Targeted drug delivery platform based on mesoporous cerium oxide nanoparticles, its preparation method and application

Abstract

This invention relates to a targeted drug delivery platform based on mesoporous cerium oxide nanoparticles, its preparation method, and its applications. The platform comprises: mesoporous cerium oxide nanoparticles coordinated with polydopamine; and mannose modified on the surface of the mesoporous cerium oxide nanoparticles. This drug delivery platform not only possesses the ability to scavenge excess ROS and alleviate oxidative stress, but also can target drugs to sites of lung inflammation, significantly improving anti-inflammatory and anti-fibrotic effects, providing new ideas and methods for the treatment of pulmonary fibrosis.

Description

Technical Field

[0001] This invention belongs to the field of biomedical material preparation, specifically relating to a targeted drug delivery platform based on mesoporous cerium oxide nanoparticles, its preparation method, and its application. Background Technology

[0002] Pulmonary fibrosis (PF) is a chronic, progressive disease characterized by the gradual replacement of lung tissue with fibrous tissue and the progressive loss of lung function. According to the World Health Organization (WHO), the global incidence of pulmonary fibrosis is increasing annually, with idiopathic pulmonary fibrosis (IPF) becoming a serious public health problem. The incidence of pulmonary fibrosis varies considerably globally, but in developed regions such as North America and Europe, the annual incidence is approximately 1 to 3 cases per 100,000 people. Epidemiological data indicate that the annual incidence and mortality rates of pulmonary fibrosis are significantly higher in the elderly population, particularly those aged 60 and older, where the mortality rate is significantly higher than in other age groups.

[0003] In China, the incidence of pulmonary fibrosis is also showing an increasing trend year by year. A 2017 study indicated that the incidence of idiopathic pulmonary fibrosis in China is approximately 1.2 per 100,000, suggesting that the potential number of pulmonary fibrosis cases may be underestimated. The chronic progression of pulmonary fibrosis and the lack of effective treatments make it a significant factor leading to lung failure, and clinically, the five-year survival rate for patients with pulmonary fibrosis is typically low, ranging from only 30% to 50%.

[0004] The pathological mechanisms of pulmonary fibrosis involve multiple steps, primarily including oxidative stress, chronic inflammatory response, excessive fibroblast activation, and collagen deposition. Excessive reactive oxygen species (ROS) play a central role in the progression of pulmonary fibrosis; oxidative stress not only promotes fibroblast proliferation and transformation but also exacerbates pulmonary inflammatory responses and the fibrotic process. Therefore, antioxidant therapy has become a key direction in current research on pulmonary fibrosis.

[0005] Current treatments for pulmonary fibrosis primarily focus on anti-fibrotic and anti-inflammatory effects. However, due to the long course and complex etiology of pulmonary fibrosis, existing drugs have limited efficacy and are prone to side effects, necessitating the development of new treatment strategies. This is especially true for idiopathic pulmonary fibrosis, a disease with an unknown etiology, where effective drugs and treatments remain lacking. With the development of nanotechnology, nanomedicine delivery systems have emerged as a new research direction for the treatment of pulmonary fibrosis due to their ability to enhance drug targeting, improve drug bioavailability, and reduce side effects.

[0006] Nanocarrier technology has received widespread attention in recent years. Nanocarriers can optimize drug delivery and release characteristics by modifying their physicochemical properties. Cerium oxide nanozymes (CeO2NPs), due to their enzyme-like activity, can effectively scavenge reactive oxygen species (ROS) generated by oxidative stress, showing great potential in the treatment of pulmonary fibrosis. Cerium oxide nanozymes have been shown to have antioxidant, anti-inflammatory, and anti-fibrotic effects by regulating redox reactions, reducing cellular oxidative damage, and inhibiting pulmonary inflammation. However, cerium oxide nanozymes exhibit poor in vivo stability, rapid distribution and clearance, and poor targeting, limiting their widespread clinical application. Therefore, modifying cerium oxide nanozymes to improve their biocompatibility, targeting, and drug delivery capacity has become an important research direction. Summary of the Invention

[0007] To address the shortcomings of cerium oxide nanozymes, the present invention aims to propose a multifunctional nanoparticle based on cerium oxide nanozymes. By modifying the surface with polydopamine, it can then complex with mannose to form a drug delivery system, which can improve the in vivo stability of cerium oxide, while also enhancing biocompatibility, targeting, and antioxidant properties.

