Polymer microspheres supported by activated metal nanoparticles, their manufacturing method, and applications.

Abstract

This invention relates to polymer microspheres supported by active metal nanoparticles, their manufacturing method, and applications, and belongs to the technical field of tumor embolization therapy. The polymer microspheres supported by active metal nanoparticles described in this invention include a polymer microsphere skeleton and active metal nanoparticles distributed inside the polymer microsphere skeleton. This invention mainly produces the target microspheres by a reversed-phase microemulsion method. The polymer microspheres supported by active metal nanoparticles produced in this invention are used in transarterial tumor embolization, embolizing tumor blood vessels and blocking nutrient supply to the tumor site, while also gradually and continuously releasing hydroxides within the epithelium to adjust the tumor microenvironment to a weakly acidic state, releasing hydrogen gas to activate the immune response, inducing hydrogen gas therapy, and amplifying the effect of embolization therapy. This demonstrates a superior combined therapeutic effect compared to embolization therapy using only polymer microspheres.

Description

Technical Field

[0001] The present invention relates to the technical field of tumor embolization therapy, and particularly to polymer microspheres carrying active metal particles, a method for manufacturing the same, and uses thereof.

Background Art

[0002] Liver cancer is one of the most common cancers in the world. In addition to common tumor treatment means (such as surgical resection, radiotherapy, chemotherapy, etc.), transarterial embolization / chemoembolization for the development of tumors in the liver has become the most common non-surgical treatment method. However, in the process of conventional embolization therapy, the lesion site is in a severe hypoxic environment, and a large number of hypoxia-tolerant liver cancer cells and tumor-derived stem cells accumulate in the liver lesion site. Therefore, the treatment of transarterial tumor embolization may end in failure. The microenvironment of excessive hypoxia is accompanied by a weakly acidic and immunosuppressive microenvironment at the same time, and further promotes the rapid proliferation of liver cancer cells. Furthermore, it is difficult to completely necrotize tumors with conventional transarterial tumor embolization. Therefore, in many cases, residual lesions of tumors may cause recurrence or metastasis after transarterial tumor embolization treatment.

[0003] Drug-loaded microspheres are a new type of embolization material that can carry chemotherapy drugs. When combined with chemotherapy drugs, they can continuously and slowly release the drugs, resulting in a high drug concentration at the tumor local area and a low drug concentration in the surrounding area, improving the effect of embolization therapy. Although such polymer drug-loaded microspheres have already been commonly used, they still cannot overcome drug resistance and tumor immune escape caused by the hypoxic environment at the tumor site. Therefore, it is necessary to develop a new type of embolization microsphere to improve the local microenvironment of transarterial tumor embolization and avoid the problem of poor chemotherapy effect due to tumor drug resistance.

Summary of the Invention

[0004] To solve the above technical problems, the present invention provides polymer microspheres supported by active metal nanoparticles, a method for producing the same, and its applications. The present invention mainly produces the target microspheres by the reversed-phase microemulsion method. Polymer microspheres supported by active metal nanoparticles produced by the present invention are used in transarterial tumor embolization, embolizing tumor blood vessels and blocking nutrient supply to the tumor site, while also gradually and continuously releasing hydroxides within the epithelium to adjust the tumor microenvironment to a weakly acidic state, releasing hydrogen gas to activate the immune response, inducing hydrogen gas therapy, and amplifying the effect of embolization therapy. This demonstrates a superior combined therapeutic effect compared to embolization therapy using only polymer microspheres.

[0005] This invention is realized by the following technical means. The first object of the present invention is to provide polymer microspheres supporting active metal nanoparticles, which include a polymer microsphere skeleton and active metal nanoparticles distributed inside the polymer microsphere skeleton. The shape of the activated metal nanoparticles is not necessarily a regular sphere, and includes, but is not limited to, regular spherical, ellipsoidal, polyhedral, or rod-shaped shapes, as well as irregular shapes such as sea urchin-shaped shapes. Therefore, the dimensions are expressed using the major axis, which is the longest dimension of the activated metal nanoparticles.

[0006] In one embodiment of the present invention, the major axis of the activated metal nanoparticles is less than or equal to the particle size of the polymer microspheres, thereby ensuring the loading efficiency and amount of activated metal nanoparticles, and making the morphology of the polymer microspheres loaded with activated metal nanoparticles more uniform, thereby ensuring the embolic effect.

[0007] In one embodiment of the present invention, the major axis of the active metal nanoparticles is 1 nm to 100 μm. In one embodiment of the present invention, the major axis of the activated metal nanoparticles is 1 nm to 0.5 μm, 0.5 μm to 2 μm, 2 μm to 5 μm, 5 μm to 10 μm, 10 μm to 15 μm, 15 μm to 20 μm, 20 μm to 25 μm, 25 μm to 30 μm, 30 μm to 35 μm, 35 μm to 40 μm, 40 μm to 45 μm, 45 μm to 50 μm, 50 μm to 55 μm, 55 μm to 60 μm, 60 μm to 65 μm, 65 μm to 70 μm, 70 μm to 75 μm, 75 μm to 80 μm, 80 μm to 85 μm, 85 μm to 90 μm, 90 μm to 95 μm, and 95 μm to 100 μm.

[0008] In one embodiment of the present invention, the particle size of the polymer microspheres is 10 μm to 500 μm. In one embodiment of the present invention, the particle size distribution of the polymer microspheres supporting the activated metal nanoparticles is 10 μm to 30 μm, 30 μm to 50 μm, 50 μm to 70 μm, 70 μm to 90 μm, 90 μm to 100 μm, 100 μm to 120 μm, 120 μm to 150 μm, 150 μm to 170 μm, 170 μm to 200 μm, 200 μm to 250 μm, 250 μm to 300 μm, 300 μm to 350 μm, 350 μm to 400 μm, 400 μm to 450 μm, and 450 μm to 500 μm.

[0009] In one embodiment of the present invention, the number of activated metal nanoparticles is one or more. In one embodiment of the present invention, the activated metal nanoparticles are one or more of the activated metal element, alloy, and metal composite structure.

[0010] In one embodiment of the present invention, the active metal element is selected from one or more of potassium, calcium, sodium, magnesium, aluminum, zinc, iron, and tin. In one embodiment of the present invention, the alloy comprises at least two metals with different reactivity.

[0011] In one embodiment of the present invention, the alloy comprises metallic magnesium and platinum. In one embodiment of the present invention, the metal composite structure comprises at least two metals with different reactivity.

[0012] In one embodiment of the present invention, the metal composite structure includes a core structure and a bonding layer bonded to the surface of the core structure, wherein the core structure is a first metal, the bonding layer is a second metal, and the first metal has different activity from the second metal. In one embodiment of the present invention, the shape of the core structure is spherical, polyhedral, porous, or rod-shaped.

[0013] In one embodiment of the present invention, the metallic activity of the first metal is stronger than the activity of hydrogen, that is, the first metal reacts with water to produce hydrogen gas. In one embodiment of the present invention, the first metal is selected from one or more of potassium, calcium, sodium, magnesium, aluminum, zinc, gallium, iron, manganese, and tin.

[0014] In one embodiment of the present invention, the metallic activity of the second metal is less than that of hydrogen. In one embodiment of the present invention, the second metal is selected from one or more of copper, silver, platinum, and gold.

[0015] In one embodiment of the present invention, the particle size distribution of the metal composite structure is uniform. In one embodiment of the present invention, the metal composite structure includes a magnesium-platinum composite structure.

[0016] A second object of the present invention is to provide a method for producing polymer microspheres supported with active metal nanoparticles. Step 1 involves preparing a polymer material into a polymer solution, Step 2 includes mixing the polymer solution described in Step 1 with activated metal nanoparticles and producing polymer microspheres supported by the microsphere manufacturing method.

[0017] In one embodiment of the present invention, the polymer material contains a functional group that is positively or negatively charged, thereby preventing the polymer microspheres from becoming electrically neutral. In one embodiment of the present invention, in step 1, the polymer material is a raw material that can be processed into microspheres.

[0018] In one embodiment of the present invention, in step 1, the polymer material is selected from one or more of gelatin, polyvinyl alcohol, chitosan, biodegradable polyurethane, silk fibroin, collagen, alginic acid, and biodegradable starch. In one embodiment of the present invention, the polymer solution includes a polymer solution capable of physical polymerization, chemical polymerization, or biological polymerization.

[0019] In one embodiment of the present invention, the chemical polymerization includes crosslinking and grafting. In one embodiment of the present invention, in step 2, the method for manufacturing the microspheres includes the inverse phase microemulsion method.

[0020] A third object of the present invention is to provide an embolization agent including polymer microspheres carrying the active metal microparticles.

[0021] In one embodiment of the present invention, the embolization agent further includes a dispersant. In one embodiment of the present invention, the dispersant is an aqueous solvent. In one embodiment of the present invention, the dispersant is selected from at least one of physiological saline, isotonic neutral buffer solution, and glucose solution.

[0022] A fourth object of the present invention is to provide a method for manufacturing an embolization agent, including the steps of drying and freezing polymer microspheres carrying active metal microparticles to obtain the embolization agent.

[0023] In one embodiment of the present invention, the drying is selected from any one of drying, freeze-drying, and air drying.

[0024] A fifth object of the present invention is to provide a tumor embolization agent including the embolization agent, and the tumor embolization agent forms an embolism at the lesion site by an administration method via arterial intervention.

[0025] A sixth object of the present invention is to provide a freeze-dried powder containing the embolic agent.

[0026] A seventh object of the present invention is to provide polymer microspheres supporting activated metal nanoparticles, wherein the activated metal nanoparticles are distributed within the polymer microspheres, or the polymer microspheres supporting the activated metal nanoparticles have a core-shell structure with the activated metal nanoparticles as the core and the polymer microspheres as the shell, the polymer microspheres covering the outer surface of the activated metal nanoparticles, or the polymer microspheres supporting the activated metal nanoparticles have voids inside the polymer microspheres, and the activated metal nanoparticles are distributed within these voids.

[0027] In one embodiment of the present invention, the activated metal nanoparticles are one or more of the activated metal element, alloy, and metal composite structure. In one embodiment of the present invention, the active metal element is selected from one or more of magnesium, aluminum, zinc, iron, manganese, tin, gallium, and indium.

