Novel microspheres using anionic polymer, and preparation method and composition thereof

Abstract

The present invention relates to novel microspheres using an anionic polymer and a preparation method therefor. The present invention provides a novel preparation method for and an optimal formulation of microspheres which are useful for the delivery of cationic drugs, are less toxic and biocompatible, and can deliver drugs to microvessels. The novel preparation method according to the present invention enables industrial mass production of microspheres of a specific size by improving the yield of microspheres of a desired size through size distribution adjustment of microsphere particles.

Description

Novel microspheres using anionic polymers, their preparation method and composition

[0001] The present invention relates to a novel microsphere and a manufacturing method using an anionic polymer.

[0002] This invention claims priority from Republic of Korea Patent Application No. 10-2023-0100331, filed August 1, 2023, and the invention claims priority from Republic of Korea Patent Application No. 10-2024-0101947, filed July 31, 2024, the entire contents of which are incorporated herein by reference.

[0003]

[0004] Liver cancer is a malignant tumor with a poor prognosis, ranking second among cancer-related deaths in Korea and third worldwide. Curative surgery is not possible for more than 70% of liver cancer patients, and even after curative resection, the risk of recurrence in other areas within five years is over 50%. The likelihood of advanced liver cancer responding to systemic chemotherapy is extremely low, less than 10%. As a non-surgical alternative, various image-guided therapies have been developed. Two representative image-guided therapies for liver cancer are transarterial chemoembolization (TACE), which involves local injection of anticancer drugs into the liver tumor via the hepatic artery and subsequent embolization of the hepatic artery. Radiofrequency ablation (RFA) is another method.

[0005] More than 95% of the blood flow to liver cancer tissue is delivered to the hepatic artery, but more than 75% of the blood flow to surrounding healthy tissue is delivered to the portal vein and only 25% is delivered to the hepatic artery, so TACE is a safe and effective treatment method that does not cause damage to normal tissue.

[0006] TACE is usually performed repeatedly three to four times a year, so the administered substances must be safe. The drugs mainly used in conventional arterial embolization include doxorubicin, cisplatin, epirubicin, mitoxantrone, and mitomycin C. Since these anticancer drugs cannot be directly dissolved in water-insoluble contrast media, they are dissolved in water-soluble contrast media such as Pamiray, and then homogeneously dispersed in a water-insoluble contrast media (iodized oil) such as Lipiodol to prepare an oil-in-water or water-in-oil emulsion. These emulsions undergo phase separation after a certain period of time, so they are prepared and used immediately before patient administration. After performing chemical embolization, gelatin sponge particles are additionally administered to promote ischemic necrosis of cancer tissue and prevent drug loss. However, the administered emulsion undergoes phase separation shortly after administration, resulting in the immediate absorption of the anticancer drug. This results in a short-term antitumor effect and potential side effects due to systemic exposure. Furthermore, most cytotoxic anticancer drugs administered in TACE are metabolized in the liver, potentially causing hepatotoxicity. For these reasons, patients experience postembolization syndrome, which hinders their daily activities and necessitates hospitalization for 2-3 days, resulting in high medical costs.

[0007] Recently, drug-eluting beads (DEBs), a selectively used chemical embolic particle, are made of non-absorbable polyvinyl alcohol (PVA) and deliver drugs by ion-exchange adsorption. Their particle size ranges from 40 to 900 μm. Compared to emulsion droplets, DEBs offer a more homogeneous and reproducible particle size distribution, and their slow drug release reduces systemic exposure. However, the drug is not fully released after administration. Less than 50% is released by day 28, and approximately 90% is released after day 90. This results in relatively slow drug accumulation in cancer tissues, resulting in high concentrations. Furthermore, the drug absorption time can exceed 1 to 2 hours, and the non-absorbable nature of DEBs means that even after complete drug release, the particles remain in the body. This can lead to necrosis of normal tissues, making repeat TACE procedures difficult and increasing the risk of cancer recurrence. Furthermore, the drug costs are higher than those of conventional TACE, placing a significant financial burden on patients.

[0008] Therefore, there is a need for the development of microspheres that are low in toxicity, biocompatible, and can deliver target drugs to microvessels quickly and efficiently.

[0009]

[0010] The present inventors have made extensive efforts to develop biocompatible microspheres that are easy to manufacture and have excellent drug-bearing capacity, and as a result, have identified a formulation optimized for the manufacture of microspheres. They have also confirmed that the yield of particles of a desired size can be improved by controlling the particle size distribution of the microspheres by adjusting the rpm of the homogenizer, thereby completing the present invention.

[0011] Accordingly, an object of the present invention is to provide a composition for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt as active ingredients.

[0012] Another object of the present invention is to provide a method for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt.

[0013] Another object of the present invention is to provide a method for preparing a composition for treating cancer.

[0014] Another object of the present invention is to provide microspheres comprising an anionic polymer and a polyvalent metal compound salt.

[0015]

[0016] However, the technical problems to be solved by the present invention are not limited to the problems mentioned above, and other problems not mentioned can be clearly understood by a person having ordinary skill in the technical field to which the present invention belongs from the description below.

[0017]

[0018] To achieve the above purpose, the present invention provides a composition for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt as active ingredients.

[0019] In addition, the present invention provides a use for producing microspheres of an anionic polymer and a polyvalent metal compound salt; or a composition comprising the same as an active ingredient.

[0020] In addition, the present invention provides a use of an anionic polymer and a polyvalent metal compound salt for the production of microspheres; or a composition comprising the same as an active ingredient.

[0021] In one embodiment of the present invention, the anionic polymer may be selected from the group consisting of, but is not limited to, carboxymethylcellulose or a salt thereof; dextran sulfate or a salt thereof; and mixtures thereof.

[0022] In another embodiment of the present invention, the microspheres may include, but are not limited to, carboxymethylcellulose and dextran sulfate.

[0023] In another embodiment of the present invention, the carboxymethylcellulose and dextran sulfate may be included in a mass ratio of 0.5 to 20:1, but is not limited thereto.

[0024] In another embodiment of the present invention, the carboxymethylcellulose and dextran sulfate may be included in a concentration ratio of 0.5 to 20:1, but is not limited thereto.

[0025] In another embodiment of the present invention, the multivalent metal compound salt is Zr 2+ , Zr 4+ , Fe 2+ , Fe 3+ , Ca 2+ , Al 3+ , Cu 2+ , and Zn 2+ It may be a salt of one or more divalent to tetravalent metal compounds selected from the group consisting of, but is not limited thereto.

[0026] In another embodiment of the present invention, the anionic polymer and the polyvalent metal compound salt may be included in a concentration ratio of 0.1 to 10:1, but is not limited thereto.

[0027] In another embodiment of the present invention, the microspheres may be loaded with a cationic drug, but are not limited thereto.

[0028] In another embodiment of the present invention, the cationic drug may be at least one selected from the group consisting of doxorubicin, procainamide, digoxin, quinidine, trimethoprim, cimetidine, vancomycin, irinotecan, daunorubicin, epirubicin, diphenhydramine, memantine, oxycodone, pyrilamine, tramadol, and pharmaceutically acceptable salts thereof, but is not limited thereto.

[0029] In another embodiment of the present invention, the cationic drug may be loaded into the microspheres at a mass ratio of cationic drug: microsphere = 1:1 to 10, but is not limited thereto.

[0030] In another embodiment of the present invention, the composition may be, but is not limited to, an emulsion formulation.

[0031] In another embodiment of the present invention, the microspheres may have a diameter of, but not limited to, 10 to 700 μm.

[0032] In another embodiment of the present invention, the microspheres may have a size distribution as follows, but is not limited thereto:

[0033] a) 10 to 50 μm;

[0034] b) 50 to 100 μm;

[0035] c) 100 to 300 μm;

[0036] d) 300 to 500 μm; and

[0037] e) 500 to 700 μm.

[0038] In another embodiment of the present invention, the microspheres may be separated according to size distribution, but are not limited thereto.

[0039] In another embodiment of the present invention, the microspheres can be used for, but are not limited to, cancer treatment.

[0040] In another embodiment of the present invention, the microspheres can be used for, but are not limited to, vascular embolization.

[0041] In addition, the present invention provides a method for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt, comprising the following steps:

[0042] A step of forming an emulsion by mixing an aqueous phase containing an anionic polymer and an oil phase containing a nonionic surfactant using a homogenizer;

[0043] A step of forming microspheres by adding the above emulsion to a multivalent metal compound salt aqueous solution; and

[0044] A step of separating microspheres of a desired size using a sieve.

[0045] In one embodiment of the present invention, the anionic polymer may be selected from the group consisting of, but is not limited to, carboxymethylcellulose or a salt thereof; dextran sulfate or a salt thereof; and mixtures thereof.

[0046] In another embodiment of the present invention, the multivalent metal compound salt is Zr 2+ , Zr 4+ , Fe 2+ , Fe 3+ , Ca 2+ , Al 3+ , Cu 2+ , and Zn 2+ It may be a salt of one or more divalent to tetravalent metal compounds selected from the group consisting of, but is not limited thereto.

[0047] In another embodiment of the present invention, the nonionic surfactant may be selected from the group consisting of polyvinyl alcohol, polyalkylene glycol, polyvinyl pyrrolidone, alkyl polyglycoside, t-octylphenoxy polyethoxy ethanol, poloxamer, polysorbate, poly(N-vinylacetamide), polyvinyl butyral, sorbitan oleate, and mixtures thereof, but is not limited thereto.

[0048] In another embodiment of the present invention, the oil phase may be selected from the group consisting of, but not limited to, mineral oil, vegetable oil, medium chain triglyceride, and mixtures thereof.

[0049] In another embodiment of the present invention, the water phase and the oil phase may be mixed in a volume ratio of 1:1 to 20, but is not limited thereto.

[0050] In another embodiment of the present invention, the emulsion may be added to the polyvalent metal compound salt aqueous solution at a volume ratio of 1:1 to 20, but is not limited thereto.

[0051] In another embodiment of the present invention, the method can control the size distribution of microspheres by controlling the rpm of the homogenizer, but is not limited thereto.

