Method for manufacturing a swelling polymer with an immobilized object

Abstract

The present invention discloses a method for manufacturing a swellable polymer having an object fixed thereto. The manufacturing method of the present invention includes a step of absorbing a solution of the object, which is dissolved or dispersed in a solvent having affinity for the swellable polymer, into a dried swellable polymer including a linker having a functional group capable of binding with the object, and unlike conventional manufacturing methods in which an object is fixed before crosslinking of the swellable polymer, the method of fixing the object after crosslinking of the polymer using the swellable polymer has the advantage of being able to fix the object quickly and with very high efficiency compared to conventional techniques.

Description

Technical Field

[0001] The present invention relates to a method for producing a swellable polymer to which an object is fixed, and more specifically, to a method for fixing an object to a swellable polymer by utilizing the absorption characteristics of the dried swellable polymer.

Background Art

[0002] A swellable polymer is a polymer that has the property of absorbing a solvent and swelling based on the water-absorbing characteristics of the polymer, and is usefully utilized in a very wide range of fields such as medicine, agriculture, and biotechnology. Generally, by utilizing the swelling behavior of the swellable polymer to carry a target substance between the cross-linked spaces of the polymer, it is used as a carrier for transmitting the target substance. At this time, a step of reacting is performed for a stable bond between the swellable polymer and the target substance.

[0003] Currently, such a step is performed before cross-linking the swellable polymer, and a method is adopted in which the polymer and the target substance are first reacted and bonded, and then the swellable polymer is cross-linked. However, this requires a lot of time, and even when a large amount of fixing substance is used, the fixing efficiency of the target substance is not good. Therefore, a method for rapidly bonding a large amount of fixing substance per unit volume of the swellable polymer is required.

[0004] On the other hand, although the mortality rate of cancer diseases tends to decrease due to early diagnosis and the development of medical technology, it is still the number one cause of death worldwide. Clinically, methods for treating cancer are roughly classified into three types: surgical therapy for locally removing cancer, radiation therapy, and chemotherapy using anticancer drugs when cancer has spread throughout the body. In particular, radiation therapy is widely used by more than 50% of cancer patients alone or in combination with chemotherapy.

[0005] Among radiation therapy methods, the therapy using a radioactive isotope has an advantage of minimizing the pain of patients because, unlike existing external radiation therapy, it does not require large equipment and is administered simply using a syringe compared to surgical operations. The radioactive isotopes currently applied are 90 Y,67 Cu, 188 Re, 177 Lu, 131 It is administered intravenously in solution. However, radioactive isotopes circulating in the body can damage normal cells and tissues due to reduced selectivity for tumor cells, causing side effects such as hormonal abnormalities and hair loss in patients.

[0006] To address these problems, a technology has recently been developed that allows for localized radiotherapy by chemically bonding (labeling) radioisotopes to biodegradable polymers in solution, and then formulating the resulting polymer-radioisotope(drug) conjugate into microparticles that can be injected while remaining in the body for an extended period (Figure 1a). However, formulating the polymer-radiodrug conjugate, which has a long half-life, into microparticles for injection into the body requires many steps and a long time, making it difficult to manufacture directly in hospitals. Furthermore, the long formulation process can lead to the disappearance of the radioactive drug and a decrease in its radioactivity. [Overview of the project] [Problems that the invention aims to solve]

[0007] The present invention is provided in response to the above-mentioned problems and needs, and aims to provide a novel method that can rapidly and easily bind a large amount of immobilized material to a swellable polymer.

[0008] The objective is to provide a method for producing a swelling polymer to which an object is immobilized, which allows for the highly efficient immobilization of an object to the swelling polymer by utilizing the absorption of the dried swelling polymer. [Means for solving the problem]

[0009] The present invention provides a method for producing an object-immobilized swelling polymer, comprising the step of absorbing the object solution, which is dissolved or dispersed in a solvent compatible with the swelling polymer, into a dried swelling polymer containing a linker having a functional group capable of binding to an object.

[0010] In the present invention, the term "object" has the same meaning as "substance to be fixed," and refers to an object to be fixed to a swellable polymer. For example, the object may be a drug used in the body for treatment and prevention. In the present invention, a swellable polymer refers to a polymer that is crosslinked and does not dissolve in a solvent, and which rapidly absorbs affinity solvents and expands in volume. For example, examples of polymers that absorb new aqueous solvents and do not dissolve in new aqueous solvents include hydrogels, such as polysaccharides like hyaluronic acid, alginates, and chitosan, polyethylene glycol (PEG), polyvinyl alcohol (PVA), gelatin, collagen, and albumin hydrogels. Examples of polymers that absorb hydrophobic solvents and are insoluble in hydrophobic organic solvents include hydrophobic polymers such as polylactide, polyglycolide, or their copolymers poly(lactide-co-glycolide) (PLGA), and may also be crosslinked polymers of PLA-diacrylate, PGA-diacrylate, or PLGA-diacrylate containing the photocrosslinking group methacarylate (MA). Preferably, the swelling polymer of the present invention may be a biocompatible polymer.

