CN108047455B - Amphiphilic hyperbranched polymer for antigen carrier and preparation method and application thereof - Google Patents

Abstract

The present invention relates to an amphiphilic hyperbranched polymer for antigen carrier, a preparation method and application thereof. Aminopyridine and EDC hydrochloride catalyzed esterification to synthesize dodecyl trithiocarbonate-acrylate, and then the initiator AIBN, dodecyl trithiocarbonate-acrylate, polyethylene glycol mono The hyperbranched polymer 2 was synthesized by hyperbranching reaction of ether acetate and octadecyl acrylate at a molar ratio of 1:10:40:10. The hyperbranched polymer can stimulate the activation of antigen-presenting cells and can be used as an antigen carrier. Development of immunoadjuvant drugs to enhance immune response after cross-linking with polypeptides.

Description

Amphiphilic hyperbranched polymer for antigen carrier, preparation method and application thereof

technical field

The invention belongs to the field of biochemistry, relates to an amphiphilic hyperbranched polymer for antigen carrier, and also relates to a preparation method and application of the compound.

Background technique

Autoimmune diseases such as multiple sclerosis are diseases caused by excessive activation of autoreactive T cells to attack their own tissues. We believe that antigen-specific treatment means targeting and eliminating autoreactive T cells in the immune system by inducing immune tolerance. treatment would be a very good strategy. The use of soluble self-epitope-specific peptides to induce immune tolerance to treat autoimmune diseases has shown a certain therapeutic effect in animal models, but many clinical trials have failed. The reasons may include the short half-life of soluble peptides and the inability to effectively stimulate antigen presentation to induce immune tolerance; soluble peptides may cause potential safety issues such as anaphylactic shock in some animal models through intravenous administration. Therefore, the strategy of developing new forms of antigen carriers to attach antigen polypeptides to enhance the ability to induce immune tolerance has application prospects in drug development for the treatment of autoimmune diseases such as multiple sclerosis.

SUMMARY OF THE INVENTION

In view of this, one of the objects of the present invention is to provide an amphiphilic hyperbranched polymer for antigen carriers; the second object of the present invention is to provide a method for preparing the amphiphilic hyperbranched polymer; the object of the present invention The third is to provide a hyperbranched polymeric polypeptide based on an amphiphilic hyperbranched polymer; the fourth purpose of the present invention is to provide a method for synthesizing a hyperbranched polymeric polypeptide; the fifth purpose of the present invention is to provide the amphiphilic hyperbranched polymer The application of the compound in the preparation of an antigen carrier to improve the antigen presenting ability; the sixth purpose of the present invention is to provide the application of the hyperbranched polymeric polypeptide in the preparation of an immune adjuvant to enhance the immunogenicity of the medicine.

To achieve the above-mentioned purpose of the invention, the present invention provides the following technical solutions:

1. An amphiphilic hyperbranched polymer for antigen carriers, the amphiphilic hyperbranched polymer is first composed of dodecyl trithiocarbonate and acrylate in 4-dimethylaminopyridine and EDC hydrochloride; Catalytic esterification was used to synthesize dodecyl trithiocarbonate-acrylate, and then the initiator AIBN, dodecyl trithiocarbonate-acrylate, poly(ethylene glycol) methyl ether acrylate and The hyperbranched polymer was synthesized by hyperbranching reaction of octadecyl acrylate under the condition of molar ratio of 1:10:40:10, which was denoted as hyperbranched polymer 2.

Preferably, the esterification reaction conditions are to react under nitrogen protection for 20 minutes, then add dichloromethane, shake at 40°C for 48 hours, wash the dichloromethane in a separation funnel, collect the organic layer after drying, and remove the dichloromethane by rotary evaporation Methane, the final product is dried under vacuum and a yellow oil is recovered.

Preferably, the hyperbranching reaction is specifically as follows: the reaction is carried out under nitrogen protection for 20 minutes, and then the oil bath is stirred at 60° C. for 18 hours. After the reaction is completed, the reaction is cooled with cold water. In order to completely remove the unreacted monomers, the product is stirred in n-hexane for 6 hours. After 2 h, the solvent was replaced, and finally the hexane was poured out, and the final product was recovered by vacuum drying.

2. The preparation method of the amphiphilic hyperbranched polymer for antigen carrier, comprising the steps of:

(1) Synthesis of dodecyl trithiocarbonate-acrylate: dodecyl trithiocarbonate and acrylate are esterified under the catalysis of 4-dimethylaminopyridine and EDC hydrochloride to synthesize dodecyl Alkyl trithiocarbonate-acrylates;

(2) Synthesis of hyperbranched polymer 1: The molar ratio of initiator AIBN, dodecyl trithiocarbonate-acrylate, poly(ethylene glycol) methyl ether acrylate and octadecyl acrylate is 1: The hyperbranched polymer used for antigen carrier was synthesized by hyperbranching reaction under the condition of 10:40:10, which was denoted as hyperbranched polymer 2.