[0052] The present invention also provides a method for preparing a composition for treating cancer, comprising the following steps:

[0053] A step of forming an emulsion by mixing an aqueous phase containing an anionic polymer and an oil phase containing a nonionic surfactant using a homogenizer;

[0054] A step of forming microspheres by adding the above emulsion to a multivalent metal compound salt aqueous solution;

[0055] A step of separating microspheres of a desired size using a sieve; and

[0056] A step of mixing a cationic drug with the above microspheres.

[0057] In addition, the present invention provides microspheres comprising an anionic polymer and a polyvalent metal compound salt.

[0058] Additionally, the present invention provides a drug delivery use of the microspheres.

[0059] The present invention also provides the use of the microspheres for the manufacture of drug delivery vehicles.

[0060] In addition, the present invention provides a composition for cationic drug delivery comprising the above microspheres and a cationic drug as active ingredients.

[0061] Additionally, the present invention provides a cationic drug delivery method comprising a step of administering the microspheres encapsulated with a cationic drug to a subject in need thereof.

[0062] In addition, the present invention provides a composition comprising the above microspheres and a cationic drug as active ingredients for use in treating vascular embolism and / or cancer.

[0063] In addition, the present invention provides a composition for vascular embolization or cancer treatment comprising the above microspheres and a cationic drug as active ingredients.

[0064] In addition, the present invention provides a method for treating vascular embolism and cancer, comprising administering to a subject a composition comprising the above microspheres and a cationic drug as active ingredients.

[0065] In addition, the present invention provides a use of a composition comprising the above microspheres and a cationic drug as active ingredients for the manufacture of a vascular embolization and cancer treatment agent.

[0066]

[0067] The present invention provides a novel method for manufacturing microspheres, which are useful for the delivery of cationic drugs, have low toxicity, are biocompatible, and can deliver drugs to microvessels, and an optimal formulation. In addition, the microsphere manufacturing method of the present invention allows the microspheres to directly embolize blood vessels by adjusting the particle size according to the size of the blood vessel through controlling the size distribution of the microsphere particles, thereby improving the yield of microspheres of a desired size, thereby enabling the mass production of microspheres of a specific size.

[0068]

[0069] Figure 1 shows an image of changes in the reaction solution when an anionic polymer and calcium chloride (CaCl2) are reacted.

[0070] Figure 2 shows an optical microscope measurement image of a solid produced through composition 2 of Example 1-2, <Reactivity Test 2>.

[0071] Figure 3 shows an image of microparticles manufactured through the reaction of an anionic polymer and iron chloride according to Composition 2 of Example 3, Table 3.

[0072] Figure 4 shows the results of measuring the particle distribution of microspheres manufactured using a bead mill using DLS (dynamic light scattering).

[0073] Figure 5 shows a TEM measurement image of microspheres manufactured using a bead mill.

[0074] Figure 6 shows a drug dissolution graph after loading a cationic drug (doxorubicin hydrochloride).

[0075] Figure 7 shows the results of measuring cell survival rate in Huh7 cells.

[0076] Figure 8 shows an optical microscope image of an emulsion of a cationic drug-loaded microsphere suspension solution and Lipiodol Ultra solution.

[0077] Figure 9 shows images of the passage through a microcatheter of each suspension using 5% sucrose and 15% mannitol as a drug-loaded aqueous suspension and a mixed emulsion with Lipiodol Ultra solution.

[0078] Figure 10 shows the appearance of microspheres manufactured in Example 8.

[0079] Figure 11 shows the appearance of doxorubicin-bound microspheres manufactured in Example 8.

[0080] Figure 12 shows the dissociation over time of the microspheres manufactured in Example 8.

[0081] Figure 13 shows the dissociation of doxorubicin-bound microspheres manufactured in Example 8 over time.

[0082] Figure 14 shows the synthesis results of CMC-DS microspheres according to the type of crosslinking agent.

[0083] Figure 15 shows the binding of doxorubicin (AD mycin) to FeCl3 or ZrOCl2 cross-linked CMC-DS microspheres.

[0084] Figure 16 shows CMC-DS microspheres with controlled size distribution.

[0085] Figure 17 shows microsphere particles formed according to the crosslinking agent concentration.

[0086] Figure 18 shows the results of confirming the stability of the W / O emulsion used in the production of microspheres.

[0087] Figures 19a and 19b show the results of microscopic observation of microspheres manufactured under 5% CMC, 0% DS conditions. Figure 19a shows microspheres with sizes of 25 to 50 and 50 to 100 um, and Figure 19b shows microspheres with sizes of 100 to 300 and 300 to 500 um.

[0088] Figures 20a to 20e show the results of microscopic observation of microspheres with different CMC:DS ratios of 100 to 300 and 300 to 500 μm in size. Figure 20a shows CMC:DS=5:1 microspheres, Figure 20b shows CMC:DS=4:1 microspheres, Figure 20c shows CMC:DS=3:1 microspheres, Figure 20d shows CMC:DS=2:1 microspheres, and Figure 20e shows CMC:DS=1:1 microspheres.

[0089] Figure 21a shows the results of checking the difference in properties before and after drug loading to confirm the drug loading ability of microspheres according to the size of the microspheres and the drug:microsphere ratio.

[0090] Figure 21b shows the results of checking the properties after drug loading to confirm the drug loading ability of microspheres according to the CMC:DS ratio.

[0091]

[0092] The microsphere according to the present invention is composed of carboxymethyl cellulose, which is an anionic polymer of the cellulose series, and dextran series with biodegradable properties, and is characterized by using a cross-linking agent to increase water stability, so that it has low toxicity and is biocompatible, and can deliver drugs to microvessels, and can obtain microspheres of a desired size, so that the size of the microspheres can be adjusted according to the size of the blood vessel so that the microspheres can directly embolize the blood vessel, and microspheres of a desired size can be provided.

[0093] Accordingly, the present invention provides a composition for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt as active ingredients.

[0094] In addition, the present invention provides a microsphere comprising an anionic polymer and a divalent metal compound salt prepared using the composition for preparing the microsphere, and according to one embodiment of the present invention, the microsphere may comprise an anionic polymer and a divalent metal compound salt, but is not limited thereto.

[0095] In the present invention, the anionic polymer may be selected from the group consisting of carboxymethylcellulose or a salt thereof; dextran sulfate or a salt thereof; and mixtures thereof, but is not limited thereto.

[0096] In the present invention, the microspheres may include, but are not limited to, carboxymethylcellulose and dextran sulfate.

[0097] In another embodiment of the present invention, the carboxymethylcellulose and dextran sulfate are mixed in a ratio of 0.5 to 20:1, 0.5 to 15:1, 0.5 to 10:1, 0.5 to 5:1, 0.5 to 4:1, 0.5 to 3:1, 0.5 to 2:1, 0.5 to 1:1, 1 to 3:1, 1.5 to 2.5:1, 1 to 5:1, 2 to 5:1, 3 to 5:1, 3.5 to 4.5:1, 4 to 5:1, 4:1, 1 to 10:1, 1 to 9:1, 3 to 9:1, 4 to 9:1, 5 to 9:1, 6 to 9 : 1, 7 to 9: 1, 8 to 9: 1, 7.5 to 8.5: 1, or 8: 1 by mass, but is not limited thereto.

[0098] In the present invention, the carboxymethylcellulose and dextran sulfate are mixed in a ratio of 0.5 to 20:1, 0.5 to 15:1, 0.5 to 10:1, 0.5 to 5:1, 0.5 to 4:1, 0.5 to 3:1, 0.5 to 2:1, 0.5 to 1:1, 1 to 3:1, 1.5 to 2.5:1, 1 to 5:1, 2 to 5:1, 3 to 5:1, 3.5 to 4.5:1, 4 to 5:1, 4:1, 1 to 10:1, 1 to 9:1, 3 to 9:1, 4 to 9:1, 5 to 9:1, 6 to 9:1, It may be included in a concentration ratio (% w / w) of 7 to 9: 1, 8 to 9: 1, 7.5 to 8.5: 1, or 8: 1, and according to one embodiment of the present invention, the carboxymethylcellulose and dextran sulfate may be included in a concentration ratio (% w / w) of 2 to 5: 1, preferably, but not limited to, a concentration ratio (% w / w) of 4: 1.

[0099] In the present invention, the carboxymethylcellulose and dextran sulfate may preferably be included in a concentration ratio of 1 to 5:1, 1 to 4:1, 1 to 3:1, 1 to 2:1, 2 to 5:1, 2 to 4:1, 2 to 3:1, 5:1, 4:1, 3:1, 2:1, 1:1, or 1:0, but is not limited thereto.

[0100] In the present invention, the multivalent metal compound salt is Zr 2+ , Zr 4+ , Fe 2+ , Fe 3+ , Ca 2+ , Al 3+ , Cu 2+ , and Zn 2+It may be one or more divalent to tetravalent metal compound salts selected from the group consisting of, and according to one embodiment of the present invention, a divalent metal compound salt, preferably, Zr 2+ More specifically, the polyvalent metal compound salt may be at least one selected from the group consisting of iron(II) chloride, iron(III) chloride, aluminum chloride, iron nitrate [Fe(NO3)3], aluminum nitrate [Al(NO3)3], iron acetate [Fe(CH3COO)3], aluminum acetate [Al(CH3COO)3], iron perchlorate [Fe(ClO4)3], zinc chloride [ZnCl2], calcium chloride [CaCl2], zirconium oxychloride [ZrOCl2·8H2O], and copper(II) nitrate trihydrate [Cu(NO3)2·3H2O], and according to one embodiment of the present invention, the polyvalent metal compound salt may be zirconium oxychloride [ZrOCl2·8H2O] or iron(III) chloride, but is not limited thereto.

[0101] According to one embodiment of the present invention, the anionic polymer is carboxymethylcellulose or a salt thereof, dextran sulfate or a salt thereof; and the polyvalent metal compound salt may be zirconium oxychloride [ZrOCl2·8H2O] or iron(III) chloride, but is not limited thereto.