[0011] Conventionally, when producing a compound in which an object is bound to such a swellable polymer, first, after fixing the object, the polymer is crosslinked and produced. However, the present invention provides a method of fixing an object after first crosslinking the polymer. The present inventors produced a crosslinked and dried swellable polymer containing a functional group or a linker capable of binding to an object, and then absorbed a solution of the object dissolved or dispersed in a solvent having an affinity for the swellable polymer, and newly found that high binding efficiency can be provided even with a small amount of the object solution. Further, the production method of the present invention has the feature that it can be produced very easily by reacting a solution containing a dried polymer and an object at the necessary site, rather than producing a polymer in which the object is fixed in advance.

[0012] In the present invention, the object may be any one selected from the group consisting of a radioisotope, a metal, an ionic compound, and an organic compound.

[0013] Preferably, the object may be a substance that provides a pharmacological effect in vivo.

[0014] The radioisotope may be any one or more selected from radioactive halogen group elements or radioactive metal elements. For example, the radioisotope is 123 I, 124 I, 125 I, 131 I, 211 At, 64 Cu, 67 Ga, 68 Ga, 44 Sc, 47 Sc, 111 In, 177 Lu, 86 Y, 90 Y, 89 Zr, 212 Bi, 213 Bi, 166 Ho, 99m Tc, 212 Pb and 225It may be any one or more selected from the group consisting of Ac, etc.

[0015] The aforementioned metal may be one or more selected from the group consisting of gold (Au), platinum (Pt), silver (Ag), aluminum (Al), copper (Cu), nickel (Ni), cobalt (Co), palladium (Pd), chromium (Cr), manganese (Mn), and alloys thereof. However, the present invention is not necessarily limited to the types of radioisotopes and metals described.

[0016] The bonding between the object and the linker can be any form of bonding that allows the object to be fixed to the swellable polymer, such as covalent bonds, hydrogen bonds, coordination bonds, non-covalent bonds, such as ionic bonds or van der Waals bonds. However, the present invention is not limited to such specific bonds. In particular, it is possible as long as the object is fixed when the swellable polymer is injected into a living organism to perform its intended function.

[0017] In one embodiment of the present invention, if the object is a metal or an ionic compound, the linker may be a ligand compound that can coordinately bond with the metal or the ionic compound.

[0018] In one embodiment of the present invention, if the object has a cationic functional group, the ligand may be a chelator. A swelling polymer on which an object produced by bonding a cationic metal and a chelate is immobilized may have, for example, the following structures of chemical formulas 1 to 3.

[0019] [ka]

[0020] In the above chemical formula 1, M is a radioactive metallic element, and n may be an integer of 1 or more.

[0021] A swelling polymer having the structure of the chemical formula 1, wherein M is specifically: 64 Cu, 67 Ga, 68 Ga, 44 Sc, 47 Sc, 111 In, 177 Lu, 86 Y, 90 Y, 89 Zr, 212 Bi, 213 Bi, 166 Ho and 99m It may be selected from the Tc group. The M may be labeled by being coordinately bonded to diethylenetriaminepentaacetic acid (DTPA) which is bonded to hyaluronic acid methacrylate (HAMA) of chemical formula 1.

[0022] [ka]

[0023] In the above chemical formula 2, M is a radioactive metal element, and n may be an integer of 1 or more.

[0024] A swelling polymer having the structure of the chemical formula 2, wherein M is specifically: 64 Cu, 67 Ga, 68 Ga, 44 Sc, 47 Sc, 111 In, 177 Lu, 86 Y, 90 Y, 89 Zr, 213 Bi, 212 Pb and 225M may be selected from the group consisting of Ac. M may be labeled by coordinating with dodecane tetraacetic acid (DOTA) bonded to hyaluronic acid methacrylate (HAMA) of chemical formula 2.

[0025] [ka]

[0026] In the above chemical formula 3, M is a radioactive metal element, and n may be an integer of 1 or more.

[0027] A swelling polymer having the structure of the chemical formula 3, wherein M is specifically: 64 Cu, 67 Ga, 68 Ga, 44 Sc, 47 Sc, 111 In, 177 Lu, 86 Y, 90 Y, 212 Bi, 213 Bi and 212 It may be selected from the group consisting of Pb. The aforementioned M may be labeled by being coordinately bonded to triazacyclononane triacetic acid (NOTA) which is bonded to hyaluronic acid methacrylate (HAMA) of chemical formula 3.

