Docetaxel-loaded SiO₂@Au@GO core–shell nanoparticles for chemo-photothermal therapy of cancer cells

Abstract

The multifunctional antitumor SiO₂@Au@GO core–shell nanoparticles (NPs) have been fabricated for serving as drug nanocarrier and photothermal inducer. The core–shell NPs exhibited high loading capacity of docetaxel (Dtxl) and pH-dependent drug release property as well as low cytotoxicity (concentration < 0.2 mg mL⁻¹). Compared with chemotherapy alone, human prostate cancer cell incubated with Dtxl-loaded SiO₂@Au@GO NPs in vitro were found to have undergone morbidity on exposure to near-infrared (NIR) light (780 nm, 5 W cm⁻²). It is expected that the multifunctional nanoparticles with unique core–shell structure will lead to new opportunities in nanomedicine.

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1. Introduction

Photothermal therapy as an encouraging approach for cancer treatment has aroused great scientific interest in nanotechnology and biomedicine due to its low damage of normal tissue and high curative effect.¹ In this treatment, light is converted into heat through photothermal agents, which would cause the local temperature to rise and result in the thermal ablation of tumor cells. To further improve the therapeutic efficacy, the combination of chemotherapy and photothermal therapy has been developed, which can realize double therapy for cancer via elevating temperature locally and delivering drugs to tumor sites simultaneously.^2,3 Generally, NIR light is the highly desirable light source in photothermal therapy, because the NIR light (700–1200 nm) is noninvasive for normal tissues and can transmit through tissues.^2,4 Thus, great efforts have been devoted to design and fabricate multifunctional antitumor NIR resonant nanomaterials, such as gold-based nanoparticles and carbon nanomaterials.

After Halas's research group first applied SiO₂@Au core–shell NPs for tumor ablation,⁵ such unique structure with metal nanoshells have received great attention in nanomedicine due to its tunable optical resonances and controlled uniform size.⁶ Additionally, the easily functionalized surface of SiO₂@Au core–shell NPs can endow them with multifunctional ability for cancer therapy, such as drug delivery, targeted therapy, cell imaging and specific identification.^3,4,7,8

Graphene is a two-dimensional nanomaterial which are potentially useful in biomedical applications, such as drug loading and delivery.^9–14 After oxidation, graphene becomes more hydrophilic due to the introduction of hydroxyl and carboxyl on its surface, which makes it well-dispersed in aqueous solution as a drug carrier. Dai and colleagues have first fabricated PEGylated nanoscale graphene oxide (GO) which was used for live cell imaging in the near-infrared and also served as a nanocarrier to load doxorubicin via π–π stacking.^15,16 It is found that the nanoscale GO derivatives are efficient nanocarriers for the loading and delivery of water-insoluble aromatic drugs, owing to large specific surface area, and noncovalent physisorption for aromatic drug molecules.

Herein, the fabrication of a multifunctional antitumor SiO₂@Au@GO core–shell NPs with the drug load/release property and photothermal efficiency is reported. In the composites, the inner cores SiO₂@Au NPs serve as photothermal inducer which has a resonance peak at 810 nm, while for outer shell GO with the size of ∼10 nm is used as drug nanocarrier to load hydrophobic antitumor drug Dtxl. The loading capacity and drug-release performance of SiO₂@Au@GO core–shell NPs were investigated. Furthermore, human prostate cancer cell DU 145 was used as a model to research the chemo-photothermal effect of SiO₂@Au@GO NPs on tumor.

2. Experimental section

2.1 Materials and characterization

Chemicals were used as received without further purification. Hydrogen tetrachloroaurate(III) hydrate (HAuCl₄·3H₂O), 3-aminopropyltrimethoxysilane (APTMS, 97%), tetraethylorthosilicate (TEOS, 98%), ammonium hydroxide (NH₃·H₂O, 28–30%) and amino–poly(ethylene glycol)–thiol (NH₂–PEG–SH, 3.4 kDa) were acquired from Sigma-Aldrich. Tetrakis (hydroxymethyl) phosphonium chloride (THPC, 80% in water) was purchased from TCI Development Co., Ltd (Shanghai, China). Sodium hydroxide (NaOH), sodium chloride (NaCl), potassium carbonate, formaldehyde (HCHO, 37–40%), methanol (≥99.5%), n-butanol (≥99.5%) and absolute ethanol (≥99.7%) were analytical-reagent grade and purchased from Sinopharm. Ultrapure water with a resistivity of 18.2 MΩ cm was provided by a Milli-Q system.

