A targeted nanoparticle and its preparation method and application

Abstract

The present invention belongs to the technical field of biomedical nanomaterials, and specifically relates to a targeted nanoparticle and a preparation method and application thereof. The targeted nanoparticle DOX@Fe-MSN-GA prepared by the present invention has a liver cancer targeting effect, carries chemotherapy drugs, improves the specificity of chemotherapy drugs, and synergizes chemotherapy and chemodynamic therapy to achieve precise treatment of liver cancer.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedical nanomaterials, and specifically relates to targeted nanoparticles for synergistic treatment of liver cancer by chemotherapy and chemodynamic therapy, and a preparation method and application thereof. Background Art

[0002] Liver cancer is one of the most complex diseases, with an average survival rate of less than 5%, ranking among the top cancer mortality rates worldwide. However, most patients are already in the advanced stage when they are first diagnosed, so it is very difficult to completely eliminate liver tumors using a single treatment method such as surgery, radiotherapy, and chemotherapy. Studies have shown that synergistic treatment of multiple cancer treatment modes can help further enhance anti-tumor efficiency. Nano drug carriers can integrate multiple treatment modes into one, and are expected to achieve a multi-modal synergistic treatment strategy.

[0003] Chemotherapy (CT) is a common and effective clinical treatment strategy for liver cancer. Common chemotherapy drugs include doxorubicin (DOX), camptothecin (CPT), paclitaxel (PTX) and cisplatin (CDDP). These chemotherapy drugs are usually subject to various limitations, such as poor water solubility and short drug circulation time in the body. In addition, the main challenges faced by chemotherapy are non-specific drug administration, low drug concentration in tumor cells, and toxic side effects caused by drug damage to normal cells. Targeted delivery of nano drug carriers can effectively solve the above problems.

[0004] Chemodynamic therapy (CDT) is an emerging strategy. CDT has the advantages of high selectivity and low toxicity, and it can use endogenous chemical energy to produce cytotoxic substances and induce cell death. CDT generally uses nanoparticles to deliver Fenton or Fenton-like reaction catalysts into tumor cells, catalyzing the decomposition of overexpressed hydrogen peroxide in tumor cells to produce hydroxyl radicals and induce cell apoptosis.

[0005] Currently, a large number of nanoparticles use galactose as a liver cancer targeting ligand to deliver drugs to the liver. However, when normal liver lesions are liver cancer, the galactose receptor is inhibited, and the targeting effect of nanoparticles with galactose as an active targeting group is reduced.

[0006] Mesoporous silica nanocarriers (MSNs) have large pore volume and specific surface area, which are conducive to efficient drug loading. The surface is easy to be functionalized and can be used for controlled release and targeted drug delivery systems, which is conducive to enhancing drug efficacy and reducing toxic side effects. Although MSNs have many advantages as nanocarriers, simple MSN drug delivery has no targeting effect, let alone CDT function. Summary of the invention

[0007] In view of the problems faced in the existing chemotherapy of liver cancer, the present invention proposes a targeted nanoparticle and a preparation method thereof. The targeted nanoparticle can synergistically treat liver cancer with chemotherapy and chemodynamic therapy. Specifically, the targeted aggregation of chemotherapy drugs at the liver cancer site can be achieved, and the drug efficacy can be exerted at the liver cancer site and the Fenton reaction can be triggered to convert endogenous H2O2 into hydroxyl radicals (·OH). ·OH is known as the most oxidative reactive oxygen species (ROS), which induces tumor cell apoptosis by damaging DNA and inactivating proteins.

[0008] The first aspect of the present invention provides a method for preparing targeted nanoparticles, comprising the steps of:

[0009] Step 1: Preparation of Fe-MSN nanocarriers;

[0010] Step 2: GA ligand is modified on the surface of Fe-MSN to prepare Fe-MSN-GA;

[0011] Step 3: Fe-MSN-GA is loaded with the anticancer drug DOX to prepare DOX@Fe-MSN-GA, which is the targeted nanoparticle.

[0012] Glycyrrhetinic acid (GA) is a natural compound isolated from licorice, which has a strong affinity for liver cancer cells. The amount of GA receptors in liver cancer tumor tissue is much higher than that in normal tissue. Therefore, the present invention uses small molecule GA as a targeting ligand targeting liver cancer cells.