[0008] The specific technical solution of the present invention is as follows:

[0009] A first aspect of the present invention provides a targeted drug delivery platform based on mesoporous cerium oxide nanoparticles, comprising:

[0010] Mesoporous cerium oxide nanoparticles coordinated with polydopamine;

[0011] Mannose modified on the surface of the mesoporous cerium oxide nanoparticles.

[0012] Polydopamine has excellent self-assembly ability and surface functionalization properties. When it is coordinated on the surface of cerium oxide nanozymes, it can synergistically enhance ROS scavenging ability, improve antioxidant capacity, improve biocompatibility, and further enhance its targeting ability through functionalization.

[0013] Mannose plays an important role in cell recognition and signal transduction. In particular, mannose can specifically bind to mannose receptors (such as mannose receptor type C, MR) on the surface of macrophages, and modifying the surface of delivery platforms with it can enhance the targeting of drug delivery systems.

[0014] The drug delivery platform described above enhances targeting through mannose modification, ensuring precise drug delivery to the lesion. It also utilizes the synergistic effect of mesoporous cerium oxide and polydopamine to enhance ROS scavenging capacity, precisely releasing drugs in the acidic microenvironment of pulmonary fibrosis, thereby improving drug utilization and reducing side effects. Furthermore, experiments have shown that cerium oxide and mannose can synergistically improve the stability of cerium oxide nanoparticles, enabling more precise drug delivery to the lung injury area and achieving better therapeutic effects.

[0015] Preferably, the mesoporous cerium oxide nanoparticles are spherical.

[0016] Preferably, the diameter of the mesoporous cerium oxide nanoparticles is 150-200 nm, and the mesopore size is 3-6 nm, which is more conducive to the coordination modification of polydopamine.

[0017] Preferably, the mesoporous cerium oxide nanoparticles are prepared by dispersing trivalent cerium salt in an aqueous ethylene glycol solution, then adding glacial acetic acid and polyvinylpyrrolidone, and carrying out a hydrothermal reaction at 150-200°C.

[0018] The trivalent cerium salt can be a nitrate, such as cerium nitrate.

[0019] Preferably, Ce 3+ Glacial acetic acid: PVP = 0.3-0.35g: 0.6-1mL: 0.6-0.8g;

[0020] Preferably, the hydrothermal reaction time is 16–24 hours;

[0021] Preferably, in the ethylene glycol aqueous solution, the volume ratio of ethylene glycol to water is 2:1.

[0022] A second aspect of the present invention provides a method for preparing the aforementioned targeted drug delivery platform, comprising the following steps:

[0023] (1) Coordinating polydopamine on the surface of mesoporous cerium oxide nanoparticles;

[0024] (2) Mannose is modified on the surface of the mesoporous cerium oxide nanoparticles obtained in step (1).

[0025] Preferably, step (1) includes:

[0026] Mesoporous cerium oxide nanoparticles were mixed with methanol and dopamine, and triethylamine was added. The mixture was refluxed at 60°C and then washed to remove free dopamine, resulting in cerium oxide nanoparticles with surface-coordinated polydopamine, denoted as P@CeO2.

[0027] Preferably, the ratio of cerium oxide nanoparticles:methanol:dopamine:triethylamine is 20 mg:2.5 mL:20 mg:20 μL;

[0028] Preferably, the reflux reaction time is 10–15 h.

[0029] Preferably, step (2) includes:

[0030] P@CeO2 was mixed and dispersed with mannose at room temperature to obtain mannose-modified cerium oxide nanoparticles with surface-coordinated polydopamine, denoted as GMP@CeO2.

[0031] Preferably, in the mixed solution, the concentration of P@CeO2 is 1-2 mg / mL and the concentration of mannose is 3-6 mg / mL;

[0032] Preferably, the dispersion time is 0.5 to 2 hours.

[0033] A third aspect of the invention also provides the application of the aforementioned targeted drug delivery platform in the preparation of drugs for pulmonary fibrosis. For example, in one specific embodiment, the targeted drug delivery platform of the present invention can load drugs such as nintedanib for the treatment of pulmonary fibrosis.

