Polymeric nanoparticles for oral administration and delivery of physiologically active substances and preparation method therefor

Abstract

The present invention relates to a polymeric nanoparticle for oral administration of physiologically active substances and a preparation method therefor and, more specifically, to a carrier or polymeric hydrogel for delivery of a physiologically active substance and a preparation method therefor, wherein the carrier or polymeric hydrogel comprises: polymeric micelles including a PEG-based block copolymer prepared by a reversible addition–fragmentation chain transfer (RAFT) radical polymerization method using a polymer that has a specific functional group at the chain end thereof and a boronic acid group introduced into the backbone thereof; and a physiologically active substance that can be encapsulated in the polymeric micelles, including a type 2 diabetes drug, insulin, an anti-obesity agent, a DPP-4 inhibitor, or an anticancer agent.

Description

Polymer nanoparticles for oral administration and delivery of bioactive substances and methods for manufacturing the same

[0001] The present invention relates to polymer nanoparticles for oral administration and delivery of physiologically active substances and a method for manufacturing the same.

[0002] More specifically, the invention relates to a carrier or polymer hydrogel for delivering a bioactive substance and a method for manufacturing the same, characterized by comprising a polymer micelle containing a PEG-based block copolymer prepared by reversible addition fragmentation chain transfer (RAFT) radical polymerization of a polymer having special functional groups at the chain ends and boronic acid groups introduced into the main chain, and a bioactive substance that can be encapsulated within the polymer micelle, such as a type 2 diabetes drug, insulin, an obesity treatment drug, a DPP-4 inhibitor, or an anticancer drug.

[0003]

[0004] The present application claims priority based on Korean Patent Application No. 10-2024-0201880 filed on December 31, 2024, and all contents disclosed in the specification and drawings of said application are incorporated into the present application.

[0005] Recently, while new drug development is important, the efficient utilization of existing commercialized drugs has become a hot topic in the field of medical science. In particular, all drugs are broadly classified into four categories based on their solubility in aqueous solutions and gastrointestinal permeability; this is referred to as the Biopharmaceutics Classification System (BCS). Meanwhile, alongside the remarkable advancements in medical science, new protein and peptide-based drugs are gaining significant attention. However, these drugs all fall under Class 4 (low solubility and low permeability) in the BCS. Despite their excellent efficacy, their low solubility inevitably leads to low efficacy and low permeability (low absorption), resulting in drug overdose. Furthermore, as these are drugs most preferred by patients for oral administration, their market potential is projected to reach 185 trillion won by 2027.

[0006]

[0007] In particular, GLP-1 receptor agonists (GLP-1 RAs), recently developed as injectable treatments for type 2 diabetes, have been repurposed as obesity treatments and reborn as expensive blockbusters in the new biopharmaceutical field; however, the high cost has emerged as a social issue in many countries. Furthermore, although the aforementioned GLP-1 RAs have obtained U.S. FDA approval (Rybelsus®) for oral formulations to improve administration convenience, their efficacy (0.4% - 1%) is reported to be very low (see Ke, Z. Ma, Q.; Ye, X.; Wang, Y.; Jin, Y.; Zhao, X.; Su, Z. Biochem. Pharmacol., 2024, 229, 116471). To address these problems, it is crucial to generalize these drugs into oral formulations and develop new formulations that can also be applied to the development of targeted anticancer therapies.

[0008]

[0009] In particular, insulin and glucagon-like peptide-1 receptor agonists (GLP-1RAs), which are required medications for patients with type 1 and type 2 diabetes, are mostly administered via injection; since this is the most critical step diabetic patients wish to avoid, a transition to oral administration is inevitable. In this regard, examples of the use of materials for nano-drug delivery tablets and capsules containing encapsulated drugs for oral administration have already been well described in other literature (see KA Rubean, M. Rafiullah, S. Jayavanth, Expert Opinion on Drug Delivery, 2015, Vol. 13, No. 2, 1-15, U.S. Patent Publication US 9,101,547 B2).

[0010]

[0011] In addition, the preparation of semaglutide, a glucagon-like peptide-1 receptor derivative used as an oral treatment for type 2 diabetes, and its mechanism of intestinal absorption are well described in the literature (see VR Aroda, J. Blonde, RE Pratley, Rev. Endocr. Metab. Disord., 2022, Vol. 23, No. 5, pp. 979-994). In particular, although oral administration of semaglutide is a patient-friendly drug, its low gastrointestinal and intestinal absorption rates and precautions are well described in the literature (Z. Ke, Q. Ma, X. Ye, Y. Wang, Y. Jin, X. Zhao, Z. Su, Biochem. Pharmacol., 2024, 229, 116471).

[0012]

[0013] In general, for use as an orally administered material, the development of substances capable of delivering drug-containing tablets or capsules from the highly acidic stomach to the end of the small intestine, where the drug is absorbed, without any degradation is inevitable. In particular, they must possess the characteristics to overcome the many barriers encountered while passing through the digestive system and deliver the drug to the required site. Many researchers have well explained these methods of overcoming obstacles through their results (see C. Damge, CP Reis, Expert Opinion on Drug Deliver 2008, 5, 45-68; CB Woitiski, RA Carvalho, AJ Ribeiro, RJ Neufeld, F. Veiga, Biodrugs 2008, 22, 223-237; A. des Rieux, V. Fievez, M. Garinot, Y.-J. Schneider, V. Preat, Journal of Controlled Release 2006, 116, 1-27).

[0014]

[0015] More specifically, among the various obstacles to be overcome in oral administration, the most difficult factor is the special ability to protect the administered drug from strong acid during passage through the stomach within about 0.5 to 4 hours, and the resolution of resistance and stability of drug carriers caused by various enzymes (see A. Sonik and R. Augustine, Advanced Drug Delivery Reviews 2016, 103, 105-120).

[0016]

[0017] In this regard, methods for preparing hydrogels and the like using natural polymers such as hyaluronic acid and dextran for use as drug delivery systems are well known in various literature (see C. Qian, T. zhang, J. Gravesande, C. Baysah, X. Song, J. Xing, International Journal of Biological Macromolecules, 2019, Vol. 123, 140-148; P. Sedova, R. Buffa, P. Silhar, L. Kovarova, H. Vagnerova, J. bednarik, I. Basarabova, L. Hejlova, I. Scigalkova, M. Simek, V. Velebny, Carbohydrate Polymers, 2019, Vol. 216, 63-71).

[0018]

[0019] Although research on various drug delivery systems is underway, efficient absorption of the drug within the small intestine is essential for effective drug efficacy through oral administration, and there is an increasing need for the development of more effective methods to achieve this.

[0020] The present invention was developed to solve the above problems and aims to provide a block copolymer capable of forming a nanocomposite with glucagon-like peptide-1 receptor agonists (GLP-1 RAs) such as exenatide, semaglutide, tirzeptide, or liraglutide, insulin, obesity treatment agents, DPP-4 inhibitors, or anticancer agents, a polymer micelle containing the block copolymer, a carrier for delivering a bioactive substance or a polymer hydrogel containing a bioactive substance that can be encapsulated within the polymer micelle, and a method for manufacturing the same.

[0021] In order to solve the above problem, the present invention provides a carrier for delivering physiologically active substances or a polymer hydrogel and a method for preparing the same by reacting a substance having an aldehyde group at the chain end with various protein drugs using an organic material having an aldehyde group, a functional polymer having an aldehyde group at the chain end of poly(ethylene glycol; PEG), a tertiary butoxy-polyethylene glycol-cyclic hydrazide (t-BuO-PEG-bP(cyclic hydrazide)) polymer, and poly(cyclic hydrazide)-b-PEG-b-poly(cyclic hydrazide) synthesized by a triblock copolymer preparation method.

[0022]

[0023] In addition, the present invention provides a method for preparing an oral drug precursor by preparing PEG-b-Poly(cyclic hydrazide) having a thiol group at the chain end and reacting it with various aldehyde organics, proteins, and peptide drugs.

[0024]

[0025] In addition, the present invention provides a method for preparing a nanogel by reacting a copolymer having a cyclic hydrazide group synthesized by a block copolymer manufacturing method with natural polymers, such as hyaluronic acid and dextran, which are oxidized to have an aldehyde group.

[0026]

[0027] In addition, the present invention provides a special PEG copolymer having an acylhydrazone group and a boronic acid group capable of a dynamic reversible reaction with the drug, which enables oral administration of injectable drugs and water-insoluble drugs, and a method for manufacturing the same, in which a drug for diabetes and obesity is encapsulated in the nanoparticle.

[0028]

[0029] In addition, the present invention provides a method for preparing a nanoscale hydrogel having an absorption enhancer group by reacting a bile acid with a functional polyethylene glycol.

[0030]

[0031] In addition, the present invention provides a method for preparing a block copolymer composed of polyethylene glycol and a homopolymer or copolymer of a monomer having a boronic acid group in its molecular structure (methyl methacrylamidophenylboronic acid: MMAPBA, etc.), and for preparing a complex with peptides and protein drugs having a primary amine group at the chain end, such as commercial insulin, exenatide, semaglutide, tirzeptide, or liraglutide.

[0032]

[0033] In addition, the present invention provides a method for preparing a stimulus-responsive smart hydrogel in which a phenylboronic acid (PBA) group is reversibly covalently bonded to a polyethylene glycol and a poly(cyclic hydrzaide) block copolymer (R-PEG-bP(CHz)x).

[0034]

[0035] In addition, the present invention provides a method for preparing complexes with peptides and protein drugs having a phenylboronic acid (PBA) group on a poly(cyclic hydrzaide) chain and a primary amine group at the end of the chain.

[0036]

[0037] In addition, the present invention provides a method for manufacturing nanoparticles by mixing complexes containing various GLP-1RAs with commercial peptides and protein drugs.

[0038] As described above, the present invention enables the preparation of a polymer hydrogel in the form of a nanocomposite with glucagon-like peptide-1 receptor agonists (GLP-1 RAs) such as exenatide, semaglutide, tirzeptide, or liraglutide, insulin, obesity treatments, DPP-4 inhibitors, or anticancer drugs by preparing a PEG-based block copolymer using a reversible addition fragmentation chain transfer (RAFT) radical polymerization method with a polymer having special functional groups at the chain ends and boronic acid groups introduced into the main chain.

[0039]

[0040] In addition, the present invention has the advantage of significantly increasing the intestinal absorption rate (~45%) of orally administered drugs by very effectively introducing bile acids (cholic acid: CLA, deoxycholic acid: DOA, glycolic acid: GCA, taurocholic acid: TCA, or folic acid: FLA, etc.) as special functional groups at the chain ends onto the surface of nano drug particles.

[0041]

[0042] In addition, in the present invention, by using folic acid (FLA) as a functional group to introduce an anticancer drug (doxorubicin; Dox) in the form of a nanocomposite, it was possible to apply this to the manufacture of a site-specific anticancer drug.