The morphologies of the samples were observed by transmission electron microscopy (TEM, JEOL 2100, Japan), field emission scanning electron microscopy (FE-SEM, Carl Zeiss Ultra Plus, Germany) and atomic force microscopy (AFM, Multimode Nanoscope, DI, USA). The ultraviolet-visible-near infrared (UV-Vis-NIR) absorption spectra were recorded by a UV-Vis-NIR spectrophotometer (Lambda 950, Perkin-Elmer, USA). Fourier transformation infrared (FTIR) spectra were recorded on a FTIR spectrometer (Vertex 70, Bruker Optics, Germany). Raman study was performed by a Raman spectrometer at an excitation wavelength of 532 nm (DXRTM, Thermo Fisher, America).

2.2 Experimental details

2.2.1 Preparation of colloidal gold nanoparticles (GNPs). A THPC gold solution composed of 1–5 nm Au colloid was prepared as described by Duff et al.¹⁷ To 180 mL of ultrapure water was added 1.2 mL of 1 M NaOH and 4 mL of 1.2 mM THPC solution (prepared by adding 48 μL of 80% THPC in water to 4 mL of ultrapure water). Then, 6.75 mL of 25 mM aqueous chloroauric acid was added quickly to the mixture, after which the solution immediately changed from colorless to a medium brown color. This gold seed solution should be stored at 4 °C for at least 2 weeks before use.

2.2.2 Preparation of APTMS-functionalized silica cores (SiO₂-APTMS). SiO₂ nanoparticle was prepared by a modified Stöber method.¹⁸ For a given SiO₂ formulation (∼220 nm), 2 mL of NH₃·H₂O was added to the mixture of methanol–butanol in a ratio of 1 : 3 (v/v). Under rapid stirring, 0.5 mL of TEOS was injected dropwise, and then the mixture reacted for 2 h. This product was centrifuged at 5000 rpm for 20 min and washed by ethanol for several times. The obtained precipitation was redispersed in 100 mL ethanol.

To prepared APTMS-functionalized silica cores, 80 μL of APTMS was added to 100 mL silica suspension. The mixture was stirred at room temperature for 24 h and then centrifuged at 5000 rpm for 20 min. The purified SiO₂-APTMS NPs were redispersed in ethanol.

2.2.3 Preparation of SiO₂-APTMS–Au. To make gold NPs attach on the surface of SiO₂-APTMS cores, 4 mL of 1 M NaCl was added to 40 mL of GNPs suspension, followed immediately by injecting 4 mL of the SiO₂-APTMS. After continuous sonication for an additional 1–2 min, the solution was allowed to reach equilibrium overnight under ambient conditions.¹⁹ The nonattached GNPs were removed by centrifuging at 2500 rpm for 10 min until the suspension was clear. The obtained SiO₂-APTMS–Au NPs were redispersed in ultrapure water. In this work, the concentration of SiO₂-APTMS–Au suspension was measured at 562 nm by UV-vis spectroscopy.

2.2.4 Preparation of plating solution (K₂CO₃/HAuCl₄). 3 mL of 25 mM HAuCl₄ aqueous solution was added to 200 mL of 1.8 mM K₂CO₃ aqueous solution. After continuous stirring for nearly 20 min, the solution changed from pale yellow to colorless. The plating solution is stirred overnight away from light, and then stored at 4 °C until use.

2.2.5 Preparation of SiO₂@Au core–shell NPs. Under sonication, 16 mL of SiO₂-APTMS–Au (562 nm 0.65 A) suspension was dispersed in 150 mL of plating solution. Then, 500 μL of HCHO was injected in one quick motion, and the mixture was vigorously stirred for 10 min. Over the course of 1–2 min, the lavender solution, which was the color of SiO₂-APTMS–Au suspension, turned blue to dark green, indicating the formation of SiO₂@Au core–shell NPs.⁶ The resulted nanoparticles were centrifuged at 1000 rpm for 15 min and redispersed in 10 mL ultrapure water. For PEGylation, 10 mL SiO₂@Au solution was mixed with 1 mL of 10⁻⁴ M NH₂–PEG–SH solution and reacted for 24 h at room temperature. The final nanoparticles were centrifuged and washed with water for several times.