[0013] The present invention uses MSN as drug-carrying nanoparticles, which have the advantages of excellent biocompatibility, high stability, strong drug-carrying capacity, and easy surface modification.

[0014] In view of the shortcoming that MSN itself has no liver cancer targeting ability, the present invention modifies GA targeting molecules on the surface of MSN to increase the enrichment of nano drug carriers in liver cancer cells, thereby enhancing the efficacy of chemotherapy drugs and reducing toxic side effects.

[0015] In view of the shortcoming that MSN itself has no CDT function, the present invention doped iron during the synthesis process of MSN. The formed Fe-MSN triggers the Fenton reaction (Fe+H2O2→Fe+OH+·OH) in the tumor microenvironment. The highly oxidizing ·OH induces tumor cell apoptosis by damaging DNA and inactivating proteins.

[0016] In step 1, specifically:

[0017] Dissolve hexadecyltrimethylammonium bromide, ferrous sulfate heptahydrate (FeSO4·7H2O), and ferric chloride hexahydrate (FeCl3·6H2O) in pure water, then add ammonia water, stir and react, then add tetraethyl silicate and ethyl acetate and stir and react, centrifuge and wash the reaction turbid liquid after the reaction, then dry it, and finally calcine at high temperature to remove the template hexadecyltrimethylammonium bromide to obtain Fe-MSN. During the synthesis of MSN carriers, iron in different valence states reacts to generate ferroferric oxide, which is doped in situ in the carrier structure, and the formed Fe-MSN triggers the Fenton reaction in the tumor microenvironment.

[0018] Preferably, in step 1, the mass ratio of each raw material is: hexadecyltrimethylammonium bromide: ferrous sulfate heptahydrate: ferric chloride hexahydrate: pure water: ammonia water: tetraethyl silicate: ethyl acetate = 0.1-10: 0.1-1: 0.2-2: 10-100: 0.3-3: 0.05-0.5: 0.5-5, the reaction temperature is 40°C, after adding ammonia water, stir and react at 300rpm for 1-3 hours, then add tetraethyl silicate and ethyl acetate, first stir vigorously at 500rpm for 1 minute, and then stir and react at 150rpm for 4-8 hours.

[0019] Preferably, in step 1, the reaction turbid liquid is first centrifugally washed with pure water until the pH value is 7, and then centrifugally washed with anhydrous ethanol for several times, which has a better dispersion effect.

[0020] Preferably, in step 1, the drying temperature is 60-80°C, and the temperature of high-temperature calcination to remove the template is 500-600°C.

[0021] In step 2, the Fe-MSN surface is modified with GA ligands, and the Fe-MSN surface is first modified with amino groups and then coupled with the carboxyl groups of GA. Specifically:

[0022] Step 2.1: reacting the Fe-MSN prepared in step 1 with a silane coupling agent, aminopropyltriethoxysilane, to obtain Fe-MSN-NH2;

[0023] Step 2.2: In an anhydrous ethanol medium, GA, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N,N-dicyclohexylcarbodiimide are stirred and reacted, and then N-hydroxysuccinimide is added and stirred and reacted to prepare a GA mixed solution;

[0024] Step 2.3 Add Fe-MSN-NH2 ethanol dispersion dropwise to the GA mixed solution obtained in step 2.2 and stir to react to obtain Fe-MSN-GA;

[0025] Preferably, in step 2.1, the reaction medium is anhydrous ethanol, the usage ratio of each raw material is: Fe-MSN: silane coupling agent = 0.1g~1g: 0.1mL~10mL, the reaction temperature is 80~90°C, and the reaction time is 4~8h.

[0026] Preferably, in the step 2.2, the mass ratio of the raw materials is: GA: 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride: N,N-dicyclohexylcarbodiimide: N-hydroxysuccinimide = 1-30: 1.2-12: 1.2-12: 3-30, GA, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N,N-dicyclohexylcarbodiimide are first stirred for reaction for 0.5-2h and then N-hydroxysuccinimide is added for reaction for 3-5h.

[0027] Preferably, in step 2.3, in the reaction solution, the mass ratio of GA to Fe-MSN-NH2 is 1-30:0.1-10, and the reaction time is 3-5h.

