Medicine, preparation method thereof, pharmaceutical composition and application

Abstract

The invention discloses a medicine, a preparation method thereof, a pharmaceutical composition and application, and relates to the technical field of pharmacy. The medicine for stopping bleeding is obtained by grafting hemostatic polypeptide VBP and polyethylene glycol-polycaprolactone, and then the medicine is loaded with quercetin to obtain the medicine composition for treating cerebral hemorrhage. Wherein the VBP is combined with hemophilia necrosis factors with exposed vascular matrix layers after cerebral hemorrhage, and the quercetin is used for breaking the blood brain barrier. The medicine provided by the invention is simple in reaction and easy to prepare, and meanwhile, the main component of the polypeptide is amino acid, so that other substances possibly harmful to the body cannot be generated. Experimental results show that the medicine and the medicine composition provided by the invention are good in biocompatibility, and can promote expression of anti-inflammatory genes and anti-inflammatory factors and inhibit expression of pro-inflammatory genes and pro-inflammatory factors; the targeting property is good and the treatment effect is obvious when the compound is used for treating mouse cerebral hemorrhage. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and in particular to a medicine, a preparation method thereof, a medicine composition and application thereof. Background Art

[0002] Intracerebral hemorrhage (ICH) is a disease in which blood flows out of the brain due to rupture of blood vessels. When cerebral blood vessels rupture, blood will overflow into the brain tissue, causing damage to the brain tissue and dysfunction. Intracerebral hemorrhage is a devastating neurological disease with extremely high mortality and disability rates. Intracerebral hemorrhage can directly cause damage to brain tissue, leading to varying degrees of neurological function loss, affecting the patient's ability to carry out daily activities, and in severe cases, it can be life-threatening.

[0003] Surgical treatment can relieve hematoma pressure and improve cerebral blood perfusion, but the risk of surgical treatment is relatively high, and the recovery of various functional indicators of patients with cerebral hemorrhage after treatment is not ideal. Traditional hemostatic drugs such as tranexamic acid and thrombin are systemic hemostatic drugs, which have no significant hemostatic effect on cerebral hemorrhage and increase the risk of thrombosis. Therefore, it is urgent to develop drugs that can target cerebral hemorrhage and promote hemostasis. Summary of the invention

[0004] In view of this, the present invention provides a drug, a preparation method, a pharmaceutical composition and application thereof. The present invention obtains a drug and a pharmaceutical composition for hemostasis of cerebral hemorrhage by grafting polyethylene glycol-polycaprolactone (mPEG-PCL) with a hemostatic polypeptide (VBP) and physically loading it with quercetin. The preparation process is simple, the drug can be targeted and delivered to the bleeding site of cerebral hemorrhage, and the drug has a good therapeutic effect.

[0005] The present invention first provides a drug having a structure of formula (I):

[0006]

[0007] wherein n is an integer of 3000 to 7000, preferably an integer of 4000 to 6000, more preferably an integer of 4500 to 5500; m is an integer of 3000 to 7000, preferably an integer of 4000 to 6000, more preferably an integer of 4500 to 5500;

[0008] R is the residue after removing any amino group from the polypeptide VBP.

[0009] The inventors of the present invention have found that the essence of cerebral hemorrhage is the rupture of the arterioles that feed the brain parenchyma. After the rupture of the blood vessels, the vascular endothelial cells are broken, exposing the vascular matrix layer containing hemophilic necrosis factor (vWF) and collagen, which provides sites for platelet adhesion and aggregation, and also provides conditions for the development of targeted nano hemostatic drugs. The vWF and collagen on the vascular matrix layer are used as potential targets for cerebral hemorrhage hemostasis, and the excellent ability of the polypeptide VBP (TRYLRIHPQSQVHQI) to accurately bind vWF without causing extensive intravascular coagulation is used to reduce the volume of hematoma after cerebral hemorrhage, prevent secondary bleeding, and transform the size of hematoma and secondary expansion of hematoma from a life-threatening topic to an advantage that can be selectively treated. mPEG-PCL nanoparticles have become an important tool in the field of medical drug delivery due to their excellent biocompatibility, adjustable drug release characteristics, targeted delivery ability and multifunctionality. It has great application potential in cancer treatment, chronic disease management and other drug delivery systems. By regulating the length and structure of the polymer chain, even zero-order release (constant release) or controlled release can be achieved, which is especially important for diseases that require long-term, stable treatment, such as cancer and chronic diseases. PEG-PCL modified by VBP can exhibit excellent hemostatic properties (VBP-PEG-PCL), and the diblock polymer can also carry fat-soluble drugs through hydrophilic and hydrophobic interactions, improve the hydrophilicity of small molecule drugs, and pass through the blood-brain barrier smoothly.