[0028] In the present invention, the chelate is DOTA, a derivative of DOTA, DOTA-GA, DO2A, HBED-CC, NOTA, p-SCN-Bn-NOTA, NODA-GA, PrP9, DTPA, CHX-A''-DTPA-isothiocyanate, CHX-A''-DTPA-maleimide, CHX-A''-DTPA-glutaric acid NHS ester. ester), DTPA derivatives (1M3B-DTPA, 1B3M-DTPA, 1B4M-DTPA, CHX-A-DTPA and CHX-B-DTPA), NETA, NE3TA, NE3TA-Bn, C-NETA, C-NE3TA, C-DOTA-isothiocyanates, MeO-DOTA-isothiocyanates, PA-DOTA-isothiocyanates, 1B4M-DTPA-isothiocyanates, 2-(4-nitrobenzyl)-cyclen, 2-(4-nitrobenzyl)-cyclam, 2-(4-benzamidobenzyl)-NOTA(2-(4-benzamid (obenzyl)-NOTA, 2-(4-nitrobenzyl)-DOTA, 2-(4-nitrobenzyl)-TETA, 1-(4-nitro)-B4MDTPA, TETA, TE2A, derivatives of TE2A (CB-TE2A, CB-TE1A1P, CB-TE2P, MM) -TE2A, DM-TE2A), cyclam, cyclam derivatives (1-(4-nitrobenzyl)-cyclam (1-(4-nitrobenzyl)-cyclam), 6-(4-nitrobenzyl)-cyclam (6-(4-nitrobenzyl)-cyclam)), CB-TE2A, CB-TE2A derivatives (3,3'-(1,4,8,11-tetraazabicyclo[6.6.2) The present invention may contain one or more of the following selected compounds: hexadecane-4,11-diyl)dipropanoic acid, CB-DO2A, CB-DO2A derivatives (3,3'-(1,4,7,10-tetraazabicyclo[5.5.2]tetradecane-4,10-diyl)dipropanoic acid, DiAmSar, SarAr, AmBaSar, derivatives of DiAmSar, AAZTA, NOPO, and TACN-TM. However, the present invention is not necessarily limited to these types.

[0029] In one embodiment, if the object has an anionic functional group, the functional group of the linker may be an aromatic compound or a heterocyclo compound, and the anionic functional group and the functional group of the linker can be bonded by an electrophilic substitution reaction. For example, the heterocyclo compound may be selected from the group consisting of imidazole, pyrrole, furan, thiophene, indole, and 3,4-dihydroxyphenyl.

[0030] As an example, a swelling polymer on which an object produced by bonding an anionic functional group with a chelate can have the structure of the following chemical formula 4.

[0031] [ka]

[0032] In the aforementioned chemical formula 4, X is a radioactive halogen element, and n may be an integer of 1 or more. X may be labeled with aminopropyl imidazole (API) bonded to hyalrunonate methacrylate (HAMA) of chemical formula 4.

[0033] In one embodiment, if the object has a functional group that can be ionized, the object can be immobilized by ionic bonding if the crosslinked swelling polymer has a functional group with the opposite charge. For example, if the object has a cationized amine group (-NH2), it can be immobilized by ionic bonding with the carboxyl group (-COOH) that becomes the anion of HAMA. Conversely, if the object has an anionized -COOH or sulfonic acid group (-SO3H), the object can be immobilized by ionic bonding by immediately incorporating the amine group into HAMA via a diacetyl reaction, or by incorporating a linker having an amine group. In particular, proteins composed of amino acids can be easily immobilized because they have -COOH and -NH2 at both ends.

[0034] In one embodiment, the object is an organic compound, the organic compound comprises one or more first functional groups, which include an amine group, a carboxyl group, a hydroxyl group, and a thiol group, and the linker may have a second functional group that can bond to the first functional group.

[0035] In one embodiment, if the organic compound contains an amine group (-NH2), the functional group of the linker may contain an aldehyde group (-CHO) that can bond to the amine group. The object can be fixed to the swellable polymer by the reaction and bonding of the amine group and the aldehyde group.

[0036] In other embodiments, if the functional group of the organic compound includes a carboxyl group (-COOH), the functional group of the linker may include an amine group (-NH2) that can bond to the carboxyl group. The object can be fixed to the swelling polymer by the reaction and bonding of the carboxyl group and the amine group.

[0037] In other embodiments, if the functional group of the organic compound includes a hydroxyl group (-OH), the functional group of the linker may include a carboxyl group (-COOH). The object can be fixed to the swellable polymer by the reaction and bonding of the hydroxyl group and the carboxyl group.

[0038] In other embodiments, if the functional group of the organic compound includes a thiol group (-SH), the functional group of the linker may include a disulfide group (-SS-). The object can be fixed to the swellable polymer by the reaction and bonding of the thiol group and the disulfide group.

[0039] On the other hand, in the present invention, the dried swelling polymer may be produced by any one of the following methods selected from freeze-drying, hot-air drying, atmospheric pressure drying, and vacuum drying, but preferably the drying is carried out by freeze-drying. Furthermore, the dried swelling polymer may be in the form of powder or aerogel.

[0040] Prior to the step of allowing the dried swelling polymer to absorb the object solution, an additional step may be performed in which a binder having functional groups capable of binding to the object, and the functional groups capable of binding to the linker of the dried swelling polymer, is reacted. However, this is not necessarily limited to this step, and may be performed selectively when necessary.

[0041] The step of allowing the dried swelling polymer to absorb the object solution may be carried out by supporting the dried swelling polymer in the object solution, but preferably by gradually dropping or adding the object solution to the dried swelling polymer so as to provide sufficient time for the dried swelling polymer to absorb the object solution. Through this absorption step, the object can be immobilized on the swelling polymer by chemically bonding with a linker that is not supported within the swelling polymer. [Effects of the Invention]

[0042] Dried swellable polymers have a porous structure and possess the property of rapidly absorbing object solutions through osmosis. The rapidly absorbed object solution has a rapidly increasing probability of meeting the functional groups of the swellable polymer, thereby enabling highly efficient bonding using a small amount of object solution.