[0102] In the present invention, the anionic polymer and the polyvalent metal compound salt may be included in a concentration ratio (% w / w) of 0.1 to 10:1, 0.3 to 10:1, 0.5 to 10:1, 0.6 to 10:1, 0.8 to 10:1, 1 to 10:1, 3 to 10:1, 5 to 10:1, 8 to 10:1, 0.5 to 8:1, 0.5 to 5:1, 0.5 to 3:1, 0.5 to 2.5:1, 0.6 to 5:1, 0.6 to 3:1, 0.6 to 2.5:1, 0.6 to 1.5:1, or 0.6 to 1:1, and according to one embodiment of the present invention, It may be included in a concentration ratio of 0.6 to 2.4: 1, preferably 0.6: 1, but is not limited thereto.

[0103] In the present invention, the microspheres may be loaded with a cationic drug, and the cationic drug may be loaded in a form combined with anions present inside and on the surface of the microspheres. That is, the microspheres according to the present invention can serve as drug delivery vehicles to deliver cationic drugs into the body.

[0104] The cationic drug may be at least one selected from the group consisting of doxorubicin, procainamide, digoxin, quinidine, trimethoprim, cimetidine, vancomycin, irinotecan, daunorubicin, epirubicin, diphenhydramine, memantine, oxycodone, pyrilamine, tramadol, and pharmaceutically acceptable salts thereof, and according to one embodiment of the present invention, may be doxorubicin, but is not limited thereto.

[0105] In the present invention, the cationic drug may be loaded into the microsphere at a mass ratio of cationic drug: microsphere = 1:1 to 10, 1:1 to 9, 1:1 to 8, 1:1 to 7, 1:1 to 6, 1:1 to 5, 1:1 to 4, 1:1 to 3, 1:1 to 2.5, 1:1 to 1.5, or 1:1, but is not limited thereto.

[0106] In the present invention, the cationic drug and microspheres may be included in a dry weight ratio of, but not limited to, 1:1 to 10, 1:1 to 9, 1:1 to 8, 1:1 to 7, 1:1 to 6, 1:1 to 5, 1:1 to 4, 1:1 to 3, 1:1 to 2.5, 1:1 to 1.5, or 1:1.

[0107] In the present invention, the microsphere may include an anionic polymer and a polyvalent metal compound salt, and the anionic polymer and the polyvalent metal compound salt may be mixed in a ratio of 0.1 to 10:1, 0.3 to 10:1, 0.5 to 10:1, 0.6 to 10:1, 0.8 to 10:1, 1 to 10:1, 3 to 10:1, 5 to 10:1, 8 to 10:1, 0.5 to 8:1, 0.5 to 5:1, 0.5 to 3:1, 0.5 to 2.5:1, 0.6 to 5:1, 0.6 to 3:1, 0.6 to 2.5:1, 0.6 to 1.5:1, or 0.6 to 1 The anionic polymer may be included in a concentration ratio (% w / w), and the carboxymethylcellulose and dextran sulfate may be included in a concentration ratio of 0.5 to 20:1, 0.5 to 15:1, 0.5 to 10:1, 0.5 to 5:1, 0.5 to 4:1, 0.5 to 3:1, 0.5 to 2:1, 0.5 to 1:1, 1 to 3:1, 1.5 to 2.5:1, 1 to 5:1, 2 to 5:1, 3 to 5:1, 3.5 to 4.5:1, 4 to 5:1, 4:1, 1 to 10:1, 1 to 9:1, 3 to The microspheres may be loaded with a cationic drug at a mass ratio or concentration ratio (% w / w) of 9:1, 4 to 9:1, 5 to 9:1, 6 to 9:1, 7 to 9:1, 8 to 9:1, 7.5 to 8.5:1, or 8:1. In addition, the microspheres may be loaded with a cationic drug, and the cationic drug may be cationic drug: microsphere = 1:1 to 10, 1:1 to 9, 1:1 to 8, 1:1 to 7, 1:1 to 6, 1:1 to 5, 1:1 to 4, 1:1 to 3, 1:1 to 2.5, 1:1 to 1.It can be loaded at a mass ratio of 5, or 1:1, but is not limited thereto.

[0108] In the present invention, the composition may be an emulsion formulation, and the emulsion formulation in the present invention may be prepared by mixing an aqueous phase to which an anionic polymer and a nonionic surfactant are added and an oil phase to which a nonionic surfactant is added using a homogenizer, but is not limited thereto.

[0109] In the present invention, the microspheres have a size of 10 to 700 μm, 10 to 600 μm, 10 to 500 μm, 10 to 400 μm, 10 to 300 μm, 10 to 200 μm, 10 to 100 μm, 10 to 50 μm, 20 to 700 μm, 20 to 600 μm, 20 to 500 μm, 20 to 400 μm, 20 to 300 μm, 20 to 200 μm, 20 to 100 μm, 20 to 50 μm, 25 to 700 μm, 25 to 600 μm, 25 to 500 μm, 25 to 400 μm, 25 to 300 μm, 25 to It may have a diameter of, but is not limited to, 200 μm, 25 to 100 μm, or 25 to 50 μm.

[0110] In the present invention, the microspheres have a size distribution as follows, so that the microspheres according to the present invention can be classified (separated) according to the diameter range as follows, that is, the size distribution, and microspheres of a specific size can be separated and used according to the intended use:

[0111] a) 10 to 50 μm, preferably 25 to 50 μm;

[0112] b) 50 to 100 μm;

[0113] c) 100 to 300 μm;

[0114] d) 300 to 500 μm; and

[0115] e) 500 to 700 μm, preferably 500 μm or more.

[0116] In the present invention, the microspheres can be used not only as drug delivery vehicles capable of delivering anticancer agents into the body, but also directly block blood vessels to block blood flow, and thus can be used for cancer treatment and / or vascular embolization, but are not limited thereto. Since the microspheres of the present invention can be separated according to particle size, vascular embolization can be performed by isolating microspheres appropriate for the size of the blood vessels at the site where vascular embolization is to be performed.

[0117] Accordingly, the present invention can provide a pharmaceutical composition for treating cancer comprising the microspheres and a cationic drug as active ingredients, or a composition for delivering a cationic drug comprising the microspheres and a cationic drug as active ingredients.

[0118] The above-mentioned angioembolization is called transarterial chemoembolization (TACE). This method treats cancer by injecting anticancer drugs into the arteries leading to the cancerous tissue and using embolic substances to block the supply of nutrients to the cancerous tissue. Furthermore, the above-mentioned angioembolization can be a treatment method in which a substance is injected into a blood vessel to stop blood flow to a specific blood vessel, thereby blocking blood flow through the vessel, in cases where the patient is suffering from excessive bleeding, vascular malformation, or tumor.

[0119] In the present invention, the cancer includes angiogenesis-dependent cancer, preferably at least one selected from the group consisting of liver cancer, ovarian cancer, breast cancer, and non-small cell lung cancer, and more preferably liver cancer.

[0120] In the present invention, by combining a cationic drug with the microspheres and then mixing them with iodized oil, which is an embolic agent, a stable emulsion can be formed, and the embolic effect can be increased by injecting the emulsion into the hepatic artery, so the composition of the present invention can further include an embolic agent, and the embolic agent can be iodized oil, but is not limited thereto.

[0121] The above iodized oil may be at least one selected from the group consisting of, but is not limited to, iodized oil derived from poppy seeds, iodized oil derived from soybeans, and ethiodol. The above iodized oil derived from poppy seeds may be used under the product name Lipiodol, but is not limited thereto.

[0122] The composition of the present invention is characterized in that it is administered within 0 to 2 hours, preferably 0 to 1 hour, after preparation, and has excellent stability as the movement of the drug within the microspheres is maintained low for a long period of time compared to the composition used in conventional chemoembolization.

[0123] In the present invention, the composition containing the microspheres and the cationic drug and the embolic material can be mixed using a mixing method using a 3-way cock, a method using ultrasound, etc., but is not limited thereto.

[0124] Additionally, the present invention provides a method for preventing or treating vascular embolism and cancer, comprising a step of administering the composition to a subject.

[0125] The content of the microspheres and cationic anticancer agent in the composition of the present invention can be appropriately adjusted depending on the symptoms of the disease, the degree of progression of the symptoms, the condition of the patient, etc., and may be, for example, 0.0001 to 99.9 wt% or 0.001 to 50 wt% based on the total weight of the composition, but is not limited thereto. The content ratio is a value based on the dry amount after removing the solvent.

[0126] In addition, the present invention provides microspheres comprising an anionic polymer and a polyvalent metal compound salt.

[0127] Additionally, the present invention provides a drug delivery use of the microspheres.

[0128] The present invention also provides the use of the microspheres for the manufacture of drug delivery vehicles.

[0129] In addition, the present invention provides a composition for cationic drug delivery comprising the above microspheres and a cationic drug as active ingredients.

[0130] Additionally, the present invention provides a cationic drug delivery method comprising a step of administering the microspheres encapsulated with a cationic drug to a subject in need thereof.

[0131] In addition, the present invention provides a composition comprising the above microspheres and a cationic drug as active ingredients for use in treating vascular embolism and / or cancer.

[0132] In addition, the present invention provides a composition for treating cancer comprising the microspheres and a cationic drug as active ingredients.

[0133] In addition, the present invention provides a method for treating vascular embolism and cancer, comprising administering to a subject a composition comprising the above microspheres and a cationic drug as active ingredients.

[0134] In addition, the present invention provides a use of a composition comprising the above microspheres and a cationic drug as active ingredients for the manufacture of a vascular embolization and cancer treatment agent.

[0135] The pharmaceutical composition according to the present invention may further comprise suitable carriers, excipients, and diluents commonly used in the manufacture of pharmaceutical compositions. The excipients may be, for example, one or more selected from the group consisting of diluents, binders, disintegrants, lubricants, adsorbents, moisturizers, film-coating materials, and controlled-release additives.

[0136] Carriers, excipients and diluents that may be included in the pharmaceutical composition according to the present invention include lactose, dextrose, sucrose, oligosaccharides, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

[0137] When formulating, it is usually prepared using diluents or excipients such as fillers, bulking agents, binders, wetting agents, disintegrants, and surfactants.