[0034] The beneficial effects of this invention are:

[0035] Compared with existing technologies, the drug delivery platform of this invention, based on cerium oxide nanozymes, utilizes polydopamine surface coordination modification to complex with mannose, forming a drug delivery system targeting macrophages. The synergistic modification of polydopamine and mannose improves the stability of cerium oxide nanoparticles; simultaneously, mesoporous cerium oxide and polydopamine synergistically enhance ROS scavenging capacity. Mannitol modification improves the targeting of cerium oxide to lung macrophages. This drug delivery platform utilizes a pH-responsive mechanism to ensure stable drug release in an acidic microenvironment. Combined with the dual ROS scavenging effects of mesoporous cerium oxide and polydopamine, it effectively mitigates oxidative stress, improves therapeutic efficacy, and ensures good biocompatibility while reducing drug toxicity, thus providing a more efficient, safe, and controllable treatment option for pulmonary fibrosis.

[0036] This drug delivery platform not only has the ability to remove excess ROS and reduce oxidative stress, but also can deliver drugs to the site of lung inflammation in a targeted manner, which can significantly improve anti-inflammatory and anti-fibrotic effects and provide new ideas and methods for the treatment of pulmonary fibrosis. Attached Figure Description

[0037] Figure 1 The particle size changes of CeO2, P@CeO2 and GMP@CeO2 dispersed in DMEM+FBS 10% solution for 48 h;

[0038] Figure 2 The results of the cytotoxicity evaluation of GMP@CeO2;

[0039] Figure 3 The particle size change of GMP@CeO2 in different dispersions over 48 hours;

[0040] Figure 4 ABTS radical scavenging rates of CeO2 and P@CeO2 at different concentrations;

[0041] Figure 5 The drug release curves for GMP@CeO2 / N at different pH values ​​are shown. Detailed Implementation

[0042] The present invention will be further illustrated below with reference to specific embodiments, but these embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in this technical field. Unless otherwise specified, the reagents and materials used in the following embodiments are commercially available.

[0043] Example 1: Preparation of the GMP@CeO2 Targeted Drug Delivery Platform

[0044] This embodiment provides a targeted drug delivery platform, GMP@CeO2, and the specific preparation method of this delivery platform is as follows:

[0045] Step (1) Preparation of mesoporous CeO2 nanospheres

[0046] Mesoporous CeO2 nanospheres were synthesized via a one-step hydrothermal method. First, 1 g of Ce(NO3)3·6H2O was added sequentially to 10 mL of deionized water and 20 mL of ethylene glycol, followed by sonication to form a homogeneous solution. Then, 1 mL of glacial acetic acid and 0.8 g of PVP were added to the resulting solution, and the mixture was subjected to a hydrothermal reaction at 180 °C in a 50 mL reactor for 20 hours. The product was washed with water and ethanol to remove any possible residual ions and organic solvents. Finally, the product was collected by centrifugation and dried at 60 °C.

[0047] Step (2) Preparation of P@CeO2

[0048] 4 mL of CeO2 (20 mg), 2.5 mL of methanol, and 2.5 mL of dopamine were mixed and transferred to a round-bottom flask. 20 μL of triethylamine was added, and the mixture was refluxed and stirred at 60 °C for 12 hours. The sample was then washed three times with methanol to remove free dopamine, yielding P@CeO2.

[0049] Step (3) Preparation of GMP@CeO2

[0050] Take 5 mL of P@CeO2 (10 mg) and 5 mL of mannose (30 mg), stir at room temperature for 3 h to obtain GMP@CeO2 solution.

[0051] Performance testing:

[0052] Test Example 1-1: Stability Test of GMP@CeO2

[0053] Appropriate amounts of CeO2 obtained in step (1) of Example 1, P@CeO2 obtained in step (2) and GMP@CeO2 obtained in step (3) were dispersed in DMEM+10% FBS complete culture medium, placed in an oven at 37°C, and DLS was detected at 1, 2, 4, 8, 12, 24 and 48 h respectively.

[0054] like Figure 1 As shown, after sequential modification with polydopamine and mannose, the particle size stability of cerium oxide particles in the culture medium was significantly improved. In particular, after chelating mannose in step (3), the particle size of the nanoparticles remained basically consistent after 50 hours in the complete culture medium. This provides the possibility for nanoparticles to circulate in vivo and reach the target site.

