A pharmaceutical composition or drug delivery system, and methods of making and using the same

Abstract

The present application relates to a kind of pharmaceutical composition or drug delivery system and its preparation method and purposes.The pharmaceutical composition or drug delivery system can target lung, effectively improve the lung delivery efficiency of drug, can be used for treating acute lung injury, pulmonary fibrosis, pulmonary tuberculosis, COVID-19 and other diseases.

Description

Technical Field

[0001] This invention relates to a pharmaceutical composition or drug delivery system, its preparation method, and its uses. The pharmaceutical composition or drug delivery system can target the lungs, effectively improving the efficiency of drug delivery to the lungs, and can be used to treat diseases such as acute lung injury, pulmonary fibrosis, tuberculosis, and COVID-19. Background Technology

[0002] Globally, the incidence and mortality rates of acute and chronic lung-related diseases remain high. Besides the efficacy and metabolic properties of the drugs themselves, the efficiency of the drug delivery system is a key factor influencing the effectiveness of drug treatment for lung-related diseases. Traditional drug delivery technologies lack targeting capabilities, have low delivery efficiency, and limited clinical efficacy; furthermore, systemic drug distribution can increase the risk of toxic side effects.

[0003] Targeted drug delivery systems can significantly improve drug delivery efficiency, increase local drug concentration in target organs / tissues, enhance therapeutic efficacy, and reduce systemic drug exposure and toxic side effects. Currently, lung-targeted drug delivery technologies can be mainly divided into two categories. One is lung inhalation drug delivery technology. However, drugs that are unstable in nebulization or easily inactivated by proteases in the airways are not suitable for lung inhalation administration. Furthermore, lung inhalation administration is not suitable for severely ill or critically ill patients who have already developed respiratory distress symptoms and are unable to breathe independently. The other category is mainly lung-targeted drug delivery technology based on molecular targets. However, these molecular targets are mostly expressed not only on lung-related cells but also on the same or other types of cells in other tissues and organs. This will lead to off-target drug delivery, reducing targeted delivery efficiency. Secondly, certain targets or cells expressing those targets (such as macrophages) play an important role in maintaining normal bodily functions; targeting such cells may affect their normal function. Furthermore, the complex interference and barrier effects of endogenous substances in the body often prevent molecular target-based lung-targeted drug delivery technologies from fully recognizing targets in the body. Sometimes, these drugs may even be excreted as exogenous toxins, failing to reach the lung lesions as designed and exert their intended therapeutic effect. Summary of the Invention

[0004] In one aspect, the present invention provides a pharmaceutical composition or pharmaceutical delivery system comprising:

[0005] (1) PLGA nanoparticles loaded with the first drug and red blood cells, or

[0006] (2) The second drug-loaded PLGA nanoparticles have their surface modified with blood group glycoprotein A antibody.

[0007] In some implementations, the red blood cells are derived from the subject's blood.

[0008] In some embodiments, the pharmaceutical composition or drug delivery system is administered to the subject via intravenous injection.

[0009] In some embodiments, the first drug-loaded PLGA nanoparticles bind to and are loaded onto the surface of erythrocytes.

[0010] In some embodiments, the second drug-loaded PLGA nanoparticles bind to and are loaded onto the surface of red blood cells in the subject after intravenous injection.

[0011] In some embodiments, the pharmaceutical composition or drug delivery system is a lung-targeting pharmaceutical composition or drug delivery system.

[0012] In some embodiments, the first drug-loaded PLGA nanoparticle or the second drug-loaded PLGA nanoparticle contains PLGA, and the ratio of lactic acid to glycolic acid is 10:90 to 90:10, preferably 10:90 to 50:50, and more preferably 25:75.

[0013] In some embodiments, the first drug-loaded PLGA nanoparticle or the second drug-loaded PLGA nanoparticle contains PLGA with a molecular weight of 5,000-100,000, preferably 10,000-50,000.

[0014] In some embodiments, the zeta potential of the first drug-loaded PLGA nanoparticle is -50 to +50 mV, preferably -30 to 0 mV, and more preferably -30 to -15 mV.

[0015] In some embodiments, the zeta potential of the second drug-loaded PLGA nanoparticle is -50 to +50 mV, preferably -30 to +30 mV, and more preferably about 0 mV.

[0016] In some embodiments, the particle size of the first drug-loaded PLGA nanoparticles is 5-1000 nm, preferably 100-500 nm, and more preferably 150-400 nm.

