A cerium vanadate targeted nanoparticle, its preparation method and application

Abstract

The present invention belongs to the technical field of anti-cancer drugs, and particularly relates to a cerium vanadate targeted nanoparticle, a preparation method thereof and an application. The cerium vanadate targeted nanoparticle provided by the present invention comprises a drug core, a protective shell layer wrapping the drug core; and a targeting material modified on the outer surface of the protective shell; the drug core comprises CeVO<subgt;4< / subgt> nanoparticles. The cerium vanadate targeted nanoparticle provided by the present invention uses CeVO<subgt;4 nanoparticles as a drug carrier. Since the CeVO<subgt;4 nanoparticles have cytochrome c oxidase-like catalytic activity and cooperate with endogenously overexpressed cytochrome c in cancer cells to reduce and consume O<subgt;2, the oxygen concentration in the tumor microenvironment is effectively reduced. The cerium vanadate targeted nanoparticle provided by the present invention enhances the targeting of the nanoparticle to cancer cells, increases the probability of the nanoparticle entering cancer cells, can activate the drug to play a role in a hypoxic environment, and effectively kills tumor cells.

Description

Technical Field

[0001] The present invention belongs to the technical field of anti-cancer drugs, and particularly relates to a cerium vanadate targeted nanoparticle, a preparation method thereof, and an application thereof. Background Art

[0002] Breast cancer is the most common malignant tumor among women, and its incidence rate shows an increasing trend year by year, seriously threatening the physical health and quality of life of women. As a malignant tumor that seriously endangers the physical and mental health of women, the incidence rate of breast cancer ranks first among female malignant tumors in China. At present, the treatment of breast cancer mainly includes surgical operation, endocrine therapy, chemotherapy, radiotherapy and other means, but the treatment effect and prognosis are not satisfactory. The development and clinical application of targeted drugs for the signaling pathways related to the occurrence and development of breast cancer have become a new research hotspot in breast cancer treatment.

[0003] The technology of nano-health materials is an emerging technology. Nanomaterials have excellent characteristics such as high intestinal absorption efficiency, high bioavailability, the ability of sustained and targeted transportation, effectiveness, good in vivo stability, and good solubility, and have been widely used in the diagnosis and treatment of diseases. Nanomaterials as drug carriers have become a research hotspot in the field of nanomedicine.

[0004] Due to the unique hypoxic environment of tumor cells, the killing of tumor cells by nano-drug delivery systems has been widely concerned. Most of the reported nano-drug delivery systems achieve the treatment effect by supplementing oxygen to relieve the tumor hypoxic environment, but the research on the aggravation of tumor cell hypoxia by nanomaterials is relatively rare. Therefore, the synthesis process is simple, and the nano-drug delivery system that aggravates tumor cell hypoxia to promote the drug effect is a challenging research hotspot. Summary of the Invention

[0005] The purpose of the present invention is to provide a cerium vanadate targeted nanoparticle, a preparation method thereof, and an application thereof. The cerium vanadate targeted nanoparticle provided by the present invention has strong targeting property, increases the probability of the nanoparticle entering cancer cells, can activate the drug to play a role in a hypoxic environment, and effectively kills tumor cells.

[0006] In order to achieve the above purpose, the present invention provides the following technical solutions:

[0007] The present invention provides a cerium vanadate targeted nanoparticle, which includes a drug core, a protective shell layer wrapping the drug core; and a targeting material modified on the outer surface of the protective shell;

[0008] The drug core includes cerium vanadate (CeVO4) nanoparticles, and a hypoxia-activated prodrug loaded on the surface of the CeVO4 nanoparticles through electrostatic adsorption;

[0009] The protective shell layer is a cationic liposome layer;

[0010] The targeting materials include mitochondrial targeting materials and tumor cell targeting materials.

[0011] Preferably, the hypoxia-activated prodrug is AQ4N.

[0012] Preferably, the mitochondrial targeting material is rhodamine 110; the tumor cell targeting material is the MUC1 nucleic acid aptamer targeting MCF-7 breast cancer cells.

[0013] Preferably, the cationic liposome layer is a lipid bilayer formed by 1,2-dioleoyl-sn-glycero-3-phosphate, dipalmitoylphosphatidylcholine, cholesterol, DSPE-PEG 5000 and (2,3-dioleoyl-propyl)-trimethylammonium chloride.

[0014] Preferably, the shape of the CeVO4 nanoparticles is cubic, and the average particle size of the CeVO4 nanoparticles is 20 nm.

[0015] The present invention provides a method for preparing the cerium vanadate targeting nanoparticles described in the above technical solution, comprising the following steps:

[0016] Mix the CeVO4 nanoparticles, the hypoxia-activated prodrug and water for electrostatic adsorption to obtain a drug core;

[0017] Mix the drug core, an organic solvent, 1,2-dioleoyl-sn-glycero-3-phosphate (DOPA), (2,3-dioleoyl-propyl)-trimethylammonium chloride (DOTAP), cholesterol (Chol), DSPE-PEG 5000 and dipalmitoylphosphatidylcholine (DPPC), and then perform solvent removal and hydration in sequence to obtain a drug core wrapped by a cationic liposome layer;

[0018] Mix the drug core wrapped by the cationic liposome layer, the mitochondrial targeting material and a solvent for the first modification to obtain a mitochondrial targeting material-modified product;

[0019] Mix the mitochondrial targeting material-modified product, the tumor cell targeting material and a solvent for the second modification to obtain the cerium vanadate targeting nanoparticles.

[0020] Preferably, the method for preparing the CeVO4 nanoparticles comprises the following steps:

[0021] Drop the cerium salt aqueous solution into the metavanadate aqueous solution for precipitation reaction to obtain the CeVO4 nanoparticles; the temperature of the precipitation reaction is 80 °C.

[0022] Preferably, the mass ratio of the CeVO4 nanoparticles to the hypoxia-activated prodrug is 1:1.

[0023] Preferably, the mass ratio of the drug core to 1,2-dioleoyl-sn-glycero-3-phosphate is 10:4; the mass ratio of the drug core to (2,3-dioleoyl-propyl)-trimethylammonium chloride is 10:12; the mass ratio of the drug core to cholesterol is 10:2; the mass ratio of the drug core to DSPE-PEG 5000 is 1:1; the mass ratio of the drug core to dipalmitoylphosphatidylcholine is 10:4;

[0024] The temperature for hydration is 60 °C, and the time is 40 - 55 min.

