A polylactic acid-based drug-loaded preparation and a preparation method thereof

Abstract

The application relates to the technical field of pharmaceutical preparations, in particular to a polylactic acid-based drug-loaded preparation and a preparation method thereof. The preparation method of the polylactic acid-based drug-loaded preparation comprises the following steps: (1) adding a polylactic acid-based carrier and fangchinese thalictrine into deionized water to obtain an aqueous phase; adding a hydrophobic drug into an organic solvent to obtain an oil phase; (2) uniformly mixing the aqueous phase and the oil phase, and then sequentially performing dialysis, ultrasonic treatment, filtration and freeze-drying to obtain the polylactic acid-based drug-loaded preparation. The polylactic acid-based drug-loaded preparation can intelligently release the effective load, and a large amount of hydrophobic drugs can be released in a short time after reaching tumor tissues, so that the treatment effect can be achieved.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical formulation technology, and in particular to a polylactic acid-based drug-loaded formulation and its preparation method. Background Technology

[0002] Most anticancer / antitumor drugs (such as paclitaxel and camptothecin) exhibit significant hydrophobicity, and their poor water solubility severely limits their clinical application. In recent years, the rapidly developing polymeric micelle drug delivery systems have provided a new strategy for solving the solubility problem of hydrophobic drugs and achieving synergistic effects while reducing toxicity. Polymer micelles are formed by the self-assembly of amphiphilic block copolymers or graft copolymers under non-covalent bonding, possessing a typical core-shell structure. Their hydrophobic core can serve as a reservoir for hydrophobic drugs, effectively improving drug solubility. Compared to traditional formulations, polymeric micelle nanomedicines exhibit superior pharmacokinetic properties, good biocompatibility, and are easily modified for multifunctionality.

[0003] Polylactic acid (PLA) is a polyester polymer synthesized from lactic acid. It possesses excellent biocompatibility, biodegradability, and mechanical properties, making it a commonly used carrier material for constructing polymeric micelle nanoparticles. However, due to its inherent hydrophobicity, hydrophilic segments are typically introduced to improve its hydrophilicity. Polyethylene glycol monomethyl ether-polylactic acid (mPEG-PLA) block copolymers are the most widely used. However, current mPEG-PLA-based polymeric micelles still have some shortcomings: such as large particle size, wide distribution, poor stability after resolvation, limited drug loading capacity, and lack of intelligent stimulus responsiveness. Therefore, it is essential to improve PLA-based carriers to address these issues. Summary of the Invention

[0004] In order to overcome the shortcomings of the prior art, one of the objectives of this invention is to provide a simple method for preparing polylactic acid-based drug-loaded formulations.

[0005] The second objective of this invention is to provide a polylactic acid-based drug delivery formulation that can intelligently respond to and release the effective load in the tumor-specific redox microenvironment, and release a large amount of hydrophobic drug in a short time after reaching the tumor tissue, thereby exerting a therapeutic effect.

[0006] One of the objectives of this invention is achieved through the following technical solution:

[0007] A method for preparing a polylactic acid-based drug-loaded formulation includes the following steps:

[0008] (1) Polylactic acid-based carrier and fenslerophane alkaloid were added to deionized water to obtain an aqueous phase; the hydrophobic drug was added to an organic solvent to obtain an oil phase;

[0009] (2) The aqueous phase and the oil phase are mixed and stirred evenly, and then dialyzed, sonicated, filtered and freeze-dried in sequence to obtain the polylactic acid-based drug-loaded formulation;

[0010] The structural formula of the polylactic acid-based carrier is as follows:

[0011] .

[0012] Preferably, the polylactic acid-based carrier is prepared as follows:

[0013] Modified polylactic acid was added to a dichloromethane / methanol solution, and then modified polyethylene glycol was added in 3-5 portions while stirring in the dark. After the reaction was completed, an iodine methanol solution was added, and the polylactic acid-based carrier was obtained after purification.

[0014] The modified polylactic acid has the following structural formula:

[0015]

[0016] The modified polyethylene glycol has the following structural formula:

[0017] .

[0018] Preferably, the mass ratio of modified polylactic acid to modified polyethylene glycol is 1:(0.8-1.5); the concentration of the iodine methanol solution is 0.03-0.05 g / mL; the stirring reaction time is 10-12 h; and the volume ratio of dichloromethane to methanol in the dichloromethane / methanol solution is 1:1.