[0028] DOX was dissolved in dimethyl sulfoxide, and the DMSO solution of DOX was dropped into the Fe-MSN-GA aqueous solution while ultrasonicating and stirring. After the dropwise addition was completed, the free DOX was removed to obtain DOX@Fe-MSN-GA.

[0029] Step 3 mainly involves encapsulating DOX in Fe-MSN-GA through the carrier mesoporous structure to form DOX@Fe-MSN-GA.

[0030] Preferably, in step 3, the mass ratio of DOX to Fe-MSN-GA is 0.1-10:0.1-10.

[0031] Preferably, in step 3, a dialysis bag with a molecular weight cutoff of 3500Da is used to dialyze and remove free DOX, which has a better removal effect.

[0032] On the other hand, the present invention also provides targeted nanoparticles prepared by the above preparation method. The nanoparticles can be used to treat liver cancer in coordination with chemotherapy and chemodynamic therapy.

[0033] In another aspect, the present invention also provides the use of the above-mentioned targeted nanoparticles in liver cancer treatment drugs.

[0034] The preparation method of the present invention can obtain a targeted nanoparticle for synergistic treatment of liver cancer with chemotherapy and chemodynamic therapy, and its beneficial effects are:

[0035] (1) The nanoparticles prepared by the present invention can carry a variety of chemotherapeutic drugs, have a large drug loading capacity, and have low toxic side effects, thereby improving the bioavailability of chemotherapeutic drugs;

[0036] (2) The nanoparticles prepared by the present invention have a liver cancer targeting effect, carry chemotherapeutic drugs, improve the specificity of chemotherapeutic drugs, and achieve precise treatment;

[0037] (3) The nanoparticles prepared by the present invention have two treatment modes: chemotherapy and chemodynamic therapy. Chemotherapy and chemodynamic therapy synergistically kill liver cancer cells, and the anti-tumor effect is more significant. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 Transmission electron microscopy image of DOX@Fe-MSN-GA;

[0039] Figure 2 The effect of DOX@Fe-MSN-GA on the activity of liver cancer cells (HepG2);

[0040] Figure 3 The effect of DOX@Fe-MSN-GA on reactive oxygen species in liver cancer cells (HepG2);

[0041] Figure 4 The distribution statistics of DOX@Fe-MSN-GA in organs of nude mice with liver cancer model;

[0042] Figure 5 The inhibitory effect of DOX@Fe-MSN-GA on the tumor size of nude mice with liver cancer model;

[0043] Figure 6 The effect of DOX@Fe-MSN-GA on the survival rate of nude mice with liver cancer model. DETAILED DESCRIPTION

[0044] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiment is a module embodiment of the present invention, rather than all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0045] Example 1

[0046] This embodiment provides a targeted nanoparticle for synergistic treatment of liver cancer with chemotherapy and chemodynamic therapy, and a preparation method thereof comprises the steps of:

[0047] Step 1: Preparation of Fe-MSN nanocarriers;

[0048] Add 0.1g hexadecyltrimethylammonium bromide (CTAB), 0.1g ferrous sulfate heptahydrate (FeSO4·7H2O), 0.2g ferric chloride hexahydrate (FeCl3·6H2O) and 10mL pure water to a beaker, stir with a mechanical stirrer and heat to 40°C to completely dissolve CTAB. At a constant temperature of 40°C, add 0.3mL ammonia water and stir at 300rpm for 2 hours, then add 0.05g tetraethyl silicate (TEOS) and 0.5g ethyl acetate, first stir vigorously at 500rpm for 1 minute, then stir at 150rpm for 6 hours, transfer the reaction turbid liquid to a centrifuge tube, centrifuge at 15000rpm for 15min, pour off the supernatant, repeatedly wash the precipitate with pure water and centrifuge until the pH of the supernatant is consistent with that of pure water, then wash the precipitate with anhydrous ethanol and centrifuge twice. Place the precipitate in a 60°C oven for drying, then calcine at 550°C to remove the template CTAB, and obtain Fe-MSN.