2.2.6 Preparation of SiO₂@Au@GO core–shell NPs. Modified Hummers method was used to prepare original GO according to our previous work.^20,21 To obtain GO nanosheet with the size of less than 20 nm, the original GO solution was cracked by ultrasonic probed at 300 W for 6 h. Under sonication, 200 μL of 1000 μL mg⁻¹ GO was added dropwise to 2 mL PEGylated SiO₂@Au solution. Then, the reaction was stand in shaking table at 25 °C for 24 h. The unreacted GO was removed by repeatedly centrifugation.

2.2.7 Photothermal heating effect of as-prepared nanoparticles. To determine the photothermal heating effect of as-prepared nanoparticles, GO, SiO₂@Au and SiO₂@Au@GO NPs in PBS (pH = 7.4) with the concentrations of 150 μg mL⁻¹ were irradiated by NIR laser (780 nm, 5 W cm⁻²) for different time periods.

2.2.8 Drug loading property of SiO₂@Au@GO NPs. SiO₂@Au@GO NPs aqueous suspension was added to Dtxl (Sanwei Pharmaceutical., Shanghai) ethanol solution with stirring for 24 h. The drug-loaded nanoparticles were collected by centrifugation and carefully washed with ethanol till the supernatant turned colorless. The resultant particles was denoted as SiO₂@Au@GO–Dtxl NPs. The amount of Dtxl loaded on SiO₂@Au@GO NPs was estimated by monitoring the concentrations of Dtxl in the initial solution and the supernatant by UV-Vis spectrometry at 230 nm. Dtxl loading capacity (LC) was estimated using eqn (1).

2.2.9 Drug release property of SiO₂@Au@GO NPs. The SiO₂@Au@GO–Dtxl NPs were dispersed in 5 mL buffer solution at various pH (5.5 and 7.4) with stirring at 37 °C. At given time intervals, the release medium was collected by centrifugation. The amount of resealed Dtxl was determined by the measurement of UV absorbance of release medium.

2.2.10 In vitro cytotoxicity assays. The cytotoxicity of SiO₂@Au@GO NPs was evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) viability assay. Human prostate cancer cell DU 145 (4 × 10³) was seeded onto 96-well plates and cultured in F-12 medium supplemented with 10% fetal bovine serum and 1% streptomycin at a humidified 5% CO₂ incubator at 37 °C. After incubation for 24 h, DU 145 cells were rinsed with prewarmed PBS (37 °C) and then incubated in 150 μL of F-12 medium contained different concentration of SiO₂@Au@GO NPs for 12 h. After incubating cells with SiO₂@Au@GO NPs, 20 μL of 0.5 mg mL⁻¹ MTT was added to each well of the plate, which was cultured for another 4 h. Then cell viability was assessed by the colorimetric measurements at 595 nm. For control group, DU 145 cells were cultured in the nanoparticle-free F-12 medium. All of the measurements were performed in quintuplicate.

2.2.11 In vitro photothermal therapy test. DU 145 cells were seeded onto 20 mm coverslips in a 6-well plate and cultured in F-12 medium with SiO₂@Au@GO–Dtxl NPs (150 μg mL⁻¹) at 37 °C with 5% CO₂ for 24 h. Then, the cells were irradiated with 5 W cm⁻² NIR light for 10 min for photothermal treatment, whereas for the control group and chemotherapy alone group, the DU 145 cells cultured in F-12 medium without and with SiO₂@Au@GO–Dtxl NPs, respectively, were not exposed to NIR light. To make the laser irradiate more cells on coverslips, the work distance of laser is kept near 4 cm from the 6-well plates. Afterwards, the cells were incubated at 37 °C for another 24 h and then rinsed with PBS buffer, followed by staining with 1.5 μM propidium iodide (PI) for 15 min at 37 °C. After staining, the cells were rinsed with PBS buffer again, and cells on the coverslips were taken out from 6-well plate for fluorescence measurement. The cell viability was estimated at 543 nm with a fluorescence optical microscope.

