Modified dendritic PAMAM (Polyamidoamine) polymer hydrogel as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of anti-tumor materials, and discloses a modified dendritic PAMAM polymer hydrogel as well as a preparation method and application thereof. The method comprises the following steps: reacting a modified dendritic PAMAM polymer with oxidized dextran in a water-containing system to obtain the modified dendritic PAMAM polymer hydrogel, the structural formula of the modified dendritic PAMAM polymer is as shown in formula I; the dendritic PAMAM is a fifth-generation polymer, and n is an integer from 1 to 128. The modified dendritic PAMAM polymer hydrogel provided by the invention is used for preparing antitumor drugs. The hydrogel disclosed by the invention has a relatively strong anti-tumor effect; small molecule drugs or antibody drugs are entrapped, so that the tumor immune microenvironment can be jointly regulated and controlled, and the anti-tumor effect is further enhanced. # imgabs0 #

Description

Technical Field

[0001] The present invention belongs to the technical field of anti-tumor materials, and in particular relates to a modified dendritic PAMAM polymer hydrogel and a preparation method and application thereof. Background Art

[0002] The immune system is involved in most physiological and pathological processes of the body, including the occurrence and development of tumors. Tumor immunotherapy provides a promising method for inhibiting or even curing tumors by activating the host immune system to specifically identify and eliminate tumor cells. Among them, immunogenic cell death (ICD) is an important mechanism that transforms the traditionally considered "silent" tumor cell death process into a key event for activating anti-tumor immune responses. When tumor cells undergo ICD under stimulation such as chemotherapy, radiotherapy, or photodynamic therapy, they release damage-associated molecular patterns such as calreticulin (CALR), adenosine triphosphate (ATP), and high-mobility group protein B1 (HMGB1), which activate dendritic cells through the "antigen exposure-danger signal transmission" cascade reaction, thereby initiating antigen-specific T cell immune responses. In the past decade, since the concept of ICD was proposed, there have been limited drugs that can induce ICD in tumor cells.

[0003] In the tumor microenvironment, tumor-associated macrophages (TAMs) are key immune cells that regulate tumor progression. The infiltration rate of TAMs in solid tumors can be as high as 50%. Among them, M2 macrophages promote tumor progression through multiple mechanisms: secreting immunosuppressive factors such as interleukin-10 (IL-10) and transforming growth factor-β (TGF-β), which inhibit T cell function; expressing pro-angiogenic factors such as vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9), which accelerate tumor metastasis; and constructing a fibrotic matrix to form a physical barrier that hinders drug penetration. Therefore, reprogramming tumor-promoting M2 TAMs into anti-tumor M1 macrophages has become an important strategy to improve the tumor immunosuppressive microenvironment and enhance the anti-tumor immune response.

[0004] However, currently, there are very limited materials that can effectively regulate the phenotypic polarization of macrophages, which greatly restricts the development of tumor immunotherapy based on TAMs reprogramming. Although traditional small molecule inhibitors can repolarize M2 macrophages to M1 macrophages, they have defects such as poor targeting and high systemic toxicity. Although gene editing technology can regulate phenotype in a targeted manner, it faces low delivery efficiency and off-target risks. Developing new materials to achieve efficient reprogramming of TAMs and induce tumor immunogenic cell death is beneficial to reversing the tumor immunosuppressive microenvironment and synergistically enhancing anti-tumor immune responses. Therefore, designing and constructing materials with macrophage reprogramming functions and ICD inducers is not only an important research direction in the field of tumor immunotherapy, but also a key breakthrough in achieving precision tumor treatment.

[0005] The modified dendritic PAMAM polymer hydrogel prepared by the present invention can not only regulate macrophages and transform macrophages from M2 anti-inflammatory type to M1 pro-inflammatory type, but also has the characteristics of an ICD inducer and has a strong anti-tumor effect. Summary of the Invention

[0006] In order to overcome the shortcomings and deficiencies of the prior art, the object of the present invention is to provide a class of modified dendritic PAMAM polymer hydrogels and their preparation methods and applications. The hydrogel of the present invention can not only regulate macrophages, causing macrophages to transform from M2 anti-inflammatory type to M1 pro-inflammatory type, but also has immune activity and can be used as a carrier to improve the anti-tumor effect. The present invention prepares a class of modified dendritic PAMAM polymer hydrogels by modifying different amounts of N-(2-hydroxyethyl)hexamethylenediamine to polyamide-amine dendrimers, which has a strong anti-tumor effect. The hydrogel of the present invention is used to prepare anti-tumor drugs. The hydrogel of the present invention further improves the anti-tumor effect by delivering small molecule drugs or antibody drugs, and the combined use of hydrogel and drugs.

