A composite nanoparticle for treating osteosarcoma and its preparation method

Abstract

The present invention discloses a composite nanoparticle for treating osteosarcoma and a preparation method thereof. The composite nanoparticle uses porous silica nanoparticles or copper-containing porous silica nanoparticles as a carrier to encapsulate ilisimol. The preparation method of the composite nanoparticle for treating osteosarcoma comprises the following steps: uniformly mixing the porous silica nanoparticles or copper-containing porous silica nanoparticles with a dimethyl sulfoxide solution of ilisimol, stirring in the dark at room temperature for 23 to 25 hours, centrifuging for 14 to 16 minutes, collecting the precipitate, and freeze-drying for 23 to 26 hours to obtain the composite nanoparticle for treating osteosarcoma. The composite nanoparticle provided by the present invention is obtained by encapsulating ilisimol using porous silica nanoparticles or copper-containing porous silica nanoparticles as a carrier. The resulting composite nanoparticle induces copper cell death in osteosarcoma, can significantly inhibit osteosarcoma cell growth, achieve osteosarcoma treatment, and prolong patient survival, providing a new direction for the treatment of osteosarcoma.

Description

Technical Field

[0001] The present invention belongs to the technical field of nanomedicines, and in particular relates to a composite nanomedicine for treating osteosarcoma and a preparation method thereof. Background Art

[0002] Osteosarcoma (OS) is an aggressive tumor caused by mesenchymal cells. It mainly occurs in adolescents. Clinical manifestations include limb pain, lumps, lameness, etc. Some patients also have fever, anemia and weight loss. It causes serious harm to the health of adolescents and has a high disability and mortality rate.

[0003] Currently, the main treatments for osteosarcoma include surgery and combined chemotherapy. However, osteosarcoma has a high propensity to metastasize, and the survival rate for patients with metastatic or recurrent osteosarcoma has remained virtually unchanged over the past 30 years, with an overall five-year survival rate of approximately 20%. Furthermore, with the increasing prevalence of chemotherapy resistance, osteosarcoma patients still face a high risk of mortality. Therefore, the development and introduction of novel treatment concepts and strategies are crucial to improving the prognosis of osteosarcoma patients. Summary of the Invention

[0004] The present invention provides a composite nanoparticle for treating osteosarcoma, which is obtained by encapsulating ilisimol with porous silica nanoparticles or copper-containing porous silica nanoparticles as carriers. The obtained composite nanoparticle induces copper death in osteosarcoma, can significantly inhibit the growth of osteosarcoma cells, achieve osteosarcoma treatment, and prolong patient survival rate, providing a new direction for the treatment of osteosarcoma. In addition, the present invention also provides a preparation method of the composite nanoparticle for treating osteosarcoma.

[0005] The first aspect of the present invention provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses porous silica nanoparticles or copper-containing porous silica nanoparticles as carriers to encapsulate ilisimol.

[0006] In one embodiment of the present invention, the porous silica nanoparticles are prepared by the following steps:

[0007] S11, adding hexadecyltrimethylammonium chloride, triethanolamine, and deionized water to a reaction vessel in sequence, and stirring at a temperature of 93 to 97° C. for 1 to 1.5 hours to obtain a mixed solution A;

[0008] S12, adding tetraethyl silicate dropwise to the mixed solution A, stirring, centrifuging, collecting the precipitate, washing, drying, and calcining to obtain the porous silica nanoparticles.

[0009] In one embodiment of the present invention, during the preparation of the porous silica nanoparticles, the mass ratio of hexadecyltrimethylammonium chloride, triethanolamine, deionized water, and tetraethyl silicate is (1.8-2.2): (0.02-0.08): (18-22): (1.308-1.495);

[0010] In one embodiment of the present invention, in step S12, tetraethyl silicate is added dropwise to the mixed solution A, stirred for 1 to 2 hours, centrifuged at 12000-13000g for 10 to 15 minutes, the precipitate is collected, washed alternately with water and anhydrous ethanol 3 to 5 times, and dried in a freeze dryer for 10 to 12 hours. The powder obtained after drying is calcined at 580 to 620°C for 5 to 7 hours to obtain the porous silica nanoparticles.

[0011] In one embodiment of the present invention, in step S12, the powder obtained after drying is placed in a muffle furnace, the heating rate is set to 1°C / min, and it is calcined at 580-620°C for 5-7 hours to remove the surfactant hexadecyltrimethylammonium chloride remaining in the preparation process, thereby obtaining the porous silica nanoparticles.

