A Magnetocatalytic Propelled Cobalt–Platinum@Graphene Navigator for Enhanced Tumor Penetration and Theranostics

Abstract

Open AccessCCS ChemistryRESEARCH ARTICLE14 Jul 2022A Magnetocatalytic Propelled Cobalt–[email protected] Navigator for Enhanced Tumor Penetration and Theranostics Liang Zhang†, Qian Dong†, Hui Zhang†, Jieqiong Xu, Shen Wang, Lufeng Zhang, Wentao Tang, Zhaoqian Li, Xin Xia, Xinqi Cai, Shengkai Li, Ruizi Peng, Zhengyu Deng, Michael J. Donovan, Long Chen, Zhuo Chen and Weihong Tan Liang Zhang† Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Qian Dong† Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Hui Zhang† Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Jieqiong Xu Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Shen Wang Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Lufeng Zhang Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Wentao Tang Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Zhaoqian Li Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Xin Xia Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Xinqi Cai Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Shengkai Li Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Ruizi Peng Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Zhengyu Deng Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Michael J. Donovan Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 , Long Chen Faculty of Science and Technology, University of Macau, Taipa, Macau 999078 , Zhuo Chen *Corresponding author: E-mail Address: [email protected] Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 and Weihong Tan Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province, Hunan University, Changsha, Hunan 410082 The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institute of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022 https://doi.org/10.31635/ccschem.021.202101219 SectionsSupplemental MaterialAboutAbstractPDF ToolsAdd to favoritesDownload CitationsTrack Citations ShareFacebookTwitterLinked InEmail Complex biological environments and multiple physiological barriers significantly impede efficient accumulation and penetration of nanomaterials within tumor tissue for therapy. In situ energy conversion of nanomotors features autonomous movements and improves cancer treatment. However, one of the key challenges is to prepare nanomotors with an adequately small size, good biocompatibility, and precise positioning. Herein, we demonstrate a simple, ultrasmall, versatile, and real-time motion guidance strategy for magnetocatalytic [email protected] navigators (MCGNs) that can enable highly efficient propulsion in the presence of H2O2 or magnetic actuation. MCGNs act as highly diffusive delivery vehicles to promote tumor tissue targeting, and the amount of drug in the tumor was three times than without navigation. By engaging movements powered through in situ energy conversion, MCGNs gain considerable propulsion to penetrate a cell's membrane and enhance intracellular delivery. Download figure Download PowerPoint Introduction In recent decades, tremendous research effort into nanomaterials and nanodevices has enabled diverse biomedical applications for the imaging, diagnosis, and treatment of intractable cancers.1–3 In this context, passive and active targeting strategies have been extensively exploited to selectively transport diagnostic and therapeutic agents to tumor sites. However, complex biological environments and multiple physiological barriers significantly impede efficient accumulation and penetration of nanomaterials within tumor tissue.4 Such environments and barriers include large distances between blood vessels in solid tumors, complicated composition of the extracellular matrix, extensive cell–cell adhesion, high interstitial fluid pressure, lack of convection, and undesirable drug metabolism and binding.5–7 Furthermore, most nanomedicines cannot actively seek tumor sites and lack a propelling force to penetrate within tumor tissue beyond their conventional diffusion limit. Nanomotors, as one kind of intelligent nanodevice, can convert chemical energy or other forms of energy into its own mechanical locomotion.8–10 Typical driving forces for nanomotor encompass bubbles,11–15 self-electrophoresis,16 light,17–19 ultrasonic waves,20–22 electric field,23,24 and magnetic field (MF).25–27 On account of their small size, autonomous movement, and large push ratio, nanomotors have shown