[0025] The present invention provides the use of the cerium vanadate-targeted nanoparticles described in the above technical solution or the cerium vanadate-targeted nanoparticles prepared by the preparation method described in the above technical solution in the preparation of anti-tumor drugs.

[0026] The present invention provides a cerium vanadate-targeted nanoparticle, which includes a drug core, a protective shell layer that wraps the drug core; and a targeting material modified on the outer surface of the protective shell; the drug core includes CeVO4 nanoparticles and a hypoxia-activated prodrug loaded on the surface of the CeVO4 nanoparticles through electrostatic adsorption; the protective shell layer is a cationic liposome layer; the targeting material includes a mitochondrial targeting material and a tumor cell targeting material. The cerium vanadate-targeted nanoparticle provided by the present invention uses CeVO4 nanoparticles as a drug carrier. Since CeVO4 nanoparticles have cytochrome c oxidase-like catalytic activity, they cooperate with endogenously overexpressed cytochrome c in cancer cells to reduce and consume O2, effectively reducing the oxygen concentration in the tumor microenvironment. The hypoxia-activated prodrug is loaded on the surface of CeVO4 nanoparticles through electrostatic adsorption and can activate the hypoxia-activated prodrug to kill tumor cells in a hypoxic environment. At the same time, to prevent drug leakage, the present invention wraps a cationic liposome layer outside the drug core to form a protective shell layer. Cytochrome c exists in mitochondria. The present invention modifies a specific material targeting mitochondria and a specific material targeting tumor cells on the outer surface of the protective shell layer, and finally forms a cerium vanadate-targeted nanoparticle. The cerium vanadate-targeted nanoparticle provided by the present invention enhances the targeting of the nanoparticle to cancer cells, increases the probability of the nanoparticle entering cancer cells, can activate the drug to play a role in a hypoxic environment, and effectively kills tumor cells. Description of the Drawings

[0027] Figure 1 It is a TEM image of cerium vanadate prepared in an embodiment of the present invention;

[0028] Figure 2AXPS spectrum analysis of Ce 3d of cerium vanadate prepared in the embodiments of the present invention;

[0029] Figure 2B V2p XPS results of cerium vanadate prepared in the embodiments of the present invention;

[0030] Figure 2C O1s XPS peak results of cerium vanadate prepared in the embodiments of the present invention;

[0031] Figure 3 XRD pattern of cerium vanadate prepared in the embodiments of the present invention

[0032] Figure 4 FT-IR spectrum of cerium vanadate prepared in the embodiments of the present invention;

[0033] Figure 5 Zeta potential diagrams of CeVO4, CeVO4@AQ4N-lip, and CeVO4@AQ4N-lip-MUCI prepared in the embodiments of the present invention;

[0034] Figure 6 UV data spectra of CeVO4, AQ4N, R-110, and CeVO4@AQ4N-R110 prepared in the embodiments of the present invention;

[0035] Figure 7 UV-Vis spectral curves showing the change over time (1 - 15 min) during the interaction of CeVO4 with Cytc and O2;

[0036] Figure 8A No O2 is produced during the interaction of CeVO4 with Cyt c - ;

[0037] Figure 8B No H2O2 is produced during the interaction of CeVO4 with Cyt c;

[0038] Figure 8C No ·OH is produced during the interaction of CeVO4 with Cyt c;

[0039] Figure 9 Fluorescence images of MDA-MB-231 cells and MCF-7 cells after co-incubation with the CeVO4@AQ4N-Lip-R110 / MUC1 nanoparticles prepared in Example 1, respectively;

[0040] Figure 10 Intracellular O2 levels reported by RDPP staining of MCF-7 cells after co-incubation with 0 μg / mL and 25 μg / mL of CeVO4, respectively;

[0041] Figure 11ACell viability of MCF-7 cells after co-incubation with different concentrations of CeVO4 respectively;

[0042] Figure 11B Cell viability of MCF-7 cells after co-incubation with different concentrations of AQ4N respectively;

[0043] Figure 11C Cell viability of MCF-7 cells after co-incubation with different concentrations of CeVO4@AQ4N respectively;

[0044] Figure 11D Cell viability of MCF-7 cells after co-incubation with different concentrations of CeVO4@AQ4N-lip-R110 respectively;

[0045] Figure 11E Cell viability of MCF-7 cells after co-incubation with different concentrations of CeVO4@AQ4N-lip-R110 / MUC1 respectively. Detailed implementation manners

[0046] The present invention provides a cerium vanadate targeted nanoparticle, which includes a drug core, a protective shell layer wrapping the drug core; and a targeting material modified on the outer surface of the protective shell;

[0047] The drug core includes CeVO4 nanoparticles, and a hypoxia-activated prodrug loaded on the surface of the CeVO4 nanoparticles through electrostatic adsorption;

[0048] The protective shell layer is a cationic liposome layer;

[0049] The targeting material includes a mitochondrial targeting material and a tumor cell targeting material.

[0050] In the present invention, unless otherwise specified, all preparation raw materials / components are commercially available products well-known to those skilled in the art.

[0051] The cerium vanadate targeted nanoparticle provided by the present invention includes a drug core. In the present invention, the drug core includes CeVO4 nanoparticles, and a hypoxia-activated prodrug loaded on the surface of the CeVO4 nanoparticles through electrostatic adsorption. The shape of the CeVO4 nanoparticles is preferably a cube, and the average particle size of the CeVO4 nanoparticles is preferably 20 nm. The hypoxia-activated prodrug is preferably AQ4N.

[0052] The cerium vanadate targeted nanoparticle provided by the present invention includes a protective shell layer wrapping the drug core. In the present invention, the protective shell layer is preferably a cationic liposome layer. The cationic liposome layer is preferably 1,2-dioleoyl-sn-glycero-3-phosphate, dipalmitoyl phosphatidylcholine, cholesterol, DSPE-PEG5000 A lipid bilayer formed together with (2,3-dioleoyl-propyl)-trimethylammonium chloride.

[0053] The cerium vanadate targeted nanoparticles provided by the present invention include a targeting material modified on the outer surface of the protective shell. In the present invention, the targeting material preferably includes a mitochondrial targeting material and a tumor cell targeting material. The mitochondrial targeting material is Rhodamine 110 (R110). The tumor cell targeting material is preferably the MUC1 nucleic acid aptamer targeting MCF-7 breast cancer cells. In the present invention, the nucleotide sequence of the MUC1 nucleic acid aptamer targeting MCF-7 breast cancer cells is as shown in SEQ ID NO.1.

