A sono / chemodynamic synergistic anti-tumor nanomedicine and its preparation method and application

Abstract

The present invention relates to the field of nanomedicine technology, and in particular to a sono / chemodynamic synergistic anti-tumor nanomedicine, its preparation method and application. The present invention provides a sono / chemodynamic synergistic anti-tumor nanomedicine, which uses hollow mesoporous MnO2 nanoparticles as a carrier and is loaded with vitamin C and sonosensitizer dihydrochlorin e6. Vc can produce abundant H2O2, providing raw materials for CDT, while MnO2, as a peroxidase-like enzyme, catalyzes H2O2 to produce O2, thereby effectively alleviating tumor hypoxia and enhancing the effect of sonodynamic therapy. In addition, GSH induces the degradation of MnO2, resulting in GSH consumption and enhanced CDT. The depletion of GSH leads to a decrease in the activity of glutathione peroxidase 4 (GPX4). The production of ROS and the decrease in GPX4 activity synergistically increase lipid peroxides (LPOs), ultimately leading to ferroptosis of tumor cells.

Description

Technical Field

[0001] The present invention relates to the technical field of nanomedicines, and in particular to a sono / chemodynamic synergistic anti-tumor nanomedicine and a preparation method and application thereof. Background Art

[0002] The information disclosed in this background technology section is only intended to enhance understanding of the overall background of the invention and should not necessarily be regarded as an admission or any form of suggestion that the information constitutes the prior art already known to those skilled in the art.

[0003] Malignant tumors have become a major public health problem that poses a serious threat to public health. Conventional treatments for malignant tumors currently include surgery, radiotherapy, chemotherapy, and immunotherapy, but these all have limitations. Therefore, non-invasive, highly targeted, and low-to-no toxicity treatment strategies are urgently needed.

[0004] Sonodynamic therapy (SDT) is a new non-invasive treatment method that has emerged in recent years. It is based on photodynamic therapy. Unlike the latter, SDT uses ultrasound instead of laser as the source of therapeutic energy, overcoming the limitations of laser penetration depth and the skin phototoxicity associated with traditional photosensitizers. Currently, it is believed that the possible mechanisms of SDT in treating tumors mainly include reactive oxygen species theory, ultrasonic cavitation effect, and thermal damage. (1) Reactive oxygen species theory: Ultrasound excites sonosensitizers to produce singlet oxygen with biological toxicity, which increases the level of reactive oxygen species inside tumor cells, leading to irreversible damage to tumor cells / tissues; (2) Ultrasonic cavitation effect: When ultrasound acts on the cell membrane surface, the sound energy generated by ultrasound can rapidly increase mechanical pressure, promote the generation of microbubbles in tissue fluid, and the microbubbles around the cells generate mechanical effects through their oscillation, instantly forming micropores in the cell membrane, thereby increasing vascular permeability and drug transport capacity; (3) Thermal damage: The absorption and conversion of ultrasonic mechanical energy induces the generation of thermal energy during its propagation, causing the tissue temperature to rise and thus leading to the death of cancer cells. As a non-invasive therapeutic strategy for killing tumor cells, SDT uses low-intensity ultrasound to activate sonosensitizers, thereby mediating the production of 1 O2 to kill tumor cells.

[0005] Unlike SDT, chemodynamic therapy (CDT) is a therapeutic strategy that utilizes the Fenton or Fenton-like reaction to catalyze the in situ reaction of hydrogen peroxide (H2O2) to produce highly cytotoxic hydroxyl radicals to kill cancer cells. CDT can occur autonomously and does not rely on external energy intervention. However, the limited endogenous reaction substrates in tumors restrict the effective implementation of CDT. When combined with SDT, the deep diffusion of nanomedicines and the amplified production of reactive oxygen species can enhance the therapeutic effect on tumors. In addition, previous research results have also shown that the introduction of ultrasound can enhance the activity of the Fenton reaction and thus amplify the efficiency of CDT. Therefore, the synergistic effect of SDT and CDT to improve treatment efficiency is of great significance and has broad clinical application prospects. Summary of the Invention

[0006] In order to overcome the above problems, the present invention provides a sono / chemodynamic synergistic anti-tumor nanomedicine and its preparation method and application.