great therapeutic potential in anticancer and antithrombic therapy.28–30 In the absence of inertial forces for macroscopic objects, propulsion of nanosized nanomotors in fluids is challenging and necessitates overcoming the low Reynolds number and ubiquitous Brownian motion. To meet these critical demands, nanomotors that are capable of leveraging local chemical fuels or external fields to generate driving forces have been proposed to achieve efficient energy harvesting and conversion.31 For example, a magnetoaerotactic bacteria has been reported to convey drug-loaded liposomes to the hypoxic regions of tumor tissue in mice.32,33 Wang et al.34 demonstrated the directed transport of magnetic polymeric drug-loaded nanomotors by investigating the effect of drugs on the propulsion speed and trajectory of such nanomotors, both theoretically and experimentally. Ghosh et al.35 have synthesized helical magnetic nanomotors that could exquisitely manipulate and cruise inside live cells, making them promising for intracellular sensing and drug delivery. However, there remains several key drawbacks that hinder potential biomedical applications of autonomous nanomotors. First, the size of most nanomotors is relatively large, >500 nm, limiting their shuttling between tissues, and the moving direction is difficult to precisely control. Second, biocompatibility and stability hinder the applications of some nanomotors in biological environments.36,37 It is of great significance to construct the carriers with small size, good biocompatibility, multiple driving forces, and the ability to navigate and target tumor tissues. The concept of multimodal self-propelling nanoparticles (NPs) using hydrolysis of H2O2 and MF has been widely used. However, due to the size, stability and biocompatibility of these nanomotors, they have not been successfully applied to solid tumors. In this work, we constructed the magnetocatalytic [email protected] navigators (MCGNs) with small diameters via a chemical vapor deposition (CVD) method. MCGNs that were uniform in size, bimetallic, highly magnetic, encased in a graphene shell, and dispersible had potential applications as carriers in tumor diagnosis and treatments (Scheme 1). Under the MF guided extravasation of small MCGNs into tumor tissue, diffusional hindrance into the interstitial matrix was lowered and penetration into the tumor parenchyma occurred. Surface PEGylation of the small NPs allowed them to diffuse smoothly into the interstitial matrix by reducing the binding, sequestration, and metabolism that hinder the transport of therapeutic agents.38 However, such MCGNs were not cleared from the tumor as rapidly as much smaller molecular species. In addition to its superior magnetic properties, the MCGNs could also be used for magnetic resonance imaging (MRI). Meanwhile, the MCGNs had an outstanding enzyme-mimicking activity over a broad pH range, which could catalyze H2O2 to produce O2 and offer another bubble driving force for the MCGNs. In addition to the CoPt core having good photothermal capabilities, the graphene shell also had photothermal conversion capability which could further enhance the therapeutic capabilities of MCGNs. Moreover, the graphene shell had superior stability and could effectively protect the inner cobalt and platinum core against corrosive environments (acid, enzymes). Furthermore, graphene has a large conjugated structure that can be loaded with various drug molecules for more therapeutic applications.39 Scheme 1 | Illustration of the MCGNs propelled navigation to enhance intracellular delivery and PTT action against tumor cells. Download figure Download PowerPoint Experimental Methods Details of the materials and instruments used are provided in the Supporting Information. Synthesis of CoxPt100-x/graphene-shell nanomotors CoxPt100-xGs were produced in a CVD system. First, fumed silica (1.0 g, Aladdin, Shanghai, China) was impregnated with H2PtCl6·6H2O (50 mg) and Co(NO3)2·6H2O (0–168 mg, see Supporting Information Table S1) in 100 mL methanol and sonicated for 2 h. The mixture was then dried at 60 °C, and the powder was ground. Typically, 0.50 g of the powder was used for methane CVD in a tube furnace. The sample grew with a methane flow of 150 cm3/min and hydrogen flow of 20 cm3/min for 5 min. After growth, the sample was etched with HF and HNO3 to dissolve the silica and the uncovered CoPt particles. The CoxPt100-xGs solid product was then