[0054] The SEQ ID NO.1 is: 5’-GCAGTTGATCCTTTGGATACCCTGG-3’.

[0055] In a specific embodiment of the present invention, the cerium vanadate targeted nanoparticles include a drug core, the drug core is CeVO4 nanoparticles and AQ4N loaded on the surface of the CeVO4 nanoparticles by electrostatic adsorption, and a protective shell layer coated on the surface of the drug core. The protective shell layer is preferably a lipid bilayer formed together with 1,2-dioleoyl-sn-glycero-3-phosphate, dipalmitoyl phosphatidylcholine, cholesterol, DSPE-PEG5000 and (2,3-dioleoyl-propyl)-trimethylammonium chloride; and Rhodamine 110 and the MUC1 nucleic acid aptamer targeting MCF-7 breast cancer cells modified on the outer surface of the protective shell layer. The cerium vanadate targeted nanoparticles provided by the specific embodiment of the present invention are denoted as CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles. Since the CeVO4 nanoparticles have cytochrome c oxidase-like catalytic activity and cooperate with the endogenously overexpressed cytochrome c in cancer cells to reduce and consume O2, the oxygen concentration in the tumor microenvironment is effectively reduced. The hypoxia-activated chemotherapeutic drug AQ4N is loaded on the surface of the CeVO4 nanoparticles by electrostatic adsorption and can be activated to generate AQ4 to kill tumor cells in a hypoxic environment. To prevent drug leakage, a cationic liposome layer DOTAP is wrapped outside CeVO4@AQ4N to form CeVO4@AQ4N-lip. Cytochrome c exists in mitochondria. Specific R110 targeting mitochondria and the MUC1 aptamer specifically targeting MCF-7 breast cancer cells are modified on the nanoparticles, and finally the nanoparticles CeVO4@AQ4N-lip-R110 / MUC1 are formed. The cerium vanadate targeted nanoparticles provided by the present invention enhance the targeting of the nanoparticles to MCF-7 breast cancer cells, increase the probability of the nanoparticles entering MCF-7 breast cancer cells, can activate the drug to play a role in a hypoxic environment, and effectively kill tumor cells.

[0056] The present invention provides a method for preparing the cerium vanadate targeted nanoparticles described in the above technical solution, comprising the following steps:

[0057] Mix CeVO4 nanoparticles, hypoxia-activated prodrug and water for electrostatic adsorption to obtain a drug core;

[0058] Mix the drug core, organic solvent, 1,2-dioleoyl-sn-glycero-3-phosphate, (2,3-dioleoyl-propyl)-trimethylammonium chloride, cholesterol, DSPE-PEG 5000 , and dipalmitoyl phosphatidylcholine, and then perform solvent removal and hydration in sequence to obtain a drug core wrapped by a cationic liposome layer;

[0059] Mix the drug core wrapped by the cationic liposome layer, mitochondrial targeting material and solvent for the first modification to obtain a product modified with mitochondrial targeting material;

[0060] Mix the product modified with mitochondrial targeting material, tumor cell targeting material and solvent for the second modification to obtain the cerium vanadate targeted nanoparticles.

[0061] The present invention mixes CeVO4 nanoparticles, hypoxia-activated prodrug and water for electrostatic adsorption to obtain a drug core.

[0062] In the present invention, the method for preparing the CeVO4 nanoparticles preferably comprises the following steps:

[0063] Drop the cerium salt aqueous solution into the metavanadate aqueous solution for precipitation reaction to obtain the CeVO4 nanoparticles. In the present invention, the metavanadate aqueous solution is preferably ammonium metavanadate aqueous solution. The molar concentration of the metavanadate aqueous solution is preferably 0.025 mmol / L. The cerium salt aqueous solution is preferably cerium nitrate aqueous solution. The molar concentration of the cerium salt aqueous solution is preferably 0.05 mmol / L. In the present invention, the molar ratio of cerium salt in the cerium salt aqueous solution to metavanadate in the metavanadate aqueous solution is preferably 1:1. The dropping is preferably dropwise addition. The precipitation reaction is carried out under oil bath conditions. The temperature of the precipitation reaction is preferably 80 °C. After the cerium salt aqueous solution is completely dropped, the present invention preferably continues to stir and react at 80 °C for 2 h. After the precipitation reaction is completed, the present invention preferably separates the solid-liquid of the obtained precipitation reaction material liquid, and the obtained solid product is washed and dried in sequence to obtain the CeVO4 nanoparticles. The solid-liquid separation is preferably centrifugation. The washing is preferably alternately washed with pure water and absolute ethanol. The drying is preferably freeze-drying.

[0064] In the present invention, the mass ratio of the CeVO4 nanoparticles to the hypoxia-activated prodrug is 1:1. The mass ratio of the hypoxia-activated prodrug to the volume of water is preferably 1 mg: 1 mL. The electrostatic adsorption is preferably carried out under the conditions of light avoidance and stirring. The temperature of the electrostatic adsorption is preferably room temperature, and the time is preferably 24 h. After the electrostatic adsorption is completed, the present invention preferably separates the obtained adsorption slurry into solid and liquid to obtain a solid product, which is successively washed and dried to obtain the drug core. The solid-liquid separation is preferably centrifugation, and the washing is preferably carried out by alternately washing with pure water and absolute ethanol.