[0007] In order to achieve the above technical objectives, the present invention adopts the following technical solutions:

[0008] The first aspect of the present invention provides a sono / chemodynamic synergistic anti-tumor nanomedicine, which uses hollow mesoporous MnO2 nanoparticles as a carrier and loads vitamin C (Vc) and sonosensitizer dihydrochlorin e6 (Ce6).

[0009] The second aspect of the present invention provides a method for preparing the above-mentioned sono / chemodynamic synergistic anti-tumor nanomedicine, comprising:

[0010] (1) Silica nanoparticles were prepared using tetraethyl orthosilicate as a raw material, and then KMnO4 solution was added to the silica nanoparticle solution to obtain SiO2@MnO2 nanoparticles. The SiO2@MnO2 nanoparticles were used as a template to remove SiO2 and obtain hollow mesoporous MnO2 nanospheres;

[0011] (2) dispersing the hollow mesoporous MnO2 nanospheres in a vitamin C solution and stirring the solution in the dark to obtain the nanodrug intermediate MnO2-Vc;

[0012] (3) The sonosensitizer dihydrochlorin e6 was modified with polyethylene glycol to obtain mPEG-Ce6. After mixing mPEG-Ce6 with the nanodrug intermediate MnO2-Vc, the mixture was stirred in the dark to obtain the nanodrug MnO2-Vc@mPEG-Ce6.

[0013] The third aspect of the present invention provides the use of the above-mentioned sono / chemodynamic synergistic anti-tumor nanomedicine in the preparation of anti-tumor drugs.

[0014] The fourth aspect of the present invention provides an anti-tumor drug composition comprising the above-mentioned sono / chemodynamic synergistic anti-tumor nanomedicine.

[0015] The beneficial effects of the present invention are:

[0016] (1) The present invention provides a nanomedicine with sonodynamic / chemodynamic synergistic anti-tumor effect, which uses hollow mesoporous MnO2 nanoparticles as carriers and is loaded with vitamin C and sonosensitizer dihydrochlorin e6. The characteristics of the tumor microenvironment are usually hypoxia, relatively insufficient hydrogen peroxide and excessive glutathione (GSH), which accelerate the progression and metastasis of tumor cells. The nanomedicine provided by the present invention can play a catalytic function in sequence according to the specific tumor microenvironment to improve the efficiency of ROS generation. Specifically, Vc can produce abundant H2O2, which provides raw materials for CDT, while MnO2, as a peroxidase-like enzyme, catalyzes H2O2 to produce O2, thereby effectively alleviating tumor hypoxia and enhancing the effect of sonodynamic therapy. In addition, GSH induces the degradation of MnO2, resulting in GSH consumption and enhanced CDT. The depletion of GSH will lead to a decrease in the activity of glutathione peroxidase 4 (GPX4). The generation of ROS and the decrease in GPX4 activity synergistically increase lipid peroxides (LPOs), ultimately leading to ferroptosis of tumor cells. Therefore, the nanomedicine provided by the present invention can enable sonodynamics and chemodynamics to synergistically fight tumors, induce ferroptosis of tumor cells, and thus inhibit tumor growth.

[0017] (2) Compared with other H2O2 precursors, Vc has good water solubility and is therefore more easily absorbed by the body, thereby improving its bioavailability. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] The accompanying drawings, which constitute a part of the present invention, are used to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations on the present invention.

[0019] Figure 1 The particle size (a) and pore size (b) of the hollow mesoporous MnO2 spheres in Example 1;

[0020] Figure 2 This is the transmission electron microscopy (TEM) image of the nanodrug MnO2-Vc@mPEG-Ce6;

[0021] Figure 3 The graph showing the change in the amount of O2 produced by the decomposition of H2O2 catalyzed by nanomedicine MnO2-Vc@mPEG-Ce6 under different concentrations (A) and different pH (B) conditions.