[0012] In one embodiment of the present invention, the copper-containing porous silica nanoparticles are prepared by the following steps:

[0013] S21, adding hexadecyltrimethylammonium chloride, triethanolamine, and deionized water to a reaction vessel in sequence, and stirring at a temperature of 93 to 97° C. for 1 to 1.5 hours to obtain a mixed solution B;

[0014] S22, copper nitrate trihydrate is dissolved in anhydrous ethanol to obtain a dark blue clear mixed solution C;

[0015] S23, uniformly mixing the mixed solution C and tetraethyl silicate to obtain a mixed solution D;

[0016] S24, adding the mixed solution D dropwise into the mixed solution B, stirring, centrifuging, collecting the precipitate, washing, drying, and calcining to obtain the copper-containing porous silica nanoparticles.

[0017] In one embodiment of the present invention, in the step of preparing copper-containing porous silica nanoparticles, the mass ratio of hexadecyltrimethylammonium chloride, triethanolamine, deionized water, copper nitrate trihydrate, and tetraethyl silicate is (1.8-2.2): (0.02-0.08): (18-22): (0.09-1.10): (1.308-1.495).

[0018] In one embodiment of the present invention, in step S22, the solvent used is anhydrous ethanol;

[0019] In step S24, the mixed solution D is added dropwise to the mixed solution B at 80°C, stirred for 4 to 6 hours, centrifuged at 12000-13000g for 10 to 15 minutes, the precipitate is collected, washed alternately with water and anhydrous ethanol for 3 to 5 times, and placed in a freeze dryer for 10 to 12 hours. The powder obtained after drying is placed in a muffle furnace, the heating rate is set to 1°C / min, and calcined at 580 to 620°C for 5 to 7 hours to remove the surfactant hexadecyltrimethylammonium chloride remaining in the preparation process, thereby obtaining the copper-containing porous silica nanoparticles.

[0020] A second aspect of the present invention provides a method for preparing a composite nanoparticle for treating osteosarcoma, comprising the following steps:

[0021] The porous silica nanoparticles or copper-containing porous silica nanoparticles are evenly mixed with a dimethyl sulfoxide solution of ilisimol, stirred in the dark at room temperature for 23 to 25 hours, centrifuged at 12000-13000g for 14 to 16 minutes, the precipitate is collected, and freeze-dried for 23 to 26 hours to obtain a composite nanoparticle for treating osteosarcoma; the obtained composite nanoparticle is porous silica nanoparticles or copper-containing porous silica nanoparticles loaded with ilisimol.

[0022] In one embodiment of the present invention, the concentration of ilisimol in dimethyl sulfoxide is 2 mM;

[0023] Calculated in mg / mL, the mass-to-volume ratio of the porous silica nanoparticles or copper-containing porous silica nanoparticles to the dimethyl sulfoxide solution of ilisimol is (4-6):(10-12).

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] 1. The composite nanoparticles provided in the embodiments of the present invention are obtained by encapsulating ilisimol with porous silica nanoparticles or copper-containing porous silica nanoparticles as carriers. The resulting composite nanoparticles induce copper cell death in osteosarcoma, significantly inhibit the growth of osteosarcoma cells, achieve osteosarcoma treatment, and prolong patient survival, providing a new direction for the treatment of osteosarcoma.

[0026] 2. The method for preparing the composite nanopharmaceutical provided in the embodiment of the present invention has a wide range of raw material sources, a simple and safe process, and is easy to implement industrial production.

[0027] Figures in the specification

[0028] Figure 1 Schematic diagram of the synthesis of the composite nanoparticles of the present invention;

[0029] Figure 2 The clearance effect of the composite nanoparticles on osteosarcoma cells 143B and HOS in Example 9;

[0030] Figure 3 The clearance effect of the composite nanoparticles on osteosarcoma cells 143B and HOS in Example 6;

[0031] Figure 4 This is the clearance effect of the composite nanoparticle agent in Example 12 on osteosarcoma cells 143B and HOS. DETAILED DESCRIPTION

[0032] In this document, the term "from a value to another value" is used as a summary to avoid listing all values ​​within the range. Therefore, a description of a specific numerical range encompasses any value within that range and any smaller numerical ranges defined by any value within that range, just as if the values ​​and smaller numerical ranges were explicitly stated in the specification.