[0043]

[0044] In addition, the present invention has the advantage of being able to significantly improve the intestinal stability and absorption rate in the small intestine of orally administered drugs, as it is possible to manufacture dynamic hydrogels by reacting PEG-based block copolymers having cyclic hydrazide or acylhydrazone with PEG functionalized with aldehyde groups and natural polymers, and by using these materials, it is possible to manufacture various types of polymer prodrugs and nanocomposites.

[0045] Figure 1 conceptually illustrates the structure of a hydrogel nanoparticle containing a drug according to one embodiment of the present invention.

[0046] Figure 2 shows the reversible gelation behavior of a hydrogel prepared according to one embodiment of the present invention.

[0047] Figure 3 shows the results of a pH-responsive anticancer drug release test of a hydrogel prepared according to one embodiment of the present invention.

[0048] FIG. 4 schematically shows hydrogel nanoparticles encapsulated with a bioactive substance prepared according to one embodiment of the present invention.

[0049] Hereinafter, the present invention will be described in detail. Terms and words used in this specification and claims should not be interpreted as being limited to their ordinary or dictionary meanings, and should be interpreted in a meaning and concept consistent with the technical spirit of the present invention, based on the principle that the inventor can appropriately define the concept of the terms to best describe his invention.

[0050]

[0051] The term “dynamic covalent bond,” used throughout this specification, means that the formation of a hydrazone is reversibly achieved through a reaction between the hydrazide functional group of a cyclic hydrazide block of a PEG block copolymer and the aldehyde (>C=O) functional group of a compound containing an aldehyde group in its molecular structure, and that the formation and separation of the covalent bond are reversibly achieved.

[0052]

[0053] In order to solve the above problem, the present invention provides a block copolymer characterized by comprising: a polyethylene glycol block having a bile acid (BA) group at the chain end; and a polymer block composed of a homopolymer or copolymer of a monomer having a boronic acid group in its molecular structure.

[0054]

[0055] According to one embodiment of the present invention, the bile acid (BA) may be any one of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA), or folic acid (FLA).

[0056]

[0057] According to one embodiment of the present invention, the block copolymer may be a block copolymer characterized by having a chemical structure represented by the following chemical formula A.

[0058] <Chemical Formula A>

[0059]

[0060] In the above chemical formula A, BA is a residue of bile acid, and the bile acid is any one of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA) or folic acid (FLA), R1 represents H- or CH3-, m is an integer from 1 to 10 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0061]

[0062] In addition, the present invention provides a polymer micelle characterized by including the block copolymer.

[0063] According to one embodiment of the present invention, the polymer micelle may contain a physiologically active substance inside.

[0064]

[0065] In addition, the present invention provides a carrier for delivering a bioactive substance or a polymer hydrogel characterized by comprising the polymer micelle; and a bioactive substance that can be encapsulated within the polymer micelle.

[0066]

[0067] According to one embodiment of the present invention, the bioactive substance may be one or more selected from type 2 diabetes drugs, insulin, obesity treatment drugs, DPP-4 inhibitors, or anticancer drugs, and the type 2 diabetes drug may be one or more selected from glucagon-like peptide-1 receptor agonists (GLP-1 RAs), exenatide, semaglutide, tirzeptide, or liraglutide having amine groups at the chain ends.

[0068]

[0069] According to one embodiment of the present invention, the carrier for delivering a bioactive substance or the polymer hydrogel may further include a block copolymer or triblock copolymer of a polymer having a polyethylene glycol block and a cyclic hydrazide as repeating units, and may also further include a compound containing two or more aldehyde groups in its molecular structure.

[0070]

[0071] According to one embodiment of the present invention, the compound containing an aldehyde group in the molecular structure may be one or more selected from aldehyde-treated polyethylene glycol, aldehyde-treated hyaluronic acid, aldehyde-treated dextran, or aldehyde-treated cellulose.

[0072]

[0073] According to one embodiment of the present invention, the performance of drug delivery and delivery of the carrier or polymer hydrogel for delivering bioactive substances can be optimized by controlling the length and composition ratio of the block copolymer of a polymer having polyethylene glycol blocks and cyclic hydrazides as repeating units, as well as the mixing ratio of compounds containing two or more aldehyde groups in their molecular structure.

[0074]

[0075] According to one embodiment of the present invention, the hydrogel includes a molecular structure capable of forming a dynamic covalent bond, so there is an advantage that it can be directly reacted and encapsulated within the hydrogel for the introduction of a receptor, etc., for drug loading or targeting of cancer cells.

[0076]

[0077] According to one embodiment of the present invention, the hydrogel contains a drug that is fixed through chemical covalent bonds or ionic bonds, but can be kinetically separated and bonded easily by external and internal stimuli, thereby enabling effective delivery of the drug without deformation.

[0078]

[0079] A carrier for delivering physiologically active substances or a polymer hydrogel according to one embodiment of the present invention and a method for manufacturing the same may use a mixture further comprising a polymer containing organic substances and drugs represented by the following chemical formulas 1 to 23.

[0080]

[0081] <Chemical Formula 1>

[0082]

[0083] In the above chemical formula 1, R1 and R2 are phenoxyaldehyde (4-hydroxybenzaldehyde) or ethylaldehyde, which may be the same or different, and n is an integer from 20 to 250 as a repeating unit.

[0084] <Chemical Formula 2>

[0085]

[0086] In the above chemical formula 2, R1 to R3 are all aldehyde groups, or two are aldehyde groups and the remainder is hydrogen.

[0087]

[0088] In addition, according to one embodiment of the present invention, in addition to the above formula 2, commercially available terephthalaldehyde, benzene-1,3,5-tricarboxyaldehyde, or isophthalaldehyde may also be used.

[0089] <Chemical Formula 3>

[0090]

[0091] In the above chemical formula 3, R is 4-hydroxybenzaldehyde, and n is a repeating unit, an integer from 5 to 110.

[0092] According to one embodiment of the present invention, the compound of Formula 3 may be used by preparing star-shaped poly(ethylene glycol): 4-arm-PEG obtained by polymerizing ethylene oxide in a polar solvent such as dimethylsulfoxide (DMSO) or dimethylformamide using pentaerythritol and KOH, and then aldehydeizing it.

[0093]

[0094] <Chemical Formula 4>

[0095]

[0096] The above chemical formula 4 represents oxidized (or aldehyde-treated) chitosan, and its molecular weight is 1.0 x 10⁵ to 2.0 x 10⁶ Daltons.

[0097]

[0098] <Chemical Formula 5>

[0099]

[0100] The above chemical formula 5 represents oxidized (or aldehyde-treated) hyaluronic acid, and its molecular weight is 1.0 x 10⁵ to 2.0 x 10⁶ Daltons.

[0101]

[0102] <Chemical Formula 6>

[0103]

[0104] The above chemical formula 6 represents an oxidized (or aldehyde-treated) dextran, and its molecular weight is 1.0 x 10⁵ to 2.0 x 10⁶ Daltons.

[0105]

[0106] <Chemical Formula 7>

[0107]

[0108] In the above chemical formula 7, m is an integer from 1 to 4 as a repeating unit, and n is an integer from 10 to 250 as a repeating unit.

[0109]

[0110] <Chemical Formula 8>

[0111]

[0112] In the above chemical formula 8, BA is a residue of bile acid, and the bile acid is any one of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA) or folic acid (FLA), k is an integer from 0 to 2 as a repeating unit, m is an integer from 1 to 3 as a repeating unit, and n is an integer from 20 to 250 as a repeating unit.

[0113]

[0114] <Chemical Formula 9>

[0115]

[0116] In the above chemical formula 9, BA is a residue of a bile acid, the bile acid being any one of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA) or folic acid (FLA), R is a hydrogen group (-H) or a methyl group (CH3-), m is an integer from 3 to 50 as a repeating unit, and n is an integer from 20 to 250 as a repeating unit.

[0117]

[0118] <Chemical Formula 10>

[0119]

[0120] In the above chemical formula 10, R1 is one of a methoxy, t-butoxy, thiol, or hydroxyl group, R2 is a methyl group (CH3-) or hydrogen (H), n is an integer from 20 to 250 as a repeating unit, and m is an integer from 3 to 20 as a repeating unit.

[0121]

[0122] <Chemical Formula 11>

[0123]

[0124] In the above chemical formula 11, R represents a methoxy, t-butoxy, or hydroxyl group, n is an integer from 20 to 250 as a repeating unit, and m is an integer from 1 to 50.

[0125]

[0126] <Chemical Formula 12>

[0127]

[0128] In the above chemical formula 12, R is one of methoxy, t-butoxy, or hydroxyl groups, and n is an integer from 20 to 250 as a repeating unit.

[0129]

[0130] <Chemical Formula 13>

[0131]

[0132] In the above chemical formula 13, R is one of methoxy, t-butoxy, or hydroxyl groups, and n is an integer from 20 to 250 as a repeating unit.

[0133]

[0134] <Chemical Formula 14>

[0135]

[0136] In the above chemical formula 14, BA is a residue of bile acid, and the bile acid is any one of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA) or folic acid (FLA), R1 represents H- or CH3-, m is an integer from 1 to 10 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0137]

[0138] <Chemical Formula 15>

[0139]

[0140] In the above chemical formula 15, R is one of a bile acid, methoxy, t-butoxy, thiol, or hydroxyl group, m is an integer from 1 to 10 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0141]

[0142] <Chemical Formula 16>

[0143]

[0144] In the above chemical formula 16, BA is a residue of bile acid, and the bile acid is any one of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA) or folic acid (FLA), R1 is one of methoxy or t-butoxy, m is an integer from 1 to 10 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0145]

[0146] <Chemical Formula 17>

[0147]

[0148] In the above chemical formula 17, R1 is either methoxy or t-butoxy, m is an integer from 1 to 10 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0149]

[0150] <Chemical Formula 18>

[0151]

[0152] In the above chemical formula 19, R is one of a methoxy, t-butoxy, thiol, or hydroxyl group, k is an integer from 1 to 5 as a repeating unit, o is an integer from 0 to 3 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0153]

[0154] <Chemical Formula 19>

[0155]

[0156] In the above chemical formula 19, R is one of a methoxy, t-butoxy, or hydroxyl group, and n is a repeating unit, an integer from 20 to 250.

[0157]

[0158] In addition, the present invention provides a carrier for delivering a bioactive substance or a polymer hydrogel characterized by comprising the polymer micelle; and a bioactive substance that can be encapsulated within the polymer micelle.

