Camptothecin conjugated polyethylene glycol acrylate hydrogel and preparation method thereof

Abstract

The present application relates to a kind of camptothecin coupling polyethylene glycol acrylate hydrogel and its preparation method.Camptothecin is coupled by acid anhydride and star multi-arm PEG, and simultaneously, polyacrylate is chemically modified, to peroxodisulfuric acid amine (APS) / tetramethyl ethylenediamine (TMEDA) oxidation-reduction system is initiated system, by active radical solution polymerization method, crosslinking polyethylene glycol polyacrylate (PEGDA) hydrogel is prepared.The gel obtained by the method described in the present application can realize long-acting sustained release of drug.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine and relates to a camptothecin-coupled polyethylene glycol acrylate hydrogel and its preparation method. Background Technology

[0002] Camptothecin is a highly effective antitumor drug that is widely used. However, its poor water solubility, short in vivo half-life, and long residual time in vivo limit its application and efficacy. Directly encapsulating the drug in a hydrogel can prolong the overall drug release time, which is beneficial for the drug to exert long-term antitumor activity in vivo.

[0003] Hydrogels are a class of biocompatible porous materials where drugs can be encapsulated within the gel during regel formation and released to the outside through the gel's pores. Traditional hydrogel drug release systems rely on passive diffusion of the drug within the gel, which is significantly affected by the drug's molecular weight and the gel's pore size. This is especially true for small-molecule chemotherapy drugs, which exhibit rapid release rates within hydrogels, often accompanied by burst release phenomena. Another approach involves covalently coupling the drug to polymer molecules and then using the drug-coupled polymer to prepare the hydrogel. This allows the drug to be chemically encapsulated within the hydrogel, preventing excessively rapid release.

[0004] Chinese patent document CN115120591A discloses a PEGylated camptothecin derivative having the following structure: The PEGylated camptothecin derivative can crosslink in situ with polyamine compounds to form an injectable gel. The drug is slowly released as the gel degrades, achieving long-term sustained-release. This PEGylated camptothecin sustained-release gel requires dissolving the PEGylated camptothecin derivative and the polyamine compound separately in buffer solution, and then mixing them using a dual-syringe. However, chemically crosslinked two-component gels are easily affected by the reaction pH and solution conditions during gel formation. Furthermore, the chemical crosslinking method requires mixing the two-component solutions before gel formation, limiting the gel's applicability. For example, when using urinary catheters, hemodialysis catheters, or vascular interventional catheters, gel formation requires a gel coating on the catheter; chemical crosslinking typically requires multiple dip-coatings, resulting in a lengthy and inconvenient process.

[0005] Free radical polymerization can control the onset time and polymerization rate of gel polymerization through ultraviolet light initiation, making gel preparation more convenient and controllable compared to chemical crosslinking methods. However, existing photocurable gels typically achieve drug loading through physical encapsulation, making it difficult to achieve long-term gel release. Furthermore, the hydrophobicity of the drug makes it difficult to disperse it uniformly within the gel, affecting coating quality. Summary of the Invention

[0006] This invention addresses the shortcomings of existing technologies by providing a novel PEGylated camptothecin sustained-release gel and its preparation method. Camptothecin and acrylate are sequentially attached to a multi-arm PEG via chemical bonding to prepare a drug-coupled polymer molecule capable of free radical polymerization. Long-term sustained release of camptothecin is achieved by adjusting the number and type of acrylate grafts.

[0007] The specific technical solution of this invention is as follows:

[0008] A method for preparing camptothecin-coupled polyethylene glycol acrylate hydrogel involves using a redox system or photoinitiation to react acrylic anhydride or alkyl-substituted acrylic anhydride (preferably C1-C6 alkyl) with a star-shaped multi-arm PEG coupled with camptothecin via free radical solution polymerization to obtain a cross-linked network polyethylene glycol polyacrylate (PEGDA) hydrogel.

[0009] The star-shaped multi-arm PEG with camptothecin coupling described in this invention can be prepared by reacting camptothecin with a dicarboxylic acid anhydride or a dicarboxylic acid and then coupling it with a star-shaped multi-arm PEG. The anhydride is selected from succinic anhydride, glutaric anhydride, adipic anhydride, pimelic anhydride, succinic anhydride, azelaic anhydride, and sebacic anhydride, and the dicarboxylic acid is HOOC(CH2)nCOOH, where n = 2-6.