[0049] Step 2: GA ligand is modified on the surface of Fe-MSN to prepare Fe-MSN-GA;

[0050] Step 2.1 Add 0.1 g of Fe-MSN to anhydrous ethanol, stir at 85°C, add 0.1 mL of silane coupling agent aminopropyltriethoxysilane (APTES), react for 6 hours, centrifuge after the reaction, wash with anhydrous ethanol, repeat the centrifugation operation 3 times to obtain Fe-MSN-NH2.

[0051] Step 2.2: Dissolve 10 mg of GA in anhydrous ethanol, add 12 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl) and 12 mg of N,N-dicyclohexylcarbodiimide (DCC), stir for 1 hour, add 30 mg of N-hydroxysuccinimide (NHS), stir and react for 4 hours to prepare a GA mixed solution;

[0052] Step 2.3: Add 1 mL of Fe-MSN-NH2 ethanol dispersion (1 mg / mL) dropwise to the GA mixture obtained in step 2.2, and continue the reaction for 4 hours. After the reaction is completed, centrifuge and wash the lower target product with anhydrous ethanol. Repeat the centrifugation operation 3 times to obtain Fe-MSN-GA.

[0053] Step 3: Dissolve 1 mg of DOX in 1 mL of dimethyl sulfoxide (DMSO), disperse 1 mg of Fe-MSN-GA in 1 mL of pure water, and drop the DMSO solution of DOX into the Fe-MSN-GA aqueous solution while stirring under ultrasound. After the addition is completed, transfer the mixed solution to a dialysis bag with a molecular weight cutoff of 3500 Da, and dialyze with pure water to remove free DOX to obtain DOX@Fe-MSN-GA, and the product is freeze-dried for storage.

[0054] Example 2

[0055] This embodiment provides a method for preparing targeted nanoparticles for synergistic treatment of liver cancer with chemotherapy and chemodynamic therapy, which is basically the same as that of Example 1, except for the feeding amounts of the following raw materials:

[0056] In step 1, 10 g of hexadecyltrimethylammonium bromide (CTAB), 1 g of ferrous sulfate heptahydrate (FeSO4·7H2O), 2 g of ferric chloride hexahydrate (FeCl3·6H2O), 100 mL of pure water, 3 mL of ammonia water, 0.5 g of tetraethyl silicate (TEOS), and 5 g of ethyl acetate are added.

[0057] In step 2, 1 g of Fe-MSN, 10 mL of silane coupling agent aminopropyltriethoxysilane (APTES), 0.3 g of GA, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl), 0.12 g of N,N-dicyclohexylcarbodiimide (DCC), 0.3 g of N-hydroxysuccinimide (NHS), and 100 mL of 1 mg / mL Fe-MSN-NH2 ethanol dispersion were prepared.

[0058] In step 3, DOX 10 mg, Fe-MSN-GA 10 mg.

[0059] Example 3

[0060] This embodiment provides a method for preparing targeted nanoparticles for synergistic treatment of liver cancer with chemotherapy and chemodynamic therapy, which is basically the same as that of Example 1, except for the feeding amounts of the following raw materials:

[0061] In step 1, 0.5 g of hexadecyltrimethylammonium bromide (CTAB), 0.2 g of ferrous sulfate heptahydrate (FeSO4·7H2O), 0.3 g of ferric chloride hexahydrate (FeCl3·6H2O), 30 mL of pure water, 2 mL of ammonia water, 0.3 g of tetraethyl silicate (TEOS), and 3 g of ethyl acetate.

[0062] In step 2, 1 g of Fe-MSN, 0.2 mL of silane coupling agent aminopropyltriethoxysilane (APTES), 0.5 g of GA, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl), 0.12 g of N,N-dicyclohexylcarbodiimide (DCC), 0.3 g of N-hydroxysuccinimide (NHS), and 100 mL of 1 mg / mL Fe-MSN-NH2 ethanol dispersion were prepared.

[0063] In step 3, DOX 10 mg, Fe-MSN-GA 30 mg.

[0064] Example 4

[0065] Particle size and drug loading rate of DOX@Fe-MSN-GA

[0066] The specific test method of the particle size test is as follows: Dynamic light scattering (DLS) is used to measure the particle size distribution of the sample. In the experiment, the particle size distribution of DOX@Fe-MSN-GA aqueous dispersion (1 mg / mL) was measured using a Malvern Zetasizer Nano-ZS 90 particle size analyzer, and the test temperature was 25°C. The results are shown in Table 1.