[0159]

[0160] According to one embodiment of the present invention, the bioactive substance may be one or more selected from type 2 diabetes drugs, insulin, obesity treatment drugs, DPP-4 inhibitors, or anticancer drugs, and the type 2 diabetes drug may be one or more selected from glucagon-like peptide-1 receptor agonists (GLP-1 RAs), exenatide having an amine group at the chain end, semaglutide, tirzeptide, or liraglutide, and may be included inside a bioactive substance delivery carrier or a polymer hydrogel while forming a polymer drug nanocomposite in the form represented by the following chemical formulas 20 to 22.

[0161]

[0162] <Chemical Formula 20>

[0163]

[0164] In the above chemical formula 20, GLP-1RA is a peptide drug and is one of a material having an amine group at the chain end, such as exenatide, semaglutide, tirzeptide, or liraglutide, or insulin, R1 is one of a methoxy, t-butoxy, thiol, or hydroxyl group, m is an integer from 1 to 10 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0165]

[0166] <Chemical Formula 21>

[0167]

[0168] In the above chemical formula 21, GLP-1RA is a peptide drug and is one of a material having an amine group at the chain end, such as exenatide, semaglutide, tirzeptide, or liraglutide, or insulin, R1 is one of a methoxy, t-butoxy, thiol, or hydroxyl group, m is a repeating unit and is an integer from 1 to 10, and n is a repeating unit and is an integer from 10 to 200.

[0169]

[0170] <Chemical Formula 22>

[0171]

[0172] In the above chemical formula 22, Dox can be selected from anticancer agents such as doxorubicin, R is one of methoxy, t-butoxy, thiol, or hydroxyl groups, k is an integer from 1 to 5 as a repeating unit, m is an integer from 0 to 3 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

[0173]

[0174] In the carrier or polymer hydrogel for delivering a bioactive substance according to one embodiment of the present invention and the method for manufacturing the same, hydrogel nanoparticles encapsulated with a bioactive substance can be manufactured from a mixture containing a polymer containing an organic substance represented by Chemical Formulas 1 to 22 and a drug, etc., as shown in Chemical Formula 23 below.

[0175]

[0176] <Chemical Formula 23>

[0177]

[0178]

[0179]

[0180]

[0181] In the above chemical formula 23, GLP-1RA is a peptide drug and is one of a material having an amine group at the chain end, such as exenatide, semaglutide, tirzeptide, or liraglutide, or insulin, and the ~~~ chain is one of PEG or aldehyde hyaluronic acid, aldehyde dextran, or aldehyde cellulose, and GCA is one selected from bile acids (BA), such as glycolic acid (GCA).

[0182]

[0183] FIG. 1 conceptually illustrates the structure of a hydrogel nanoparticle containing a drug according to one embodiment of the present invention. According to one embodiment of the present invention, the structure of the hydrogel nanoparticle is possible to include a 4-arm PEG (tetrafunctionalized 4-arm PEG) having four functional groups, a heterobifunctional PEG, a bifunctional PEG, and a bile acid or folate, along with a type 2 diabetes drug, insulin, an obesity treatment drug, a DPP-4 inhibitor, or an anticancer drug.

[0184]

[0185] Figure 2 shows the reversible gelation behavior of a hydrogel prepared according to one embodiment of the present invention.

[0186] (cyclic hydrazide) according to one embodiment of the present invention 2.5 -PEG1.7K-(cyclic hydrazide) 2.5 ((CHz) 2.5 -PEG1.7K-(CHz) 2.5A solution of 0.12 g of ) dissolved in 1.0 mL of DMSO, a solution of 0.8 g of aldehyde-PEG4K-aldehyde (OHCBz-PEG4K-BzCHO) prepared by the same method as Preparation Example 8 dissolved in 1.5 mL of DMSO, and a solution of 70.2 g of t-BuO-PEG5K-(CHz) prepared similarly by dissolving in 1 mL of DMSO were added to a 25 mL vial and mixed by shaking. Subsequently, the pH was adjusted to 8.0 using PBS (pH = 7.2) solution and NaOH (0.1 M) aqueous solution, and the vial temperature was maintained at 70 ℃ for 3 hours. Gelation occurred during this process. Then, H2SO4 aqueous solution (0.1 M) was added to adjust the vial pH to 4.0, forming a clear purple solution (sol), and the formation of a gel within the vial was observed again by adjusting the pH to 8.0 with NaOH aqueous solution.

[0187]

[0188]

[0189] Figure 3 shows the results of a pH-responsive anticancer drug release test of a hydrogel prepared according to one embodiment of the present invention. The binding of the hydrazone according to one embodiment of the present invention enabled the release of an anticancer drug under conditions of pH 4.5, and the result could be observed as a change in color as shown in the photograph.

[0190]

[0191] FIG. 4 schematically illustrates hydrogel nanoparticles containing a bioactive substance prepared according to one embodiment of the present invention. It conceptually shows the release of the bioactive substance contained inside to the outside of the hydrogel nanoparticles due to external stimuli such as changes in pH or temperature.

[0192]

[0193] As described above, the present invention enables the preparation of a polymer hydrogel in the form of a nanocomposite with glucagon-like peptide-1 receptor agonists (GLP-1 RAs) such as exenatide, tirzeptide, or liraglutide, insulin, obesity treatments, DPP-4 inhibitors, or anticancer drugs by preparing a PEG-based block copolymer using a reversible addition fragmentation chain transfer (RAFT) radical polymerization method with a polymer having special functional groups at the chain ends and boronic acid groups introduced into the main chain.

[0194]

[0195] In addition, the present invention has the advantage of significantly increasing the intestinal absorption rate (~45%) of orally administered drugs by very effectively introducing bile acids (cholic acid: CLA, deoxycholic acid: DOA, glycolic acid: GCA, taurocholic acid: TCA, or folic acid: FLA, etc.) as special functional groups at the chain ends onto the surface of nano drug particles.

[0196]

[0197] In addition, in the present invention, by using folic acid (FLA) as a functional group to introduce an anticancer drug (doxorubicin; Dox) in the form of a nanocomposite, it was possible to apply this to the manufacture of a site-specific anticancer drug.

[0198]

[0199] In addition, the present invention has the advantage of being able to significantly improve the intestinal stability and absorption rate in the small intestine of orally administered drugs, as it is possible to manufacture dynamic hydrogels by reacting PEG-based block copolymers having cyclic hydrazide or acylhydrazone with PEG functionalized with aldehyde groups and natural polymers, and by using these materials, it is possible to manufacture various types of polymer prodrugs and nanocomposites.

[0200]

[0201] In addition, the present invention provides a method for manufacturing a polymer hydrogel, characterized by comprising the steps of: preparing a first block copolymer composed of an ethylene glycol monomer block having a bile acid (BA) group at the chain end and a monomer block having a boronic acid group; preparing a second block copolymer composed of an ethylene glycol monomer block and a cyclic hydrizide block; and forming polymer micelles from a mixture of the first block copolymer, the second block copolymer, and a compound containing an aldehyde group within a molecular structure capable of forming a dynamic covalent bond with the cyclic hydrizide block of the second block copolymer.

[0202]

[0203] According to one embodiment of the present invention, the step of preparing the first block copolymer may include: preparing a macroinitiator comprising a polyethylene glycol block; polymerizing a monomer having a boronic acid group in its molecular structure with the macroinitiator; and introducing a bile acid (BA) group to the end of the polyethylene glycol block.

[0204]

[0205] According to one embodiment of the present invention, the bile acid (BA) may be one or more of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA), or folic acid (FLA).

[0206]

[0207] According to one embodiment of the present invention, the step of forming the polymer micelles may include the step of further mixing a bioactive substance into the mixture, and the bioactive substance may be one or more selected from type 2 diabetes drugs, insulin, obesity treatment drugs, DPP-4 inhibitors, or anticancer drugs.

[0208]

[0209] According to one embodiment of the present invention, the step of preparing the second block copolymer may include: preparing a block copolymer of polyethylene glycol and poly(N-phenylmaleimide); and reducing the poly(N-phenylmaleimide) block using hydrizine.

[0210]

[0211] According to one embodiment of the present invention, the compound containing an aldehyde group in the molecular structure may be one or more selected from aldehyde-treated polyethylene glycol, aldehyde-treated hyaluronic acid, aldehyde-treated dextran, or aldehyde-treated cellulose.

[0212]

[0213] Hereinafter, preferred embodiments of the present invention are described in detail with reference to the accompanying drawings, so that those skilled in the art can easily implement them. In particular, the technical concept, core components, and operation of the present invention are not limited by this. Furthermore, the content of the present invention may be implemented in various different forms of equipment and is not limited to the embodiments and examples described herein.

[0214]

[0215] Hereinafter, preferred embodiments of the present invention are described in detail with reference to the accompanying drawings, so that those skilled in the art can easily implement them. In particular, the technical concept, core components, and operation of the present invention are not limited by this. Furthermore, the content of the present invention may be implemented in various different forms of equipment and is not limited to the embodiments and examples described herein.

[0216]

[0217] <Example 1> Preparation of t-BuO-PEO

[0218] Poly(ethylene glycol: PEG) was prepared under high vacuum or high pressure. First, the necessary potassium t-butoxide (t-BuO₂) was prepared under an argon stream. -+K) was injected as an initiator into a high-pressure reactor (2 L), and 20 mL of dimethylformamide (DMF) was injected. Afterward, the argon gas inside the reactor was completely removed using a vacuum pump. The temperature of the reactor was gradually raised to 50 o While raising the temperature to C, ethylene oxide (EO) monomer gas (molecular weight: 3,400 g / mol) was injected and stirred until 2 atmospheres were achieved. After stopping the EO injection and completing the reaction, the product was discharged through a valve to solidify. The t-BuO-PEO thus prepared was dissolved in methanol and precipitated in an excess amount of diethyl ether to obtain a white powder. The molecular weight of the obtained polymer was GPC and 1 The molecular weight was 3,400 g / mol according to 1H NMR analysis, and the molecular weight distribution was 1.07 according to GPC analysis.

[0219]

[0220] <Example 2> Preparation of MeO-PEG

[0221] Sodium methoxide (CH3O) in a manner similar to that in Example 1 above -+ Na) initiator was injected into the high-pressure reactor (2 L), and after injecting 20 mL of DMF, the argon gas inside the reactor was completely removed using a vacuum pump. The temperature of the reactor was gradually raised to 50 o While raising the temperature to C, ethylene oxide (EO) monomer gas (molecular weight: 3,400 g / mol) was injected and stirred until 2 atmospheres were achieved. After stopping the EO injection and completing the reaction, the mixture was discharged through a valve to solidify. The MeO-PEG thus prepared was dissolved in methanol and precipitated in an excess amount of diethyl ether to obtain a white powder. The molecular weight of the obtained polymer was GPC and 1 The molecular weight was 3,400 g / mol according to 1H NMR analysis, and the molecular weight distribution was 1.06 according to GPC analysis.