[0138] As additives of the liquid formulation according to the present invention, water, dilute hydrochloric acid, dilute sulfuric acid, sodium citrate, monostearate sucrose, polyoxyethylene sorbitol fatty acid esters (twin esters), polyoxyethylene monoalkyl ethers, lanolin ethers, lanolin esters, acetic acid, hydrochloric acid, ammonia water, ammonium carbonate, potassium hydroxide, sodium hydroxide, prolamine, polyvinylpyrrolidone, ethylcellulose, sodium carboxymethylcellulose, etc. can be used.

[0139] The syrup according to the present invention may include a solution of white sugar, other sugars, or sweeteners, and may also include a fragrance, a coloring agent, a preservative, a stabilizer, a suspending agent, an emulsifier, a viscosity modifier, and the like, as needed.

[0140] Purified water may be used in the emulsion according to the present invention, and emulsifiers, preservatives, stabilizers, fragrances, etc. may be used as needed.

[0141] The suspension according to the present invention may include suspending agents such as acacia, tragacanth, methylcellulose, carboxymethylcellulose, sodium carboxymethylcellulose, microcrystalline cellulose, sodium alginate, hydroxypropylmethylcellulose, HPMC 1828, HPMC 2906, and HPMC 2910, and, if necessary, surfactants, preservatives, stabilizers, colorants, and fragrances may be used.

[0142] The injection according to the present invention includes a solvent such as distilled water for injection, 0.9% sodium chloride injection, Ringer's injection, dextrose injection, dextrose + sodium chloride injection, PEG, lactated Ringer's injection, ethanol, propylene glycol, non-volatile oils - sesame oil, cottonseed oil, peanut oil, soybean oil, corn oil, ethyl oleate, isopropyl myristate, and benzene benzoate; a solubilizing agent such as sodium benzoate, sodium salicylate, sodium acetate, urea, urethane, monoethylacetamide, butazolidine, propylene glycol, tween, nitrile acid amide, hexamine, and dimethylacetamide; a buffer such as weak acids and their salts (acetic acid and sodium acetate), weak bases and their salts (ammonia and ammonium acetate), organic compounds, proteins, albumin, peptone, and gums; It may include isotonic agents such as sodium chloride; stabilizers such as sodium bisulfite (NaHSO3), carbon dioxide gas, sodium metabisulfite (Na2S2O3), sodium sulfite (Na2SO3), nitrogen gas (N2), and ethylenediaminetetraacetic acid; oxidizing agents such as sodium bisulfite 0.1%, sodium formaldehyde sulfoxylate, thiourea, disodium ethylenediaminetetraacetic acid, and acetone sodium bisulfite; analgesics such as benzyl alcohol, chlorobutanol, procaine hydrochloride, glucose, and calcium gluconate; and suspending agents such as sodium cis-methylenediamine, sodium alginate, Tween 80, and aluminum monostearate.

[0143] The pharmaceutical composition according to the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment. The effective dosage level can be determined based on factors including the type and severity of the patient's disease, drug activity, drug sensitivity, administration time, administration route and excretion rate, treatment period, concurrently used drugs, and other factors well known in the medical field.

[0144] The pharmaceutical composition according to the present invention can be administered as an individual therapeutic agent or in combination with other therapeutic agents. It can be administered sequentially or simultaneously with conventional therapeutic agents, or in single or multiple doses. Taking all of the above factors into account, it is important to administer an amount that achieves maximum efficacy with minimal side effects. This amount can be readily determined by those skilled in the art to which the present invention pertains.

[0145] The pharmaceutical composition of the present invention is determined according to the type of drug as an active ingredient along with various related factors such as the disease to be treated, route of administration, age, sex, weight of the patient, and severity of the disease.

[0146] In the present invention, the term "subject" means a subject requiring treatment for a disease, and more specifically, may be a mammal such as a human or non-human primate, mouse, rat, dog, cat, horse, and cow, but is not limited thereto.

[0147] In the present invention, “administration” means providing a predetermined composition of the present invention to a subject by any appropriate method.

[0148] In the present invention, “prevention” means any action that suppresses or delays the onset of a target disease, “treatment” means any action that improves or beneficially changes a target disease and its metabolic abnormality symptoms by administering a pharmaceutical composition according to the present invention, and “improvement” means any action that reduces a parameter related to a target disease, for example, the severity of a symptom, by administering a composition according to the present invention.

[0149]

[0150] The present invention also provides a kit for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt. The kit may further include instructions describing a method for producing the microspheres of the present invention.

[0151] In addition, the present invention provides a drug delivery kit comprising the microspheres; and / or a cancer prevention or treatment kit comprising microspheres encapsulating an anticancer agent.

[0152] In the present invention, the "kit" is not limited in specific form or type, and a kit of a form commonly used in the art may be used, and in addition to the microspheres of the present invention, any component necessary to achieve the purpose of the present invention may be further included. For example, in addition to the microspheres of the present invention, a storage solution, an administration tool, a drug, an administration suspension, etc. may be additionally included, and an instruction manual containing information regarding the microspheres of the present invention may be additionally included.

[0153]

[0154] In addition, the present invention provides a method for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt, comprising the following steps:

[0155] (S1) A step of forming an emulsion by mixing an aqueous phase containing an anionic polymer and an oil phase containing a nonionic surfactant using a homogenizer;

[0156] (S2) a step of forming microspheres by adding the above emulsion to a multivalent metal compound salt aqueous solution; and

[0157] (S3) A step of separating microspheres of a desired size using a sieve.

[0158] Matters concerning the above composition can be applied to the present manufacturing method.

[0159] The above step (S1) may include, but is not limited to, one or more selected from the group consisting of steps a) to c).

[0160] The above step (S2) may include, but is not limited to, steps d) and / or e).

[0161] The above step (S3) may include, but is not limited to, the following step f).

[0162] The method for manufacturing the above microspheres may include, but is not limited to, a step of stirring between each step so that each component is uniformly mixed.

[0163] In the present invention, the emulsion may be in a W / O form, but is not limited thereto.

[0164] The above (W / O) emulsion may have a (volume) ratio of the water phase to the oil phase of, but is not limited to, 1:1 to 20, 1:1 to 15, 1:1 to 10, 1:1 to 8, 1:1 to 5, 1:1 to 4, 1:1 to 3, 1:1 to 2, 1:3 to 15, 1:3 to 10, 1:3 to 8, 1:3 to 5, 1:3 to 4, 1:4 to 10, 1:4 to 8, 1:4 to 5, 1:20, 1:15, 1:10, 1:9, 1:8, 1:7, 1:6, 1:5, 1:4, 1:3, 1:2, or 1:1. In the present invention, the higher the ratio of the oil phase, the better the emulsion formation can be achieved, but the optimal ratio for using the oil phase to a minimum may preferably be 1:4.

[0165] In the present invention, in the step (S2), the emulsion may be added to the polyvalent metal compound salt aqueous solution at a volume ratio of 1:1 to 20, 1:1 to 15, 1:1 to 10, 1:1 to 7.5, 1:1 to 5, 1:1 to 2.5, 1:1 to 2, 1:2 to 15, 1:2 to 10, 1:2 to 7.5, 1:2 to 5, 1:6 to 10, 1:6 to 9, 1:6 to 7.5, 1:7.5 to 9, 1:20, 1:15, 1:10, 1:9, 1:8, 1:7.5, 1:7, 1:6, 1:5, 1:4, 1:3, 1:2, or 1:1, and according to one embodiment of the present invention, It can be added in a volume ratio of 1:7.5, but is not limited thereto.

[0166] According to one embodiment of the present invention, it was confirmed that the size distribution of the microspheres produced in step (S1) can be controlled by adjusting the rpm of the homogenizer that mixes the water phase and the oil phase. Therefore, the microsphere production method of the present invention is characterized by being able to control the particle distribution of the microspheres by adjusting the rpm of the homogenizer.

[0167] According to one embodiment of the present invention, the microspheres separated using a 500, 300, 100, 53, and / or 25 μm sieve in the step (S3) may have a size distribution as follows, but is not limited thereto, and the size distribution may change depending on the type of sieve used.

[0168] a) 10 to 50 μm, preferably 25 to 50 μm;

[0169] b) 50 to 100 μm;

[0170] c) 100 to 300 μm;

[0171] d) 300 to 500 μm; and

[0172] e) 500 to 700 μm, preferably 500 μm or more.

[0173] In the present invention, the method may further include a step of purifying and washing the separated microspheres using a sieve, but is not limited thereto.

[0174] In the present invention, purification of the microspheres may be performed using a centrifuge or a filter, but is not limited thereto.

[0175] In the present invention, the washing may be selected from the same organic solvent or the group of solvents mentioned above, but is not limited thereto.

[0176] In the present invention, the washing may be performed using, but is not limited to, an aqueous solvent selected from the group consisting of distilled water, water for injection, sucrose, mannitol, sodium chloride, etc. dissolved therein after the organic solvent washing.

[0177] The method for manufacturing microspheres of the present invention may specifically include one or more of the following steps, or may consist of the following steps, but is not limited thereto:

[0178] a) a step of dissolving one or more anionic polymers and nonionic surfactants in the water;

[0179] b) a step of dissolving a nonionic surfactant in the oil phase;

[0180] c) a step of forming an emulsion by stirring the water phase of a) and the oil phase of b) using a homogenizer;

[0181] d) A step of forming microspheres by adding the emulsion of step c) to a crosslinking agent aqueous solution (aqueous solution of a polyvalent metal compound salt) in which a crosslinking agent (aqueous solution of a polyvalent metal compound salt) is dissolved in an aqueous ethanol solution.

[0182] e) Additional stirring step in the above step d),

[0183] f) a step of separating microspheres according to particle size using a 500, 300, 100, 53, and / or 25 μm sieve in the solution of step e);

[0184] g) a step of separating and / or removing residual organic solvent from the microspheres using an aqueous solution;

[0185] h) Step of removing moisture by placing it in the oven.

[0186] The above steps a) to c) may be performed in any suitable order.

[0187] Additionally, the above steps are not necessarily limited to the order described above.

[0188] In the present invention, the water may be deionized water or distilled water, but is not limited thereto.

[0189] In the present invention, the oil phase may be selected from the group consisting of mineral oil, vegetable oil, medium-chain triglyceride (MCT) and mixtures thereof, and according to one embodiment of the present invention, the oil phase may be medium-chain triglyceride, but is not limited thereto.