[0067] The specific test methods for drug loading and encapsulation efficiency are as follows:

[0068] (1) Preparation of standard solution: Accurately weigh the DOX standard, dissolve it in DMSO, and prepare a standard solution of 5-100 μg / ml;

[0069] (2) The standard solution and DOX@Fe-MSN-GA aqueous dispersion (1 mg / mL) were detected by UV-Vis method, and linear regression was performed with the absorbance of DOX in the standard solution as the ordinate and the concentration (μg / mL) as the abscissa to obtain the linear regression equation. The absorbance value ratio of the sample solution to be tested was substituted into the linear regression equation to calculate the DOX content of DOX@Fe-MSN-GA, thereby calculating the content of the drug in the carrier.

[0070] The drug loading and encapsulation efficiency were calculated according to the following formula. The test was performed three times in parallel and the average values ​​were taken. The drug loading and encapsulation efficiency results are shown in Table 1 below.

[0071]

[0072]

[0073] Table 1 Test results of various embodiments

[0074] project Average particle size PDI Encapsulation efficiency Drug loading rate Example 1 237.84±10.14nm 0.233 24.46±1.68％ 19.65±0.54％ Example 2 273.95±16.56nm 0.545 25.6±2.67％ 20.42±2.14％ Example 3 217.28±13.61nm 0.228 89.26±1.06％ 22.93±1.03％

[0075] Table 1 shows that the particle size of DOX@Fe-MSN-GA prepared in each example is between 200 and 300 nm, but the PDI of Example 2 is relatively large and the dispersibility is slightly poor. The drug loading rate of each example is similar, but the encapsulation rate of Example 3 is the best.

[0076] Example 5

[0077] Effect of DOX@Fe-MSN-GA on the activity of hepatocellular carcinoma cells (HepG2)

[0078] Using DOX@Fe-MSN-GA prepared in Example 3 as a sample, HepG2 cells were cultured at 5×10 5The cells were inoculated in a 24-well plate at a density of 100 μL per well and cultured in a 37°C, 5% CO2 incubator for 12 h. 200 μL of DOX@Fe-MSN-GA was added to co-incubate with HepG2 cells for 24 h, washed twice with PBS to remove free drugs, 1 mL of 10% CCK-8 solution was added to each well, incubated in a 37°C, 5% CO2 incubator for a period of time, and the OD value was measured at 450 nm using a multifunctional microplate reader.

[0079] DOX@Fe-MSN-GA was dissolved in sterile PBS, and then the DOX@Fe-MSN-GA solution was diluted with complete culture medium to a final DOX concentration of 5, 10, 20, and 50 μg / mL. HepG2 cells were cultured at 8×10 cells per well. 3 The cells were inoculated into a 96-well plate at a density of 100 μL. After culturing in the incubator for 24 hours, the original culture medium was discarded, and 100 μL of the prepared DOX@Fe-MSN-GA solution was added to each well. Six parallel wells were set for each concentration, and a blank control group was set up. After continuing to culture in the incubator for 24 hours, 1 mL of 10% CCK-8 solution was added to each well, and the cells were incubated in a 37°C, 5% CO2 incubator for a period of time. The OD value was measured at 450 nm using a multifunctional microplate reader.

[0080]

[0081] OD0: OD value of blank group; OD1: OD value of experimental group; OD2: OD value of control group

[0082] like Figure 2 As shown in the figure, the higher the DOX concentration, the lower the survival rate of HepG2 cells. Compared with the DOX group, the cytotoxicity of DOX@Fe-MSN-GA was significantly enhanced, which indicates that DOX@Fe-MSN-GA triggers the Fenton reaction (Fe+H2O2→Fe+OH+·OH) in the tumor microenvironment. The strongly oxidizing ·OH induces tumor cell apoptosis by damaging DNA and inactivating proteins, and synergizes with the chemotherapeutic effect of DOX to exert in vitro antitumor activity.

