An embolic microsphere capable of sustained drug release and controlled degradation rate, and its preparation method.

Abstract

This invention proposes an embolic microsphere capable of sustained drug release and controlled degradation rate, and its preparation method. The method includes: dissolving a dispersant in an oil phase liquid and stirring at a first preset temperature to obtain a dispersion; dissolving gelatin in water at a second preset temperature to obtain a gelatin solution; dissolving a compound with negatively charged groups in water, adding an alkali solution and an initiator under ice bath conditions to obtain a modified solution; adding the modified solution to the gelatin solution and stirring to obtain an aqueous prepolymer solution, which is then added dropwise to the dispersion for premixing; adding a crosslinking agent and a catalyst; and directly grafting the negatively charged groups onto the gelatin molecules via low-temperature free radical polymerization to obtain an intermediate product; and then performing post-treatment to obtain embolic microspheres. This invention eliminates the need for non-degradable materials, enabling the direct grafting of negatively charged groups onto the gelatin surface, resulting in embolic microspheres with highly efficient drug loading capacity, excellent long-term sustained drug release, and controlled degradation rate.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to a microsphere-type embolic agent, and more particularly to an embolic microsphere capable of sustained drug release and controlled degradation rate, and its preparation method. Background Technology

[0002] Currently, commonly used solid embolization agents include microsphere embolization agents and particulate embolization agents. Among them, embolization microspheres are a common clinical treatment for tumor diseases. The principle is to embolize blood vessels at the affected site with microspheres, cutting off the nutrient supply to tumor cells, causing them to die due to lack of nutrition, thereby achieving a therapeutic effect. Depending on the material, embolization microspheres can be divided into biodegradable and non-biodegradable types. Non-biodegradable microspheres accumulate in the body after embolization and produce toxicity, while biodegradable microspheres can automatically degrade into harmless products. Ideally, biodegradable embolization microspheres allow for controlled degradation time, enabling blood vessel recanalization after degradation and allowing for repeated embolization treatments. They gradually degrade under the action of relevant enzymes in the body, slowly releasing the drug.

[0003] Currently, the main materials of commercially available embolization microspheres are poly(methyl methacrylate), PMMA, polyvinyl alcohol (PVA), polyethylene glycol (PEG), or a combination of these three materials, all of which are non-degradable. While some products also use gelatin, the carriers that enable their drug delivery function are all non-degradable. For example, drug-loadable embolization microspheres are made by using gelatin, polyethylene glycol derivatives, and drug-loadable monomers as the aqueous phase materials and forming a microemulsion with an oil phase containing an emulsifier, followed by cross-linking with an aldehyde cross-linking agent and a photoinitiator. In this case, the polyethylene glycol derivative is grafted onto the gelatin, and the drug-loadable monomer is grafted onto the polyethylene glycol derivative. This results in limited drug loading and release capacity, and the polyethylene glycol derivative is non-degradable. This method requires grafting intermediate carriers for drug delivery, such as polyethylene glycol derivatives, onto gelatin. The process is complex, and these intermediate carriers are non-degradable, making repeated treatment impossible and posing a risk of inflammation or ectopic embolism. In addition, this method requires the use of specific photoinitiators, which are susceptible to light, have poor stability, and cause environmental pollution. The synthesis process is difficult and the production efficiency is low.

[0004] In addition, another method involves dissolving a mixture of gelatin and drugs in water, adding a cross-linking agent during stirring, and then spray drying to obtain embolization microspheres of different particle sizes. However, these embolization microspheres only achieve drug encapsulation through physical means, resulting in uneven particle size and, in particular, an inability to achieve sustained drug release, thus failing to achieve long-term treatment and failing to meet the needs of practical applications. Summary of the Invention

[0005] This invention proposes an embolic microsphere capable of sustained drug release and controlled degradation rate, and its preparation method. The embolic microsphere and its preparation method provided by this invention enable the direct grafting of negatively charged groups onto the surface of gelatin molecules via low-temperature free radical polymerization without the need for an intermediate carrier, thereby achieving drug loading. This results in embolic microspheres with highly efficient drug loading performance, enabling slow drug release and targeted drug delivery. Furthermore, it eliminates the need for non-degradable materials, achieving complete degradation. This allows for repeated treatment while avoiding inflammation and ectopic embolism. The obtained embolic microspheres exhibit excellent long-term sustained drug release and allow for controlled degradation rate control.

[0006] To address the aforementioned technical problems, one embodiment of the present invention provides a method for preparing embolic microspheres capable of sustained drug release and controlled degradation rate, comprising at least the following steps:

[0007] The dispersant is dissolved in the oil phase liquid and heated to a first preset temperature, and stirred to obtain a dispersion.

[0008] Gelatin is dissolved in water at a second preset temperature to obtain a gelatin solution;

[0009] A compound with a negatively charged group is dissolved in water, and an alkaline solution and an initiator are added under ice bath conditions to obtain a modified solution.

[0010] The modified liquid is added to the gelatin solution and stirred to obtain an aqueous prepolymer solution;

[0011] After the aqueous prepolymer is added dropwise to the dispersion for premixing, a crosslinking agent and a catalyst are added for crosslinking and catalysis. The negatively charged groups are directly grafted onto the gelatin molecules by low-temperature free radical polymerization to obtain an intermediate product.

[0012] The intermediate product is post-processed to obtain embolic microspheres.

[0013] In one embodiment of the present invention, the low-temperature free radical polymerization is a polymerization reaction that occurs at a temperature of 30°C to 40°C.

[0014] In one embodiment of the present invention, the compound with a negatively charged group includes at least one of a compound with a carboxylic acid group or a sulfonic acid group, wherein the carboxylic acid group is derived from acrylic acid, methacrylic acid or other carboxylic acid compounds or carboxylate compounds with a carboxyl group and a double bond, and the sulfonic acid group is derived from one or more of 2-acrylamido-2-methylpropanesulfonic acid or other sulfonic acid compounds or sulfonate compounds with a sulfonate group and a double bond.