[0190] In the present invention, the mineral oil is also called mineral oil, and is a by-product produced in the process of refining crude oil, and is an oil whose main components are alkane and paraffin.

[0191] In the present invention, the vegetable oil may be an oil derived from nuts, seeds, grains, etc., and may include, for example, peanut oil, soybean oil, coconut oil, or olive oil.

[0192] In the present invention, the medium-chain triglyceride is a representative alternative fat produced by hydrolyzing palm oil and palm oil, fractionating caprylic acid and capric acid, and then esterifying (or combining) these with glycerol. It is a medium-chain fatty acid with a small molecular structure, usually 8 to 10 carbon atoms.

[0193] In the present invention, the nonionic surfactant may be selected from the group consisting of polyvinyl alcohol, polyalkylene glycol, polyvinyl pyrrolidone, alkyl polyglycoside, t-octylphenoxy polyethoxy ethanol, poloxamer, polysorbate (preferably polysorbate 20 (Tween 20)), poly(N-vinylacetamide), polyvinyl butyral, sorbitan oleate (Span 80), and mixtures thereof, and according to one embodiment of the present invention, the nonionic surfactant may be selected from the group consisting of polysorbate 20 and / or sorbitan The nonionic surfactant that can be dissolved in the water phase with the anionic polymer can be polysorbate 20, and the nonionic surfactant that can be dissolved in the oil phase can be sorbitan oleate, but is not limited thereto.

[0194] In the present invention, the polyalkylene glycol may be any of the conventional polyalkylene glycol series known in the art without limitation, but it is preferable to use a polyethylene glycol series.

[0195] In the present invention, the poloxamer is a nonionic triblock copolymer in which hydrophilic ethylene oxide is bonded to both ends of a hydrophobic propylene oxide, and has the characteristic of being temperature sensitive, so that it can be converted into a sol and a gel depending on the concentration and temperature, and the properties of the poloxamer change depending on the ratio of polyoxypropylene and polyoxyethylene. The above poloxamer is poloxamer 101, poloxamer 105, poloxamer 105 benzoate, poloxamer 108, poloxamer 122, poloxamer 123, poloxamer 124, poloxamer 181, poloxamer 182, poloxamer 182 dibenzoate, poloxamer 183, poloxamer 184, poloxamer 185, poloxamer 188, poloxamer 212, poloxamer 215, poloxamer 217, poloxamer 231, poloxamer 234, poloxamer 235, poloxamer 237, poloxamer 238, poloxamer 282, poloxamer 284, poloxamer 288, It may be selected from the group consisting of, but is not limited to, poloxamer 331, poloxamer 333, poloxamer 334, poloxamer 335, poloxamer 338, poloxamer 401, poloxamer 402, poloxamer 403, and poloxamer 407.

[0196] In the present invention, the alkyl polyglycoside is, for example, C8- 22 Alkyl glycosides, more specifically, but not limited to, may be selected from the group consisting of caprylyl glucoside, coco-glucoside, decyl glucoside, lauryl glucoside, methyl coco glucoside, and myristyl glucoside.

[0197] In the present invention, the organic solvent may be at least one selected from the group consisting of alkyl alcohols, ketones, acetonitrile, tetrahydrofuran, and mixtures thereof, but is not limited thereto.

[0198] The above alkyl alcohols may be alcohols having 1 to 6 carbon atoms, and examples thereof include, but are not limited to, methanol, ethanol, alcohol, propanol, butanol, pentanol, and hexanol.

[0199] The above ketones may be organic solvents having the chemical formula of RCOR, and may include, but are not limited to, acetone, methyl ethyl ketone, methyl butyl ketone, cyclohexanone, methyl isobutyl ketone, diisobutyl ketone, etc.

[0200] In addition, the present invention provides a method for preparing a composition for treating vascular embolism and cancer, comprising the following steps:

[0201] A step of forming an emulsion by mixing an aqueous phase containing an anionic polymer and an oil phase containing a nonionic surfactant using a homogenizer;

[0202] A step of forming microspheres by adding the above emulsion to a multivalent metal compound salt aqueous solution;

[0203] A step of separating microspheres of a desired size using a sieve; and

[0204] A step of mixing a cationic drug with the above microspheres.

[0205] The step of mixing the cationic drug with the microspheres may be performed by ultrasonic treatment, but is not limited thereto.

[0206] The method for preparing the composition for vascular embolism and cancer treatment may further include, but is not limited to, a step of mixing the mixed composition of the microspheres and the cationic drug with an embolic material.

[0207] The mixing step with the above-mentioned coloring matter can be performed using a mixing method using a 3-way cock, a method using ultrasound, etc., but is not limited thereto.

[0208] The term "ultrasound" used in the present invention generally refers to sound waves exceeding the frequency of sound waves that can be heard by the human ear, which is 16 Hz to 20 kHz, and high-intensity focused ultrasound introduces focused ultrasound that provides continuous, high-intensity ultrasonic energy to a focus, and can produce instantaneous thermal effects (65-100℃), cavitation effects, mechanical effects, and sonochemical effects depending on the energy and frequency. Ultrasound is not harmful when passing through human tissue, but high-intensity ultrasound that forms a focus generates enough energy to cause coagulation necrosis and thermocautery effects regardless of the type of tissue.

[0209] In the present invention, the ultrasonic waves refer to sound waves having a frequency higher than the audible frequency range of 16 Hz to 20 kHz.

[0210] The present invention also provides a method for preparing a cationic drug delivery composition comprising the following steps:

[0211] A step of forming an emulsion by mixing an aqueous phase containing an anionic polymer and an oil phase containing a nonionic surfactant using a homogenizer;

[0212] A step of forming microspheres by adding the above emulsion to a multivalent metal compound salt aqueous solution;

[0213] A step of separating microspheres of a desired size using a sieve; and

[0214] A step of mixing a cationic drug with the above microspheres.

[0215]

[0216] In this specification, “active ingredient” means an ingredient that exhibits the intended activity alone or can exhibit the intended activity together with a carrier or the like that is inactive in itself.

[0217] Throughout the specification of the present invention, when a part is said to "include" a certain component, unless specifically stated otherwise, this does not exclude other components, but rather means that other components may be included. The terms "about," "substantially," and the like, as used throughout the specification of the present invention, are used in a meaning at or near the numerical value when manufacturing and material tolerances inherent to the meaning stated are presented, and are used to prevent unscrupulous infringers from unfairly exploiting disclosures that state precise or absolute values ​​to aid understanding of the present invention.

[0218] Throughout the specification of the present invention, the term "combination thereof" included in the expressions in the Makushi format means a mixture or combination of one or more selected from the group consisting of the components described in the expressions in the Makushi format, and means including one or more selected from the group consisting of the components.

[0219] The use of the term "above" and similar referential terms in this specification (especially in the claims) may refer to both the singular and the plural. Furthermore, when a range is described, it is intended to include individual values ​​within the range (unless otherwise stated), and is equivalent to describing each individual value constituting the range in the detailed description. Finally, unless the order of the steps constituting the method is explicitly stated or otherwise stated to the contrary, the steps may be performed in any suitable order. The present invention is not necessarily limited to the order in which the steps are described. The use of any examples or exemplary terms (e.g., "for example," etc.) is merely intended to further illustrate the technical idea and is not intended to limit the scope of the invention unless otherwise defined by the claims. Furthermore, those skilled in the art will recognize that various modifications, combinations, and variations may be made within the scope of the appended claims or their equivalents, depending on design conditions and factors.

[0220] While terms such as "first" and "second" may be used to describe various components, these components should not be limited by these terms. These terms are used solely to distinguish one component from another. For example, without departing from the scope of the present invention, a first component may be referred to as a "second component," and similarly, a second component may also be referred to as a "first component." The term "and / or" includes a combination of multiple related items described herein or any of multiple related items described herein.

[0221] The present invention is susceptible to various modifications and embodiments. Specific embodiments are illustrated in the drawings and described in detail in the detailed description. However, this is not intended to limit the present invention to specific embodiments, but rather to encompass all modifications, equivalents, and alternatives falling within the spirit and technical scope of the present invention. In describing the present invention, detailed descriptions of related known technologies will be omitted if they are deemed to obscure the gist of the present invention.

[0222]

[0223]

[0224] Hereinafter, preferred examples are presented to aid in understanding the present invention. However, the following examples are provided solely to facilitate a better understanding of the present invention, and the scope of the present invention is not limited by the following examples.

[0225]

[0226] [Example]

[0227] Example 1. Test to confirm the reactivity between polymer and crosslinking agent

[0228] To compare the reactivity between anionic polymers and calcium chloride, the reactivity was tested by adding calcium chloride in the presence of CMC (Carboxymethyl cellulose).

[0229]

[0230] Example 1-1. Reactivity Test 1

[0231] 2 mL of 0.2% CMC and 2 mL of 0.2% carboxymethyl dextran sodium (hereinafter referred to as CMD) were added to a 20 mL container, and 0.5 mL of 10% calcium chloride was added while stirring. No solids were formed after checking for 1 hour.

[0232]

[0233] Example 1-2. Reactivity Test 2

[0234] The reactivity was confirmed by increasing the concentration and conducting a test in the same manner as <Reactivity Test 1> for each composition in Table 1 below.

[0235] As shown in Figure 1, there was no change in the case of composition 1, and compositions 2 and 3 were changed into suspensions.

[0236] In addition, as shown in Fig. 2, the optical microscope analysis image of the reaction solid of composition 2 of Table 1 confirmed that it was gelled and not granulated.

[0237] According to the above results, it was confirmed that calcium chloride is not suitable as a cross-linking agent for producing nanoparticles.

[0238]

[0239]

[0240] Example 2. Preparation of microparticles using W / O emulsion

[0241] According to the composition ratios in Table 2 below, CMC, iron chloride (FeCl3) solution, and DS (dextran sulfate) powder were prepared, respectively.