[0033] In this application, unless otherwise stated, the use of “or” means “and / or.” In the case of multiple dependent claims, use of “or” is used only in the alternative to refer to more than one of the preceding independent or dependent claims.

[0034] Unless otherwise indicated, as used in accordance with this disclosure, the following terms shall be understood to have the following meanings:

[0035] It should be noted that the scientific and technical terms and abbreviations used in the present invention have the meanings commonly understood by those skilled in the art. The following lists some of the terms and abbreviations used in the present invention:

[0036] The term "treat," when used in reference to treating, for example, a condition or disease, refers to alleviating and / or eliminating one or more symptoms of the condition or disease, and / or delaying the progression of one or more symptoms of the condition or disease, and / or reducing the incidence or severity of one or more symptoms of the condition or disease, and / or preventing the condition or disease. The term treat may refer to prophylactic treatment, which includes delaying the onset of the condition or disease or preventing the onset of the condition or disease.

[0037] The terms "porous silica nanoparticles" and "porous silica nanoparticles" are used interchangeably and refer to nanostructures having a porous particle core.

[0038] The first aspect of the present invention provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses porous silica nanoparticles or copper-containing porous silica nanoparticles as carriers to encapsulate ilisimol.

[0039] The surface of the porous silica nanoparticles or copper-containing porous silica nanoparticles has multiple pores, which are used to encapsulate ilisimol; ilisimol is transported into tumor cells through the silica nanoparticles or copper-containing porous silica nanoparticles to achieve the treatment of osteosarcoma, and copper death-inducing therapy is introduced into the treatment of osteosarcoma.

[0040] The composite nanoparticles for treating osteosarcoma provided by the present invention have excellent osteosarcoma therapeutic effects.

[0041] The porous silica nanoparticles are prepared by the following steps:

[0042] S11, adding hexadecyltrimethylammonium chloride, triethanolamine, and deionized water to a reaction vessel in sequence, and stirring at a temperature of 93 to 97° C. for 1 to 1.5 hours to obtain a mixed solution A;

[0043] S12, adding tetraethyl silicate dropwise to the mixed solution A, stirring, centrifuging, collecting the precipitate, washing, drying, and calcining to obtain the porous silica nanoparticles.

[0044] Wherein, in the preparation process of the porous silica nanoparticles, the mass ratio of hexadecyltrimethylammonium chloride, triethanolamine, deionized water, and tetraethyl silicate is (1.8-2.2): (0.02-0.08): (18-22): (1.308-1.495);

[0045] In step S12, tetraethyl silicate is added dropwise to the mixed solution A, stirred for 1 to 2 hours, centrifuged at 12000-13000g for 10 to 15 minutes, the precipitate is collected, washed alternately with water and anhydrous ethanol 3 to 5 times, and placed in a freeze dryer for 10 to 12 hours. After drying, the resulting powder is calcined at 580 to 620°C for 5 to 7 hours to obtain the porous silica nanoparticles.

[0046] In step S12, the powder obtained after drying is placed in a muffle furnace, the heating rate is set to 1°C / min, and it is calcined at 580-620°C for 5-7 hours to remove the surfactant hexadecyltrimethylammonium chloride remaining in the preparation process, thereby obtaining the porous silica nanoparticles.

[0047] The copper-containing porous silica nanoparticles are prepared by the following steps:

[0048] S21, adding hexadecyltrimethylammonium chloride, triethanolamine, and deionized water to a reaction vessel in sequence, and stirring at a temperature of 93 to 97° C. for 1 to 1.5 hours to obtain a mixed solution B;

[0049] S22, copper nitrate trihydrate is dissolved in anhydrous ethanol to obtain a dark blue clear mixed solution C;

[0050] S23, uniformly mixing the mixed solution C and tetraethyl silicate to obtain a mixed solution D;

[0051] S24, adding the mixed solution D dropwise into the mixed solution B, stirring, centrifuging, collecting the precipitate, washing, drying, and calcining to obtain the copper-containing porous silica nanoparticles.

[0052] Among them, in the preparation step of the copper-containing porous silica nanoparticles, the mass ratio of hexadecyltrimethylammonium chloride, triethanolamine, deionized water, copper nitrate trihydrate, and tetraethyl silicate is (1.8-2.2): (0.02-0.08): (18-22): (0.09-1.10): (1.308-1.495).