[0010] Therefore, the present invention selects to graft the hemostatic polypeptide VBP with mPEG-PCL for the treatment of cerebral hemorrhage. After cerebral hemorrhage, the vWF exposed in the vascular matrix layer provides a binding site for the polypeptide VBP. The main component of the polypeptide is amino acid, and no other substances that may be harmful to the body are produced. The click chemistry reaction process is simple, and the water-soluble small molecule toxic products can be removed by dialysis after grafting to mPEG-PCL.

[0011] In some specific implementations of the present invention, the drug having the structure of formula (I) is preferably the following structure:

[0012]

[0013] Here, n is an integer from 3000 to 7000, and m is an integer from 3000 to 7000.

[0014] The present invention provides a pharmaceutical composition, comprising: the above-mentioned drug and quercetin or a quercetin derivative. In order to improve the secondary damage caused by cerebral hemorrhage, the present invention also uses a physical loading method to encapsulate quercetin in the hydrophobic segment of the above-mentioned drug. Quercetin is a widely existing natural flavonoid with multiple pharmacological effects. Its excellent antioxidant stress resistance and fat solubility can penetrate the blood-brain barrier and inhibit the release of ROS. In the present invention, the pharmaceutical composition can be prepared according to Figure 1 The synthesis route shown is: alkynyl-polyethylene glycol-polycaprolactone (ALK-PEG-PCL) is combined with the polypeptide VBP through a click reaction of the alkynyl and azide groups, and then quercetin is physically loaded to obtain the pharmaceutical composition of the present invention.

[0015] In some specific implementations of the present invention, the mass ratio of quercetin or quercetin derivatives to the drug in the pharmaceutical composition is 1: (10-20), preferably 1: (10-18), and more preferably 1: (10-15). In some specific implementations, the particle size of the pharmaceutical composition is 50-80 nm, preferably 55-80 nm, and more preferably 60-80 nm. In some specific implementations, the encapsulation efficiency of quercetin or quercetin derivatives in the pharmaceutical composition is 80%-95%; the drug loading is 5%-10%.

[0016] The present invention also provides a method for preparing the above-mentioned drug, comprising: subjecting the azide-modified polypeptide VBP to a click reaction with the alkynyl-polycaprolactone-polyethylene glycol shown in formula (II) to obtain the drug shown in formula (I);

[0017]

[0018] Wherein, n is an integer from 3000 to 7000, and m is an integer from 3000 to 7000;

[0019] R is the residue after removing any amino group from the polypeptide VBP.

[0020] Compared with traditional organic synthesis methods, the click reaction of azide and alkyne groups usually does not require the use of toxic or metal catalysts and has higher biocompatibility; at the same time, the reaction conditions are mild and can be carried out at a lower temperature, which reduces the difficulty of operation and helps to reduce the generation of by-products. Therefore, the present invention selects the polypeptide VBP modified with an azide group to undergo a click reaction with alkyne-polycaprolactone-polyethylene glycol to obtain the drug of the present invention.

[0021] The present invention has no limitation on the source of the azide-modified polypeptide VBP, and those skilled in the art can synthesize or purchase it by themselves. For example, an amino acid residue containing an azide group can be introduced at a specific position during polypeptide synthesis; or an azide group can be connected to a synthesized polypeptide chain by chemical modification.

[0022] In some specific implementations of the present invention, the azide-modified polypeptide VBP has the following structure, which cannot cover all azide-modified polypeptide VBPs described in the present invention;

[0023]

[0024] In some specific implementations, the molar ratio of the azide-modified polypeptide VBP to the alkynyl-polycaprolactone-polyethylene glycol for click reaction is 1:(800-1200), preferably 1:(900-1100), and more preferably 1:(950-1050). In some specific implementations, the click reaction time is 10-20 hours, preferably 10-18 hours, and more preferably 10-16 hours; the temperature is 20-50°C, preferably 25-40°C, and more preferably 30-40°C.