[0028] In one embodiment of the present invention, the activated metal element is selected from one or more of magnesium, aluminum, and zinc, and the major axis of the activated metal nanoparticles is 1 nm to 150 μm. In one embodiment of the present invention, the active metal element is selected from one or more of iron, manganese, tin, gallium, and indium, and the major axis of the active metal nanoparticles is 1 nm to 5000 nm.

[0029] In one embodiment of the present invention, the particle size of the polymer microspheres is 10 μm to 500 μm. In one embodiment of the present invention, the activated metal nanoparticles are an alloy, and the major axis of the activated metal nanoparticles is 1 nm to 150 μm.

[0030] In one embodiment of the present invention, the alloy is a binary alloy and / or a ternary alloy. In one embodiment of the present invention, the alloy comprises metallic manganese and platinum, and metallic aluminum and gallium.

[0031] In one embodiment of the present invention, the active metal nanoparticles have a metal composite structure, and the major axis of the active metal nanoparticles is 1 nm to 150 μm. In one embodiment of the present invention, the metal composite structure includes a manganese-platinum composite structure and an aluminum-gallium composite structure. In one embodiment of the present invention, the polymer microspheres supporting the activated metal nanoparticles contain a functional group that is positively or negatively charged.

[0032] An eighth object of the present invention is to provide polymer drug-supported microspheres containing polymer microspheres supporting the above-mentioned active metal nanoparticles and a drug, wherein the drug is supported within the polymer microspheres supporting the active metal nanoparticles.

[0033] In one embodiment of the present invention, a drug is embedded or adsorbed onto the surface or inside of polymer microspheres supporting active metal nanoparticles by physical means. In one embodiment of the present invention, the drug and the polymer microspheres supporting the active metal nanoparticles are bound together by electrostatic action.

[0034] In one embodiment of the present invention, the polymer microspheres supporting the activated metal nanoparticles are composed of one or more polymer materials such as gelatin, polyvinyl alcohol, chitosan, biodegradable polyurethane, silk fibroin, collagen, alginic acid, and biodegradable starch.

[0035] In one embodiment of the present invention, the polymer microspheres supporting the active metal nanoparticles contain a functional group R that is positively or negatively charged, and the drug contains a functional group that is charged opposite to the functional group R. In one embodiment of the present invention, the functional group R is one or more of a carboxyl group, an aldehyde group, a sulfonic acid group, a cyano group, and a nitroamino group, and the drug is selected from one of epirubicin, pirarubicin, irinotecan, gemcitabine, and aspirin.

[0036] In one embodiment of the present invention, the functional group R is an amino group, and the drug is selected from either sodium alendronate or methotrexate.

[0037] In one embodiment of the present invention, the polymer microspheres supporting the activated metal nanoparticles may also be modified with positively charged polymer molecules, which include, but are not limited to, polyallylamine hydrochloride, polyethyleneimine, polyvinylpyrrolidone, cellulose, carboxyl-modified cellulose, and polyvinyl alcohol.

[0038] By modifying polymer microspheres supporting active metal nanoparticles with positively charged polymer molecules, the charge level of the polymer microspheres supporting the active metal nanoparticles can be increased, thereby improving the mass of the drug supported by the polymer microspheres supporting the active metal nanoparticles per unit solid phase volume.

[0039] In one embodiment of the present invention, the polymer microspheres supporting the active metal nanoparticles may also be modified with negatively charged polymer molecules, and the negatively charged polymer molecules include, but are not limited to, polyacrylic acid, polyethylene glycol, polylactic acid, polyglycolic acid, and polylactic acid-glycolic acid copolymers.

[0040] By modifying polymer microspheres supporting active metal nanoparticles with negatively charged polymer molecules, the charge level of the polymer microspheres supporting the active metal nanoparticles can be increased, thereby improving the mass of the drug supported by the polymer microspheres supporting the active metal nanoparticles per unit solid phase volume.

[0041] In one embodiment of the present invention, the mass of the drug supported by the polymer microspheres carrying the active metal nanoparticles per unit solid phase volume is 0 to 30 mg / mL. Preferably, the mass of the drug supported by the polymer microspheres carrying the active metal nanoparticles per unit solid phase volume is 0 mg / mL or more.

[0042] In one embodiment of the present invention, the mass of the drug supported by the polymer microspheres carrying the active metal nanoparticles per unit solid phase volume is greater than 0 mg / mL, and the mass of the drug supported by the polymer microspheres carrying the active metal nanoparticles per unit solid phase volume is 30 mg / mL or less.

[0043] The ninth object of the present invention is to provide a method for producing polymer drug-carrying microspheres. Step 1 involves preparing a polymer material into a polymer solution, Step 2 includes mixing the polymer solution described in Step 1 with activated metal nanoparticles and producing polymer microspheres supported by the microsphere manufacturing method. Step 3 includes mixing the polymer microspheres supported by the activated metal nanoparticles produced in Step 2 with a drug solution and shaking to obtain the polymer drug-supported microspheres.

[0044] In one embodiment of the present invention, in step 1, the polymer material is a raw material that can be processed into microspheres. In one embodiment of the present invention, in step 1, the polymer material is selected from one or more of gelatin, polyvinyl alcohol, chitosan, biodegradable polyurethane, silk fibroin, collagen, alginic acid, and biodegradable starch.

[0045] In one embodiment of the present invention, the polymer solution includes a polymer solution capable of physical polymerization, chemical polymerization, or biological polymerization. In one embodiment of the present invention, the chemical polymerization includes crosslinking and grafting.

[0046] In one embodiment of the present invention, in step 2, the method for producing the microspheres includes a reversed-phase microemulsion method. In one embodiment of the present invention, in step 2, the reversed-phase microemulsion method is realized by microfluidic technology, allowing for better control of the size of the microspheres.

[0047] In one embodiment of the present invention, the polymer microspheres supporting the obtained activated metal nanoparticles are further subjected to a solidification treatment. In one embodiment of the present invention, the solidification treatment is selected from one or more of the following: photosolidification, thermal solidification, X-ray solidification, ultrasonic solidification, and cryosolidification, and the photosolidification is solidification by ultraviolet irradiation.

[0048] In one embodiment of the present invention, the solidification treatment is achieved by adding a solidifying agent to a polymer solution, the solidifying agent comprising methacrylic anhydride, 1-ethyl-(3-dimethylaminopropyl)carbodiimide, and the like. In one embodiment of the present invention, the polymer solution further contains polymer molecules that are positively or negatively charged.

[0049] The negatively charged polymer molecule is selected from one or more of the following: polyacrylic acid, polyethylene glycol, polylactic acid, polyglycolic acid, and polylactic acid-glycolic acid copolymer. The positively charged polymer molecule is selected from one or more of the following: polyallylamine, polyethyleneimine, polyvinylpyrrolidone, cellulose, carboxyl-modified cellulose, and polyvinyl alcohol.

[0050] By modifying a polymer material by adding positively or negatively charged polymer molecules to a polymer solution, the charge level of the produced polymer microspheres can be increased, thereby improving the rate and amount of drug loading by the polymer microspheres through electrostatic adsorption.

[0051] Polymer microspheres supported with the activated metal nanoparticles described in the present invention have the following beneficial effects. (1) The polymer microspheres supported by the active metal nanoparticles provided by the present invention can achieve vascular embolization at the lesion site, thereby blocking nutrient supply and achieving the effect of embolization therapy. In particular, when the active metal nanoparticles come into contact with the slightly acidic environment of the lesion site, they slowly react and can continuously generate hydrogen gas. The acidic microenvironment of a tumor can cause a series of negative symptoms such as tumor metastasis, tumor angiogenesis, and enhanced drug resistance of the tumor. The polymer microspheres supported by the active metal nanoparticles provided by the present invention react in the acidic microenvironment of the tumor to produce hydroxide and hydrogen gas. Of these, the hydroxide can adjust the pH of the tumor site, improve the acidic microenvironment of the tumor, and reduce the probability of tumor metastasis, angiogenesis, drug resistance, etc. Furthermore, the generated hydrogen gas selectively removes highly cytotoxic reactive oxygen species [hydroxyl radicals (·OH) and peroxonitrite ions (ONOO-)], disrupts the intracellular redox balance, induces apoptosis in tumor cells, modulates inflammatory responses, and thereby exerts a therapeutic effect on tumors. Simultaneously, hydrogen molecules can suppress mitochondrial function in tumors, inhibit normal cellular energy metabolism, and cause tumor cell death. Moreover, it can retain ROS necessary for signal transduction in normal cells, and has low toxicity and side effects. The polymer microspheres supported by the active metal nanoparticles described in this invention can gently and continuously generate hydrogen gas and hydroxides, possessing the dual functions of improving the tumor microenvironment and hydrogen gas therapy.

[0052] (2) The method for producing polymer microspheres supported by activated metal nanoparticles as described in the present invention is simple, allows for mass production, and employs a microfluidic process to enable controllable and uniform production of the product. The product is stable in a dry state and easy to store and transport.

[0053] (3) The tumor embolic agent described in the present invention combines the characteristics of embolization with high molecular weight microspheres and hydrogen gas therapy, and has superior therapeutic effect compared to blank polyvinyl alcohol (PVA) embolic microspheres. Furthermore, since the administration process of the formulation itself is carried out in an aqueous phase environment, it has higher applicability and safety, and is biodegradable, so it gradually decomposes after fully exerting its embolization and hydrogen generation functions, and has superior biosafety.

[0054] (4) The polymer microspheres supporting the activated metal nanoparticles described in the present invention can have a core-shell structure, and drugs can be supported on a shell layer made of polymer material by electrostatic adsorption to form polymer drug-supported microspheres supporting the activated metal nanoparticles. The polymer drug-supported microspheres generate hydrogen gas and release drugs very slowly in a neutral environment, avoiding toxicity and side effects to the normal tissues of the body, and thus providing better safety.

[0055] (5) In the slightly acidic environment of the tumor site, the polymer drug-carrying microspheres carrying the active metal nanoparticles can generate hydroxide and hydrogen gas, which regulates the tumor microenvironment and inflammatory response. At the same time, the physical vibrations caused by gas generation increase the release rate of the carried drug, enabling the combined use of chemotherapy and embolic immunotherapy and achieving a superior therapeutic effect of arterial chemoembolization.