[0053] Wherein, in step S22, the solvent used is anhydrous ethanol;

[0054] In step S24, the mixed solution D is added dropwise to the mixed solution B at 80°C, stirred for 4 to 6 hours, centrifuged at 12000-13000g for 10 to 15 minutes, the precipitate is collected, washed alternately with water and anhydrous ethanol for 3 to 5 times, and placed in a freeze dryer for 10 to 12 hours. The powder obtained after drying is placed in a muffle furnace, the heating rate is set to 1°C / min, and calcined at 580 to 620°C for 5 to 7 hours to remove the surfactant hexadecyltrimethylammonium chloride remaining in the preparation process, thereby obtaining the copper-containing porous silica nanoparticles.

[0055] A second aspect of the present invention provides a method for preparing a composite nanoparticle for treating osteosarcoma, comprising the following steps:

[0056] The porous silica nanoparticles or copper-containing porous silica nanoparticles are evenly mixed with a dimethyl sulfoxide solution of ilisimol, stirred in the dark at room temperature for 23 to 25 hours, centrifuged at 12000-13000g for 14 to 16 minutes, the precipitate is collected, and freeze-dried for 23 to 26 hours to obtain a composite nanoparticle for treating osteosarcoma; the obtained composite nanoparticle is porous silica nanoparticles or copper-containing porous silica nanoparticles loaded with ilisimol.

[0057] The concentration of ilisimol in dimethyl sulfoxide is 2 mM; in terms of mg / mL, the mass volume ratio of the porous silica nanoparticles or copper-containing porous silica nanoparticles to the dimethyl sulfoxide solution of ilisimol is (4-6): (10-12).

[0058] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only intended to illustrate the present invention and are not intended to limit the scope of protection of the present invention. Improvements and adjustments made by those skilled in the art in accordance with the present invention in actual applications still fall within the scope of protection of the present invention.

[0059] Example 1

[0060] This embodiment provides a method for preparing porous silica nanoparticles, comprising the following steps:

[0061] Dissolve 1.8g of hexadecyltrimethylammonium chloride and 0.02g of triethanolamine in 18mL of deionized water and stir at 93-97°C for 1-1.5 hours. Add 1.4mL of tetraethyl silicate dropwise to the mixed solution of hexadecyltrimethylammonium chloride and triethanolamine, stir for 1-2 hours, and centrifuge (13000g for 12 minutes) to collect the product. Rinse with water and ethanol 3-5 times. After the final centrifugation, dry the solid in a freeze dryer for 12 hours. The dried powder is placed in a muffle furnace, heated at a rate of 1°C / min, and calcined at 600°C for 5 hours to remove any residual surfactant from the preparation process.

[0062] Example 2

[0063] This embodiment provides a method for preparing porous silica nanoparticles, comprising the following steps:

[0064] Dissolve 2.2g of hexadecyltrimethylammonium chloride and 0.08g of triethanolamine in 22mL of deionized water and stir at 93-97°C for 1-1.5 hours. Add 1.6mL of tetraethyl silicate dropwise to the mixture of hexadecyltrimethylammonium chloride and triethanolamine, stir for 1-2 hours, and centrifuge (13000g for 12 minutes) to collect the product. Rinse with water and ethanol 3-5 times. After the final centrifugation, dry the solid in a freeze dryer for 11 hours. The dried powder is placed in a muffle furnace, heated at a rate of 1°C / min, and calcined at 620°C for 7 hours to remove any residual surfactant from the preparation process.

[0065] Example 3

[0066] This embodiment provides a method for preparing porous silica nanoparticles, comprising the following steps:

[0067] Dissolve 2g of hexadecyltrimethylammonium chloride and 0.06g of triethanolamine in 20mL of deionized water and stir at 93-97°C for 1-1.5 hours. Add 1.5mL of tetraethyl silicate dropwise to the mixture of hexadecyltrimethylammonium chloride and triethanolamine, stir for 1-2 hours, and centrifuge (13,000g for 12 minutes) to collect the product. Rinse with water and ethanol 3-5 times. After the final centrifugation, dry the solid in a freeze dryer for 10 hours. The dried powder is placed in a muffle furnace, heated at a rate of 1°C / min, and calcined at 600°C for 6 hours to remove any residual surfactant from the preparation process.