2.2.12 Cell viability assay. Cytotoxicity of DU 145 cells under different treatments was investigated using the MTT assay. First, DU 145 cells were cultured in 96-well plates with 150 μL of F-12 media at a density of 4 × 10³ cells per well for 48 h. GO, SiO₂@Au@GO, SiO₂@Au@GO–Dtxl and Dtxl with a certain concentration were each added separately into wells and cultured for 24 h. For thermal therapy, corresponding wells were irradiated with a 780 nm NIR laser (5 W cm⁻²) for 10 min. Then, the culture media was then replaced with fresh F-12 media. After incubation for another 24 h, the cellular viability of the resulting cancer cells was confirmed by MTT assay.

The concentration for GO, SiO₂@Au@GO and SiO₂@Au@GO–Dtxl NPs is 150 μg mL⁻¹. SiO₂@Au@GO–Dtxl NPs applied in all the test were the samples that SiO₂@Au@GO stirred with 800 μg mL⁻¹ of Dtxl ethanol solution for 24 h. According to the loading capacity and release rate of Dtxl on SiO₂@Au@GO as shown in Fig. 7, the concentration of Dtxl is determined as 3 μg mL⁻¹.

3. Results and discussion

3.1 Preparation and characterization of SiO₂@Au@GO core–shell NPs

The synthesis procedure for SiO₂@Au@GO NPs is shown in Fig. 1. The SiO₂@Au NPs were prepared as previously reported.⁶ Through electrostatic interaction, GO with nanometers, less than 10 nm, coated on the surface of SiO₂@Au NPs modified by SH–PEG–NH₂ to form the double shell and multifunctional SiO₂@Au@GO core–shell nanocomposites. In composites, the inner core SiO₂@Au NPs serve as photothermal inducer to realize the ablation of tumor and the outer shell GO NPs can carry hydrophilic/hydrophobic anti-carcinogen to achieve chemotherapy for tumor treatment. To obtain the resultant SiO₂@Au@GO NPs with high monodispersity in this multistep reactions, it should strictly control the quality of nanoparticles fabricated in every step, such as uniform size and high stability.

Fig. 1 Schematic illustration of the fabrication process of Dtxl-loaded SiO₂@Au@GO core–shell nanostructures.

Fig. 2 presents the SEM images of monodisperse silica nanosphere with sizes ranging from 140 nm to 620 nm prepared by a modified Stöber method.¹⁸ The size of silica can be precisely controlled by adjusting the ratio of NH₃·H₂O and TEOS (Table S1†). In traditional Stöber method, except that usually using ethanol as the solvent to produce silica, it is also mentioned that using methanol as solvent displays fastest reaction rates and produce smallest silica spheres while the result of using butanol is just the opposite. From this point, we propose to substitute the mixture of butanol and methanol for ethanol solution. It is assumed that methanol contributes to the quick formation of massive silica seeds, while butanol mainly takes charge of seeds regrowth. Although this method doesn't overcome the main limitation of Stöber process in preparation of small particles with the size less than 100 nm, it is more simple and efficient than multiple regrowth approach²² and significantly balances the nucleation and growth rates, which greatly improves the monodispersity and uniformity of silica spheres.

Fig. 2 SEM images of monodisperse silica spheres with different size. The average size of (a) 143 nm, (b) 222 nm, (c) 419 nm, (d) 500 nm, (e) 590 nm, and (f) 620 nm. The scale bar is 400 nm.

It can be seen from Fig. 3a and b, the spherical GNPs with uniform size possess high monodispersity and stability in aqueous solution and only exhibit a broad shoulder near 520 nm, rather than a striking absorbance peak, due to the small size (∼3 nm) which can be observed form the TEM image.¹⁷ This negatively charged gold seeds can attach on the surface of SiO₂-APTMS NPs through the electrostatic interaction with terminal amine group, which were used as the nucleation site for the formation of gold shell. As shown in Fig. 3c, the gold seeds uniformly and densely distributes on the surface of silica without obvious aggregation, which extremely avail to acquire contiguous and smooth gold shell. The gold shell was formed by the further reductive growth of gold seeds on the surface of SiO₂-APTMS–Au NPs along with the color changed from purple to green (inset in Fig. 3c and d). The fabricated SiO₂@Au core–shell NPs in our work are with a core size of 222 nm and shell thickness of 32 nm. In addition, from the SEM images in Fig. S1,† it is found that most gold shell is quite complete and relatively smooth. To improve the stability and coated by GO NPs, the SiO₂@Au NPs were modified with HS–PEG–NH₂ through the formation of Au–SH bond.²³