[0015] In one embodiment of the present invention, the crosslinking agent includes at least one of acetal crosslinking agents, amide crosslinking agents, or iridoid crosslinking agents; the acetal crosslinking agent includes one or more combinations of formaldehyde, glutaraldehyde, or n-butyraldehyde; the amide crosslinking agent includes one or more combinations of N,N-methylenebisacrylamide, ethylene glycol dimethacrylate, or diethylenetriamine; and the iridoid crosslinking agent includes one or more combinations of genipin, oleuropein, or aucubin.

[0016] In one embodiment of the present invention, the catalyst comprises one or a combination of several of N,N-dimethylaniline, sodium bisulfite, tetraethyleneimine, tetramethylethylenediamine, thiol, ferrous chloride, or silver nitrate.

[0017] In one embodiment of the present invention, the dispersant includes one or more of Span 20, Span 80, Tween 20, Tween 80, PEG-40 or PEG-20, and the oil phase liquid includes one or more of liquid paraffin, glycerin, dimethyl silicone oil, white oil, vegetable oil or petroleum ether.

[0018] In one embodiment of the present invention, the alkaline solution includes at least one of sodium hydroxide, ammonia, or potassium hydroxide, and the initiator includes one or a combination of several of ammonium persulfate, potassium persulfate, hydrogen peroxide, diacyl peroxide, benzoyl peroxide, or azobisisobutyronitrile.

[0019] In one embodiment of the present invention, the mass ratio of the compound with the negatively charged group to the gelatin is 1:(2~4); and / or

[0020] The mass ratio of the catalyst to the gelatin is 1:(4~20); and / or

[0021] The mass ratio of the crosslinking agent to the gelatin is 1:(4~20).

[0022] In one embodiment of the present invention, the first preset temperature is 30°C to 40°C;

[0023] The second preset temperature is 38℃~45℃.

[0024] In one embodiment of the present invention, the manufacturing method further includes:

[0025] The aqueous prepolymer is added dropwise to the dispersion and sheared at 250 rpm to 350 rpm for 10 to 60 minutes.

[0026] The catalyst was added dropwise, and the reaction was continued for 150 to 600 minutes with stirring at 250 to 350 rpm; and

[0027] Then add the crosslinking agent and stir at 250 rpm to 350 rpm for 2 to 60 minutes to obtain the intermediate product.

[0028] In one embodiment of the present invention, the post-processing includes:

[0029] After obtaining the intermediate product, the intermediate product is filtered, and a solvent containing a surfactant is added for washing for a first preset time. Then, the product is rinsed with pure water and filtered, and then rinsed with water for injection to collect the intermediate phase of the embolization microspheres.

[0030] The embolization microspheres are pre-frozen in the intermediate phase and then freeze-dried for a second preset time to obtain the embolization microspheres.

[0031] In one embodiment of the present invention, the first preset time is 10 minutes to 120 minutes; the second preset time is 60 hours to 260 hours.

[0032] In one embodiment of the present invention, the surfactant includes Tween 80.

[0033] An embodiment of the present invention also provides an embolic microsphere capable of sustained drug release and controlled degradation rate, obtained by the preparation method described above. The embolic microsphere is obtained by polymerizing gelatin molecules and a compound containing negatively charged groups and cross-linking them with a cross-linking agent; wherein: negatively charged groups are directly grafted onto the surface of the gelatin molecules.

[0034] In one embodiment of the present invention, the negatively charged group includes at least one of a sulfonic acid group or a carboxylic acid group; the crosslinking agent includes at least one of an acetal crosslinking agent, an amide crosslinking agent, or a cycloalkenyl ether crosslinking agent.

[0035] In summary, the embolic microspheres with sustained-release drug delivery and controlled degradation rate proposed in this invention, along with their preparation method, enable the direct grafting of negatively charged groups onto the surface of gelatin molecules via low-temperature free radical polymerization. This eliminates the need for conventional drug-loading intermediates, especially non-degradable ones, resulting in fully degradable embolic microspheres with excellent biocompatibility. After degradation, the blood vessels can be reopened, allowing for repeated embolization treatments without inducing inflammatory reactions or ectopic embolization. Furthermore, the embolic microspheres obtained through this method exhibit highly efficient drug loading capacity and can achieve slow drug release, enabling targeted drug delivery while simultaneously achieving complete degradation. By controlling the type of cross-linking agent, the embolic microspheres achieve excellent long-term drug release, and the degradation rate can be controlled. The degradation time can be tailored to specific needs, and functional monomers with different cross-linking strengths can be designed and synthesized to meet the practical application requirements of different lesion scenarios, providing patients with more treatment options. Furthermore, according to the technical solution of the present invention, the number of grafted negatively charged groups formed on the embolization microspheres can be controlled, thereby allowing the drug loading to be controlled according to the needs of the condition. Moreover, the preparation process is simple, the manufacturing process is safe and easy to control, and the cost is reduced. Attached Figure Description

[0036] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0037] Figure 1 This is a diagram illustrating the drug delivery mechanism of embolization microspheres in one embodiment of the present invention.

[0038] Figure 2 This is a diagram illustrating the drug delivery mechanism of the embolization microspheres in Example 1.

[0039] Figure 3 This is a diagram illustrating the drug delivery mechanism of the embolization microspheres in Example 2.

[0040] Figure 4 This is a diagram illustrating the drug delivery mechanism of the embolization microspheres in Examples 3-5.

[0041] Figure 5 This is a microscope image of embolic microspheres prepared using the method of the present invention in one embodiment of the present invention.