[0019] Preferably, the preparation process of the modified polylactic acid is as follows:

[0020] Polylactic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine were added to dichloromethane and stirred at room temperature for 3-5 hours. Then, 3,3'-dithiobis(propionylhydrazine) was added and the reaction was continued for 48-60 hours. After the reaction was completed, the mixture was purified to obtain the modified polylactic acid.

[0021] Preferably, the mass ratio of polylactic acid, 3,3'-dithiobis(propionylhydrazine), EDC hydrochloride, N-hydroxysuccinimide, and triethylamine is 1:(0.036-0.072):(0.03-0.058):(0.018-0.035):(0.015-0.03); and the number average molecular weight of the polylactic acid is 10,000.

[0022] Preferably, the modified polyethylene glycol is prepared by the following method:

[0023] S1. Add L-cysteine ​​and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide to N,N-dimethylformamide, then add 4-dimethylaminopyridine and stir for 0.5-1 h, then add polyethylene glycol and react at 40-50 °C for 20-24 h. After the reaction is complete, purify to obtain intermediate 1.

[0024] The structural formula of intermediate 1 is as follows:

[0025] ;

[0026] S2. Add 5,7-dodecadiyneic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine to N,N-dimethylformamide, stir at 45-50°C for 0.5-1 h, then add intermediate 1 and continue stirring for 48-60 h. After the reaction is complete, purify to obtain the modified polyethylene glycol.

[0027] Preferably, in step S1, the mass ratio of polyethylene glycol, L-cysteine, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, and 4-dimethylaminopyridine is 1:(0.02-0.03):(0.03-0.04):(0.02-0.03); and in step S2, the amount ratio of intermediate 1, 5,7-dodecanoic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine is 1:(0.04-0.07):(0.03-0.06):(0.02-0.04):(0.02-0.03).

[0028] Preferably, in step (1), the mass ratio of polylactic acid-based carrier, fenestrated thalidomide, and hydrophobic drug is 100:(4-8):(12-15); the concentration of the polylactic acid-based carrier in the aqueous phase is 5-10 mg / mL; the concentration of the hydrophobic drug in the oil phase is 2-5 mg / mL; the hydrophobic drug is selected from one of paclitaxel, docetaxel, anastrozole, and letrozole; and the organic solvent is selected from one of methanol, ethanol, acetonitrile, acetone, dichloromethane, and dimethyl sulfoxide.

[0029] Preferably, the stirring time in step (2) is 3-8 hours; the dialysis time is 24-48 hours; and the ultrasonic treatment time is 10-15 minutes.

[0030] The second objective of this invention is achieved by the following technical solution:

[0031] A polylactic acid-based drug-loaded formulation was prepared using the above-described method.

[0032] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0033] 1. This invention uses polylactic acid as a hydrophobic backbone and modifies polyethylene glycol through a thiol-disulfide bond exchange reaction to construct a redox-responsive polymer drug-loaded formulation. This formulation intelligently responds to the tumor-specific redox microenvironment, releasing its effective load and releasing large amounts of hydrophobic drugs, such as paclitaxel, shortly after reaching tumor tissue to exert a therapeutic effect. Specifically, when the drug-loaded formulation reaches tumor lesions with high GSH (glutathione) / ROS (free radical) levels, the disulfide bonds break, triggering the disintegration of the hydrophobic core, achieving controlled drug release, thereby improving therapeutic efficacy, reducing systemic toxicity, and enhancing drug bioavailability. To further endow the drug-loaded formulation with targeting and antioxidant capabilities for lesions, this invention also uses 5,7-dodecadiyneic acid to hydrophobically modify PEG: 5,7-dodecadiyneic acid contains a conjugated diyne structure (–C≡C–C≡C–), which has free radical affinity, thereby improving the targeting of the drug-loaded formulation to lesions. It can also undergo free radical addition with excess ROS in the lesion area, rapidly clearing ROS and alleviating oxidative stress; the rigid diyne skeleton of 5,7-dodecadiyneic acid can also restrict the movement of the core chain segments, keeping the micelles in a dense structure in the blood circulation and reducing premature drug release; upon reaching the target site, the increased ROS triggers the breakage of disulfide bonds, the rigid barrier is removed, and the drug is rapidly released, achieving a "silent-burst" delivery.

[0034] 2. This invention also introduces fenestrated thalidomide into the polylactic acid matrix. The cationic group in this molecule can form an electrostatic interaction with the carboxyl anion of 5,7-dodecanediyne in the polylactic acid matrix. At the same time, its hydrophobic parent nucleus can generate π-π stacking and hydrophobic interaction with hydrophobic drugs. The two work together to effectively expand the micelle core space, improve the compatibility of the micro-region with the drug, thereby increasing the drug loading and improving the stability of the drug load.

