Embolism microsphere, preparation method thereof and drug-loaded embolism microsphere

Abstract

The invention relates to an embolism microsphere, a preparation method thereof and a drug-loaded embolism microsphere, and belongs to the technical field of embolism microspheres. The embolization microsphere comprises a three-dimensional network structure formed by cross-linking polymerization of a polyhydroxy water-soluble high-molecular polymer, a water-soluble cross-linking agent, a first water-soluble monomer and a second water-soluble monomer, wherein the first water-soluble monomer comprises a sulfonic acid group or a carboxylic acid group and at least one polymerizable double bond, and the second water-soluble monomer comprises a cis vicinal diol group and at least one polymerizable double bond. The embolism microsphere contains a sulfonic acid group or a carboxylic acid group and a cis vicinal diol group, so that the embolism microsphere can simultaneously load a positively charged drug and a drug containing a boric acid structure, the problem of drug resistance caused by long-time application of a single type of drug can be solved, and the tumor treatment effect is effectively improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of embolization microspheres, and particularly relates to an embolization microsphere, a preparation method thereof, and a drug-loaded embolization microsphere. BACKGROUND

[0002] At present, the embolization microspheres on the market mainly use polyvinyl alcohol as the skeleton material of the microspheres, graft a monomer with a polymerizable double bond on the molecular chain of the polyvinyl alcohol, then polymerize and crosslink the monomer with a monomer with an ionic functional group, to form a charged embolization microsphere, so as to realize the drug loading function, and then realize the dual effects of embolization and chemotherapy.

[0003] However, the existing embolization microspheres can generally only load some positively charged drugs, and these positively charged drugs, such as anthracycline drugs, are prone to drug resistance in long-term application. Therefore, the single drug loading and drug resistance result in poor tumor treatment effect of the existing drug-loaded embolization microspheres. SUMMARY

[0004] In view of the deficiencies of the prior art, the present application provides an embolization microsphere, a preparation method thereof, and a drug-loaded embolization microsphere, to improve the problem that the existing embolization microspheres can only load one type of drug and are prone to drug resistance, resulting in poor tumor treatment effect. In a first aspect, the embodiments of the present application provide an embolization microsphere, which comprises a three-dimensional network structure formed by crosslinking polymerization of a polyhydroxy water-soluble high molecular polymer, a water-soluble crosslinking agent, a first water-soluble monomer and a second water-soluble monomer; wherein the first water-soluble monomer comprises a sulfonic acid group or a carboxylic acid group and at least one polymerizable double bond, and the second water-soluble monomer comprises a cis- vicinal diol group and at least one polymerizable double bond. In the above technical solution, the embolization microsphere in the present application is obtained by crosslinking copolymerization of the first water-soluble monomer comprising a sulfonic acid group or a carboxylic acid group and the second water-soluble monomer comprising a cis- vicinal diol group. The introduction of the sulfonic acid group or the carboxylic acid group enables the embolization microsphere to load positively charged drugs through electrostatic interaction, and the introduction of the cis- vicinal diol group enables the embolization microsphere to load drugs containing a boronic acid structure through a reversible esterification reaction, so as to improve the problem of drug resistance caused by long-term application of a single type of drug. Therefore, the embolization microsphere can combine embolization therapy and chemotherapy of two types of drugs, and synergistically improve the treatment effect on tumors.

[0005] In a second aspect, the embodiments of the present application provide a preparation method of the embolization microsphere provided in the first aspect of the present application, which comprises the following steps: S1: grafting a water-soluble crosslinking agent on a polyhydroxy water-soluble high molecular polymer to obtain a modified polymer aqueous solution; S2: mixing the modified polymer aqueous solution, the first water-soluble monomer and the second water-soluble monomer, and the initiator to obtain an aqueous phase solution; S3: adding the aqueous phase solution into the oil phase solution to form a water-in-oil inverse suspension polymerization system after mixing, and obtaining the embolization microspheres after reaction.

[0006] In the technical solution, the polyhydroxy water-soluble polymer is used as a skeleton material, the hydroxyl group is used for graft modification to introduce a crosslinking reaction site, and then the first water-soluble monomer including a sulfonic acid group or a carboxylic acid group and the second water-soluble monomer including a cis-adjacent diol group are crosslinked and copolymerized to introduce the sulfonic acid group or the carboxylic acid group and the cis-adjacent diol group into the embolization microspheres. The introduction of the sulfonic acid group or the carboxylic acid group enables the embolization microspheres to load a positively charged drug through electrostatic interaction, and the introduction of the cis-adjacent diol group enables the embolization microspheres to load a drug containing a boric acid structure through a reversible esterification reaction, thereby improving the drug resistance problem caused by long-term use of a single drug. Thus, the embolization microspheres can combine embolization therapy and chemotherapy of two types of drugs to synergistically improve the treatment effect on tumors.