[0056] (6) The drug loading process of polymer drug-loaded microspheres supported by the active metal nanoparticles described in the present invention is simple, and the drug loading efficiency is high, making it convenient for actual clinical use.

[0057] To make the contents of the present invention easier to understand, the present invention will be described in more detail below with reference to the drawings, based on specific embodiments of the present invention. [Brief explanation of the drawing]

[0058] [Figure 1] Figure 1 is a schematic diagram illustrating the production of polymer microspheres supporting activated metal nanoparticles using the microfluidic engineering technique described in Example 3. [Figure 2]Figure 2 is an optical microscope image of polymer microspheres supported with the activated metal nanoparticles described in Example 2, which were manufactured by the method described in Example 2. [Figure 3] Figure 3 shows the nuclear magnetic resonance hydrogen spectrum of gelatin protein grafted with methacrylic anhydride (MA) in Example 2. [Figure 4] Figure 4 is an optical microscope image of polymer microspheres supported with the activated metal nanoparticles described in Example 3, which were manufactured by the method described in Example 3. [Figure 5] Figure 5 is an optical microscope image of polymer microspheres supported with the activated metal nanoparticles described in Example 4, which were manufactured by the method described in Example 5. [Figure 6-a] Figure 6-a is a DSA image of the hepatic artery in Example 7 before embolization with polymer microspheres carrying active metal nanoparticles. [Figure 6-b] Figure 6-b is a DSA image of the hepatic artery after embolization with polymer microspheres carrying active metal nanoparticles in Example 7. [Figure 7] Figure 7 is a signal diagram for detecting the embolic site using ultrasound imaging in Example 7. [Figure 8] Figure 8 is a statistical table summarizing the tumor size changes in each group of rabbits in Example 8. [Figure 9] Figure 9 is an optical microscope image of polymer microspheres supported with active metal nanoparticles produced in Examples 9.1A to 9.1E. [Figure 10] Figure 10 is an optical microscope image of polymer microspheres supported with active metal nanoparticles produced in Examples 9.2A to 9.2E. [Figure 11] Figure 11 is an optical microscope image of polymer microspheres supported with active metal nanoparticles produced in Examples 9.3A to 9.3D. [Figure 12] Figure 12 is an optical microscope image of polymer microspheres supported with active metal nanoparticles produced in Examples 9.4A to 9.4D. [Figure 13]Figure 13 is an optical microscope image of polymer microspheres supported with active metal nanoparticles produced in Examples 9.5A to 9.5E. [Figure 14] Figure 14 is an optical microscope image of the GelMA-PAA-Mg microspheres produced in Example 9.6. [Figure 15] Figure 15 shows the release curves of the drug (EPI) released by polymer drug-carrying microspheres in various buffer solutions in Example 10.2. [Figure 16] Figure 16 is an optical microscope image of polymer microspheres (polyvinyl alcohol microspheres) supported with active metal nanoparticles (magnesium nanoparticles) as described in Example 13. [Figure 17] Figure 17 is a statistical chart summarizing the relative viability of cells after treatment with different samples in Example 15. [Figure 18] Figure 18 is a confocal image of a cross-section of a tumor cell in Example 16, where the fluorescence signal originates from the chemotherapeutic drug released from the high molecular weight drug-carrying microspheres. [Figure 19] Figure 19 shows the tumor volume of a mouse H22 subcutaneous tumor model after injection of high molecular weight drug-carrying microspheres into the tumor of a mouse in Example 17. [Figure 20] Figure 20 shows photographs of the auricular artery of a New Zealand rabbit before and after embolization. [Modes for carrying out the invention]

[0059] The present invention will be further described below with reference to the attached drawings and specific embodiments, so that those skilled in the art may better understand and implement the present invention. However, the embodiments described are not intended to limit the present invention.

[0060] To explain the technical solutions of the above invention in more detail, specific examples are given below to demonstrate the technical effects. It should be emphasized that these examples are for the purpose of illustrating the present invention, but do not limit the scope of the present invention.

[0061] As set forth herein and in the claims, unless the context clearly indicates an exception, terms such as “one,” “a kind,” and / or “the” are not limited to the singular but may include the plural. The terms “includes” and “contains” merely indicate the inclusion of clearly identified elements, and these elements do not constitute an exclusive enumeration; the apparatus may include other elements. The term “one embodiment” means “at least one embodiment,” and the term “other embodiment” means “at least one other embodiment.” Definitions of other terms are set forth below.

[0062] As described herein, the activated metal nanoparticles are usually spherical, but may also be ellipsoidal or irregular in shape, and unless otherwise specified, the dimensions are all defined by the major axis, the major axis of the spherical nanoparticles being equal to the particle size.

[0063] Description of experimental materials: Polyacrylic acid (PAA, MW1800) and lithium phenyl-2,4,6-trimethylbenzoylphosphinate were both purchased from Sigma-Aldrich.

[0064] Example 1: Polymer microspheres supported with activated metal nanoparticles. Example 1.1: A polymer microsphere supporting activated metal nanoparticles, wherein the polymer is methacrylic anhydride-modified gelatin protein, and each polymer microsphere supporting the activated metal nanoparticles contains one or more magnesium nanoparticles. The average particle size of the internal magnesium nanoparticles is approximately 20 μm, and the methacrylic anhydride-modified gelatin protein uniformly coats the surface of the magnesium nanoparticles. The methacrylic anhydride-modified gelatin protein has methacrylic anhydride side chains and possesses photosensitive solidification properties, contributing to the stability of the product. As shown in Figure 2, the particle size of the polymer microspheres supporting the active metal nanoparticles is approximately 70 μm. They have a smooth appearance, uniform magnesium nanoparticle support inside, relatively uniform morphology, good dispersibility, no aggregation, and are convenient for embolization.

[0065] Because ionic bonds are in place between the surface of magnesium nanoparticles and methacrylic anhydride gelatin protein, the supported state is stable. When methacrylic anhydride gelatin protein microspheres supporting magnesium nanoparticles are applied to tumor embolization, the magnesium nanoparticles react in the acidic microenvironment of the tumor, and their size gradually decreases. Due to the presence of ionic bonding, the magnesium nanoparticles are less likely to leak out of the high molecular weight methacrylic anhydride gelatin protein spheres. As a result, the magnesium nanoparticles remain at the embolization site due to the action of the high molecular weight methacrylic anhydride gelatin protein outer shell, allowing them to exert a sustained effect.

[0066] Example 1.2: Polymer microspheres supported by activated metal nanoparticles, wherein the polymer is methacrylic anhydride-modified gelatin protein, and each polymer microsphere supporting the activated metal nanoparticles contains one or more magnesium-platinum alloy nanoparticles. The particle size of the magnesium-platinum alloy nanoparticles is 35 μm, and the particle size of the polymer microspheres is 200 μm. Because the metal alloy nanoparticles contain two types of metals with different reactivity, they have a better hydrogen gas generation rate in aqueous solutions or acidic microenvironments, and the effects of hydrogen gas therapy can be achieved even with a lower content of activated metal nanoparticles.

[0067] Example 1.3: Polymer microspheres supported by activated metal nanoparticles, wherein the polymer is methacrylic anhydride-modified gelatin protein, and each polymer microsphere supporting the activated metal nanoparticles contains one or more magnesium-platinum composite structures. The particle size of the magnesium-platinum composite structure is 35 μm, and magnesium is substituted onto the surface of the platinum core by a proto-cell reaction. The particle size of the polymer microspheres is 200 μm. The magnesium-platinum composite structure also has better hydrogen gas generation efficiency because the two metals with different reactivity have better hydrogen gas generation efficiency, so the effect of hydrogen gas therapy can be achieved even with a lower content of activated metal nanoparticles. Furthermore, the metal composite structure has better manufacturing controllability, resulting in better controllability of the manufacturing process and metal content.

[0068] Example 2: A method for producing polymer microspheres supported by the activated metal nanoparticles described in Example 1.1. The manufacturing method of this embodiment includes the steps of mixing an oily outer phase with an aqueous inner phase containing a polymer and magnesium fine particles, stirring the mixture, and then irradiating it with an ultraviolet lamp to solidify the mixed phase, thereby obtaining a product of methacrylic anhydride gelatin protein microspheres supported with magnesium fine particles. The product can be separated from the solvent phase by methods such as centrifugation.

[0069] As shown in Figure 1, the specific manufacturing process involves first modifying gelatin protein to obtain gelatin protein grafted with methacrylic anhydride (MA). Specifically, 2.5 g of gelatin protein is dissolved in 25 mL of DPBS solution, heated to 60°C to dissolve completely, then 2 mL of methacrylic anhydride is added, and the mixture is reacted in a 50°C water bath with stirring for 3 hours. After that, dialysis is performed in a 40°C water bath, and freeze-drying is performed to obtain methacrylated gelatin protein (GelMA). Figure 3 is the nuclear magnetic resonance hydrogen spectrum of the methacrylic anhydride (MA) grafted gelatin protein in Example 2.

[0070] Furthermore, 100 mg of methacrylate-oxidized gelatin protein and 5 mg of lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) were weighed, dissolved in 1 mL of deionized water, and heated to 60°C to completely dissolve them to obtain solution a. 100 mg of magnesium nanoparticles were weighed and dispersed in 1 mL of solution a to obtain solution b. Simultaneously, 10 mL of mineral oil containing 2% Span 80 was prepared to form liquid phase c. During the stirring of liquid phase c, solution b is added dropwise, stirring is continued for approximately 10 minutes after the addition is complete, and the product is solidified by irradiation with an ultraviolet lamp to obtain polymer microspheres supported by the activated metal nanoparticles of Example 1.1.

[0071] Figure 2 is a microscopic view of polymer microspheres supported by activated metal nanoparticles, manufactured in Example 2, observed using a Leica optical microscope. As can be seen from the figure, the particle size distribution of the microspheres is 30 μm to 100 μm. Since the particle size of the raw material magnesium nanoparticles is approximately 20 μm, each polymer microsphere supported by activated metal nanoparticles shown in the figure contains one or more magnesium nanoparticles. This allows for a balance between embolic effect, hydrogen gas generation efficiency, and duration of action, thereby enhancing the combined effect of embolic-hydrogen gas therapy.