washed thoroughly and collected through centrifugation. To further investigate the influence of Co/Pt ratio to magnetic and catalytic properties, CoxPt100-xGs with different Co/Pt ratios ( Supporting Information Table S1) were prepared and characterized by inductively coupled plasma mass spectrometry (ICP-MS; Optima 8000; PerkinElmer, Massachusetts, America). We chose Co64Pt36Gs, which had the best catalytic performance and magnetic properties. Functionalization by C18-PEG We added MCGNs to 30 mL octadecyl-polyethylene glycol solution (1 mg/mL) and sonicated them for 1 h. We used centrifugation at 12,000g for 60 min to remove any aggregates. We added 30 mL water to the centrifuge product and washed three times to remove excess octadecyl-polyethylene glycol. In vitro photothermal study To investigate the photothermal property of MCGNs, 0.5 mL of MCGNs (1.0, 0.5, 0.25, and 0.1 mM) and phosphate-buffered saline (PBS) solutions were put into 1.5 mL microtubes and exposed to an 808 nm laser with a power density of 1.0 W/cm2 for 8 min. During the irradiation, an infrared (IR) thermal camera was used to record the temperature change of different samples every 1 min. Similarly, we investigated the photothermal property of MCGNs under different laser powers (0.25, 0.5, 1.0, and 2.0 W/cm2). To study their photostability, 0.5 mL of MCGNs (1.0 mM) were irradiated by an 808 nm laser at 1.0 W/cm2 for 8 min and then naturally cooled for 12 min. This heating and cooling cycle was repeated five times to evaluate the photostability of MCGNs. Cell viability Hela cells were utilized for the viability tests. In brief, 5000 Hela cells per well were seeded into 96-well plates for 24 h (37 °C, 5% CO2). Then, MCGNs (0, 0.2, 0.4, 0.6, 0.8, 1, and 1.5 mM) were added into the Hela cells for another 24 h. After that, Hela cells were washed with dulbecco phosphate-buffered saline (dPBS) twice and incubated with cell counting kit-8 (CCK-8) solution (10 μL) for 2 h. Finally, a microplate reader was used to measure the absorbance of each at 450 nm. Intracellular O2 generation analysis under different pH [(Ru(dpp)3)]Cl2 (RDPP) was used to detect the intracellular production of O2 by confocal laser scanning microscope (CLSM) imaging. In the presence of O2 molecules, the fluorescence of [(Ru(dpp)3)]Cl2, an O2 sensing probe, is strongly quenched. In brief, Hela cells (1 × 105) were seeded into culture dishes for 24 h (37 °C in 5% CO2), then H2O2 (100 μM) was added to the cells for 4 h. After that, cells were washed with dPBS twice and treated with RDPP (5 μM) for another 4 h. Subsequently, Hela cells were further incubated with MCGNs (200 μM) under different pH and hypoxic environment for another 12 h. Finally, cells were collected and the intracellular fluorescence was measured via a CLSM with excitation from a 488 nm laser. Raman imaging of MCGNs in living cells Raman imaging was used to measure the endocytosis of MCGNs into living cells. Hela cells were seeded on dishes for 24 h (37 °C, 5% CO2), followed by incubation with MCGNs under different conditions for another 3 h. After that, the cells were washed with dPBS twice, collected and fixed on a slide for Raman mapping with an excitation from a 633 nm laser. In vivo tumor therapy All healthy BALB/c mice (female, 4–6 weeks old, 18–20 g) were purchased from the Hunan SLRC Laboratory Animal Co., Ltd (Changsha, China) and used under protocols approved by the Institutional Animal Care and Use Committee of Hunan University. To develop the subcutaneous tumor model, 4T1 cells (2 × 106), suspended in 100 μL of PBS, were subcutaneously injected into the right flank of each female BALB/c mouse. When the tumor volume reached 80–100 mm3, the mice were divided randomly into 12 groups consisting of five mice in each group. 4T1 tumor-bearing mice were injected with 100 μL of normal saline (groups 1, 2, 3, and 4) or 30 mg kg−1 MCGNs (groups 5, 6, 7, 8, 9, 10, 11, and 12) through the tail vein. After injection, the mice (groups 9, 10, 11, and 12) were placed under strong MF. After 6 h post injection, the mice (groups 3, 4, 5, 6, 11, and 12) the mice were injected subcutaneously with 50 μL 0.003% H2O2 in normal saline. After 12 h postinjection, mice were irradiated with the 808 nm laser at 1 Wcm−2 for 8 min. Subsequently, tumor growth was recorded by measuring the tumor's perpendicular diameter using a caliper estimated by employing the following equation: volume = (tumor length) × (tumor