[0035] 3. The method for preparing polylactic acid-based drug-loaded formulations provided by this invention is simple and easy to mass-produce. Furthermore, the resulting drug-loaded formulations are regularly spherical with small particle size and narrow particle size distribution, exhibiting good colloidal stability and making them suitable for biomedical applications such as intravenous administration. Attached Figure Description

[0036] Figure 1 The infrared spectrum of the modified polyethylene glycol prepared in Example 4 of this invention;

[0037] Figure 2 SEM image of the polylactic acid-based carrier prepared in Example 7 of this invention;

[0038] Figure 3 The infrared spectrum of the polylactic acid-based support prepared in Example 7 of this invention;

[0039] Figure 4This is a SEM image of the polylactic acid-based drug-loaded formulation prepared in Example 1 of the present invention. Detailed Implementation

[0040] The present invention will be further described below with reference to the accompanying drawings and specific embodiments. It should be noted that, without conflict, the various embodiments or technical features described below can be arbitrarily combined to form new embodiments. Specific conditions not specified in the embodiments shall be performed according to conventional conditions or conditions recommended by the manufacturer. Unless otherwise specified, all reagents or instruments used are conventional products obtained through commercial channels.

[0041] Preparation Example 1

[0042] This preparation example provides a modified polylactic acid, prepared by the following method:

[0043]

[0044] According to the ratio of polylactic acid, 3,3'-dithiobis(propionylhydrazine), EDC hydrochloride, N-hydroxysuccinimide, triethylamine, and dichloromethane of 100 mg: 5 mg: 4 mg: 2 mg: 2 mg: 4 mL, polylactic acid (Mn=10000), EDC hydrochloride (CAS: 107-06-2), N-hydroxysuccinimide (CAS: 6066-82-6), and triethylamine were added to dichloromethane and stirred at room temperature for 4 h to obtain activated polylactic acid. Then, 3,3'-dithiobis(propionylhydrazine) (CAS: 50906-77-9) was added and the reaction was stirred for another 52 h. After the reaction was completed, the solid product was precipitated with ice-cold diethyl ether, washed with cold methanol, and then dried under vacuum to obtain the modified polylactic acid.

[0045] Preparation Example 2

[0046] This preparation example provides a modified polylactic acid, prepared by the following method:

[0047] According to the following ratio of polylactic acid, 3,3'-dithiobis(propionylhydrazine), EDC hydrochloride, N-hydroxysuccinimide, triethylamine, and dichloromethane (100 mg: 3.6 mg: 3 mg: 1.8 mg: 1.5 mg: 3 mL), polylactic acid (Mn=10000), EDC hydrochloride, N-hydroxysuccinimide, and triethylamine were added to dichloromethane and stirred at room temperature for 5 h to obtain activated polylactic acid. Then, 3,3'-dithiobis(propionylhydrazine) was added and the reaction was stirred for another 48 h. After the reaction was completed, the solid product was precipitated with ice-cold diethyl ether, washed with cold methanol, and then dried under vacuum to obtain the modified polylactic acid.

[0048] Preparation Example 3

[0049] This preparation example provides a modified polylactic acid, prepared by the following method:

[0050] According to the following ratio of polylactic acid, 3,3'-dithiobis(propionylhydrazine), EDC hydrochloride, N-hydroxysuccinimide, triethylamine, and dichloromethane (100 mg: 7.2 mg: 5.8 mg: 3.5 mg: 3 mg: 5 mL), polylactic acid (Mn=10000), EDC hydrochloride, N-hydroxysuccinimide, and triethylamine were added to dichloromethane and stirred at room temperature for 3 h to obtain activated polylactic acid. Then, 3,3'-dithiobis(propionylhydrazine) was added and the reaction was stirred for another 60 h. After the reaction was completed, the solid product was precipitated with ice-cold diethyl ether, washed with cold methanol, and then dried under vacuum to obtain the modified polylactic acid.