[0017] In some embodiments, the second drug-loaded PLGA nanoparticles, before being modified with blood group glycoprotein A antibody, have a particle size of 5-1000 nm, preferably 100-500 nm, and more preferably 150-400 nm.

[0018] In some embodiments, the pharmaceutical composition or drug delivery system further comprises a pharmaceutically acceptable carrier, such as saline solution.

[0019] In some embodiments, the drug loaded on the first drug-loaded PLGA nanoparticle or the second drug-loaded PLGA nanoparticle is a drug that can treat lung-related diseases, such as methylprednisolone, pirfenidone, or gentianin.

[0020] Another aspect of the present invention provides a method for preparing the pharmaceutical composition or drug delivery system of the present invention, comprising:

[0021] 1) Add the drug to the organic solution of PLGA and emulsify it using ultrasound;

[0022] 2) Optionally, the product obtained in 1) is added to a solution of the surfactant (e.g., an aqueous solution) and stirred;

[0023] 3) Remove the organic solvent from the product obtained in 1) or 2), separate and wash to obtain the first drug-loaded PLGA nanoparticles;

[0024] 4) Combine the first drug-loaded PLGA nanoparticles with red blood cells, or co-incubate the first drug-loaded PLGA nanoparticles with blood group glycoprotein A antibody to obtain second drug-loaded PLGA nanoparticles with the surface modified by blood group glycoprotein A antibody.

[0025] In some embodiments, the surfactant solution is selected from: poloxamer (e.g., poloxamer P188), polyvinyl alcohol, cetyltrimethylammonium bromide, and any combination thereof.

[0026] Another aspect of the present invention provides a method for delivering the pharmaceutical composition or pharmaceutical delivery system of the present invention, comprising:

[0027] 1) Collect red blood cells from the subject;

[0028] 2) The first drug-loaded PLGA nanoparticles, resuspended in physiological saline, were co-incubated with erythrocytes to allow the first drug-loaded PLGA nanoparticles to bind to and be loaded onto the surface of the erythrocytes.

[0029] 3) The product obtained in 2) is administered intravenously to the subject, or

[0030] Another aspect of the present invention provides a method for delivering the pharmaceutical composition or pharmaceutical delivery system of the present invention, comprising: directly intravenously delivering the second drug-loaded PLGA nanoparticles to a subject.

[0031] Another aspect of the present invention provides the use of the pharmaceutical composition or drug delivery system described herein in the preparation of medicaments for treating acute lung injury, pulmonary fibrosis, tuberculosis, and COVID-19.

[0032] In some embodiments, the subject of the present invention is a mammal, including bovines, equines, sheep, suidae, canines, felines, rodents, primates, etc., such as humans, cats, dogs, horses, cattle, sheep, or pigs.

[0033] definition

[0034] Unless otherwise defined below, all technical and scientific terms used herein are intended to have the same meaning as commonly understood by one of ordinary skill in the art. References to technical terms herein refer to techniques commonly understood in the art, including variations or equivalent substitutions of techniques that are obvious to one of ordinary skill in the art. While it is believed that the following terms will be well understood by one of ordinary skill in the art, the following definitions are set forth to better explain the invention.

[0035] The terms “including,” “comprising,” “having,” “containing,” or “involving,” as used herein, and their other variations thereof, are inclusive or open-ended and do not exclude other unlisted elements or method steps.

[0036] As used herein, the term “about” means within an acceptable standard error, such as ±0.05, ±0.1, ±0.2, ±0.3, ±1, ±2, or ±3, as would be considered by one of ordinary skill in the art. Unless otherwise apparent from the context, all numerical values ​​provided herein are modified by the term “about”.

[0037] As used herein, the term "pharmaceuticalally acceptable carrier" refers to a diluent, excipient, vehicle, or medium that is administered co-administered with a therapeutic agent and is suitable, to the extent of reasonable medical judgment, for contact with human and / or other animal tissues without excessive toxicity, irritation, allergic reactions, or other problems or complications commensurate with a reasonable benefit / risk ratio.

[0038] Pharmaceutically acceptable carriers that can be used in the pharmaceutical compositions or drug delivery systems described in this invention include, but are not limited to, sterile liquids such as water, physiological saline and glucose solutions, and especially various liquids suitable for injection.