width)2/2. Results and Discussion Advanced characterization of MCGNs CoPt nanocrystal was specifically chosen as a catalase mimic because of easy preparation, excellent catalytic ability in a broad pH range, smaller size, good dispersity, and stability.40,41 We prepared MCGNs by a CVD method,42,43 and Figure 1a illustrates MCGNs synthesis and surface functionalization. The influence of different atomic ratio of Co and Pt on MCGNs' enzyme-mimic activity and magnetic properties were initially studied ( Supporting Information Table S1 and Figure S1). Notably, the MCGNs exhibited both superior catalytic activity and magnetic resonance (MR) T2 relaxivities when the atomic ratio of Co/Pt was approximately 3∶2. Hence, the MCGNs with this atomic ratio of Co/Pt were used for further studies. We obtained stable aqueous suspensions of MCGNs by noncovalent functionalization with C18-PEG (octadecyl-polyethylene glycol, molecular weight of ∼4670) molecules. The hydrocarbon chains of C18-PEG were adsorbed onto the graphene shells via van der Waals and hydrophobic interactions, whereas the hydrophilic PEG chain extended into the aqueous phase to impart solubility. The functionalized MCGNs were highly stable in H2O, dPBS, cell culture solution, and serum against aggregation for 7 days, and the concentrated solution of the MCGNs maintained good dispersibility after being stored for 1 year at room temperature ( Supporting Information Figure S2). The transmission electron microscopy (TEM) image revealed that nanocrystals were uniform with an average diameter of 3.5 ± 1.2 nm ( Supporting Information Figure S3). High-resolution TEM image (Figure 1b) clearly showed one- or two-layer graphene shells overcoating the core nanocrystals. From the representative scanning TEM (STEM) image and its corresponding Pt and Co elemental maps (Figure 1c), we observed that both Co and Pt were distributed in each individual NP. The energy dispersive X-ray (EDX) analysis ( Supporting Information Figure S4) and the X-ray photoelectron spectroscopy (XPS) results ( Supporting Information Figure S5) showed detectable signals of Pt and Co from these samples. The powder X-ray diffraction (XRD) patterns of the MCGNs could be indexed to Pt(111), Pt(200), Pt(220), Pt(311), Co(100), Co(002), Co(101), Co(110), and graphite diffractions of a CoPt fcc structure ( Supporting Information Figure S6). The diffraction peaks were positioned between the standard peaks of Pt and Co, demonstrating the formation of the bimetallic phase of Co and Pt. Raman spectroscopy measurements identified a graphene carbon G peak at ∼1600 cm−1, a disordered D peak at ∼1330 cm−1, and a 2D peak at ∼2670 cm−1 (Figure 1d), providing further evidence for the graphene shells. The MCGNs demonstrated excellent corrosion resistance in aqua regia, while the CoPt NPs without graphene coating were completely dissolved in the aqua regia within 3 min, turning the solution green (Figure 1e). TEM, XPS, and XRD characterization results ( Supporting Information Figure S7) of MCGNs treated with aqua regia were consistent with that of the original MCGNs. The MCGNs maintained good magnetic properties even after being immersed in aqua regia for 35 days, and the mass was only reduced by less than 10%. In addition, MCGNs were a high relaxivity MRI contrast agent. The r2 relaxivity of MCGNs was 537.4 mM−1s−1, and as concentrations increased, T2 shortening resulted in a significant loss in signal intensity noticeable on T2-weighted images (Figure 1f). Additionally, dynamic light scattering (DLS) data showed that MCGNs had good dispersity with an average hydrodynamic diameter of approximately 10 nm ( Supporting Information Figure S8), with negative charges ( Supporting Information Figure S9). Figure 1 | (a) Illustration showing the process of MCGNs synthesis and surface functionalization. (b) Typical TEM image of MCGNs. Scale bar is 5 nm. (c) The high-angle annular dark-field (HAADF)-STEM image of MCGNs and corresponding TEM element mappings of the Co K-edge and Pt K-edge signals. Scale bars are 5 nm. (d) Raman spectrum (excitation 633 nm) of MCGNs, showing the G, D, and 2D bands of graphene carbon. (e) weight and of MCGNs or CoPt NPs in aqua regia at different T2 measurements and T2-weighted images of MCGNs. effect of MCGNs. temperature of and MCGNs of 0.25, 0.5, and 1.0 are the corresponding thermal images of the solutions of 1.0 MCGNs under the laser at the power density of W/cm2 