[0042] Figure 6 This is a microscope image of embolized microspheres after drug loading in one embodiment of the present invention. Detailed Implementation

[0043] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0044] It should be understood that the invention can be embodied in various forms and should not be construed as being limited to the embodiments set forth herein. Rather, providing these embodiments will make the disclosure thorough and complete, and will fully convey the scope of the invention to those skilled in the art.

[0045] The technical solution of the present invention will be further described in detail below with reference to the embodiments. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0046] This invention proposes an embolic microsphere capable of sustained drug release and controlled degradation rate. The embolic microsphere is obtained by polymerizing gelatin molecules, compounds containing negatively charged groups, and cross-linking them with a cross-linking agent. The gelatin molecules are grafted with negatively charged groups, including at least one of sulfonic acid groups or carboxylic acid groups. The cross-linking agent includes at least one of acetal cross-linking agents, amide cross-linking agents, or iridoid cross-linking agents. By selecting gelatin as the main material, completely degradable embolic microspheres can be obtained, which are harmless to the human body after degradation. By grafting negatively charged groups onto the surface of gelatin molecules, the embolic microspheres possess highly efficient drug loading capacity. By controlling the type of cross-linking agent, the embolic microspheres exhibit good long-term sustained drug release effects, and the degradation rate of the embolic microspheres can be controlled. Embolistic microspheres can be customized to meet different application requirements.

[0047] In one embodiment of the present invention, the acetal crosslinking agent includes one or more combinations of formaldehyde, glutaraldehyde, or n-butyraldehyde; the amide crosslinking agent includes one or more combinations of N,N-methylenebisacrylamide, ethylene glycol dimethacrylate, or diethylenetriamine; and the iridoid crosslinking agent includes one or more combinations of genipin, oleuropein, or aucubin. In another embodiment of the present invention, gelatin can also be replaced with natural polymer materials such as polylactic acid, chitosan, alginate, or soluble starch to ensure that the obtained embolic microspheres can be degraded.

[0048] This invention provides a method for preparing embolic microspheres capable of sustained drug release and controlled degradation rate. The method comprises at least the following steps: dissolving a dispersant in an oil phase liquid and heating it to a first preset temperature, then stirring to obtain a dispersion; dissolving gelatin in water at a second preset temperature to obtain a gelatin solution; dissolving a compound with negatively charged groups in water, and adding an alkali solution and an initiator under ice bath conditions to obtain a modified solution; adding the modified solution to the gelatin solution and stirring to obtain an aqueous prepolymer solution; adding the aqueous prepolymer solution dropwise to the dispersion, premixing, then adding a crosslinking agent and a catalyst for crosslinking and catalysis, directly grafting the negatively charged groups onto the gelatin molecules via low-temperature free radical polymerization to obtain an intermediate product; and post-processing the intermediate product to obtain embolic microspheres.

[0049] In one embodiment of the present invention, the dispersant includes, for example, one or a combination of several dispersants such as Span 20, Span 80, Tween 20, Tween 80, PEG-40, or PEG-20, and the oil phase liquid includes, for example, one or a combination of several materials such as liquid paraffin, glycerin, dimethyl silicone oil, white oil, vegetable oil, or petroleum ether. In one embodiment of the present invention, for example, 0.1g to 10g of the dispersant is dispersed in 1000ml of the oil phase liquid, and the process is carried out, for example, in a reaction vessel or a stirred tank, and the first preset temperature is, for example, 30°C to 40°C, and the mixture is continuously stirred, for example, at 200rpm / min to 250rpm / min, to improve the uniformity of the obtained dispersion.

[0050] In one embodiment of the present invention, the second preset temperature is, for example, 38°C to 45°C. The solvent water is, for example, purified water or water for injection, and the mass ratio of gelatin to water is, for example, (1~5):10.

[0051] In one embodiment of the present invention, the compound with a negatively charged group is, for example, at least one of compounds including a carboxylic acid group or a sulfonic acid group. The carboxylic acid group is derived from a carboxylic acid compound or carboxylate compound containing a carboxyl group and a double bond, such as acrylic acid or methacrylic acid. The sulfonic acid group is derived from one or more combinations of sulfonic acid compounds or sulfonate compounds containing a sulfonate group and a double bond, such as 2-acrylamido-2-methylpropanesulfonic acid. The solvent water is selected, for example, purified water or water for injection. The mass ratio of the compound with the negatively charged group to water is, for example, (0.1~30):20. After the compound with the negatively charged group is dissolved, an alkaline solution and an initiator are added under ice bath conditions to obtain a modified solution. The alkaline solution includes, for example, at least one of sodium hydroxide, ammonia, or potassium hydroxide. The initiator includes one or more combinations of ammonium persulfate, potassium persulfate, hydrogen peroxide, diacyl peroxide, benzoyl peroxide, or azobisisobutyronitrile. In one embodiment of the present invention, for example, 0.1 ml to 10 ml of 7.5 mol / L sodium hydroxide solution is added under an ice bath, followed by 0.01 g to 5 g of potassium persulfate. Using an ice bath releases the heat generated by the reaction, improving the safety of the synthesis and ensuring the performance of the modified solution.