[0022] Figure 4The ESR signals (A) of ·OH generated by different treatment groups were detected by using DMPO as a capture agent. 1 ESR signal generated by O2 (B);

[0023] Figure 5 The consumption of GSH (1mM) by nanodrug MnO2-Vc@mPEG-Ce6 at different concentrations;

[0024] Figure 6 This is a graph showing the uptake and phagocytosis test of nanomedicine MnO2-Vc@mPEG-Ce6 by tumor cells (PC-3 cells);

[0025] Figure 7 The cell killing ability experiment of tumor cells (PC-3) under different experimental conditions; from left to right: 1) control group; 2) MnO2 (200ppm) group; 3) Ce6 (200ppm) + ultrasound (1w / cm 2 , 3min) group; 4) MnO2-Vc@mPEG-Ce6 (200ppm) group; 5) MnO2-Vc@mPEG-Ce6 (200ppm) + ultrasound (1w / cm 2 , 3min) group results;

[0026] Figure 8 Fluorescence images of tumor-bearing mice at different time points after intravenous injection of MnO2-Vc@mPEG-Ce6 (A) and in vitro photographs and fluorescence images of the heart, liver, spleen, lung, kidney, and tumor of mice 24 hours after intravenous injection (B);

[0027] Figure 9 Schematic diagram of the operation of tumor-bearing mice injected with MnO2-Vc@mPEG-Ce6 via the tail vein and treated with ultrasound within 24 hours (A); curves showing the changes in mouse body weight (B) and tumor volume (C) over time after treatment with different methods;

[0028] Figure 10 The GPX4 immunohistochemistry images of mouse tumor sections after treatment with different methods are as follows: 1) control group; 2) MnO2 (200 ppm) group; 3) Ce6 (200 ppm) + ultrasound (1 w / cm 2 , 3min) group; 4) MnO2-Vc@mPEG-Ce6 (200ppm) group; 5) MnO2-Vc@mPEG-Ce6 (200ppm) + ultrasound (1w / cm 2 , 3min) group results. DETAILED DESCRIPTION

[0029] It should be noted that the following detailed descriptions are exemplary and intended to provide further explanation of the present invention. Unless otherwise specified, all technical and scientific terms used in the present invention have the same meanings as those commonly understood by those skilled in the art to which the present invention belongs.

[0030] It should be noted that the terms used herein are only for describing specific embodiments and are not intended to limit the exemplary embodiments according to the present invention. As used herein, unless the context clearly indicates otherwise, the singular form is intended to include the plural form. In addition, it should be understood that when the terms "comprise" and / or "include" are used in this specification, they indicate the presence of features, steps, operations, devices, components and / or combinations thereof.

[0031] The first typical embodiment of the present invention provides a sono / chemodynamic synergistic anti-tumor nanomedicine, which uses hollow mesoporous MnO2 nanoparticles as a carrier and loads vitamin C (Vc) and sonosensitizer dihydrochlorin e6 (Ce6).

[0032] In one or more embodiments, the MnO2 nanoparticles are MnO2 nanospheres;

[0033] Preferably, the particle size of the MnO2 nanospheres is about 190 nm; and the pore size is about 3.9 nm.

[0034] A second typical embodiment of the present invention provides a method for preparing the above-mentioned sono / chemodynamic synergistic anti-tumor nanomedicine, comprising:

[0035] (1) Silica nanoparticles were prepared using tetraethyl orthosilicate as a raw material, and then KMnO4 solution was added to the silica nanoparticle solution to obtain SiO2@MnO2 nanoparticles. The SiO2@MnO2 nanoparticles were used as a template to remove SiO2 and obtain hollow mesoporous MnO2 nanospheres;

[0036] (2) dispersing the hollow mesoporous MnO2 nanospheres in a vitamin C solution and stirring the solution in the dark to obtain the nanodrug intermediate MnO2-Vc;

[0037] (3) The sonosensitizer dihydrochlorin Ce6 was modified with polyethylene glycol to obtain mPEG-Ce6. After mixing mPEG-Ce6 with the nanodrug intermediate MnO2-Vc, the mixture was stirred in the dark to obtain the nanodrug MnO2-Vc@mPEG-Ce6.