[0242] First, DS powder was dissolved in CMC solution, mixed with 1% polyvinyl alcohol (PVA) solution at room temperature, and then added dropwise to medium chain triglyceride (MCT) oil used as the oil phase, and an emulsion was prepared using a sonicator. FeCl3 was added dropwise to the emulsion and further stirred to prepare particles. To obtain the formed particles, centrifugation was performed at 4°C and 15,000 rpm for 10 minutes, and insoluble matter and MCT oil were removed using ethanol. Ethanol and purified water were sequentially separated using a centrifuge under the same conditions, and finally redispersed in 2 mL of purified water.

[0243]

[0244] As a result, as the CMC concentration increased and the sonication time increased, particle formation was well achieved.

[0245]

[0246] Example 3. Confirmation of yield according to microparticle production using W / O emulsion

[0247] For scale-up, a homogenizer was used instead of an ultrasonic disperser. Using a homogenizer (rpm: 17,000), particles were manufactured at room temperature using the same manufacturing method according to the composition ratios shown in Table 3 below, and the yields for each composition ratio were compared. Finally, each redispersed suspension was freeze-dried, and the mass was measured after drying.

[0248] As shown in Fig. 3, through the image of the solid produced through composition 2 of Table 3, it was found that particles were formed when iron chloride was used rather than when calcium chloride was used.

[0249] As shown in Table 3, it was confirmed that the highest yield was obtained when CMC and DS were mixed at a mass ratio of 2:1.

[0250]

[0251]

[0252] Example 4. Manufacturing of microspheres through additional nanoprocessing after manufacturing microparticles using W / O emulsion.

[0253] Microparticles were manufactured using the following method. To prepare the aqueous solution, 2 g of PVA and 1 g of CMC were each dissolved in 100 mL of distilled water. Then, 0.5 g of DS was measured, added to the CMC solution, and dissolved. To prepare the cross-linking agent solution, 0.5 g of iron(III) chloride (FeCl3) was dissolved in 100 mL of distilled water.

[0254] To prepare a water-on-water emulsion, 800 mL of MCT oil was added to a 2-L beaker, and the entire water phase solution was added while stirring at 17,000 rpm at room temperature using a homogenizer. After stirring for 10 minutes under the same conditions, the cross-linking agent solution was added dropwise and stirred for an additional approximately 30 minutes.

[0255] Microparticles were isolated using the following experimental method: The reaction solution was transferred to a separatory funnel and allowed to stand for 2 hours. The lower layer (aqueous phase) containing yellow solids in the separatory funnel was collected and centrifuged to generate a yellow particle pellet. The supernatant was discarded, and the white material was dissolved and removed using ethanol. Care was taken not to affect the lower layer pellet. 15 mL of ethanol was added, and the pellet was redispersed using a vortexer and an ultrasonic disperser. This process was repeated until no clumps were visible. Then, the washing process was sequentially repeated with distilled water and sucrose (5%) solution in addition to ethanol to obtain microparticles. Finally, the particles were redispersed in a 5% sucrose solution.

[0256] A bead mill (Minicer, Netzsch) was used to manufacture microspheres. The washed particles were diluted in a 5% sucrose solution to manufacture microspheres in an amount of approximately 500 mL. The bead mill was milled using 0.2 mm beads at a pump speed of 20 rpm, a flow rate of 5 m / s, and approximately 10°C to produce small particles. After replacing the beads with 0.1 mm beads, additional milling was performed at a pump speed of 10 rpm, a flow rate of 4 m / s, and the same temperature.

[0257] As shown in Fig. 4, the DLS (dynamic light scattering) measurement results showed that the particle distribution was 50 nm to 460 nm.

[0258] Additionally, as shown in Fig. 5, it was confirmed that the particles were well formed into a spherical shape.

[0259]

[0260] Example 5. Preparation and dissolution test of microspheres loaded with doxorubicin hydrochloride

[0261] Doxorubicin hydrochloride was completely dissolved in a 5% sucrose solution at a concentration of 25 mg / mL. The completely dissolved doxorubicin hydrochloride sucrose solution was added to 1.25 mL of a 5% sucrose solution containing 60 mg of microspheres dispersed therein to adjust the ratio of doxorubicin hydrochloride: particles = 25 mg: 60 mg, and the mixture was sonicated (bath type sonication) for an initial 1 minute, followed by additional mixing for approximately 30 minutes.

[0262] 500 μL of the above doxorubicin hydrochloride-loaded microsphere suspension was added to a Cellu Sep membrane (MW: 15,000, Width: 32 mm, Wall thickness: 24 μm) and both sides were closed with clips. 50 mL of buffer (PVA 0.4%, Tween 0.1% in lactic acid pH=2.7, 37°C) was placed in a vial, stirred at 200 rpm, and the temperature was increased to 37°C. The membrane sample was added and a dissolution test was performed. The absorbance of samples taken at each point was measured using a UV spectrometer (wavelength: 475 nm) to confirm the amount of dissolution.

[0263] As a result, as shown in Fig. 6, the doxorubicin release rate was found to be excellent, at approximately 70%.

[0264]

[0265] Example 6. In vitro cytotoxicity evaluation

[0266] To evaluate the toxicity of the manufactured microspheres, a viability assay was performed in Huh7 hepatoma cells. Three groups were evaluated: doxorubicin hydrochloride (dox), doxorubicin-loaded particles (032d), and non-doxorubicin-loaded particles (032).

[0267] As shown in Figure 7, it was confirmed that particles loaded with the same drug as doxorubicin hydrochloride itself exhibited a cancer cell killing effect in a concentration-dependent manner, whereas particles not loaded with the drug showed no killing effect regardless of concentration and thus no toxicity.

[0268]

[0269] Example 7. Preparation of a mixed emulsion with Lipiodol and confirmation of passage through a microcatheter.

[0270] A vial containing 50 mg of doxorubicin hydrochloride (Ildong Adriamycin RDF) was filled with 1.25 mL of Omnipaq 300 (GE Healthcare) to dissolve the drug. The entire amount was taken up using a syringe, and the solution was added to a solution containing 60 mg of microspheres dispersed in 2 mL of a 5% sucrose solution to prepare a suspension in which the drug was loaded onto nanoparticles. In addition, 10 mL of Lipiodol Ultra solution (Guerbet Korea) was prepared in a separate syringe. The two solutions were placed in a 3-way cock and the syringes were alternately pushed to prepare an emulsion. An image of the formed emulsion is shown in Fig. 8.

[0271] A microcatheter was connected to the last end of a three-way stopcock containing the emulsion prepared above, and the syringe was pressed to pass the solution. A microcatheter passage test was performed to compare the results with an emulsion prepared by replacing the 5% sucrose solution with a 15% mannitol solution using the same method.

[0272] As a result, as shown in Fig. 9, in both cases, the microcatheter was passed through.

[0273] Therefore, it was confirmed that the emulsion prepared by the method of the present invention can be used for embolization by passing through a microcatheter.

[0274]

[0275] Example 8. Preparation of microspheres (I) by combining CMC (Carboxymethylcellulose), DS (dextran sulfate), and doxorubicin.

[0276] CMC-DS microspheres were prepared by an ionic cross-linking method. First, 5 g of CMC and 1.25 g of DS were dissolved in 95 g of deionized water. The CMC and DS solutions were stirred at room temperature for 2 hours. As a cross-linking agent, 2.5 g of zirconium oxychloride octahydrate (ZrOCl2·8H2O) was completely dissolved in 100 g of deionized water (DI) at room temperature. The CMC and DS mixture was slowly added dropwise to the ZrOCl2·8H2O solution. The CMC and DS microspheres were cross-linked for 2 hours and washed with deionized water. The appearance of the CMC and DS microparticles was analyzed under a microscope. As a result, transparent microspheres were observed, as shown in Figure 10. To bind doxorubicin hydrochloride to the microspheres, a doxorubicin hydrochloride solution (Drug product) was mixed with the microspheres (N=13) at a concentration of 2 mg / mL and stored overnight at room temperature. The appearance of the doxorubicin-bound microspheres was observed under a microscope. As a result, as shown in Figure 11, the doxorubicin-bound microspheres were confirmed to have turned red.

[0277]

[0278] Example 9. Dissociation of microspheres and doxorubicin-bound microspheres

[0279] Empty microspheres and doxorubicin-conjugated microspheres were dissociated in phosphate-buffered saline (PBS, pH 7.4). The microspheres were each stored in a vial containing 100 mL of deionized water, and the vials were stored at room temperature for 7 days to observe the dissociation of the microspheres.

[0280] As a result, as shown in FIGS. 12 and 13, it was confirmed that both empty microspheres and doxorubicin-bound microspheres dissociated over time.

[0281]

[0282] Example 10. Synthesis of CMC-DS microspheres dependent on cross-linking agent.

[0283] CMC-DS microspheres were synthesized using various cross-linking agents. The method for preparing the microspheres followed the procedure described in Example 8, except for the cross-linking agent solution. All cross-linking experiments were performed independently.

[0284] First, 5 g of CMC and 5 g of DS were dissolved in 100 mL of deionized water. 1 g of ZnCl, 0.834 g of FeCl (0.5%), 0.834 g of FeCl (0.5%), 2.5 g of CaCl, and 2.5 g of ZrOCl2·8H2O were dissolved in 100 mL of deionized water, respectively. As a crosslinking agent, ZnCl2, FeCl 2, FeCl3, CaCl2, and ZrOCl2·8H2O were applied to CMC-DS microspheres, respectively. The CMC-DS mixture was added dropwise to each cross-linking agent and stirred for 2 h. After cross-linking, the CMC-DS microspheres were washed with deionized water. The appearance of each CMC-DS microsphere was observed under a microscope.

[0285] As a result, as shown in Fig. 14, FeCl3 and ZrOCl2·8H2O were effectively bound to DS through electrostatic bonding. On the other hand, ZnCl2 and FeCl 2,CaCl2 was not effective in forming the CMC-DS microsphere surface, so the CMC-DS microspheres were not maintained and were separated or all merged.

[0286]

[0287] Example 11. Doxorubicin binding to CMC-DS microspheres cross-linked with FeCl3 or ZrOCl2

[0288] To test the binding of AD mycin (doxorubicin hydrochloride) to CMC-DS microspheres, FeCl3 or ZrOCl2 cross-linked CMC-DS microspheres were evaluated. 1 mL of AD mycin (2 mg / mL) was added to each microsphere and incubated for 6 h. AD mycin binding was confirmed by a change in apparent color.