[0043] Furthermore, according to the present invention, it is possible to label objects in moisture at sites requiring swellable polymers (for example, hospitals), and although it is a single-step process, it is possible to label objects with high efficiency, improving user convenience and providing a groundbreaking reduction in object waste. In addition, when radioactive isotopes are used as objects, it has the advantage of being able to label them on-site in a short time while minimizing the decrease in the activity of the radioactive isotopes.

[0044] Furthermore, the manufacturing method proposed in this invention has the advantage of being applicable not only to a single substance, but also to the labeling of a wide variety of drugs, from organic and low molecular weight compounds (chemicals) that can be supported during the absorption process, to high molecular weight peptides, gene therapy agents, and protein drugs. In addition, it can be used not only for particles but also for various formulations such as films, sponges, and patches, giving it the potential for application in diverse fields. [Brief explanation of the drawing]

[0045] [Figure 1] The drawings illustrate a method for producing a swelling polymer with an object immobilized by the present invention, showing a) a conventional method and b) the method of the present invention. [Figure 2] This drawing compares the fixing efficiency of object (131I) in Example 1 using the manufacturing method of the present invention and in Comparative Example 1 using a conventional manufacturing method. [Figure 3] This diagram shows the fixation efficiency of object (131I) depending on the concentration of the oxidizing agent and the absorption time in the manufacturing method of the present invention. [Figure 4] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 5a] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 5b] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 6a] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 6b] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 6c] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 6d] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 6e] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 7a] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 7b] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 8a] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 8b] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 9a] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Figure 9b] This is a drawing illustrating one embodiment of the linker-object coupling in the manufacturing method of the present invention. [Modes for carrying out the invention]

[0046] Embodiments of the present invention will be described in detail below with reference to the accompanying drawings. The present invention can be modified in various ways and has various embodiments, and specific embodiments are illustrated in the drawings and described in detail in the text. However, this is not intended to limit the present invention to any particular disclosure, but should be understood to include all modifications, equivalents, or substitutes that fall within the spirit and technical scope of the present invention. In the description of each drawing, similar reference numerals indicate similar components.

[0047] The terms used in this application are used solely to describe specific embodiments and are not intended to limit the invention. Singular expressions include plural expressions unless they are clearly different in context. In this application, terms such as “includes” or “having” are intended to specify the existence of features, stages, operations, components, parts, or combinations thereof described in the specification, and should be understood not to preemptively exclude the possibility of the existence or addition of one or more other features, stages, operations, components, parts, or combinations thereof.

[0048] Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as those generally understood by a person of ordinary skill in the art to which this invention pertains. Terms as defined in commonly used dictionaries should be interpreted as having the meaning consistent with their meaning in the context of the relevant art, and not as ideal or overly formal unless expressly defined herein.

[0049] Hereinafter, a method for producing a swelling polymer in which the object of the present invention is fixed will be specifically described with reference to the drawings of the present invention.

[0050] Figure 1 is a diagram illustrating a method for producing a swellable polymer with an immobilized object according to the present invention, comparing a) a conventional method for producing a polymer with an immobilized object and b) the method according to the present invention.

[0051] Referring to Figure 1, a) Conventional methods for producing polymers with immobilized objects involve reacting a linker having functional groups capable of binding to the object with the polymer to first bond the object, and then crosslinking it. Conventional methods have the disadvantage of being difficult to use easily on-site because they require many steps and a long time, which may reduce the activity of the object. To solve this, methods that increase the concentration of the object and reduce the reaction time are generally used, but this is difficult because the object does not dissolve above a certain concentration that exceeds the solubility. In addition, conventional methods have the disadvantage that the object is bonded to the polymer with very low efficiency.

[0052] On the other hand, b) the manufacturing method of the present invention includes reacting a swellable polymer with a linker having a functional group capable of binding to an object, binding them together, drying the polymer, and then absorbing the resulting polymer with an object solution dissolved or dispersed in a solvent compatible with the swellable polymer. In the manufacturing method of the swellable polymer of the present invention, the absorption step can relatively increase the local concentration within the particles of the swellable polymer so that the object can bind to the reaction groups (linkers) within the swellable polymer particles, thus having the advantage of binding the object in a short time. Furthermore, as long as there is a dried swellable polymer containing a linker capable of binding to an object and an object solution dissolved or dispersed in a solvent compatible with the swellable polymer, a swellable polymer with immobilized objects can be easily manufactured at any site, and it has the advantage of providing a very high binding rate in a short time, and process efficiency can also be improved.

[0053] The method for producing a swelling polymer in which the object of the present invention is immobilized will be described in more detail below with reference to specific examples and comparative examples. However, the examples of the present invention are only a part of the embodiments of the present invention, and the scope of the present invention is not limited to the examples below.