[0039] The dosing regimen can be adjusted to provide the optimal required response. For example, a single bolus injection can be administered, several fractions can be administered over time, or the dose can be proportionally reduced or increased as indicated by the urgency of the treatment situation. It should be noted that dosage values ​​can vary depending on the type and severity of the condition to be alleviated, and may include single or multiple doses. To further understand, for any given individual, the specific dosing regimen should be adjusted over time based on individual needs and the professional judgment of the person administering the composition or supervising its administration.

[0040] The amount of the drug described in this invention administered will depend on the individual being treated, the severity of the condition or illness, the rate of administration, the disposal of the compound, and the prescribing physician's judgment. Generally, the effective dose is from about 0.0001 to about 100 mg per kg of body weight per day, for example, from about 0.01 to about 10 mg / kg / day (single or divided doses). In some cases, dose levels not exceeding the lower limit of the foregoing range may be sufficient, while in other cases, a larger dose may still be used without causing any harmful side effects, provided that the larger dose is first divided into several smaller doses for administration throughout the day.

[0041] Unless otherwise stated, as used herein, the term “treatment” means to reverse, alleviate, or inhibit the progression of a disease or condition or one or more symptoms of such a disease or condition to which such term is applied, or to prevent such a disease or condition or one or more symptoms of such a disease or condition. Attached Figure Description

[0042] Figure 1 This is a transmission electron microscope image of the methylprednisolone-PLGA nanoparticles prepared in Example 1;

[0043] Figure 2 Transmission electron microscopy (TEM) image of the methylprednisolone-PLGA nanoparticles prepared in Example 2;

[0044] Figure 3 This is a transmission electron microscope image of the methylprednisolone-PLGA nanoparticles prepared in Example 3;

[0045] Figure 4 Transmission electron microscopy (TEM) image of the methylprednisolone-PLGA nanoparticles prepared in Example 4;

[0046] Figure 5 Scanning electron microscope images of the methylprednisolone-PLGA nanoparticle-erythrocyte complex prepared in Example 10. The top left, top right, bottom left, and bottom right images are the methylprednisolone-PLGA nanoparticle-erythrocyte complexes prepared in Examples 1, 2, 3, and 4, respectively.

[0047] Figure 6 The adhesion rate of nanoparticles prepared in Examples 1, 2, 3, 4 and 7 to erythrocytes;

[0048] Figure 7 The distribution of drug concentrations in the lungs and livers of rats after intravenous administration of the nanoparticles prepared in Examples 1, 2, 3, 4 and 7, and the methylprednisolone-PLGA nanoparticle-erythrocyte complex prepared in Example 10;

[0049] Figure 8The drug concentration distribution in the lungs and liver of rats after intravenous administration of the phytol-PLGA nanoparticles prepared in Example 6 and the phytol-PLGA nanoparticles modified with blood group glycoprotein A antibody.

[0050] Figure 9 The distribution of fluorescence signals in the lungs and liver of mice after intravenous injection of Cy7.5-NP or RBC-Cy7.5-NP. Detailed Implementation

[0051] The present invention will now be described in conjunction with embodiments and test examples. It should be understood that the following embodiments and test examples are for illustrative purposes only and should not be construed as limiting the scope of the invention. Where specific conditions are not specified in the embodiments and test examples, they are performed under conventional conditions or conditions recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0052] Unless otherwise stated, the raw materials and reagents used in the following examples are commercially available products or can be prepared by known methods.

[0053] Example 1: Preparation of methylprednisolone-PLGA nanoparticles

[0054] 30 mg of PLGA (lactic acid and glycolic acid in a 75:25 ratio, molecular weight 15000) was weighed and dissolved in 1 ml of dichloromethane. 3 mg of methylprednisolone was weighed and added to the PLGA solution to dissolve. The methylprednisolone-PLGA solution was then added dropwise to 10 ml of 1% poloxamer P188 aqueous solution at a rate of 8 mL / h. The mixture was stirred continuously at 800 r / min. After the addition was complete, the mixture was placed in a fume hood and stirred for 6 h to remove the organic solvent. The mixture was centrifuged at 14000 r / min for 20 min and washed three times. The prepared methylprednisolone-PLGA nanoparticles had a particle size of 205.14 nm and a Zeta potential of -25.36 mV. Transmission electron microscopy images are shown below. Figure 1 .