[0051] Preparation Example 4

[0052] This preparation example provides a modified polyethylene glycol, prepared by the following method:

[0053]

[0054] S1. According to the ratio of polyethylene glycol, L-cysteine, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, 4-dimethylaminopyridine, and N,N-dimethylformamide of 300 mg: 7 mg: 9 mg: 8 mg: 12 mL, L-cysteine ​​and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (CAS: 1892-57-5) were added to N,N-dimethylformamide, then 4-dimethylaminopyridine was added and stirred for 0.8 h, then polyethylene glycol (Mn=6000) was added, and the reaction was carried out at 45 °C for 22 h. The reaction was terminated by adding deionized water, dialyzed with deionized water, and freeze-dried to obtain intermediate 1;

[0055] S2. According to the ratio of intermediate 1, 5,7-dodecadienoic acid, EDC hydrochloride, N-hydroxysuccinimide, triethylamine, and N,N-dimethylformamide, 5,7-dodecadienoic acid (CAS: 28393-04-6), EDC hydrochloride, N-hydroxysuccinimide, and triethylamine to N,N-dimethylformamide, the mixture was stirred at 45°C for 0.8 h. Then, intermediate 1 was added and the mixture was stirred for 48 h. After the reaction was completed, the solid product was precipitated with ice-cold diethyl ether, washed successively with n-hexane, diethyl ether, and anhydrous ethanol, and dried under vacuum to obtain the modified polyethylene glycol.

[0056] Preparation Example 5

[0057] This preparation example provides a modified polyethylene glycol, prepared by the following method:

[0058] S1. According to the ratio of polyethylene glycol, L-cysteine, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, 4-dimethylaminopyridine, and N,N-dimethylformamide of 300 mg: 6 mg: 8.5 mg: 6.5 mg: 10 mL, L-cysteine ​​and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide were added to N,N-dimethylformamide, then 4-dimethylaminopyridine was added and stirred for 0.5 h, then polyethylene glycol (Mn=6000) was added, and the reaction was carried out at 40 °C for 24 h. The reaction was terminated by adding deionized water, dialyzed with deionized water, and freeze-dried to obtain intermediate 1;

[0059] S2. According to the ratio of intermediate 1, 5,7-dodecadienoic acid, EDC hydrochloride, N-hydroxysuccinimide, triethylamine, and N,N-dimethylformamide, 5,7-dodecadienoic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine were 150 mg: 6.8 mg: 5.8 mg: 3.5 mg: 3 mg: 25 mL, 5,7-dodecadienoic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine were added to N,N-dimethylformamide and stirred at 50 °C for 0.5 h. Then, intermediate 1 was added and the reaction was stirred for 60 h. After the reaction was completed, the solid product was precipitated with ice-cold diethyl ether, and then washed successively with n-hexane, diethyl ether, and anhydrous ethanol. The product was then dried under vacuum to obtain the modified polyethylene glycol.

[0060] Preparation Example 6

[0061] This preparation example provides a modified polyethylene glycol, prepared by the following method:

[0062] S1. According to the ratio of polyethylene glycol, L-cysteine, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, 4-dimethylaminopyridine, and N,N-dimethylformamide of 300 mg: 8.5 mg: 11 mg: 9 mg: 15 mL, L-cysteine ​​and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide were added to N,N-dimethylformamide, then 4-dimethylaminopyridine was added and stirred for 1 h, then polyethylene glycol (Mn=6000) was added, and the reaction was carried out at 50 °C for 20 h. The reaction was terminated by adding deionized water, dialyzed with deionized water, and freeze-dried to obtain intermediate 1;

[0063] S2. According to the ratio of intermediate 1, 5,7-dodecadienoic acid, EDC hydrochloride, N-hydroxysuccinimide, triethylamine, and N,N-dimethylformamide, 5,7-dodecadienoic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine to N,N-dimethylformamide, the mixture was stirred at 45°C for 1 h, and then intermediate 1 was added and stirred for 48 h. After the reaction was completed, the solid product was washed successively with n-hexane, diethyl ether, and anhydrous ethanol, and then dried under vacuum to obtain the modified polyethylene glycol.

[0064] Preparation Example 7

[0065] This preparation example provides a polylactic acid-based carrier, the structural formula of which and its preparation method are as follows:

[0066]

[0067] With modified polylactic acid, modified polyethylene glycol, and dichloromethane / methanol solution in a ratio of 5 mg:4.5 mg:1 mL, the modified polylactic acid prepared in Preparation Example 1 was added to the dichloromethane / methanol solution (v / v=1:1), and then the modified polyethylene glycol prepared in Preparation Example 4 was added in four portions. The mixture was stirred at room temperature in the dark for 11 h, and then a methanol solution of iodine with a concentration of 0.04 g / mL was added until the system turned a stable light yellow color. The solid product was then precipitated with ice-cold diethyl ether, redissolved with dimethyl sulfoxide, dialyzed for 3 days, and then freeze-dried to obtain the polylactic acid-based carrier.