[0289] As shown in Fig. 15, AD mycin was bound to both FeCl3-crosslinked CMC-DS microspheres and ZrOCl2-crosslinked CMC-DS microspheres, and it was confirmed that more than 90% of AD mycin was loaded into the microspheres.

[0290]

[0291] Example 12. Size distribution control

[0292] CMC-DS microspheres were fabricated by controlling the size distribution. The production of CMC-DS microspheres for size distribution control was accomplished through emulsification and filtration. First, 2% CMC and 0.5% DS were dissolved in water. The CMC and DS mixture was added dropwise to MCT oil containing 4% SPAN 80. The aqueous phase (W) and oil phase (O) ratio was 25%, and the mixture was homogenized at 3000 rpm for 10 min. This CMC-DS emulsion (W / O) was further added dropwise to a 2.5% solution of ZrOCl2·8H2O, a cross-linking agent, at a rate of 3 mL / min. The cross-linking process was performed by stirring at 200 rpm. The CMC-DS microspheres were sequentially filtered through sieves with pore sizes of 500 μm, 300 μm, 100 μm, and 53 μm. The microspheres filtered through each sieve were optically visualized and shown in Figure 16.

[0293]

[0294] Example 13. Optimization of microsphere formulation

[0295] In the process of optimizing the formulation of microspheres, the raw materials used were as shown in Table 4, the devices used were as shown in Table 5, and the analytical instruments used were as shown in Table 6.

[0296]

[0297]

[0298]

[0299]

[0300] Example 13-1. Optimization of ZrOCl2 cross-linking agent concentration

[0301] As shown in Table 7, microspheres were manufactured by adding the aqueous phase dropwise to the ZrOCl2 cross-linking agent at concentrations of 2.5, 5, and 10% (w / w) using a syringe (using a 26G syringe needle). When the aqueous phase was added, the cross-linking agent was stirred using an orbital shaker at 150 rpm. The appearance of the manufactured microspheres was visually confirmed to confirm the appropriate cross-linking agent concentration.

[0302]

[0303] As a result, as shown in Fig. 17, when the CMC / DS concentrations were each 5%, it was determined that a cross-linking agent concentration of 10% (w / w%) or more was suitable for particle formation.

[0304]

[0305] Example 13-2. Stability of W / O emulsion

[0306] For the homogeneous production of microspheres, the stability of the W / O emulsion can be an important factor, and the stability of the emulsion can be controlled by a surfactant.

[0307] Therefore, in order to secure an emulsion formulation suitable for microsphere production (W / O / W), the stability of the emulsion was confirmed according to the addition of surfactants Span 80 and Twen 20 under the conditions of Table 8.

[0308] The HLB (Hydrophile-Lipophile Balance) of condition 1 was calculated as follows:

[0309] HLB = 16.7(Tween 20 HLB) x 0.1 + 4.3(Span 80 HLB) x 0.1 = 2.1

[0310]

[0311] Theoretically, a low HLB (2.0–6.0) is known to be suitable for W / O emulsions. As a result of observing the degree of emulsion stability visually and microscopically, as shown in Figure 18, the experimental results also confirmed that the emulsion under Condition 1 was more stable than the emulsion under Condition 2, consistent with the theory.

[0312]

[0313] Example 13-3. Confirmation of the feasibility of manufacturing microspheres without DS.

[0314] Water (W): 5% CMC, 0% DS, 10% Tween 20 added to distilled water

[0315] Oil (O): 10% Span 80 added to MCT oil

[0316] The above-mentioned water (W) and oil (O) phases were each mixed in a W:O = 30 mL:120 mL ratio using a homogenizer at 1000 rpm for 10 minutes to prepare a W / O emulsion. 30 mL of a solution containing 10% ZrOCl2·8H2O added to 90% EtOH aqueous solution was placed in a 100 mL beaker and stirred using a mechanical stirrer at 300 rpm to prepare a cross-linking agent aqueous solution. 0.5 mL of the prepared W / O emulsion was pipetted into the cross-linking agent aqueous solution, and then particles were obtained and washed by size using sieves of 500, 300, 100, 53, and 25 μm. Each particle obtained by size was placed in an oven at 60 degrees to remove moisture.

[0317] As a result of observing the manufactured microspheres under a microscope, as shown in FIGS. 19a and 19b, it was found that when particles were manufactured through W / O emulsion manufacturing compared to the dropping method, it was easier to manufacture spherical microspheres by using a micro-sized emulsion manufactured with a homogenizer.

[0318] Therefore, it was confirmed that the above microsphere manufacturing method can control the size of the microspheres manufactured according to the W emulsion size of the W / O emulsion by controlling the stirring speed (RPM) of the homogenizer.

[0319]

[0320] Example 13-4. Preparation of microspheres according to CMC:DS ratio

[0321] <CMC:DS = 5:1 마이크로스피어 제조>

[0322] W: 5% CMC, 1% DS, 10% Tween 20 added to distilled water

[0323] O: Add 10% Span 80 to MCT oil

[0324] The above-mentioned water (W) and oil phase (O) were each mixed in a W:O = 30 mL:120 mL ratio using a homogenizer at 1000 rpm for 10 minutes to prepare a W / O emulsion. 30 mL of a solution containing 10% ZrOCl2·8H2O added to a 90% EtOH aqueous solution was placed in a 100 mL beaker and stirred using a mechanical stirrer at 180 rpm to prepare a cross-linking agent aqueous solution. 4.0 mL of the prepared W / O emulsion was pipetted into the cross-linking agent aqueous solution, and then particles were obtained and washed by size using sieves of 500, 300, 100, 53, and 25 μm. Each particle obtained by size was placed in an oven at 60 degrees to remove moisture.

[0325]

[0326] <CMC:DS = 4:1 마이크로스피어 제조>

[0327] W: 4.8% CMC, 1.2% DS, 10% Tween 20 added to distilled water

[0328] O: Add 10% Span 80 to MCT oil

[0329] The above-mentioned water (W) and oil phase (O) were each mixed in a W:O = 30 mL:120 mL ratio using a homogenizer at 1000 rpm for 10 minutes to prepare a W / O emulsion. 30 mL of a solution containing 10% ZrOCl2·8H2O added to a 90% EtOH aqueous solution was placed in a 100 mL beaker and stirred using a mechanical stirrer at 180 rpm to prepare a cross-linking agent aqueous solution. 4.0 mL of the prepared W / O emulsion was pipetted into the cross-linking agent aqueous solution, and then particles were obtained and washed by size using sieves of 500, 300, 100, 53, and 25 μm. Each particle obtained by size was placed in an oven at 60 degrees to remove moisture.

[0330]

[0331] <CMC:DS = 3:1 마이크로스피어 제조>

[0332] W: 4.5% CMC, 1.5% DS, 10% Tween 20 added to distilled water

[0333] O: Add 10% Span 80 to MCT oil

[0334] The above-mentioned water (W) and oil phase (O) were each mixed in a W:O = 30 mL:120 mL ratio using a homogenizer at 1000 rpm for 10 minutes to prepare a W / O emulsion. 30 mL of a solution containing 10% ZrOCl2·8H2O added to a 90% EtOH aqueous solution was placed in a 100 mL beaker and stirred using a mechanical stirrer at 180 rpm to prepare a cross-linking agent aqueous solution. 4.0 mL of the prepared W / O emulsion was pipetted into the cross-linking agent aqueous solution, and then particles were obtained and washed by size using sieves of 500, 300, 100, 53, and 25 μm. Each particle obtained by size was placed in an oven at 60 degrees to remove moisture.

[0335]

[0336] <CMC:DS = 2:1 마이크로스피어 제조>

[0337] W: 4.0% CMC, 2.0% DS, 10% Tween 20 added to distilled water

[0338] O: Add 10% Span 80 to MCT oil

[0339] The above-mentioned water (W) and oil phase (O) were each mixed in a W:O = 30 mL:120 mL ratio using a homogenizer at 1000 rpm for 10 minutes to prepare a W / O emulsion. 30 mL of a solution containing 10% ZrOCl2·8H2O added to a 90% EtOH aqueous solution was placed in a 100 mL beaker and stirred using a mechanical stirrer at 180 rpm to prepare a cross-linking agent aqueous solution. 4.0 mL of the prepared W / O emulsion was pipetted into the cross-linking agent aqueous solution, and then particles were obtained and washed by size using sieves of 500, 300, 100, 53, and 25 μm. Each particle obtained by size was placed in an oven at 60 degrees to remove moisture.

[0340]

[0341] <CMC:DS = 1:1 마이크로스피어 제조>

[0342] W: 3.0% CMC, 3.0% DS, 10% Tween 20 added to distilled water

[0343] O: Add 10% Span 80 to MCT oil

[0344] The above-mentioned water (W) and oil phase (O) were each mixed in a W:O = 30 mL:120 mL ratio using a homogenizer at 1000 rpm for 10 minutes to prepare a W / O emulsion. 30 mL of a solution containing 10% ZrOCl2·8H2O added to a 90% EtOH aqueous solution was placed in a 100 mL beaker and stirred using a mechanical stirrer at 180 rpm to prepare a cross-linking agent aqueous solution. 4.0 mL of the prepared W / O emulsion was pipetted into the cross-linking agent aqueous solution, and then particles were obtained and washed by size using sieves of 500, 300, 100, 53, and 25 μm. Each particle obtained by size was placed in an oven at 60 degrees to remove moisture.

[0345] As a result of observing microspheres manufactured at different CMC:DS ratios under a microscope, as shown in Figures 20a to 20e, when the CMC:DS ratio was 1:1, no microspheres were formed. Therefore, it was determined that the CMC:DS ratio should be 2:1 or higher for microspheres to be easily formed.

[0346]

[0347] Example 13-5. Drug loading capacity according to CMC:DS ratio

[0348] The microspheres manufactured in Example 13-4 at different CMC:DS ratios were classified by size (100-300 μm). The classified microspheres were placed in a 60-degree oven to completely remove moisture, weighed, and then re-adjusted to a concentration of 1 mg / mL by adding distilled water. Doxorubicin drug 0.25, 0.5, 1, and 2 mg were each dissolved in 2 mL of distilled water. 2 mg / 2 mL of microspheres and 0.25, 0.5, 1, or 2 mg / 2 mL of drug were each mixed and vortexed for 30 seconds. After 7 days of drug loading, 50 uL of the supernatant was sampled, diluted in 1950 uL of distilled water, and the drug loading rate was measured by measuring UV-vis absorbance, and the drug loading capacity was calculated.