[0007] The purpose of the present invention is achieved through the following technical solutions:

[0008] A method for preparing a modified dendritic PAMAM polymer hydrogel comprises the following steps: reacting a modified dendritic PAMAM polymer with oxidized dextran in an aqueous system to obtain a modified dendritic PAMAM polymer hydrogel;

[0009] The structural formula of the modified dendritic PAMAM polymer is: In the structural formula, -NH- comes from the -NH2 at the end of the dendritic PAMAM polymer.

[0010] The dendritic PAMAM is a fifth generation polymer.

[0011] When the PAMAM is the fifth generation PAMAM, n is an integer of 1 to 128.

[0012] The n represents the number of N-(2-hydroxyethyl)hexamethylenediamine grafted onto the surface of PAMAM.

[0013] The PAMAM is the fifth generation PAMAM, and n is an integer of 1 to 128, preferably 40 to 80 (such as 40, 60 and 80), more preferably 50 to 80, and further preferably 50 to 70.

[0014] The oxidized dextran is obtained by oxidizing the hydroxyl groups of dextran into aldehyde groups, such as oxidizing the hydroxyl groups of dextran into aldehyde groups by sodium periodate.

[0015] The molecular weight of the dextran is 40,000-70,000 Da.

[0016] The oxidation degree of the oxidized dextran is 20-40%.

[0017] The mass ratio of the modified dendritic PAMAM polymer to the oxidized dextran is 8:1 to 1:20, preferably 1:(1.5 to 3).

[0018] The preparation method comprises the following steps: preparing a modified dendritic PAMAM polymer and oxidized dextran into separate solutions to obtain a modified dendritic PAMAM polymer solution and an oxidized dextran solution; and mixing the two solutions to react and obtain a hydrogel. The concentration of the modified dendritic PAMAM polymer solution is 30-100 mg / mL, preferably 45-55 mg / mL; and the concentration of the oxidized dextran solution is 50-150 mg / mL, preferably 95-105 mg / mL.

[0019] The solvent of the modified dendritic PAMAM polymer solution is water or PBS buffer solution.

[0020] The solvent of the oxidized dextran solution is water or PBS buffer solution.

[0021] The volume ratio of the modified dendritic PAMAM polymer solution to the oxidized dextran solution is 4:1 to 1:4, preferably 1:1.

[0022] The hydrogel is used to prepare an anti-tumor drug that can regulate macrophages and transform the macrophages from an M2 anti-inflammatory type to an M1 pro-inflammatory type.

[0023] The tumor in the anti-tumor drug refers to colorectal cancer, breast cancer, melanoma, liver cancer and / or glioma.

[0024] Furthermore, the hydrogel is used to prepare an anti-tumor drug that can regulate macrophages, causing them to transform from an M2 anti-inflammatory type to an M1 pro-inflammatory type, and can also induce immunogenic cell death in tumor cells.

[0025] An anti-tumor drug that can regulate macrophages, transform macrophages from M2 anti-inflammatory type to M1 pro-inflammatory type and induce immunogenic cell death of tumor cells, including the above-mentioned hydrogel.

[0026] The drugs also include small molecule drugs or antibody drugs. The hydrogel is used not only as a carrier but also in combination with small molecule drugs or antibody drugs to induce tumor immunogenic cell death to exert anti-tumor effects and to regulate macrophages to kill tumor cells.

[0027] The small molecule drugs or antibody drugs include αPD-1 antibodies and αCD47 antibodies.

[0028] The hydrogel is used as a preparation for delivering small molecule drugs or antibody drugs, especially drugs for tumor immunotherapy.

[0029] The hydrogel of the present invention is used by injection or by freeze-drying before implantation.

[0030] The tumor medicine of the present invention is used by injection or by freeze-drying before implantation.

[0031] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0032] (1) The present invention provides a class of hydrogels prepared from modified dendritic PAMAM polymers for use in anti-tumor drugs; the hydrogels of the present invention can not only regulate macrophages, causing macrophages to transform from M2 anti-inflammatory type to M1 pro-inflammatory type, but also induce immunogenic cell death of tumor cells, and have a strong anti-tumor effect.