[0007] In a third aspect, the embodiments of the present application provide a drug-loaded embolization microsphere, which comprises the embolization microsphere provided in the first aspect of the present application and a loaded drug; wherein the loaded drug comprises at least one of a first type of drug or a second type of drug; the first type of drug comprises a positively charged drug, and the second type of drug comprises a drug containing a boric acid structure.

[0008] In the technical solution, since the embolization microspheres simultaneously have the sulfonic acid group or the carboxylic acid group and the cis-adjacent diol group, the sulfonic acid group or the carboxylic acid group can load the first type of drug, i.e., the positively charged drug, through electrostatic interaction, and the cis-adjacent diol group can load the second type of drug, i.e., the drug containing the boric acid structure, through a reversible esterification reaction to form a dynamic covalent bond, thereby enabling the drug-loaded embolization microspheres to be efficiently applied to tumor treatment. BRIEF DESCRIPTION OF DRAWINGS

[0009] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings needed in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present application, and therefore should not be considered as limiting the scope. For those skilled in the art, other related drawings can also be obtained without creative labor on the basis of these drawings.

[0010] Figure 1 The process flow chart of the preparation method of the embolization microsphere provided in the embodiments of the present application.

[0011] Figure 2 The drug release curve of the drug-loaded embolization microsphere in Embodiment 1 of the present application.

[0012] Figure 3 Fig. 1 is a graph showing the results of a cell activity test provided by an embodiment of the present application. DETAILED DESCRIPTION

[0013] Hereinafter, specific embodiments of embolization microspheres, a method for preparing embolization microspheres, and drug-loaded embolization microspheres of the present application are described in detail with appropriate reference to the accompanying drawings. However, there will be cases where unnecessary detailed descriptions are omitted. For example, there will be cases where detailed descriptions of matters well known in the art, redundant descriptions of substantially identical structures are omitted. This is to avoid the following description from becoming unnecessarily lengthy and to facilitate understanding by those skilled in the art. In addition, the accompanying drawings and the following description are provided so that those skilled in the art can fully understand the present application, and are not intended to limit the subject matter recited in the claims.

[0014] Note that "and / or" in the present application, such as "feature 1 and / or feature 2", means any of "feature 1 alone", "feature 2 alone", and "feature 1 plus feature 2".

[0015] In addition, in the description of the present application, unless otherwise specified, "two or more" in "one or more" means two or more; the range of "value a to value b" includes both end values "a" and "b", and "unit of measurement" in "value a to value b + unit of measurement" represents the "unit of measurement" of both "value a" and "value b".

[0016] Currently marketed embolization microspheres mainly use polyvinyl alcohol as the backbone material of the microspheres. A monomer with a polymerizable double bond is grafted onto the molecular chain of polyvinyl alcohol, and then the monomer is polymerized and crosslinked with a monomer having an ionic functional group to form a charged embolization microsphere, thereby realizing the drug loading function, and further realizing the dual action of embolization and chemotherapy. However, the existing drug-loaded embolization microspheres can usually only load some positively charged drugs, and these positively charged drugs, such as anthracycline drugs, are prone to drug resistance in long-term use, including cross-resistance between drugs of this class, and cross-resistance to vinblastine and vincristine, resulting in poor treatment effect on tumors. Therefore, it is urgent to combine other treatment methods to enhance long-term effective treatment of tumor cells.

[0017] Boron Neutron Capture Therapy (BNCT) is an effective treatment for tumors. In BNCT, a compound containing isotope boron-10 with strong affinity for cancer cells is introduced into the body, rapidly accumulated in cancer cells, and then a thermal neutron slowed down by normal tissue is used to react with boron-10 to release alpha particles and lithium atoms, thereby killing tumor cells. Alpha particles are extremely powerful rays that can effectively kill tumor cells, but their killing radius is only about 10 μm, which is equivalent to the diameter of a cell, so they can only kill tumor cells that undergo nuclear reactions, and have no effect on surrounding normal tissues and cells. However, to achieve good therapeutic effect, the following two conditions must be met: (1) the concentration of the drug in the tumor after administration is 3-4 times that of the normal tissue; (2) the concentration of boron in the tumor tissue needs to reach 20 μg / g. In order to meet these two conditions, a suitable means is also needed to deliver boron drugs to the vicinity of the tumor tissue. The drug commonly used for BNCT treatment is 1,4-dihydroxyboron phenylalanine (BPA), which contains a special boronic acid group. BPA can enrich boron-10 as a boron carrier, which is beneficial to the realization of radiotherapy on tumor cells through BNCT.