[0025] Based on the advantages of the drug of the present invention and the ability of quercetin or quercetin derivatives to break the blood-brain barrier, the present invention combines the two to obtain the above-mentioned pharmaceutical composition, and provides a method for preparing the pharmaceutical composition. The preparation method provided by the present invention cannot cover all situations, and those skilled in the art can choose according to their needs. The protection scope of the present invention is not affected by this preparation method.

[0026] The above-mentioned drugs and quercetin or quercetin derivatives are dissolved in a solvent and stirred vigorously, ultrapure water is added during the stirring process, and then dialyzed using a dialysis bag with a molecular weight cutoff of 3000-7000Da to obtain the pharmaceutical composition.

[0027] The drug provided by the present invention can accurately bind to vWF, but will not cause extensive intravascular coagulation; at the same time, the pharmaceutical composition can break the blood-brain barrier and smoothly transport the drug to the brain, so the present invention applies the above-mentioned drug and pharmaceutical composition to the field of treating cerebral hemorrhage. The present invention has no limitation on the specific mode of application, and those skilled in the art can choose according to their needs.

[0028] In summary, the present invention combines the polypeptide VBP with mPEG-PCL nanoparticles by means of a click reaction, and utilizes the specific binding of the polypeptide VBP with vWF in the vascular matrix layer to achieve hemostasis. At the same time, mPEG-PCL as a carrier has excellent biocompatibility, adjustable drug release characteristics, targeted delivery capabilities and multifunctionality. On this basis, the present invention combines VBP-modified mPEG-PCL with quercetin or quercetin derivatives by means of physical loading, which can effectively break the blood-brain barrier and be applied to the treatment of cerebral hemorrhage. The experimental results show that the drugs and pharmaceutical compositions provided by the present invention have good biocompatibility, can inhibit the expression of anti-inflammatory genes and anti-inflammatory factors, and promote the expression of pro-inflammatory genes and pro-inflammatory factors; they have good targeting when used for the treatment of mouse cerebral hemorrhage, and the therapeutic effect is significant. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 Schematic diagram of the preparation of Qu-VBP-PEG-PCL;

[0030] Figure 2 This is the characterization result diagram of VBP-PEG-PCL;

[0031] Figure 3 This is the characterization result diagram of Q u-VBP-PEG-PCL;

[0032] Figure 4 The data graph is a cytotoxicity result graph of different concentrations of Qu-VBP-PEG-PCL;

[0033] Figure 5 This is a data graph of phalloidin staining and live-dead staining results;

[0034] Figure 6 Data graphs showing the results of reactive oxygen species staining and EDU-488 staining;

[0035] Figure 7 This is the data diagram of the gene expression results detected by RT-qPCR and Elisa;

[0036] Figure 8 This is a data graph of the in vivo experimental results of cerebral hemorrhage in mice;

[0037] Fig. 9 This is a data graph showing the results of an in vitro experiment on cerebral hemorrhage in mice. DETAILED DESCRIPTION

[0038] It should be understood that the expression "one or more of..." includes each of the items recited after the expression individually and in various different combinations of two or more of the recited items, unless otherwise understood from the context and usage. The expression "and / or" in combination with three or more recited items should be understood to have the same meaning, unless otherwise understood from the context.

[0039] The use of the terms "comprising", "having" or "containing", including their grammatical synonyms, should generally be understood as open and non-restrictive, for example not excluding other unrecited elements or steps, unless otherwise specifically stated or otherwise understood from the context.

[0040] It should be understood that the order of steps or the order in which certain actions are performed is not important as long as the present invention remains operable. In addition, two or more steps or actions may be performed simultaneously.

[0041] The use of any and all examples or exemplary language, such as "for example" or "including", herein is intended only to better illustrate the invention and does not limit the scope of the invention unless otherwise claimed. No language in this specification should be construed as indicating that any non-claimed element is essential to the practice of the invention.