[0033] (2) The hydrogel of the present invention is an injectable hydrogel. The in vivo injectable hydrogel of the present invention can increase the accumulation of drugs at the tumor site, continuously and slowly release drugs, and local administration can induce a systemic anti-tumor immune response;

[0034] (3) The hydrogel of the present invention can encapsulate small molecule drugs or antibody drugs to jointly regulate the tumor immune microenvironment and enhance the anti-tumor effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 is the H NMR spectrum of C7A-CD1 in deuterated chloroform (CDCl3);

[0036] Figure 2 For G5P-C7A 40 (G5-40), G5P-C7A 60 (G5-60) and G5P-C7A 80 H NMR spectrum (CDCl3) of (G5-80);

[0037] Figure 3 For CT26 cells and G5P-C7A 40 、G5P-C7A 60 and G5P-C7A 80 (G5-80) Average fluorescence intensity of calreticulin (CALR) expression on the surface of CT26 cells after 4 hours of incubation; G4P-C7A 60 : The fourth generation PAMAM replaces the fifth generation PAMAM in Example 1, and other conditions are the same as in Example 1;

[0038] Figure 4 For G5P-C7A 60In vitro induction of macrophage repolarization, (A) normalized mRNA expression level, (B) CD86 expression;

[0039] Figure 5 For G5P-C7A 60 Spectra of storage modulus (G') and loss modulus (G") of Gel as a function of frequency;

[0040] Figure 6 For G5P-C7A 60 Storage modulus (G') and loss modulus (G") of Gel as a function of strain;

[0041] Figure 7 Optical images and quantitative analysis of G5P-C7A at different time points after subcutaneous injection of hydrogel into mice 60 Remaining weight curves of Gel at 1 hour, 7 days, 14 days, and 21 days;

[0042] Figure 8 is the average tumor growth curve of CT26 tumor-bearing mice after injection of different drugs;

[0043] Figure 9 The primary tumor on the right side of the mouse was implanted with freeze-dried hydrogel, while the distal tumor on the left side of the mouse was not implanted with drugs; the average growth curve of the primary tumor on the right side of the mouse (a) and the average growth curve of the distal tumor on the left side (b);

[0044] Figure 10 Semi-quantitative analysis curve of bioluminescence signal intensity of tumor recurrence after injection of different drugs (a) and survival curve of mice (b). DETAILED DESCRIPTION

[0045] The present invention will be further described in detail below with reference to specific examples, but the embodiments of the present invention are not limited thereto.

[0046] Example 1:

[0047] (1) Activation of N-(2-hydroxyethyl)hexamethylenediamine

[0048] First, 1.2011 g of N,N'-carbonyldiimidazole (CDI) was mixed with 20.0 mL of anhydrous dichloromethane; 0.403 g of N-(2-hydroxyethyl)hexamethylenediamine was dissolved in 10.0 mL of anhydrous dichloromethane and then added dropwise to the CDI solution. The mixture was reacted at 25°C for 12 hours, then washed three times with saturated brine and dried over anhydrous magnesium sulfate. The solvent in the product was then removed by vacuum pump to obtain a pale yellow oily product, C7A-CDI, with a yield of 78.3%. The H-NMR spectrum of C7A-CDI is shown in Figure 2. Figure 1 shown.

[0049] (2) Synthesis of polymer materials modified with N-(2-hydroxyethyl)hexamethylenediamine

[0050] G5P-C7A 60 The synthesis of is described as an example. First, 25.1 mg of the fifth generation PAMAM was dissolved in 1 mL of anhydrous dimethyl sulfoxide (DMSO). Then, 20.5 mg of N-(2-hydroxyethyl)hexamethylenediamine activated by CDI was dissolved in 0.5 mL of anhydrous DMSO and slowly added to the above solution. Thereafter, the reaction vessel was sealed and reacted at a constant temperature of 40°C for 24 hours. After the reaction, the product was purified using a dextran gel column (LH-20) with methanol as the mobile phase, and the product was monitored and collected using thin layer chromatography spot plate technology. The synthesized polymer material is named according to the number of grafts of N-(2-hydroxyethyl)hexamethylenediamine. Subsequently, the product was subjected to a vacuum rotary evaporator to remove the solvent to obtain a light yellow product G5P-C7A. 60 , and its yield reached 72.3%. G5P-C7A 40 and G5P-C7A 80 Preparation method and G5P-C7A 60 Similar. G5P-C7A 40 、G5P-C7A 60 and G5P-C7A 80 The results of H NMR spectrum are as follows Figure 2 shown.