for different photothermal effect of MCGNs. Download figure Download PowerPoint We further measured the photothermal of MCGNs under of a laser using as the control. After 8 min of at 1.0 the temperature of only by 2.0 °C, while the temperature of 1.0 MCGNs by We also the temperature of and CoPt the the temperature of MCGNs was °C than that of CoPt without graphene ( Supporting Information Figure When the MCGNs was 0.1 the temperature also by °C (Figure We then investigated the photothermal of 1.0 MCGNs under different laser When the laser intensity was 0.25, 0.5, 1.0, and 2.0 the temperature by and °C, (Figure Supporting Information S1 showed when the laser intensity was 0.5 heating of 1.0 MCGNs occurred. The MCGNs had excellent photothermal even after five of 8 min at 1.0 W/cm2 (Figure Magnetocatalytic propelling ability of MCGNs To evaluate the catalytic effect of MCGNs, we H2O2 after the addition of MCGNs. In pH and more than of H2O2 was by MCGNs within min (Figure We further investigated MCGNs could generate a amount of at a low H2O2 under when at pH and most H2O2 was into by with the MCGNs (Figure of the of MCGNs was catalytic generation of MCGNs had excellent catalytic activity in a pH ( Supporting Information Figure The catalytic activity of the MCGNs was even after 5 addition of H2O2 (Figure The superior enzyme-mimic activity from the graphene shells the electron from the CoPt to the graphene which the electron density of the graphene surface and the enzyme-mimic activity on the graphene We further studied magnetocatalytic propelling properties of MCGNs. of MCGNs moving at different times under an external MF were by in different concentrations of H2O2 The direction of MCGNs in H2O2 was When exposed to an the nanomotors in the direction of the MF. The H2O2 the speed of the MCGNs under the MF. When the of H2O2 increased, the speed of the MCGNs the of H2O2 from to the speed of MCGNs from to When the of H2O2 was and the MF was the speed of the MCGNs times than that without the MF ( Supporting Information Figure When MF increased, the speed of the MCGNs and The MF to different of Supporting Information Figure When the MF was approximately the speed of the reached which a superior magnetocatalytic propelling capability ( Supporting Information Figure of MCGNs moving at different times under a MF were by a in a H2O2 solution Supporting Information Figure and of MCGNs showed the motion of the catalytic magnetic when guided under an external MF. Figure 2 | (a) of H2O2 in under different pH and (b) O2 generation with MCGNs in under different pH and (c) catalytic ability of MCGNs with addition of (d) Intracellular O2 by at different times after MCGNs treatment. (e) pH were to the normal and tumor conditions and to investigate the pH of the O2 generation Download figure Download PowerPoint To further the catalytic activity of MCGNs in a the intracellular O2 of the Hela cells were with as the O2 In a hypoxic strong fluorescence was observed in Hela cells after with the When MCGNs were the fluorescence The intensity of to in 6 h and in 12 h O2 generation within these (Figure Furthermore, we the intracellular O2 generation at pH and (Figure pH and the intensity of was significantly the O2 generation in a pH that graphene could we the using [email without catalytic properties. Under the MCGNs could catalyze the production of O2 from H2O2 and the fluorescence of the whereas the fluorescence signal in the [email protected] ( Supporting Information Figure results clearly demonstrated that MCGNs could convert H2O2 to O2 through in living cells under a of MCGNs performance After we the magnetocatalytic we further studied the by cells and the penetration of solid under an external The MF was the and provided the magnetic driving The produced O2 by H2O2 provided another driving which the accumulation of MCGNs in the and a temperature laser was therapy tumor cells through and when the temperature approximately °C, while the is the at a The of the MCGNs cell was investigated ( Supporting Information Figure significant was even at a of MCGNs as high as 1.0 which the used in the following photothermal Then, we studied the in the ability of MCGNs to cells under different propelling In the presence of the number of MCGNs the cell was than that without added Under the external the number of MCGNs the cell was significantly than that without an external MF. When the MF and H2O2 were at the the number of MCGNs