[0055] Example 2: Preparation of methylprednisolone-PLGA nanoparticles

[0056] 30 mg of PLGA (lactic acid and glycolic acid in a 50:50 ratio, molecular weight 15000) was weighed and dissolved in 1 ml of dichloromethane. 3 mg of methylprednisolone was weighed and added to the PLGA solution to dissolve. The methylprednisolone-PLGA solution was then added dropwise to 10 ml of 1% poloxamer P188 aqueous solution at a rate of 8 mL / h. The mixture was stirred continuously at 800 r / min. After the addition was complete, the mixture was placed in a fume hood and stirred for 6 h to remove the organic solvent. The mixture was centrifuged at 14000 r / min for 20 min and washed three times. The prepared methylprednisolone-PLGA nanoparticles had a particle size of 201.39 nm and a Zeta potential of -20.65 mV. Transmission electron microscopy images are shown below. Figure 2 .

[0057] Example 3: Preparation of methylprednisolone-PLGA nanoparticles

[0058] 30 mg of PLGA (lactic acid and glycolic acid in a 25:75 ratio, molecular weight 15000) was weighed and dissolved in 1 ml of dichloromethane. 3 mg of methylprednisolone was weighed and added to the PLGA solution to dissolve. The methylprednisolone-PLGA solution was then added dropwise to 10 ml of 1% poloxamer P188 aqueous solution at a rate of 8 mL / h. The mixture was stirred continuously at 800 r / min. After the addition was complete, the mixture was placed in a fume hood and stirred for 6 h to remove the organic solvent. The mixture was centrifuged at 14000 r / min for 20 min and washed three times. The prepared methylprednisolone-PLGA nanoparticles had a particle size of 203.57 nm and a Zeta potential of -17.28 mV. Transmission electron microscopy images are shown below. Figure 3 .

[0059] Example 4: Preparation of methylprednisolone-PLGA nanoparticles

[0060] 30 mg of PLGA (lactic acid and glycolic acid in a 10:90 ratio, molecular weight 15000) was weighed and dissolved in 1 ml of dichloromethane. 3 mg of methylprednisolone was weighed and added to the PLGA solution to dissolve. The methylprednisolone-PLGA solution was then added dropwise to 10 ml of 1% poloxamer P188 aqueous solution at a rate of 8 mL / h. The mixture was stirred continuously at 800 r / min. After the addition was complete, the mixture was placed in a fume hood and stirred for 6 h to remove the organic solvent. The mixture was centrifuged at 14000 r / min for 20 min and washed three times. The prepared methylprednisolone-PLGA nanoparticles had a particle size of 198.56 nm and a Zeta potential of -16.34 mV. Transmission electron microscopy images are shown below. Figure 4 .

[0061] Example 5: Preparation of pirfenidone-PLGA nanoparticles

[0062] Weigh 10 mg of pirfenidone and dissolve it in 0.5 mL of ultrapure water. Add this solution dropwise to 10 mL of a 10 mg / mL PLGA (lactic acid and glycolic acid ratio of 25:75, molecular weight 30000) solution in dichloromethane, and sonicate for 30 seconds. Then, add the resulting emulsion to 10 mL of a 5% PVA (polyvinyl alcohol) aqueous solution, sonicate for 30 seconds, and continue stirring for 4 hours to evaporate the organic solvent. Collect the nanoparticles by centrifugation at 14000 rpm for 20 minutes.

[0063] Example 6: Preparation of Bryophyllum-PLGA Nanoparticles

[0064] Weigh 20 mg of PLGA (lactic acid and glycolic acid ratio of 25:75, molecular weight 10000) and 4 mg of terpinene and dissolve them in 4 ml of acetone. Add the terpinene-PLGA solution dropwise to 20 ml of an aqueous solution containing 1% poloxamer P188 while stirring at 800 r / min. After the addition is complete, place the solution in a fume hood and continue stirring for 6 h to remove the organic solvent. Centrifuge at 14000 r / min for 20 min and wash three times to obtain the final terpinene-PLGA nanoparticles. The prepared terpinene-PLGA nanoparticles have a particle size of 165.34 nm and a Zeta potential of -26.37 mV.

[0065] Weigh 20 mg of PLGA (lactic acid and glycolic acid ratio of 25:75, molecular weight 10000) and 4 mg of *Tetrapanax papyriferus* and dissolve them in 4 ml of acetone. Add the *Tetrapanax papyriferus*-PLGA solution dropwise to 20 ml of an aqueous solution containing 1% poloxamer P188 and 0.005% CTAB (cetyltrimethylammonium bromide) while stirring at 800 r / min. After the addition is complete, place the solution in a fume hood and continue stirring for 6 h to remove the organic solvent. Centrifuge at 14000 r / min for 20 min and wash three times to obtain the final product. The prepared *Tetrapanax papyriferus*-PLGA nanoparticles have a particle size of 169.12 nm and a Zeta potential of +21.61 mV.