[0042] In addition, the numerical ranges and parameters used to define the present invention are approximate values, and the relevant values ​​in the specific embodiments have been presented as accurately as possible. However, any numerical value inherently inevitably contains standard deviations due to individual test methods. Therefore, unless otherwise expressly stated, it should be understood that all ranges, quantities, values ​​and percentages used in this disclosure are modified by "about". Here, "about" generally means that the actual value is within plus or minus 10%, 5%, 1% or 0.5% of a specific value or range.

[0043] The present invention is further described below in conjunction with the following examples. The protection scope of the present invention is not limited by the following examples.

[0044] Example 1

[0045] To a 50 mL glass round-bottom flask, 20 mg (0.01 mmol) of VBP-N3, 2 mL of dimethylformamide (DMF), and 102.09 mg (10.21 mmol) of alkynyl-PEG were added in sequence. 5000 -PCL 5000 (ALK-PEG-PCL, purchased from Changchun Institute of Applied Chemistry, Chinese Academy of Sciences), stirred at 37°C for 12 hours for reaction. After the reaction was completed, dialyzed for three days using a dialysis bag with a molecular weight cutoff of 5000Da, with water replaced every 6 hours, and finally freeze-dried to obtain the product VBP-PEG-PCL.

[0046] The elemental content and functional groups of the product VBP-PEG-PCL were analyzed by elemental analysis, gel permeation chromatography (GPC) and Fourier transform infrared spectroscopy (FI-TR). Figure 2 Show. Figure 2 middle, Figure 2A is the FI-TR analysis result, where 1670 is the C=O characteristic absorption peak of amide; Figure 2 B is the elemental analysis result, where the nitrogen content in VBP-PEG-PCL is 3.19%; Figure 2 C and Figure 2 D is the result of gel chromatography analysis. The molecular weight of the product VBP-PEG-PCL calculated from the test results is 12000Da. The above data confirm that VBP-PEG-PCL was successfully prepared.

[0047] The VBP-N3 used in this example was commissioned to be synthesized by Qiangyao Biotechnology Co., Ltd., and its structure is shown below.

[0048]

[0049] Example 2

[0050] 0.7 mg of quercetin, 9.3 mg of VBP-PEG-PCL (prepared in Example 1) and 3 mL of dimethylformamide (DMF) were added to a 50 mL glass round-bottom flask, and 5 mL of ultrapure water was added during vigorous stirring. The mixture was dialyzed for 12 hours using a dialysis bag with a molecular weight cutoff of 5000 Da, and the water was changed every two hours. Finally, the final product Qu-VBP-PEG-PCL was obtained by lyophilization.

[0051] The final product Qu-VBP-PEG-PCL was analyzed by UV absorption spectroscopy, particle size analysis and potential measurement. Figure 3 shown. Figure 3 middle Figure 3 A is the UV absorption spectrum test result, from which it can be seen that the quercetin loading is 6.8% and the encapsulation efficiency is 87.5%; Figure 3 B is the particle size test result. The particle size of the final product Qu-VBP-PEG-PCL after loading quercetin is 67.96±2.27nm; Figure 3 C is the potential test result. After loading quercetin, the potential change of the final product Qu-VBP-PEG-PCL is -4.99±0.14. The above data confirm that Qu-VBP-PEG-PCL was successfully prepared.

[0052] Comparative Example 1

[0053] 0.7 mg of quercetin, 9.3 mg of ALK-PEG-PCL and 3 mL of dimethylformamide (DMF) were added to a 50 mL glass round-bottom flask. 5 mL of ultrapure water was added during vigorous stirring. The mixture was dialyzed for 12 hours using a dialysis bag with a molecular weight cutoff of 5000 Da, with the water replaced every two hours. Finally, the final product Qu-PEG-PCL was obtained by lyophilization.

[0054] The difference between this comparative example and Example 2 is that the carrier for loading quercetin is changed from VBP-PEG-PCL to ALK-PEG-PCL.

[0055] Comparative Example 2

[0056] The difference between this comparative example and Example 1 is that after the product VBP-PEG-PCL is obtained, quercetin is not loaded, that is, the final product VBP-PEG-PCL of this comparative example is obtained.

