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10 1039@C7NR08747K
10 1039@C7NR08747K
10 1039@C7NR08747K
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Nanoscale
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Zhao, X. Yi, L. Zhu, F. Ge, X. Mou, L. Chen, L. Sun and K. Yang, Nanoscale, 2018, DOI:
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DOI: 10.1039/C7NR08747K
Nanoscale
Paper
While radiotherapy (RT) is commonly used in clinic for cancer treatment, the therapeutic efficiency is not satisfactory
owing to the existence of the hypoxia tumor microenvironment which seriously affect the efficiency of RT. Herein, we
design polyethylene glycol (PEG) modified reduced nano-graphene oxide-manganese dioxide (rGO-MnO2-PEG)
nanocomposites to trigger oxygen generation from H2O2 to reduce the tumor hypoxic microenvironments. We use
radioisotope, 131I labeled rGO-MnO2-PEG as therapeutic agents for in vivo tumor radioisotope therapy (RIT), achieving
excellent tumor killing and further enhancing the therapeutic efficiency of RIT. More importantly, dissolution of MnO2 in
acidic condition and the redox process during catalytic pathway of H2O2 decomposition in cellular microenvironment direct
to the production of enormous amount of Mn2+ which has been used as contrast agents for magnetic resonance imaging
(MRI). Our proposed work provides a strategy to trigger oxygen formation via internal stimulus to enhance imaging guided
RIT efficiency.
This journal is © The Royal Society of Chemistry 20xx J. Name., 2017, 00, 1-3 | 1
laser irradiation and the γ rays from the 131I labeled RGO-PEG spectroscopy (FTIR) analysis (Supporting information Figure
were significantly inhibited compared to that of the control S3). After PEGylation, the size of rGO-MnO2-PEG became
groups. Therefore, we for the first time successfully designed smaller compared to uncoated rGO-MnO2 as revealed from
graphene-based nanoplatform for combination of photothermal TEM and atomic force microscope (AFM) imaging
therapy and radiotherapy in cancer treatment. However, it has measurements (Figure 2a&d). The sizes distribution of rGO-
not been yet reported that the use of nano-graphene for imaging MnO2 before and after PEGylation were measured to be 80 nm
guided radiotherapy simultaneously with hypoxia overcoming. and 24 nm, respectively by dynamic light scattering (DLS)
In our work, we design PEG modified 131I labelled, assay (Figure 2f). The results were also corroborated with the
reduced nano-graphene oxide and manganese dioxide TEM and AFM images.
nanocomposites (131I-rGO-MnO2-PEG) nanocomposites as MnO2 nanoparticles have been widely considered as agents
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therapeutic agents for overcoming hypoxic microenvironments to trigger the oxygenation in acidic condition. The catalytic
and enhancing radioisotope therapy (RIT) of cancer (Figure 1). pathway of MnO2 for the decomposition of H2O2 in acidic pH is
The rGO-MnO2-PEG nanocomposites significantly improve the shown by the following equation:
oxygen formation in the solid tumor sites in the presence of MnO2 +2H+ → Mn2+ + H2O + 1/2O2↑
2 | J. Name., 2017, 00, 1-3 This journal is © The Royal Society of Chemistry 20xx
toxic effect on 4T1 cells at our tested concentrations, indicating pH 5.8, we were encouraged to use these nanocomposites for
the biocompatibility of rGO-MnO2 nanocomposites with tumor MR imaging. Similar to gamma imaging, the mice of
PEGylation. In addition, unlike to free 131I which was unable to bearing 4T1 tumor model were i.v. injected with rGO-MnO2-
kill tumor cells effectively at our tested dose range, 131I-rGO- PEG (10 mg/kg) and imaged by a 3.0-T clinical MR scanner
MnO2-PEG with different radioactivity doses induced (Bruker Biospin Corporation, Billerica, MA, USA) at different
significant death of cancer cell (Figure 4b). Therefore, 131I- time points after injection. The rGO-MnO2-PEG
rGO-MnO2-PEG exhibited obvious toxicity to cancer cells, nanocomposites demonstrated strong T1-weighted in
likely owing to the ease penetration of 131I-rGO-MnO2-PEG comparison with the positive MR imaging of tumor tissues after
nanocomposites into the cells. Additionally, 4T1 cancer cells 1h of post-intravenous administration. The signal intensity was
were incubated with Cy5.5 labelled rGO-MnO2-PEG gradually increased with time and reached in a maximum
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nanocomposites for 6 h, and then imaged by confocal intensity in 6 h. This phenomenon could be explained as when
fluorescence microscopy. As expected, obvious red signal light rGO-MnO2-PEG reached to the tumor tissue via the typical
of Cy5.5 were observed in the cytoplasm (Figure 2d), EPR effect, MnO2 NPs disintegrated into Mn2+ under the
indicating that rGO-MnO2-PEG nanocomposites could easily specific triggers of acidic environment and generated huge
This journal is © The Royal Society of Chemistry 20xx J. Name., 2017, 00, 1-3 | 3
made 131I-rGO-MnO2-PEG plus DMXAA showed obvious cell The Preparation and modification of rGO-MnO2
apoptosis and necrosis compared to other control groups nanocomposites
(Supporting information Figure S5c). Furthermore, the major Firstly, graphene-oxide (GO) was manufactured using the
organs of mice treated with 131 I-rGO-MnO2-PEG or PBS were modified Hummer’s method as followed in our previous work.