[0052] In one embodiment of the present invention, the modified liquid is added to a gelatin solution, stirred until homogeneous, and bubble-removing is performed, for example, by ultrasonication, to obtain an aqueous prepolymer. The mass ratio of the compound with negatively charged groups to gelatin is, for example, 1:(2~4). After obtaining the aqueous prepolymer, it is added dropwise to a dispersion for premixing. Premixing is performed, for example, by shearing at 250 rpm / min~350 rpm / min for 10 min~60 min to ensure homogeneous mixing of the aqueous prepolymer and dispersion. A crosslinking agent and a catalyst are then added for crosslinking and catalysis. The negatively charged group is grafted onto the gelatin molecules via low-temperature free radical polymerization to obtain an intermediate product. The volume ratio of the aqueous prepolymer to the dispersion is, for example, (10~15):100. The crosslinking agent and catalyst can be added simultaneously or intermittently. For example, after adding the crosslinking agent and catalyst simultaneously, the reaction continues for 150 min~600 min to obtain the intermediate product. In another embodiment of the present invention, for example, after adding the catalyst dropwise, the mixture is stirred at 250 rpm / min to 350 rpm / min and reacted for 150 min to 600 min, then a crosslinking agent is added, and the mixture is stirred at 250 rpm / min to 350 rpm / min for 2 min to 60 min to obtain an intermediate product. The particle size of the drug-loaded microspheres can be controlled by controlling the stirring speed.

[0053] In one embodiment of the present invention, the mass ratio of crosslinking agent to gelatin is, for example, 1:(4~20), and the mass ratio of catalyst to gelatin is, for example, 1:(4~20). Low-temperature free radical polymerization is a polymerization reaction that occurs at a temperature of 30°C to 40°C. In this embodiment, the temperature during low-temperature free radical polymerization is controlled by controlling the first temperature, the second temperature, and ice bath treatment. In other embodiments, the temperature during low-temperature free radical polymerization is controlled, for example, by using a water bath or temperature control equipment. In the present invention, the initiator can lower the activation energy of the reaction between the compound with negatively charged groups and gelatin, stimulating free radical polymerization between the two. This allows the reaction to occur at a lower temperature. That is, the low-temperature free radical polymerization system allows negatively charged groups to be directly grafted onto gelatin molecules, crosslinked into microspheres by the crosslinking agent, and catalyzed by the catalyst to obtain intermediate products. By controlling the polymerization temperature, the problem of poor stability of gelatin at high temperatures and easy gelation at low temperatures is solved, thus achieving the modification of gelatin. This invention can directly graft negatively charged groups onto gelatin molecules, thereby improving the drug loading and release capacity of embolization microspheres, while avoiding the use of non-degradable materials such as polyethylene glycol, and achieving complete degradation of embolization microspheres.

[0054] In one embodiment of the present invention, the catalyst includes, for example, one or more combinations of N,N-dimethylaniline, sodium bisulfite, tetraethyleneimine, tetramethylethylenediamine, thiols, ferrous chloride, or silver nitrate; the crosslinking agent includes at least one of acetal crosslinking agents, amide crosslinking agents, or iridoid crosslinking agents; the acetal crosslinking agent includes one or more combinations of formaldehyde, glutaraldehyde, or n-butyraldehyde; the amide crosslinking agent includes one or more combinations of N,N-methylenebisacrylamide, ethylene glycol dimethacrylate, or diethylenetriamine; and the iridoid crosslinking agent includes one or more combinations of genipin, oleuropein, or aucubin. When an amide crosslinking agent is selected, the amide crosslinking agent and gelatin can undergo a crosslinking reaction under the action of the catalyst. By controlling the mass ratio of compounds with negatively charged groups to gelatin, the number of negatively charged groups grafted onto the formed embolic microspheres can be controlled. This allows for control of drug loading according to the patient's condition. Furthermore, by controlling the type and ratio of cross-linking agents, the drug release rate and degradation rate of the embolic microspheres can be controlled, enabling the preparation of customizable embolic microspheres to meet different application needs.

[0055] In one embodiment of the present invention, after obtaining the intermediate product, the post-processing includes: filtering the intermediate product, adding a solvent containing a surfactant for washing for a first preset time, rinsing with pure water and filtering, then rinsing with water for injection, and collecting the embolization microsphere intermediate phase; pre-freezing the embolization microsphere intermediate phase, and then freeze-drying for a second preset time to obtain the embolization microspheres. Specifically, for example, the solvent and oil phase liquid are removed by filtration through a sieve, and the filtered solid is placed in a beaker or other container, a solvent containing a surfactant is added for washing for a first preset time, an appropriate amount of purified water is used for rinsing, and the solvent is removed again by filtration through a sieve, followed by rinsing with water for injection, and collecting the embolization microsphere intermediate phase. The solvent containing the surfactant includes, for example, Tween 80 solution, and stirring is performed during the washing process to improve washing efficiency and effectively remove any residual oil phase liquid and residual catalyst that may remain on the surface of the intermediate product. The first preset time is, for example, 10 min to 120 min.

[0056] In one embodiment of the present invention, after collecting the mesophase of the embolic microspheres, the mesophase is transferred to a freeze-drying box for pre-freezing. After pre-freezing, it is freeze-dried for a second preset time to obtain the embolic microspheres. The pre-freezing temperature is, for example, -20°C, the freeze-drying temperature is, for example, -40°C, and the second preset time is, for example, 60h to 260h. Freeze-drying ensures the stability of the embolic microspheres, protects their activity, and yields high-quality embolic microspheres.

[0057] The present invention will be explained in more detail below by referring to embodiments, which should not be construed as limiting. Appropriate modifications can be made within the scope of the present invention, and all such modifications fall within the technical scope of the present invention.

[0058] Example 1

[0059] Dissolve 1g of Span 80 in 1000ml of liquid paraffin, add it to a 2L reactor, preheat at 30℃, and stir at 230r / min to obtain a dispersion.

[0060] Add 40g of gelatin to 100ml of purified water and stir at 40℃ to dissolve, thus obtaining a gelatin solution.

[0061] Dissolve 20g of 2-acrylamido-2-methylpropanesulfonic acid in 20ml of purified water, add 10ml of 7.5mol / L sodium hydroxide solution under an ice-water bath, and then add 5g of potassium persulfate and stir to dissolve, thus obtaining the modified solution.