[0072] Example 3: A method for producing polymer microspheres supported by the activated metal nanoparticles described in Example 1.1. This embodiment differs from Example 2 in that it is manufactured using microfluidic technology. Based on the methacrylic anhydride gelatin and magnesium nanoparticles obtained in Example 2, a subsequent step involves using microfluidic technology to produce polymer microspheres supporting the product's active metal nanoparticles.

[0073] Solution a is obtained by weighing 100 mg of methacrylic anhydride-modified gelatin and 5 mg of lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP), dissolving them in 1 mL of deionized water, and heating to 60°C to ensure complete dissolution. 100 mg of magnesium nanoparticles are weighed and dispersed in the 1 mL of solution a to form the internal phase. Mineral oil containing 2% Span 80 is used as the external phase. The internal and external phases are flowed through a microfluidic chip, with the internal phase flow rate set to 5 μL / min to 20 μL / min and the external phase flow rate to 100 μL / min to 500 μL / min. The resulting product is immediately irradiated with an ultraviolet lamp (395 nm) to solidify, thereby obtaining gelatin protein polymer microspheres containing magnesium nanoparticles internally, i.e., polymer microspheres supporting the active metal nanoparticles described in Example 1.1.

[0074] Figure 4 is an optical microscope image of polymer microspheres produced by the method described in Example 3. As is clear from the figure, the polymer microspheres supported by the activated metal nanoparticles have a smooth surface, good dispersibility, a uniform particle size of approximately 100 μm, and are stable in a neutral aqueous dispersant.

[0075] Example 4: Polymer microspheres supported with activated metal nanoparticles. This embodiment provides polymer microspheres supported by active metal nanoparticles, where the polymer is polycaprolactone and the metal is magnesium. As shown in Figure 5, their morphology under optical microscopy is smooth, with a uniform particle size of approximately 60 μm, meeting the particle size requirements necessary for embolusing peripheral blood vessels. They can be used for vascular embolization at lesion sites, blocking nutrient supply to tumors and suppressing tumor growth. Furthermore, the polymer microspheres supported by the active metal nanoparticles contain one or more magnesium nanoparticles with a particle size of 1 μm or less, and because they have a larger specific surface area, these polymer microspheres supported by the active metal nanoparticles have a higher hydrogen generation efficiency.

[0076] Example 5: A method for producing polymer microspheres supported by the activated metal nanoparticles described in Example 4. The polymer microspheres supported by the activated metal nanoparticles in this embodiment include the preparation of a solution of magnesium nanoparticles with a particle size of 1 μm or less and a PVA aqueous phase using microfluidic technology.

[0077] 100 mg of polycaprolactone (PCL) is weighed and dissolved in 1 mL of dichloromethane, then 20 mg of nano-magnesium spheres (Nano-Mg, size less than 1 μm) are added, and the resulting solution system is used as the inner phase. A 3% PVA (Type 1788, alcohol decomposition rate 87%-89%) solution was prepared and used as the outer phase. The internal and external phases are flowed through a microfluidic chip, with the internal phase flow rate set to 5 μL / min to 20 μL / min and the external phase flow rate to 100 μL / min to 500 μL / min. The resulting mixed phase is stirred overnight to completely volatilize the dichloromethane and precipitate and solidify the polycaprolactone, thereby obtaining polycaprolactone polymer microspheres (PCL-mgMSs) containing nanomagnesium spheres internally, i.e., polymer microspheres supporting the active metal nanoparticles described in the present invention.

[0078] Example 6: Tumor embolic agent. The tumor embolic agent provided in this embodiment includes polymer microspheres supported by active metal nanoparticles as described in Example 1.1, Example 1.2, Example 1.3, or Example 4. It is stored in a dry state under nitrogenous conditions, and when in use, the polymer microspheres supported by the active metal nanoparticles are dispersed in an aqueous solvent such as physiological saline, isotonic neutral buffer, glucose solution, or other biocompatible solution, and delivered to the site to be embolic via an intervention catheter to perform embolic therapy.

[0079] Example 7: Verification experiment of arterial embolization and gas generation effects of polymer microspheres supported with active metal nanoparticles in a liver tumor model. A New Zealand rabbit model of hepatoma in situ was constructed. After successful model construction, polymer microspheres carrying the active metal nanoparticles described in Example 1.1 were dispersed in physiological saline. Under the guidance of DSA contrast, these microspheres were implanted into the hepatic artery via catheter intervention to achieve embolization.

[0080] Figure 6 shows DSA images before (Figure 6-a) and after (Figure 6-b) embolization of the hepatic artery with polymer microspheres carrying active metal nanoparticles. Clearly, a significant change is observed in the vascular image at the indicated site, indicating that blood flow is blocked due to embolization of the vessel. This demonstrates that the polymer microspheres carrying active metal nanoparticles described in Example 1.1 have a hepatic artery embolization effect, and verifies that the polymer microspheres carrying active metal nanoparticles described in this application can be used for hepatic artery embolization.

[0081] Approximately 8 hours after interventional embolization, the gas generation status at the lesion site was detected by ultrasound. Figure 7 shows the signal diagram for detecting the embolic site using ultrasound imaging, and the three groups are as follows.

[0082] Example 7.1: This was a blank control group and did not receive interventional embolization therapy. Example 7.2: A group of polyvinyl alcohol microspheres, commercially available polyvinyl alcohol microspheres, brand Alcon, with a size of 150 μm to 300 μm, and a dosage of 5 mg to 8 mg, determined based on the actual embolic endpoint of each sample. Example 7.3: This is a polymer microsphere supported with the activated metal nanoparticles described in Example 1.1, with a particle size of approximately 70 μm and a dose of approximately 10 mg, which is determined based on the actual embolic endpoint of each sample.

[0083] The indicated area in Figure 7 is the liver tumor site. Since the ultrasound signal originates from the gas at the detection site, a stronger signal indicates a larger amount of gas. Observation revealed that no gas was generated at the liver tumor site in the samples from Examples 7.1 and 7.2, whereas a gas signal was clearly detected at the liver tumor site in the sample from Example 7.3. This indicates that the supported active metal nanoparticles reacted in the physiological environment to generate hydrogen gas. This verifies that the polymer microspheres supported by the active metal nanoparticles described in the present invention can generate gas at the embolic site, meaning that the polymer microspheres supported by the active metal nanoparticles described in the present invention have the potential to synergistically perform embolization therapy, tumor microenvironment modification, and hydrogen gas therapy.

[0084] Example 8: Experimental study on the therapeutic effect of polymer microspheres supported with active metal nanoparticles in a liver cancer model of New Zealand rabbits. A New Zealand rabbit model of hepatoma in situ was constructed. After successful model construction, polymer microspheres carrying the active metal nanoparticles described in Example 1.1 were dispersed in physiological saline. Under the guidance of DSA contrast, these microspheres were implanted into the hepatic artery via catheter intervention to achieve embolization. The groups for embolization administration are as follows:

[0085] Example 8.1: This was a blank control group and did not receive interventional embolization therapy. Example 8.2: This is a group of polyvinyl alcohol microspheres, using commercially available polyvinyl alcohol microspheres (Brand Alcon, specification 150 μm to 300 μm), with a dosage of approximately 5 mg to 8 mg, determined based on the actual embolic endpoint of each sample. Example 8.3: A polymer microsphere supported with the activated metal nanoparticles described in Example 1.1, with a particle size of approximately 70 μm, and a dose of approximately 10 mg, determined based on the actual embolic endpoint of each sample.

[0086] The day of embolization was counted as day 1, and CT scans were performed on New Zealand rabbits on days 0, 7, and 14 to observe tumor size. Figure 8 is a statistical table summarizing the changes in tumor size in each group of rabbits. The results showed that the growth of liver cancer in the New Zealand rabbits in Example 8.3 was suppressed, indicating that the polymer microspheres supported by the active metal nanoparticles described in Example 1.1 have a superior therapeutic effect compared to commercially available polyvinyl alcohol microspheres. In other words, the polymer microspheres supported by the active metal nanoparticles described in the present invention have a superior therapeutic effect.

[0087] Example 9.1: Polymer microspheres supported with activated metal nanoparticles The method for producing polymer microspheres supported by active metal nanoparticles according to this embodiment includes a step of using methacrylate-oxidized gelatin protein (GelMA) and magnesium nanoparticles produced in Example 2 as raw materials and employing a reversed-phase microemulsion method, and the specific production method is as follows.

[0088] Solution a was obtained by weighing different masses of methacrylate-oxidized gelatin protein (GelMA) and 5 mg of lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP), adding them to 1 mL of deionized water, and heating to 60°C to dissolve them completely. Solution b was obtained by weighing 100 mg of magnesium nanoparticles with an average major diameter of 20 μm and dispersing them in 1 mL of solution a. Simultaneously, 10 mL of mineral oil containing 1% Span 80 was prepared to form liquid phase c. Solution b was added dropwise to liquid phase c while stirring, and stirring was continued for 2-3 minutes after the addition was complete, with a stirring speed of approximately 300 r / min throughout the entire process. After stirring was complete, the product was irradiated with an ultraviolet lamp for 3-5 minutes to solidify, and washed 3-5 times by centrifugation to obtain polymer microspheres supported by active metal nanoparticles. The amounts of methacrylate-oxidized gelatin protein added in Examples 9.1A to 9.1E are shown in Table 1. [Table 1]

[0089] The results of observing the morphology of the manufactured microspheres using an optical microscope are shown in Figure 9. In Example 9.1A, when the GelMA concentration was 2.5%, the active metal nanoparticles were not effectively coated. However, as the GelMA concentration increased, the thickness of the polymer coating layer on the surface of the active metal nanoparticles increased, and GelMA microspheres carrying magnesium nanoparticles with a core-shell structure were formed, and the particle size of the product was clearly larger even to the naked eye. From these results, it was found that by selecting a GelMA concentration of 2.5% or higher in the manufacturing process, the active metal nanoparticles can be effectively loaded, and effective coating of metal nanoparticles by methacrylate-oxidized gelatin protein (GelMA) is achieved, which facilitates subsequent drug loading and concomitant embolization therapy.

