A targeted nanotherapeutic agent, its preparation method and application

Abstract

This invention belongs to the field of pharmaceutical technology, specifically relating to a targeted nanotherapeutic agent, its preparation method, and its application. This invention provides a method for preparing a targeted nanotherapeutic agent, which involves preparing composite nanoparticles of water-soluble / water-insoluble drugs using supercritical carbon dioxide antisolvent technology. Then, layer-by-layer self-assembly technology is used to modify the surface of the composite nanoparticles with polycations and hyaluronic acid. This invention combines supercritical carbon dioxide antisolvent technology and layer-by-layer self-assembly technology to prepare targeted nanotherapeutic agents, achieving the co-delivery of multiple drugs (including water-soluble or water-insoluble drugs), while all drugs exhibit high encapsulation efficiency and also possess the function of targeting tumor sites.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a targeted nanotherapeutic agent, its preparation method, and its application. Background Technology

[0002] Malignant tumors seriously affect human health. Although researchers have developed chemical or natural drugs to treat tumors, low solubility and poor bioavailability remain serious problems affecting the efficacy of drug treatment for most drugs.

[0003] Nanoparticles possess a large specific surface area, exhibiting higher drug solubility and faster dissolution rates, effectively improving drug bioavailability. Furthermore, their nanoscale size provides potential for tumor targeting. Supercritical carbon dioxide antisolvent technology is a promising method for preparing micro / nanoparticles, combining the drug solubility of organic solvents with that of supercritical carbon dioxide. It has a certain research foundation in polymer-drug nanoparticle preparation and drug miniaturization. However, polymer-drug nanoparticles prepared using supercritical carbon dioxide antisolvent technology often exhibit low encapsulation efficiency, and the morphology and properties of the particles are limited by the polymer's own structure. Moreover, supercritical carbon dioxide antisolvent technology can only improve drug solubility and cannot adapt to the complex in vivo environment, severely limiting its subsequent applications. Currently, drugs with only a single therapeutic effect cannot meet the needs of tumor treatment, while nanocarriers that simultaneously load multiple drugs and ensure high encapsulation efficiency for all drugs are difficult to prepare. Summary of the Invention

[0004] In view of this, the purpose of this invention is to provide a targeted nanotherapeutic agent, its preparation method, and its application. This invention combines supercritical carbon dioxide antisolvent technology and layer-by-layer self-assembly technology to prepare a targeted nanotherapeutic agent that can simultaneously load multiple drugs, all of which exhibit high encapsulation efficiency.

[0005] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0006] This invention provides a method for preparing targeted nanotherapeutic agents, comprising the following steps:

[0007] Dissolve the water-soluble drug and the water-insoluble drug to obtain a mixed solution;

[0008] The mixed solution was prepared into composite nanoparticles using a supercritical carbon dioxide antisolvent method.

[0009] The composite nanoparticles are mixed with a solution containing polycations to obtain composite nanoparticles encapsulated by polycations.

[0010] The polycation-encapsulated composite nanoparticles are mixed with a solution containing hyaluronic acid to obtain the targeted nanotherapeutic agent.

[0011] Preferably, the mass ratio of the polycation-encapsulated composite nanoparticles to hyaluronic acid is 0.1–4:0.1–10.

[0012] Preferably, the temperature for mixing the polycation-encapsulated composite nanoparticles with the hyaluronic acid-containing solution is 2–30°C, and the time is 2–24 h.

[0013] Preferably, the polycation comprises one or more of polyornithine, polylysine, polyethyleneimine, polyarginine, and chitosan.

[0014] Preferably, the water-soluble drug includes a photosensitizer, and the photosensitizer includes indocyanine green.

[0015] Preferably, the water-insoluble drug comprises alkaloid compounds, including paclitaxel and / or camptothecin.

[0016] Preferably, the dissolved reagent includes an organic solvent.

[0017] Preferably, the organic solvent includes at least two of dichloromethane, acetone, chloroform, hexafluoroisopropanol, ethanol, methanol, dimethyl sulfoxide, and N,N-dimethylformamide.