[0222]

[0223] <Example 3> Preparation of 4-arm PEG

[0224] For the synthesis of 4-arm PEG in a manner similar to Example 1, pentaerythritol (4 x OH) (10 mmol), K2CO3 (0.1 mmol), and 20 mL of DMF were injected into a 2 L reactor under an argon stream, and the argon gas inside the reactor was completely removed using a vacuum pump to create a vacuum. Polymerization was carried out in the same manner as in Example 1. The 4-arm PEG thus prepared was dissolved in methanol and precipitated in an excess amount of diethyl ether to recover the powder. The molecular weight of the obtained polymer (MPEE4K) was determined by GPC and 1 The molecular weight was 4,100 g / mol according to H NMR analysis, and the molecular weight distribution was 1.08 according to GPC analysis.

[0225]

[0226] <Example 4> Preparation of t-BuO-PEG3.4k-P(N-PMI)3

[0227] 17 g (0.005 mol) of the sample obtained in Example 1 and 3.5 g (0.025 mol) of potassium carbonate (K2CO3) (FW = 138.205) were injected into a 1 L three-necked flask, and 300 mL of acetone was added. Then, 2.60 g (0.015 mol) of N-phenylmaleimide (N-PMI) (FW = 173.17 g / mol) was dissolved in 50 mL of acetone and injected into the main reactor. At room temperature (25 o C) The reaction was carried out for 24 hours under an argon gas stream with stirring. To terminate the reaction, 5 mL of 0.1 M HCl methanol solution was added and stirred for about 2 hours, after which the mixture was precipitated in excess diethyl ether to obtain a pale pink powder, which was dried at room temperature in a vacuum oven for 48 hours. The yield was over 99 wt% (t-BuO-PEG3.4k-P(N-PMI)3).

[0228]

[0229] <Example 5> Preparation of t-BuO-PEG3.4K-b-Poly(CHz)3

[0230] t-BuO-PEG3.4K-b-poly(N-PMI)3(M prepared in Example 4 n 10 g (= 3,920 g / mol) was injected into a 1 L round-necked three-necked flask, 200 mL of dimethylsulfoxide (DMSO) was injected into the reactor, and the reactor temperature was set to 120°C under an argon stream. o The temperature was raised to C and completely dissolved. After injecting 18.2 mL of hydrazine hydrate (FW = 50.06; H2N-NH2·H2O; 50~60 wt%) ([H2N-NH2] / [N-PMI] = 8 / 1; 10.01 g), 120 o The reaction was carried out at C for 24 hours, and then 3.0 mL of HCl (1.0 M) was injected using a syringe under an argon stream and reacted for another 24 hours. Afterward, the reaction temperature was lowered to room temperature, 40 mL of THF was injected, and the reaction was terminated. The solution was converted to a concentrated solution using an evaporator and precipitated in excess diethyl ether. The precipitate was dried in a vacuum oven at room temperature for 24 hours and then stored under an inert gas atmosphere. The yield was over 98% (t-BuO-PEG3.4K-b-Poly(CHz)3).

[0231]

[0232] <Example 6> P(N-PMI) 2.5 -b-PEG4K-bP(N-PMI) 2.5 manufacturing

[0233] Commercial HO-PEG-OH (M n20 g of potassium carbonate (K2CO3; FW = 138.205) and 1.5 g of Dongnam Synthetic Co., Ltd. were injected into a 1 L three-necked flask, and 350 ml of acetone was added. Under an argon stream, the reactor temperature was set to 40 o The temperature was raised to C to completely dissolve the reactants, and then the reactor temperature was lowered back to room temperature. Subsequently, 4.33 g (0.025 mol) of N-phenylmaleimide (N-PMI) (FW = 173.17 g / mol) was dissolved in 50 ml of acetone and injected into the main reactor. The reaction was carried out for 48 hours at room temperature under an argon atmosphere. To terminate the reaction, 2 mL of HCl / methanol (1 / 5 mL / mL) was injected into the reactor and stirred for 2 hours. The mixture was then precipitated in excess diethyl ether to obtain a pink powder. The powder was dried in a vacuum oven at room temperature for 48 hours. The yield was over 98%.

[0234]

[0235] <Example 7> P(CHz) 2.5 -b-PEG4K-bP(CHz) 2.5 manufacturing

[0236] P(N-PMI) prepared in Example 6 2.5 -b-PEG4K-bP(N-PMI) 2.5 (M n 10 g (= 4,866 g / mol) was measured and injected into a 1 L three-necked round-bottom flask in a manner similar to Example 5, and 200 mL of DMSO was injected into the reactor. Under an argon stream, the reactor temperature was set to 120 o The temperature was raised to C and completely dissolved. After adding 18.2 mL of hydrazine hydrate (FW = 50.06; H2N-NH2·H2O; 50~60 wt%) ([H2N-NH2] / [N-PMI] = 8 / 1; 10.01 g), 120 oThe reaction was carried out at C for 24 hours, and then 3.0 mL of HCl (1.0 M) was injected using a syringe under an argon stream and reacted for another 24 hours. Afterward, the reaction temperature was lowered to room temperature, 40 mL of THF was injected, and the reaction was terminated. The solution was converted to a concentrated solution using an evaporator and precipitated in excess diethyl ether (P(CHz) 2.5 -b-PEG4K-bP(CHz) 2.5 ; pale yellow -> contact with air -> dark purple). The dark purple precipitate was dried in a vacuum oven at room temperature for 24 hours and then stored under inert gas. The yield was over 98%.

[0237]

[0238] <Example 8> Preparation of Tosyl-PEG2K-Tosyl

[0239] A solution of 10 g of HO-PEG2K-OH (0.005 mol; 0.01 mol; [OH] base) and 2.0 g of K2CO3 (0.0145 mol) dissolved in 70 mL of dichloromethane (DCM) was injected into a 100 mL dropping funnel and set up in a 1 L round-necked three-necked flask. 160 mL of DCM was injected into the reactor, 55.18 g of p-toluenesulfonyl chloride (FW = 190.65) (0.29 mol; 20 eq) was injected into the flask under an argon stream, and triethylamine (TEA: FW = 101.193; d= 0.7255; 5.63 ml; 4.04 mol) was injected into the reactor and thoroughly dissolved. The polymer solution in the set dropping funnel was injected into the reactor (10 mL / min). A heating mantle was used for temperature control, and the reaction was carried out while stirring with a magnetic stirring bar. The reactor temperature was 40 oThe reaction was carried out for 48 hours under an argon stream while maintaining the temperature. After terminating the reaction by lowering the reactor temperature to room temperature, the reaction mixture was transferred to a separating funnel, washed twice with a saturated aqueous solution of ammonium chloride, and then separated into a DCM solution to extract the TEA complex once with an aqueous solution of NaCl. Subsequently, the DCM solution was placed in a 1 L flask, and an appropriate amount of anhydrous magnesium sulfate (MgSO4) was added to absorb moisture from the solution. The DCM solution was then filtered, and a portion of the DCM in the filtrate was evaporated using a rotary evaporator and precipitated in an excess amount of diethyl ether (1.0 L). After filtration, the powder was dried in a vacuum oven at room temperature for at least 24 hours. The yield was over 98 wt%. The tosylation rate was over 99 mol% (Tosyl-PEG2K-Tosyl).

[0240]

[0241] <Example 9> Preparation of OHC-Bz-PEG2K-Bz-CHO

[0242] 4-hydroxybenzaldehyde (FW = 122.12; 10 eq.; 0.61 g), potassium carbonate (FW = 138.205; 20 eq.; 1.332 g), and triethylamine (FW = 101.19; d = 0.7255 g / mL; 20 eq.; 1.4 mL) were placed in a 500 mL three-necked round-bottom flask reactor and completely dissolved in 200 mL of dimethylformamide (DMF), and the reactor temperature was raised to 130°C using a heating mantle. o It was raised to C. As in Example 8, the prepared tosylate-PEG2K-tosylate (M n2.0 g of (= 4,000 g / mol) was completely dissolved in 50 mL of dimethylformamide (DMF) in a 250 mL dropping funnel and added to the reactor. It was slowly injected under an argon stream (10 mL / min) and reacted for 24 hours. The reaction was terminated by lowering the reactor temperature to room temperature. After filtering the reaction solution, the DMF was distilled using a rotary evaporator, and 100 mL of dichloromethane was added to completely dissolve it. The salt was then extracted twice using an aqueous ammonium chloride solution. The DCM was washed again with an aqueous NaCl solution, filtered, and precipitated in an excess of diethyl ether to obtain a white powder. After drying this in a vacuum oven at room temperature for 48 hours, 1.8 g of white powder was obtained. Its functionalization rate was over 98 mol% ( 1 H NMR analysis) (OHC-Bz-PEG2K-Bz-CHO).

[0243]

[0244] <Example 10> Preparation of Aldehyded Hyaluronic Acid

[0245] Add 100 mL of distilled water to a three-necked flask reactor (for 1 L) wrapped in black cloth, then add commercial hyaluronic acid (hyaluronic acid; M n = 1.4 × 10 51.0 g (0.00071 mmol) of Da was added and stirred to dissolve thoroughly (10 mg / mL). Sodium periodate (FW: 213.89 g / mol; NaIO4) was prepared as a 5 mL aqueous solution (0.145 mmol = 200 x [HA]), slowly added to the reactor at room temperature using a separatory funnel, and the reaction was carried out for 2 hours. Subsequently, 10 mL of ethylene glycol was added to terminate the reaction, and the reaction was concluded by stirring for 1 hour. To purify the reaction product, impurities were removed by dialyzing in distilled water using a dialysis membrane (MWCO 10,000) for 3 days, followed by freeze-drying. The results were then confirmed by FT-IR spectroscopy, and the degree of oxidation was determined as 100 units of glucose. The degree of aldehydeation was obtained by titration with hydroxylamine hydrochloride. The analysis showed that the aldehyde content was approximately 0.41 g (yield 41%).

[0246]

[0247] <Example 11> Preparation of Aldehyded Dextran

[0248] dextran (M in a manner similar to Example 10) n Alaldehyde conversion of (= 40,000 Da) was carried out. First, 5.0 g (0.125 mmol) of dextran was injected into the reactor and thoroughly dissolved in 200 mL of distilled water. 6.6 g (0.031 mol) of NaIO4 (~250 x [Dex]) The substance was dissolved in 20 mL of distilled water and slowly injected using a funnel at room temperature, then reacted for 8 hours while stirring. Approximately 2.2 mL of ethylene glycol was injected into the reactor to terminate the oxidation reaction, followed by dialysis (MWCO 7 kDa) for 3 days to purify the substance. After freeze-drying, the yield was measured. The yield was measured using the same analytical method as in Example 10. The measured results showed that the substance contained approximately 38 units of aldehyde groups per 100 units of glucose.