[0018] In addition, Bortezomib is a revolutionary proteasome inhibitor anti-tumor drug containing a unique boronic acid group. It inhibits the proteasome by reversibly binding to the threonine hydroxyl group of the β5 subunit of the proteasome, specifically inhibits the chymotrypsin-like activity of the 26S proteasome, blocks the ubiquitin-proteasome pathway, causes misfolded protein accumulation, activates apoptosis signals (such as inducing NOXA expression), inhibits the NF-κB pathway, down-regulates anti-apoptotic proteins (such as BCL-2), enhances the sensitivity of tumor cells to chemotherapy, and achieves the effect of chemotherapy on tumor cells.

[0019] Therefore, there is a need for an embolization microsphere that can simultaneously load positively charged drugs and Bortezomib and / or 1,4-dihydroxyboron phenylalanine to improve the therapeutic effect on tumors.

[0020] Therefore, there is a need for an embolization microsphere that can simultaneously load positively charged drugs and Bortezomib and / or 1,4-dihydroxyboron phenylalanine to improve the therapeutic effect on tumors. In the present application, by copolymerizing the multi-hydroxyl water-soluble polymer with the first water-soluble monomer and the second water-soluble monomer, the sulfonic acid group or the carboxylic acid group and the cis- vicinal diol group are simultaneously introduced into the embolization microspheres. The sulfonic acid group or the carboxylic acid group is negatively charged, and can load some positively charged drugs, such as anthracycline drugs, through electrostatic interaction. The two hydroxyl groups of the cis- vicinal diol group are on adjacent carbon atoms and in cis configuration, and can undergo a reversible esterification reaction with the boronic acid group in a compound containing a boronic acid structure to form a dynamic covalent bond, so as to load drugs containing a boronic acid structure, such as bortezomib and / or 1,4-dihydroxyborophenylalanine; after loading bortezomib, the chemotherapeutic effect of different curative effects can be achieved; after loading 1,4-dihydroxyborophenylalanine, the radiotherapy of boron neutron capture therapy can be introduced to achieve the synergistic therapeutic effect of chemotherapy and radiotherapy. Thus, the embolization microspheres can combine embolization therapy and the chemotherapy and radiotherapy effects brought by the two drugs carrying the boronic acid group, improve the drug resistance problem caused by the long-term use of a single drug, and synergistically improve the therapeutic effect on tumors.

[0021] Figure 1 For the process flow chart of the preparation method of the embolization microspheres provided in the embodiments of the present application, please refer to Figure 1 The preparation method of the embolization microspheres includes the following steps: S1: grafting a water-soluble crosslinking agent on a multi-hydroxyl water-soluble polymer to obtain a modified polymer aqueous solution.

[0022] The multi-hydroxyl water-soluble polymer can introduce active groups that can participate in polymerization on the polymer molecular chain by grafting the water-soluble crosslinking agent, which not only retains water solubility but also provides a connection site for subsequent crosslinking copolymerization with the first water-soluble monomer and the second water-soluble monomer.

[0023] In some embodiments, the multi-hydroxyl water-soluble polymer includes at least one of polyvinyl alcohol, sodium alginate, sodium hyaluronate, sodium carboxymethyl cellulose or water-soluble chitosan. These high molecular polymers contain a large number of hydroxyl groups and have good water solubility, which can adapt to the aqueous system, and can be grafted with the crosslinking agent to realize crosslinking and curing, and also have good biocompatibility.

[0024] In some embodiments, the water-soluble crosslinking agent includes at least one aldehyde group, acetal or hemiacetal group and at least one polymerizable double bond. The aldehyde group, acetal or hemiacetal group in the water-soluble crosslinking agent can undergo acetalization / hemiacetalization reaction with the hydroxyl group in the multi-hydroxyl water-soluble polymer to form a stable grafted crosslinking structure; and the polymerizable double bond can make the modified polymer and the two water-soluble monomers undergo free radical copolymerization under the action of a subsequent initiator, thereby forming a stable three-dimensional network structure.

[0025] Further, the water-soluble crosslinking agent includes at least one of N-(2,2-dimethoxy)-2-methyl acrylamide, N-(1-hydroxy-2,2-dimethoxyethyl)-2-propenamide, N-(2,2-dimethoxy)-2-propenamide, N-(1-hydroxy-2,2-dimethoxyethyl)-2-methyl acrylamide, N-(2,2-dimethoxy)-2-methyl acrylate, N-(1-hydroxy-2,2-dimethoxyethyl)-2-propenoate, N-(2,2-dimethoxy)-2-propenoate or N-(1-hydroxy-2,2-dimethoxyethyl)-2-methyl acrylate.

[0026] In some embodiments, the water-soluble crosslinking agent accounts for 2% to 6% of the mass percentage of the polyhydroxyl water-soluble high molecular polymer. The amount of the water-soluble crosslinking agent in the appropriate range is conducive to generating moderate crosslinking degree, giving the embolization microspheres good mechanical strength and compression elasticity.

[0027] By way of example, the water-soluble crosslinking agent accounts for any one value or a range value between any two values of 2%, 3%, 4%, 5% or 6% of the mass percentage of the polyhydroxyl water-soluble high molecular polymer.