[0066] Weigh 20 mg of PLGA (lactic acid and glycolic acid ratio of 25:75, molecular weight 10000) and 4 mg of tebufenozide and dissolve in 4 ml of acetone. Add the tebufenozide-PLGA solution dropwise to 20 ml of an aqueous solution containing 1% poloxamer P188 and 0.0027% CTAB. Stir continuously while adding, at a stirring speed of 800 r / min. After the addition is complete, place in a fume hood and continue stirring for 6 h to remove the organic solvent. Centrifuge at 14000 r / min for 20 min and wash three times to obtain the final product. The prepared tebufenozide-PLGA nanoparticles have a particle size of 160.89 nm and a Zeta potential of -0.0057 mV.

[0067] Three different charged nanoparticles (0.01 mol each) were resuspended in PBS buffer, and 0.1 mol of carbodiimide (EDC) and 0.3 mol of N-hydroxysuccinimide (NHS) were added respectively. The mixtures were incubated at 4°C with shaking for 30 minutes. After centrifugation and washing, the nanoparticles were resuspended in PBS buffer, and 0.01 mol of GPAA (purchased from Thermo Fisher, 14-5921-82) was added. The mixtures were incubated at 4°C with shaking overnight. After centrifugation and washing, the GPAA-PLGA nanoparticles modified with GPAA were obtained.

[0068] Example 7: Preparation of Methylprednisolone-PLGA Nanoparticles Modified with Blood Group Glycoprotein A Antibody

[0069] A suitable amount (0.01 mol) of the methylprednisolone-PLGA nanoparticles prepared in Example 3 was resuspended in PBS buffer, and 0.1 mol of carbodiimide (EDC) and 0.3 mol of N-hydroxysuccinimide (NHS) were added. The mixture was incubated at 4°C with shaking for 30 minutes. After centrifugation and washing, the nanoparticles were resuspended in PBS buffer, and 0.01 mol of glycoprotein A antibody (GPAA) was added. The mixture was incubated at 4°C with shaking overnight. After centrifugation and washing, methylprednisolone-PLGA nanoparticles modified with glycoprotein A antibody (GPAA-methylprednisolone-PLGA) were obtained.

[0070] Example 8: Preparation of pirfenidone-PLGA nanoparticles modified with blood group glycoprotein A antibody

[0071] A suitable amount (0.01 mol) of the pirfenidone-PLGA nanoparticles prepared in Example 5 was resuspended in PBS buffer, and 0.1 mol of carbodiimide (EDC) and 0.3 mol of N-hydroxysuccinimide (NHS) were added. The mixture was incubated at 4°C with shaking for 30 minutes. After centrifugation and washing, the nanoparticles were resuspended in PBS buffer, and 0.01 mol of blood group glycoprotein A antibody (GPAA) was added. The mixture was incubated at 4°C with shaking overnight. After centrifugation and washing, pirfenidone-PLGA nanoparticles modified with blood group glycoprotein A antibody were obtained.

[0072] Example 9: Preparation of hematocrit cells

[0073] Whole blood was collected in a heparin sodium vacuum tube and centrifuged at 1300×g to remove the supernatant and intermediate white blood cells, platelets, etc. The lower red blood cells were gently mixed with 4 times the volume of sterile physiological saline solution, centrifuged at 850×g for 5 min three times, and the supernatant was removed to obtain the washed hematocrit red blood cells, which were then placed in a refrigerator at 4 ℃ for later use.

[0074] Example 10: Preparation of Methylprednisolone-PLGA Nanoparticle-Red Blood Cell Complex

[0075] Take 200 µL of the hematocrit red blood cells prepared in Example 9, add a methylprednisolone-PLGA nanoparticle solution (equivalent to 800 µg of PLGA) prepared in Example 1, 2, 3, or 4, resuspended in sterile physiological saline, and incubate at 37°C for 30 min. Centrifuge at 100 g for 5 min, remove the supernatant, and wash three times with sterile physiological saline. The resulting complex is shown in the scanning electron microscope image below. Figure 5 .

[0076] Example 11: Preparation of fluorescent dye cyt7.5-PLGA nanoparticles

[0077] Weigh 30 mg of PLGA (lactic acid and glycolic acid in a 75:25 ratio, molecular weight 15000) and 1 mg of fluorescent dye cyt7.5 and dissolve them in 1 ml of acetone. Add the above solution dropwise to 10 ml of 1% poloxamer P188 aqueous solution at a rate of 10 mL / h while stirring at 800 r / min. After the addition is complete, place the solution in a fume hood and continue stirring for 6 h to remove the organic solvent. Centrifuge at 14000 r / min for 20 min and wash three times to obtain the final product.