[0249]

[0250] <Example 12> Preparation of Aldehyded Cellulose

[0251] Cellulose was oxidized in the same manner as in Example 10. First, 2 g of cellulose (Sigma-Aldrich: #098K0157) was introduced into a reactor containing 200 mL of distilled water, and 55 o The mixture was well dispersed at C while stirring. To this, 5.28 g (0.0247 mol; 2 x [glucose units]) dissolved in 100 mL of distilled water was slowly added to the reactor and reacted for 2 hours while stirring (dark state). To terminate the reaction, 2 mL of ethylene glycol was injected and reacted for 2 hours while stirring. The reaction product was purified by dialysis (MWCO 7 kDa) for 3 days, freeze-dried, and the yield was measured. The prepared aldehyde-modified cellulose (0.62) was dissolved in distilled water. The yield was approximately 31%.

[0252]

[0253] <Example 13> Preparation of Hydrogel 1

[0254] P(CHz) obtained by the same reaction as in Example 7 2.5 -PEG4K-P(CHz) 2.5 (M nA solution of 0.12 g of (= 2,400 g / mol) dissolved in 1.0 mL DMSO, a solution of 0.8 g of aldehyde-PEG4K-aldehyde (OHC-Bz-PEG4K-Bz-CHO) prepared by the same method as in Example 9 dissolved in 1.5 mL DMSO, and a solution of 70.2 g of t-BuO-PEG5K-(CHz) prepared similarly to Example 4 dissolved in 1 mL DMSO were added to a 25 mL vial and mixed by shaking. The pH was adjusted to 8.0 using PBS (pH = 7.2) solution and NaOH (0.1 M) aqueous solution, and the vial temperature was maintained at room temperature for 3 hours. Gelation occurred during this time. Then, H2SO4 aqueous solution (0.1 M) was added to adjust the vial pH to 4.0 to obtain a clear purple solution. Finally, the gel was prepared again in the vial by adjusting the pH to 8.0 with NaOH aqueous solution.

[0255]

[0256] Figure 2 shows the reversible gelation behavior of a hydrogel prepared according to one embodiment of the present invention.

[0257]

[0258] <Example 14> Preparation of Hydrogel 2

[0259] In a manner similar to Example 13, a solution of 0.1 g of ALD-dextran prepared in Example 11 dissolved in 1.5 mL of DMSO and P(CHz) prepared in Example 7 2.5 -PEG4K-P(CHz) 2.5 Solutions of 0.21 g each dissolved in 2.0 mL of DMSO were mixed in a 25 mL vial, and the reversible gel-sol formation was confirmed by adjusting the pH to 8 and pH 4 using an aqueous solution of NaOH (0.1 M) and an aqueous solution of H2SO4 (0.1 M).

[0260]

[0261] <Example 15> Preparation of Hydrogel 3

[0262] In a manner similar to Example 13, a solution of 0.1 g of the ALD-cellulose prepared in Example 11 dissolved in 1.5 mL of DMSO and the (CHz) prepared in Example 7 2.5 -PEG4K-(CHz) 2.5 Solutions of 0.2 g each dissolved in 2.0 mL of DMSO were mixed in a 25 mL vial, and the reversible gel-sol formation was confirmed by adjusting the pH to 8 and pH 4 using an aqueous solution of NaOH (0.1 M) and a solution of H2SO4 (0.1 M).

[0263]

[0264] <Example 16> Preparation of HO-PEG4.0K-poly(N-PMI)3

[0265] 5.0 grams of t-BuO-PEG4.0K-poly(N-PMI)3 (molecular weight: 5,000 g / mol), prepared by the same process as in Example 5, were injected into a three-necked round-bottom flask (1 L), and 300 mL of methanol was injected under a nitrogen stream similar to Example 7. After stirring well for about 1 hour, 25 mL of a 0.1 molar HCl solution was injected using a dropper, and then under a nitrogen stream 80 o The temperature was raised to C and stirred well for 4 hours. After the reaction was complete, the pink solution was precipitated in an excess amount of diethyl ether and filtered to obtain approximately 4.9 g of purple powder (HO-PEG4.0K-poly(N-PMI)3).

[0266]

[0267] <Example 17> Preparation of Tosyl-PEG4.0K-P(N-PMI)3

[0268] Using 2 g of the sample prepared in Example 16, 1.98 g of PEG4.0K-P(N-PMI)3 having a tosyl group at the chain end was prepared in the same manner as in Example 8 (Tosyl-PEG4.0K-P(N-PMI)3).

[0269]

[0270] <Example 18> Preparation of HS-PEG4.0K-P(CHz)3

[0271] 1.5 g (0.3 mmol) of the sample prepared as in Example 17 was thoroughly dissolved in 150 mL of dimethylformamide (DMF) in a 1 L three-necked flask reactor, potassium thioacetate (FW: 114.21, 0.17 g in 15 mL of DMF) was injected, and 85 o The reaction was carried out at C under an argon stream for 24 hours, and after the reaction was complete, the mixture was precipitated in excess diethyl ether at room temperature and filtered to obtain a light purple powder (1.5 g). The powder was then dried in a vacuum oven, and 1.2 g was dissolved again in 100 mL of DMF using a 1 L beaker. Hydrazide and HCl / methanol (1.0 N solution) were added to the beaker ([HCl] / [H2N-NH2] > 8 / 1, mol / mol) 80 o Deacetylation and the Gabriel process (amine synthesis) using phthalimide were carried out simultaneously by stirring at C for 6 hours, then precipitated in excess diethyl ether and dried to obtain a 1.2 g dark purple gel sample (HS-PEG4.0K-P(CHz)3).

[0272]

[0273] <Example 19> HS-PEG3.4K-P(CHz) 3-x -P(CHz-PBA) x manufacturing

[0274] HS-PEG3.4K-P(CHz)3(M prepared by a method similar to that in Example 18 n = 3,600 g / mol. GPC & 1 ¹H NMR analysis) Inject 3.6 g (1.0 mmol) into a 1 L three-necked flask and inject approximately 100 mL of DMF to 70 oIt was thoroughly dissolved at C. Then, 3-formylphenylboronic acid (fPBA; FW = 149.94; 0.3 g; 2.0 mmol) was thoroughly dissolved in 50 mL of DMF, injected into the reactor, and after adjusting the pH to 7.4 with PBS (pH 7.4), 70 o The reaction was carried out at C for 24 hours with stirring. After the reaction was complete, 3.5 g of a pink gel sample was obtained by precipitating in excess diethyl ether at room temperature (HS-PEG3.4K-P(CHz) 3-x -P(CHz-PBA) x ).

[0275]

[0276] <Example 20> Preparation of a Carrier for Anticancer Drug Delivery 1

[0277] 3.6 g of a sample prepared by a method similar to Example 19 was reacted by injecting Doxorubicin (Boryung Pharmaceutical; Dox; FW = 579.98) ([CHz] / [Dox] = 1-3, mol / mol) to obtain 3.54 g of a pink gel sample (HS-PEG3.4K-P(CHz) 3-x -P(CHz-Dox) x )

[0278]

[0279] <Example 21> Preparation of a Carrier for Anticancer Drug Delivery 2

[0280] 0.025 g of the sample prepared in Example 20 was thoroughly dissolved in 5 mL of DMSO, 5 mL of PBS (0.01 M, pH 7.4) was added, the mixture was placed in a 25 mL beaker, and stirred well for 2 hours using a magnetic stirring bar. Dialysis membrane (MWCO 3.5K, Fisher Scientific; SnakeSkin) TMDMSO and unreacted Dox were completely removed using dialysis tubing and PBS (0.01 M, pH 7.4) solution, and the micelle structure was confirmed. Then, 20 mL of distilled water was added to prepare various solutions with micelle concentrations of 0.5–2.0 mg / mL.

[0281]

[0282] <Example 22> (Preparation of t-BuO-PEG3.4K-O-CO-CH=CH-COOH and preparation of mPEG4K-O-CO-CH=CH-COOH

[0283] t-BuO-PEG3.4K-O prepared by the same method as in Example 1 -+ K (3.4 g) and commercial CH3O-PEG4.0K-OH (mPEG4K-OH; 4.0 g) were each injected into separate 1 L three-necked flasks, 150 mL of DMF was added under an argon stream and stirred for 30 minutes to completely dissolve, 0.03 mol (4.15 g) of K2CO3 was injected into each reactor and stirred well; after 15 minutes, maleic anhydride (FW: 98.057, 2.94 g) was added to the reactor, and the reactor temperature was set to 60 o The temperature was raised to C and reacted for 12 hours. After adding a 0.01 mol HCl solution and reacting for 10 minutes, the reaction was terminated after cooling to room temperature, the excess diethyl ether was precipitated, and after filtration, dried in a vacuum oven at room temperature for 48 hours to prepare 3.35 g and 3.95 g of white powder, respectively (t-BuO-PEG3.4K-O-CO-CH=CH-COOH and mPEG4K-O-CO-CH=CH-COOH).

[0284]

[0285] <Example 23> Preparation of mPEG4K-O-Bz-maleimide

[0286] mPEG4K-OH (4.0 g) used in Example 22 was added to a 1 L three-necked flask, and under an argon stream, mPEG4K-tosyl (3.91 g) was prepared in the same manner as in Example 8. Using 3.5 g of the dried sample, and in a manner similar to Example 9, a solution prepared by mixing and dissolving N-(4-hydroxyphenyl)maleimide (FW=189.17, 0.38 g), K2CO3 (FW=138.205, 0.553 g), and triethylamine (FW=101.19, d=0.7255, 1.0 mL) in 20 mL of DMF in a 1 L three-necked flask was injected using a syringe under an argon stream; 3.5 g of the sample was dissolved in 30 mL of DMF and slowly injected into the reactor through a dropping funnel (10 mL / min), and the reactor temperature was raised to 120°C using a heating mantle. o The temperature was raised to C and the reaction was carried out for 12 hours. After lowering the reaction temperature to room temperature and terminating the reaction, the solution was filtered, and 90% of the DMF was distilled using an evaporator. Then, 100 mL of dichloromethane (DCM) was added to dissolve the solution, and some salts were extracted with an aqueous ammonium chloride solution (0.5 M, 50 mL). The DCM solution was then washed with an aqueous NaCl solution, filtered, precipitated in excess diethyl ether, and filtered to obtain a white powder. The powder was dried in a vacuum oven at room temperature for 48 hours to obtain 3.45 g of a white powder having a maleimide group at the chain end (mPEG4K-O-Bz-maleimide).