[0065] After obtaining the drug core, the present invention mixes the drug core, an organic solvent, 1,2-dioleoyl-sn-glycero-3-phosphate (DOPA), (2,3-dioleoyl-propyl)-trimethylammonium chloride (DOTAP), cholesterol (Chol), DSPE-PEG 5000 , and dipalmitoyl phosphatidylcholine (DPPC), and then successively removes the solvent and hydrates them to obtain a drug core encapsulated by a cationic liposome layer. In the present invention, the mass ratio of the drug core to DOPA is preferably 10:4. The mass ratio of the drug core to DOTAP is preferably 10:12. The mass ratio of the drug core to Chol is preferably 10:2. The mass ratio of the drug core to DSPE-PEG 5000 is preferably 1:1. The mass ratio of the drug core to DPPC is preferably 10:4. The organic solvent preferably includes ethanol and / or chloroform, and the dehydrated ethanol is preferably absolute ethanol. When the organic solvent is preferably absolute ethanol and / or chloroform, the volume ratio of the absolute ethanol to the chloroform is preferably 5:6. The mixing preferably includes the following steps: dissolving the drug core in a first part of the organic solvent to obtain a drug core solution; dissolving DOPA in a second part of the organic solvent to obtain a DOPA solution; ultrasonically mixing the drug core solution and the DOPA solution to obtain a mixed solution; dissolving DOTAP, cholesterol (Chol), DSPE-PEG 5000 and dipalmitoyl phosphatidylcholine (DPPC) in the remaining organic solvent to obtain a mixed solution; stirring and mixing the mixed solution and the mixed solution. The first part of the organic solvent is preferably absolute ethanol, and the second part of the organic solvent is preferably chloroform. The remaining organic solvent is preferably chloroform. The time of the ultrasonic mixing is preferably 30 min. The time of the stirring and mixing is preferably 24 h. The specific implementation manner of removing the solvent is preferably rotary evaporation under pressure. The temperature of removing the solvent is preferably 40-45 °C. The time of removing the solvent is preferably 2-4 h. The material obtained by removing the solvent is preferably mixed with pure water for hydration. The hydration is preferably carried out under a water bath condition. The temperature of the hydration is preferably 60 °C, and the time of the hydration is preferably 40-55 min.

[0066] In the present invention, the cationic liposome layer is obtained through two-step modification. The first step is the interaction between the drug core CeVO4@AQ4N and DOPA. Due to the coupling of the phosphate group in DOPA with the metal ion cerium ion in the nanoparticle and the hydrophobicity between DOPA molecules, the drug core CeVO4@AQ4N is further wrapped by a monolayer of DOPA to form a hydrophobic surface. The second step is to utilize the hydrophobic / hydrophobic interaction between DOPA and DPPC / Chol / DSPE-PEG 5000 / DOTAP to form a self-assembled cationic lipid bilayer on the surface of CeVO4@AQ4N. Among them, Chol acts as a lipid excipient to stabilize the lipid bilayer structure, DSPE-PEG 5000 confers long-circulation characteristics to the nanoparticles and increases the blood circulation time of the liposome, and DPPC acts as an auxiliary lipid to form the liposome. Therefore, the present invention uses DOPA, DOTAP, Chol, DSPE-PEG 5000 and DPPC to wrap the drug core to form a cationic liposome layer.

[0067] [[ID=!0]]After obtaining the drug core wrapped by the cationic liposome layer, the present invention mixes the drug core wrapped by the cationic liposome layer, the mitochondrial targeting material and a solvent for the first modification to obtain a mitochondrial targeting material-modified product. In the present invention, the solvent is preferably dimethyl sulfoxide (DMSO) and water. The mixing preferably includes the following steps: dispersing the drug core wrapped by the cationic liposome layer in a part of the solvent to obtain a drug core solution wrapped by the cationic liposome layer; dissolving the mitochondrial targeting material in the remaining solvent to obtain a mitochondrial targeting material solution; and mixing the drug core solution wrapped by the cationic liposome layer and the mitochondrial targeting material solution. The part of the solvent is preferably water. The remaining solvent is preferably DMSO. The temperature of the first modification is preferably room temperature, the first modification is preferably carried out under stirring conditions, and the stirring time is preferably 24 h.

[0068] After obtaining the modified product of the mitochondrial targeting material, the present invention mixes the modified product of the mitochondrial targeting material, the tumor cell targeting material and a solvent for a second modification to obtain the cerium vanadate targeted nanoparticles. In the present invention, the solvent is preferably water. The mixing preferably includes the following steps: dissolving the tumor cell targeting material in the solvent to obtain a tumor cell targeting material solution; dispersing the modified product of the mitochondrial targeting material in the tumor cell targeting material solution. The concentration of the tumor cell targeting material solution is preferably 2 OD. The second modification preferably includes ultrasonic treatment and shaking treatment in sequence. The time of the ultrasonic treatment is preferably 1 min, and the temperature is preferably 25 °C. The shaking treatment is preferably carried out under dark conditions, the temperature of the shaking treatment is preferably room temperature, and the time is preferably 24 h. After the second modification is completed, the present invention preferably separates the solid and liquid of the obtained second modification material liquid, and the obtained solid product is washed in sequence to obtain the cerium vanadate targeted nanoparticles. The solid-liquid separation is preferably centrifugation, and the washing is preferably carried out with pure water. The washing preferably removes the unloaded MUC1.

[0069] The present invention preferably disperses the prepared cerium vanadate targeted nanoparticles in a neutral PBS solution for storage.

[0070] The present invention provides the use of the cerium vanadate targeted nanoparticles described in the above technical solution or the cerium vanadate targeted nanoparticles prepared by the preparation method described in the above technical solution in the preparation of anti-tumor drugs.

[0071] In the present invention, the anti-tumor drug is preferably an anti-breast cancer drug.

[0072] In order to further illustrate the present invention, the technical solutions provided by the present invention will be described in detail below in conjunction with examples, but they should not be construed as limiting the protection scope of the present invention.

[0073] The materials and reagents used in the following examples include:

[0074] Rhodamine 110 (R110), AQ4N, ammonium metavanadate, cerium nitrate hexahydrate, disodium hydrogen phosphate (Na2HPO4), sodium dihydrogen phosphate (NaH2PO4), porcine heart cytochrome c ≥ 95%, sodium dithionite (Na2S2O4), Amplex Red, horseradish peroxidase (HRP), hydrogen peroxide (H2O2), [Ru(dpp)3]Cl2 dye, WST-1 dye, terephthalic acid (TPA), dimethyl sulfoxide (DMSO), (2,3-dioleoyl-propyl)-trimethylammonium chloride (DOTAP), DSPE-PEG 5000Dipalmitoylphosphatidylcholine (DPPC), 1,2-dioleoyl-sn-glycero-3-phosphate (DOPA), and chloroform were purchased from Macklin. MUC1 aptamer (nucleotide sequence: 5'-GCAGTTGATCCTTTGGATACCCTGG-3'), N-hydroxysulfosuccinimide sodium salt (NHS), and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) were purchased from Sangon Biotech (Shanghai) Co., Ltd. Anhydrous ethanol was purchased from Sinopharm. DAPI was purchased from Beyotime.

[0075] MTT cell proliferation and cytotoxicity assay kits were purchased from Sangon Biotech. Fetal bovine serum was provided by Solebio. MCF-7 breast cancer cells were obtained from Wuhan Pronocell Life Science Co., Ltd.