[0090] Example 9.2: Polymer microspheres supporting activated metal nanoparticles of different sizes The method for producing polymer microspheres supported by activated metal nanoparticles according to this embodiment involves using methacrylate-oxidized gelatin protein (GelMA) and magnesium nanoparticles produced in Example 2 as raw materials, and employing a reversed-phase microemulsion method to produce polymer microspheres supported by activated metal nanoparticles according to this embodiment. The specific production method is the same as in Example 9.1, with the differences being as follows: The GelMA concentration in solution a is 10%, the average major axis of the magnesium nanoparticles is different, the major axis of the magnesium nanoparticles used is as shown in Table 2, and the span 80 concentration is 1%. [Table 2]

[0091] The results of observing the morphology of the microspheres produced using an optical microscope are shown in Figure 10. Increasing the size (major axis) of magnesium nanoparticles affects the loading efficiency by the polymer. The larger the major axis of the magnesium nanoparticles, the larger the size of the polymer microspheres, and the thinner the polymer layer becomes, resulting in a smaller number of magnesium nanoparticles contained in a single microsphere. When the average major axis of the magnesium nanoparticles exceeds 150 μm (see the corresponding test results in Example 9.2D (average major axis of magnesium nanoparticles is approximately 190 μm) and Example 9.2E (average major axis of magnesium nanoparticles is approximately 240 μm)), the polymer layer coated on the surface of the nanoparticles becomes noticeably thinner. In actual use, this may lead to excessively rapid degradation of the magnesium nanoparticles or a decrease in the embolization effect, and simultaneously be detrimental to subsequent drug loading. Verification results show that when the major axis of the active metal nanoparticles is 150 μm or less, polymer microspheres produced by the reversed-phase microemulsion method are relatively ideal and convenient for subsequent use.

[0092] Example 9.3: Polymer microspheres carrying activated metal nanoparticles produced at different surfactant concentrations The method for producing polymer microspheres supported by activated metal nanoparticles according to this embodiment involves using methacrylate-oxidized gelatin protein (GelMA) and magnesium nanoparticles produced in Example 2 as raw materials, and employing a reversed-phase microemulsion method to produce polymer microspheres supported by activated metal nanoparticles according to this embodiment. The specific production method is the same as in Example 9.1, with the differences being as follows: The GelMA concentration in solution a is 10%, the average major diameter of the magnesium nanoparticles is 20 μm, and the concentrations of span 80 are 0.5%, 1%, 2%, and 5%, respectively. [Table 3]

[0093] The results of observing the morphology of the microspheres produced using an optical microscope are shown in Figure 11. In Example 9.3A, the concentration of Span 80 used in the manufacturing process was lower, resulting in larger polymer microspheres, approximately 300 μm in size, and a greater number of active metal nanoparticles supported within a single polymer microsphere. When the concentration of Span 80 was higher (Example 9.3D), the size of the polymer microspheres was smaller, approximately 100 μm, and the number of active metal nanoparticles supported within a single polymer microsphere decreased slightly to approximately 1-3. From these results, it can be seen that the size of the microspheres produced in Examples 9.3A to 9.3D is ideal, less than 500 μm, and meets the size requirements for embolization therapy. In actual use, the size of the polymer microspheres and the actual number of supported active metal nanoparticles can be controlled by adjusting the concentration of Span 80 as needed.

[0094] Example 9.4: Polymer microspheres carrying activated metal nanoparticles produced at different stirring speeds The method for producing polymer microspheres supported by activated metal nanoparticles according to this embodiment involves using methacrylate-oxidized gelatin protein (GelMA) and magnesium nanoparticles produced in Example 2 as raw materials, and employing a reversed-phase microemulsion method to produce polymer microspheres supported by activated metal nanoparticles according to this embodiment. The specific production method is the same as in Example 9.1, with the differences being as follows: the GelMA concentration in solution a is 10%, the average major diameter of the magnesium nanoparticles is 20 μm, the concentration of span 80 is 1%, and the stirring speed is different. The stirring speeds corresponding to each embodiment are shown in Table 4. [Table 4]

[0095] The results of observing the morphology of the microspheres produced using an optical microscope are shown in Figure 12. The lower the stirring speed, the larger the size of the polymer microspheres. For example, the particle size of the microspheres produced in Example 9.4A was approximately 200-500 μm, and the number of active metal nanoparticles supported within a single polymer microsphere was greater. The size of the polymer microspheres produced in Examples 9.4A to 9.4D gradually decreased, to approximately 100-200 μm, and the number of active metal nanoparticles supported within a single polymer microsphere decreased slightly to approximately 1-5 particles. From these results, it was found that increasing the stirring speed reduces the particle size of the product. At the same time, the size of the microspheres produced at different stirring speeds was 500 μm or less, satisfying the size requirements for embolization therapy. Therefore, in actual use, the size of the polymer microspheres and the actual number of supported active metal nanoparticles can be controlled by adjusting the stirring speed as needed. This indicates that there is some room for adjustment in the production method, and the adjustment method is simple, making it advantageous to adjust the parameters as needed to produce products of corresponding specifications.

[0096] Example 9.5: Polymer microspheres supporting activated metal nanoparticles produced with different stirring times The method for producing polymer microspheres supported by activated metal nanoparticles according to this embodiment involves using methacrylate-oxidized gelatin protein (GelMA) and magnesium nanoparticles produced in Example 2 as raw materials, and employing a reversed-phase microemulsion method to produce polymer microspheres supported by activated metal nanoparticles according to this embodiment. The specific production method is the same as in Example 9.1, with the differences being as follows: The GelMA concentration in solution a is 10%, the average major diameter of the magnesium nanoparticles is 20 μm, the concentration of span 80 is 1%, the stirring speed is 300 r / min, and the stirring times corresponding to each embodiment are shown in Table 5. [Table 5]

[0097] The results of observing the morphology of the microspheres produced using an optical microscope are shown in Figure 13. From the results, it was found that the thickness of the polymer layer in the products of Examples 9.5A to 9.5E was directly reduced. This indicates that the shorter the stirring time, the thicker the polymer layer formed on the surface of the active metal nanoparticles, and the greater the number of active metal nanoparticles supported within a single polymer microsphere. Conversely, as the stirring time is extended, the polymer layer formed on the surface of the active metal nanoparticles gradually thins, and the number of active metal nanoparticles supported within a single polymer microsphere decreases. Therefore, although the stirring time has a relatively small effect on the size of the polymer microspheres, it affects parameters such as the loading ratio of polymer to active metal nanoparticles, and may further affect the drug loading amount and drug release effect. In actual use, the stirring time can be flexibly adjusted as needed to produce the desired product.

[0098] Example 9.6: Polymer microspheres supporting polymer-modified active metal nanoparticles In this example, polymer microspheres supported by polymer-modified active metal nanoparticles were produced using polyacrylic acid (PAA), an anionic polymer, as an example. The specific production method is as follows: Using methacrylate-oxidized gelatin protein (GelMA) and magnesium nanoparticles produced in Example 2 as raw materials, the specific production method is the same as in Example 9.1, the difference being that an additional 5 mg of polyacrylic acid (PAA), 100 mg of GelMA, and 5 mg of LAP were weighed and dissolved in 1 mL of deionized water to produce solution a. The results of observing the morphology of the methacrylate-oxidized gelatin protein microspheres supported by polyacrylic acid-modified magnesium nanoparticles (defined in this specification as GelMA-PAA-Mg microspheres) using an optical microscope are shown in Figure 14. The microspheres of the product have a particle size of approximately 50-200 μm and a regular spherical shape. The polymer microsphere layer and metal nanoparticles can be clearly distinguished, indicating that the methacrylate-oxidized gelatin protein layer effectively coats the magnesium nanoparticles and can further support drug loading.

[0099] Example 9.7: Production of polymer drug-carrying microspheres In this example, polymer drug-supported microspheres were produced using polymer microspheres supporting activated metal nanoparticles manufactured in Examples 9.1 to 9.6 as raw materials, and the specific manufacturing method is as follows: Polymer microspheres supporting activated metal nanoparticles were dispersed in an aqueous solvent, and polymer microspheres not supporting activated metal nanoparticles were removed by centrifugation (100 r / min, 3 min). The purified polymer microspheres supporting activated metal nanoparticles were added to a drug solution containing epirubicin hydrochloride (EPI) (2 mg / mL, 2 mL), shaken inverted up and down in a shaker for 3 to 5 minutes, and the supernatant was removed again by centrifugation (100 r / min, 3 min) to obtain polymer drug-supported microspheres (GelMA-Mg-EPI microspheres) that successfully supported EPI.

[0100] Observation of the product morphology using an optical microscope revealed no significant change in the morphology of the microspheres after drug loading, and the change in particle size was also smaller. This indicates that the microspheres obtained by the above manufacturing method have good stability. Observation showed that when the polymer drug-loaded microspheres produced in this example were left in a neutral buffer (physiological saline) for one day under room temperature conditions, the drug loading remained stable, and no significant drug release was observed. This indicates that the method for producing polymer drug-loaded microspheres described in the present invention is simple, highly reproducible, and the product is relatively stable.

[0101] Example 10: Tumor microenvironment responsiveness test of polymer drug-carrying microspheres Example 10.1: Hydrogen release of polymer drug-carrying microspheres at different pH levels Using methacrylate-oxidized gelatin protein microspheres (defined herein as GelMA-Mg microspheres) supported with magnesium nanoparticles having a methacrylate-oxidized gelatin protein concentration of 10% in solution a prepared in Example 9.1C as a raw material, high molecular weight drug-supported microspheres (defined herein as GelMA-Mg-EPI microspheres) supported with epirubicin hydrochloride are prepared by referring to the method described in Example 9.7.

[0102] Simultaneously, methacrylate-oxidized gelatin protein microspheres without supported active metal nanoparticles (defined herein as GelMA microspheres) were produced as a control, and the method for producing them is as follows: The method was carried out with reference to the method in Example 9.2, the only difference being that the step of adding active metal nanoparticles to solution a to produce solution b was omitted, and GelMA microspheres were produced. Using the above GelMA microspheres as a raw material, methacrylate-oxidized gelatin protein microspheres supported with epirubicin hydrochloride (defined herein as GelMA-EPI microspheres) were produced with reference to the method in Example 9.7.