[0028] In some embodiments, the step S1 includes: dissolving the polyhydroxyl water-soluble high molecular polymer in water to obtain a polymer aqueous solution; sequentially adding the water-soluble crosslinking agent and the acid catalyst to the polymer aqueous solution, and reacting at 20°C to 30°C for 10h to 15h to obtain a modified polymer aqueous solution. By first dissolving the polyhydroxyl water-soluble high molecular polymer in water to expose the reaction sites (i.e. hydroxyl groups), and then sequentially adding the water-soluble crosslinking agent and the acid catalyst to catalyze the grafting reaction in an acidic environment, the water-soluble crosslinking agent is effectively grafted onto the polymer molecular chain, and active sites (i.e. polymerizable double bonds) are introduced for subsequent polymerization to form microspheres.

[0029] By way of example, the reaction temperature can be 20°C, 22°C, 25°C, 28°C, 30°C, etc.; and the reaction time can be 10h, 12h, 14h, 15h, etc.

[0030] Further, the mass percentage of the polyhydroxyl water-soluble high molecular polymer in the polymer aqueous solution is 15% to 25%, for example, 15%, 18%, 20%, 22%, 25%, etc. The moderate concentration of the polyhydroxyl water-soluble high molecular polymer is conducive to the full dissolution of the polymer and provides sufficient hydroxyl reaction sites, and makes the reaction system maintain an appropriate viscosity, which is conducive to improving the efficiency and uniformity of the grafting reaction.

[0031] Further, the temperature for dissolving the polyhydroxyl water-soluble high molecular polymer in water is 90°C to 100°C, for example, 90°C, 92°C, 95°C, 100°C, etc.

[0032] Further, the acid catalyst includes at least one of hydrochloric acid, sulfuric acid, nitric acid, or p-toluenesulfonic acid. Still further, the acid catalyst is hydrochloric acid.

[0033] Further, the acid catalyst accounts for 20% to 40% of the mass percentage of the polyhydroxyl water-soluble polymer, such as 20%, 25%, 30%, 35%, 40%, etc.

[0034] S2: mixing the modified polymer aqueous solution, the first water-soluble monomer and the second water-soluble monomer, and the initiator to obtain an aqueous phase solution; wherein the first water-soluble monomer includes a sulfonic acid group or a carboxylic acid group and at least one polymerizable double bond, and the second water-soluble monomer includes a cis vicinal diol group and at least one polymerizable double bond.

[0035] In some embodiments, the first water-soluble monomer includes at least one of 2-acrylamido-2-methylpropanesulfonic acid sodium, allyl sulfonic acid sodium, methacryl sulfonic acid sodium, or methacrylic acid sodium. The 2-acrylamido-2-methylpropanesulfonic acid sodium, the allyl sulfonic acid sodium, the methacryl sulfonic acid sodium contain a sulfonic acid group, and the methacrylic acid sodium contains a carboxylic acid group, and all have good water solubility, polymerization activity, and biocompatibility.

[0036] In some embodiments, the first water-soluble monomer accounts for 5% to 15% of the mass percentage of the modified polymer aqueous solution. The proportion of the first water-soluble monomer in the appropriate range is conducive to introducing more sulfonic acid groups or carboxylic acid groups in the embolization microspheres, while being conducive to maintaining the structural stability of the embolization microspheres.

[0037] By way of example, the first water-soluble monomer accounts for 5%, 8%, 10%, 12%, 15%, or any range between any two of the values.

[0038] In some embodiments, the second water-soluble monomer includes at least one of mannitol acrylate, talitol acrylate, gulitol acrylate, galactitol acrylate, sorbitol acrylate, iditol acrylate, or alitol acrylate. These monomers all contain a cis vicinal diol group and a polymerizable double bond, and have good water solubility and biocompatibility.

[0039] In some embodiments, the second water-soluble monomer accounts for 5% to 15% of the mass percentage of the modified polymer aqueous solution. The proportion of the second water-soluble monomer in the appropriate range is conducive to introducing more cis vicinal diol groups in the embolization microspheres after polymerization, while being conducive to maintaining the structural stability of the embolization microspheres.

[0040] For example, the second water-soluble monomer accounts for 5%, 8%, 10%, 12%, 15%, or a range between any two of the values.

[0041] In some embodiments, step S2 further comprises adding a chain regulator with at least two polymerizable double bonds. In the presence of two drug-loaded monomers, steric hindrance exists, which affects the drug loading capacity of the embolization microspheres. In the embodiments of the present application, the chain regulator is added to participate in free radical copolymerization, and the two polymerizable double bonds can be covalently combined with the first water-soluble monomer and the second water-soluble monomer respectively to form a bridge, thereby adjusting the covalent crosslinking distance between the first water-soluble monomer and the second water-soluble monomer, and further adjusting the steric hindrance between the two water-soluble monomers, so as to improve the drug loading rate of each of the two water-soluble monomers, and further improve the drug loading capacity of the embolization microspheres.