[0076] The experimental instruments used in the following examples include:

[0077] Zeta potential measurements were performed at 25°C using a laser particle size analyzer (Zetasizer Nano ZS90). The material morphology was characterized by transmission electron microscopy (JEM2100). The chemical and electronic states of the elements were determined by X-ray diffractometer (Rigaku smartlab SE, Japan). X-ray photoelectron spectroscopy (XPS) analysis was performed on an Axis Ultra DLD. FT-IR analysis was performed using an infrared spectrometer (iS50). UV-visible spectra were recorded on an Agilent Cary60 spectrophotometer. Fluorescence data analysis was performed using a fluorescence spectrophotometer (Hitachi FL-4700). MTT assays were performed on a microplate reader (Thermo Fisher K3 touch). Fluorescence imaging of cells was performed using a fluorescence microscope (Nikon Ti2-E).

[0078] The cell culture methods involved in the following embodiments include:

[0079] The MCF-7 breast cancer cells used in the experiment were obtained from Wuhan Punosai Life Science Co., Ltd. and cultured in a constant temperature and humidity chamber at 37°C, 95% humidity, and 5% CO2. The cell culture medium used was DMEM supplemented with 10% fetal bovine serum and 1% double antibody (penicillin-streptomycin).

[0080] Example 1

[0081] The preparation method of the cerium vanadate targeted nanoparticles provided in this embodiment comprises the following steps:

[0082] First step: Dissolve 1 mmol of NH4VO3 in 40 mL of pure water and heat it in an oil bath to 80 °C. Dissolve 1 mmol of Ce(NO3)3·6H2O in 20 mL of pure water and then add it dropwise to the NH4VO3 aqueous solution. Continue stirring at 80 °C for 2 h. Centrifuge the obtained precipitate, wash it alternately with pure water and absolute ethanol for multiple times, and freeze-dry it at -60 °C to obtain CeVO4 nanoparticles.

[0083] Second step: Dissolve 5 mg of AQ4N in 5 mL of pure water. Weigh 5 mg of CeVO4 nanoparticles and add them to the AQ4N aqueous solution, and stir in the dark for 24 h. Centrifuge the obtained precipitate, wash it alternately with pure water and absolute ethanol for multiple times, and dry it at 60 °C to obtain CeVO4@AQ4N.

[0084] Third step: Dissolve 10 mg of CeVO4@AQ4N in 5 mL of absolute ethanol, and dissolve 4 mg of DOPA in 2 mL of chloroform. Ultrasonically mix the two for 30 min. Weigh 12 mg of DOTAP, 2 mg of Chol, 10 mg of DSPE-PEG 5000 and 4 mg of DPPC and dissolve them in 4 mL of chloroform. Stir all the above solutions overnight, remove the organic solvents by rotary evaporation under reduced pressure at 40 - 45 °C for 2 - 4 h, add 1 mL of pure water, and hydrate it in a water bath at 60 °C for 40 - 55 min to obtain CeVO4@AQ4N encapsulated by a cationic liposome layer (denoted as CeVO4@AQ4N-lip).

[0085] Fourth step: Dissolve 10 mg of R-110 in 10 mL of DMSO to obtain a high-concentration stock solution of R-110. Mix the hydrated 10 mg / mL CeVO4@AQ4N-lip aqueous solution and the R110 stock solution in a volume ratio of 1:1, and stir for 24 h to obtain CeVO4@AQ4N-Lip-R110.

[0086] Fifth step: Mix 1 mL of the above-prepared CeVO4@AQ4N-lip-R110 with 100 μL of MUC1 aqueous solution (containing 2 OD), and ultrasonically treat it for 1 min. Gently shake it in the dark at 25 °C for 24 h. Then, centrifuge and collect the obtained CeVO4@AQ4N-lip-R110 / MUC1, wash it several times with water to remove the unloaded MUC1. Finally, disperse CeVO4@AQ4N-lip-R110 / MUC1 in a neutral PBS solution.

[0087] Test example

[0088] (1) Fluorescence detection

[0089] Fluorescence measurements of the partially reduced oxygen species that may be produced by the interaction of CeVO4 with Cyt c were performed to investigate the ability of CeVO4-based cytochrome c oxidases to catalyze O2 without producing partially reduced oxygen species (H2O2, ·OH). CeVO4 was synthesized according to Step 1 of Example 1. Each test sample was diluted to 100 μL with PBS buffer for subsequent analysis. Fluorescence spectra of ·OH and H2O2 were recorded from 335 nm to 600 nm using an F-4700 fluorescence spectrophotometer, using a xenon lamp as the excitation source and an excitation wavelength of 315 nm.

[0090] (2) MTT assay to determine the cytotoxicity of nanoparticles

[0091] MCF-7 breast cancer cells (100 μL, 2×10 6 cells·mL -1 ) were inoculated in 96-well plates and cultured for 24 h. -1 ) and incubated for 24 hours. Then, 10 μL of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) solution (5 mg mL -1 ) and incubated for 4 hours. After rinsing with PBS to remove NPs, 100 μL of DMSO was used to dissolve the formaldehyde crystal deposits in each well. The cells were shaken at low speed for 10 minutes on a shaker. The cells were measured at 490 nm using a microplate reader, and the cell viability was calculated based on the detection data.

[0092] (3) Fluorescence microscopy detected that nanoparticles exacerbated hypoxia in cells

[0093] MCF-7 breast cancer cells were cultured for 24 hours to detect intracellular hypoxia. Various concentrations of CeVO4 nanoparticles were then added and cultured under hypoxic conditions for 24 hours. The cells were then incubated with 0.1 μM tris(4,7-diphenyl-1,10-phenanthroline)ruthenium(II) dichloride (RDPP) dye for 4 hours. After washing the cells three times with PBS to remove excess RDPP, fluorescence intensity was observed using a fluorescence microscope.

[0094] (4) Fluorescence microscopy to detect the targeting of nanoparticles

[0095] MDA-MB-231 and MCF-7 breast cancer cells were inoculated and cultured for 24 hours for targeting analysis. CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles were then added and incubated for 1.5 hours. Finally, cell nuclei were stained with 4',6-diamidino-2-phenylindole (DAPI), and fluorescence intensity was observed under a fluorescence microscope.