Fig. 3 (a) The TEM image and (b) UV-Vis-NIR absorption spectra of GNPs. TEM images of (c) SiO₂-APTMS–Au NPs and (d) SiO₂@Au core–shell NPs. The inset in (a)–(c) is the photo of GNPs, SiO₂-APTMS–Au NPs and SiO₂@Au core–shell NPs aqueous solution, respectively.

GO was usually used as supporter to carry molecular or NPs, especially drugs loading for tumor treatment, due to the π–π conjugation interaction between GO sheets and antitumor drug. Additionally, the carbonaceous GO should be more biocompatible than other nanomaterials, since carbon constitutes the backbone of biomolecules.²⁵ To obtain GO with nanometers which can form outer shell on the surface of SiO₂@Au NPs, the raw GO particles were cracked by ultrasonic probe. The raw GO were mostly 10–300 nm in lateral width, whereas the GO sheets after sonication were ∼5–10 nm according to the AFM characterization (Fig. S2†).

Obviously, the outer shell evolution have a significant effect on the absorbance of as-prepared nanoparticles, as shown in Fig. 4. The absorption peak of SiO₂-APTMS–Au NPs at 520 nm which evidently red-shift to 805 nm after the formation of gold shell. Although the thickness of GO shell coated on SiO₂@Au was only less than 5 nm, it still made the absorption peak a little red-shifting due to the plasma resonance of noble metals is sensitive to the contact medium.²⁴ In addition, the position of resonance is also decided by the ratio of core size and shell thickness, which can be visually estimated by the color of solution, as show in Fig. S3.†

Fig. 4 The magnified TEM image of (a) SiO₂-APTMS–Au NPs, (b) SiO₂@Au NPs, and (c) SiO₂@Au@GO NPs. (d) The UV-Vis-NIR absorption spectra of SiO₂-APTMS–Au NPs, SiO₂@Au NPs and SiO₂@Au@GO NPs.

The reaction process were monitored through characterizing samples by FTIR and Raman spectra. Fig. 5a showed the FTIR spectra of GO, SiO₂@Au–SH–PEG–NH₂ and SiO₂@Au@GO NPs. The peak of –OH (∼3413 cm⁻¹), C=O (∼1737 cm⁻¹) and C=C (∼1629 cm⁻¹) were attributed to the functional groups in GO nanosheets.²¹ For SiO₂@Au–SH–PEG–NH₂ NPs, SiO₂ NPs exhibited a peak at 1103 cm⁻¹ which was caused by the stretching vibration of Si–O bonds, and the peaks around ∼2842 cm⁻¹ implied the strong –CH₂– vibrations in the PEG chains.¹⁵ In addition, the signature of –NH in PEG were also observed at 3410 cm⁻¹ and 1650 cm⁻¹, consistent with the modification of HS–PEG–NH₂ on the surface of SiO₂@Au NPs.²⁶ Compared with SiO₂@Au–SH–PEG–NH₂ NPs, the FTIR spectra of SiO₂@Au@GO showed an additional peak of –OH in GO nanosheets and some differences in fingerprint region below 1000 cm⁻¹. To further confirm the formation of SiO₂@Au@GO core–shell NPs, Raman measurement was performed on all as-prepared nanoparticles. As shown in Fig. 5b, both GO and SiO₂@Au@GO exhibit characteristic D band (1597 cm⁻¹) and G band (1358 cm⁻¹) with a D/G intensity ratio of ∼1.00 and 0.875, respectively. The decrease of D/G ratio implies a reduction in the degree of disorder in GO.^13,27 For SiO₂@Au–SH–PEG–NH₂ NPs, the values of peaks for SiO₂, 564 cm⁻¹ and 1089 cm⁻¹, is consistent well with the result of FTIR measurement, which are significantly weaken for SiO₂@Au@GO NPs as labeled with arrow.