[0068] Preparation Example 8

[0069] This preparation example provides a polylactic acid-based carrier, which is prepared by the following method:

[0070] With modified polylactic acid and modified polyethylene glycol dichloromethane / methanol solution in a ratio of 10 mg:8 mg:1.5 mL, the modified polylactic acid prepared in Preparation Example 2 was added to the dichloromethane / methanol solution (v / v=1:1), and then the modified polyethylene glycol prepared in Preparation Example 5 was added in three portions. The mixture was stirred at room temperature in the dark for 10 h, and then iodine in methanol solution with a concentration of 0.03 g / mL was added until the system turned a stable light yellow color. The solid product was then precipitated with ice-cold diethyl ether, redissolved with dimethyl sulfoxide, dialyzed for 3 days, and then freeze-dried to obtain the polylactic acid-based carrier.

[0071] Preparation Example 9

[0072] This preparation example provides a polylactic acid-based carrier, which is prepared by the following method:

[0073] With a modified polylactic acid and modified polyethylene glycol dichloromethane / methanol solution in a ratio of 10 mg:15 mg:3 mL, the modified polylactic acid prepared in Preparation Example 3 was added to the dichloromethane / methanol solution (v / v=1:1), and then the modified polyethylene glycol prepared in Preparation Example 6 was added in 5 portions. The mixture was stirred at room temperature in the dark for 12 h, and then a methanol solution of iodine with a concentration of 0.05 g / mL was added until the system turned a stable light yellow color. The solid product was then precipitated with ice-cold diethyl ether, redissolved with dimethyl sulfoxide, dialyzed for 3 days, and then freeze-dried to obtain the polylactic acid-based carrier.

[0074] Example 1

[0075] This embodiment provides a method for preparing a polylactic acid-based drug-loaded formulation, comprising the following steps:

[0076] (1) According to the mass ratio of polylactic acid-based carrier, fennthrine, and paclitaxel of Preparation Example 7, the polylactic acid-based carrier and fennthrine were added to deionized water to obtain an aqueous phase, and the concentration of polylactic acid-based carrier in the aqueous phase was 8 mg / mL; paclitaxel was added to dichloromethane to obtain an oil phase, and the concentration of paclitaxel in the oil phase was 4 mg / mL;

[0077] (2) Mix the aqueous phase and the oil phase, stir for 5 hours, then transfer to a dialysis bag, dialyze in deionized water for 36 hours, then sonicate for 12 minutes, filter through a 0.22 μm microporous membrane, and freeze dry to obtain the polylactic acid-based drug-loaded formulation.

[0078] This embodiment also provides a polylactic acid-based drug-loaded formulation, which is prepared using the above-described preparation method.

[0079] Example 2

[0080] This embodiment provides a method for preparing a polylactic acid-based drug-loaded formulation, comprising the following steps:

[0081] (1) According to the mass ratio of polylactic acid-based carrier, fennthrine, and paclitaxel of Preparation Example 8, the polylactic acid-based carrier and fennthrine were added to deionized water to obtain an aqueous phase, and the concentration of polylactic acid-based carrier in the aqueous phase was 5 mg / mL; paclitaxel was added to dichloromethane to obtain an oil phase, and the concentration of paclitaxel in the oil phase was 2 mg / mL;

[0082] (2) Mix the aqueous phase and the oil phase, stir for 3 hours, then transfer to a dialysis bag, dialyze in deionized water for 24 hours, then sonicate for 10 minutes, filter through a 0.22 μm microporous membrane, and freeze dry to obtain the polylactic acid-based drug-loaded formulation.

[0083] This embodiment also provides a polylactic acid-based drug-loaded formulation, which is prepared using the above-described preparation method.

[0084] Example 3

[0085] This embodiment provides a method for preparing a polylactic acid-based drug-loaded formulation, comprising the following steps:

[0086] (1) According to the mass ratio of polylactic acid-based carrier, fennthrine, and paclitaxel of Preparation Example 9, polylactic acid-based carrier and fennthrine were added to deionized water to obtain an aqueous phase, and the concentration of polylactic acid-based carrier in the aqueous phase was 10 mg / mL; paclitaxel was added to dichloromethane to obtain an oil phase, and the concentration of paclitaxel in the oil phase was 5 mg / mL;

[0087] (2) Mix the aqueous phase and the oil phase, stir for 8 hours, then transfer to a dialysis bag, dialyze in deionized water for 48 hours, then sonicate for 15 minutes, filter through a 0.22 μm microporous membrane, and freeze dry to obtain the polylactic acid-based drug-loaded formulation.