[0042] In some embodiments, the chain regulator comprises an acrylate compound with at least two polymerizable double bonds. This type of chain regulator has good polymerization activity and water solubility, and is easy to adjust the covalent crosslinking distance between the first water-soluble monomer and the second water-soluble monomer according to the length of the molecular chain segment, while adjusting the strength and elasticity of the embolization microspheres.

[0043] Further, the chain regulator comprises at least one of ethylene glycol diacrylate, propylene glycol diacrylate, butanediol diacrylate, or pentanediol diacrylate. These chain regulators all contain a flexible alkane chain segment and two polymerizable double bonds, the length of the alkane chain segment is different, the distance between the two polymerizable double bonds is different, and the influence on the structure and drug loading capacity of the embolization microspheres is also different. By selecting an appropriate chain regulator, the drug loading capacity can be effectively improved while maintaining the structural strength and elasticity of the embolization microspheres.

[0044] In some embodiments, the chain regulator accounts for 2.5% to 7.5% of the mass percentage of the modified polymer aqueous solution, for example, 2.5%, 3.5%, 5.5%, 7.5%, or a range between any two of the values. The proportion of the chain regulator in the appropriate range is conducive to providing sufficient crosslinking sites, effectively adjusting the steric hindrance of the first water-soluble monomer and the second water-soluble monomer, and at the same time reducing the influence on the overall strength and elasticity of the embolization microspheres.

[0045] In some embodiments, the chain regulator accounts for 25% to 55% of the mass percentage of the first water-soluble monomer, for example, 25%, 35%, 45%, 55%, or a range between any two of the values.

[0046] In some embodiments, the chain regulator accounts for 25% to 55% of the mass percentage of the second water-soluble monomer, for example, 25%, 35%, 45%, 55%, or a range between any two of the values.

[0047] The proportion of the chain regulator is in a suitable range, which is conducive to effectively regulating the steric hindrance of the first water-soluble monomer and the second water-soluble monomer, while reducing the influence on the copolymerization of the monomers themselves, so as to balance the steric regulation and drug loading efficiency.

[0048] In some embodiments, the initiator includes at least one of ammonium persulfate, sodium persulfate, or potassium persulfate.

[0049] In some embodiments, the initiator accounts for 0.1% to 0.6% of the mass percentage of the modified polymer aqueous solution, for example, any one value or a range value between any two values of 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, or 0.6%.

[0050] S3: The aqueous phase solution is added to the oil phase solution, and after mixing, a water-in-oil reverse suspension polymerization system is formed, and embolization microspheres are obtained after reaction.

[0051] In the process, the aqueous phase solution serves as the dispersed phase, the oil phase solution serves as the continuous phase, the modified polymer in the aqueous phase solution serves as the basic skeleton, the first water-soluble monomer provides sulfonic acid groups or carboxylic acid groups, and the second water-soluble monomer provides cis- vicinal diol groups. Under the action of the initiator, the modified polymer, the first water-soluble monomer, and the second water-soluble monomer undergo free radical copolymerization, and the crosslinking agent realizes intermolecular crosslinking, and finally the embolization microspheres are formed.

[0052] In some embodiments, the reaction is carried out under stirring, the temperature of the reaction is 50°C to 80°C, the time is 3h to 15h, and the stirring rate is 200rpm to 800rpm.

[0053] For example, the temperature of the reaction can be any one value or a range value between any two values of 50°C, 60°C, 70°C, or 80°C; the time can be any one value or a range value between any two values of 3h, 5h, 10h, 12h, or 15h; and the stirring rate can be any one value or a range value between any two values of 200rpm, 300rpm, 500rpm, 600rpm, or 800rpm.

[0054] In some embodiments, the preparation method of the oil phase solution includes: dissolving an oily dispersant in an oily solvent to obtain the oil phase solution.

[0055] In the process, the oily dispersant includes at least one of Tween-60, Tween-80, Span-60, Span-80, OP-10, OP-20, or cellulose acetate butyrate, and the oily solvent includes at least one of butyl acetate, ethyl acetate, methyl acetate, or propyl acetate.

[0056] Further, the mass percentage of the oily dispersant in the oil phase solution is 2% to 6%, for example, any one value or a range value between any two values of 2%, 3%, 4%, 5%, or 6%.

[0057] Further, the solubility temperature of the oily dispersant is 30°C to 50°C, for example, any one value or a range value between any two values of 30°C, 35°C, 40°C, 45°C, or 50°C.