[0038] In one or more embodiments, in step (1), the molar ratio of silica nanoparticles to KMnO4 is 2.5 to 3:1, preferably 2.85:1.

[0039] In one or more embodiments, in step (1), the concentration of the KMnO4 solution is 0.015 to 0.03 mmol / L, preferably 0.02 mmol / L.

[0040] In one or more embodiments, in step (1), SiO2@MnO2 nanoparticles are obtained by ultrasound, and the ultrasonic power is 140-160W, preferably 150W; and the time is 5-7h, preferably 6h.

[0041] In one or more embodiments, in step (1), Na2CO3 is used to etch away SiO2, and the concentration of Na2CO3 is 0.015 to 0.03 mmol / L, preferably 0.02 mmol / L; the etching temperature is 55 to 65°C, preferably 60°C; the etching time is 10 to 15 hours, preferably 12 hours; and the stirring rate during etching is 120 to 200 rpm, preferably 150 rpm.

[0042] In one or more embodiments, in step (2), the mass ratio of hollow mesoporous MnO2 nanospheres to vitamin C is 0.03 to 0.01:1, preferably 0.05:1.

[0043] In one or more embodiments, in step (2), the concentration of vitamin C in the vitamin C solution is 1 to 3 g / L, preferably 2 g / L.

[0044] In one or more embodiments, in step (2), the temperature of the reaction under stirring in the dark is room temperature, and the stirring time is 10 to 15 hours, preferably 12 hours.

[0045] In one or more embodiments, in step (3), the mass ratio of Ce6 to MnO2 is 8 to 12:1, preferably 10:1.

[0046] In one or more embodiments, in step (3), the temperature of the reaction under stirring in the dark is room temperature, and the stirring time is 20 to 30 hours, preferably 24 hours.

[0047] A third typical embodiment of the present invention provides the use of the above-mentioned sono / chemodynamic synergistic anti-tumor nanomedicine in the preparation of anti-tumor drugs.

[0048] A fourth typical embodiment of the present invention provides an anti-tumor drug composition comprising the above-mentioned sono / chemodynamic synergistic anti-tumor nanomedicine.

[0049] In one or more embodiments, the anti-tumor pharmaceutical composition further includes drugs related to tumor treatment.

[0050] Preferably, the tumor treatment-related drug is an anti-tumor drug, an immune adjuvant, a checkpoint inhibitor or an antigen protein.

[0051] In one or more embodiments, the tumor includes benign tumors and / or malignant tumors; the malignant tumors include solid tumors and hematological tumors.

[0052] It should be noted that the term "tumor" as used herein, as known to those skilled in the art, includes benign and / or malignant tumors. Benign tumors are defined as excessive cell proliferation that is incapable of forming aggressive, metastatic tumors in the body. Conversely, malignant tumors are defined as cells with multiple cellular and biochemical abnormalities that are capable of developing systemic diseases (e.g., metastasis to distant organs).

[0053] Solid tumors such as breast, bladder, bone, brain, central and peripheral nervous system, colon, endocrine glands (such as thyroid and adrenal cortex), esophagus, endometrium, germ cells, head and neck, kidney, liver, lung, larynx and hypopharynx tumors, mesothelioma, ovary, pancreas, prostate, rectum, kidney, small intestine, soft tissue, testicle, stomach, skin (such as melanoma), ureter, vagina and vulva tumors. Malignant tumors include hereditary cancers, such as retinoblastoma and Wilms tumor (Wilms tumor). In addition, malignant tumors include primary tumors in the organs and corresponding secondary tumors (tumor metastasis) in distal organs. Hematologic malignancies such as aggressive and indolent forms of leukemia and lymphoma, i.e. non-Hodgkin's disease, chronic and acute myeloid leukemia (CML / AML), acute lymphocytic leukemia (ALL), Hodgkin's disease, multiple myeloma and T-cell lymphoma. Also included are myelodysplastic syndromes, plasmacytomas, tumor-like syndromes, cancers of unknown primary site, and AIDS-related malignancies.