[0096] Results and Discussion

[0097] (1) Design and synthesis of CeVO4@AQ4N-lip-R110 / MUC1 prepared in Example 1. The nanomaterials were characterized by other techniques such as transmission electron microscopy (TEM), X-ray diffraction (XRD), and ultraviolet-visible spectrophotometer (UV-vis). The results showed that CeVO4@AQ4N-Lip-R110 / MUC1 nanoparticles were successfully synthesized in Example 1. First, CeVO4@AQ4N was synthesized on the basis of the synthesis of CeVO4 nanoparticles, and CeVO4@AQ4N-Lip-R110 / MUC1 was obtained by surface modification of it. Figure 1 TEM image of cerium vanadate prepared in Example 1. In Figure 1 , TEM data showed that the synthesized cerium vanadate nanoparticles were cubic (≈20 nm). Figure 2A , Figure 2B and Figure 2C are the XPS spectrum analysis of cerium vanadate prepared in Example 1. In Figure 2A X-ray photoelectron spectroscopy (XPS) data, signals on the Ce 3d orbit could be fitted to Ce 3+ and Ce 4+ , indicating its mixed valence state characteristics. Correspondingly, Figure 2B V2p XPS in 5+ showed the presence of V 4+ and V Figure 2C The O1s spectral peaks in Figure 1 , Figure 2A , Figure 2B and Figure 2C were around 529.9 and 531.8 eV, corresponding to the lattice oxygen and oxygen of hydroxyl ions of CeVO4, respectively. Figure 1 , Figure 2A , Figure 2B and Figure 2C The results of Figure 3 proved the successful synthesis of CeVO4 nanoparticles in Example 1. XRD analysis plays an important role in studying the crystal structure of substances. Figure 3 XRD pattern of cerium vanadate prepared in Example 1. Figure 3 In Figure 4 FT-IR spectrum of cerium vanadate prepared in Example 1. Figure 4FT-IR spectra of CeVO4 nanoparticles: There are obvious infrared peaks at 445 and 776 cm -1 −1, corresponding to the stretching vibrations of Ce-O and V-O bonds respectively. Shallow bands of O-H stretching and bending corresponding to water molecules adsorbed on the surface of CeVO4 nanoparticles are observed at 3400 and 1600 cm -1 −1. The infrared characteristic peaks are consistent with the literature reports, which also proves the successful preparation of CeVO4 nanoparticles in Example 1. Figure 5 Zeta potential diagrams of CeVO4, CeVO4@AQ4N-lip, and CeVO4@AQ4N-lip-MUCI prepared in Example 1. Figure 5 The zeta potential results show that the zeta potential of CeVO4 in aqueous solution is negative, -4.86. After wrapping it with a cationic liposome layer, the potential of the material changes significantly, showing +17.36. Since the MUC1 aptamer has a negative potential, the zeta potential value becomes +6.32 after connecting the aptamer, and the positive potential value decreases. Figure 6 UV absorption data spectra of CeVO4, AQ4N, R-110, and CeVO4@AQ4N-R110 in Example 1. Figure 6 The UV absorption data show that the UV absorption characteristic peaks of CeVO4, AQ4N, and R110 are at 275, 498, and 610 nm respectively, and CeVO4@AQ4N-R110 simultaneously has the characteristic peaks of these three substances at 275, 498, and 610 nm, indicating the successful preparation of CeVO4@AQ4N-R110 in Example 1.

[0098] (2) Ability of CeVO4-like cytochrome c oxidase to catalyze the reduction of O2 to produce water

[0099] The cytochrome c oxidase-like activity of CeVO4 was studied by UV / visible spectroscopy. Ferrous cytochrome c (ferrous Cyt c) was obtained by adding 0.4 g of Na2S2O4 to 10 μM / mL of porcine heart cytochrome c for reduction. Ferrous Cyt c has light absorption bands at 414 nm and 550 nm. In the presence of the CeVO4 nanozyme, electrons transfer from ferrous Cyt c to oxygen, and Cyt c is oxidized, resulting in a blue shift of the light absorption band at 414 nm to 409 nm and a significant decrease in the peak intensity of the light absorption at 550 nm. Figure 7 Absorption spectral curves showing the change with time (1 - 15 min) when CeVO4 catalyzes the interaction between ferrous Cyt c and O2. As Figure 7As shown, CeVO4 catalyzes the interaction between Cytc and O2. As time progresses (1 - 15 min), the light absorption band of ferrous Cyt c blue-shifts from 414 nm to 409 nm, and the light absorption intensity gradually decreases. Similarly, the light absorption intensity at another characteristic peak of ferrous Cyt c at 505 nm also gradually decreases with the prolongation of time. This indicates that CeVO4 can catalyze the reaction between ferrous Cyt c and O2, consuming O2.

[0100] When the electron-donating efficiency of Cyt c is not high, the stepwise reduction of O2 generates O2−, H2O2, and ·OH through 1, 2, and 3 electron reductions respectively. Therefore, an ultraviolet / visible spectrophotometer and a fluorescence spectrophotometer were used to analyze the possible products generated. Based on the WST-1 method, it was determined whether O2− was produced when O2 underwent 1e reduction. - As shown in, when O2− exists, the dye (2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt) (WST-1) can react with O2− to generate formazan, which has a maximum ultraviolet absorption peak at approximately 440 nm. However, when CeVO4 interacts with Cyt c, there is no maximum absorption peak at 440 nm, indicating that no O2− is produced. In addition, the method of specifically binding H2O2 with the fluorescent red dye (Amplex Red) to produce fluorescence was also used to exclude the possibility of releasing H2O2 through 2e reduction (as shown in). When H2O2 exists, horseradish peroxidase (HRP) interacts with Amplex Red to obtain the fluorescent product resorufin, which has a strong fluorescence intensity when measured at an excitation wavelength of 571 nm. However, when CeVO4, Cyt c, and CeVO4 + Cyt c interact with HRP and Amplex Red respectively, almost no fluorescent product resorufin is produced. There are significant differences between the three experimental groups without fluorescent products and the control group with fluorescent products (n = 3, ****P < 0.0001), indicating that no H2O2 is produced in the three experimental groups of CeVO4, Cyt c, and CeVO4 + Cyt c. To further verify the possibility of ·OH generation, terephthalic acid (TPA) was used to monitor its fluorescence change. TPA reacts with ·OH to generate 2-hydroxyterephthalic acid, which emits fluorescence at 425 nm under 315 nm light excitation. - as Figure 8A shown, when O2− exists - , the dye (2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt) (WST-1) can react with O2− - to generate formazan, which has a maximum ultraviolet absorption peak at approximately 440 nm. However, when CeVO4 interacts with Cyt c, there is no maximum absorption peak at 440 nm, indicating that no O2− is produced. - In addition, the method of specifically binding H2O2 with the fluorescent red dye (Amplex Red) to produce fluorescence was also used to exclude the possibility of releasing H2O2 through 2e reduction (as shown in Figure 8B ). When H2O2 exists, horseradish peroxidase (HRP) interacts with Amplex Red to obtain the fluorescent product resorufin, which has a strong fluorescence intensity when measured at an excitation wavelength of 571 nm. However, when CeVO4, Cyt c, and CeVO4 + Cyt c interact with HRP and Amplex Red respectively, almost no fluorescent product resorufin is produced. There are significant differences between the three experimental groups without fluorescent products and the control group with fluorescent products (n = 3, ****P < 0.0001), indicating that no H2O2 is produced in the three experimental groups of CeVO4, Cyt c, and CeVO4 + Cyt c. To further verify the possibility of ·OH generation, terephthalic acid (TPA) was used to monitor its fluorescence change. TPA reacts with ·OH to generate 2-hydroxyterephthalic acid, which emits fluorescence at 425 nm under 315 nm light excitation. Figure 8C The reaction between the ·OH radical generated by treating H2O2 with Fe 2+ showed an increase in fluorescence, but no increase in fluorescence was observed when CeVO4 interacted with Cyt c, indicating that no ·OH was produced when CeVO4 interacted with Cyt c. Figure 8ANo O2 is generated when CeVO4 interacts with Cyt c - 。 Figure 8B No H2O2 is generated when CeVO4 interacts with Cyt c. Figure 8C No ·OH is generated when CeVO4 interacts with Cyt c. The above results confirm that CeVO4 catalyzes Cyt c to completely reduce O2 to water without generating partially reduced oxygen (O2 - 、H2O2 and ·OH).