[0010] The star-shaped multi-arm PEG coupled with camptothecin has the following structure:

[0011] Where n represents a positive integer from 1 to 10, preferably a positive integer from 2 to 6. PEG is star-shaped multi-arm polyethylene glycol (uncapped, with hydroxyl groups at the free ends).

[0012] Preferably, the star-shaped multi-arm polyethylene glycol has 2-8 arms and a single-arm molecular weight of 1000-5000 Da.

[0013] The preparation method can be referred to the method disclosed in Chinese patent document CN115120591A.

[0014] The redox system initiator described in this invention is preferably an ammonium disulfate (APS) / tetramethylethylenediamine (TMEDA) redox system. The photoinitiator is a water-soluble photoinitiator, preferably 2-hydroxy-4'-(2-hydroxyethoxy)-2-methylphenylacetone (I2959) or lithium phenyl (2,4,6-trimethylbenzoyl)phosphate (LAP).

[0015] The solvent in the reaction system of this invention is an aqueous solution.

[0016] Preferably, the reaction temperature is 4-60°C and the reaction time is 1 second to 30 minutes.

[0017] Preferably, the molar ratio of the star-shaped multi-arm PEG of the coupled camptothecin to the dicarboxylic acid anhydride or dicarboxylic acid is 1:1.1-7, more preferably 1:2-6.

[0018] Another objective of this invention is to provide a camptothecin-coupled polyethylene glycol acrylate hydrogel, prepared using the method described in this invention.

[0019] Furthermore, the present invention also provides the application of the camptothecin-coupled polyethylene glycol acrylate hydrogel in the preparation of adhesives for the preparation of antitumor drugs.

[0020] Advantages of this invention:

[0021] Crosslinking mechanisms of chemical hydrogels are classified into two categories: free radical polymerization and chemical condensation reactions. Free radical polymerization, with its broad adaptability and resistance to pH and other conditions, is a widely adopted method for hydrogel crosslinking. Initiation systems for free radical polymerization include redox initiation and photoinitiation. In this invention, the polyethylene glycol system coupled with camptothecin drug molecules achieves controllable polymerization and gelation through redox and photoinitiation systems. This invention researches and develops gelation methods, and through formulation optimization, successfully prepares drug-coupled gels using an APS / TEMED initiation system, photoinitiator I2959, and LAP. Furthermore, by adjusting the grafting number of camptothecin and the gel properties, 30%-80% of the drug is released within one week, demonstrating superior long-term sustained-release effects compared to existing physical encapsulation methods (CN115120591, 80% release in 24 hours, complete release in 3 days). Attached Figure Description

[0022] Figure 1 1H NMR spectra of PEG-CPT and PEG-CPT-methacrylate.

[0023] Figure 2 1H NMR spectra of PEG-CPT and PEG-CPT-acrylate.

[0024] Figure 3 Drug release curves of PEG-CPT-methacrylate and PEG-CPT-acrylate polymerized gels.

[0025] Figure 4 In vitro swelling curves of PEG-CPT-methacrylate and PEG-CPT-acrylate polymerized gels.

[0026] Figure 5 Gel-coated conduit prepared by ultraviolet light curing. Detailed Implementation

[0027] The specific steps of the present invention are illustrated below through examples, but are not limited to these examples.

[0028] Unless otherwise stated, the terms used in this invention generally have the meanings commonly understood by those skilled in the art.

[0029] The present invention will now be described in further detail with reference to specific embodiments and data. It should be understood that these embodiments are merely illustrative of the invention and are not intended to limit the scope of the invention in any way.

[0030] Example 1: Synthesis of Carboxycamptothecin CPT-OOC(CH2)3COOH

[0031]

[0032] Camptothecin (CPT-OH, 342 mg, 3 mmol) and glutaric anhydride (348 mg, 1 mmol) were weighed into a 50 mL flask. Then, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU, 240 μL, 1.5 mmol) and 15 mL of dichloromethane (DCM) were added. The mixture was stirred at room temperature for 1 h. The reaction was quenched with approximately 50 mL of water, resulting in a large amount of white precipitate. The white precipitate was filtered and washed three times with water, then dried in a 60 °C oven for 1 h to obtain 368 mg of a pale yellow solid, with a yield of 79.7%. The above steps were repeated twice more, yielding 387 mg and 358 mg respectively, with yields of 83.8% and 77.5%.