[0018] The present invention also provides a targeted nanotherapeutic agent obtained by the preparation method described above, wherein the encapsulation rate of the drug in the targeted nanotherapeutic agent is 85-98%.

[0019] The present invention also provides the application of the targeted nanotherapeutic agents described above in the preparation of drugs for tumor diagnosis and / or treatment.

[0020] This invention provides a method for preparing targeted nanotherapeutic agents, comprising the following steps:

[0021] Dissolve the water-soluble drug and the water-insoluble drug to obtain a mixed solution;

[0022] The mixed solution was prepared into composite nanoparticles using a supercritical carbon dioxide antisolvent method.

[0023] The composite nanoparticles are mixed with a solution containing polycations to obtain composite nanoparticles encapsulated by polycations.

[0024] The polycation-encapsulated composite nanoparticles are mixed with a solution containing hyaluronic acid to obtain the targeted nanotherapeutic agent.

[0025] Compared with the prior art, the present invention has the following beneficial effects:

[0026] This invention prepares composite nanoparticles of water-soluble / water-insoluble drugs using supercritical carbon dioxide antisolvent technology, and then modifies the surface of the composite nanoparticles with polycations and hyaluronic acid using layer-by-layer self-assembly technology. This invention combines supercritical carbon dioxide antisolvent technology and layer-by-layer self-assembly technology to prepare targeted nanotherapeutic agents, achieving the co-delivery of multiple drugs (including water-soluble or water-insoluble drugs), while all drugs have high encapsulation efficiency and also have the function of targeting tumor sites.

[0027] Furthermore, the preparation method of the targeted nanotherapeutic agent of the present invention is simple to operate, has a short operation time, and has good prospects for industrial application.

[0028] The present invention also provides targeted nanotherapeutic agents prepared by the preparation method described above, which exhibit ultra-high encapsulation efficiency and drug loading for drugs of different properties (water-soluble and water-insoluble drugs), with an encapsulation efficiency of 85-98%, solving the problem of low encapsulation efficiency of traditional drug loading technology, and has great application prospects and important significance.

[0029] The present invention also provides the application of the targeted nanotherapeutic agent described in the above technical solution. The targeted nanotherapeutic agent of the present invention is loaded with a photosensitizer and has excellent fluorescence imaging performance, which can effectively target tumors. The data of the examples show that the targeted nanotherapeutic agent of the present invention can effectively target the CD44 receptor and is expected to achieve targeted combination therapy for tumors. Attached Figure Description

[0030] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0031] Figure 1 The infrared spectrum of the indocyanine green / paclitaxel composite nanoparticles in Example 1 is shown below.

[0032] Figure 2 This is a scanning electron microscope image of the targeted nanotherapeutic agent loaded with indocyanine green and paclitaxel in Example 1;

[0033] Figure 3 The image shows the encapsulation efficiency of the targeted nanotherapeutic agents indocyanine green and paclitaxel loaded with indocyanine green and paclitaxel in Example 1.

[0034] Figure 4 This is a graph showing the uptake results of tumor cells;

[0035] Figure 5Images of animals at different time periods;

[0036] Figure 6 Fluorescence images of major organs of the animal 24 hours after injection. Detailed Implementation

[0037] This invention provides a method for preparing targeted nanotherapeutic agents, comprising the following steps:

[0038] Dissolve the water-soluble drug and the water-insoluble drug to obtain a mixed solution;

[0039] The mixed solution was prepared into composite nanoparticles using a supercritical carbon dioxide antisolvent method.

[0040] The composite nanoparticles are mixed with a solution containing polycations to obtain composite nanoparticles encapsulated by polycations.

[0041] The polycation-encapsulated composite nanoparticles are mixed with a solution containing hyaluronic acid to obtain the targeted nanotherapeutic agent.

[0042] Unless otherwise specified, all materials and equipment used in this invention are commercially available products in the field.

[0043] This invention dissolves water-soluble drugs and water-insoluble drugs to obtain a mixed solution.