Fig. 5 (a) The FTIR and (b) Raman spectra of GO NPs, SiO₂@Au–SH–PEG–NH₂ NPs and SiO₂@Au@GO NPs.

3.2 Photothermal heating effect of as-prepared nanoparticles

Fig. 6 shows the photothermal heating curves of GO, SiO₂@Au and SiO₂@Au@GO solution (PBS, pH = 7.4). When irradiated by 780 nm NIR laser at the power intensity of 5 W cm⁻², the solution temperature increased from 25.4 to 46.8 °C with 10 min with the concentration of SiO₂@Au at 150 μg mL⁻¹. After coated by GO, there show unobvious change on photothermal heating curves of SiO₂@Au@GO solution, of which the temperature increased from 25.8 to 47.4 °C. In strong contrast, the temperature of GO solution under the same laser irradiation remained below 37 °C with 10 min.²⁸ The results indicate that SiO₂@Au core–shell NPs are the dominated photothermal agent for localized hyperthermia cancer therapy, and GO NPs would contribute more for drug loading.

Fig. 6 Photothermal heating curves of GO, SiO₂@Au and SiO₂@Au@GO solution (PBS, pH = 7.4) with the concentration of 150 μg mL⁻¹ irradiated by NIR at the power intensity of 5 W cm⁻².

3.3 Docetaxel loading and release properties of SiO₂@Au@GO core–shell NPs

The loading of Dtxl on SiO₂@Au@GO NPs was studied in different initial Dtxl concentrations with respect of the same concentration of SiO₂@Au@GO (500 μg mL⁻¹), as shown in Fig. 7a. With the increase of initial Dtxl concentration, the loading capacity of Dtxl increases from 1 mg mg⁻¹ to 2.16 mg mg⁻¹ at the Dtxl concentration of 1.5 mg mL⁻¹. Furthermore, the loading ratio was linearly correlated to the concentration of Dtxl, which is of practical significance for drug-loaded nanocarriers in clinical. The interaction between SiO₂@Au@GO NPs and Dtxl are mainly ascribed to the π–π staking and hydrophobic effect, and there are also hydrogen bonding between them.

Fig. 7 (a) The loading capacity of Dtxl on SiO₂@Au@GO NPs in different initial Dtxl concentrations, and (b) the release of Dtxl on SiO₂@Au@GO NPs at different pH values.

Due to the neutral environment of blood circulation system and the acidic condition in cellular endosome, the drug release efficiency of SiO₂@Au@GO NPs was investigated at pH = 7.4 and 5.5, respectively. Fig. 7b displays that the cumulative release of Dtxl from SiO₂@Au@GO NPs is higher at acidic condition than that at neutral environment. About 65% of Dtxl were released at pH = 5.5 with 72 h, while 44% at pH = 7.4. The pH-dependent drug release of SiO₂@Au@GO NPs could be mainly caused by the increased hydrophilicity of Dtxl at acidic condition, which weakened the π–π stacking and hydrophobic interactions between them, resulting in the easier dissociation of Dtxl from SiO₂@Au@GO nanocarrier. Similar drug release behavior was reported previously for GO.¹⁰ It has been reported that the tumor microenvironment is mildly acidic with a pH range of 5.8–7.1²⁹ and the intracellular environment is even more acidic, at ∼5.0 pH.³⁰ The pH-sensitive release of Dtxl from SiO₂@Au@GO NPs is important in the clinical setting, which can greatly enhance the therapeutic effect.¹⁰

3.4 In vitro cytotoxicity of SiO₂@Au@GO core–shell NPs

To evaluate the cytotoxicity of as-prepared nanoparticles, the human prostate cancer cell DU 145 were incubated in cells culture medium contained various concentrations of SiO₂@Au@GO NPs at 37 °C with 5% CO₂ for 24 h, and then cell viability was assessed by MTT assay. It can be seen from Fig. 8 that the cell viability slowly decreases with the increasing concentrations of SiO₂@Au@GO NPs. However, it is believed that the solutions of SiO₂@Au@GO with a low concentration (<0.2 mg mL⁻¹) can be considered to have low cytotoxicity. The low-toxic indicates that SiO₂@Au@GO NPs are an ideal drug nanocarrier and photothermal agent in biological applications.