[0078] Experimental Example 1: Adhesion rate of nanoparticles to red blood cells

[0079] Test method:

[0080] Take 200 µL of hematocrit red blood cells prepared in Example 9 and add them to the GPAA-methylprednisolone-PLGA nanoparticle solution prepared in Example 7 (equivalent to 800 µg of PLGA), resuspended in sterile physiological saline. Incubate at 37°C for 30 min. Centrifuge at 100 g for 5 min, and take 200 μL of the suspension before centrifugation and 200 μL of the suspension after centrifugation and washing, and add 800 μL of acetonitrile to each. Sonicate for 1 min and vortex mix for 3 min. Determine the methylprednisolone content in the above samples by HPLC. The methylprednisolone content in the sample before centrifugation is recorded as W. 总药量 The methylprednisolone content in the sample after centrifugation and washing is denoted as W. RBC According to the adhesion rate of red blood cells = W RBC / W 总药量 *100% calculation of the adhesion rate between nanoparticles and red blood cells.

[0081] In another example, the methylprednisolone-PLGA nanoparticles from Example 10 were incubated with red blood cells at 37°C before centrifugation and after centrifugation and washing. The adhesion rate between the nanoparticles and red blood cells was determined using the same method.

[0082] Experimental results:

[0083] The adhesion rates of the nanoparticles prepared in Examples 1, 2, 3, 4, and 7 to erythrocytes are as follows: Figure 6As shown, the adhesion rate of nanoparticles prepared with PLGA at a lactic acid to glycolic acid ratio of 75:25 (Example 1) to erythrocytes was significantly higher than that of nanoparticles prepared with PLGA at ratios of 50:50 (Example 2), 25:75 (Example 3), and 10:90 (Example 4) to erythrocytes. Figure 5 Scanning electron microscopy (SEM) images of the nanoparticle-erythrocyte complex show that the nanoparticles prepared with PLGA in a lactic acid to glycolic acid ratio of 75:25 have a stronger binding force with erythrocytes, thus exhibiting a higher erythrocyte adhesion rate. The SEM images also show that the nanoparticles prepared with PLGA in a lactic acid to glycolic acid ratio of 75:25 (Example 1) adhere to erythrocytes in significantly higher quantities than those prepared with PLGA in lactic acid to glycolic acid ratios of 50:50 (Example 2), 25:75 (Example 3), and 10:90 (Example 4).

[0084] The methylprednisolone-PLGA nanoparticles modified with blood group glycoprotein A antibody (GPAA-methylprednisolone-PLGA) prepared in Example 7 had an adhesion rate of 88.27% to erythrocytes, significantly higher than that in Example 3. The binding of GPAA to blood group glycoprotein A on the surface of erythrocytes enhanced the binding efficiency of GPAA-methylprednisolone-PLGA to erythrocytes.

[0085] Experimental Example 2: Lung Targeting Efficiency of Nanoparticle-Red Blood Cell Composition

[0086] Experimental Methods: Twenty-seven male SD rats were randomly divided into 9 groups. Methylprednisolone-PLGA nanoparticle solutions prepared in Examples 1, 2, 3, and 4 (group names: NP1, NP2, NP3, NP4); GPAA-methylprednisolone-PLGA nanoparticles prepared in Example 7 (group name: NP7); and complexes formed by methylprednisolone-PLGA nanoparticles and erythrocytes prepared in Examples 1, 2, 3, and 4 (Example 10, group names: RBC-NP1, RBC-NP2, RBC-NP3, RBC-NP4) were administered via tail vein injection. The dosage was 750 μg / rat (based on the active ingredient methylprednisolone). Rats were sacrificed 2 hours after administration, and lung and liver samples were collected, weighed, and the concentration of methylprednisolone in the tissues was determined.

[0087] The test results are shown in Figure 7 The results show the drug concentration distribution in the lungs and liver of rats after intravenous administration of the nanoparticles prepared in Examples 1, 2, 3, 4 and 7, as well as the methylprednisolone-PLGA nanoparticle-erythrocyte complex prepared in Example 10.