[0033] Example 2: Synthesis of the eight-arm polyethylene glycol-camptothecin coupling compound PEG-CPT

[0034]

[0035] CPT-OOC(CH2)3COOH (190 mg, 2 eq.), eight-arm polyethylene glycol (PEG, 20 kDa, 4.0 g, 1.0 eq.), carbodiimide hydrochloride (EDCI, 6.94 g, 200 eq.), and p-dimethylaminopyridine (DMAP, 5.30 g, 240 eq.) were placed in a round-bottom flask, and 30 mL of dichloromethane solvent was added. The reaction mixture was stirred overnight at room temperature. The mixture was then extracted, washed twice with dilute hydrochloric acid, three times with saturated brine, and precipitated three times in diethyl ether. The final product yielded 1.84 g, with a yield of 44.1%. The degree of substitution of PEG-CPT was determined by UV-Vis spectrophotometry, showing an average of 1.7 CPT functional groups substituted on each eight-arm polyethylene glycol arm.

[0036] Example 3 Synthesis of PEG-CPT-methacrylate

[0037]

[0038] PEG(CPT) 1.7 (OH) 6.3 1.82 g (0.0912 mmol), methacrylic anhydride (354 mg, 2.296 mmol), triethylamine (Et3N) (349 mg, 3.449 mmol), and p-dimethylaminopyridine (DMAP) (58 mg, 0.475 mmol) were placed in a round-bottom flask, and 10 mL of dichloromethane solvent was added. The reaction mixture was stirred at room temperature for 3 h. The resulting solution was concentrated and precipitated three times in diethyl ether. The final product was 1.57 g, with a yield of 86.3%. The ¹H NMR spectra of PEG-CPT and PEG-CPT-methacrylate are shown below. Figure 1 As shown.

[0039] Example 4: Synthesis of the eight-arm polyethylene glycol-camptothecin coupling compound PEG-CPT

[0040]

[0041] CPT-OOC(CH2)3COOH (123.5 mg, 2 eq.), PEG (3.10 g, 1 eq.), EDCI (2.55 g, 100 eq.), and DMAP (2.09 g, 120 eq.) were placed in a round-bottom flask, and 30 mL of dichloromethane solvent was added. The reaction mixture was stirred overnight at room temperature. The product was then extracted, washed twice with dilute hydrochloric acid, twice with saturated brine, and precipitated three times in diethyl ether. The final product yielded 2.24 g, with a yield of 69.6%. 1 ¹H NMR was used to determine the degree of substitution of PEG-CPT, and on average, each eight-arm polyethylene glycol was substituted with 1.5 CPT functional groups.

[0042] Example 5 Synthesis of PEG-CPT-acrylate

[0043]

[0044] PEGCPT 1.5 (OH) 6.5 (1.8 g, 0.0902 mmol), triethylamine (309 mg, 3.032 mmol), and DAMP (51 mg, 0.417 mmol) were added sequentially to the reaction flask. The mixture was then placed in an ice-water bath. Diluted acrylic anhydride (255 mg, 2.022 mmol) was added through a constant-pressure dropping funnel. After stirring in an ice-water bath for 4 hours, the mixture was allowed to settle three times with ice-cold ether. The product was then dried under vacuum to yield 1.47 g. The 1H NMR spectra of PEG-CPT and PEG-CPT-acrylate are shown below. Figure 2 As shown.

[0045] Example 6: Preparation of drug-loaded gel and determination of drug release

[0046] Redox systems initiate gel polymerization:

[0047] The PEG-CPT-methacrylate or PEG-CPT-acrylate prepared in Examples 3 and 5 were respectively prepared into 20% (w / w) solutions using 0.1M PBS. APS initiator was added to a final concentration of 2 mg / ml, followed by TEMED accelerator to a final concentration of 2 μl / ml. The mixture was then placed in a 37°C water bath, and the curing time and gel mass were recorded. Ten times the gel volume of PBS solution was added, and the sustained-release solution was collected at 37°C using a shaker. Extracts were collected at 2 h, 6 h, 1 d, 2 d, 4 d, and 5 d, and the CPT content in the extracts was determined using a standard UV absorption curve of camptothecin to obtain the sustained-release curve. Figure 3 The extraction solution was changed at each time point, and the gel mass at each time point was recorded. The ratio of the gel mass at different time points to the initial gel mass was calculated to obtain the swelling curve. Figure 4 ).