[0287]

[0288] <Example 24> Preparation of tBuO-PEG-PrBr

[0289] t-BuO-PEG3.4K-O prepared by the same method as in Example 1 -+K (3.4 g, 0.01 mol) was injected into a 1 L three-necked flask under an argon stream, 300 mL of dichloromethane was added, and the mixture was thoroughly dissolved. 0.01 mol (2.15 g) of 2-bromopropionyl bromide (PrBr) was injected using a dropping funnel, and the mixture was reacted at room temperature for 24 hours. The solution was precipitated in excess diethyl ether, filtered, and dried in a vacuum oven for 48 hours to obtain 3.36 g of white powder (t-BuO-PEG3.4K-CO-CH(CH3)-Br:tBuO-PEG-PrBr).

[0290]

[0291] <Example 25> Preparation of Macroinitiator-1 (RMI-1)

[0292] tBuO-PEG-PrBr (3.0 g) prepared in Example 24 was injected into a 1 L three-necked flask, and 200 mL of purified THF was injected under an argon stream to dissolve it well. Then, 0.05 mol (8.01 g) potassium ethyl xanthogenate (KEX) was dissolved in 20 mL of THF and injected into a reactor under an argon stream using a dropping funnel, and 25 o The reaction was carried out at C for 24 hours ([KEX] / [PrBr] = 5 / 1, mol / mol). After the reaction, the solution was precipitated in diethyl ether, filtered, dissolved in toluene and filtered again, precipitated in excess diethyl ether, and dried to obtain 2.96 g of a pale yellow powder (RAFT macroinitiator; RMI-1) (t-BuO-PEG3.4K-CO-CH(CH3)-S-CS-O-CH2-CH3; RMI).

[0293]

[0294] <Example 26> Preparation of t-BuO-PEG3.4K-bP(MAPBA)3-block copolymer

[0295] The RAFT radical polymerization initiator (RMI-1) prepared in Example 25 was injected into a 1 L three-necked plastic container, and 150 mL of DMF was added to completely dissolve it. Then, 3-(methacrylamido)phenylboronic acid (MAPBA) monomer ([MAPBA] / [RMI] = 3 / 1, mol / mol, 20 mL DMF) was injected into the reactor, and the reaction temperature was set to 90 o After raising the temperature to C, polymerization was carried out under an argon stream for 24 hours, α,α'-azoisobutyronitrile (AIBN) ([RMI] / [AIBN] = 1 / 5, mol / mol) was injected to terminate the reaction, and a white t-BuO-PEG3,4K-bP(MAPBA)3-block copolymer was prepared by precipitating in excess diethyl ether. At this time, P(MAPBA) n The molecular weight could be controlled according to the [MAPBA] / [RMI] ratio.

[0296]

[0297] <Example 27> Preparation of HO-PEG3.4K-bP(MAPBA)3

[0298] 1 g of the block copolymer prepared in Example 26 was dissolved in 100 mL of methanol in a 1 L three-necked flask, and then a 1 M HCl solution was added and stirred well at room temperature for 6 hours. The reaction solution was precipitated in an excess amount of diethyl ether to prepare white HO-PEG3.4K-bP(MAPBA)3.

[0299]

[0300] <Example 28> Preparation of tosyl-PEG3.4K-bP(MAPBA)3

[0301] 1 g of the dried sample from Example 27 and 100 mL of dichloromethane (DCM) were injected into a three-necked flask and dissolved by stirring well, and 1.2 g of the block copolymer tosyl-PEG3.4K-bP(MAPBA)3 having tosylate groups at the chain ends was prepared in a manner similar to Example 22.

[0302]

[0303] <Example 29> Preparation of DCA-PEG3.4K-bP(MAPBA)3

[0304] 1 g of the copolymer prepared as in Example 28 was injected into a 1 L three-necked flask in 100 mL of dimethylsulfoxide (DMSO), and 60°C was placed under an argon stream. o Dissolve well at C, and again, dissolve 0.005 mol of sodium deoxycholate (DCA; FW: 414.55) and 1.38 g of K2CO3 in 20 mL of DMSO in a mixture beforehand, slowly inject the solution into the reactor through a dropping funnel, and 120 o The temperature was raised to C and the reaction was carried out for 24 hours. After the reaction was finished, DMSO was partially distilled, 20 mL of DCM was added and dissolved, and the solution was precipitated in an excess amount of diethyl ether to obtain 0.98 g of white powder (DCA-PEG3.4K-bP(MAPBA)3).

[0305]

[0306] <Example 30> Preparation of TCA-PEG3.4K-bP(MAPBA)3

[0307] 0.95 g of white powder (TCA-PEG3.4K-bP(MAPBA)3) was obtained using 51.0 g of Tosyl-PEG3.4K-bP(MAPBA) with chain-terminal tosylate groups introduced, prepared in the same manner as in Example 29, and using taurocholic acid sodium salt hydrate (TCA: anhydrous FW = 537.68), a type of bile acid, in the same way, using the same molar ratio as in Example 30.

[0308]

[0309] <Example 31> Preparation of GCA-PEG3.4K-bP(MAPBA)3

[0310] Using 31.0 g of Tosyl-PEG3.4K-bP(MAPBA)3 with chain-terminal tosylate groups introduced, prepared in the same manner as in Example 29, and using glycocholic acid, sodium salt (GCA: FW = 487.60), a type of bile acid, in the same way, and reacting under the same reaction conditions using the same molar ratio as in Example 30, 0.95 g of white powder (GCA-PEG3.4K-bP(MAPBA)3) was obtained.

[0311]

[0312] <Example 32> Preparation of GCA-PEG3.4K-bP(N-PMI)3

[0313] Using 1 g of the t-BuO-PEG3.4K-bP(N-PMI)3 sample prepared as in Example 3, 30.98 g of the block copolymer GCA-PEG3.4K-bP(N-PMI) functionalized with glycocholic acid, sodium salt (GCA) was prepared in the same manner as in Examples 29, 30, and 31.

[0314]

[0315] <Example 33> Preparation of GCA-PEG3.4K-bP(CHz)3

[0316] 0.95 g of a block copolymer, GCA-PEG3.4K-bP(CHz)3 having a cyclic acylhydrazone group, was prepared by deimidating 1.0 g of a sample prepared in the same manner as in Example 32 in a manner similar to Example 4.

[0317]

[0318] <Example 34> Preparation of GCA-PEG3.4K-b-[P(CHz)-co-P(CHz-PBA)2]

[0319] 1.0 g of the sample (GCA-PEG3.4K-bP(CHz)3) prepared by the same method as in Example 33 was injected into a 1 L three-necked flask (100 mL, DMSO), and 40 under an argon stream o Dissolve well at C, adjust pH to 7.4 using PBS (0.1 M), dissolve 3-formylphenylboronic acid (0.25 g) ([CHz] / [fPBA] = 1-3 mol / mol) in 10 mL of DMSO and inject into the reactor using a syringe, and 40 o Reacted at C for 5 hours and precipitated in excess diethyl ether to obtain 0.95 g of dark purple powder (GCA-PEG3.4K-b-[P(CHz)-co-P(CHz-PBA)2]).

[0320]

[0321] <Example 35> Preparation of t-BuO-PEG3.4K-O-CO-C(CH3)2-Br

[0322] t-BuO-PEG3.4K-O prepared by the same method as in Example 1 -+Inject K (3.4 g, 0.001 mol) into a 1 L three-necked flask (200 mL: tetrahydrofuran; THF), add α-bromoisobutyryl bromide (BuBr; FW = 229.90, 0.01 mol) into 30 mL of THF and dissolve by stirring well, inject into a reactor, and under an argon stream for 25 o The reaction was carried out at C for 24 hours. After the reaction, it was precipitated in excess diethyl ether to obtain 3.26 g of white powder (t-BuO-PEG3.4K-O-CO-C(CH3)2-Br).

[0323]

[0324] <Example 36> Preparation of Macroinitiator-2 (RMI-2)

[0325] 2 g of bromine-termianted PEG3.4K (t-BuO-PEG3.4K-BuBr) obtained in Example 35 was placed in a three-necked plastic flask (150 mL THF), potassium ethyl xanthogenate ([BuBr] / [K] = 1 / 5, mol / mol) was added to 20 mL THF and completely dissolved, injected into a three-necked flask using a syringe, and reacted for 24 hours under an argon stream. The finished reaction product was precipitated in excess diethyl ether, filtered, and dried in a vacuum oven to obtain 1.9 g of a pale yellow powder (RAFT macroinitiator; RMI-2) (t-BuO-PEG3.4K-O-CO-C(CH3)2-S-CS-O-CH2-CH3; RMI-2).

[0326]

[0327] <Example 37> PEG-bP(MAPBA) 2~10 manufacturing

[0328] Using the reversible addition fragmentation chain transfer (RAFT) macroinitiator (RMI-2) prepared as in Example 36, 3-(methacrylamido)phenylboronic acid (MAPBA) monomer in dimethylformamide (DMF) solvent 90 in the same manner as in Example 26 o After RAFT radical polymerization under an argon stream at C for 24 hours, α,α'-azoisobutyronitrile (AIBN; [RMI] / [AIBN] = 1 / 5, mol / mol) was injected to terminate the reaction, the temperature inside the reactor was lowered to room temperature to terminate the polymerization, the polymerization solution was precipitated in excess diethyl ether, and dried to obtain PEG-bP(MAPBA) 2~10 was prepared. At this time, the molecular weight of MAPBA could be controlled ([MAPBA] / [RMI] = 2-10).

[0329]

[0330] <Example 38> Preparation of Macroinitiator-3 (mRMI-3) and mPEG4K-bP(MAPBA)3

[0331] 3.92 g of mPEG-macroinitiator (mRMI-3) mPEG4K-O-CO-CH(CH3)-Br) was prepared using commercial methoxy-PEG4K-OH (mPEG4K, 4.0 g) prepared or purchased in the same manner as in Example 2 and K2CO3 (FW = 138.205 g / mol, 0.7 g) in a manner similar to Example 25, and mPEG4K-bP(MAPBA)3 was prepared by polymerizing MAPBA in the same manner as in Examples 26 and 27.

[0332]

[0333] <Example 39> Preparation of Macroinitiator-4 (mRMI-4) and mPEG4K-bP (MAPBA)3

[0334] mPEG-macroinitiator (RAFT macroinitiator; mRMI-4) mPEG4K-O-CO-C(CH3)2-Br was prepared in a manner similar to Examples 36 and 37 using commercial methoxy-PEG4K-OH (mPEG4K, 3.0 g) prepared in the same manner as in Example 38, and mPEG4K-bP(MAPBA)3 was prepared by polymerizing MAPBA in the same manner as in Example 38 (mPEG4K-bP(MAPBA)3).

[0335]

[0336] <Example 40> Preparation of mPEG4K-bP(N-PMI)3

[0337] Commercial mPEG4K-OH (4.0 g), similar to that in Example 2, was injected into a 1 L three-necked flask, K2CO3 (1.382 g, 0.01 mol) was injected, 250 mL of acetone was injected, and the mixture was completely dissolved under an argon stream at room temperature. Polymerization was carried out by adding N-phenylmaleimide (N-PMI) (FW; 173.17, 0.003 mol) in the same manner as in Example 4 to synthesize 34.8 g of mPEG4K-bP(N-PMI) with a molecular weight of 4,520 g / mol.