[0088] On the one hand, nanoparticle-erythrocyte complexes prepared with PLGA in different lactate-glycolic acid ratios exhibit different lung-targeting properties. RBC-NP3 (a nanoparticle-erythrocyte complex prepared with PLGA in a lactate-glycolic acid ratio of 25:75) shows better lung-targeting drug delivery, with significantly higher drug concentrations in the lungs 2 hours after administration compared to RBC-NP1 (a nanoparticle-erythrocyte complex prepared with PLGA in a lactate-glycolic acid ratio of 75:25), RBC-NP2 (a nanoparticle-erythrocyte complex prepared with PLGA in a lactate-glycolic acid ratio of 50:50), and RBC-NP4 (a nanoparticle-erythrocyte complex prepared with PLGA in a lactate-glycolic acid ratio of 10:90). Furthermore, RBC-NP3 can significantly reduce drug distribution in the liver. In contrast, RBC-NP4 shows poorer lung-targeting efficacy, and its drug distribution in the liver is not significantly different from that of nanoparticles alone. Therefore, the nanoparticle-erythrocyte complex prepared by PLGA with a lactic acid to glycolic acid ratio of 25:75 has excellent lung-targeting drug delivery capability, and its targeting performance is significantly better than that of nanoparticle-erythrocyte complex prepared by PLGA with other lactic acid to glycolic acid ratios.

[0089] On the other hand, the methylprednisolone-PLGA nanoparticles modified with blood group glycoprotein A antibody prepared in Example 7, after intravenous injection, exhibited lung-targeting performance comparable to RBC-NP3. Although the GPAA-methylprednisolone-PLGA nanoparticles were not incubated with erythrocytes in vitro to prepare nanoparticle-erythrocyte complexes, they still possessed excellent lung-targeting ability, with significantly higher drug concentrations in the lungs after administration compared to unmodified NP3 nanoparticles. GPAA-methylprednisolone-PLGA nanoparticles can actively target erythrocytes via GPAA, binding to blood group glycoprotein A on the surface of erythrocytes to complete nanoparticle-erythrocyte complexation in vivo, exhibiting lung-targeting performance similar to in vitro prepared nanoparticle-erythrocyte complexes. Compared to the method of in vitro preparation and reinfusion of nanoparticle-erythrocyte complexes, this method is simpler and more convenient, reducing the processes of blood cell collection and incubation, avoiding the potential risk of blood cell contamination, and reducing damage to erythrocytes caused by the in vitro preparation process.

[0090] Experimental Example 3: Lung Targeting Efficiency of Nanoparticle-Red Blood Cell Complex

[0091] Experimental Methods: Eighteen male SD rats were randomly divided into 6 groups. The rats were administered either the styrax-PLGA nanoparticle solution prepared in Example 6 (groups named NP+, NP-, and NP0 according to their charges) or the styrax-PLGA nanoparticle solution modified with blood group glycoprotein A antibody (groups named GPAA-NP+, GPAA-NP-, and GPAA-NP0 according to their charges) via tail vein injection at a dose of 900 μg / rat. Two hours after administration, the rats were sacrificed, and lung and liver samples were collected, weighed, and the concentration of styrax-PLGA nanoparticles in the tissues was determined.

[0092] The test results are shown in Figure 8 The results show the drug concentrations in the lungs and livers of rats after intravenous administration of the *Tetrandrine-PLGA* nanoparticles prepared in Example 6 and the *Tetrandrine-PLGA* nanoparticles modified with blood group glycoprotein A antibody. The results showed that the unmodified nanoparticles were mainly taken up by the liver after intravenous injection, resulting in a lower drug concentration in the lungs. However, the distribution of uncharged neutral nanoparticles (NP0) in the liver was lower than that of positively or negatively charged nanoparticles. Intravenous injection of nanoparticles modified with blood group glycoprotein A antibody significantly improved lung targeting performance, with significantly higher local drug concentrations in the lungs compared to the unmodified nanoparticles. Furthermore, the drug concentration in the lungs of the neutral nanoparticles modified with blood group glycoprotein A antibody was significantly higher than that of the charged nanoparticles modified with blood group glycoprotein A antibody; while the drug concentration in the liver was significantly lower than that of the charged nanoparticles modified with blood group glycoprotein A antibody.