[0057] Test Example 1

[0058] The Qu-VBP-PEG-PCL prepared in Example 2 was dissolved in water to form a 0.009375 mM to 0.3 mM solution, which was recorded as the experimental group; at the same time, an equal volume of PBS buffer solution was recorded as the control group. 3 Mouse hippocampal neuron cells (HT22) were seeded in a 96-well plate at a density of 100 μL and incubated for one day. The next day, 20 μL of the control group and different concentrations of the experimental group solution were added, and the optimal concentration of Qu-VBP-PEG-PCL for the toxic effect on mouse hippocampal neuron cells (HT22) was determined using CCK-8 after overnight incubation. The results are shown in Figure 4 As shown, it can be seen that the cytotoxicity of the experimental group and the control group solutions to HT22 cells is not much different, which means that the toxicity of Qu-VBP-PEG-PCL solutions of different concentrations to mouse cells is relatively small, indicating that Qu-VBP-PEG-PCL has good biocompatibility.

[0059] Test Example 2

[0060] HT22 cells were cultured at 2×10 4 The cells were seeded at a density of 1.50 μg / mL in a 24-well plate. After one day of incubation, 200 μL of PBS buffer solution was added to form the control group. The other groups were added with 200 μL of 100 μM hydrogen peroxide (H2O2) to simulate the oxidative stress state after cerebral hemorrhage. The group without drug was marked as the ICH group, the group with VBP-PEG-PCL was marked as the ICH+VBP-PEG-PCL group, and the group with Qu-VBP-PEG-PCL was marked as the

[0061] ICH+Qu-VBP-PEG-PCL group.

[0062] The above groups of samples were tested using phalloidin staining and live-dead staining, respectively. The specific methods are as follows:

[0063] Phalloidin staining: Wash each group of sample cells or tissue sections twice with PBS buffer solution, fix the cells with 3.7% formaldehyde solution at room temperature for 20 minutes, and then wash them with 0.1% Triton X-100 in PBS buffer solution 4 times, 5 minutes each time. Dilute Actin-Tracker Red with 0.1% Triton X-100 in PBS buffer solution at a ratio of 1:200 and use it as the staining working solution. Add 200 μL of the staining working solution to each well and incubate at room temperature in the dark for 30 minutes. Finally, wash 2 to 4 times with PBS buffer solution containing 0.1% Triton X-100 for 5 minutes each time, and then observe with a fluorescence microscope.

[0064] Live-dead staining: remove the culture medium from each group of samples, wash the cells once with PBS buffer solution, add 250 μL of 1:1000 diluted Calcein AM / PI detection working solution to each well, incubate at 37°C in the dark for 30 min, and observe the staining effect under a fluorescence microscope.

[0065] The results are as follows Figure 5 As shown, Figure 5 A is the result of phalloidin staining. Figure 5 B is the live-dead staining result. Figure 5 C is the quantitative result of live-dead staining. It can be seen that, except for the ICH group, the cytotoxicity of the ICH+VBP-PEG-PCL group and the ICH+Qu-VBP-PEG-PCL group is not much different from that of the Control group, which indicates that the VBP-PEG-PCL and Qu-VBP-PEG-PCL provided by the present invention have good biocompatibility.

[0066] Test Example 3

[0067] The same method as in Experimental Example 2 was used to set up each group, and the DCFH-DA fluorescent probe was used to stain the samples for active oxygen. The results are as follows: Figure 6 A and Figure 6 As shown in B, Figure 6 A is the result of reactive oxygen species immunofluorescence staining. Figure 6 B is the quantitative result of reactive oxygen species immunofluorescence staining, from which it can be seen that compared with the ICH group and the ICH+VBP-PEG-PCL group, the test results of the ICH+Qu-VBP-PEG-PCL group can significantly inhibit the release of ROS and maintain the normal physiological activity of cells.