[0054] In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0055] Example 1:

[0056] Formation of manganese dioxide (MnO2) nanoparticles:

[0057] First, silicon dioxide (SiO2) nanoparticles were prepared using tetraethyl orthosilicate (TEOS) as the raw material. 0.75 mL of TEOS was quickly added to a mixture containing ethanol (25 mL), ultrapure water (0.5 mL), and ammonia (1.8 mL). The mixture was stirred at 40°C and 150 rpm for 12 hours to produce 80±5 nm SiO2 nanoparticles. Then, 47 mL of KMnO4 solution (0.02 mmol / L) was slowly added dropwise to the SiO2 nanoparticle solution under ultrasonic conditions at 150 W / 40 kHz. After 6 hours of ultrasonic reaction, the solution was centrifuged three times (14,800 rpm, 10 minutes) and the supernatant was discarded to obtain a precipitate. SiO2@MnO2 nanoparticles were precipitated by washing and centrifugation. In order to remove SiO2, the prepared SiO2@MnO2 nanoparticles were mixed with Na2CO3 (2mmol / L) and reacted under constant magnetic stirring conditions of 60° and 150rpm for 12h. The precipitate was then washed by centrifugation and dissolved in 40mL of ultrapure water to obtain 2mg / mL hollow mesoporous MnO2 spheres. The particle size and pore size of the hollow mesoporous MnO2 spheres are shown in Figure 2. Figure 1 As shown, from Figure 1 It can be seen that the average hydrodynamic diameter of the hollow mesoporous MnO2 spheres is about 190.1nm; using the BJH method to analyze the desorption branch curve, it was determined that the pore size distribution of the MnO2 nanoparticles is approximately concentrated at 3.9nm.

[0058] Example 2

[0059] Preparation of intermediate MnO2-Vc:

[0060] The prepared MnO2 nanoparticles (0.6 mg) were added to 5 mL of ultrapure water containing 10 mg of Vc, and then the mixture was sonicated for 5 min and stirred in the dark for 2 h to obtain a MnO2-Vc solution.

[0061] Example 3

[0062] Preparation of nanomedicine MnO2-Vc@mPEG-Ce6:

[0063] In order to improve the water solubility and physiological stability of Ce6, it was modified with PEG to obtain mPEG-Ce6.

[0064] 60 mg of Ce6 dissolved in 10 mL of DMSO was stirred with 19 mg of EDAC (1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride) and 12 mg of NHS (N-hydroxysuccinimide) and 200 mg of mPEG for 24 hours. The resulting solution was lyophilized, and the lyophilized solid was mixed with chloroform to remove unreacted Ce6. The precipitate was then removed by filtration, and the solvent was removed on a rotary evaporator to obtain mPEG-Ce6. The relative molecular mass of mPEG is 2000.

[0065] 1 mL of the prepared mPEG-Ce6 solution was mixed with 5 mL of the MnO2-Vc solution, ultrasonicated for 5 minutes, stirred in the dark for 24 hours, and then centrifuged to remove the supernatant. This resulted in the MnO2-Vc@mPEG-Ce6 nanoparticles. The resulting product was suspended in 10 mL of ultrapure water and stored at 4°C.

[0066] Transmission electron microscopy (TEM) was used to characterize the morphology of the nanomedicine MnO2-Vc@mPEG-Ce6. Figure 2 As shown, the prepared nanoparticles have spherical morphology, porous structure and good dispersibility.