[0103] GelMA-Mg-EPI microspheres were placed in buffers of different pH levels, and the formation of bubbles in the buffers was observed. Additionally, GelMA microspheres and GelMA-EPI microspheres were placed in acidic buffers at pH 5.5, respectively, and the formation of bubbles in the buffers was observed. The results are shown in Table 7. [Table 6]

[0104] As shown in the results in Table 7, the GelMA-Mg-EPI microspheres reacted vigorously in acidic buffer, indicating that the magnesium nanoparticles supported within them react with hydrogen ions in the solution under a weakly acidic environment, generating active metal hydroxides and hydrogen gas, thereby causing bubbles to form. At the same time, the reaction of GelMA-Mg-EPI microspheres was very weak (almost non-reactive) at a neutral pH (7.4), and since the pH of normal human tissue is 7.4, the reaction of GelMA-Mg-EPI microspheres in normal human tissue is very weak, generating almost no hydrogen gas and significantly reducing side effects on normal tissue. On the other hand, the pH of the tumor microenvironment is clearly weakly acidic (pH approximately 6.4), and at this pH value, GelMA-Mg-EPI microspheres react vigorously, generating a large amount of hydrogen gas, thus realizing tumor microenvironment-responsive hydrogen gas therapy. At the same time, GelMA microspheres and GelMA-EPI microspheres reacted slowly in an acidic buffer at pH 5.5, indicating that neither the empty gelatin protein microspheres nor the supported drugs possessed acid-responsive hydrogen release capabilities, and therefore no bubbles were generated.

[0105] Example 10.2 Acid-responsive drug release capacity test of polymer drug-carrying microspheres Referring to the method described in Example 9.1, methacrylate-oxidized gelatin protein microspheres (defined herein as GelMA-PAA microspheres) that do not support polyacrylic acid-modified active metal nanoparticles were prepared, with the following differences: Additionally, 5 mg of polyacrylic acid (PAA), 100 mg of GelMA, and 5 mg of LAP were weighed and dissolved in 1 mL of deionized water to prepare solution a, while simultaneously omitting the step of adding active metal nanoparticles to solution a to prepare solution b.

[0106] Referring to the method described in Example 9.7, high molecular weight drug-supported microspheres (defined herein as GelMA-PAA-Mg-EPI microspheres and GelMA-PAA-EPI microspheres) were manufactured using GelMA-PAA-Mg microspheres and GelMA-PAA-EPI microspheres as raw materials, with the amounts of drug (EPI) added during the manufacturing process being 2 mg / mL and 2 mL, respectively. The above microspheres were left to stand in ultrapure water, phosphate buffer (PBS), and pH 6.4 acidic buffer, respectively, and the supernatants were collected at different time points. The drug content was detected by ultraviolet spectrophotometer, and the amount of drug released was evaluated based on this, as shown in Figure 15. Activated metal nanoparticles react in an acidic buffer to generate hydrogen gas, thereby promoting the release of the supported drug. GelMA-PAA-Mg-EPI microspheres clearly released EPI in an acidic buffer at pH 6.4, demonstrating clear acid-responsive drug release. Drug release in the acidic buffer at pH 6.4 was significantly faster than that of GelMA-PAA-EPI microspheres, effectively realizing tumor microenvironment-responsive chemotherapy. At the same time, the drug release behavior of both in ultrapure water and phosphate buffer was similar, but both were very slow. This indicates that GelMA-PAA-EPI and GelMA-PAA-Mg-EPI release drugs extremely slowly at a neutral pH (7.4), thereby reducing toxicity and side effects to normal human tissue.

[0107] Example 11: Drug-carrying performance test of polymer drug-carrying microspheres Example 11.1: Drug loading rate test of polymer drug-carrying microspheres Polymer microspheres supporting a fixed solid phase volume of active metal nanoparticles are mixed with 2 mL of an aqueous solution containing epirubicin hydrochloride (1 mg / mL). The mixture is shaken in a shaker for 3-5 minutes while inverting it up and down. After completion, it is centrifuged again (100 r / min, 3 min), and the supernatant solution is collected. The mass of epirubicin hydrochloride remaining in it is detected. Subtracting the mass of epirubicin hydrochloride remaining in the supernatant solution after manufacturing from the known mass of epirubicin hydrochloride added gives the actual mass of epirubicin hydrochloride supported. The formula for calculating the drug loading rate is as follows. Drug loading rate = Mass of epirubicin hydrochloride actually loaded / Mass of epirubicin hydrochloride added × 100%.

[0108] The test results for the types of microspheres and their drug-carrying rates related to this embodiment are shown in Table 8. [Table 7] The test results in Table 8 show that the drug loading rate in Example 11.1.4 was clearly higher than in Example 11.1.3. The introduction of PAA significantly improved the loading efficiency of the microspheres for cationic drugs. Furthermore, polymer microspheres containing active metal nanoparticles showed a higher drug loading rate within the same drug loading time. This is because the introduction of active metal nanoparticles increased the specific surface area of ​​the polymer microspheres, resulting in a higher drug loading rate.

[0109] Example 11.2: Effect of polymer modification on drug loading rate of polymeric metal microspheres As described in Example 11.1, modification with anionic polymers, such as polyacrylic acid (PAA), can significantly improve the drug loading rate of polymer microspheres. This example further explores the effect of the amount of PAA added during the microsphere manufacturing process on the drug loading rate of microspheres, using PAA as an example. GelMA-PAA microspheres were manufactured with reference to the method described in Example 9.1, with the following differences: In addition, different masses of PAA, 100 mg of GelMA, and 5 mg of LAP were weighed and dissolved in 1 mL of deionized water to produce solution a. At the same time, the step of adding activated metal nanoparticles to solution a to produce solution b was omitted, and the amounts of PAA added were 5 mg, 10 mg, 20 mg, and 50 mg, respectively. After the manufacturing process was completed, the charge status of the microspheres of the product was measured using a zeta potential meter. A certain solid phase volume (0.5 mL) of microspheres was mixed with 2 mL of an aqueous solution containing epirubicin hydrochloride (1 mg / mL), shaken in a shaker while inverting it up and down for 3 to 5 minutes, and then centrifuged again (100 r / min, 3 min). The supernatant solution was collected, and the mass of epirubicin hydrochloride remaining therein was detected to calculate the drug load. The results of the zeta potential measurement and the drug load calculation are shown in Table 9. [Table 8]

[0110] As shown in the test results, the amount of PAA added during the microsphere manufacturing process has less effect on the drug-loading performance of the product. Since there is no significant change in the negative charge value or drug-loading rate with respect to the amount of PAA added, it can be reasonably inferred that good drug-loading effects can be achieved within the above-mentioned addition range.

[0111] Example 11.3: Effect of drug input amount on drug-carrying capacity of microspheres GelMA-PAA-Mg microspheres (0.5 ml) of the same solid phase volume were each mixed with 2 ml of epirubicin hydrochloride solution of different concentrations. The mixture was shaken in a shaker for 3-5 minutes while inverting it up and down, and then centrifuged again (100 r / min, 3 min). The supernatant solution was collected, and the amount of drug-carrying mass was calculated by detecting the mass of epirubicin hydrochloride remaining in it. The calculation results are shown in Table 10. [Table 9]

[0112] As shown in the drug-carrying amount detection results, as the concentration of epirubicin hydrochloride increases, the amount of drug carried by GelMA-PAA-Mg microspheres also increases accordingly. This explains that the polymer drug-carrying microspheres produced in this invention can handle drug-carrying conditions at higher concentrations and higher doses, and that their drug-carrying rate is stable, resulting in an increase in drug-carrying amount with increasing drug concentration. At the same time, this also explains that the polymer drug-carrying microspheres produced in this invention have a large drug-carrying potential per unit volume. In this embodiment, none of the drug-carrying amounts exceeded the carrying load of the polymer drug-carrying microspheres, which is advantageous in actual use for increasing the drug dose during embolization and enhancing the effect of chemotherapy. It should be noted that because the reaction time is relatively short and the drug-carrying of the microspheres has not reached saturation, the amount of drug-carrying also increases with increasing chemical content, and the amount of drug-carrying in saturated microspheres is even higher.

[0113] Example 11.4: Maximum drug-carrying capacity test of polymer drug-carrying microspheres The test results of Example 11.3 revealed that the drug loading of the polymer drug-carrying microspheres did not reach saturation at the EPI concentration. Furthermore, to test the maximum drug-carrying capacity of microspheres per unit solid phase volume, a constant solid phase volume (0.5 mL) of GelMA-PAA-Mg microspheres produced by the method described in Example 9.6 was mixed with 2 mL of high-concentration epirubicin hydrochloride. The mixture was shaken for 10 minutes while inverting it up and down in a shaker. After completion, centrifugation was performed again (100 r / min, 3 min), and the supernatant solution was collected. The mass of residual epirubicin hydrochloride was detected to evaluate the maximum drug-carrying capacity of microspheres per unit solid phase volume. The calculation formula is as follows. Maximum drug-carrying capacity of microspheres per unit solid phase volume = Maximum drug-carrying capacity / Volume of microspheres in solid phase

[0114] Using the method described above, the drug-carrying range of different microspheres was tested, and the test results are shown in Table 11. [Table 10] Tests revealed that when the total amount of epirubicin hydrochloride added exceeded 16 mg, the drug-carrying capacity of polymer microspheres supported with 0.5 mL of active metal nanoparticles gradually reached saturation. Calculations showed that the maximum drug-carrying capacity of the microspheres described in Example 11.4.2 was approximately 30 mg / mL, meaning the drug-carrying capacity range for GelMA-PAA-Mg microspheres was 0-30 mg / mL. This drug-carrying capacity was significantly improved compared to Example 11.4.1, further explaining the effect of polymer modification in increasing the charge of the microspheres. The drug-carrying capacity of the above polymer drug-carrying microspheres is high, meeting the needs of clinical use, and combined with the function of the active metal nanoparticles, a better therapeutic effect can be expected.