[0068] EDU-488 staining was used to evaluate the ability of each group of samples to maintain cell proliferation in the ROS environment. The specific method was as follows: 2X (20μM) EdU working solution was prepared, then preheated to 37°C, and added to a 24-well plate in an equal volume of culture medium, so that the final EdU concentration in the 24-well plate became 1X, and the cells were incubated for 24 hours. After EdU labeling of cells, the culture medium was removed, and 1mL of 4% paraformaldehyde was added, fixed at room temperature for 15 minutes, the fixative was removed, and the cells were washed 3 times with 1mL of washing solution (3% BSA solution) per well, each time for 3 to 5 minutes. The washing solution was removed, and 1mL of permeabilization solution (0.3% Triton X-100 in PBS buffer solution) was used per well, and incubated at room temperature for 10 to 15 minutes. The permeabilization solution was removed, and the cells were washed 1 to 2 times with 1mL of washing solution per well, each time for 3 to 5 minutes. Then add 0.5mL Click reaction solution to each well, shake the culture plate gently to ensure that the reaction mixture can evenly cover the sample, incubate at room temperature in the dark for 30 minutes, remove the Click reaction solution, wash with washing solution 3 times, 3 to 5 minutes each time, and finally detect with a fluorescence microscope. Figure 6 C and Figure 6 D, where Figure 6 C is a staining electron microscope photo. Figure 6 D is the staining quantitative result, from which it can be seen that compared with the ICH group and the ICH+VBP-PEG-PCL group, the ICH+Qu-VBP-PEG-PCL group can also maintain the cell viability of HT22 well in the ROS environment and improve the proliferation ability of HT22.

[0069] Test Example 4

[0070] The same method as in Experimental Example 2 was used to set up each group of samples, and the expression of pro-inflammatory genes and anti-inflammatory genes, pro-inflammatory factors and anti-inflammatory factors in microglia (BV-2) was detected by RT-qPCR and Elisa. The results are as follows Figure 7 As shown, Figure 7 A~ Figure 7 C respectively show the expression results of anti-inflammatory genes CD206, YM1 / 2 and TGF-β; Figure 7 D~ Figure 7 F are the expression results of pro-inflammatory genes IFN-γ, CD16 / 32, and iNOS; Figure 7 G and Figure 7 H are the expression results of pro-inflammatory factors TNF-α and IL-1β; Figure 7 I and Figure 7 J are the expression results of anti-inflammatory factors IL-10 and TGF-β. Figure 7It can be seen from the test results that the ICH+Qu-VBP-PEG-PCL group can effectively reduce the expression of pro-inflammatory genes and pro-inflammatory factors in BV-2, and increase the expression of anti-inflammatory genes and anti-inflammatory factors, which indicates that Qu-VBP-PEG-PCL can effectively reverse the polarization of BV-2 to M1 phenotype, promote M2 phenotype polarization, and enhance the anti-inflammatory effect.

[0071] Test Example 5

[0072] Using 6-8 week old C57BL / 6J male mice, 0.075U / 0.5μL collagenase IV was dissolved in PBS buffer solution and injected into the right basal ganglia with slight changes. First, the drilling position was selected at 0.2mm in front of the anterior bregma, 2.2mm to the right of the midline, and 3.5mm ventral; then, 0.075U collagenase IV was dissolved in 0.5μL PBS buffer solution and injected within 5min with the help of a microinfusion pump; in addition, the drilling hole was sealed with bone wax after slowly withdrawing it within 2 minutes to avoid backflow. PBS buffer solution, ALK-PEG at a concentration of 10mg / ml were injected at 6 hours, 24 hours, and 48 hours after the completion of the model. 5000 -PCL 5000 , Qu-PEG-PCL, VBP-PEG-PCL and Qu-VBP-PEG-PCL, recorded as ICH group, mPEG-PCL group, Qu-PEG-PCL group, VBP-PEG-PCL group and Qu-VBP-PEG-PCL group, and a sham operation (sham) group was set up at the same time.