[0067] Example 4

[0068] A portable dissolved oxygen meter was used to measure the O2 content released by the nanomedicine MnO2-Vc@mPEG-Ce6 and H2O2 under different conditions to verify that MnO2-Vc@mPEG-Ce6 can effectively alleviate tumor hypoxia. Figure 3 As shown in Figure A, after adding MnO2-Vc@mPEG-Ce6 to the H2O2 solution, O2 was produced, and the amount of dissolved O2 increased with the increase of MnO2-Vc@mPEG-Ce6 concentration, indicating that MnO2-Vc@mPEG-Ce6 has catalyst-like activity and can catalyze the decomposition of H2O2 to produce O2. At the same time, in another set of experiments, the concentration of MnO2-Vc@mPEG-Ce6 was fixed at 200ppm and the pH value of the solution was changed. The results are shown in Figure A. Figure 3 B shows that under acidic conditions, MnO2-Vc@mPEG-Ce6 is more efficient in catalyzing H2O2 to produce O2. In the acidic and hypoxic environment within the tumor, the addition of MnO2-Vc@mPEG-Ce6 can significantly increase the O2 production rate.

[0069] Example 5

[0070] Electron spin resonance (ESR) was performed in 25 mM NaHCO3 / CO2 buffer using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a capture agent to verify the Mn 2+ The ·OH generation capacity. Figure 4As shown in A, a typical ·OH signal (1:2:2:1) was found. Since GSH has the ability to remove ROS, Mn 2+ The peak of +H2O2+GSH group is weakened. 2,2,6,6-tetramethyl-4-piperidone (TEMP) is used as a trapping agent and detected by ESR. 1 The production of O2. Figure 4 As shown in B, in all cases exposed to ultrasound (1W / cm 2 , 10 minutes) group were observed to have typical 1 The O2 signal (1:1:1) was significantly enhanced after H2O2 treatment. The results showed that the nanomedicine MnO2-Vc@mPEG-Ce6 had a high ROS production.

[0071] Example 5

[0072] 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) was used to detect the ability of MnO2-Vc@mPEG-Ce6 to consume GSH. Different concentrations of MnO2-Vc@mPEG-Ce6 (0, 50, 100, 200, 300, 400 and 500 μg / mL) were co-incubated with GSH for 30 minutes, and the consumption of GSH (1 mM) by MnO2-Vc@mPEG-Ce6 was measured. Then, DTNB (500 μM) was added and the UV-visible spectrum was recorded at a wavelength of 300-550 nm. The results are shown in Figure 2. Figure 5 As shown in Figure 3, the absorption peak intensity at 412 nm decreases with the increase of MnO2-Vc@mPEG-Ce6 concentration. The results indicate that MnO2-Vc@mPEG-Ce6 can act as a GSH depleting agent, which can completely degrade MnO2 in cancer cells by GSH.

[0073] Example 6

[0074] The uptake of MnO2-Vc@mPEG-Ce6 by PC-3 cells was observed using a fluorescence microscope. MnO2-Vc@mPEG-Ce6 was co-incubated with the fluorescent agent FITC overnight to connect MnO2-Vc@mPEG-Ce6 with the fluorescent agent FITC. Then, MnO2-Vc@mPEG-Ce6 containing FITC was added to the culture medium, and the fluorescence signal of FITC was observed under a fluorescence microscope to evaluate the entry of nanomedicine into the cells after 0, 1, 2 and 4 hours. Figure 6 As shown, PC-3 cells can effectively take up the nanodrug MnO2-Vc@mPEG-Ce6 in a time-dependent manner.

[0075] Example 7

[0076] The CAM / PI double staining method was used to detect the toxicity of the nanodrug MnO2-Vc@mPEG-Ce6 on PC-3 cells. In order to analyze the in vitro anti-tumor effect of MnO2-Vc@mPEG-Ce6, the survival and death of PC-3 cells were detected using a CAM / PI double staining kit. PC-3 cells were seeded into six-well plates and allowed to adhere to the wall, and then cultured in a 37°C incubator for 24 hours. The cells were then divided into five treatment groups: 1) control group; 2) MnO2 (200 ppm) group; 3) Ce6 (200 ppm) + ultrasound (1 w / cm 2 , 3min) group; 4) MnO2-Vc@mPEG-Ce6 (200ppm) group; 5) MnO2-Vc@mPEG-Ce6 (200ppm) + ultrasound (1w / cm 2 After the different treatments, the cells were stained with CAM / PI for 15 minutes according to the manufacturer's instructions. Finally, the fluorescence signal of each group of cells was observed under a fluorescence microscope to determine the cell viability. Figure 7 As shown in the figure, the nanodrug MnO2-Vc@mPEG-Ce6 has the strongest killing ability on PC-3 cells under ultrasound.

