Homochiral Metal−Organic Frameworks for Asymmetric Heterogeneous Catalysis. Chem. Rev. 2012, 112, 1196−1231. (3) Keskin, S.; Kizilel, S. Biomedical Applications of Metal Organic Frameworks. Ind. Eng. Chem. Res. 2011, 50, 1799−1812. (4) Zhang, T.; Lin, W. Metal−organic Frameworks for Artificial Photosynthesis and Photocatalysis. Chem. Soc. Rev. 2014, 43, 5982− 5993. (5) James, S. L. Metal-organic frameworks. Chem. Soc. Rev. 2003, 32, 276−288. (6) McKinlay, A. C.; Morris, R. E.; Horcajada, P.; Férey, G.; Gref, R.; Couvreur, P.; Serre, C. BioMOFs: Metal−Organic Frameworks for Biological and Medical Applications. Angew. Chem., Int. Ed. 2010, 49, 6260−6266. (7) Wang, C.; Liu, D.; Lin, W. Metal−Organic Frameworks as A Tunable Platform for Designing Functional Molecular Materials. J. Am. Chem. Soc. 2013, 135, 13222−13234. (8) Horcajada, P.; Gref, R.; Baati, T.; Allan, P. K.; Maurin, G.; Couvreur, P.; Férey, G.; Morris, R. E.; Serre, C. Metal−Organic Frameworks in Biomedicine. Chem. Rev. 2012, 112, 1232−1268. (9) Horcajada, P.; Chalati, T.; Serre, C.; Gillet, B.; Sebrie, C.; Baati, T.; Eubank, J. F.; Heurtaux, D.; Clayette, P.; Kreuz, C. Porous Metal− organic-framework Nanoscale Carriers as A Potential Platform for Drug Delivery and Imaging. Nat. Mater. 2010, 9, 172−178. (10) Wang, H. N.; Yang, G. S.; Wang, X. L.; Su, Z. M. pH-induced Different Crystalline Behaviors in Extended Metal−organic Frameworks based on the Same Reactants. Dalton Trans. 2013, 42, 6294− 6297. (11) Cooper, L.; Hidalgo, T.; Gorman, M.; Lozano-Fernández, T.; Simón-Vázquez, R.; Olivier, C.; Guillou, N.; Serre, C.; Martineau, C.; Taulelle, F. A Biocompatible Porous Mg-gallate Metal−organic Framework as An Antioxidant Carrier. Chem. Commun. 2015, 51, 5848−5851. (12) Liu, K. G.; Zhu, Z.; Wang, X. Y.; Gonçalves, D.; Zhang, B.; Hierlemann, A.; Hunziker, P. Microfluidics-based Single-step Preparation of Injection-ready Polymeric Nanosystems for Medical Imaging and Drug Delivery. Nanoscale 2015, 7, 16983−16993. (13) Wong, P. T.; Chen, D. X.; Tang, S. Z.; Yanik, S.; Payne, M.; Mukherjee, J.; Coulter, A.; Tang, K.; Tao, K.; Sun, K.; Baker, J. R., Jr;

Figure 11. Drug release profile for DDS Fe-MIL-53-NH2-FA-5-FAM/ 5-FU in PBS buffer solution at pH 7.4 (A) and 5 (B).

as a contrast agent for MRI. Additionally, such DDS is also found to generate better toxicity to cancer cells (MGC-803 cells) due to the targeted 5-FU release, and release studies show that the 5-FU release behavior can last for more than 20 h. In conclusion, this paper provides a brief and efficacious method to explore MOFs as a new targeted drug delivery system which generates significantly enhanced antitumor efficacy. These results provide a proof of concept for a new mode of drug delivery, and we fully expect to see this drug delivery system make its presence known in the area of biological applications.

■

ASSOCIATED CONTENT

S Supporting Information *

Supporting Information for this article is available on the The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsami.6b14795. Chemical structure, fluorescence imaging, TGA curves, XRD patterns, N2 adsorption−desorption isotherms, and absorbance of 5-FU in PBS buffer solution (PDF)

■

REFERENCES

AUTHOR INFORMATION

Corresponding Authors

*E-mail: [email protected]. *E-mail: [email protected]. ORCID

Zhiliang Liu: 0000-0003-3917-6014 Notes

The authors declare no competing financial interest. 3461


Research Article


superparamagnetic iron oxide nanoparticles. J. Mater. Chem. B 2016, 4, 3969−3981. (33) Liu, Y.; Kang, N.; Lv, J.; Zhou, Z. J.; Zhao, Q. L.; Ma, L. C.; Chen, Z.; Ren, L.; Nie, L. M. Deep Photoacoustic/Luminescence/ Magnetic Resonance Multimodal Imaging in Living Subjects Using High-Efficiency Upconversion Nanocomposites. Adv. Mater. 2016, 28, 6411−6419.