[0101] (3) Targeting study of CeVO4@AQ4N-lip-R110 / MUC1

[0102] To study the targeting of CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles to MCF-7 breast cancer cells, MDA-MB-231 breast cancer cells were used as a control experimental group. Since the hypoxia-activated prodrug AQ4N in the nanoparticles prepared in Example 1 has red fluorescence, the targeting of the nanoparticles was studied based on the red fluorescence of the drug. Figure 9 Figure A in shows the fluorescence images of MDA-MB-231 cells co-incubated with CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles for 1 h, 2 h, and 3 h, respectively. Figure 9 Figure B in shows the fluorescence images of MCF-7 cells co-incubated with CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles for 1 h, 2 h, and 3 h, respectively. Figure 9 The scale bars in both are: 20 μm. By Figure 9 comparing the MDA-MB-231 breast cancer cells in Figure A in Figure 9 with the MCF-7 breast cancer cells in Figure B in, it was found that within the same time period, the red fluorescence intensity of MCF-7 breast cancer cells was significantly stronger than that of MDA-MB-231 breast cancer cells. The experimental data show that more CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles entered MCF-7 breast cancer cells than MDA-MB-231 breast cancer cells. This is because the MUC1 aptamer of CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles targets MCF-7 breast cancer cells, resulting in more nanoparticles entering MCF-7 breast cancer cells than MDA-MB-231 breast cancer cells. The results show that CeVO4@AQ4N-lip-R110 / MUC1 nanoparticles have targeting to MCF-7 breast cancer cells.

[0103] (4) Study on the exacerbation of tumor hypoxia by CeVO4 nanoparticles

[0104] To further study the consumption of oxygen in cancer cells by nanomaterials, in this invention, MCF-7 cells were treated with CeVO4 nanozymes at different concentrations (0 μg / mL, 25 μg / mL), and the judgment was made based on the oxygen quenching fluorescence of RDPP. Figure 10 Figure A in Figure 10 shows the intracellular O2 levels reported by RDPP staining after MCF-7 cells were co-incubated with CeVO4 at 0 μg / mL respectively. Figure 10 Figure B in Figure 10 shows the intracellular O2 levels reported by RDPP staining after MCF-7 cells were co-incubated with CeVO4 at 25 μg / mL respectively. Figure 10 The scale bar in both is: 50 μm. From Figure 10 From the fluorescence data in Figure A in Figure 10 , it can be seen that when CeVO4 at 0 μg / mL was co-incubated with MCF-7 cells, the fluorescence intensity of RDPP was not shown, indicating that the oxygen in the cells quenched the fluorescence of RDPP. And when CeVO4 at 25 μg / mL was co-incubated with MCF-7 cells ( Figure 10 Figure B in Figure 10 ), obvious red fluorescence could be seen, indicating that the fluorescence of RDPP was not quenched, that is, under the catalysis of CeVO4 nanozyme, the intracellular oxygen level decreased, thus exacerbating the hypoxia of tumor cells.