[0051] Example 2

[0052] A type of modified dendritic PAMAM polymer hydrogel (G5P-C7A 60 The preparation method of Gel hydrogel comprises the following steps: using water to 60 and oxidized dextran Dex-CHO (dextran molecular weight 50000-60000Da; the oxidation degree of the oxidized dextran is 33%) were prepared into solutions to obtain G5P-C7A 60 solution (50 mg / mL) and Dex-CHO solution (100 mg / mL); the two solutions were mixed in equal volumes to obtain a hydrogel (G5P-C7A 60 Gel hydrogel). G5P-C7A 40 Hydrogel and G5P-C7A 80 Hydrogel and G5P-C7A 60 Gel hydrogel is similar.

[0053] Example 3: G5P-C7A 60 Induces immunogenic tumor cell death

[0054] CT26 cells were treated with G5P-C7A 40 (3μM), G5P-C7A 60 (3μM and 5μM), G5P-C7A 80 (3 μM) and OXA (200 μM) were treated for 4 hours. The expression of CALR on the cell membrane was quantified by flow cytometry. Figure 3 As shown in the flow cytometry results, OXA can promote the expression of CALR on the cell surface to a certain extent, G5P-C7A 60 (5 μM) and G5P-C7A 80 (3μM) significantly enhanced the exposure of CALR on the cell membrane. 40 (3μM), G5P-C7A 60 (3μM and 5μM), G5P-C7A 80 The expression of CALR on the cell surface of CT26 cells treated with OXA (3μM) and OXA (200μM) increased by 4-fold, 3-fold, 10-fold, 31-fold and 2-fold, respectively.

[0055] G5P-C7A 60 Gel hydrogel, G5P-C7A 80 The hydrogels were also able to induce immunogenic cell death in tumors.

[0056] Figure 3 For CT26 cells and G5P-C7A 40 、G5P-C7A 60 and G5P-C7A 80 The mean fluorescence intensity of calreticulin (CALR) expression on the surface of CT26 cells after 4 hours of incubation.

[0057] Example 4: Monocytes were extracted from mouse bone marrow and induced for 5 days with 10 ng / mL M-CSF to obtain mature BMDMs. The cells were then digested and harvested, seeded into well plates, and incubated for 24 hours with 20 ng / mL IL-4 to induce BMDM differentiation into the M2 phenotype. The old culture medium was removed, and fresh blank or drug-containing complete culture medium was added and incubated for 12 hours (RT-PCR) or 24 hours (flow cytometry). Figure 4 As shown in A, the results of RT-qPCR experiments showed that G5P-C7A 60 It can upregulate the expression of macrophage CD80, THFA, IFNB, and IL1B genes and downregulate the expression of CD206. Flow cytometry results showed that ( Figure 4 B), G5P-C7A 60It can activate M2 macrophages and significantly upregulate the expression of macrophage activation marker CD86. The above experimental results show that G5P-C7A 60 It can directly regulate macrophages, causing them to transform from M2 anti-inflammatory type to M1 pro-inflammatory type.

[0058] G5P-C7A 60 Gel hydrogel can also regulate macrophages, causing them to transform from M2 anti-inflammatory type to M1 pro-inflammatory type.

[0059] Figure 4 For G5P-C7A 60 In vitro induction of macrophage repolarization, (A) normalized mRNA expression levels, (B) CD86 expression.

[0060] Example 5: G5P-C7A 60 Gel preparation and rheological properties testing

[0061] 30 μL G5P-C7A 60 The (50 mg / mL) aqueous solution was mixed with 30 μL of Dex-CHO aqueous solution (100 mg / mL), and the mixture was immediately transferred to the plate of the rheometer. 60 The mechanical properties of the gel were evaluated by measuring the oscillation frequency using a parallel plate with a diameter of 8 mm. Figure 5 As shown in the figure, its storage modulus (G') remains basically constant in the frequency range of 0.01 to 10 Hz, while the loss modulus (G") is frequency dependent. In addition, G' is always greater than G' in the entire frequency range, indicating that G5P-C7A 60 Gel hydrogel has a cross-linked network structure. Figure 6 As shown, further strain sweep results show that the storage modulus (G') and loss modulus (G") of the gel remain almost unchanged until the strain approaches 80%, indicating that the hydrogel has the potential to withstand relatively large elastic deformation. However, G' and G" intersect at 125%, which is a critical point indicating the collapse of the hydrogel network.