Choi, S. K. Modular Integration of Upconverting NanocrystalDendrimer Composites for Folate Receptor-Specific NIR Imaging and Light-Triggered Drug Release. Small 2015, 11, 6078−6090. (14) Chen, W. H.; Luo, G. F.; Lei, Q.; Cao, F. T.; Fan, J. X.; Qiu, W. X.; Jia, H. Z.; Hong, S.; Fang, F.; Zeng, X.; Zhuo, R. X.; Zhang, X. Z. Rational Design of Multifunctional Magnetic Mesoporous Silica Nanoparticle for Tumor-targeted Magnetic Resonance Imaging and Precise Therapy. Biomaterials 2016, 76, 87−101. (15) Song, W. Y.; Di, W. H.; Qin, W. P. Synthesis of mesoporoussilica-coated Gd2O3: Eu@silica particles as cell imaging and drug delivery agents. Dalton Trans. 2016, 45, 7443−7449. (16) Shen, B. B.; Ma, Y.; Yu, S. Y.; Ji, C. H. Smart Multifunctional Magnetic Nanoparticle-Based Drug Delivery System for Cancer Thermo-Chemotherapy and Intracellular Imaging. ACS Appl. Mater. Interfaces 2016, 8, 24502−24508. (17) Della Rocca, J.; Liu, D. M.; Lin, W. B. Nanoscale Metal− Organic Frameworks for Biomedical Imaging and Drug Delivery. Acc. Chem. Res. 2011, 44, 957−968. (18) Carné, A.; Carbonell, C.; Imaz, I.; Maspoch, D. Nanoscale Metal−organic Materials. Chem. Soc. Rev. 2011, 40, 291−305. (19) Lin, W. B.; Rieter, W. J.; Taylor, K. M. L. Modular Synthesis of Functional Nanoscale Coordination Polymers. Angew. Chem., Int. Ed. 2009, 48, 650−658. (20) Spokoyny, A. M.; Kim, D.; Sumrein, A.; Mirkin, C. A. Infinite Coordination Polymer Nano- and Microparticle Structures. Chem. Soc. Rev. 2009, 38, 1218−1227. (21) Serre, C.; Mellot-Draznieks, C.; Surblé, S.; Audebrand, N.; Filinchuk, Y.; Férey, G. Role of Solvent-Host Interactions That Lead to Very Large Swelling of Hybrid Frameworks. Science 2007, 315, 1828−1831. (22) Zhang, P. C.; Lock, L. L.; Cheetham, A. G.; Cui, H. G. Enhanced Cellular Entry and Efficacy of Tat Conjugates by Rational Design of the Auxiliary Segment. Mol. Pharmaceutics 2014, 11, 964− 973. (23) Prabaharan, M.; Grailer, J. J.; Pilla, S.; Steeber, D. A.; Gong, S. Q. Folate-conjugated Amphiphilic Hyperbranched Block Copolymers based on Boltorn®H40, Poly(l-lactide) and Poly(ethylene glycol) for Tumor-targeted Drug Delivery. Biomaterials 2009, 30, 3009−3019. (24) Sudimack, J.; Lee, R. J. Targeted Drug Delivery via the Folate Receptor. Adv. Drug Delivery Rev. 2000, 41, 147−162. (25) Mansoori, G. A.; Brandenburg, K. S.; Shakeri-Zadeh, A. A Comparative Study of Two Folate-Conjugated Gold Nanoparticles for Cancer Nanotechnology Applications. Cancers 2010, 2, 1911−1928. (26) Stella, B.; Arpicco, S.; Peracchia, M. T.; Desmaële, D.; Hoebeke, J.; Renoir, M.; D’Angelo, J.; Cattel, L.; Couvreur, P. Design of Folic Acid-Conjugated Nanoparticles for Drug Targeting. J. Pharm. Sci. 2000, 89, 1452−1464. (27) Carlsson, M.; Gustavsson, M.; Hu, G. Z.; Murén, E.; Ronne, H. A Ham1p-Dependent Mechanism and Modulation of the Pyrimidine Biosynthetic Pathway Can Both Confer Resistance to 5-Fluorouracil in Yeast. PLoS One 2013, 8, e52094. (28) Mojardín, L.; Botet, J.; Quintales, L.; Moreno, S.; Salas, M. New Insights into the RNA-Based Mechanism of Action of the Anticancer Drug 5′-Fluorouracil in Eukaryotic Cells. PLoS One 2013, 8, e78172. (29) Li, Y. T.; Tang, J. L.; He, L. C.; Liu, Y.; Liu, Y. L.; Chen, C. Y.; Tang, Z. Y. Core−Shell Upconversion Nanoparticle@Metal−Organic Framework Nanoprobes for Luminescent/Magnetic Dual-Mode Targeted Imaging. Adv. Mater. 2015, 27, 4075−4080. (30) Wu, Q.; Cheng, Q. Q.; Yuan, S. M.; Qian, J. C.; Zhong, K.; Qian, Y. F.; Liu, Z. Y. A Cell-penetrating Protein Designed for Bimodal Fluorescence and Magnetic Resonance Imaging. Chem. Sci. 2015, 6, 6607−6613. (31) Sun, Y. Q.; Wang, D. D.; Zhao, T. X.; Jiang, Y. N.; Zhao, Y. Q.; Wang, C. X.; Sun, H. C.; Yang, B.; Lin, Q. Fluorescence-Magnetism Functional EuS Nanocrystals with Controllable Morphologies for Dual Bioimaging. ACS Appl. Mater. Interfaces 2016, 8, 33539−33545. (32) Lam, T.; Avti, P. K.; Pouliot, P.; Tardif, J. C.; Rhéaume, E.; Lesage, F.; Kakkar, A. Magnetic resonance imaging/fluorescence dual modality protocol using designed phosphonate ligands coupled to 3462


Controllable Synthesis of a Smart Multifunctional Nanoscale Metal

Recommend Documents