[0338]

[0339] <Example 41> Preparation of mPEG4K-bP(CHz)3

[0340] 34.0 g of the sample mPEG4K-bP(N-PMI) obtained in Example 40 was deamidated in the same manner as in Example 19 to produce 33.5 g of mPEG4K-bP(CHz).

[0341]

[0342] <Example 42> t-BuO-PEG3.4K-b-[P(CHz) 3-x -co-P(CHz-GCA) x Manufacturing of ]

[0343] 3.4 g of t-BuO-PEG3.4K-bP(CHz)3 (tP(CHz)3), prepared in the same manner as in Example 5, was reacted with glycocholic acid, sodium salt (GCA; FW = 487.60) ([tP(CHz)] / [GCA] = 1-3, mol / mol) under conditions similar to those in Example 19 (pH 7.4) to produce a pink t-BuO-PEG3.4K-b-[P(CHz) 3-x -co-P(CHz-GCA) x ] 3.5 g of powder was prepared.

[0344]

[0345] <Example 43> t-BuO-PEG3.4K-b-[P(CHz) 3-x -co-P(CHz-TCA) x Manufacturing of ]

[0346] Under the same reaction conditions as in Example 42, the reaction with taurocholic acid sodium salt hydrate (TCA; FW = 537.68) ([tP(CHz) / [TCA] = 1-3, mol / mol) instead of GCA was carried out under conditions similar to those in Example 19 to produce the pink powder t-BuO-PEG3.4K-b-[P(CHz) 3-x -co-P(CHz-TCA) x ] obtained 3.5 g.

[0347]

[0348] <Example 44>t-BuO-PEG3.4K-b-[P(CHz)-co-P(CHz-FLA) 1-2 Manufacturing of ]

[0349] Under the same reaction conditions as Example 42, the reaction with folic acid (FLA; FW = 441.40) ([tP(CHz) / [FLA] = 1-2, mol / mol) instead of GCA was carried out under conditions similar to Example 19 to produce a pink powder t-BuO-PEG3.4K-b-[P(CHz)-co-P(CHz-FLA) 1-2 ] 3.6 g was obtained.

[0350]

[0351] <Example 45> mPEG4K-b-[P(CHz) 3-x -co-P(CHz-GCA) x ], mPEG4K-b-[P(CHz) 3-x -co-P(CHz-TCA) x ] and mPEG4K-b-[P(CHz) 3-x -co-P(CHz-FLA) x Manufacture of ])

[0352] 34.0 g of mPEG4K-P(CHz) prepared as in Example 41 was reacted under the same conditions as the GCA, TCA, and FLA used in Examples 42, 43, and 44 to quantitatively prepare the pink related powders, respectively.

[0353] (mPEG4K-b-[P(CHz) 3-x -co-P(CHz-GCA) x ],

[0354] mPEG4K-b-[P(CHz) 3-x -co-P(CHz-TCA) x ],

[0355] mPEG4K-b-[P(CHz) 3-x -co-P(CHz-FLA) x ]).

[0356]

[0357] <Example 46> Preparation of t-BuO-PEG3.4K-O-Bz-maleimide

[0358] t-butoxy-PEG3.4K-O prepared by the same method as in Example 1 -+3.4 g of K was injected into a 1 L three-necked flask containing 200 mL of dichloromethane (DCM) and dissolved while stirring well. Then, trimethylamine (1.182 g; 0.02 mol) was injected into the reactor and stirred well. t-butoxy-PEG3.4K-tosyl (3.25 g) was prepared in the same manner as in Example 8, and using 3.0 g of the dried sample, in a manner similar to Example 9, a solution prepared by mixing and dissolving N-(4-hydroxyphenyl)maleimide (FW=189.17, 0.38 g), K2CO3 (FW=138.205, 0.553 g), and triethylamine (FW=101.19, d=0.7255, 1.0 mL) in 20 mL of DMF in a 1 L three-necked flask was injected using a syringe under an argon stream; 3.0 g of the sample was dissolved in 30 mL of DMF and slowly injected into the reactor through a dropping funnel (10 mL / min), and the reactor temperature was raised to 120°C using a heating mantle. o The temperature was raised to C and the reaction was carried out for 12 hours. After lowering the reaction temperature to room temperature and terminating the reaction, the solution was filtered, and 90% of the DMF was distilled using an evaporator. Then, 100 mL of dichloromethane (DCM) was added to dissolve the solution, and some salts were extracted with an aqueous ammonium chloride solution (0.5 M, 50 mL). The DCM solution was then washed with an aqueous NaCl solution, filtered, precipitated in excess diethyl ether, and filtered to obtain a white powder. The powder was dried in a vacuum oven at room temperature for 48 hours to obtain 2.9 g of a white powder having a maleimide group at the chain end (t-BuO-PEG3.4K-O-Bz-maleimide).

[0359]

[0360] <Example 47> Preparation of t-BuO-PEG3.4K-O-CO-CH2-CH(S-CO-CH3)(COOH)

[0361] 3.4 g of the sample (t-BuO-PEG3.4K-maleate) prepared in the same manner as in Example 22 was placed into a 1 L three-necked flask injected with 150 mL of DMF under an argon stream, the pH was adjusted to 7.4 using PBS (pH 7.4), and 60 o Raise temperature to °C, and after thorough dissolution, thioacetic acid (FW = 76.12 g / mol, d = 1.08 g / cm³) 3 1.0 mL of ) was injected into the reactor using a syringe and reacted for 24 hours (Micheal thiol-ene addition). The reaction was terminated by lowering the temperature to room temperature, and after partially removing DMF by distillation using an evaporator, 50 mL of methanol was added, the mixture was precipitated in excess diethyl ether and filtered, and after drying in a vacuum oven, 3.35 g of a light yellow powder was obtained (t-BuO-PEG3.4K-O-CO-CH2-CH(S-CO-CH3)(COOH)).

[0362]

[0363] <Example 48> Preparation of hetero-trifunctional PEG (HO-PEG3,4K-O-CO-CH2-CH(SH)(COOH): hetero-trifunctional PEG)

[0364] 3.0 g of the sample prepared in Example 47 was placed in a 500 mL beaker, 50 mL of 7 methanol was added, 2 mL of HCl (0.1 M) was injected while stirring, and deacetylation and deprotection of the t-butoxy group were carried out at room temperature for 6 hours while stirring. The solution was then precipitated in excess diethyl ether, filtered, and dried in a vacuum oven to produce 2.9 g of a light yellow powder (HO-PEG3.4K-O-CO-CH2-CH(SH)(COOH); hetero-trifunctional PEG).

[0365]

[0366] <Example 49> Preparation of Carriers for Protein and Peptide Precursor Delivery 1

[0367] 4.0 g of the sample (mPEG4K-O-CO-CH=CH-COOH) prepared as in Example 22 was placed in a 1 L three-necked flask, 150 mL of DMF was injected under an argon stream, and 40 o Sonication was performed at C. 0.61 g of cysteine ​​(FW = 121.16 g / mol), a protein and peptide precursor, was sonicated in a 50 mL beaker injected with 10 mL of DMF, injected using a dropping funnel, adjusted to pH 7.4 (basic), and 40 o The reaction was carried out for 2 hours under sonication conditions. After terminating the reaction and filtering out the unreacted material, the excess diethyl ether was precipitated, and after filtration and drying in a vacuum oven, 1 ¹H NMR analysis confirmed that there was more than 98 mol% of Michael addition reaction (mPEG4K-O-CO-CH2-C(Cysteine)(COOH)).

[0368]

[0369] <Example 50> Preparation of a Carrier for Anticancer Drug Delivery 3

[0370] Sample prepared similarly to Example 33 (FLA-PEG3.4K-bP(CHz)3) (M n = 3,600 g / mol; 3.6 g) was added to a 1 L three-necked flask, and after injecting 100 mL of DMF under an argon stream, 40 o The mixture was heated to C and thoroughly dissolved. 1.74 g, 1.16 g, and 0.58 g of doxorubicin (Dox: FW = 579.98 g / mol) ([CHz] / [Dox] = 1~3, mol / mol) were dissolved in 10 mL DMF in 50 mL vials, respectively. After injecting into a reactor using a syringe and reacting for 24 hours, the mixtures were precipitated in excess diethyl ether at room temperature to obtain light pink powders (FLA-PEG3.4K-b-[P(CHz) 3-x -co-P(CHz-Dox) x ]).

[0371]

[0372] <Example 51> Preparation of a Carrier for Anticancer Drug Delivery 4

[0373] Sample prepared in the same manner as in Example 41 (mPEG4K-bP(CHz)3;M n = 4,100 g / mol) 4.1 g was reacted with Dox in a manner similar to Example 50 to obtain light pink powder, 4.0 g, 3.9 g, and 4.1 g respectively (mPEG4K-b-[P(CHz) 3-x -co-P(CHz-Dox) x ]).

[0374]

[0375] <Example 52> Preparation of a Carrier for Anticancer Drug Delivery 5

[0376] The samples prepared in Examples 50 and 51 were thoroughly dissolved in 50 mL of DMSO, 5 mL of PBS (0.01 M, pH 7.4) was added, the mixture was placed in a 250 mL beaker, and stirred well for 2 hours using a magnetic stirring bar. Then, the solution was dialysis membrane (MWCO 3.5K, Fisher Scientific; SnakeSkin) TM DMSO and unreacted Dox were completely removed using dialysis tubing and PBS (0.01 M, pH 7.4) solution, dissolved in 5 mL of ethanol, and precipitated with excess diethyl ether to obtain 2.0 g of pink powder, which was dried in a vacuum oven at room temperature and used to prepare micelles.

[0377]

[0378] <Example 53> Preparation of an Insulin Delivery Carrier 1

[0379] GCA-PEG3.4K-bP(MAPBA)5 prepared as in Example 31 0.36 g of mPEG4K-bP(MAPBA) prepared as in Example 39 and 33.69 g of mPEG4K-bP (MAPBA) (1:9, mol / mol) were injected into a 1 L three-necked flask with 50 mL of pH 12 NaOH aqueous solution and thoroughly dissolved. Then, pH 2 HCl insulin aqueous solution was injected using a dropping funnel at a rate of 6 drops / min, and the mixture was stirred well using a magnetic stirring bar until a turbid solution was formed to form micelles. The salts in this solution were separated using a dialysis bag (MWCO 3 kD), and the pH was adjusted to 7.4 using distilled water and PBS (pH 7.4) to prepare a solution with a concentration of 0.60 mg / mL. At this time, the insulin concentration was analyzed by UV-Vis spectrum measurement after decomposing the micelles by adjusting the pH of the prepared micelles to 10.