53
obtained for H&E staining after 15 days of injection. No The obtained GO was dissolved in deionized (DI) water for
obvious toxicity was found of the 131I-rGO-MnO2-PEG treated next experiments. For preparation of rGO-MnO2
group contrasted with the control group. (Supporting nanocomposites, 10 mg of potassium permanganate (KMnO 4)
information Figure S6). Moreover, the body weight of mice were added into the 10 ml of water-solution containing 10 mg
received different groups showed no apparent change of GO under magnetic stirring at room temperature. After 5 min,
(Supporting information Figure S5a&b). 2 ml of borane morpholine complex (5 mg/ml) solution was
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Such excellent therapeutic efficiency of 131I-rGO-MnO2- added dropwise into the above mixture to trigger the formation
PEG with or without DMXAA was attributed to overcome the of MnO2 nanoparticles on the surface of reduced GO. The
tumor hypoxic microenvironments via the decomposition of above compound was reacted at room temperature overnight.
MnO2 in the presence of H2O2. In order to further verify the The yielded rGO-MnO2 nanocomposites were collected and
Conclusions 131
I labelling
In summary, we have developed rGO-MnO2 nanocomposites In this work, radionuclide 131I, which has been commonly used
with PEG modification as multifunctional therapeutic agents in clinic, was used to radiolabel rGO-MnO2-PEG to study its in
for multimodal imaging and enhanced RIT of cancer via vivo behaviour. The detailed process of radiolabelling could be
overcoming tumor hypoxic microenvironments. The significant found in our previously published protocol.50, 57 The
amount of the oxygen formation was possible due to the radiolabelling yield of 131I-rGO-MnO2-PEG was measured to
decomposition of MnO2 from rGO-MnO2-PEG. 131I be 60% after purification. The radiolabelling stability was
radiolabelled rGO-MnO2-PEG possessed long time blood tested in serum at 37°C.
circulation and high tumor accumulation via intravenous
injection into mice. After the decomposition of MnO2 Cell experiments
nanoparticles, the released Mn2+ ions from rGO-MnO2-PEG 4T1 murine breast cancer cells were cultured following the
nanocomposites acted as strong T1-weighted MR contrast standard method.58 For cytotoxicity analysis, 4T1 cells pre-
agent. Meanwhile, tumor hypoxia had been reduced remarkably seeded in 96-well plate were treated with different
through rGO-MnO2-PEG triggered oxygen formation in the concentrations of rGO-MnO2-PEG, 131I and 131I-rGO-MnO2-
presence of H2O2 in the solid tumor. With the help of reversing PEG for 24 h. The cells relative viabilities were tested by
tumor hypoxic microenvironments, 131I-rGO-MnO2-PEG CCK8 (cell counting kit-8). To investigate the cell uptake of
exhibited significant inhibition of tumor growth. Importantly, nanocomposites, 4T1 cells incubated with Cy5.5 labelled rGO-
DMXAA we used significantly caused the tumor vascular MnO2-PEG-NH2 at different time points (0.5, 2, 4, 6 h) were
disruption, and increase tumor uptake of 131I-rGO-MnO2-PEG washed with phosphate buffered saline (PBS) for two or three
nanocomposites, further enhancing the therapeutic efficiency of times, and then visualized by confocal fluorescence microscope
RIT. Therefore, our study provided an appropriate strategy to (OLYMPUS).
overcome tumor hypoxia and enhance RIT efficiency, further
highlighting the promising application prospect of graphene- In vitro oxygen generation
based functional nanocomposites in tumor diagnosis and We investigated the O2 generation ability of rGO-MnO2-PEG
treatment nanocomposite (100 µM of [Mn2+]) in a H2O2 solution (100
µM). Amount of dissolved oxygen generation was measured by
Experimental Section
4 | J. Name., 2017, 00, 1-3 This journal is © The Royal Society of Chemistry 20xx
an oxygen probe (JPBJ-608 portable Dissolved Oxygen Meters, group, (6) 131I-rGO-MnO2-PEG plus DMAXX (20 mg/kg)
Shanghai REX Instrument Factory). treated group (200 µCi of 131I corresponding to 10 mg/kg rGO-
MnO2-PEG). Those nanomaterials were i.v. injected at day 0, 4,
In vitro MRI measurement 8. The tumor volumes were measured using a caliper every
rGO-MnO2-PEG solutions with different manganese other day, and calculated the volume to tumor according our
concentrations treated with pH 7.4 or pH 5.8 were scanned previously published method.52
under a 3.0-T clinical MR scanner (Bruker Biospin Corporation,
Billerica, MA, USA) at room temperature. After acquiring the
T1-weighted MR images, the signal intensity values were Acknowledgements
measured for each sample in the region of interest (ROI). T This work was partially supported by National Natural
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Relaxation rates r1 (1/T1) was calculated from T1 values at Science Foundation of China (81471716, 81302383, 81672430,
different Mn concentrations. 31400861, 11575124), the National Natural Science Foundation
of Jiangsu Province (BK20140320), Fund for Young medical
Tumor model Scholars of Juangsu province (QNRC2016232), a Project
This journal is © The Royal Society of Chemistry 20xx J. Name., 2017, 00, 1-3 | 5
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