[0068] Example 4

[0069] This embodiment provides a method for preparing copper-containing porous silica nanoparticles, comprising the following steps:

[0070] Dissolve 1.8g of hexadecyltrimethylammonium chloride and 0.02g of triethanolamine in 18mL of deionized water and stir at 93-97°C for 1.5 hours. Weigh 0.09g of copper nitrate trihydrate and dissolve thoroughly in anhydrous ethanol with stirring to obtain a dark blue clear solution. Mix the dark blue clear solution with 1.4mL of tetraethyl silicate and add it dropwise to the mixed solution of hexadecyltrimethylammonium chloride and triethanolamine at 80°C. Stir for 4-6 hours. Collect the product by centrifugation (12000-14000g, 10-15 minutes), rinse with water and ethanol 3-5 times, and dry the resulting solid in a freeze dryer for 11 hours. The resulting powder is placed in a muffle furnace, heated at a rate of 1°C / min, and calcined at 620°C for 5-7 hours to remove any residual surfactant from the preparation process.

[0071] Example 5

[0072] This embodiment provides a method for preparing copper-containing porous silica nanoparticles, comprising the following steps:

[0073] Dissolve 2.2g of hexadecyltrimethylammonium chloride and 0.08g of triethanolamine in 22mL of deionized water and stir at 93-97°C for 1.2 hours. Weigh 1.10g of copper nitrate trihydrate and dissolve thoroughly in anhydrous ethanol, yielding a dark blue, clear solution. Mix the dark blue, clear solution with 1.6mL of tetraethyl silicate and add it dropwise to the hexadecyltrimethylammonium chloride and triethanolamine solution at 80°C. Stir for 4-6 hours. Collect the product by centrifugation (12,000-14,000g, 10-15 minutes), rinse with water and ethanol 3-5 times, and dry the resulting solid in a freeze dryer for 10-12 hours. The resulting powder is placed in a muffle furnace, heated at a rate of 1°C / min, and calcined at 600°C for 5-7 hours to remove any residual surfactant from the preparation process.

[0074] Example 6

[0075] This embodiment provides a method for preparing copper-containing porous silica nanoparticles, comprising the following steps:

[0076] Dissolve 2g of hexadecyltrimethylammonium chloride and 0.06g of triethanolamine in 20mL of deionized water and stir at 93-97°C for 1.4 hours. Weigh 1.00g of copper nitrate trihydrate and dissolve thoroughly in anhydrous ethanol to obtain a dark blue clear solution. Mix the dark blue clear solution with 1.5mL of tetraethyl silicate and add it dropwise to the mixed solution of hexadecyltrimethylammonium chloride and triethanolamine at 80°C. Stir for 4-6 hours. Collect the product by centrifugation (12000-14000g, 10-15 minutes), rinse with water and ethanol 3-5 times, and dry the resulting solid in a freeze dryer for 10-12 hours. The resulting powder is placed in a muffle furnace, heated at a rate of 1°C / min, and calcined at 610°C for 5-7 hours to remove any residual surfactant from the preparation process.

[0077] Example 7

[0078] This embodiment provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses porous silica nanoparticles as a carrier to encapsulate ilisimol.

[0079] The composite nanoparticles for treating osteosarcoma in this embodiment are prepared by the following method:

[0080] 4 mg of the porous silica prepared in Example 1 was evenly mixed with 10 mL of a dimethyl sulfoxide solution (2 mM) of ilisimol, stirred in the dark for 23-25 ​​h at room temperature, and the porous silica loaded with ilisimol and the copper-containing porous silica were separated from the solution by high-speed centrifugation (centrifugal speed of 13000 g, centrifugation time of 11-16 min), and the precipitate was collected, freeze-dried for 23-26 h, and stored in a refrigerator at 3-5 ° C for use. At this point, the porous silica loaded with ilisimol was prepared.

[0081] Example 8

[0082] This embodiment provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses porous silica nanoparticles as a carrier to encapsulate ilisimol.

[0083] The composite nanoparticles for treating osteosarcoma in this embodiment are prepared by the following method:

[0084] 5 mg of the porous silica prepared in Example 1 was evenly mixed with 11 mL of a dimethyl sulfoxide solution (2 mM) of ilisimol, stirred in the dark at room temperature for 23-25 ​​h, and the porous silica loaded with ilisimol and the copper-containing porous silica were separated from the solution by high-speed centrifugation (centrifugal speed of 13000 g, centrifugation time of 11-16 min), and the precipitate was collected, freeze-dried for 23-26 h, and stored in a refrigerator at 3-5 ° C for use. At this point, the porous silica loaded with ilisimol was prepared.