[0054] [Example 1] To synthesize hyaluronic acid methacrylate (HAMA), which was used in the production of a swelling polymer, 1 g of hyaluronic acid was placed in 10 mL of deionized water at 25°C and dissolved using a stirrer at 1000 RPM (rotation per minute) for 1 hour to prepare a solution. Then, the temperature of the solution was lowered to 4°C, and 1.6 g of methacrylic anhydride was added while adjusting the pH of the solution to 9 using 1 M NaOH, and the mixture was reacted for 24 hours to produce a mixture. To remove unreacted material from the mixture, the mixture was placed in a cellulose membrane (molecular weight cut-off: 3500) and a dialysis process was performed for 96 hours, with the deionized water being replaced every 8 hours. The dialyzed reaction product was freeze-dried to obtain crosslinkable HAMA. Next, an object (radioactive iodine) was added to the HAMA. 131 A reaction was carried out to incorporate a reaction group (linker, API) that binds to I). For this purpose, HAMA polymer (1 g) was added to phosphate-buffered saline (PBS) (100 mL) with a pH of 7.4 and dissolved using a stirrer at 1000 RPM for 1 hour to prepare a polymer solution. Next, 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) (900 mg) and N-Hydroxysuccinimide (1100 mg) were added to the polymer solution and stirred at 1000 RPM for 1 hour. After that, API (600 mg) dissolved in 40 mL of PBS was added to the mixture and stirred for 24 hours to obtain the reaction product. To remove unreacted material from the mixture, the mixture was placed in a cellulose membrane (molecular weight cut-off: 3500) and a dialysis process was carried out for 96 hours, with the deionized water being replaced every 8 hours. The dialyzed reaction product was freeze-dried to obtain HAMA-API with the API bonded to it.

[0055] Next, in order to produce a swellable polymer capable of fixing objects, HAMA-API (50 mg) and the photoinitiator lithium phenyl(2,4,6-trimethylbenzoylphosphinate, LAP) (1 mg) were dissolved in deionized water (1 mL), and then dispersed on oil to a size of several tens of microns, and then subjected to an intensity of 300 mW / cm². 2 Crosslinked polymers in hydrogel particle form were produced by irradiating with 365nm light for 60 seconds. Subsequently, the crosslinked polymers were washed twice with ethanol (ethyl alcohol) and deionized water to remove impurities (oil, photoinitiator, etc.), and the crosslinked polymers dispersed in deionized water were freeze-dried to form the object. 131 A swellable polymer capable of immobilizing I) was obtained. A vial containing the dried swellable polymer in particulate form was filled with chloramine T (0.5 mg) and 1 mCi concentration Na. 131 After adding 100 μL of aqueous solution I and vortexing for 10 minutes, sodium metabisulfite solution (10 mg / mL, 10 μL) was added to quench the labeling of the radioactive iodine. The supernatant was removed, and after washing with additional ethanol and deionized water, a polymer was finally produced in which the radioactive iodine was fixed and swollen.

[0056] Comparative Example 1 Comparative Example 1 of the present invention was prepared using a conventional manufacturing method. 240 mg of HA was dissolved in 20 mL of deionized water at 25°C to prepare the first solution. The pH of the first solution was then adjusted to pH 9 using 1 M NaOH at 4°C. Then, 420 mg of methacrylic anhydride was added, and the mixture was prepared by stirring for 24 hours. The mixture was then dialyzed with deionized water every 8 hours for 96 hours, and the dialyzed product was freeze-dried to obtain hyaluronic acid methacrylate (HAMA). Next, N-(3-Aminopropyl)-imidazole was reacted with HAMA to bond API to HAMA via an amidation reaction between the carboxyl group of HAMA and the amine group of API. Specifically, HAMA (240 mg) was dissolved in phosphate-buffered saline (PBS) (40 mL, pH 7.4) at 25°C for 4 hours. Then, EDC (230 mg, 1.2 mmol) and NHS (276 mg, 2.4 mmol) were added and the mixture was stirred for 30 minutes to activate the carboxyl group of HAMA. Subsequently, API (150 mg, 1.2 mmol) dissolved in PBS (10 mL) was added dropwise, and the reaction was carried out for 12 hours. The resulting mixture was dialyzed with PBS and freeze-dried to obtain HAMA bound to API (HAMA-API).

[0057] Next, to immobilize the iodine object on HAMA-API, chloramine T was used as a catalyst (oxidizing agent) for iodation of the imidazole ring of HAMA-API. HAMA-API (24 mg) was dissolved in 500 μL of PBS (pH 7.4) in a 1.5 mL centrifuge tube, and 100 μL of Na was added. 131I was added for 10 minutes. Next, chloramine T solution (5 mg / mL, 10 μL) dissolved in PBS (pH 7.4) was added to the mixture and vortexed for 10 minutes, then sodium metabisulfite solution (10 mg / mL, 10 μL) was added to quench the iodation. Ethyl alcohol (1 mL) was added to the reaction to remove the residue, and the mixture was centrifuged at 5000 RPM for 20 minutes. The supernatant was removed, and iodine-labeled HAMA (I-HAMA or 131 I obtained I-HAMA.