[0062] The modified liquid was added to the gelatin solution, stirred until homogeneous, and ultrasonically removed to remove air bubbles, thus obtaining an aqueous prepolymer solution.

[0063] The aqueous prepolymer was added dropwise to the dispersion at 30°C using a peristaltic pump. After shearing at 300 r / min for 50 min, 5 g of N,N-methylenebisacrylamide was added, followed by 10 g of tetramethylethylenediamine. The reaction was continued for 300 min to obtain the intermediate product.

[0064] The intermediate product was filtered through a sieve to remove the oil phase liquid and solvent. The solid was placed in a beaker, and an appropriate amount of Tween 80 solution was added and stirred for 60 min. The mixture was then rinsed with purified water and filtered through a sieve. The microspheres were then rinsed with an appropriate amount of water for injection and sieved again. The microspheres in the sieve were collected to obtain the embolic microsphere intermediate phase. The embolic microsphere intermediate phase was transferred to a lyophilization chamber and pre-frozen at -20℃. After pre-freezing, it was freeze-dried at -40℃ for 200 h to obtain the embolic microspheres.

[0065] Example 2

[0066] Dissolve 1g of Span 80 in 1000ml of liquid paraffin, add it to a 2L reactor, preheat at 35℃, and stir at 230r / min to obtain a dispersion.

[0067] Add 40g of gelatin to 100ml of purified water and stir at 40℃ to dissolve, thus obtaining a gelatin solution.

[0068] Dissolve 20g of acrylic acid in 20ml of purified water, add 10ml of 7.5mol / L sodium hydroxide solution under an ice-water bath, and then add 5g of potassium persulfate and stir to dissolve, thus obtaining the modified solution.

[0069] The modified liquid was added to the gelatin solution, stirred until homogeneous, and ultrasonically removed to remove air bubbles, thus obtaining an aqueous prepolymer solution.

[0070] The above aqueous prepolymer was added dropwise to the above dispersion at 35°C using a peristaltic pump. After shearing at 300 r / min for 50 min, 5 g of N,N-methylenebisacrylamide was added, followed by 10 g of tetramethylethylenediamine. The reaction was continued for 300 min to obtain the intermediate product.

[0071] The intermediate product was filtered through a sieve to remove the oil phase and solvent. The solid was added to a beaker, and an appropriate amount of Tween 80 solution was added and stirred for 60 min. The mixture was then rinsed with purified water and filtered through a sieve. The microspheres were then rinsed with an appropriate amount of water for injection and sieved again. The microspheres in the sieve were collected to obtain the embolic microsphere intermediate phase. The embolic microsphere intermediate phase was transferred to a lyophilization chamber and pre-frozen at -20℃. After pre-freezing, it was freeze-dried at -40℃ for 200 h to obtain the embolic microspheres.

[0072] Example 3

[0073] Dissolve 1g of Span 80 in 1000ml of liquid paraffin, add it to a 2L reactor, preheat at 40℃, and stir at 230r / min to obtain a dispersion.

[0074] Add 40g of gelatin to 100ml of purified water and stir at 40℃ to dissolve, thus obtaining a gelatin solution.

[0075] Dissolve 10g of 2-acrylamido-2-methylpropanesulfonic acid and 10g of acrylic acid in 20ml of purified water. Add 10ml of 7.5mol / L sodium hydroxide solution under an ice-water bath, and then add 5g of potassium persulfate and stir to dissolve, thus obtaining the modified solution.

[0076] The modified liquid was added to the gelatin solution, stirred until homogeneous, and ultrasonically removed to remove air bubbles, thus obtaining an aqueous prepolymer solution.

[0077] The above aqueous prepolymer was added dropwise to the above dispersion at 40°C using a peristaltic pump. After shearing at 300 r / min for 50 min, 5 g of N,N-methylenebisacrylamide was added, followed by 10 g of tetramethylethylenediamine. The reaction was continued for 300 min to obtain the intermediate product.

[0078] The intermediate product was filtered through a sieve to remove the oil phase liquid and solvent. The solid was added to a beaker, and an appropriate amount of Tween 80 solution was added and stirred for 60 min. The mixture was then rinsed with purified water and filtered through a sieve. The microspheres were then rinsed with an appropriate amount of water for injection and sieved again. The microspheres in the sieve were collected to obtain the embolic microsphere intermediate phase. The embolic microsphere intermediate phase was transferred to a lyophilization chamber and pre-frozen at -20℃. After pre-freezing, it was freeze-dried at -40℃ for 200 h to obtain the embolic microspheres.

[0079] Example 4

[0080] Dissolve 1g of Span 80 in 1000ml of liquid paraffin, add it to a 2L reactor, preheat at 40℃, and stir at 230r / min to obtain a dispersion.

[0081] Add 40g of gelatin to 100ml of purified water and stir at 40℃ to dissolve, thus obtaining a gelatin solution.

[0082] Dissolve 10g of 2-acrylamido-2-methylpropanesulfonic acid and 10g of acrylic acid in 10ml of purified water. Add 10ml of 7.5mol / L sodium hydroxide solution under an ice-water bath, and then add 5g of potassium persulfate and stir to dissolve, thus obtaining the modified solution.

[0083] The modified liquid was added to the gelatin solution, stirred until homogeneous, and ultrasonically removed to remove air bubbles, thus obtaining an aqueous prepolymer solution.

[0084] The above aqueous prepolymer was added dropwise to the above dispersion at 40°C using a peristaltic pump. After shearing at 300 r / min for 50 min, 10 g of tetramethylethylenediamine was added, and the reaction was continued for 300 min. Then, an aqueous formaldehyde solution containing 5 g of formaldehyde was added, and the mixture was stirred for 30 min.