[0073] Then, the targeted hemostatic performance and anti-inflammatory ability of the above-mentioned samples were verified in mice. The changes of mouse hematoma were observed by brain tissue hematoma content and dynamic speckle imaging. The permeability of the blood-brain barrier of mice was evaluated by Evans blue staining. The enrichment of each group of samples in the brain hemorrhage part of mice was observed by in vivo imaging. The results are as follows: Figure 8 As shown, Figure 8 A is the hematoma content of mouse cerebral hemorrhage. Figure 8 B Evans blue staining results. Figure 8 C is the in vivo imaging of mice at 6h, 12h, and 24h after administration. Figure 8 D is the in vitro imaging of mouse brain tissue at 6h, 12h, and 24h after administration. Figure 8 E is the dynamic speckle imaging of mice at 0, 1, 3, and 7 days after intracerebral hemorrhage. Figure 8 F is the quantitative analysis of blood flow recovery degree using dynamic speckle imaging. Figure 8 G is the dynamic speckle imaging vascular diameter quantification. Figure 8 As can be seen in A, compared with the other groups, the intracranial hematoma of mice treated with Qu-VBP-PEG-PCL group was significantly reduced; Figure 8B shows that the blood-brain barrier permeability of mice treated with Qu-VBP-PEG-PCL group was significantly restored; Figure 8 C and Figure 8 The in vivo imaging of mice in D showed that the samples in the Qu-VBP-PEG-PCL group had good targeting ability, and the drug was still highly enriched at the injury site 24 hours after administration; Figure 8 The dynamic speckle results of E to G showed that the cerebral surface blood flow of mice treated with Qu-VBP-PEG-PCL group was significantly restored on the 7th day, which had a positive effect on the prognosis of cerebral hemorrhage.

[0074] After reaching the treatment cycle, the treated mice were euthanized, and the brain tissues were removed for CD163 / Iba-1 / dapi staining and CD206 / Iba-1 / dapi staining to evaluate the polarization state of microglia after intracerebral hemorrhage. HE staining and Nissl body staining were also performed. The results are shown in Fig. 9 As shown, Fig. 9 A is CD163 / Iba-1 / dapi immunofluorescence staining of brain tissue; Fig. 9 B is CD206 / Iba-1 / dapi immunofluorescence staining of brain tissue; Fig. 9 C is the fluorescence quantification of CD163 / Iba-1 / dapi in brain tissue; Fig. 9 D is the fluorescence quantification of CD206 / Iba-1 / dapi in brain tissue; Fig. 9 E is HE staining of brain tissue; Fig. 9 F is Nissl body staining of brain tissue. Fig. 9 A and Fig. 9 The CD163 / Iba-1 / dapi staining results of brain tissue in C showed that the microglia of mice treated with Qu-VBP-PEG-PCL group were less polarized toward M1 and more polarized toward M2 phenotype ( Fig. 9 B and Fig. 9 D) At the same time Fig. 9 HE staining results of E and Fig. 9 The Nissl body staining results of F showed that the hematoma and intracranial neuronal damage were significantly reduced in the mice treated with Qu-VBP-PEG-PCL group after treatment.

[0075] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A drug, characterized in that Having the structure of formula (I): Wherein, n is an integer from 3000 to 7000, and m is an integer from 3000 to 7000; R is the residue after removing any amino group from the polypeptide VBP.

2. The drug according to claim 1, characterized in that It has the following structure: Here, n is an integer from 3000 to 7000, and m is an integer from 3000 to 7000.

3. A pharmaceutical composition, characterized in that include: The drug according to claim 1 or claim 2 and quercetin or a quercetin derivative.

4. The pharmaceutical composition according to claim 3, characterized in that The mass ratio of the quercetin or quercetin derivative to the drug is 1:(10-20).

5. The pharmaceutical composition according to claim 3, characterized in that The particle size is 50~80nm.

6. The pharmaceutical composition according to claim 3, characterized in that The encapsulation rate of the quercetin or quercetin derivative is 80-95%; the drug loading is 5-10%.

7. A method for preparing a drug, characterized in that: include: A click reaction is performed between the azide-modified polypeptide VBP and the alkynyl-polycaprolactone-polyethylene glycol represented by formula (II) to obtain the drug represented by formula (I); Wherein, n is an integer from 3000 to 7000, and m is an integer from 3000 to 7000; R is the residue after removing any amino group from the polypeptide VBP.

8. The preparation method according to claim 7, characterized in that: The molar ratio of the azide-modified polypeptide VBP to the alkynyl-polycaprolactone-polyethylene glycol for click reaction is 1:(800-1200).

9. The preparation method according to claim 7, characterized in that: The click reaction time is 10 to 20 hours, and the temperature is 20 to 50°C.

10. Use of the drug according to claim 1, the drug according to claim 2, the pharmaceutical composition according to any one of claims 3 to 6, or the drug prepared by any one of the preparation methods of claims 7 to 9 in the field of treating cerebral hemorrhage.

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