[0044] In this invention, the water-soluble drug preferably includes a photosensitizer, and the photosensitizer preferably includes indocyanine green.

[0045] In this invention, the water-insoluble drug preferably includes alkaloid compounds, and the alkaloid compounds preferably include paclitaxel and / or camptothecin.

[0046] In this invention, the dissolving reagent preferably includes an organic solvent, which preferably includes at least two of dichloromethane, acetone, chloroform, hexafluoroisopropanol, ethanol, methanol, dimethyl sulfoxide, and N,N-dimethylformamide, more preferably a mixture of ethanol and dichloromethane. The volume ratio of ethanol to dichloromethane in the mixture is preferably 1 to 2:1. These two organic solvents can simultaneously dissolve the water-soluble drug and the water-insoluble drug, and each drug has a lower solubility in one of the organic solvents. This ensures that the drug has a high supersaturation during mixing, rather than being diluted, which can effectively achieve subsequent nucleation.

[0047] In this invention, the concentration of the water-soluble drug in the mixed solution is preferably 0.1 to 1 mg / mL, more preferably 0.5 mg / mL.

[0048] In this invention, the concentration of the water-insoluble drug in the mixed solution is preferably 0.1 to 1 mg / mL, more preferably 0.25 mg / mL.

[0049] In this invention, the dissolution preferably includes method one or method two, and method one preferably includes the following steps:

[0050] A water-soluble drug is dissolved in a first organic solvent to obtain a first organic solution; the concentration of the water-soluble drug in the first organic solution is 0.1–10 mg / mL, preferably 1 mg / mL;

[0051] A water-insoluble drug is dissolved in a second organic solvent to obtain a second organic solution; the concentration of the water-insoluble drug in the second organic solution is 0.1–10 mg / mL, preferably 0.5 mg / mL;

[0052] The first organic solution and the second organic solution are mixed to obtain the mixed solution.

[0053] The present invention does not have any special requirements for the method by which the water-soluble drug is dissolved in the first organic solvent or the method by which the water-insoluble drug is dissolved in the second organic solvent; methods commonly used by those skilled in the art can be used.

[0054] In this invention, the volume ratio of the first organic solution to the second organic solution is preferably 2-8:2-8, and more preferably 1-2:1.

[0055] In this invention, the types of the first organic solvent and the second organic solvent are preferably the same as the types of the dissolved reagents described above, and will not be repeated here.

[0056] The preferred method two includes the following steps:

[0057] The water-soluble drug and the water-insoluble drug are dissolved in the first organic solvent and then mixed with the second organic solvent.

[0058] In this invention, the preferred ratio of the water-soluble drug, the water-insoluble drug, the first organic solvent, and the second organic solvent is 10 mg: 5 mg: 14 mL: 7 mL.

[0059] In this invention, the types of the first and second organic solvents are preferably the same as the types of reagents dissolved as described above, and will not be repeated here. In a specific embodiment of this invention, the first organic solvent is preferably ethanol, and the second organic solvent is preferably dichloromethane.

[0060] After obtaining the mixed solution, the present invention uses the supercritical carbon dioxide antisolvent method to prepare the mixed solution into composite nanoparticles.

[0061] In this invention, the parameters of the supercritical carbon dioxide antisolvent method include: the temperature is preferably 35-50℃; the pressure is preferably 8-15MPa, more preferably 9-11MPa; and the injection flow rate of the mixed solution is preferably 0.2-5mL / min, more preferably 0.75-1mL / min.

[0062] In this invention, after the injection of the mixed solution is completed, it is preferable to further perform rinsing and depressurization in sequence, and the rinsing time is preferably 2 hours.

[0063] In this invention, the particle size of the composite nanoparticles is preferably 10-200 nm.

[0064] After obtaining the composite nanoparticles, the present invention mixes the composite nanoparticles with a solution containing polycations to obtain composite nanoparticles encapsulated by polycations.

[0065] In this invention, the polycation in the polycation-containing solution preferably includes one or more of polyornithine, polylysine, polyethyleneimine, polyarginine, and chitosan. When the polycation is preferably chitosan, the polycation-containing solution preferably also includes acetic acid, and the volume concentration of acetic acid in the solution is preferably 2%.