[0058] Figure 2 shows the product of Example 1 manufactured by the manufacturing method of the present invention. 131 I Fixed efficiency and Comparative Example 1 manufactured by a conventional manufacturing method 131 This is a diagram for comparing fixed efficiency.

[0059] Referring to Figure 2, when manufactured using a conventional solution-based manufacturing method, the object is 131 While the fixation efficiency of I is very low at approximately 39%, when manufactured using the manufacturing method (freeze-dried) utilizing the swellable polymer of the present invention, 131 The fixing efficiency of I is approximately 68%, confirming that it provides significantly higher object fixing efficiency compared to conventional technologies.

[0060] Figure 3 shows the manufacturing process of Example 1 produced by the manufacturing method of the present invention, a) the concentration of the oxidizing agent chloramine T 131 I) Immobilization efficiency and b) Dried HAMA-API swelling polymer 131 Absorption reaction time of solution I 131 This is a diagram showing the fixed efficiency of I.

[0061] Referring to Figure 3, a) the concentration of the oxidizing agent used increases, 131 It can be confirmed that the fixation efficiency of I increases, and b) HAMA-API swelling polymer 131When absorbing solution I, approximately 65% ​​or more per minute 131 It can be confirmed that I is fixed. Therefore, the manufacturing method of the present invention is a method that can fix an object to a swellable polymer very quickly with just a simple step of reacting a solution containing the object with a dried swellable polymer, thereby minimizing the decrease in the activity of the object and allowing for easy and rapid manufacturing in any location.

[0062] [Example 2] HAMA swelling polymer with immobilized cationic metal A reaction was carried out to incorporate a chelating agent (linker, DTPA) that binds to an object (cationic radioisotope) into HAMA. For this purpose, 0.7 g of HAMA polymer was added to 30 mL of phosphate-buffered saline (PBS) with a pH of 7.4 and dissolved at 1000 RPM for 1 hour using a stirrer to prepare a polymer solution. Then, 0.35 g of 4-(4,6-Dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM) was added to the polymer solution and stirred at 1000 RPM for 6 hours. After that, 0.42 g of p-NH2-Bn-DTPA dissolved in 2 mL of PBS was added to the mixture and stirred for 18 hours to obtain the reaction product. To remove unreacted material from the mixture, the mixture was placed in a cellulose membrane and a dialysis process was carried out for 72 hours, with the deionized water being replaced every 12 hours. The dialyzed reaction product was freeze-dried to obtain HAMA-DTPA bound to DTPA.

[0063] Next, in order to produce a swellable polymer capable of fixing objects, the HAMA-DTPA (50 mg) and the photoinitiator LAP (10 mg) were dissolved in deionized water (1 mL), and then dispersed on oil to a size of several tens of microns, and then subjected to a strength of 7 W / cm². 2Crosslinked polymers in hydrogel particle form were produced by irradiating with 365 nm light for 60 seconds. Subsequently, the crosslinked polymers were washed three times with ethanol (ethyl alcohol) and deionized water to remove impurities (oil, photoinitiator, etc.). The crosslinked polymers dispersed in deionized water were then freeze-dried to obtain a swellable polymer capable of immobilizing an object (cationic radioisotope). A vial containing the dried swellable polymer in particle form (3 mg) was mixed with a 5-6 mCi concentration ZrCl4 (100 μL) aqueous solution and reacted for 1 hour with a stirrer at 27°C and 15000 RPM. The supernatant was removed and washed with additional deionized water to finally produce a swellable polymer with immobilized radioactive zirconium.

[0064] As shown in Figure 4, the fixation efficiency of zirconium ions by swelling polymers can be confirmed, and it was confirmed that swelling polymers to which DTPA, which can fix cationic metals, is bonded have a fixation efficiency of approximately 90%.

[0065] [Example 3] HAMA swelling polymer with immobilized doxorubicin Figure 5a is a diagram showing the manufacturing process of Example 3, which was manufactured using the manufacturing method of the present invention.

[0066] Referring to Figure 5a, Example 3 of the present invention is a method for immobilizing a fixable material having a cationized amine group and a swelling polymer having an anionized carboxyl group by ionic bonding. In a container filled with a hyaluronic acid swelling polymer swollen in deionized water and a freeze-dried hyaluronic acid swelling polymer, 1 mL (5 mg / mL) of doxorubicin solution was absorbed into the fixable material having the amine group. During the absorption process, the solvent was adjusted to a pH (5-5.5) in which the two substances could be ionized, and the reaction was carried out with a stirrer for 1 hour.

[0067] After the reaction, the swelling polymer was precipitated using a centrifuge at 3000 RPM for 3 minutes, and the supernatant that did not participate in the reaction was obtained. The absorbance at 490 nm was analyzed using a UV spectrometer. As shown in Figure 5b, compared to the solution-based production method, doxorubicin immobilized on the swelling polymer showed a loading efficiency (LE) of over 70%.

[0068] [Example 4] HAMA swelling polymer with immobilized cationic drug Figures 6a and 6b are diagrams showing the manufacturing process of Example 4, which was manufactured using the manufacturing method of the present invention.