[0058] The embodiment of the present application also provides a drug-loaded embolization microsphere, which comprises the embolization microsphere provided in the first aspect of the embodiment of the present application and a drug; and the drug comprises at least one of a first drug or a second drug; the first drug comprises a positively charged drug; and the second drug comprises a drug containing a boric acid structure.

[0059] In some embodiments, the positively charged drug comprises at least one of an anthracycline drug, a camptothecin drug, a vinca alkaloid drug, or a nucleoside drug.

[0060] The anthracycline drug can comprise at least one of doxorubicin (adriamycin), aclarubicin, epirubicin, idarubicin, valrubicin; the camptothecin drug can comprise at least one of irinotecan and topotecan; the vinca alkaloid drug can comprise at least one of vinorelbine and vindesine; and the nucleoside drug can comprise at least one of gemcitabine and capecitabine.

[0061] In some embodiments, the drug containing a boric acid structure comprises at least one of bortezomib, ixazomib, or 1,4-dihydroxyborophenylalanine.

[0062] To make the purpose, technical scheme and advantages of the embodiments of the present application more clear, the technical scheme in the embodiments of the present application will be described clearly and completely below. If the specific conditions are not indicated in the embodiments, the conventional conditions or the conditions suggested by the manufacturer are adopted. If the reagents or instruments are not indicated by the manufacturer, they are all conventional products that can be purchased in the market.

[0063] Embodiment 1 The embodiment provides an embolization microsphere, and a preparation method thereof comprises the following steps: (1) Preparation of modified polymer aqueous solution 100 g of polyvinyl alcohol (a polyhydroxy water-soluble polymer) is added to 500 mL of water, and the temperature is raised to 95°C for stirring and dissolution to obtain a polymer aqueous solution; 3.00 g of N-(2,2-dimethoxy)-2-methyl acrylamide (a water-soluble crosslinking agent) is added to the polymer aqueous solution, and after stirring uniformly, 30 mL of hydrochloric acid (an acid catalyst) is added, and the stirring reaction is continuously carried out at 25°C for 15 h to obtain a modified polymer aqueous solution.

[0064] (2) Preparation of aqueous phase solution 10 g of 2-acrylamide-2-methylpropanesulfonic acid (first water-soluble monomer), 10 g of mannitol acrylate monomer (second water-soluble monomer), 0.3 g of potassium persulfate (initiator), and 100 g of modified polymer aqueous solution were stirred and mixed uniformly to obtain the aqueous phase solution.

[0065] (3) Preparation of oil phase solution 20 g of cellulose acetate butyrate (oil dispersant) was added to 500 mL of butyl acetate (oil solvent), and the mixture was stirred and dissolved at 35°C to form a uniform solution, thereby obtaining the oil phase solution.

[0066] (4) Preparation of embolization microspheres Under the condition of stirring at 500 rpm, the aqueous phase solution was slowly added to the oil phase solution to form a water-in-oil inverse suspension polymerization system. After the addition was completed, the reaction system was heated to 65°C, and a certain amount of catalyst tetramethyl ethylenediamine was continuously added to the reaction system to perform inverse suspension polymerization. After 8 h, the reaction was stopped, and the reaction system was allowed to stand and separate into layers. The oil phase was separated, and the microspheres were collected and repeatedly washed and sieved to obtain the embolization microspheres.

[0067] Example 2 The embolization microspheres provided in this example differ from those of Example 1 in that: In step (2), 5 g of ethylene glycol diacrylate (chain regulator) was further added.

[0068] Example 3 The embolization microspheres provided in this example differ from those of Example 2 in that: The chain regulator is propylene glycol diacrylate.

[0069] Example 4 The embolization microspheres provided in this example differ from those of Example 2 in that: The chain regulator is butanediol diacrylate.

[0070] Example 5 The embolization microspheres provided in this example differ from those of Example 2 in that: The chain regulator is pentanediol diacrylate.

[0071] Performance test and result analysis 1. Mechanical properties The blank embolization microspheres with a particle size range of 100 μm to 300 μm in Examples 1 to 5 were taken, and the strength, elasticity, and catheter passability of the microspheres were tested. The test results are shown in Table 1. The specific test methods are as follows: (1) Strength and elasticity: spread on a glass slide, place under the probe of a texture analyzer (TA.XT Plus C), select Hold Compression mode for testing.

[0072] (2) Catheter passability: mix the embolization microspheres in normal saline, use a syringe to suck the well-mixed microsphere solution, connect a 2.6F microcatheter, and inject at a speed of 1 mL / min, and observe whether the microspheres pass smoothly.

[0073] Table 1 Test results of mechanical properties of embolization microspheres

[0074] As can be seen from Table 1, the embolization microspheres prepared in Examples 1-5 all have high strength and elasticity, and the strength can be greater than 70 g and the compression elasticity is greater than 40%. And from the comparison of Examples 1, 2-5, it can be seen that the strength and elasticity of the embolization microspheres can be adjusted by adjusting the length of the alkane chain segment of the chain regulator, and as the length of the alkane chain segment of the chain regulator shortens, the strength and elasticity of the microspheres will increase.