[0085] Example 9

[0086] This embodiment provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses porous silica nanoparticles as a carrier to encapsulate ilisimol.

[0087] The composite nanoparticles for treating osteosarcoma in this embodiment are prepared by the following method:

[0088] 6 mg of the porous silica prepared in Example 1 was evenly mixed with 12 mL of a dimethyl sulfoxide solution (2 mM) of ilisimol, stirred in the dark at room temperature for 23-25 ​​h, and the porous silica loaded with ilisimol and the copper-containing porous silica were separated from the solution by high-speed centrifugation (centrifugal speed of 13000 g, centrifugation time of 11-16 min), and the precipitate was collected, freeze-dried for 23-26 h, and stored in a refrigerator at 3-5 ° C for use. At this point, the porous silica loaded with ilisimol was prepared.

[0089] Example 10

[0090] This embodiment provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses copper-containing porous silica nanoparticles as a carrier to encapsulate ilisimol.

[0091] The composite nanoparticles for treating osteosarcoma in this embodiment are prepared by the following method:

[0092] 4 mg of the copper-containing porous silica prepared in Example 4 was evenly mixed with 10 mL of a dimethyl sulfoxide solution (2 mM) of ilisimol, stirred in the dark for 23-25 ​​h at room temperature, and the porous silica loaded with ilisimol and the copper-containing porous silica were separated from the solution by high-speed centrifugation (centrifugal speed of 13000 g, centrifugation time of 11-16 min), and the precipitate was collected, freeze-dried for 23-26 h, and then stored in a refrigerator at 3-5 ° C for use. At this point, copper-containing porous silica loaded with ilisimol was prepared.

[0093] Example 11

[0094] This embodiment provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses copper-containing porous silica nanoparticles as a carrier to encapsulate ilisimol.

[0095] The composite nanoparticles for treating osteosarcoma in this embodiment are prepared by the following method:

[0096] 5 mg of the copper-containing porous silica prepared in Example 4 was evenly mixed with 11 mL of a dimethyl sulfoxide solution (2 mM) of ilisimol, stirred in the dark for 23-25 ​​h at room temperature, and the porous silica loaded with ilisimol and the copper-containing porous silica were separated from the solution by high-speed centrifugation (centrifugal speed of 13000 g, centrifugation time of 11-16 min), and the precipitate was collected, freeze-dried for 23-26 h, and stored in a refrigerator at 3-5 ° C for use. At this point, copper-containing porous silica loaded with ilisimol was prepared.

[0097] Example 12

[0098] This embodiment provides a composite nanoparticle for treating osteosarcoma. The composite nanoparticle uses copper-containing porous silica nanoparticles as a carrier to encapsulate ilisimol.

[0099] The composite nanoparticles for treating osteosarcoma in this embodiment are prepared by the following method:

[0100] 6 mg of the copper-containing porous silica prepared in Example 4 was evenly mixed with 12 mL of a dimethyl sulfoxide solution (2 mM) of ilisimol, stirred in the dark for 23-25 ​​h at room temperature, and the porous silica loaded with ilisimol and the copper-containing porous silica were separated from the solution by high-speed centrifugation (centrifugal speed of 13000 g, centrifugation time of 11-16 min), and the precipitate was collected, freeze-dried for 23-26 h, and stored in a refrigerator at 3-5 ° C for use. At this point, copper-containing porous silica loaded with ilisimol was prepared.

[0101] Test example

[0102] The clearance effect of the composite nanoparticle prepared in Example 12 on osteosarcoma cells 143B and HOS was verified.

[0103] The specific test verification steps are as follows:

[0104] 1. Add 4000-5000 143B or HOS cells per well in 100 μL of culture medium. Culture and stimulate with the appropriate material for 24 hours, as needed.

[0105] 2. Add 10 μl of CCK-8 solution to each well. Use wells containing the corresponding amount of cell culture medium and CCK-8 solution but no cells as blank controls.

[0106] 3. Continue incubating in the cell culture incubator for 2-3 hours.

[0107] 4. Measure the absorbance at 450 nm.