[0069] Referring to Figure 6a, Example 4 of the present invention was produced by utilizing the bonding of a fixable material having an amine group and a linker containing an aldehyde group. First, in order to provide a linker containing an aldehyde group to a hyaluronic acid methacrylate (HAMA) swelling polymer to which a methacrylate group containing an aldehyde group is bonded, HAMA was reacted with sodium periodate (Na₂O₄) for 24 hours, then crosslinked and freeze-dried to produce a dried HAMA-aldehyde swelling polymer. Subsequently, the dried HAMA-aldehyde swelling polymer was reacted with 5-aminofluorescein as the fixable material having an amine group. During the absorption process, the amine group of 5-aminofluorescein reacted with the aldehyde group of the HAMA-aldehyde swelling polymer to produce a HAMA swelling polymer to which a cationic drug was bound.

[0070] Furthermore, in Figure 6b, in order to provide a linker containing an aldehyde group to the HAMA swelling polymer, HAMA was reacted with 3-amino-1,2-propanediol, followed by crosslinking and freeze-drying to produce a swelling polymer to which the linker was bonded. Subsequently, sodium periodate (NaI04) was reacted, and then freeze-dried to produce a dried HAMA-aldehyde swelling polymer. The dried HAMA-aldehyde swelling polymer was then subjected to a step in which a 5-aminofluorescein solution was absorbed as a fixative having an amine group, thereby bonding the amine group of 5-aminofluorescein to the aldehyde group of the dried HAMA-aldehyde swelling polymer. This resulted in the production of HAMA swellable polymers in which cationic drugs were immobilized by the bonding of amine groups and aldehyde groups.

[0071] After the reaction, the swellable polymer was precipitated using a centrifuge at 3000 RPM for 3 minutes. The supernatant that did not participate in the reaction was obtained and its fluorescence at 520 nm was analyzed using a fluorescence plate reader. As shown in Figures 6c and 6d, 5-aminofluorescein immobilized on the swellable polymer showed a loading efficiency (LE) of approximately 20% compared to HAMA without a labeled linker. Figure 6e is a diagram for comparing the 5-AFL immobilization efficiency of Comparative Example 4, which was produced by the manufacturing method of Example 4, which was produced by the manufacturing method of the present invention. Referring to Figure 6e, it can be confirmed that when produced by a conventional solution-based manufacturing method, the immobilization efficiency of the object 5-AFL is very low at approximately 2%, while when produced by the freeze-dried manufacturing method using the swellable polymer of the present invention, the immobilization efficiency of 5-AFL is approximately 21%, which is significantly higher than the conventional technique.

[0072] [Example 5] HAMA swelling polymer with immobilized mercaptopurine (6-Mercaptopurine) Figure 7 is a diagram showing the manufacturing process of Example 5, which was manufactured using the manufacturing method of the present invention.

[0073] Referring to Figure 7, Example 5 of the present invention was produced by utilizing the bonding of a fixable material having a disulfide (-SS-, disulfide group) with a linker containing a thiol group (thiol, -SH). To bond the linker containing a thiol group to the HAMA swelling polymer, (2-pyridylthio)cysteamine hydrochloride was reacted, followed by crosslinking and freeze-drying to produce a dried HAMA-thiol swelling polymer. Subsequently, the dried HAMA-thiol swelling polymer was absorbed into the polymer using a solution containing mercaptopurine, a drug containing a disulfide group. During this absorption process, the disulfide group of mercaptopurine bonded to the thiol group of the HAMA-thiol swelling polymer, producing a HAMA swelling polymer with immobilized mercaptopurine.

[0074] [Example 6] Chlorambucyl-immobilized HAMA swelling polymer Figure 8 is a diagram illustrating the manufacturing process of a HAMA swelling polymer with immobilized chlorambucil produced by the manufacturing method of the present invention.

[0075] Referring to Figure 8, the polymer was manufactured by utilizing the bonding of an object containing a carboxyl group with a linker containing an amine group. First, to provide a linker containing an amine group (-NH2), cystamine was reacted with the HAMA polymer, followed by crosslinking and freeze-drying to produce a swollen polymer to which the linker was bonded. Subsequently, a solution containing chlorambucil, a drug containing a carboxyl group, was absorbed into the swollen polymer to which the linker was bonded. During this absorption process, the carboxyl group of chlorambucil bonded to the amine group of the swollen polymer, producing a swollen polymer to which chlorambucil was bonded.

[0076] [Example 7] Paclitaxel immobilized in HAMA swelling polymer Figure 9 is a diagram showing the manufacturing process of HAMA swelling polymer in which paclitaxel, produced by the manufacturing method of the present invention, is immobilized.

[0077] Referring to Figure 9, the polymer was manufactured by utilizing the bonding of an object containing a hydroxyl group with a linker containing a carboxyl group. First, to provide a linker containing a carboxyl group to the HAMA polymer, alanine was reacted and bonded, then crosslinked and freeze-dried to produce a swollen polymer to which the linker was bonded. Subsequently, the swollen polymer to which the linker was bonded was absorbed into the polymer using a solution containing paclitaxel, a drug containing a hydroxyl group (-OH). During this absorption process, the hydroxyl group of paclitaxel bonded to the carboxyl group of the swollen polymer, producing a swollen polymer to which paclitaxel was immobilized.