[0088] This embodiment also provides a polylactic acid-based drug-loaded formulation, which is prepared using the above-described preparation method.

[0089] Comparative Example 1

[0090] The difference between this comparative example and Example 1 is that polylactic acid-polyethylene glycol is used instead of the polylactic acid-based carrier prepared in Example 7.

[0091] The preparation method of polylactic acid-polyethylene glycol is as follows: Polylactic acid, polyethylene glycol, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine are prepared in a mass ratio of 100 mg: 120 mg: 5.8 mg: 3.5 mg: 3 mg. Under nitrogen protection, polylactic acid (Mn=10000), EDC hydrochloride, and N-hydroxysuccinimide are dissolved in dichloromethane and stirred for 0.8 h to obtain a mixed solution with a polylactic acid concentration of 35 mg / mL. Then, triethylamine and a dichloromethane solution of polyethylene glycol (Mn=6000) with a concentration of 20 mg / mL are added. The mixture is reacted at room temperature for 24 h. After the reaction is complete, the product is precipitated with ice-cold diethyl ether, redissolved with dimethyl sulfoxide, dialyzed with deionized water, and freeze-dried to obtain polylactic acid-polyethylene glycol.

[0092] Comparative Example 2

[0093] The difference between this comparative example and Example 1 is that no fenestrated alkaloid was added in step (1).

[0094] Experimental Example 1

[0095] The modified polyethylene glycol obtained in Preparation Example 4 was analyzed by Fourier transform infrared spectroscopy (FT-IR), and the results are as follows: Figure 1As shown; the polylactic acid-based support prepared in Example 7 was characterized by SEM and analyzed by Fourier transform infrared spectroscopy (FT-IR), and the results are as follows. Figure 2-3 As shown in Figure 4, the polylactic acid-based drug-loaded formulation obtained in Example 1 was characterized by SEM.

[0096] Figure 1 The images show the infrared spectra of the modified polyethylene glycol obtained in Preparation Example 4. Curve 1 is the infrared spectrum of unmodified polyethylene glycol, curve 2 is the infrared spectrum of intermediate 1, and curve 3 is the infrared spectrum of modified polyethylene glycol. (Observation) Figure 1 It is known that, compared to polyethylene glycol, intermediate 1 is at 3485 cm⁻¹ -1 2570cm -1 1646cm -1 The presence of characteristic absorption peaks for amino NH stretching vibration, -SH, and NH bending vibration at 3500 cm⁻¹ indicates the successful preparation of intermediate 1; the modified polyethylene glycol shows a peak at 3500 cm⁻¹. -1 2570cm -1 2240cm -1 Characteristic absorption peaks for carboxyl hydroxyl groups, -SH, and C≡C appear at 1646 cm⁻¹; -1 and 1548cm -1 The presence of a characteristic absorption peak for amide bonds indicates that the modified polyethylene glycol was successfully prepared.

[0097] Figure 2 This is a SEM image of the polylactic acid-based carrier prepared in Example 7. Figure 3 The images show the infrared spectra of the polylactic acid-based support prepared in Example 7, where curve a is the infrared spectrum of polylactic acid and curve b is the infrared spectrum of the polylactic acid-based support. (Observation) Figure 3 It is known that, compared to polylactic acid, polylactic acid-based carriers have a lower density at 2235 cm⁻¹. -1 1635cm -1 632cm -1 530cm -1 The presence of characteristic absorption peaks for C≡C, NH, CS, and SS indicates that the polylactic acid-based support was successfully prepared.

[0098] Figure 4 This is a SEM image of the polylactic acid-based drug-loaded formulation prepared in Example 1. (Observation) Figure 4 It can be seen that the obtained polylactic acid-based drug-loaded formulation is in the shape of a regular sphere.

[0099] Experiment Example 2

[0100] The preparations obtained in Examples 1-3 and Comparative Examples 1-2 were reconstituted and diluted with phosphate buffer at pH 7.4 to a clinical drug concentration of 0.5-2 mg / mL, and stored at 4°C in the dark. The particle size was measured using a laser particle size analyzer after 0 months, 1 month, and 3 months of storage. The results are shown in Table 1.

[0101] Table 1

[0102]

[0103] As shown in Table 1, compared with Comparative Example 2, which omits the alkaloid fenestrate, the polylactic acid-based drug-loaded formulation obtained in this invention has better stability at 4°C.