[0105] (5) Investigation of the cytotoxicity of CeVO4@AQ4N-lip-R110 / MUC1

[0106] To further study the cytotoxic effect of nanomaterials on cancer cells, in this invention, an MTT detection kit was used to detect and record the cytotoxicity of CeVO4, AQ4N, and CeVO4@AQ4N-lip-R110 / MUC1. Figure 11A shows the cell survival rates after MCF-7 cells were co-incubated with different concentrations of CeVO4 respectively. Figure 11B shows the cell survival rates after MCF-7 cells were co-incubated with different concentrations of AQ4N respectively. Figure 11C shows the cell survival rates after MCF-7 cells were co-incubated with different concentrations of CeVO4@AQ4N respectively. Figure 11D shows the cell survival rates after MCF-7 cells were co-incubated with different concentrations of CeVO4@AQ4N-lip-R110 respectively. Figure 11E shows the cell survival rates after MCF-7 cells were co-incubated with different concentrations of CeVO4@AQ4N-lip-R110 / MUC1 respectively. From Figure 11AIt can be seen from the data in [[]] that after adding CeVO4 at different concentrations (0 μg / mL, 25 μg / mL, 50 μg / mL, 100 μg / mL, 200 μg / mL), with the increase in the concentration of CeVO4, the cell viability is better. This is because the CeVO4 nanozyme catalyzes the interaction between Cytc and O2 in cells without generating partially reduced oxygen (O2 - , H2O2 and ·OH), but directly generates H2O, indicating that CeVO4 has good biosafety. Figure 11B It was observed from the data in [[]] that with the increase in the concentration of AQ4N (0 μg / mL, 25 μg / mL, 50 μg / mL, 100 μg / mL, 200 μg / mL), the cell viability was not significantly different from that of CeVO4. This is because the slightly hypoxic environment of the cells themselves cannot fully activate the hypoxia-activated prodrug AQ4N to exert its effect, so the cell survival rate is still relatively high. Figure 11C It can be seen that when CeVO4 and the hypoxia drug AQ4N act together, with the increase in the concentration of CeVO4@AQ4N (0 μg / mL, 25 μg / mL, 50 μg / mL, 100 μg / mL, 200 μg / mL), the cell viability decreases significantly. This is because under the action of CeVO4 nanoparticles, the oxygen in the cells is consumed, thereby activating the hypoxia-activated prodrug AQ4N to exert its effect, damaging DNA and killing tumor cells. With the increase in the concentration of CeVO4@AQ4N-lip-R110 (0 μg / mL, 25 μg / mL, 50 μg / mL, 100 μg / mL, 200 μg / mL), Figure 11D the cell viability of [[]] is compared with Figure 11C the cell viability of [[]] and is effectively reduced. Since cytochrome c in cells mainly aggregates in mitochondria, after connecting R110 targeting mitochondria, CeVO4@AQ4N-lip-R110 accumulates in mitochondria, enabling CeVO4 to act with cytochrome c in mitochondria, thereby increasing the oxygen consumption, exacerbating the hypoxic environment of tumor cells, enabling the hypoxia-activated prodrug AQ4N to exert its efficacy to a greater extent, damaging DNA and killing tumor cells. It can be seen from Figure 11E that with the continuous increase in the concentration of CeVO4@AQ4N-lip-R110 / MUC1 prepared in Example 1 (0 μg / mL, 25 μg / mL, 50 μg / mL, 100 μg / mL, 200 μg / mL), compared with Figure 11D the cell viability decreases. This is because MUCI is an aptamer targeting MCF-7 and is not toxic to cells, but it increases the probability of nanoparticles entering cells.

[0107] In summary, the present invention provides a CeVO4@AQ4N-lip-R110 / MUC1 nanoparticle for treating tumor cells based on a hypoxic environment. The MCF-7 breast cancer cells are specifically targeted by the MUC1 aptamer, enabling the nanoparticles to smoothly enter the cells. R110 targets the mitochondria, and the interaction between CeVO4 and Cytc in the mitochondria consumes the oxygen in the cells, exacerbating the hypoxia in the tumor cells, thereby activating the hypoxia-activated prodrug AQ4N to be converted into the toxic AQ4, damaging the DNA, and ultimately causing cell death.

[0108] The research work of this invention is funded by the Natural Science Foundation of Shandong Province (Project No.: ZR2023JQ004), and the project name is: Nanoprobe and Cell Imaging.

[0109] Although the above embodiments have described the present invention in detail, they are only a part of the embodiments of the present invention, rather than all embodiments. Other embodiments can also be obtained based on these embodiments without creative efforts, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A cerium vanadate targeted nanoparticle, characterized in that, It includes a drug core, a protective shell layer wrapping the drug core; and a targeting material modified on the outer surface of the protective shell; The drug core includes cerium vanadate nanoparticles, and a hypoxia-activated prodrug loaded on the surface of the cerium vanadate nanoparticles by electrostatic adsorption, and the hypoxia-activated prodrug is AQ4N; The protective shell layer is a cationic liposome layer, and the cationic liposome layer is a lipid bilayer formed by 1,2-dioleoyl-sn-glycero-3-phosphate, dipalmitoylphosphatidylcholine, cholesterol, DSPE-PEG 5000 and (2,3-dioleoyl-propyl)-trimethylammonium chloride; The targeting material includes a mitochondrial targeting material and a tumor cell targeting material. The mitochondrial targeting material is rhodamine 110; the tumor cell targeting material is a MUC1 nucleic acid aptamer targeting MCF-7 breast cancer cells, and the nucleotide sequence of the MUC1 nucleic acid aptamer targeting MCF-7 breast cancer cells is shown in SEQ ID NO.

1.

2. The cerium vanadate targeted nanoparticles according to claim 1, characterized in that, The cerium vanadate nanoparticles are in the shape of a cube, and the average particle size of the cerium vanadate nanoparticles is 20 nm.

3. The preparation method of the cerium vanadate targeted nanoparticles according to claim 1 or 2, characterized in that, It includes the following steps: Mix cerium vanadate nanoparticles, a hypoxia-activated prodrug and water for electrostatic adsorption to obtain a drug core; Mix the drug core, organic solvent, 1,2-dioleoyl-sn-glycero-3-phosphate, (2,3-dioleoyl-propyl)-trimethylammonium chloride, cholesterol, DSPE-PEG 5000 , and dipalmitoylphosphatidylcholine, and then successively remove the solvent and hydrate to obtain a drug core wrapped by a cationic liposome layer; Mix the drug core wrapped by the cationic liposome layer, the mitochondrial targeting material and a solvent for the first modification to obtain a mitochondrial targeting material modified product; Mix the mitochondrial targeting material modified product, the tumor cell targeting material and a solvent for the second modification to obtain the cerium vanadate targeted nanoparticles.

4. The preparation method according to claim 3, characterized in that, The preparation method of the cerium vanadate nanoparticles includes the following steps: Drop a cerium salt aqueous solution into a metavanadate aqueous solution for a precipitation reaction to obtain the cerium vanadate nanoparticles; the temperature of the precipitation reaction is 80 °C.

5. The preparation method according to claim 3, characterized in that, The mass ratio of the cerium vanadate nanoparticles to the hypoxia-activated prodrug is 1:

1.

6. The preparation method according to claim 3, characterized in that, The mass ratio of the drug core to 1,2-dioleoyl-sn-glycero-3-phosphate is 10:4; the mass ratio of the drug core to (2,3-dioleoyl-propyl)-trimethylammonium chloride is 10:12; the mass ratio of the drug core to cholesterol is 10:2; the mass ratio of the drug core to DSPE-PEG 5000 is 1:1; the mass ratio of the drug core to dipalmitoylphosphatidylcholine is 10:4; The temperature of hydration is 60 °C, and the time is 40 - 55 min.

7. Use of the cerium vanadate targeted nanoparticles according to claim 1 or 2 or the cerium vanadate targeted nanoparticles prepared by the preparation method according to any one of claims 3 - 6 in the preparation of an anti-tumor drug.

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