[0380]

[0381] <Example 53> Preparation of Hydrogel 4

[0382] Sample in Example 7 (P(CHz) 2.5 -PEG3.4K-P(CHz) 2.5 ) (PCHz; 3.5 g), Sample 9 (OHC-PEG2K-CHO) (PCHO; 4.0 g), Sample 31 (GCA-PEG3.4K-P(MAPBA)3) (GPPMPBA; 0.9 g), commercial 1,3,5-triformylbenzene [Bz(CHO)3] (BzCHO; 0.054 g), Sample 39 mPEG4K-bP(MAPBA)3 (mPPMPBA; 1.45 g), and PVA (4.0 kDa; 0.01 g) were injected into a 1 L three-necked flask in 100 mL ethanol / water (9 / 1, v / v) using PBS (pH 7.4), and 30 oIt was completely dissolved at C. After separately preparing 0.1 g liraglutide with 4 mL of PBS (pH 7.4) and 1 mL of 10 wt% sucrose solution (0.1 g / 5 mL), it was slowly injected using a dropping funnel under an argon stream, and the 1 M HCl aqueous solution was adjusted to pH 6.0 using a syringe, and 37 o The solution was stirred using a magnetic stirring bar to form a suspension (gel) at C (30 min). The solution was then transferred back into a dialysis tube (Spectra / Por ⓡ Using a 7 MWCO 2000, unencapsulated polymers, liraglutides, and organic matter were separated and purified in a pH 6.0 ethano1 / water solution (250 mL), precipitated in excess diethyl ether, filtered, dried in a vacuum oven at room temperature, and stored in a refrigerator. When used, it was used as a 15 wt% aqueous solution.

[0383]

[0384] <Example 54> Preparation of Hydrogel 5

[0385] Sample in Example 7 (P(CHz) 2.5 -PEG3.4K-P(CHz) 2.5 ) (PCHz; 3.5 g), sample from Example 11 (Dextran-CHO; 40 kDa; [ALD] / [Glucose] = 38 / 100 units / units) (DCHO; 0.596 g), sample from Example 30 (TCA-PEG3.4K-bP(MAPBA)3) (TPPMPBA; 0.9 g), sample from Example 39 mPEG4K-bP(MAPBA)3 (mPPMPBA; 1.45 g), and PVA (4.0 kDa; 0.01 g) were injected into a 1 L three-necked flask in 100 mL ethanol / water (9 / 1, v / v) using PBS (pH 7.4), and 30 oIt was completely dissolved at C. After separately preparing 0.1 g liraglutide with 4 mL of PBS (pH 7.4) and 1 mL of 10 wt% sucrose solution (0.1 g / 5 mL), it was slowly injected using a dropping funnel under an argon stream, and the 1 M HCl aqueous solution was adjusted to pH 6.0 using a syringe, and 37 o The solution was stirred using a magnetic stirring bar to form a suspension (gel) at C (30 min). The solution was then transferred back into a dialysis tube (Spectra / Por ⓡ Using a 7 MWCO 2000, unencapsulated polymers, liraglutides, and organic matter were separated and purified in a pH 6.0 ethano1 / water solution (250 mL), precipitated in excess diethyl ether, filtered, dried in a vacuum oven at room temperature, and stored in a refrigerator. When used, it was used as a 15 wt% aqueous solution.

[0386]

[0387] <Example 55> Preparation of Hydrogel 6

[0388] Sample HS-PEG3.4K-b-[P(CHz)-co-P(CHz-PBA)2] (SPCHzPBA; 0.36 g) from Example 19, sample mPEG4K-b-[P(CHz)-co-P(CHz-TCA)2] (mPPCHTCA; 0.41 g) from Example 45, and sample mPEG4K-O-CO-CH=CH-COOH (mPMALT; 8.2 g) from Example 22 were injected into a three-necked flask, and under an argon stream, 60 mL of ethanol / distilled water (9 / 1, v / v) was added to 40 oDissolve well at C, and separately prepare 0.1 g liraglutide with 4 mL of PBS (pH 7.4) and 1 mL of 10 wt% sucrose solution (0.1 g / 5 mL), then slowly inject using a dropping funnel under an argon stream, adjust the pH of the PBS (pH 7.4) solution to 7.4 using a syringe, and 37 o The solution was stirred using a magnetic stirring bar at C (4 hours). Then, the solution was transferred back into a dialysis tube (Spectra / Por ⓡ Using a 7 MWCO 2000, unencapsulated polymers, liraglutides, and organic matter were separated and purified in a pH 7.4 ethano1 / water solution (250 mL), precipitated in excess diethyl ether, filtered, dried in a vacuum oven at room temperature, and stored in a refrigerator.

[0389]

[0390] <Example 56> Preparation of Hydrogel 7

[0391] In Example 33, 30.36 g of sample GCA-PEG3.4K-bP(CHz) and exenatide acetate (Ex-4; > 98%, MW 4246 Da) were mixed with Zn 2+ Inject the solution into 20 mL of dichloromethane (DCM), and 4 o Nanocomplexes were prepared by stirring at C for 12 hours (GCA:Ex; 1:1, 10:1 by weight ratio). Subsequently, trehalose (2%, w / v) was introduced into the reactor (0.43%, w / v) to prevent aggregation of the nanocomplexes, and after stabilization by stirring well for 2 hours, the mixture was precipitated in an excess of diethyl ether to obtain the powder. The prepared solid powder was -20 o C. Stored frozen. Enteric capsules were used for use.

[0392]

[0393] <Example 57> Preparation of Hydrogel 8

[0394] 0.036 g of sample FLA-PEG3.4K-b-[P(CHz)-co-P(CHz-Dox)2] from Example 50 and sample (mPEG4K-bP(CHz)3;M) prepared in the same manner as in Example 41 n 0.41 g (= 4,100 g / mol), 30.36 g of the sample HO-PEG3.4k-bP(MAPBA) from Example 27 was injected into a 1 L three-necked flask, 80 mL of anhydrous DMSO was added under an argon stream, and completely dissolved at room temperature. Then, Dox (0.11 g mg, 0.22 mmol) and triethylamine (TEA; 184 mL, 1.32 mmol) were dissolved in 20 mL of anhydrous DMSO and injected into the reactor using a syringe, and reacted at room temperature for 24 hours. Using a dialysis bag (MWCO; 3,500), 37 PBS pH 7.4 aqueous solution o After removing prodrugs and substances not involved in micelle formation in C, and removing a portion of DMSO using an evaporator, the mixture was precipitated in excess diethyl ether to obtain a pink powder. For separate use, it was dispersed in distilled water (1.0 mg / mL).

Claims

1. A polyethylene glycol block containing a bile acid (BA) group at the chain end; and A block copolymer characterized by comprising a polymer block composed of a homopolymer or copolymer of monomers having boronic acid groups in their molecular structure.

2. In Claim 1, A block copolymer characterized in that the above bile acid (BA) is any one of cholic acid, deoxycholic acid, glycolic acid, or taurocholic acid.

3. In Claim 1, The above block copolymer is a block copolymer characterized by having a chemical structure represented by the following chemical formula A: <Chemical Formula A> In the above chemical formula A, BA is a residue of bile acid, and the bile acid is any one of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA) or folic acid (FLA), R1 represents H- or CH3-, m is an integer from 1 to 10 as a repeating unit, and n is an integer from 10 to 200 as a repeating unit.

4. A polymer micelle characterized by comprising the block copolymer of any one of claims 1 to 3.

5. Polymer micelles according to claim 4; and A carrier for delivering a bioactive substance, characterized by comprising a bioactive substance that can be encapsulated within the above-mentioned polymer micelle.

6. Polymer micelles according to claim 4; and A polymer hydrogel characterized by comprising a bioactive substance that can be encapsulated within the polymer micelles.

7. In Claim 6, A polymer hydrogel characterized by further comprising a block copolymer or a triblock copolymer of a polymer having polyethylene glycol blocks and cyclic hydrazides as repeating units.

8. In Claim 6, A polymer hydrogel characterized in that the above carrier further comprises a compound containing two or more aldehyde groups in its molecular structure.

9. In Claim 6, A polymer hydrogel characterized in that the above-mentioned bioactive substance is one or more selected from type 2 diabetes drugs, insulin, obesity treatment drugs, DPP-4 inhibitors, or anticancer drugs.

10. In Claim 9, A polymer hydrogel characterized in that the above-mentioned type 2 diabetes drug is one or more selected from glucagon-like peptide-1 receptor agonists (GLP-1 RAs), exenatide, semaglutide, tirzeptide, or liraglutide having amine groups at the chain ends.

11. A step of preparing a first block copolymer composed of an ethylene glycol monomer block having a bile acid (BA) group at the chain end and a monomer block having a boronic acid group; A step of preparing a second block copolymer composed of an ethylene glycol monomer block and a cyclic hydrizide block; and A method for preparing a polymer hydrogel, characterized by comprising the step of forming polymer micelles from a mixture comprising the first block copolymer, the second block copolymer, and a compound containing an aldehyde group within a molecular structure capable of forming a dynamic covalent bond to the cyclic hydrizide block of the second block copolymer.

12. In Claim 11, The step of manufacturing the first block copolymer is, A step of preparing a macroinitiator comprising a polyethylene glycol block; A step of polymerizing a monomer having a boronic acid group in its molecular structure using the above macroinitiator; and A method for preparing a polymer hydrogel characterized by including the step of introducing a bile acid (BA) group to the end of the polyethylene glycol block.

13. In Claim 12, A method for preparing a polymer hydrogel characterized in that the above bile acid (BA) is one or more of cholic acid (CLA), deoxycholic acid (DOA), glycolic acid (GCA), taurocholic acid (TCA), or folic acid (FLA).

14. In Claim 11, A method for manufacturing a polymer hydrogel, characterized in that the step of forming the polymer micelles includes the step of further mixing a bioactive substance into the mixture.

15. In Claim 14, A method for manufacturing a polymer hydrogel characterized in that the above-mentioned bioactive substance is one or more selected from type 2 diabetes drugs, insulin, obesity treatment drugs, DPP-4 inhibitors, or anticancer drugs.

16. In Claim 11, The step of manufacturing the second block copolymer is, A step of preparing a block copolymer of polyethylene glycol and poly(N-phenylmaleimide); and A method for preparing a polymer hydrogel characterized by including the step of reducing the above-mentioned poly(N-phenylmaleimide) block using hydrizine.

17. In Claim 11, A method for preparing a polymer hydrogel characterized in that the compound containing an aldehyde group within the molecular structure is one or more selected from aldehyde-treated polyethylene glycol, aldehyde-treated hyaluronic acid, aldehyde-treated dextran, or aldehyde-treated cellulose.

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