[0108] The results are as follows Figure 2-Figure 4 As shown, Figure 2 The clearance effect of the composite nanoparticles on osteosarcoma cells 143B and HOS in Example 9; Figure 3 The clearance effect of the composite nanoparticles on osteosarcoma cells 143B and HOS in Example 6; Figure 4 This is the clearance effect of the composite nanoparticle agent in Example 12 on osteosarcoma cells 143B and HOS.

[0109] Depend on Figure 2-Figure 4 It can be seen that the composite nanoparticle prepared in Example 12 has an excellent clearance effect on osteosarcoma cells 143B and HOS, and the drug obtained by copper-containing porous silica loaded with ilisimol has achieved a "1+1>2" therapeutic effect on osteosarcoma.

[0110] The above disclosure is merely a preferred embodiment of the present invention. The preferred embodiment does not exhaustively describe all details, nor does it limit the invention to the specific embodiments described. Obviously, many modifications and variations are possible based on the contents of this specification. These embodiments are selected and described in detail in this specification to better explain the principles and practical applications of the present invention, thereby enabling those skilled in the art to better utilize the present invention. The present invention is limited only by the claims and their full scope and equivalents.

[0111] Under the guidance of the present invention and the above-mentioned embodiments, it is easy for those skilled in the art to foresee that the raw materials or their equivalent substitutes, the processing methods or their equivalent substitutes listed or exemplified in the present invention can realize the present invention, and the upper and lower limit values ​​and interval values ​​of the parameters of the raw materials and processing methods can realize the present invention. The embodiments are not listed one by one here.

Claims

1. A composite nanoparticle for treating osteosarcoma, characterized in that: The composite nanoparticles use copper-containing porous silica nanoparticles as carriers to encapsulate ilisimol. The copper-containing porous silica nanoparticles are prepared by the following steps: S21, adding hexadecyltrimethylammonium chloride, triethanolamine, and deionized water to a reaction vessel in sequence, and stirring at a temperature of 93-97° C. for 1-1.5 h to obtain a mixed solution B; S22, copper nitrate trihydrate is dissolved in anhydrous ethanol to obtain a dark blue clear mixed solution C; S23, uniformly mixing the mixed solution C and tetraethyl silicate to obtain a mixed solution D; S24, adding the mixed solution D dropwise to the mixed solution B, stirring, centrifuging, collecting the precipitate, washing, drying, and calcining to obtain the copper-containing porous silica nanoparticles; In the preparation step of the copper-containing porous silica nanoparticles, the mass ratio of hexadecyltrimethylammonium chloride, triethanolamine, deionized water, copper nitrate trihydrate, and tetraethyl silicate is (1.8-2.2): (0.02-0.08): (18-22): (0.09-1.10): (1.308-1.495).

2. The composite nanoparticles for treating osteosarcoma according to claim 1, characterized in that: In step S22, the solvent used is anhydrous ethanol; In step S24, the mixed solution D is added dropwise to the mixed solution B at 80°C, stirred for 4-6 hours, centrifuged at 12000-13000g for 10-15 minutes, the precipitate is collected, washed alternately with water and anhydrous ethanol for 3-5 times, and dried in a freeze dryer for 10-12 hours. The powder obtained after drying is placed in a muffle furnace, the heating rate is set to 1°C / min, and the temperature is set to 580-620°C for calcination for 5-7 hours to remove the surfactant hexadecyltrimethylammonium chloride remaining in the preparation process, thereby obtaining the copper-containing porous silica nanoparticles.

3. A method for preparing the composite nanoparticle drug for treating osteosarcoma according to claim 1, characterized in that: The following steps are involved: The copper-containing porous silica nanoparticles and the dimethyl sulfoxide solution of ilisimol are mixed evenly, stirred in the dark at room temperature for 23 to 25 hours, centrifuged at 12000-13000g for 14 to 16 minutes, the precipitate is collected, and freeze-dried for 23 to 26 hours to obtain a composite nanoparticle for treating osteosarcoma; the obtained composite nanoparticle is copper-containing porous silica nanoparticles loaded with ilisimol.

4. The method for preparing the composite nanoparticle drug for treating osteosarcoma according to claim 3, characterized in that: The concentration of ilisimol in dimethyl sulfoxide was 2 mM; In terms of mg / mL, the mass-to-volume ratio of the copper-containing porous silica nanoparticles to the dimethyl sulfoxide solution of ilisimol is (4-6): (10-12).

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