[0085] The intermediate product was filtered through a sieve to remove the oil phase liquid and solvent. The solid was added to a beaker, and an appropriate amount of Tween 80 solution was added and stirred for 60 min. The mixture was then rinsed with purified water and filtered through a sieve. The microspheres were then rinsed with an appropriate amount of water for injection and sieved again. The microspheres in the sieve were collected to obtain the embolic microsphere intermediate phase. The embolic microsphere intermediate phase was transferred to a lyophilization chamber and pre-frozen at -20℃. After pre-freezing, it was freeze-dried at -40℃ for 200 h to obtain the embolic microspheres.

[0086] Example 5

[0087] Dissolve 1g of Span 80 in 1000ml of liquid paraffin, add it to a 2L reactor, preheat at 40℃, and stir at 230r / min to obtain a dispersion.

[0088] Add 40g of gelatin to 100ml of purified water and stir at 40℃ to dissolve, thus obtaining a gelatin solution.

[0089] Dissolve 10g of 2-acrylamido-2-methylpropanesulfonic acid and 10g of acrylic acid in 20ml of purified water. Add 10ml of 7.5mol / L sodium hydroxide solution under an ice-water bath, and then add 5g of potassium persulfate and stir to dissolve, thus obtaining the modified solution.

[0090] The modified liquid was added to the gelatin solution, stirred until homogeneous, and ultrasonically removed to remove air bubbles, thus obtaining an aqueous prepolymer solution.

[0091] The above aqueous prepolymer was added dropwise to the above dispersion at 40°C using a peristaltic pump. After shearing at 300 r / min for 50 min, 10 g of tetramethylethylenediamine was added, and the reaction was continued for 300 min. Then, an aqueous solution of genipin acetic acid containing 5 g of genipin acetic acid was added, and the mixture was stirred for 30 min.

[0092] The intermediate product was filtered through a sieve to remove the oil phase liquid and solvent. The solid was added to a beaker, and an appropriate amount of Tween 80 solution was added and stirred for 60 min. The mixture was then rinsed with purified water and filtered through a sieve. The microspheres were then rinsed with an appropriate amount of water for injection and sieved again. The microspheres in the sieve were collected to obtain the embolic microsphere intermediate phase. The embolic microsphere intermediate phase was transferred to a lyophilization chamber and pre-frozen at -20℃. After pre-freezing, it was freeze-dried at -40℃ for 200 h to obtain the embolic microspheres.

[0093] Comparative Example 1

[0094] Comparative Example 1 uses embolic microspheres manufactured by Whitecon UK Ltd., model number: DC2V103; specification: 100-300μm.

[0095] In this invention, in Examples 1-5 and Comparative Example 1, the drug loading and release properties and degradation properties of the embolization microspheres were tested, and the test results are shown in Tables 1 and 2.

[0096] In one embodiment of the present invention, the drug loading and release test is conducted, for example, using the cationic drug doxorubicin. The drug loading steps include: measuring 100 mg of embolic microspheres into each of three 10 ml amber vials using a 2 ml disposable syringe; transferring 3 ml of 15 mg / ml Doxorubicin solution into three amber vials using a pipette, sealing with rubber stoppers, shaking and inverting, and starting timing; shaking and inverting the vials every 5 minutes for the first 30 minutes, and then shaking and inverting the vials intermittently thereafter; at 0.5 h, taking 20 μl of the supernatant from each vial, placing it in a clean amber vial, adding 4.98 ml of purified water, mixing well, and testing its ultraviolet absorbance. During the drug loading process, a small amount of hydrochloric acid (HCl) can also be added, such as 20 μl of 37% hydrochloric acid, to adjust the pH value during the drug loading process, thereby improving the solubility, stability, and pharmacological activity of the drug.

[0097] In one embodiment of the present invention, the drug release step includes: assembling a top-mounted stirrer, a polytetrafluoroethylene (PTFE) stirring paddle, a 1000ml three-necked flask, and a heat-collecting constant-temperature magnetic stirrer, and adding an appropriate amount of purified water; covering the exposed parts of the three-necked flask with tin foil at a water bath temperature of 37°C, while preheating the prepared 0.15M physiological saline solution in a 37°C water bath; filtering the drug-loaded embolization microspheres through filter paper to remove unloaded Doxorubicin solution, transferring all the embolization microspheres to the three-necked flask, adding 900ml of preheated 0.15M physiological saline solution, and starting the timing; taking 2.5ml of dissolution medium from the three-necked flask at 0min, 5min, 10min, 0.5h, 1.5h, 3.0h, 24h, and 48h respectively, and adding 2.5ml of 0.15M physiological saline solution to the flask. The absorbance of Doxorubicin solutions with concentrations of 100.0 μg / ml, 10.00 μg / ml, and 1.000 μg / ml was measured at a wavelength of 480 nm to create a standard curve of drug concentration versus absorbance.

[0098] In one embodiment of the present invention, the degradation test includes: weighing 100 mg of embolic microspheres, placing them in a constant weight bottle and weighing them, then drying them in an oven at 105°C for 2 hours, and weighing the total weight after constant weight; taking 100 mg of the dried sample and placing it in a test tube, adding 10 ml of buffer solution (phosphate solution with a pH of 7.4), sealing the test tube with aluminum foil, and marking it with the corresponding sample batch number. Two sets are performed for each sampling point. The samples are placed in a water bath at 37±1°C, and the samples are taken out at 3, 7, 14, 21, 30, and 60 days respectively for constant weight testing, and the rate of change in the weight of the embolic microspheres corresponding to the degradation time is recorded.

[0099] Table 1. Drug loading and release rates of embolic microspheres in Examples 1-5 and Comparative Example 1

[0100]

[0101] Table 2 shows the degradation rates of the embolic microspheres in Examples 1-5 and Comparative Example 1.