[0062] Figure 5 For G5P-C7A 60 The storage modulus (G') and loss modulus (G") of Gel vary with frequency.

[0063] Figure 6 For G5P-C7A 60 The storage modulus (G') and loss modulus (G") of gel vary with strain.

[0064] Example 6: In vivo hydrogel degradation evaluation

[0065] 100 mg / mL Dex-CHO and 50 mg / mL G5P-C7A 60 Prepare G5P-C7A by mixing equal volumes at room temperature 60 Then 50 μL of G5P-C7A 60 The hydrogel was injected into the back of BALB / c mice. The mice were then treated 1 hour, 7 days, 14 days, and 21 days after injection. The remaining hydrogel in each mouse was photographed and its weight was recorded. Figure 7 As shown, G5P-C7A 60 One week after the Gel injection, the weight dropped by about 22%, 50% after 14 days, and about 18% after 21 days.

[0066] Figure 7 Optical images and quantitative analysis of G5P-C7A at different time points after subcutaneous injection of hydrogel into mice 60 Remaining weight curves of the gel at 1 hour, 7 days, 14 days, and 21 days.

[0067] Example 7: G5P-C7A 60 Gel in vivo anti-tumor therapy

[0068] To establish a CT26 colorectal cancer subcutaneous tumor model, CT26 cells were first cultured to the logarithmic growth phase and then digested with 0.25% trypsin. After digestion, complete medium containing serum was added to terminate the digestion, and the cells were washed three times with PBS. Afterwards, the cells were counted and the cell concentration was adjusted to 1×10 7 / mL. Subsequently, 1×10 6 CT26 cell suspension.

[0069] When the volume of mouse CT26 tumor reached 50-100 mm 3 At the same time, mice were randomly divided into six treatment groups and intratumorally administered with: PBS, G5P-C7A 60 (G5P-C7A 60 :10mg / kg), G5P-C7A 60 Gel (hydrogel, G5P-C7A 60 :10mg / kg), G5P-C7A 60 Gel (hydrogel, G5P-C7A 60 : 40mg / kg) and G5P-C7A 60 Gel (hydrogel, G5P-C7A 60 : 60mg / kg). Figure 8As shown, local administration of free G5P-C7A 60 Tumors in mice treated with G5P-C7A showed a delay in tumor growth, while 60 Mice treated with Gel showed a more significant inhibitory effect on tumor growth.

[0070] Figure 8 Figure 2 is the average tumor growth curve of CT26 tumor-bearing mice after injection of different drugs.

[0071] Example 8: Bilateral tumor model

[0072] To establish a syngeneic bilateral tumor model of CT26 colorectal cancer, CT26 tumor cells cultured to the logarithmic growth phase were first digested with 0.25% trypsin. After digestion, complete medium containing serum was added to terminate the digestion, and the cells were washed three times with PBS. The cells were then counted and the cell density was adjusted to 1×10 7 Afterwards, 1×10 6 CT26 tumor cell suspension was used to establish a primary tumor model. Four days before treatment, the same amount of CT26 tumor cell suspension was injected subcutaneously on the left side of the back of the mouse as a distal tumor. When the size of the primary tumor reached 50-100 mm 3 BALB / c mice bearing bilateral CT26 tumors were divided into the following four groups: PBS, G5P-C7A 60 &αPD-1: Free G5P-C7A 60 +αPD-1, G5P-C7A 60 Gel (hydrogel prepared in Example 2) and G5P-C7A 60 &αPD-1Gel:G5P-C7A 60 +αPD-1 hydrogel. Among them, 100mg / mL Dex-CHO and 50mg / mL G5P-C7A 60 Prepare G5P-C7A by mixing equal volumes at room temperature 60 Gel hydrogel, G5P-C7A 60 The dosage of the drug was 10 mg / kg, and the dosage of αPD-1 antibody used in each mouse was 100 μg. The αPD-1 antibody was loaded on G5P-C7A 60 The hydrogel was freeze-dried and then implanted into the tumor site. After the primary tumor was treated, the size of the primary tumor and distal tumors were measured every two days.