[0115] Example 12: Microspheres carrying anionic drugs Step 1) Preparation of 1-ethyl-(3-dimethylaminopropyl)carbodiimide-modified gelatin protein: Gelatin protein grafted with 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) is prepared according to the method described in Example 2, with the amount of crosslinking agent EDC used being 10% to 15% of the gelatin. Specifically, 2.5 g of gelatin protein is dissolved in 25 mL of DPBS solution, heated to 60°C to dissolve completely, then 0.3 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride is added, and the mixture is reacted in a 50°C water bath with stirring for 3 hours. After that, dialysis is performed in a 40°C water bath, and freeze-drying is performed to obtain 1-ethyl-(3-dimethylaminopropyl)carbodiimide-modified gelatin protein (GelEDC). Step 2) Preparation of GelEDC-Mg microspheres: 1-ethyl-(3-dimethylaminopropyl)carbodiimide-gelatin protein microspheres (GelEDC-Mg microspheres) supported with magnesium nanoparticles are prepared according to the method described in Example 9.1, wherein the concentration of methacrylate-oxidized gelatin protein in solution a is 10%.

[0116] Observation of the morphology and particle size of the product microspheres using an optical microscope reveals that when the average particle size of the supported magnesium nanoparticles is in the range of 20-150 μm, the average particle size of the resulting GelEDC-Mg microspheres is 10-500 μm, enabling further drug loading and local sustained release.

[0117] Gelatin is a molecule with the properties of an amphoteric polyelectrolyte; it is normally electrically neutral in neutral solutions, but often becomes positively charged in acidic solutions. By introducing a negatively charged functional group into gelatin protein through modification with methacrylic anhydride, the modified gelatin protein becomes negatively charged and can support cationic drugs through electrostatic adsorption (see Example 11). As described in this example, by introducing a positively charged functional group into gelatin protein through modification with 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, the modified gelatin protein becomes positively charged and can further support anionic drugs through electrostatic adsorption.

[0118] Furthermore, referring to the method described in Example 9.6, magnesium microparticle-supported 1-ethyl-(3-dimethylaminopropyl)carbodiimide-gelatin protein microspheres modified with anionic polymers were prepared, the difference being that an additional 5 mg of polyallylamine hydrochloride (anionic polymer, PAH), 100 mg of GelEDC, and 5 mg of LAP were weighed and dissolved in 1 mL of deionized water to prepare solution a, thereby obtaining PAH-modified magnesium microparticle-supported 1-ethyl-(3-dimethylaminopropyl)carbodiimide-gelatin protein microspheres (GelEDC-PAH-Mg microspheres).

[0119] Alendronate sodium is loaded onto the GelEDC-Mg microspheres and GelEDC-PAH-Mg microspheres produced in this example, and the drug load is tested.

[0120] The drug concentration in the supernatant is detected, and the maximum drug-carrying capacity of GelEDC-PAH-Mg microspheres per unit solid phase volume is calculated. The test results show that when the amount of alendronate sodium added during the reaction process exceeds 15 mg (0.5 mL of solid phase microspheres, 2 mL of ALN with a concentration of 7.5 mg / mL), drug loading in the GelEDC-PAH-Mg microspheres reaches saturation, with a maximum drug-carrying capacity of approximately 25 mg / mL. Therefore, the drug-carrying capacity range of GelEDC-PAH-Mg microspheres is 0-25 mg / mL. By adjusting parameters such as the polymer modification ratio and the ratio of polymers to metal microparticles, the drug-carrying capacity can be further increased. Flexible parameter adjustment allows for the production of products with various specifications, meeting clinical needs.

[0121] Example 13: Polyvinyl alcohol drug-loaded microspheres carrying epirubicin hydrochloride In this example, polymer microspheres supported with activated metal nanoparticles are manufactured. The polymer is methacrylic anhydride polyvinyl alcohol, and the activated metal nanoparticles are magnesium nanoparticles. Each polymer microsphere supporting the activated metal nanoparticles contains one or more magnesium nanoparticles. The average major diameter of the magnesium nanoparticles is 20 μm. The specific manufacturing process is as follows.

[0122] First, polyvinyl alcohol (PVA) is modified. Specifically, 2.5 g of PVA is dissolved in 25 mL of ultrapure water, heated to 90°C, and stirred for 2 hours to ensure complete dissolution. Then, 5 mL of methacrylic anhydride is added, the pH of the solution is adjusted to approximately 8, and the reaction is carried out at room temperature for 48 hours with stirring. Subsequently, precipitation is performed with ice-cold ethanol, the precipitate is collected, washed three times with ultrapure water, and freeze-dried to obtain methacrylic anhydride-modified polyvinyl alcohol (PVAMA).

[0123] Furthermore, using the method described in Example 10.1 as a reference, methacrylate-oxidized polyvinyl alcohol microspheres (PVAMA-Mg microspheres) supporting magnesium nanoparticles were produced, the only difference being that methacrylate-oxidized gelatin protein was replaced with an equal amount of methacrylate-oxidized polyvinyl alcohol.

[0124] The results of observing the particle size of the PVAMA-Mg microspheres produced as described above using an optical microscope are shown in Figure 16. The particle size of the produced PVAMA-Mg microspheres is approximately 65 μm, and on average, each microsphere contains 1 to 2 magnesium nanoparticles.

[0125] Furthermore, using the method described in Example 9.6 as a reference, PAA-modified methacrylate-oxidized polyvinyl alcohol microspheres (PVAMA-PAA-Mg microspheres) were produced, and their loading effect on cationic drugs (epirubicin hydrochloride) was also verified. From the test results, it was found that the drug loading rate was close to that of GelMA-PAA-Mg microspheres, and the drug loading range per unit solid phase volume was 0 to 28 mg / mL.

[0126] Example 14: Results of drug-loading tests of polymer drug-carrying microspheres Based on the methods described in Examples 12 and 13, polymer metal microspheres supported with active metal nanoparticles based on different polymer materials and different types of active metal nanoparticles were manufactured, and different drugs were loaded onto them by electrostatic adsorption depending on their charge state. Furthermore, the maximum drug load was improved through modification with a charged polymer. The types of polymer materials, solidifying agents, types of active metal nanoparticles, sizes of active metal nanoparticles, particle size of polymer microspheres supported with active metal nanoparticles, types of polymers, types of supported drugs, and drug loads related to this example are shown in Table 12. [Table 11]

[0127] The activity of metallic iron is relatively low compared to metals such as magnesium, aluminum, and zinc, and the rate at which it reacts in acidic solutions to produce hydrogen gas is relatively slow. However, as its size decreases, the reaction in acidic buffers becomes significantly faster, increasing the rate of hydrogen gas generation and drug release, thus enabling better hydrogen gas chemotherapy. At the same time, the reduced microsphere size increases the specific surface area, and its drug-carrying capacity also increases accordingly.

[0128] The above test results show that the drug load of the polymer drug-carrying microspheres described in this embodiment varies little and is not affected by factors such as the type of polymer material, the type of solidifying agent, the type of active metal nanoparticles, the size of the active metal nanoparticles, the particle size of the polymer microspheres carrying the active metal nanoparticles, the type of polymer, and the type of supported drug. This further explains the versatility of the polymer drug-carrying microspheres obtained by this manufacturing method, and demonstrates that good drug-carrying effects can be achieved for a wide variety of drugs.

[0129] Example 15: Cytotoxicity test of polymer drug-carrying microspheres In this example, the in vitro killing activity of polymer drug-carrying microspheres against tumor cells was verified. The relative EPI content per unit solid volume of microspheres was measured by the method described in Example 11.1, and the relative magnesium content per unit solid volume of microspheres was quantified by ICP-OES. Cells were seeded in a 96-well plate, and after the cells adhered, different microspheres were added to the cell medium and cultured with the cells for 24 hours. The killing effect of the microspheres on cells was then detected by the thiazole blue colorimetric method (MTT). The specific experimental groups are as follows.

[0130] Example 15.1: Blank control group, Example 15.2: GelMA-PAA microsphere group, Example 15.3: GelMA-PAA-Mg microsphere group, Mg added amount: 0.3 mg / well, Example 15.4: GelMA-PAA-EPI microsphere group, EPI added amount: 10 μg / well, Example 15.5: GelMA-PAA-Mg-EPI microsphere group, Mg added: 0.3 mg / well, EPI added: 10 μg / well, The killing effect was tested on three different tumor cell lines: mouse liver cancer cells (Hepa1-6), mouse mammary cancer cells (4T1), and mouse colon cancer cells (CT26) from Examples 15.1 to 15.5. The relative viability of the corresponding cells is shown in Figure 17. The results show that the cell viability of the culture group in Example 15.5 was significantly lower than that of the culture groups in Examples 15.3 and 15.4, and that the relative viability against different tumor cell lines was close. This explains that the magnesium nanoparticles and high molecular weight drug-carrying microspheres loaded with epirubicin exhibited higher antitumor activity and are applicable to various tumor types.

[0131] Example 16: Study of drug release and penetration of polymer drug-carrying microspheres in solid tumors We simulated the drug release and penetration of macromolecular drug-carrying microspheres in tumors using a 3D tumor cell model. The experimental method is as follows:

[0132] A 3D tumor cell model of the CT26 cell line was constructed according to standard procedures. The constructed CT26 tumor cells were cultured in cell medium containing different microspheres, and after 24 hours and 48 hours, the tumor cells were washed three times with phosphate-buffered water (PBS), fixed with 4% paraformaldehyde, and the cell nuclei were stained and positioned with 4',6-diamidino-2-phenylindole (DAPI). The fluorescence distribution within the tumor cells was then observed using a confocal microscope (Figure 18).

[0133] Epirubicin hydrochloride (EPI) is a fluorescent drug that, when excited by light at a wavelength of 488 nm, emits red light at a wavelength of approximately 594 nm. Confocal images showed that the red fluorescence intensity of epirubicin hydrochloride in tumor cells treated with GelMA-PAA-Mg-EPI was significantly higher than in the other two groups, and clear co-localization was observed between the fluorescence signal of the cell nucleus and the red fluorescence of epirubicin hydrochloride. This explains that the drug carried by GelMA-PAA-Mg-EPI effectively penetrated into the tumor cells and entered the tumor cells themselves. Furthermore, the fluorescence signal of epirubicin hydrochloride inside tumor cells was stronger with GelMA-PAA-Mg-EPI compared to GelMA-PAA-EPI, which explains that GelMA-PAA-Mg-EPI promotes the penetration of more drug molecules. GelMA-PAA-Mg-EPI can significantly improve drug penetration in tumor cells and drug uptake by tumor cells. It is expected that the drug-carrying microspheres described in this invention can improve the penetration of chemical drugs into tumor tissue at the patient's lesion site, promote the entry of chemical drugs into tumor cells, and enhance their effects.