[0077] Example 8

[0078] First, a PC-3 prostate cancer subcutaneous tumor model was established in BALB / c mice. C-3 cells (1×10 6 ) were suspended in 0.2 mL of normal saline and injected subcutaneously into the left abdomen of mice to establish a tumor model. 3 At the same time, mice were randomly assigned to different treatment groups to prepare for in vivo experiments. First, Cy5.5-loaded MnO2-Vc@mPEG-Ce6 nanoparticles were synthesized to evaluate the biodistribution of MnO2-Vc@mPEG-Ce6 in vivo. 50 μL of Cy5.5-labeled MnO2-Vc@mPEG-Ce6 solution was injected into the tail vein of mice, and the fluorescence images of mice were collected within 24 hours after injection. BALB / c mice were dissected 24 hours after injection, and the fluorescence images of organs and tumors were collected. Figure 8 As shown in Figure A, the fluorescence intensity of the tumor gradually increased over time, completely concentrating at the tumor site 12 hours after injection, and then decreased due to in vivo metabolism (a separate color scale is used due to the higher fluorescence intensity at 0.5 hours). This indicates that the distribution of MnO2-Vc@mPEG-Ce6 is time-dependent, which may be due to enhanced tumor permeability and retention. Tumors and major organs were also removed 24 hours after injection for ex vivo imaging. Figure 8B shows that the tumors of mice treated with MnO2-Vc@mPEG-Ce6 showed the strongest Cy5.5 fluorescence signal, while other resected organs no longer had fluorescence, indicating that MnO2-Vc@mPEG-Ce6 nanoparticles were still enriched in the tumor site 24 h after injection.

[0079] Example 9

[0080] When the tumor size reaches about 100 mm 3 Afterwards, PC-3 tumor-bearing mice were randomly divided into five treatment groups (n=5) and injected with 100 μL of therapeutic drugs through the tail vein, divided into 1) control group; 2) MnO2 (200 ppm) group; 3) Ce6 (200 ppm) + ultrasound (1 w / cm 2 , 3min) group; 4) MnO2-Vc@mPEG-Ce6 (200ppm) group; 5) MnO2-Vc@mPEG-Ce6 (200ppm) + ultrasound (1w / cm 2 The concentration of the drug solution was set at 200 ppm. The body weight and tumor volume of the mice were recorded within 21 days after the different treatments. Figure 9 A shows that the tumors in the control group grew rapidly, while the tumor growth in the Ce6+ultrasound group and the MnO2-Vc@mPEG-Ce6+ultrasound group was inhibited to a certain extent. Among them, the tumor inhibition effect of the MnO2-Vc@mPEG-Ce6+ultrasound group was the best. At the end of the experiment, the weight of the mice was relatively stable without significant changes, and no statistical difference was found between the groups ( Figure 9 B). This indicates that MnO2-Vc@mPEG-Ce6 is biologically safe and has no significant toxic effects on animals. Tumor images of different treatment groups on day 21 further confirm the effectiveness of the synergistic treatment of this nanomedicine.

[0081] Example 10

[0082] To further evaluate the effects of different treatments on tumor tissue, GPX4 antibody was used to stain the sections of tumor tissue removed from each group. Figure 10 As shown, compared with the other treatment groups, the MnO2-Vc@mPEG-Ce6+ultrasound group had the least positive cells (brown), indicating that GPX4 protein expression was downregulated, which proved that the nanodrug MnO2-Vc@mPEG-Ce6 caused ferroptosis of tumor cells through the GPX4 pathway.

[0083] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention shall be included within the scope of protection of the present invention.