[0073] G5P-C7A 60&αPD-1Gel: 100 mg / mL Dex-CHO aqueous solution (dextran molecular weight 50000-60000 Da; the oxidation degree of the oxidized dextran is 33%) and 50 mg / mL G5P-C7A 60 The aqueous solution was mixed at room temperature in equal volumes, and then αPD-1 antibody was added to obtain G5P-C7A. 60 &αPD-1Gel;G5P-C7A 60 The dosage of the drug was 10 mg / kg, and the dosage of αPD-1 antibody used in each mouse was 100 μg.

[0074] like Figure 9 As shown, G5P-C7A 60 &αPD-1Gel not only effectively inhibited the growth of primary tumors, but also inhibited the growth of distal tumors. 60 and αPD-1 sustained release in hydrogels, G5P-C7A 60 The primary tumor and distal tumor of the &αPD-1Gel group were significantly smaller than those of the other groups. 60 &αPD-1 group (free G5P-C7A 60 +αPD-1) had a distal tumor inhibition rate of 43%, while G5P-C7A 60 The distal tumor inhibition rate of the Gel group was 60%. 60 The αPD-1Gel group significantly delayed tumor growth, and the inhibition rate of distal tumors reached 82%. This shows that the αPD-1 antibody loaded on G5P-C7A 60 hydrogel, and the combined use effectively stimulated a systemic anti-tumor response.

[0075] Figure 9 The primary tumor on the right side of the mouse was implanted with freeze-dried hydrogel, while the distal tumor on the left side of the mouse was not implanted with drugs; the average growth curve of the primary tumor on the right side of the mouse (a) and the average growth curve of the distal tumor on the left side (b).

[0076] Example 9: Evaluation of the effect of hydrogel in GL261 brain tumor postoperative model

[0077] To establish an orthotopic GL261 brain tumor mouse model, 3 μL of luciferase-expressing GL261-luc cells (2 × 10 cells per mouse) were injected into the brains of C57BL / 6 mice. 5 The injection site was 2 mm to the right, 1 mm in front, and 2 mm deep from the fontanelle. On the 8th day after tumor implantation, 90% of the glioblastoma was removed by surgery, and different drugs were given in the resection cavity: PBS group, G5P-C7A 60&aCD47 group (free G5P-C7A 60 and aCD47;G5P-C7A 60 :10mg / kg; αCD47: 50μg / per), G5P-C7A 60 Gel group (hydrogel, G5P-C7A 60 :10mg / kg), G5P-C7A 60 &αCD47Gel group (G5P-C7A 60 : 10 mg / kg; αCD47: 50 μg / per). Among them, αCD47 antibody was loaded on G5P-C7A 60 After intraperitoneal injection of D-luciferin (150 mg / kg) into mice, the size of GL261 brain tumors was monitored by an in vivo imaging system, and the survival of the mice was also monitored. Figure 10 As shown in the figure, the bioluminescence signal decreased significantly after tumor surgical resection. The mice in the control surgery group experienced rapid tumor recurrence, and all mice died on the 40th day after tumor implantation, with a median survival of 37 days and a significant loss of body weight. 60 In the &αCD47 group, tumor recurrence was delayed, but all mice died on the 50th day after tumor implantation, with a median survival of 44 days. 60 &αCD47 group, G5P-C7A 60 Gel was more effective in inhibiting the proliferation of residual tumors and delaying recurrence, increasing the median survival to 60 days. 60 The &αCD47 Gel group showed a slow tumor recurrence rate and significantly prolonged median survival time, with a survival rate of 50%, which was a better effect. 60 &αCD47 Gel hydrogel has the potential to inhibit the recurrence of malignant glioblastoma as an auxiliary means after surgery.

[0078] G5P-C7A 60 &αCD47 Gel: 100 mg / mL Dex-CHO aqueous solution (dextran molecular weight 50000-60000 Da; the oxidation degree of the oxidized dextran is 33%) and 50 mg / mL G5P-C7A 60 Equal volumes of aqueous solutions were mixed at room temperature, and then αCD47 was added to obtain G5P-C7A. 60 &αCD47 Gel;

[0079] G5P-C7A 60 The dosage of the drug was 10 mg / kg, and the dosage of αCD47 used in each mouse was 50 μg.