[0075] 2. Drug loading performance Mix 2 mL of embolization microspheres with a particle size of 100-150 μm with 8 mL of doxorubicin solution (Dox, 20 mg / mL), stir at 25°C for 1 min, test the residual amount of doxorubicin in the supernatant by high performance liquid chromatography, and then calculate the doxorubicin loading capacity of the embolization microspheres; then mix with 8 mL of bortezomib solution (Bor, 20 mg / mL), stir at 25°C for 1 min, test the residual amount of bortezomib in the supernatant by high performance liquid chromatography, and then calculate the bortezomib loading capacity of the embolization microspheres. The test results are shown in Table 2.

[0076] Table 2 Test results of drug loading of embolization microspheres

[0077] As can be seen from Table 2, the embolization microspheres in Examples 1-5 can simultaneously load doxorubicin and bortezomib, i.e. can simultaneously load two types of drugs, and have high drug loading capacity and drug loading efficiency. From the comparison of Examples 1, 2-4, it can be seen that as the length of the alkane chain segment of the chain regulator increases, the drug loading capacity of the two drugs increases due to the decrease in steric hindrance between the two drug loadable monomers.

[0078] 3. Drug release performance The drug-loaded microspheres corresponding to the above-mentioned embodiment 1 were placed in a dialysis bag, and then the dialysis bag was placed in a centrifuge tube, physiological saline (20 mL) was added to the centrifuge tube, the centrifuge tube was sealed and placed on a constant temperature oscillator (37.0°C) at a speed of 100 rpm; sample was taken every time, and after dilution of 0.1 mL solution at different time points, the concentrations of doxorubicin and bortezomib solution were tested by high performance liquid chromatography, and the cumulative release amount of the drug-loaded embolization microspheres was calculated. The cumulative release rate calculation formula is as follows: ; In the formula, Q is the cumulative release rate of the drug (%), M D is the mass of the loaded drug, n is the number of drug taking, C n is the sample concentration of the nth sample, V is the total volume of the release liquid, C i is the sample concentration of the ith sample, V i is the sample volume of the ith sample, C0 and V0 are both 0.

[0079] Figure 2 is the drug release curve diagram of the drug-loaded embolization microspheres in the present application embodiment 1, from Figure 2 It can be seen that the doxorubicin and bortezomib in the drug-loaded embolization microspheres can be slowly released, and the cumulative release rate is 79% in 16 days.

[0080] 4. Cell activity test MTT colorimetric method was used to determine the cell activity. The cells were divided into complete culture medium (recorded as blank group), embolization microspheres in embodiment 1 (recorded as EM group), pure doxorubicin (1 μg / mL, recorded as Dox group), pure bortezomib (1 μg / mL, recorded as Bor group), embolization microspheres loaded with doxorubicin (the content of Dox is 1 μg / mL, recorded as Dox@EM group), embolization microspheres loaded with bortezomib (the content of Bor is 1 μg / mL, recorded as Bor@EM group), and embolization microspheres loaded with doxorubicin and bortezomib (the content of Dox is 1 μg / mL, and the content of Bor is 1 μg / mL, recorded as Dox+Bor@EM group), three parallel holes were set for each group, and the ultrasonic condition was 0.6 W / cm 2 , 60 s. HepG2 liver cancer cells were inoculated in a 96-well plate at a quantity of 8×10 3 After the cells adhered, the blank group and the experimental group were treated for 24 h, the culture medium was removed, and after the MTT solution (0.5 mg / mL) was cultured at 37°C for 4 h, 100 μL of dimethyl sulfoxide was used to dissolve the MTT crystal, and the absorbance was measured at 490 nm using a cell imaging microplate detector (Bio-Tek Cytation5).

[0081] Figure 3 is the cell activity test result diagram provided by the present application embodiment, fromFigure 3 It can be seen that embolization microspheres simultaneously loaded with doxorubicin and bortezomib (Dox+Bor@EM group) have a stronger tumor cell killing effect than embolization microspheres loaded with doxorubicin alone (Dox@EM group) or embolization microspheres loaded with bortezomib alone (Bor@EM group).

[0082] The embodiments described above are part of, but not all, embodiments of the present application. The detailed description of the embodiments of the present application is not intended to limit the scope of the present application, but merely represents selected embodiments of the present application. Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative work are within the scope of protection of the present application.

Claims

1. An embolizing microsphere, characterized by, The three-dimensional network structure is formed by cross-linking polymerization of a polyhydroxy water-soluble high-molecular polymer, a water-soluble cross-linking agent, a first water-soluble monomer and a second water-soluble monomer; wherein the first water-soluble monomer comprises a sulfonic acid group or a carboxylic acid group and at least one polymerizable double bond, and the second water-soluble monomer comprises a cis-adjacent diol group and at least one polymerizable double bond.