[0066] In this invention, the concentration of the polycation in the polycation-containing solution is preferably 0.05 to 4 mg / mL, more preferably 0.1 to 1 mg / mL.

[0067] In this invention, the preferred ratio of the composite nanoparticles to the polycation-containing solution is 4-6 mg: 10-100 mL.

[0068] In this invention, the preferred mass ratio of the composite nanoparticles to the polycation is 4-6:1-100.

[0069] In this invention, the polycation-containing solution preferably also includes a sodium salt, which preferably includes one or more of sodium chloride, sodium sulfate and sodium phosphate, more preferably sodium chloride. The concentration of sodium chloride in the polycation-containing solution is preferably 0.5 to 1 mol / L. Indocyanine green in the water-soluble drug is a sodium salt, and the sodium ions in the sodium salt can reduce the dissolution of indocyanine green.

[0070] In this invention, the composite nanoparticles are preferably mixed with a solution containing polycations by sequentially performing ultrasound and stirring. This invention does not have special requirements for the power and time of the ultrasound, as long as the composite nanoparticles are evenly dispersed. The stirring time is preferably 4 hours.

[0071] In this invention, the composite nanoparticles are preferably further subjected to centrifugation and washing after being mixed with the polycation-containing solution, and the washing is preferably done with ultrapure water. When the polycation is preferably chitosan, the washing is preferably further performed with a 2% (v / v) acetic acid solution before the ultrapure water washing.

[0072] After obtaining the polycation-encapsulated composite nanoparticles, the present invention mixes the polycation-encapsulated composite nanoparticles with a solution containing hyaluronic acid to obtain the targeted nanotherapeutic agent.

[0073] In this invention, the polycation-encapsulated composite nanoparticles are first dispersed in water to obtain a polycation-encapsulated composite nanoparticle dispersion, and then mixed with a solution containing hyaluronic acid. The concentration of the polycation-encapsulated composite nanoparticle dispersion is preferably 0.1-4 mg / mL, more preferably 2 mg / mL.

[0074] In this invention, the concentration of hyaluronic acid in the hyaluronic acid-containing solution is preferably 0.1 to 10 mg / mL, more preferably 4 mg / mL.

[0075] In this invention, the volume ratio of the polycation-encapsulated composite nanoparticle dispersion to the hyaluronic acid-containing solution is preferably 1:1.

[0076] In this invention, the mass ratio of the polycation-encapsulated composite nanoparticles to hyaluronic acid is preferably 0.1–4:0.1–10, and more preferably 1:2.

[0077] In this invention, the solution containing hyaluronic acid preferably also includes one or more of sodium chloride, sodium sulfate and sodium phosphate, more preferably sodium chloride, and the concentration of sodium chloride in the solution containing hyaluronic acid is preferably 0.5 to 1 mol / L.

[0078] In this invention, the mixing temperature of the polycation-encapsulated composite nanoparticles with the hyaluronic acid-containing solution is preferably 2 to 30°C, more preferably 4°C; the mixing time is preferably 2 to 24 hours, more preferably 4 hours.

[0079] In this invention, the preferred method for mixing the polycation-encapsulated composite nanoparticles with the hyaluronic acid-containing solution is to sequentially perform ultrasonication and stirring, wherein the ultrasonication temperature is preferably 0–4°C and the stirring time is preferably 4 h.

[0080] In this invention, the polycation-encapsulated composite nanoparticles are mixed with a solution containing hyaluronic acid, and preferably then centrifuged and freeze-dried sequentially to obtain a freeze-dried powder of a targeted nanotherapeutic agent. The freeze-drying temperature is preferably -80°C, and the time is preferably 24 hours.

[0081] The present invention also provides a targeted nanotherapeutic agent obtained by the preparation method described above, wherein the encapsulation rate of the drug in the targeted nanotherapeutic agent is 85-98%.