[0102]

[0103] Please see Figure 1 As shown, in one embodiment of the present invention, multiple negatively charged groups 20 are grafted onto the surface of the embolization microspheres 10. The negatively charged groups 20 can be of the same type or a combination of multiple negatively charged groups. The number of negatively charged groups 20 on the surface of the embolization microspheres 10 can be controlled according to the proportion of raw materials used in the manufacturing process. During drug loading, the drug 30 is carried on the embolization microspheres 10 via the negatively charged groups 20. During use, after reaching a specific location, it gradually degrades under the action of relevant enzymes in vivo, slowly releasing the drug 30, embolizing the microspheres 10, and causing its degradation. Please refer to [link to relevant documentation]. Figures 2 to 4 The diagrams shown are the mechanism diagrams for drug loading of the embolic microspheres in Examples 1-5, respectively. Figure 4 The diagrams illustrate the drug loading mechanisms in Examples 3-5. As can be seen from the figures, when the surface of the embolic microspheres is grafted with the same type of negatively charged group, or a combination of multiple negatively charged groups, the resulting embolic microspheres can all be loaded with drugs. The embolic microspheres obtained by this invention possess controllable degradability and high-efficiency drug loading capacity, exhibit good biocompatibility, and can reduce post-interventional treatment side effects in patients, providing more options for interventional therapy.

[0104] Please see Figures 5 to 6 As shown, in one embodiment of the present invention, the obtained embolization microspheres have good sphericity, smooth surface, and uniform particle size. After drug loading, the drug can be uniformly distributed on the embolization microspheres, indicating that the negatively charged groups are uniformly distributed on the embolization microspheres, thus achieving uniform drug loading.

[0105] Please refer to Table 1. Comparing Examples 1-5 and Comparative Example 1, it can be seen that the embolic microspheres obtained through the technical solution of this application can improve the drug loading capacity of the embolic microspheres and achieve sustained drug release. Comparing Examples 1-3, it can be seen that when the negatively charged group is a sulfonic acid group, the drug loading capacity increases relative to the carboxylic acid group, and the drug release rate is slower. When the embolic microspheres are simultaneously grafted with sulfonic acid groups and carboxylic acid groups, the drug loading capacity further increases, and the drug release rate further decreases. Comparing Examples 3 and 5, it can be seen that when different crosslinking agents are used to obtain embolic microspheres, and the same amount of sulfonic acid groups and carboxylic acid groups are simultaneously grafted onto the gelatin in the embolic microspheres, the drug loading capacity and drug release rate are similar, indicating that the type of crosslinking agent has a weak effect on the drug loading capacity, and the drug loading capacity is mainly controlled by the different types and proportions of negatively charged groups. Therefore, controlling the type and proportion of negatively charged groups on embolic microspheres can control drug loading and release rate, thereby achieving different drug loading capacities. Simultaneously, it can achieve slow drug release, resulting in targeted drug delivery. This allows for the preparation of customizable embolic microspheres to meet the needs of different drug loading and release rates. Furthermore, the obtained embolic microspheres exhibit good biocompatibility; after degradation, blood vessels can be recanalized, allowing for repeated embolization treatments without inducing inflammatory responses, providing patients with more treatment options.

[0106] Please refer to Table 2. Comparing Examples 1-5 and Comparative Example 1, it can be seen that when the same crosslinking agent is selected, the degradation rate of the embolic microspheres is similar; when different crosslinking agents are selected, the degradation rate of the embolic microspheres is different. Therefore, by selecting different crosslinking agents, the degradation rate of the embolic microspheres can be controlled, thereby allowing for desired control of their degradation time. Functional monomers with different crosslinking strengths can be designed and synthesized according to actual needs to meet the practical application requirements of different lesion scenarios.

[0107] In summary, this invention proposes an embolic microsphere capable of sustained drug release and controlled degradation rate, along with its preparation method. By selecting gelatin as the drug-carrying substrate and employing a low-temperature free radical polymerization system, negatively charged groups can be directly grafted onto the surface of gelatin molecules, eliminating the need for conventional drug-carrying intermediates, especially non-degradable ones. This results in fully degradable embolic microspheres with excellent biocompatibility. After degradation, the blood vessels can be reopened, allowing for repeated embolization treatments without inducing inflammatory reactions or ectopic embolization. Furthermore, the embolic microspheres obtained through this method exhibit highly efficient drug loading capacity and can achieve slow drug release, enabling targeted drug delivery while simultaneously achieving complete degradation. By controlling the type of cross-linking agent, the embolic microspheres achieve excellent long-term sustained drug release, and the degradation rate can be controlled. The degradation time can be tailored to specific needs, and functional monomers with different cross-linking strengths can be designed and synthesized to meet the practical application requirements of different lesion scenarios, providing patients with more treatment options. Furthermore, according to the technical solution of the present invention, the number of grafted negatively charged groups formed on the embolization microspheres can be controlled, thereby allowing the drug loading to be controlled according to the needs of the condition. Moreover, the preparation process is simple, the manufacturing process is safe and easy to control, and the cost is reduced.

[0108] The above description is merely a preferred embodiment of this application and an explanation of the technical principles used. Those skilled in the art should understand that the scope of the invention involved in this application is not limited to the technical solutions formed by a specific combination of the above-mentioned technical features, but should also cover other technical solutions formed by any combination of the above-mentioned technical features or their equivalent features without departing from the inventive concept. For example, technical solutions formed by replacing the above-mentioned features with technical features with similar functions disclosed in this application (but not limited to) each other.

[0109] Apart from the technical features described in the specification, the other technical features are known to those skilled in the art. To highlight the innovative features of this invention, the other technical features will not be described in detail here.