[0048] Photoinitiation system initiates gel polymerization:

[0049] The PEG-CPT-methacrylate or PEG-CPT-acrylate prepared in Examples 3 and 5 were respectively prepared into 20% solutions with 0.1M PBS. Photoinitiator I2959 or LAP was added to a final concentration of 5 mg / ml. Gel polymerization was initiated using an LED UV lamp (wavelength 365 nm). The curing time after mixing was recorded, and the gel mass was recorded after curing. Ten times the gel volume of PBS solution was added, and the sustained-release solution was collected at 37°C in a shaker. Extracts were collected at 2 h, 6 h, 1 d, 2 d, 4 d, and 5 d, and the CPT content in the extracts was tested using the standard UV absorption curve of camptothecin to obtain the sustained-release curve. Figure 3 The extraction solution was changed at each time point, and the gel mass at each time point was recorded. The ratio of the gel mass at different time points to the initial gel mass was calculated to obtain the swelling curve. Figure 4 ).

[0050] As shown in Table 1, both the UV initiation system and the APS / TEMED initiation system successfully initiated gel polymerization. Higher acrylate substitution rates resulted in faster gelation; acrylates exhibited higher reactivity than methacrylates, leading to significantly faster gelation.

[0051] The drug release curves of the PEG-CPT-methacrylate and PEG-CPT-acrylate polymerized gels are as follows: Figure 3 As shown, the in vitro swelling curve of the gel is as follows. Figure 4 As shown.

[0052] The results showed that the substitution rate, the number and type of acrylate grafts all significantly affected the release of camptothecin. The slowest drug release was observed when the CPT substitution number of the acrylate gel was 4 and when the acrylate substitution number was 3. The gel swelled between 40% and 120% within 24 hours, then reached an equilibrium period, maintaining its morphology without significant changes. Figure 4 ).

[0053] Table 1: Gel Preparation Table

[0054]

[0055] Example 7: Application of the PEG-CPT-methacrylate described in this invention

[0056] Refer to Table 1 of Example 6 to prepare gel 6 solution. The solution is coated onto the surface of the nanofiber catheter by dip coating. The coating thickness can be increased by multiple dip coatings. The coating is cured by ultraviolet light irradiation (wavelength 365nm) to obtain a catheter with camptothecin drug-loaded gel coating.

Claims

1. A method for preparing camptothecin-coupled polyethylene glycol acrylate hydrogel, characterized in that... Cross-linked network polyethylene glycol polyacrylate hydrogels were prepared by reacting acrylic anhydride or alkyl-substituted acrylic anhydride with star-shaped multi-arm PEG coupled with camptothecin via free radical solution polymerization, initiated by redox system or photoinitiation.

2. The preparation method according to claim 1, characterized in that... The star-shaped multi-arm PEG coupled with camptothecin is prepared by reacting camptothecin with a dicarboxylic acid anhydride or a dicarboxylic acid and then coupling it with a star-shaped multi-arm PEG. The anhydride is selected from succinic anhydride, glutaric anhydride, adipic anhydride, pimelic anhydride, octanoic anhydride, azelaic anhydride or sebacic anhydride, and the dicarboxylic acid is HOOC(CH2)nCOOH, where n = 2-6.

3. The preparation method according to claim 2, characterized in that... The molar ratio of the star-shaped multi-arm PEG of the coupled camptothecin to the dicarboxylic acid anhydride or dicarboxylic acid is 1:1.1-7.

4. The preparation method according to claim 1, characterized in that... The star-shaped multi-arm PEG coupled with camptothecin has the following structure: Where n represents a positive integer from 1 to 10, and PEG is star-shaped multi-arm polyethylene glycol.

5. The preparation method according to claim 1, characterized in that... The star-shaped multi-arm polyethylene glycol has 2-8 arms, and the molecular weight of each arm is 1000-5000 Da.

6. The preparation method according to claim 1, characterized in that... The alkyl-substituted acrylic anhydride is methacrylic anhydride.

7. The preparation method according to claim 1, characterized in that... The redox system is an ammonium disulfate / tetramethylethylenediamine redox system; the photoinitiation uses water-soluble photoinitiators 2-hydroxy-4'-(2-hydroxyethoxy)-2-methylphenylacetone or lithium phenyl (2,4,6-trimethylbenzoyl)phosphate, and is initiated under ultraviolet light.

8. The preparation method according to claim 1, characterized in that... The reaction temperature is 4-60℃, and the reaction time is 1 second to 30 minutes.

9. A camptothecin-coupled polyethylene glycol acrylate hydrogel, characterized in that... It is prepared by the method described in any one of claims 1-8.

10. The application of the camptothecin-coupled polyethylene glycol acrylate hydrogel according to claim 9 in the preparation of antitumor drugs.

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