Claims

1. A sono / chemodynamic synergistic anti-tumor nanomedicine, characterized in that: The nanomedicine uses hollow mesoporous MnO2 nanoparticles as a carrier, loaded with vitamin C and sonosensitizer dihydrochlorin e6; The method for preparing the sono / chemical dynamic synergistic anti-tumor nanomedicine comprises: (1) Silica nanoparticles were prepared using tetraethyl orthosilicate as raw material, and then KMnO4 solution was added to the silica nanoparticle solution to obtain SiO2@MnO2 nanoparticles. Using SiO2@MnO2 nanoparticles as a template, SiO2 was removed to obtain hollow mesoporous MnO2 nanospheres; (2) Dispersing the hollow mesoporous MnO2 nanospheres in the vitamin C solution, stirring and reacting in the dark to obtain the nanodrug intermediate MnO2-Vc; (3) The sonosensitizer dihydrochlorin e6 was modified with polyethylene glycol to obtain mPEG-Ce6. After mixing mPEG-Ce6 with the nanodrug intermediate MnO2-Vc, the mixture was stirred in the dark to obtain the nanodrug MnO2-Vc@mPEG-Ce6.

2. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: The MnO2 nanoparticles are MnO2 nanospheres.

3. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 2, characterized in that: The particle size of the MnO2 nanospheres is about 190nm; the pore size is about 3.9nm.

4. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (1), the molar ratio of silica nanoparticles to KMnO4 is 2.5-3:

1.

5. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 4, characterized in that: The molar ratio of silica nanoparticles to KMnO4 was 2.85:

1.

6. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (1), the concentration of the KMnO4 solution is 0.015~0.03mmol / L.

7. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 6, characterized in that: The concentration of KMnO4 solution is 0.02mmol / L.

8. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (1), the ultrasonic power for obtaining SiO2@MnO2 nanoparticles is 140~160 W; the time is 5~7 h.

9. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 8, characterized in that: The ultrasonic power for obtaining SiO2@MnO2 nanoparticles was 150 W and the time was 6 h.

10. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (1), Na2CO3 is used to etch and remove SiO2, and the concentration of Na2CO3 is 0.015~0.03mmol / L; the etching temperature is 55~65℃; the etching time is 10~15h; and the stirring rate during etching is 120~200rpm.

11. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 10, characterized in that: Na2CO3 was used to etch away SiO2, and the concentration of Na2CO3 was 0.02 mmol / L. The etching temperature was 60 ℃, the etching time was 12 h, and the stirring rate during etching was 150 rpm.

12. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (2), the mass ratio of hollow mesoporous MnO2 nanospheres to vitamin C is 0.03~0.1:

1.

13. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 12, characterized in that: The mass ratio of hollow mesoporous MnO2 nanospheres to vitamin C is 0.05:

1.

14. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (2), the concentration of vitamin C in the vitamin C solution is 1-3 g / L.

15. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 14, characterized in that: The concentration of vitamin C in the vitamin C solution is 2 g / L.

16. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (2), the reaction temperature is room temperature and the stirring time is 10 to 15 hours.

17. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 16, characterized in that: The reaction temperature was kept at room temperature and the stirring time was 12 h.

18. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (3), the mass ratio of Ce6 to MnO2 is 8~12:

1.

19. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 18, characterized in that: The mass ratio of Ce6 to MnO2 is 10:

1.

20. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 1, characterized in that: In step (3), the reaction temperature is room temperature and the stirring time is 20 to 30 h.

21. The sono / chemodynamic synergistic anti-tumor nanomedicine according to claim 20, characterized in that: The reaction temperature was kept at room temperature and the stirring time was 24 h.

22. Use of the sono / chemodynamic synergistic anti-tumor nanomedicine according to any one of claims 1 to 21 in the preparation of anti-tumor drugs.

23. An anti-tumor pharmaceutical composition, characterized in that: The invention relates to a sono / chemodynamic synergistic anti-tumor nanomedicine according to any one of claims 1 to 21.

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