Claims

1. A method for preparing embolic microspheres capable of sustained drug release and controlled degradation rate, characterized in that, At least comprising the following steps: Dissolve the dispersing agent in the oil phase liquid and heat to a first preset temperature, stirring to obtain a dispersion liquid; Dissolve the gelatin in water at a second preset temperature to obtain a gelatin solution; Dissolve the compound with negative group in water, and add alkali and initiator under ice bath to obtain a modified liquid; Add the modified liquid to the gelatin solution and stir to obtain a water phase pre-polymer liquid; After pre-mixing by dropwise adding the water phase pre-polymer liquid to the dispersion liquid, add the catalyst and crosslinking agent at the same time or add the catalyst first and then add the crosslinking agent to control the degradation rate of the prepared microspheres, the negative group is directly grafted on the gelatin molecule in the form of low temperature free radical polymerization, and an intermediate product is obtained, wherein the low temperature free radical polymerization is a polymerization reaction occurring at a temperature of 30-40℃; Post-treat the intermediate product to obtain embolization microspheres, The embolization microspheres are microspheres obtained by polymerizing the gelatin molecules and the compound with negative group and crosslinking with a crosslinking agent; wherein the surface of the gelatin molecule is directly grafted with a negative group.

2. The method for preparing sustained-release drug and controlled-degradation-rate embolic microspheres according to claim 1, characterized in that, The compound with negative group includes at least one of a compound with a carboxylic acid group or a sulfonic acid group, the carboxylic acid group is from one or more of acrylic acid, methacrylic acid, or other carboxylic acid compounds with carboxylate and double bond, or carboxylic acid salt compounds, and the sulfonic acid group is from one or more of 2-acrylamido-2-methylpropane sulfonic acid or other sulfonic acid compounds with sulfonate and double bond, or sulfonic acid salt compounds.

3. The method for preparing sustained-release drug and controlled-degradation-rate embolic microspheres according to claim 1, characterized in that, The crosslinking agent includes at least one of an acetal crosslinking agent, an amide crosslinking agent, or a secoiridoid crosslinking agent; the acetal crosslinking agent includes one or a combination of formaldehyde, glutaraldehyde, or n-butyl aldehyde; the amide crosslinking agent includes one or a combination of N,N-methylene bisacrylamide, ethylene glycol dimethacrylate, or diethylene triamine; and the secoiridoid crosslinking agent includes one or a combination of genipin, oleuropein, or aucubin.

4. The method for preparing embolic microspheres with sustained-release drug and controlled degradation rate according to claim 1, characterized in that, The catalyst includes one or a combination of N,N-dimethylaniline, sodium bisulfite, tetraethylenepentamine, tetramethyl ethylenediamine, thiol, ferrous chloride, or silver nitrate.

5. The method for preparing sustained-release drug and controlled-degradation-rate embolic microspheres according to claim 1, characterized in that, The dispersing agent includes one or a combination of Span 20, Span 80, Tween 20, Tween 80, PEG-40, or PEG-20, and the oil phase liquid includes one or a combination of liquid paraffin, glycerol, dimethyl silicone oil, white oil, vegetable oil, or petroleum ether.

6. The method for preparing sustained-release drug and controlled-degradation-rate embolic microspheres according to claim 1, characterized in that, The alkali solution includes at least one of sodium hydroxide, ammonia, or potassium hydroxide, and the initiator includes one or a combination of ammonium persulfate, potassium persulfate, hydrogen peroxide, diacyl peroxide, benzoyl peroxide, or azobisisobutyronitrile.

7. The preparation method of the embolization microspheres according to claim 1, wherein, The mass ratio of the compound with negative group to the gelatin is 1: (2-4); and / or The mass ratio of the catalyst to the gelatin is 1: (4-20); and / or The mass ratio of the crosslinking agent to the gelatin is 1: (4-20).

8. The method of claim 1, wherein the first preset temperature is 30-40°C, and the second preset temperature is 38-45°C. The first preset temperature is 30-40°C. The second preset temperature is 38-45°C.

9. The method for preparing the sustained-release drug and the embolic microspheres with controlled degradation rate according to claim 1, characterized in that, The method further comprises: adding the water phase pre-polymer droplets to the dispersion liquid, shearing at 250-350 rpm for 10-60 minutes; adding the catalyst, continuing to stir at 250-350 rpm for 150-600 minutes; and adding the cross-linking agent, stirring at 250-350 rpm for 2-60 minutes to obtain an intermediate product.

10. The method for preparing the sustained-release drug and the embolic microspheres with controlled degradation rate according to claim 1, characterized in that, The post-treatment comprises: after obtaining the intermediate product, filtering the intermediate product, adding a solvent containing a surfactant to clean for a first preset time, then washing and filtering with pure water, and then washing with water for injection, collecting the embolism microspheres intermediate phase; pre-freezing the embolism microspheres intermediate phase, freeze-drying for a second preset time to obtain the embolism microspheres.

11. The method for preparing the sustained-release drug and the embolic microspheres with controlled degradation rate according to claim 10, characterized in that, The first preset time is 10-120 minutes, and the second preset time is 60-260 hours.

12. The method for preparing the sustained-release drug and the embolic microspheres with controlled degradation rate according to claim 10, characterized in that, The surfactant comprises Tween 80.

13. A drug release-controllable embolism microsphere which can release a drug and control a degradation rate, characterized by comprising: The embolism microspheres are obtained by the method of any one of claims 1-11, and are microspheres obtained by polymerization of gelatin molecules and a compound containing a negative group and cross-linking with a cross-linking agent, wherein the gelatin molecules are directly grafted with a negative group on the surface.

14. The sustained release drug and controlled degradation rate embolizing microspheres according to claim 13, wherein, The negative group comprises at least one of a sulfonic acid group or a carboxylic acid group, and the cross-linking agent comprises at least one of an acetal cross-linking agent, an amide cross-linking agent, or a seciridoid cross-linking agent.

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