[0078] As shown in the examples, a dried swellable polymer containing a linker having a functional group that can bond to an object produced by the manufacturing method of the present invention can be produced by crosslinking and drying the polymer and then bonding the linker, or by bonding the linker to the polymer first, and then crosslinking and drying.

[0079] Although preferred embodiments of the present invention have been described above with reference to the present invention, those skilled in the art should understand that the present invention can be modified and altered in various ways without departing from the spirit and scope of the invention as described in the claims.

[0080] The abbreviations used in this specification are as follows:

[0081] DOTA 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid

[0082] DOTA-GA 2-(4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl)pentanedioic acid

[0083] DO2A 1,4,7,10-tetraazacyclododecane-1,7-diacetic acid

[0084] HBED-CC 3,3'-(3,3'-(ethane-1,2-diylbis((carboxymethyl)azanediyl))bis(methylene)bis(4-hydroxy-3,1-phenylene))dipropanoic acid

[0085] NOTA 1,4,7-triazonane-1,4,7-triacetic acid

[0086] p-SCN-Bn-NOTA 2-(4-isothiocyanatobenzyl)-NOTA

[0087] NODA-GA 2-(4,7-bis(carboxymethyl)-1,4,7-triazonan-1-yl)pentanedioic acid

[0088] PrP9 1,4,7-triazonane-1,4,7-tri(2-carboxyethyl)methyl phosphinic acid

[0089] DTPA Diethylenetriamine-pentaacetic acid

[0090] p-SCN-CHX-A’’-DTPA 2S-(4-isothiocyanatobenzyl)cyclohexyldiethylenetriamine-pentaacetic acid

[0091] p-SCN-CHX-B’’-DTPA 2R-(4-isothiocyanatobenzyl)cyclohexyldiethylenetriamine-pentaacetic acid

[0092] NETA (2-(4,7-bis(carboxymethyl)-1,4,7-triazonan-1-yl)ethylazane-diacetic acid

[0093] NE3TA 7-(2-(carboxymethylamino)ethyl)-1,4,7-triazonane-1,4-diacetic acid

[0094] NE3TA-Bn (7-(2-(benzyl(carboxymethyl)amino)ethyl)-1,4,7-triazonane-1,4-diacetic acid

[0095] C-NETA 2,2’-(2-(4,7-bis(carboxymethyl)-1,4,7-triazonan-1-yl)-(2-(4-isothiocyanatobenzyl))ethylazanediyl)diacetic acid

[0096] C-NE3TA 2,2’-(7-(2-(carboxymethylamino)-2-(4-isothiocyanatobenzyl)ethyl)-1,4,7-triazonane-1,4-diyl)diacetic acid

[0097] C-DOTA 2-(4-nitrobenzyl)-DOTA

[0098] 1B3M-DTPA 1-benzyl-3-methyldiethylenetriamine-pentaacetic acid

[0099] 1B4M-DTPA 1-benzyl-4-methyldiethylenetriamine-pentaacetic acid

[0100] 1M3B-DTPA 1-methyl-3-benzyldiethylenetriamine-pentaacetic acid

[0101] TETA 1,4,8,11-tetraaza-cyclotetradecane-1,4,8,11-tetraacetic acid

[0102] TE2A 1,4,8,11-tetraazacyclo-tetradecane-1,8-diacetic acid

[0103] CB-TE2A 1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4,11-diacetic acid

[0104] CB-DO2A 1,4,7,10-tetraazabicyclo[5.5.2]tetradecane-4,10-diacetic acid

[0105] DiAmSar 3,6,10,13,16,19-hexaazabicyclo[6.6.6]icosane-1,8-diamine

[0106] SarAr N-(4-aminobenzyl)-3,6,10,13,16,19-hexaazabicyclo[6.6.6]icosane-1,8-diamine

[0107] AmBaSar 4-((8-amino-3,6,10,13,16,19-hexaazabicyclo[6.6.6]icosan-1-ylamino)methyl)benzoic acid

[0108] AAZTA 6-amino-6-methylperhydro-1,4-diazepinetetraacetic acid

Claims

1. To prepare a crosslinked polymer in the form of hydrogel particles by irradiating a polymer containing hyaluronic acid methacrylate (HAMA) that includes a linker having a functional group capable of chemically bonding with radioisotopes with light, The crosslinked polymer is freeze-dried. To prepare an aqueous solution containing the radioisotope dissolved or dispersed in water, and A method for producing a carrier containing a polymer to which a radioisotope has been chemically bonded, comprising absorbing the aqueous solution into the freeze-dried, crosslinked polymer to chemically bond the radioisotope to the polymer.

2. The aforementioned radioactive isotope has an anionic functional group, The functional group of the linker is a heterocyclo compound, The manufacturing method according to claim 1, wherein the anionic functional group and the functional group of the linker undergo an electrophilic substitution reaction.

3. The production method according to claim 2, wherein the heterocyclo compound is selected from the group consisting of imidazole, pyrrole, furan, thiophene, indole, and 3,4-dihydroxyphenyl.

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