[0349] The results of confirming the properties before and after drug loading according to the size of the microspheres based on CMC:DS=5:1 are shown in Fig. 21a, the drug loading amount according to the CMC:DS ratio is shown in Table 9, and the results of confirming the properties after drug loading according to the CMC:DS ratio are shown in Fig. 21b.

[0350]

[0351]

[0352] Example 14. Optimization of the microsphere manufacturing process.

[0353] Example 14-1. Confirmation of microsphere size distribution according to W / O emulsion conditions.

[0354] The size distribution of microspheres manufactured according to the homogenizer conditions of the W / O emulsion was confirmed. Specifically, the size distribution of the microspheres was confirmed as follows.

[0355] W: 5% CMC, 1% DS, 10% Tween 20 added to distilled water

[0356] O: Add 10% Span 80 to MCT oil

[0357] The above-mentioned water (W) and oil (O) phases were each homogenized at a ratio of W:O=30 mL:120 mL at 1000 rpm or 2000 rpm for 10 minutes to prepare a W / O emulsion. 30 mL of a solution of 10% ZrOCl2·8H2O added to 90% EtOH aqueous solution was placed in a 100 mL beaker and stirred at 180 rpm using a mechanical stirrer to prepare a cross-linking agent aqueous solution. 4.0 mL of the prepared W / O emulsion was pipetted into the cross-linking agent aqueous solution, and then microspheres were obtained and washed by size using sieves of 500, 300, 100, 53, and 25 μm. Each microsphere obtained by size was placed in a 60 degree oven to remove moisture, and then the weight of each microsphere by size was measured to confirm the size distribution.

[0358] As a result, as shown in Tables 10 and 11, it was confirmed that a large amount of microspheres of 100 to 500 μm in size were produced under 1000 rpm conditions, and a large amount of microspheres of 25 to 100 μm in size were produced under 2000 rpm conditions. From this, it was found that the size of the microspheres was controlled depending on the rpm conditions of the homogenizer of the W / O emulsion. The weight measurement results of the microspheres by size according to the rpm of the homogenizer are shown in Tables 10 and 11.

[0359]

[0360]

[0361]

[0362] Example 14-2. Confirmation of microsphere yield according to CMC:DS ratio

[0363] As shown in Table 12, after manufacturing the microspheres, the weight of the manufactured microspheres was measured to confirm the yield of microspheres according to the CMC:DS ratio.

[0364] As a result, as shown in Table 13, when 4 mL of W / O emulsion was injected for crosslinking under each condition, an average of 70.2 mg of microspheres could be obtained, and the deviation in the amount of microspheres obtained under each condition was confirmed to be 4.74 mg.

[0365] The yield of microspheres according to the CMC:DS ratio is shown in Table 13.

[0366]

[0367]

[0368]

[0369] As described above, the novel microspheres using the anionic polymer according to the present invention were manufactured by injecting a W / O emulsion containing CMC and DS into a crosslinking agent aqueous solution, and the manufactured microspheres exhibited size distributions of 25-50, 50-100, 100-300, and 300-500 um. In addition, in order to find the optimal formulation for manufacturing the microspheres of the present invention, the CMC / DS ratio was divided into 5:1, 4:1, 3:1, 2:1, and 1:1 to manufacture microspheres, and as a result, it was confirmed that the CMC / DS ratio of 2:1 or higher is necessary for easy manufacturing of spherical particles, and as a result of loading Doxorubicin, a cationic drug, onto the microspheres and checking the loading ability according to the CMC / DS ratio, it was confirmed that 0.3771, 0.5611, 0.5112, 0.4488, and 0.3490 mg of drug were loaded per 1 mg of particles for 5:1, 4:1, 3:1, 2:1, and 1:1, respectively. Therefore, it was determined that the CMC / DS ratio of 4:1 is optimal in terms of ease of manufacturing microspheres and drug loading ability.

[0370] Furthermore, it was confirmed that the particle distribution of the microspheres can be controlled by adjusting the rpm conditions of the homogenizer used in the production of the W / O emulsion in the production process of the microspheres of the present invention.

[0371]

[0372] The foregoing description of the present invention is provided for illustrative purposes only. Those skilled in the art will readily appreciate that the present invention can be readily modified into other specific forms without altering the technical spirit or essential characteristics of the present invention. Therefore, the embodiments described above should be understood as illustrative in all respects and not restrictive.

[0373]

[0374] The present invention provides a novel method for manufacturing microspheres, which are useful for the delivery of cationic drugs, have low toxicity, are biocompatible, and can deliver drugs to microvessels, and an optimal formulation. In addition, the microsphere manufacturing method of the present invention allows the microspheres to directly embolize blood vessels by adjusting the particle size according to the size of the blood vessel through controlling the size distribution of the microsphere particles, and improves the yield of microspheres of a desired size, thereby enabling the mass production of microspheres of a specific size, and thus has industrial applicability.

Claims

1. A composition for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt as active ingredients.

2. In paragraph 1, A composition characterized in that the anionic polymer is selected from the group consisting of carboxymethylcellulose or a salt thereof; dextran sulfate or a salt thereof; and mixtures thereof.

3. In paragraph 1, A composition characterized in that the above microspheres comprise carboxymethylcellulose and dextran sulfate.

4. In paragraph 3, A composition characterized in that the carboxymethylcellulose and dextran sulfate are included in a mass ratio of 0.5 to 20:

1.

5. In paragraph 3, A composition characterized in that the carboxymethylcellulose and dextran sulfate are included in a concentration ratio of 0.5 to 20:

1.

6. In paragraph 1, The above multivalent metal compound salt is Zr 2+ , Zr 4+ , Fe 2+ , Fe 3+ , Ca 2+ , Al 3+ , Cu 2+ , and Zn 2+ A composition characterized in that it is a salt of one or more divalent to tetravalent metal compounds selected from the group consisting of:

7. In paragraph 1, A composition characterized in that the anionic polymer and the polyvalent metal compound salt are included in a concentration ratio of 0.1 to 10:

1.

8. In paragraph 1, A composition characterized in that the above microspheres are loaded with a cationic drug.

9. In paragraph 8, A composition characterized in that the cationic drug is at least one selected from the group consisting of doxorubicin, procainamide, digoxin, quinidine, trimethoprim, cimetidine, vancomycin, irinotecan, daunorubicin, epirubicin, diphenhydramine, memantine, oxycodone, pyrilamine, tramadol, and pharmaceutically acceptable salts thereof.

10. In paragraph 8, A composition characterized in that the cationic drug is loaded into the microspheres at a mass ratio of 1:1 to 10.

11. In paragraph 1, A composition characterized in that the composition is in the form of an emulsion.

12. In paragraph 1, A composition characterized in that the above microspheres have a diameter of 10 to 700 μm.

13. In paragraph 1, A composition characterized in that the above microspheres have a size distribution as follows: a) 10 to 50 μm; b) 50 to 100 μm; c) 100 to 300 μm; d) 300 to 500 μm; and e) 500 to 700 μm.

14. In paragraph 13, A composition characterized in that the above microspheres are separated according to size distribution.

15. In paragraph 1, A composition characterized in that the above microspheres are used for cancer treatment.

16. In paragraph 1, A composition characterized in that the above microspheres are used in vascular embolization.

17. A method for producing microspheres comprising an anionic polymer and a polyvalent metal compound salt, comprising the following steps: A step of forming an emulsion by mixing an aqueous phase containing an anionic polymer and an oil phase containing a nonionic surfactant using a homogenizer; A step of forming microspheres by adding the above emulsion to a multivalent metal compound salt aqueous solution; and A step of separating microspheres of a desired size using a sieve.

18. In paragraph 17, A method characterized in that the anionic polymer is selected from the group consisting of carboxymethylcellulose or a salt thereof; dextran sulfate or a salt thereof; and mixtures thereof.

19. In paragraph 17, A method characterized in that the nonionic surfactant is selected from the group consisting of polyvinyl alcohol, polyalkylene glycol, polyvinyl pyrrolidone, alkyl polyglycoside, t-octylphenoxy polyethoxy ethanol, poloxamer, polysorbate, poly(N-vinylacetamide), polyvinyl butyral, sorbitan oleate, and mixtures thereof.

20. In paragraph 17, A method, characterized in that the oil phase is selected from the group consisting of mineral oil, vegetable oil, medium chain triglyceride, and mixtures thereof.

21. In paragraph 17, A method characterized in that the above-mentioned water phase and oil phase are mixed in a volume ratio of 1:1 to 20.

22. In paragraph 17, The above multivalent metal compound salt is Zr 2+ , Zr 4+ , Fe 2+ , Fe 3+ , Ca 2+ , Al 3+ , Cu 2+ , and Zn 2+ A method characterized in that it is a salt of one or more divalent to tetravalent metal compounds selected from the group consisting of.

23. In paragraph 17, A method characterized in that the above emulsion is added to a multivalent metal compound salt aqueous solution at a volume ratio of 1:1 to 20.

24. In paragraph 17, The above method is characterized in that the size distribution of microspheres is controlled by controlling the rpm of the homogenizer.

25. A method for preparing a composition for treating cancer, comprising the following steps: A step of forming an emulsion by mixing an aqueous phase containing an anionic polymer and an oil phase containing a nonionic surfactant using a homogenizer; A step of forming microspheres by adding the above emulsion to a multivalent metal compound salt aqueous solution; A step of separating microspheres of a desired size using a sieve; and A step of mixing a cationic drug with the above microspheres.

26. Microspheres comprising an anionic polymer and a polyvalent metal compound salt.

27. Use of an anionic polymer and a polyvalent metal compound salt; or a composition containing the same as an active ingredient for producing microspheres.

28. Use of an anionic polymer and a polyvalent metal compound salt for the production of microspheres; or a composition comprising the same as an active ingredient.

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