[0083] Example 6

[0084] Effects of DOX@Fe-MSN-GA on reactive oxygen species in hepatoma cells (HepG2)

[0085] Using DOX@Fe-MSN-GA prepared in Example 3 as a sample, HepG2 cells were cultured at 4×10 5The cells were inoculated at a density of 100 cells / well in a 6-well plate and cultured under appropriate conditions (37°C, 5% CO2) for 24 hours. The original culture medium was removed, and 1 mL of culture medium containing DOX@Fe-MSN-GA was added, and the culture medium was incubated for 2 hours, 4 hours, and 8 hours respectively. The culture medium was discarded, and the original culture medium was replaced with fresh DCFH-DA (10 μM) culture medium for 30 minutes. The cells were then digested and collected, and the fluorescence intensity of ROS was measured by flow cytometry. The greater the fluorescence intensity, the higher the content of ROS.

[0086] The results show that the fluorescence intensity of intracellular ROS increases with time. In addition, as the concentration of DOX@Fe-MSN-GA increases, the content of intracellular ROS also increases, indicating that DOX@Fe-MSN-GA can affect the content of intracellular ROS. This is because as the cell's intake of DOX@Fe-MSN-GA increases, iron accumulation increases, causing the Fenton reaction to become stronger and the production of ROS also increases.

[0087] Example 7

[0088] Distribution statistics of DOX@Fe-MSN-GA in organs of nude mice with hepatocellular carcinoma

[0089] The DOX@Fe-MSN-GA prepared in Example 3 was used as the sample.

[0090] 4-week-old BALB / c nude mice were selected, carefully raised and observed for one week. After the nude mice were stable, HepG2 cell suspension was inoculated with 1×10 7 The cells were inoculated at a density of 100 μg / cm2 on the right posterior side of the nude mouse back until the tumor diameter was about 100 mm. 3 The administration started at 1:00 p.m. Cy5@Fe-MSN-GA (Cy5 replaced DOX) was intravenously injected into tumor-bearing nude mice. After 3h, 6h, 12h, and 24h of administration, the nude mice were anesthetized and the distribution of nanoparticles in the nude mice was detected by the small animal in vivo imaging system.

[0091] Figure 4 The fluorescence intensity statistics of the heart, liver, spleen, lung, kidney and tumor sites in nude mice show that the relative fluorescence intensity of the tumor site is significantly higher than that of other sites, indicating that the nanoparticles of the present invention have obvious targeting to the tumor site.

[0092] Example 8

[0093] Antitumor effect of DOX@Fe-MSN-GA in nude mouse model of liver cancer

[0094] The DOX@Fe-MSN-GA prepared in Example 3 was used as the sample.

[0095] The nude mice with tumor volumes that met the requirements were randomly divided into 3 groups, 5 mice in each group, namely PBS group, DOX group, and DOX@Fe-MSN-GA group. 100 μL of different drugs (PBS, DOX (10 mg / kg), DOX@Fe-MSN-GA (DOX10 mg / kg)) were injected into the tail vein, and then 100 μL of the corresponding drugs were injected into the tail vein again on the 4th, 7th, 10th, and 13th days. The health status of the nude mice was observed daily, and the length and diameter of the nude mouse tumor were measured every other day to monitor the tumor volume (tumor volume = length × width 2 ×0.5), and the 28-day survival rate of the experimental nude mice was calculated.

[0096] result( Figure 5 and Figure 6 ) showed that compared with the PBS group, the DOX group and the DOX@Fe-MSN-GA group showed anti-tumor effects. Compared with the DOX group, the DOX@Fe-MSN-GA group had a stronger inhibitory effect on tumor growth, which was due to the targeting of the carrier and the triggering of the Fenton reaction to produce more ROS to kill tumor cells. In addition, DOX@Fe-MSN-GA can significantly improve the survival rate of tumor-bearing nude mice.

[0097] In summary, DOX@Fe-MSN-GA is a targeted nanoparticle for the synergistic treatment of liver cancer with chemotherapy and chemodynamic therapy.

[0098] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions provided according to the embodiments of the present invention, rather than to limit them. Although the present invention has been described in detail with reference to the aforementioned embodiments, it can be understood by those of ordinary skill in the art that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principles and spirit provided according to the embodiments of the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles provided according to the embodiments of the present invention should be included in the protection scope provided according to the embodiments of the present invention.