[0082] In this invention, the encapsulation efficiency of the drug in the targeted nanotherapeutic agent is preferably greater than 90%. This invention combines supercritical carbon dioxide antisolvent technology and layer-by-layer self-assembly technology to prepare targeted nanotherapeutic agents, achieving co-delivery of multiple drugs with high encapsulation efficiency for all drugs.

[0083] The present invention also provides the application of the targeted nanotherapeutic agents described above in the preparation of drugs for tumor diagnosis and / or treatment.

[0084] In this invention, the tumor treatment preferably includes tumor photothermal therapy, photodynamic therapy, sonodynamic therapy or chemotherapy, and the targeted nanotherapeutic agent of this invention can effectively target the CD44 receptor.

[0085] To further illustrate the present invention, the targeted nanotherapeutic agents, their preparation methods, and applications of the present invention are described in detail below with reference to the accompanying drawings and embodiments, but these should not be construed as limiting the scope of protection of the present invention.

[0086] Example 1

[0087] An ethanol solution of indocyanine green (1 mg / mL, 5 mL) and a dichloromethane solution of paclitaxel (0.5 mg / mL, 5 mL) were prepared and mixed thoroughly to obtain a mixed solution. This mixed solution was pumped into a supercritical carbon dioxide antisolvent reactor under the following conditions: flow rate of the mixed solution 1 mL / min, pressure 9 MPa, and temperature 35 °C. After rinsing for 2 hours, indocyanine green / paclitaxel composite nanoparticles were obtained.

[0088] Six mg of indocyanine green / paclitaxel composite nanoparticles were added to a 1M sodium chloride solution containing polyarginine (0.1 mg / mL, 10 mL), ultrasonically dispersed, and stirred for 4 hours. The nanoparticles were then washed twice by centrifugation with ultrapure water. An aqueous dispersion (2 mg / mL, 1 mL) of the polycation-coated indocyanine green / paclitaxel composite nanoparticles was added to a 1M sodium chloride solution containing hyaluronic acid (4 mg / mL, 1 mL), ultrasonically dispersed, and stirred at 4°C for 4 hours to obtain a targeted nanotherapeutic agent loaded with indocyanine green and paclitaxel.

[0089] Figure 1 The infrared spectrum of the indocyanine green / paclitaxel composite nanoparticles obtained in Example 1 is shown at 1411 cm⁻¹. -1 It has a characteristic peak at 1718 cm⁻¹ belonging to the sulfonic acid group of ICG. -1The presence of a distinct characteristic peak at this point indicates that the peak is attributed to the stretching vibration of the ester group in the paclitaxel molecule, proving the successful preparation of indocyanine green / paclitaxel composite nanoparticles.

[0090] Figure 2 The image shows a scanning electron microscope (SEM) image of the targeted nanotherapeutic agent loaded with indocyanine green and paclitaxel obtained in Example 1. This demonstrates that the targeted nanotherapeutic agent constructed using the supercritical carbon dioxide fluid antisolvent method combined with the layer-by-layer self-assembly method has good dispersibility and a uniform spherical structure.

[0091] Figure 3 The image shows the encapsulation efficiency of the targeted nanotherapeutic agent loaded with indocyanine green and paclitaxel obtained in Example 1. The encapsulation efficiency of indocyanine green is 98.4%, and that of paclitaxel is 97.7%. The encapsulation efficiency of both indocyanine green and paclitaxel is greater than 90%.

[0092] Example 2

[0093] 10 mg of indocyanine green and 5 mg of paclitaxel were dissolved in 14 mL of ethanol. After complete dissolution, 7 mL of dichloromethane was added and mixed thoroughly to obtain a mixed solution. The mixed solution was pumped into a supercritical carbon dioxide antisolvent reactor. The reaction conditions were: flow rate of the mixed solution 0.75 mL / min, pressure 11 MPa, and temperature 35 °C. After rinsing for 2 hours, indocyanine green / paclitaxel composite nanoparticles were obtained.