[0055] Test Example 1-2 GMP@CeO2 Cytotoxicity Test

[0056] The cytotoxicity of GMP@CeO2 on MLE-12 cells was evaluated using the CCK-8 assay. The specific procedures were as follows: First, MLE-12 cells were seeded at a density of 5000 cells / well in 96-well plates and incubated overnight in a CO2 incubator. Then, the original culture medium was aspirated and replaced with fresh complete culture medium containing different concentrations of GMP@CeO2 (0-800 μg / mL), with five replicates for each concentration. Cells were then incubated for 24 h. After incubation, the cells were washed once with PBS, and 100 μL of fresh culture medium (containing 10% CCK-8) was added to each well. The cells were incubated in an incubator for a period of time. Finally, the absorbance at a wavelength of 450 nm was detected and recorded using an ELISA reader. The cell viability was calculated using the following formula: Cell viability (%) = (Absorbance of experimental group - Absorbance of blank group) / (Absorbance of negative control group - Absorbance of blank group) × 100%.

[0057] like Figure 2 As shown, when the concentration of GMP@CeO2 nanoparticles reached 800 μg / mL, the cell viability of MLE-12 cells was over 95%, indicating that high concentrations of GMP@CeO2 nanoparticles did not affect cell proliferation. This demonstrates that the delivery vector has good biocompatibility.

[0058] Test Examples 1-3: pH-responsive GMP@CeO2

[0059] Appropriate amounts of GMP@CeO2 were dispersed in H2O, PBS (pH 7.4, pH 6.5, pH 5.5), and DMEM+10% FBS complete culture media, respectively, and placed in an oven at 37℃. DLS was detected at 1, 2, 4, 8, 12, 24, and 48 h.

[0060] like Figure 3 As shown, the particle size of nanoparticles changed significantly over time under slightly acidic conditions, indicating that the structural integrity of nanoparticles is disrupted under acidic conditions, which is beneficial for drug release after reaching lung tissue.

[0061] Test Examples 1-4: Antioxidant Capacity of GMP@CeO2

[0062] First, prepare the ABTS stock solution (7.4 mmol / L, 0.4 mL): Take 0.0045 g of ABTS and 1.1025 mL of distilled water (MW = 548.7); prepare the K2S2O8 stock solution (2.6 mmol / L, 1.43 mL): Take 0.0025 g of K2S2O8 and add 3.575 mL of distilled water (MW = 270.32). Mix the two solutions and let them stand in the dark at room temperature for 12 hours. Then dilute the solution 50 times with anhydrous ethanol. For A0 value detection: Take 1.6 mL of the ABTS solution and 0.6 mL of anhydrous ethanol, mix thoroughly, and measure the absorbance at 734 nm. For A value detection: Take 1.6 mL of the ABTS test solution and mix thoroughly with 0.6 mL of CeO2 and P@CeO2 at gradient concentrations, and measure the absorbance at 734 nm. Repeat the experiment independently three times.

[0063] ROS clearance rate = (A0 - A) / A0 × 100%

[0064] like Figure 4 As shown, at the same concentration, P@CeO2 at 100 μg / mL achieved a scavenging rate of over 90% for ABTS free radicals, while CeO2 at 200 μg / mL did not reach this level. This demonstrates that coordination with polydopamine can significantly enhance the antioxidant capacity of cerium oxide nanoparticles. Simultaneously, it can reduce the accumulation of cerium oxide in vivo, improving the biosafety of the nanoparticles.

[0065] In summary, the targeted drug delivery platform of this invention significantly improves particle stability by modifying the cerium oxide surface with mannose and polydopamine. The resulting particles also exhibit good biocompatibility, remaining stable in vivo. Furthermore, the surface coordination modification with polydopamine significantly enhances the antioxidant properties of the cerium oxide nanozyme, while the mannose complexation improves targeting. The delivery platform of this invention can stably deliver drugs to lung tissue via systemic circulation, where their structural integrity disintegrates under acidic conditions, achieving targeted drug release.

[0066] The following experiments, using specific drugs, illustrate the application of this drug delivery platform in the preparation of drugs for pulmonary fibrosis.

[0067] Example 2 Drug Loading Application - Preparation of GMP@CeO2 / N

[0068] Dissolve 15 mg of nintedanib in 2 mL of ultrapure water, and then add it dropwise to the GMP@CeO2 solution obtained in step (3) of Example 1 while stirring. Incubate overnight, centrifuge, and collect the product.

[0069] The loading of nintedanib was detected at 390 nm using a UV spectrophotometer. The encapsulation efficiency (1) and drug loading efficiency (2) were calculated according to the following formulas, where W1 is the actual drug loading, W2 is the total amount of drug added, and W3 is the total amount of carrier and drug.

[0070] EE (%) = W1 / W2 × 100% (1)

[0071] DLC (%) = W1 / W3 × 100% (2)

[0072] The encapsulation efficiency and drug loading rate of nintedanib by GMP@CeO2 were finally calculated to be 66.7% and 18.2%, respectively, which indicates that the nanocarrier has excellent drug loading performance.