Fig. 8 Cytotoxicity detection via MTT assay for DU 145 cells treated with different concentrations of SiO₂@Au@GO NPs for 24 h.

3.5 Chemo-photothermal therapy based on SiO₂@Au@GO core–shell NPs

To investigate the chemo-photothermal treatment, DU 145 cells were treated with SiO₂@Au@GO–Dtxl NPs with and without NIR irradiation, respectively. Then, the cell viability was measured by staining cells with PI fluorescent dye. The fluorescence microscope images of cells treated with laser only (Fig. 9c and d) show similar result with control group (Fig. 9a and b), indicating the laser with 780 nm at 5 W cm⁻² (work distance, 4 cm) is harmless for cells. Compared with control groups, although cells were exposed to SiO₂@Au@GO–Dtxl NPs, traditional chemotherapy alone has a little killing effect on DU 145 cells due to the insufficient release of Dtxl, as shown in Fig. 9e and f. However, cells treated with SiO₂@Au@GO–Dtxl NPs under NIR irradiation changed significantly on morphology and were totally stained by PI (Fig. 9g and h), indicating the death of cancer cells. This result also effectively confirms that the SiO₂@Au@GO–Dtxl NPs fabricated in our work can realize chemo-photothermal treatment on DU 145 cancer cells through NIR irradiation.

Fig. 9 Fluorescence microscope images of DU 145 cells after stained with PI (a, c, e and g) in dark field and (b, d, f and h) in bright field; (a) and (b) the control group without any treatment; (c) and (d) cells treated only with laser; cells incubated with SiO₂@Au@GO–Dtxl treated (e and f) without and (g and h) with NIR irradiation. The scale bar is 20 μm.

In order to further evaluate the therapeutic effect of SiO₂@Au@GO–Dtxl NPs, cell viability with different treatments was measured. The quantitative evaluation of cell viability was performed via the MTT assay, as shown in Fig. 10. The results showed that SiO₂@Au@GO–Dtxl with NIR irradiation cause the highest rate of cell death compared to a single treatment of Dtxl (chemotherapy) or SiO₂@Au@GO with NIR irradiation (photothermal therapy). Limited death was observed by SiO₂@Au@GO–Dtxl treatment alone, which could be caused by the insufficient release of drugs due to the π–π conjugation interaction between GO and Dtxl. These results demonstrated that the combined chemotherapy and photothermal treatments were more efficiency on cancer treament. Dtxl has been reported to have a hyperthermia-enhanced cytotoxicity.³¹ Thus, this synergistic effect was probably ascribed to enhanced cytotoxicity of Dtxl at elevated temperature, whereas a higher heat sensitivity for the cells exposed to Dtxl than for the cells not exposed to Dtxl.⁷

Fig. 10 Viability of DU 145 cells with different treatment.

4. Conclusions

In summary, we fabricated Dtxl-loaded SiO₂@Au@GO NPs that serve as both drug nanocarrier and photothermal agent for chemo-photothermal treatment on human prostate cancer cell DU 145. The SiO₂@Au@GO NPs with low cytotoxicity (concentration < 0.2 mg mL⁻¹) showed high loading capacity of docetaxel and pH-dependent drug release property. Compared with chemo- and photothermal therapy alone, the combined treatment have higher killing effect on DU 145 cells. Last but not least, we still need to optimize SiO₂@Au@GO NPs to realize the targeted therapy on tumor cells, which can further reduce side effects of chemotherapy. Taken together, all results demonstrate the as-prepared SiO₂@Au@GO core–shell NPs have great potential utility for the combination of chemotherapy and photothermal therapy on tumor in clinical trials.

Acknowledgements

This work was supported by National Natural Science Foundation of China (No. 81172191) and the Key Basic Research Foundation of Shanghai Committee of Science and Technology of China (No. 14JC1491200). We also acknowledge support from the Instrumental Analysis Center of Shanghai Jiao Tong University and the Center for Advanced Electronic Materials and Devices of Shanghai Jiao Tong University.

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Footnote

† Electronic supplementary information (ESI) available. See DOI: 10.1039/c6ra03886g

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