[0104] Experimental Example 3

[0105] Accurately weigh 2 mg of the preparations from Examples 1-3 and Comparative Examples 1-2, transfer them to a 2 mL volumetric flask, add acetonitrile to dissolve, sonicate for 15 min, and make up to volume. Filter through a 0.22 μm microporous membrane to obtain the sample for testing.

[0106] The chromatographic conditions were as follows: mobile phase: water = 70:30; flow rate: 1 mL / min; injection volume: 20 μL; detection wavelength: 227 nm; column temperature: 30 ℃. The content of paclitaxel (PTX) in the sample was determined, and the encapsulation efficiency and drug loading were calculated. Specifically, encapsulation efficiency (%) = PTX content in the formulation / drug dosage × 100%; drug loading (%) = PTX content in the formulation / total mass of the formulation × 100%. The results are shown in Table 2.

[0107] Table 2

[0108]

[0109] As shown in Table 2, compared with Comparative Example 2, which omits the use of fenestrated thalidomide, the polylactic acid-based drug-loaded formulation obtained in this invention has a higher encapsulation efficiency and drug loading.

[0110] Analysis of the experimental data in Table 1-2 shows that when fenestrated thalidomide is introduced into the polylactic acid matrix, the cationic groups in the molecule can form electrostatic interactions with the carboxyl anion of 5,7-dodecanediyne in the polylactic acid matrix. At the same time, its hydrophobic parent nucleus can generate π-π stacking and hydrophobic interactions with hydrophobic drugs. The synergy of these two factors can effectively expand the micelle core space, improve the compatibility of the micro-region with the drug, thereby increasing the drug loading capacity and improving the stability of the drug load.

[0111] Experiment Example 4

[0112] The formulations of Examples 1-3 and Comparative Examples 1-2 were placed in dialysis bags, and then added to phosphate buffer solution with pH=7.4. Different concentrations of glutathione (GSH) were added as release media. The dialysis system was placed in a constant temperature shaking incubator at 37°C. 1 mL of release media was collected at 1 h, 5 h, 10 h, 20 h, 24 h, and 48 h (and an equal amount of release media was added). 50 μL of the released medium was injected into HPLC to determine the paclitaxel content in the released solution and calculate the cumulative release amount. The chromatographic conditions were the same as in Experiment 3. The results are shown in Table 3.

[0113] Table 3

[0114]

[0115] As shown in Table 3, compared to Comparative Example 1, which uses polylactic acid-polyethylene glycol instead of the polylactic acid-based carrier prepared in Preparation Example 7, the polylactic acid-based drug-loaded formulation obtained in this invention can intelligently respond to and release the effective load in the tumor-specific redox microenvironment. It can release a large amount of paclitaxel shortly after reaching the tumor tissue, thus exerting a therapeutic effect. This is because this invention uses polylactic acid as a hydrophobic framework and grafts modified polyethylene glycol through a thiol-disulfide bond exchange reaction to construct a redox-responsive polymer drug-loaded formulation. Specifically, when the drug-loaded formulation reaches tumor lesions with high GSH (glutathione) / ROS (free radical) levels, the disulfide bonds break, triggering the disintegration of the hydrophobic core, achieving controlled drug release, thereby improving therapeutic efficacy, reducing systemic toxicity, and enhancing drug bioavailability. To further endow the drug-loaded formulation with targeting and antioxidant capabilities for lesions, this invention also uses 5,7-dodecadiyneic acid to hydrophobically modify PEG: 5,7-dodecadiyneic acid contains a conjugated diyne structure (–C≡C–C≡C–), which has free radical affinity, thereby improving the targeting of the drug-loaded formulation to lesions. It can also undergo free radical addition with excess ROS in the lesion area, rapidly clearing ROS and alleviating oxidative stress; the rigid diyne skeleton of 5,7-dodecadiyneic acid can also restrict the movement of the core chain segments, keeping the micelles in a dense structure in the blood circulation and reducing premature drug release; upon reaching the target site, the increased ROS triggers the breakage of disulfide bonds, the rigid barrier is removed, and the drug is rapidly released, achieving a "silent-burst" delivery.

[0116] The above embodiments are merely preferred embodiments of the present invention and should not be construed as limiting the scope of protection of the present invention. Any non-substantial changes and substitutions made by those skilled in the art based on the present invention shall fall within the scope of protection claimed by the present invention.