[0094] 0.1 g of chitosan was dissolved in 100 mL of 2% (v / v) acetic acid solution, and 2.926 g (0.05 mol) of sodium chloride was added to obtain a chitosan-sodium chloride solution. 4 mg of indocyanine green / paclitaxel composite nanoparticles were added to the above chitosan-sodium chloride solution, ultrasonically dispersed, and stirred for 4 hours. The nanoparticles were then washed twice by centrifugation with 2% (v / v) acetic acid solution and twice by centrifugation with ultrapure water. An aqueous dispersion (2 mg / mL, 1 mL) of polycationically coated indocyanine green / paclitaxel composite nanoparticles was added to a 1 M sodium chloride solution containing hyaluronic acid (4 mg / mL, 1 mL). After ultrasonic dispersion, the nanoparticles were stirred at 4 °C for 4 hours to obtain a targeted nanotherapeutic agent loaded with indocyanine green / paclitaxel. The encapsulation efficiency of indocyanine green was 98.6%, and that of paclitaxel was 96.4%. The encapsulation efficiencies of both indocyanine green and paclitaxel were greater than 90%.

[0095] Comparative Example 1

[0096] This invention sets out indocyanine green-paclitaxel composite nanoparticles (particles without combined layer-by-layer self-assembly technology) prepared by supercritical carbon dioxide antisolvent method as Comparative Example 1. The preparation method is as follows: An ethanol solution of indocyanine green (1 mg / mL, 5 mL) and a dichloromethane solution of paclitaxel (0.5 mg / mL, 5 mL) are prepared and mixed thoroughly to obtain a mixed solution. The mixed solution is pumped into a supercritical carbon dioxide antisolvent reactor, and the reaction conditions are: flow rate of the mixed solution is 1 mL / min, pressure is 9 MPa, and temperature is 35℃; after rinsing for 2 hours, indocyanine green / paclitaxel composite nanoparticles are obtained.

[0097] Comparative Example 2

[0098] After preparing indocyanine green / paclitaxel composite nanoparticles using the same method as Comparative Example 1, 6 mg of the indocyanine green / paclitaxel composite nanoparticles were added to a 1M sodium chloride solution containing polyarginine (0.1 mg / mL, 10 mL). The nanoparticles were ultrasonically dispersed and stirred for 4 hours. The nanoparticles were then washed twice by centrifugation with ultrapure water to obtain polyarginine-coated indocyanine green / paclitaxel composite nanoparticles, which are non-targeted modified nanotherapeutic agents co-loaded with indocyanine green and paclitaxel.

[0099] Example 3

[0100] Cellular uptake experiments were conducted on the targeted nanotherapeutic agent co-loaded with indocyanine green and paclitaxel prepared in Example 1, using indocyanine green / paclitaxel composite nanoparticles constructed using supercritical carbon dioxide antisolvent technology as a control. The operation steps are as follows:

[0101] 4T1 cells were seeded in laser confocal microscopy dishes. Non-targeted nano-modified nano-therapeutic agents co-loaded with indocyanine green and paclitaxel, and targeted nano-therapeutic agents co-loaded with indocyanine green and paclitaxel were co-incubated with 4T1 cells for 6 hours. For the CD44 receptor blocking group, 10 mg / mL hyaluronic acid medium solution was prepared, added to 4T1 cells, and co-incubated for 1 hour. After washing, targeted nano-therapeutic agents co-loaded with indocyanine green and paclitaxel were added and co-incubated for 6 hours. The cells were then rinsed and fixed. The cell nuclei were stained with DAPI, and the cell uptake effect was observed and cell images were captured using laser confocal microscopy.