[0134] Example 17: Study on the tumor-suppressing effect of polymer drug-carrying microspheres on H22 subcutaneous tumors Female Balb / c mice were purchased from Changzhou Cavens Laboratory Animal Co., Ltd. and reared according to the procedures approved by our school's Laboratory Animal Center. First, the tumor-suppressing effect of high molecular weight drug-carrying microspheres will be investigated in a mouse hepatocyte subcutaneous tumor model (H22 subcutaneous tumor). The experimental method is as follows.

[0135] A mouse liver cancer H22 subcutaneous tumor model was constructed by inoculating BALB / c mice with H22 tumor cells in the lower right dorsal region. The tumor volume was 300-400 mm². 3 Once the tumors have grown, the tumor-bearing mice will be randomly divided into four groups of seven mice each. Treatment will be administered by injecting microspheres into the tumors. This day will be designated as day 0, and the increase in tumor volume will be monitored every two days. After the tumor volume measurements are completed, the mice will be euthanized, and the tumor tissue will be collected. Necrosis and proliferation of the tumor sites will be observed using H&E immunohistochemical staining and Ki67 immunofluorescence staining. The specific experimental group divisions and treatment doses are as follows.

[0136] Example 17.1: GelMA-PAA microsphere group, Example 17.2: GelMA-PAA-Mg microsphere group, injection dose calculated by magnesium mass, 1.5 mg / animal. Example 17.3: GelMA-PAA-EPI microsphere group, injection dose calculated by EPI mass, 50 μg / animal. Example 17.4: GelMA-PAA-Mg-EPI microsphere group, injection dose: magnesium mass 1.5 mg / animal, EPI mass 50 μg / animal. Figure 19 shows the statistical results of tumor volume in tumor-bearing mice after drug administration. The tumor volume of the mice in group 17.4 was clearly smaller than that of the other groups, indicating that the therapeutic effect of GelMA-PAA-Mg-EPI microspheres administered by subcutaneous injection was the best and significantly superior to that of the other microspheres. At the same time, group 17.4 showed the most obvious cell necrosis in H&E immunohistochemical staining and the lowest tumor growth activity in Ki67 immunofluorescence staining. These results jointly demonstrate that high molecular weight drug-carrying microspheres containing active metal nanoparticles improve the tumor embolization therapeutic effect. Considering the results of multiple examples together, it is speculated that high molecular weight microspheres carrying active metal nanoparticles improve the acidic microenvironment at the tumor site, reduce drug resistance of tumor cells, and exert a synergistic effect with chemical drugs. Furthermore, the generation of hydrogen gas promotes the release and penetration of the carried chemotherapeutic drug, increasing the local concentration and utilization rate of chemical drug molecules and enhancing the antitumor effect.

[0137] Example 18: New Zealand rabbit auricular artery embolization experiment To test the arterial embolization effect of the polymer drug-carrying microspheres described in this invention, GelMA-PAA-Mg microspheres and GelMA-PAA microspheres were injected into the auricular arteries of anesthetized New Zealand rabbits, respectively. The rabbits' ears were photographed and recorded at different time points after injection. Figure 20 shows photographs of the auricular arteries of New Zealand rabbits before and after embolization. At 24 hours post-embolization, blood flow in the embolic area of ​​the rabbit ears of the GelMA-PAA-Mg microsphere embolization group and the GelMA-PAA microsphere embolization group had not recovered, and a decrease in blood supply was observed. This indicates that the embolization effect of the GelMA-PAA-Mg microsphere embolization group and the GelMA-PAA microsphere embolization group was good. The introduction of active metal nanoparticles contributed to the improvement of the embolization effect, and the duration of embolization was longer. In TACE treatment, the duration of embolization and the embolization effect have a significant impact on the treatment outcome. The more sufficient and longer the embolization, the more effectively nutrient transport to the tumor site is blocked, enhancing the tumor-killing effect, and thereby resulting in a better treatment effect. The above results indicate that the polymer drug-carrying microspheres described in the present invention have a good embolic effect on the auricular artery of rabbits. Clearly, the above embodiments are merely illustrative examples for clarity and do not limit the embodiments.

[0138] While the above disclosure discusses embodiments of the invention that are currently considered useful through various examples, these details are for illustrative purposes only, and it should be understood that the attached claims are not limited to the disclosed embodiments, but rather encompass all modifications and equivalent combinations that are substantially and in scope to the embodiments of the present application.

[0139] Similarly, note that in order to simplify the disclosure of this application and to aid in the understanding of embodiments of one or more inventions, the preceding descriptions of embodiments of this application may combine many features into a single embodiment, drawing, or description thereof. However, this method of disclosure does not mean that the subject matter of this application requires more features than those described in the claims. In fact, the features of an embodiment are fewer than all the features of a single embodiment described above.

[0140] In some embodiments, numerical values ​​are used to describe the quantities of components and attributes. It should be understood that in some cases, these numerical values ​​used in the description of such embodiments are modified by the modifiers “approximately,” “about,” or “roughly.” Unless otherwise specified, “approximately,” “about,” or “roughly” indicate that the numerical values ​​may vary by ±. Accordingly, in some embodiments, the numerical parameters used in the specification and claims are approximations, and these approximations may vary depending on the required features of each individual embodiment.

[0141] Finally, it should be understood that the embodiments described herein are merely illustrative of the principles of the embodiments herein. Other variations may also fall within the scope of this application. Therefore, alternative configurations of the embodiments herein, for illustrative purposes only and not as limitations, may be considered consistent with the teachings herein. Accordingly, the embodiments herein are not limited to those expressly introduced and described herein.

Claims

1. A polymer microsphere carrying active metal nanoparticles, characterized in that it comprises a polymer microsphere skeleton and active metal nanoparticles distributed inside the polymer microsphere skeleton.

2. The polymer microsphere according to claim 1, characterized in that the major axis of the activated metal nanoparticles is less than or equal to the particle size of the polymer microsphere.

3. The polymer microsphere according to claim 2, characterized in that the major axis of the activated metal nanoparticles is 1 nm to 100 μm.

4. The polymer microsphere according to claim 2, characterized in that the particle size of the activated metal nanoparticles is 10 μm to 500 μm.

5. The polymer microsphere according to claim 1, characterized in that the number of activated metal nanoparticles is one or more.

6. The polymer microsphere according to claim 1, characterized in that the activated metal nanoparticles are one or more of the activated metal element, alloy, and metal composite structure.

7. The polymer microsphere according to claim 6, characterized in that the active metal element is selected from one or more of magnesium, aluminum, zinc, iron, and tin.

8. The polymer microsphere according to claim 6, characterized in that the alloy contains at least two metals with different reactivity.

9. The polymer microsphere according to claim 6, characterized in that the metal composite structure contains at least two metals with different reactivity.

10. Step 1 involves preparing a polymer material into a polymer solution, A method for producing polymer microspheres supported by activated metal nanoparticles, characterized by comprising: step 2, mixing the polymer solution described in step 1 with activated metal nanoparticles and producing polymer microspheres supported by the activated metal nanoparticles by a method for producing microspheres, according to any one of claims 1 to 9.

11. The manufacturing method according to claim 10, characterized in that in step 1, the polymer material is selected from one or more of gelatin, polyvinyl alcohol, chitosan, biodegradable polyurethane, silk fibroin, collagen, alginic acid, and biodegradable starch.

12. The manufacturing method according to claim 10, characterized in that in step 2, the method for producing the microspheres includes a reversed-phase microemulsion method.

13. An embolizing agent characterized by comprising polymer microspheres supporting activated metal nanoparticles as described in any one of claims 1 to 9.

14. A polymer microsphere carrying activated metal nanoparticles, comprising a polymer microsphere and activated metal nanoparticles, wherein the activated metal nanoparticles are distributed within the polymer microsphere, the polymer microsphere carrying the activated metal nanoparticles has a core-shell structure with the activated metal nanoparticles as the core and the polymer microsphere as the shell, the polymer microsphere covering the outer surface of the activated metal nanoparticles, or the polymer microsphere carrying the activated metal nanoparticles has voids inside the polymer microsphere, and the activated metal nanoparticles are distributed within these voids.

15. The polymer microspheres supported by the activated metal nanoparticles according to claim 14, characterized in that the activated metal nanoparticles are one or more of the activated metal element, an alloy, and a metal composite structure.

16. The polymer microspheres supported by the activated metal nanoparticles according to claim 15, characterized in that the activated metal element is selected from one or more of magnesium, aluminum, zinc, iron, manganese, tin, gallium, and indium.

17. The active metal element is selected from one or more of magnesium, aluminum, zinc, iron, and manganese, and the major axis of the active metal nanoparticles is 1 nm to 150 μm, characterized in that the polymer microspheres supported by the active metal nanoparticles according to any one of claims 15 or 16.

18. The polymer microspheres supported by the activated metal nanoparticles according to claim 15 or 16, characterized in that the activated metal element is selected from one or more of iron, manganese, tin, gallium, and indium, and the major axis of the activated metal nanoparticles is 1 nm to 5000 nm.

19. The polymer microspheres supported by the activated metal nanoparticles according to any one of claims 14 to 18, characterized in that the particle size of the polymer microspheres supported by the activated metal nanoparticles is 10 μm to 500 μm.

20. The polymer microspheres supporting the activated metal nanoparticles are characterized in that they contain a functional group that is positively or negatively charged, as described in any one of claims 14 to 18.

21. A polymer drug-carrying microsphere, comprising a polymer microsphere carrying an active metal nanoparticle as described in any one of claims 1 to 9 or any one of claims 14 to 20, and a drug, wherein the drug is carried within the polymer microsphere carrying the active metal nanoparticle.

22. The polymer drug-supported microsphere according to claim 21, characterized in that the mass of the drug in the polymer microsphere supporting the active metal nanoparticles per unit solid phase volume is 0 to 30 mg / mL.

23. The method for producing the polymer material according to claim 10, characterized in that the polymer material contains a functional group that is positively or negatively charged.

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