Claims

1. A method for preparing a polylactic acid-based drug-loaded formulation, characterized in that, Includes the following steps: (1) Polylactic acid-based carrier and fenslerophane alkaloid were added to deionized water to obtain an aqueous phase; the hydrophobic drug was added to an organic solvent to obtain an oil phase; (2) The aqueous phase and the oil phase are mixed and stirred evenly, and then dialyzed, sonicated, filtered and freeze-dried in sequence to obtain the polylactic acid-based drug-loaded formulation; The structural formula of the polylactic acid-based carrier is as follows: ； The polylactic acid-based carrier is prepared as follows: Modified polylactic acid was added to a dichloromethane / methanol solution, and then modified polyethylene glycol was added in 3-5 portions. The reaction was carried out under light and stirred. After the reaction was completed, an iodine methanol solution was added, and the polylactic acid-based carrier was obtained after purification. The modified polylactic acid has the following structural formula: The modified polyethylene glycol has the following structural formula: 。 2. The method for preparing the polylactic acid-based drug-loaded formulation according to claim 1, characterized in that, The mass ratio of modified polylactic acid to modified polyethylene glycol is 1:(0.8-1.5); the concentration of the iodine methanol solution is 0.03-0.05 g / mL; the stirring reaction time is 10-12 h; and the volume ratio of dichloromethane to methanol in the dichloromethane / methanol solution is 1:

1.

3. The method for preparing the polylactic acid-based drug-loaded formulation according to claim 2, characterized in that, The preparation process of the modified polylactic acid is as follows: Polylactic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine were added to dichloromethane and stirred at room temperature for 3-5 hours. Then, 3,3'-dithiobis(propionylhydrazine) was added and the reaction was continued for 48-60 hours. After the reaction was completed, the mixture was purified to obtain the modified polylactic acid.

4. The method for preparing the polylactic acid-based drug-loaded formulation according to claim 3, characterized in that, The mass ratio of polylactic acid, 3,3'-dithiobis(propionylhydrazine), EDC hydrochloride, N-hydroxysuccinimide, and triethylamine is 1:(0.036-0.072):(0.03-0.058):(0.018-0.035):(0.015-0.03); the number average molecular weight of the polylactic acid is 10,000.

5. The method for preparing the polylactic acid-based drug-loaded formulation according to claim 2, characterized in that, The modified polyethylene glycol is prepared as follows: S1. Add L-cysteine ​​and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide to N,N-dimethylformamide, then add 4-dimethylaminopyridine and stir for 0.5-1 h, then add polyethylene glycol and react at 40-50 °C for 20-24 h. After the reaction is complete, purify to obtain intermediate 1. The structural formula of intermediate 1 is as follows: ； S2. Add 5,7-dodecadiyneic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine to N,N-dimethylformamide, stir at 45-50°C for 0.5-1 h, then add intermediate 1 and continue stirring for 48-60 h. After the reaction is complete, purify to obtain the modified polyethylene glycol.

6. The method for preparing the polylactic acid-based drug-loaded formulation according to claim 5, characterized in that, In step S1, the mass ratio of polyethylene glycol, L-cysteine, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, and 4-dimethylaminopyridine is 1:(0.02-0.03):(0.03-0.04):(0.02-0.03); in step S2, the ratio of intermediate 1,5,7-dodecanoic acid, EDC hydrochloride, N-hydroxysuccinimide, and triethylamine is 1:(0.04-0.07):(0.03-0.06):(0.02-0.04):(0.02-0.03).

7. The method for preparing the polylactic acid-based drug-loaded formulation according to claim 1, characterized in that, In step (1), the mass ratio of polylactic acid-based carrier, fenestrated thalidomide alkaloid, and hydrophobic drug is 100:(4-8):(12-15); the concentration of the polylactic acid-based carrier in the aqueous phase is 5-10 mg / mL; the concentration of the hydrophobic drug in the oil phase is 2-5 mg / mL; the hydrophobic drug is selected from one of paclitaxel, docetaxel, anastrozole, and letrozole; and the organic solvent is selected from one of methanol, ethanol, acetonitrile, acetone, dichloromethane, and dimethyl sulfoxide.

8. The method for preparing the polylactic acid-based drug-loaded formulation according to claim 7, characterized in that, The stirring time in step (2) is 3-8 hours; the dialysis time is 24-48 hours; and the ultrasonic treatment time is 10-15 minutes.

9. A polylactic acid-based drug delivery formulation, characterized in that, It is prepared by the preparation method according to any one of claims 1-8.

Images