[0102] Experimental results of cellular uptake are as follows Figure 4As shown (left column: nuclear staining of 4T1 cells; middle column: intracellular fluorescence localization of ICG; right column: superimposed image of the left and middle columns), the first row shows the cellular uptake of indocyanine green and paclitaxel composite nanoparticles prepared with supercritical carbon dioxide as an antisolvent; the second row shows the non-targeted modified nanoparticles of indocyanine green and paclitaxel modified with polyarginine; the third row shows the cellular uptake of the targeted modified nanoparticles of indocyanine green and paclitaxel co-loaded with indocyanine green and paclitaxel; the fourth row shows the cellular uptake of the targeted modified nanoparticles of indocyanine green and paclitaxel co-loaded with CD44 blocking. Within the same time frame, the nanoparticle group without self-assembly modification showed no fluorescence, while the intracellular fluorescence of the targeted modified group was significantly higher than that of the untargeted modified nanoparticle group, with most particles distributed in the cytoplasm. For the CD44 receptor blocking group, the intracellular fluorescence of the targeted nanoparticles of indocyanine green and paclitaxel co-loaded with indocyanine green was observed to be very weak, and the uptake rate was significantly reduced, demonstrating the targeting and fluorescence stability of the nanoparticles.

[0103] Example 4

[0104] The targeted nanotherapeutic agent co-loaded with indocyanine green and paclitaxel obtained in Example 1, along with indocyanine green / paclitaxel composite nanoparticles constructed using supercritical carbon dioxide antisolvent technology, were dispersed in PBS solution. 100 μL of the indocyanine green / paclitaxel composite nanoparticles and the targeted nanotherapeutic agent co-loaded with indocyanine green and paclitaxel were injected intravenously via the tail vein at a concentration of 1 mg / kg of indocyanine green. Fluorescence signals from different sites in mice were collected at different injection times using a small animal in vivo efficacy system. Mice were euthanized 24 hours after injection, and the fluorescence signal intensity of their major organs and tumor tissues was analyzed. Animal imaging results at different time points are shown below. Figure 5 As shown, the fluorescence images of the animal's major organs 24 hours after injection are as follows. Figure 6 As shown, ( Figure 5 and Figure 6 The top image shows indocyanine green / paclitaxel composite nanoparticles constructed using supercritical carbon dioxide antisolvent technology, while the bottom image shows targeted nanotherapeutic agents co-loaded with indocyanine green and paclitaxel. It can be seen that the targeted nanotherapeutic agents co-loaded with indocyanine green and paclitaxel can effectively target tumor sites, while the untargeted nanoparticles are distributed throughout the body, demonstrating the targeting ability of the nanotherapeutic agents.

[0105] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, not all embodiments. People can obtain other embodiments based on the present invention without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A method for preparing a targeted nanotherapeutic agent, characterized in that, Includes the following steps: Prepare 5 mL of an ethanol solution with an indocyanine green concentration of 1 mg / mL and 5 mL of a dichloromethane solution with a paclitaxel concentration of 0.5 mg / mL. Mix the two solutions thoroughly to obtain a mixed solution. Pump the mixed solution into a supercritical carbon dioxide antisolvent reactor. The reaction conditions are: flow rate of the mixed solution of 1 mL / min, pressure of 9 MPa, and temperature of 35 °C. After rinsing for 2 hours, indocyanine green / paclitaxel composite nanoparticles are obtained. 6 mg of indocyanine green / paclitaxel composite nanoparticles were added to 10 mL of 1 M sodium chloride solution containing 0.1 mg / mL polyarginine. After ultrasonic dispersion, the mixture was stirred for 4 hours and washed twice by centrifugation with ultrapure water to obtain polycation-encapsulated indocyanine green / paclitaxel composite nanoparticles. 1 mL of an aqueous dispersion of polycation-encapsulated indocyanine green / paclitaxel composite nanoparticles at a concentration of 2 mg / mL was added to 1 mL of a 1M sodium chloride solution containing 4 mg / mL hyaluronic acid. After ultrasonic dispersion, the mixture was stirred at 4°C for 4 hours to obtain a targeted nanotherapeutic agent loaded with indocyanine green and paclitaxel.

2. The targeted nanotherapeutic agent obtained by the preparation method according to claim 1, characterized in that, The targeted nanotherapeutic agent had an indocyanine green encapsulation rate of 98.4% and a paclitaxel encapsulation rate of 97.7%.

3. The use of the targeted nanotherapeutic agent according to claim 2 in the preparation of medicaments for tumor diagnosis and / or treatment.

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