[0093] Experimental Example 4: Lung Targeting of Nanoparticle-Red Blood Cell Composition

[0094] Experimental Method: The fluorescent dye cyt7.5-PLGA nanoparticles (cyt7.5-NP) prepared in Example 11 were resuspended in sterile physiological saline. This was added to 200 µL of the hematocrit erythrocytes prepared in Example 9 and incubated at 37°C for 30 min. After centrifugation at 100 g for 5 min, the supernatant was removed, and the mixture was washed three times with sterile physiological saline to obtain the cyt7.5-PLGA nanoparticle-erythrocyte complex (RBC-cyt7.5-NP).

[0095] Eighteen ICR mice (30±2g) were randomly divided into three groups and intravenously injected with PBS (negative control), Cy7.5-NP, and RBC-Cy7.5-NP, respectively. Mice were sacrificed 0.5h and 2h after administration, and the heart, liver, spleen, lung, and kidney were harvested for imaging analysis (excitation wavelength 745nm; emission wavelength 800nm) to determine the fluorescence distribution intensity of each organ.

[0096] Test results: such as Figure 9As shown, after intravenous injection of Cy7.5-NP, the nanoparticles mainly accumulated in the liver, with no obvious fluorescent signal observed in the lungs. After intravenous injection of RBC-Cy7.5-NP, the liver remained the primary distribution organ for the nanoparticles, but significant fluorescent signals appeared in the lungs. This indicates that the nanoparticle-erythrocyte composition possesses lung-targeted drug delivery capabilities and can significantly promote local drug accumulation in the lungs.

Claims

1. A pharmaceutical composition or drug delivery system comprising first drug-loaded PLGA nanoparticles and red blood cells, and a pharmaceutically acceptable carrier, wherein the first drug-loaded PLGA nanoparticles contain PLGA in a lactic acid to glycolic acid ratio of 25:

75.

2. The pharmaceutical composition or drug delivery system of claim 1, wherein the red blood cells are derived from the blood of the subject.

3. The pharmaceutical composition or drug delivery system of claim 1 or 2, administered to a subject by intravenous injection.

4. The pharmaceutical composition or drug delivery system according to claim 1 or 2, wherein... The first drug-loaded PLGA nanoparticles bind to erythrocytes and are loaded onto the surface of erythrocytes.

5. The pharmaceutical composition or drug delivery system according to claim 1 or 2, wherein it is a pharmaceutical composition or drug delivery system targeting the lungs.

6. The pharmaceutical composition or drug delivery system according to claim 1 or 2, characterized by one or more of the following features: i) The first drug-loaded PLGA nanoparticles contain PLGA with a molecular weight of 5,000-100,000; ii) The zeta potential of the first drug-loaded PLGA nanoparticle is -50 ~ +50 mV; iii) The particle size of the first drug-loaded PLGA nanoparticles is 5~1000 nm; iv) The pharmaceutically acceptable carrier is physiological saline.

7. The pharmaceutical composition or drug delivery system of claim 6, characterized by one or both of the following features: i) The first drug-loaded PLGA nanoparticles contain PLGA with a molecular weight of 10,000-50,000; ii) The zeta potential of the first drug-loaded PLGA nanoparticle is -30 ~ 0mV.

8. The pharmaceutical composition or drug delivery system of claim 7, characterized by one or both of the following features: i) The zeta potential of the first drug-loaded PLGA nanoparticles is -30 ~ -15mV; ii) The particle size of the first drug-loaded PLGA nanoparticles is 150~400 nm.

9. The pharmaceutical composition or pharmaceutical delivery system of claim 1 or 2, wherein the pharmaceutical product is a medicament for treating lung-related diseases.

10. The pharmaceutical composition or drug delivery system of claim 9, wherein the drug is methylprednisolone, pirfenidone, or senna.

11. A method for preparing the pharmaceutical composition or drug delivery system according to any one of claims 1-10, comprising: 1) Add the drug to the organic solution of PLGA and emulsify it using ultrasound; 2) Remove the organic solvent from the product obtained in the previous step, separate and wash to obtain the first drug-loaded PLGA nanoparticles.

12. The method of claim 11, further comprising: Before step 2), add the product obtained in step 1) to the surfactant solution and stir.

13. The method of claim 12, wherein the solution of the surfactant is an aqueous solution of the surfactant.

14. The method of claim 12, wherein the surfactant solution is selected from: poloxamer, polyvinyl alcohol, hexadecyltrimethylammonium bromide, and any combination thereof.

15. The method of claim 14, wherein the poloxamer is poloxamer P188.

16. Use of the pharmaceutical composition or drug delivery system according to any one of claims 1-7 in the preparation of a medicament for treating acute lung injury, pulmonary fibrosis, pulmonary tuberculosis or COVID-19.

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