[0080] Figure 10 Semi-quantitative analysis curve of bioluminescence signal intensity of tumor recurrence after injection of different drugs (a) and survival curve of mice (b).

Claims

1. A method for preparing a modified dendritic PAMAM polymer hydrogel, characterized by: The following steps are involved: The modified dendritic PAMAM polymer and oxidized dextran are reacted in an aqueous system to obtain a modified dendritic PAMAM polymer hydrogel; The structural formula of the modified dendritic PAMAM polymer is Formula I: In the structural formula, -NH- comes from the -NH2 at the end of the dendritic PAMAM polymer; The dendritic PAMAM is a fifth-generation polymer; When the PAMAM is the fifth generation PAMAM, n is an integer from 1 to 128; The n represents the number of N-(2-hydroxyethyl)hexamethylenediamine grafted onto the surface of PAMAM.

2. The method for preparing the modified dendritic PAMAM polymer hydrogel according to claim 1, wherein: The PAMAM is the fifth generation PAMAM, and n is an integer from 40 to 80; The oxidized dextran is obtained by oxidizing the hydroxyl groups of dextran into aldehyde groups; The molecular weight of the dextran is 40,000-70,000 Da; The oxidation degree of the oxidized dextran is 20-40%; The mass ratio of the modified dendritic PAMAM polymer to the oxidized dextran is 8:1 to 1:

20.

3. The method for preparing the modified dendritic PAMAM polymer hydrogel according to claim 2, wherein: The mass ratio of the modified dendritic PAMAM polymer to oxidized dextran is 1:(1.5-3); The n is an integer of 50 to 80.

4. The method for preparing the modified dendritic PAMAM polymer hydrogel according to claim 1, characterized in that: The specific steps are: preparing a modified dendritic PAMAM polymer and oxidized dextran into solutions respectively to obtain a modified dendritic PAMAM polymer solution and an oxidized dextran solution; mixing the two solutions to react to obtain a hydrogel; The concentration of the modified dendritic PAMAM polymer solution is 30-100 mg / mL; the concentration of the oxidized dextran solution is 50-150 mg / mL; The solvent of the modified dendritic PAMAM polymer solution is water or PBS buffer solution; The solvent of the oxidized dextran solution is water or PBS buffer solution; The volume ratio of the modified dendritic PAMAM polymer solution to the oxidized dextran solution is 4:1 to 1:

4.

5. An application of the modified dendritic PAMAM polymer hydrogel obtained by the preparation method according to any one of claims 1 to 4, characterized in that: The modified dendritic PAMAM polymer hydrogel is used for preparing anti-tumor drugs.

6. The use according to claim 5, characterized in that: The anti-tumor drug is an anti-tumor drug that can regulate macrophages to transform macrophages from M2 anti-inflammatory type to M1 pro-inflammatory type, or an anti-tumor drug that can both regulate macrophages to transform macrophages from M2 anti-inflammatory type to M1 pro-inflammatory type and induce immunogenic cell death of tumor cells; The modified dendritic PAMAM polymer hydrogel is used by injection or freeze-drying and then implanting; The tumor drug is used by injection or by freeze-drying and then implanting.

7. The use according to claim 5, characterized in that: The tumor in the anti-tumor drug refers to colorectal cancer, breast cancer, melanoma, liver cancer and / or glioma.

8. An anti-tumor drug, characterized in that: The invention comprises a modified dendritic PAMAM polymer hydrogel obtained by the preparation method according to any one of claims 1 to 4.

9. The anti-tumor drug according to claim 8, characterized in that: It also includes small molecule drugs or antibody drugs; the modified dendritic PAMAM polymer hydrogel is used as a carrier of small molecule drugs or antibody drugs and is also used in combination with small molecule drugs or antibody drugs for anti-tumor effects; The small molecule drugs or antibody drugs include αPD-1 antibodies and αCD47 antibodies.

10. The anti-tumor drug according to claim 8, characterized in that: The anti-tumor drug is an anti-tumor drug that can regulate macrophages and transform them from M2 anti-inflammatory type to M1 pro-inflammatory type, or an anti-tumor drug that can both regulate macrophages and transform them from M2 anti-inflammatory type to M1 pro-inflammatory type and induce immunogenic cell death of tumor cells.

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