[0073] Test Example 2-1: Drug Release Curve of GMP@CeO2 / N

[0074] First, 2 mL of GMP@CeO2 / N solution (1 mg / mL) was added to a dialysis bag and immersed in 20 mL of phosphate-buffered saline (PBS) containing 0.5% Tween 80 (pH = 5.5, 6.5, 7.4). Then, it was transferred to a constant-temperature vibrating screen (37°C, 100 rpm) for incubation. Afterward, 2 mL of PBS solution was taken at different time points and 2 mL of fresh PBS solution was added. The absorbance of the solution taken at 390 nm was measured, and the drug release rate was calculated according to equation (3).

[0075] Drug release (%) = (Cn × V0 + ∑C n-1 ×V1) / m0×100%(3)

[0076] Where Cn is the concentration of the nth sample, V0 is the total volume of the buffer solution, V1 is the volume of each sample, and m0 represents the total amount of drug in the sample.

[0077] like Figure 5As shown, GMP@CeO2 / N is released faster in acidic environments (pH 5.5 and pH 6.5) than in neutral environments, indicating that GMP@CeO2 / N can be well released in pulmonary fibrosis tissue. Furthermore, the drug release rate reached over 60% in acidic environments after 12 hours, demonstrating effective drug release upon reaching the target site, improving bioavailability, and reducing toxicity in other tissues.

[0078] In summary, it can be seen that the drug delivery platform of the present invention can effectively load drugs and stably deliver them to lung tissues in systemic circulation, where they disintegrate in an acidic environment, achieving targeted drug release with a good release rate. The drug delivery platform of the present invention can be used in the preparation of lung therapeutic drugs.

[0079] The technical features of the embodiments described above can be combined arbitrarily. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as the combination of these technical features does not contradict each other, it should be considered within the scope of this specification. It should be noted that those skilled in the art can make several modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention. Therefore, the protection scope of this patent should be determined by the appended claims.

Claims

1. A targeted drug delivery platform based on mesoporous cerium oxide nanoparticles, characterized in that, include: Mesoporous cerium oxide nanoparticles coordinated with polydopamine; Mannose modified on the surface of the mesoporous cerium oxide nanoparticles; The diameter of the mesoporous cerium oxide nanoparticles is 150~200nm, and the mesopore size is 3~6nm; The mesoporous cerium oxide nanoparticles were prepared by dispersing trivalent cerium salt in an aqueous ethylene glycol solution, then adding glacial acetic acid and polyvinylpyrrolidone, and carrying out a hydrothermal reaction at 150-200°C; the raw materials were prepared in the following proportions: Ce 3+ : Glacial acetic acid: PVP = 0.3~0.35g: 0.6~1mL: 0.6~0.8g; The hydrothermal reaction time is 16~24h; The preparation method of the targeted drug delivery platform includes the following steps: (1) Coordinating polydopamine on the surface of mesoporous cerium oxide nanoparticles; (2) The surface of the mesoporous cerium oxide nanoparticles obtained in step (1) is modified with mannose; Step (1) includes: Mesoporous cerium oxide nanoparticles were mixed with methanol and dopamine, and triethylamine was added. The mixture was refluxed at 60°C, and then the free dopamine was removed by washing to obtain cerium oxide nanoparticles with surface-coordinated polydopamine, denoted as P@CeO2. The cerium oxide nanoparticles: methanol: dopamine: triethylamine = 20 mg: 2.5 mL: 20 mg: 20 μL; The reflux reaction time is 10-15 hours; Step (2) includes: P@CeO2 was mixed and dispersed with mannose at room temperature to obtain mannose-modified cerium oxide nanoparticles with surface-coordinated polydopamine, denoted as GMP@CeO2. In the mixed solution, the concentration of P@CeO2 is 1~2 mg / mL, and the concentration of mannose is 3~6 mg / mL; The dispersion time is 0.5~2h.

2. The targeted drug delivery platform according to claim 1, characterized in that, The mesoporous cerium oxide nanoparticles are spherical.

3. The targeted drug delivery platform according to claim 1 or 2, characterized in that, In the ethylene glycol aqueous solution, the volume ratio of ethylene glycol to water is 2:

1.

4. The application of the targeted drug delivery platform as described in any one of claims 1-3 in the preparation of drugs for pulmonary fibrosis.

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