2. The embolizing microspheres according to claim 1, characterized in that, The first water-soluble monomer comprises at least one of 2-acrylamide-2-methylpropane sulfonic acid sodium, allyl sulfonic acid sodium, methyl acrylate sulfonic acid sodium or sodium methacrylate; And / or, the second water-soluble monomer comprises at least one of mannitol acrylate, talitol acrylate, gulitol acrylate, galactitol acrylate, sorbitol acrylate, iditol acrylate or alitol acrylate; And / or, the polyhydroxy water-soluble high-molecular polymer comprises at least one of polyvinyl alcohol, sodium alginate, sodium hyaluronate, sodium carboxymethyl cellulose or water-soluble chitosan; And / or, the water-soluble cross-linking agent comprises at least one aldehyde group, acetal or hemiacetal group and at least one polymerizable double bond; optionally, the water-soluble cross-linking agent comprises at least one of N-(2,2-dimethoxy)-2-methyl acrylamide, N-(1-hydroxy-2,2-dimethoxyethyl)-2-acrylamide, N-(2,2-dimethoxy)-2-acrylamide, N-(1-hydroxy-2,2-dimethoxyethyl)-2-methyl acrylamide, N-(2,2-dimethoxy)-2-methacrylate, N-(1-hydroxy-2,2-dimethoxyethyl)-2-acrylate, N-(2,2-dimethoxy)-2-acrylate or N-(1-hydroxy-2,2-dimethoxyethyl)-2-methacrylate.

3. The embolizing microspheres according to claim 1, wherein, The embolization microspheres further comprise a chain regulator with at least two polymerizable double bonds, and the chain regulator, the first water-soluble monomer and the second water-soluble monomer are covalently cross-linked and polymerized to form the three-dimensional network structure.

4. The embolizing microspheres according to claim 3, characterized in that, The chain regulator is an acrylate compound with at least two polymerizable double bonds; Optionally, the chain regulator comprises at least one of ethylene glycol diacrylate, propylene glycol diacrylate, butanediol diacrylate or pentanediol diacrylate.

5. A method of producing embolizing microspheres as claimed in any one of claims 1 to 4, characterized by, The method comprises the following steps: S1: grafting a water-soluble cross-linking agent on a polyhydroxy water-soluble high-molecular polymer to obtain a modified polymer aqueous solution; S2: mixing the modified polymer aqueous solution, a first water-soluble monomer and a second water-soluble monomer and an initiator to obtain an aqueous phase solution; S3: adding the aqueous phase solution into an oil phase solution to form a water-in-oil reverse phase suspension polymerization system after mixing, and obtaining embolization microspheres after reaction.

6. The preparation method according to claim 5, characterized in that, The water-soluble cross-linking agent accounts for 2% to 6% of the mass percentage of the polyhydroxy water-soluble high-molecular polymer; And / or, the first water-soluble monomer accounts for 5% to 15% of the mass percentage of the modified polymer aqueous solution; And / or, the second water-soluble monomer accounts for 5% to 15% of the mass percentage of the modified polymer aqueous solution.

7. The preparation method according to claim 5, characterized in that, The first water-soluble monomer comprises at least one of sodium 2-acrylamido-2-methylpropanesulfonate, sodium allylsulfonate, sodium methacrylsulfonate or sodium methacrylate; And / or, the second water-soluble monomer comprises at least one of mannitol acrylate, talitol acrylate, gulitol acrylate, galactitol acrylate, sorbitol acrylate, iditol acrylate or alitol acrylate.

8. The preparation method according to claim 5, characterized in that, The step S2 further comprises adding a chain regulator with at least two polymerizable double bonds; Optionally, the chain regulator comprises an acrylate compound with at least two polymerizable double bonds; Optionally, the chain regulator comprises at least one of ethylene glycol diacrylate, propylene glycol diacrylate, butylene glycol diacrylate or pentanediol diacrylate.

9. The production method according to claim 8, characterized by, The chain regulator accounts for 2.5% to 7.5% of the mass percentage of the modified polymer aqueous solution; And / or, the chain regulator accounts for 25% to 55% of the mass percentage of the first water-soluble monomer; And / or, the chain regulator accounts for 25% to 55% of the mass percentage of the second water-soluble monomer.

10. A drug-loaded embolizing microsphere, characterized in that, The embolization microspheres comprise at least one of the first drug or the second drug; The first drug comprises a positively charged drug; optionally, the positively charged drug comprises at least one of an anthracycline drug, a camptothecin drug, a vinca alkaloid drug or a nucleoside drug; The second drug comprises a drug containing a boronic acid structure; optionally, the drug containing a boronic acid structure comprises at least one of bortezomib, ixazomib or 1,4-dihydroxyborophenylalanine.

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