Claims

1. A method for preparing targeted nanoparticles, characterized in that: Includes steps: Step 1: Preparation of Fe-MSN nanocarriers; Dissolve hexadecyltrimethylammonium bromide, ferrous sulfate heptahydrate, and ferric chloride hexahydrate in pure water, then add ammonia water, stir and react, then add tetraethyl silicate and ethyl acetate, stir and react, centrifuge and wash the reaction turbid liquid after the reaction, then dry it, and finally calcine at high temperature to remove the template hexadecyltrimethylammonium bromide, so as to obtain Fe-MSN; Step 2: The surface of Fe-MSN is modified with glycyrrhetinic acid ligand to obtain Fe-MSN-glycyrrhetinic acid; Step 2.1: reacting the Fe-MSN prepared in step 1 with a silane coupling agent, aminopropyltriethoxysilane, to obtain Fe-MSN-NH2; Step 2.2: In an anhydrous ethanol medium, glycyrrhetinic acid, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N,N-dicyclohexylcarbodiimide are stirred for reaction, and then N-hydroxysuccinimide is added and stirred for reaction to prepare a glycyrrhetinic acid mixed solution; Step 2.3: Add the Fe-MSN-NH2 dispersion dropwise to the glycyrrhetinic acid mixture obtained in step 2.2, and stir to react to obtain Fe-MSN-glycyrrhetinic acid; Step 3: Fe-MSN-glycyrrhetinic acid is loaded with the anticancer drug DOX to obtain DOX@Fe-MSN-glycyrrhetinic acid; DOX was dissolved in dimethyl sulfoxide, and the DMSO solution of DOX was dropped into the Fe-MSN-glycyrrhetinic acid aqueous solution while ultrasonically stirring. After the addition was completed, the free DOX was removed to obtain DOX@Fe-MSN-glycyrrhetinic acid.

2. The preparation method according to claim 1, characterized in that: In the step 1, the mass ratio of the raw materials is: hexadecyltrimethylammonium bromide: ferrous sulfate heptahydrate: ferric chloride hexahydrate: pure water: ammonia water: tetraethyl silicate: ethyl acetate = 0.1~10: 0.1~1: 0.2~2: 10~100: 0.3~3: 0.05~0.5: 0.5~5, the reaction temperature is 40°C, after adding ammonia water, stirring at 300rpm for 1~3 hours, then adding tetraethyl silicate and ethyl acetate, first stirring vigorously at 500rpm for 0.5~2 minutes, and then stirring at 150rpm for 4~8 hours.

3. The preparation method according to claim 1, characterized in that: In the step 1, the reaction turbid liquid is first centrifuged and washed with pure water until the pH value is 7, and then centrifuged and washed several times with anhydrous ethanol.

4. The preparation method according to claim 1, characterized in that: In the step 1, the drying temperature is 60-80°C, and the temperature for high-temperature calcination to remove the template is 500-600°C.

5. The preparation method according to claim 1, characterized in that: In the step 2.1, the reaction medium is anhydrous ethanol, the usage ratio of each raw material is: Fe-MSN: silane coupling agent = 0.1g~1g: 0.1mL~10mL, the reaction temperature is 80~90°C, and the reaction time is 4~8h.

6. The preparation method according to claim 1, characterized in that: In the step 2.2, the mass ratio of the raw materials is: glycyrrhetinic acid: 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride: N,N-dicyclohexylcarbodiimide: N-hydroxysuccinimide = 1-30: 1.2-12: 1.2-12: 3-30, Glycyrrhetinic acid, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N,N-dicyclohexylcarbodiimide were stirred and reacted for 0.5-2 h, and then N-hydroxysuccinimide was added and reacted for 3-5 h.

7. The preparation method according to claim 1, characterized in that: In the step 2.3, in the reaction solution, the mass ratio of glycyrrhetinic acid to Fe-MSN-NH2 is 1-30:0.1-10, and the reaction time is 3-5h.

8. The preparation method according to claim 1, characterized in that: In the step 3, the mass ratio of DOX to Fe-MSN-glycyrrhetinic acid is 0.1-10:0.1-10.

9. The targeted nanoparticles prepared according to the preparation method according to any one of claims 1 to 8.

10. Use of the nanoparticles according to claim 9 in preparing drugs for treating liver cancer.

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