3. A hyperbranched polymeric polypeptide containing the amphiphilic hyperbranched polymer for antigen carriers, wherein the hyperbranched polymeric polypeptide is decomposed into polythiol by butylamine aminolysis by the hyperbranched polymer 2 under nitrogen protection, Then, under nitrogen protection, thiol alkene is gradually added to convert sulfhydryl to thioether and then polymerized with a polypeptide. The amino acid sequence of the polypeptide is shown in SEQ ID NO. 2.

4. The method for synthesizing the hyperbranched polymeric polypeptide comprises the following steps: adding the polypeptide with the amino acid sequence shown in SEQ ID NO.2 to the hyperbranched polymer 2 and butylamine in a mass ratio of 1:40:0.1 into water, then Packed into a dialysis bag with a molecular weight cut-off of 3500 Da, dialyzed and purified to obtain a hyperbranched polymeric polypeptide.

5. The application of the amphiphilic hyperbranched polymer for antigen carrier in the preparation of a medicament for antigen carrier to improve antigen presenting ability.

6. The application of the hyperbranched polymeric polypeptide in the preparation of a drug for enhancing immunogenicity of an immune adjuvant.

The beneficial effects of the present invention are as follows: the present invention discloses an amphiphilic hyperbranched polymer, and by controlling the addition amount of octadecyl acrylate in the synthesis of the hyperbranched polymer, a hyperbranched polymer with good hydrophilicity is obtained, and the obtained hyperbranched polymer It can stimulate the activation of antigen-presenting cells (including BMDC cells, macrophages) and enhance immunogenicity, so its hyperbranched polymer can be used as an antigen carrier, and can activate mouse T cells after cross-linking with OVA polypeptide, indicating that the immunogen Therefore, the hyperbranched polymeric polypeptides HB-OVA-1 and HB-OVA-2 have potential applications as immune adjuvants.

Description of drawings

In order to make the purpose, technical solutions and beneficial effects of the present invention clearer, the present invention provides the following drawings for description:

Figure 1 shows the synthetic route of amphiphilic hyperbranched polymer and polypeptide cross-linking.

Figure 2 shows the ¹ H NMR spectrum analysis of the structures of each compound (a) DMATC, (b) DMATC-acrylate, (c) hyperbranched polymer 1, (d) hyperbranched polymer 2, (e) hyperbranched polymer 1-acrylate , (f) hyperbranched polymer 2-acrylate.

Figure 3 is a water contact angle analysis of hydrophobicity (a: hyperbranched polymer 1; b: hyperbranched polymer 2; c: PEG control, n=3).

Figure 4 shows the uptake of hyperbranched polymer polypeptides HB-MOG-1 and HB-MOG-2 in bone marrow-derived antigen-presenting cells (rhodamine-labeled HB-MOG-1 and HB- MOG-2 stimulated BMDC cells for 3 hours, (a) flow analysis of the fluorescence intensity of rhodamine phagocytosis by cells; (b) quantitative analysis of rhodamine fluorescence intensity; (c) confocal microscopy analysis of rhodamine fluorescence in cells after stimulation for 3 hours distribution in ).

Figure 5 shows the activation of bone marrow-derived antigen-presenting cells stimulated by hyperbranched polymer polypeptides HB-MOG-1 and HB-MOG-2 ((a): from left to right: PBS, soluble MOG polypeptide, HB-MOG-1 , HB-MOG-2 stimulated BMDC cells, and flow cytometry analysis of the proportion of cells co-expressing antigen presentation-related molecule MHCII and costimulatory molecule CD86; (b): Comparison of the percentage of cells double positive for MHCII and costimulatory molecule CD86.

Figure 6 shows the results of the activation of OVA antigen-specific DO11.10 mouse T cells induced by antigen presentation of hyperbranched polymer polypeptides HB-OVA-1 and HB-OVA-2 (from left to right, OVA polypeptide, HB-OVA- 1. The expression of activation molecules CD44 and CD69 in DO11.10 mouse spleen-derived naive T cells stimulated by HB-OVA-2; a, c: Flow cytometry analysis of CD4+CD44+ T cells after stimulation of DO11.10 mouse T cells frequency; b, d: flow analysis of the frequency of CD4+CD69+ T cells after T cell stimulation in DO11.10 mice).

Figure 7 shows the therapeutic effects of hyperbranched polymers HB-MOG-1 and HB-MOG-2 on EAE disease in mice ((a) EAE mice were injected with 40ug MOG-containing MOG polypeptide, HB on the 15th, 17th and 19th day of the onset of the disease by tail vein -MOG-1, HB-MOG-2 or placebo PBS, the disease course of mice was scored; (b) HE staining results of mouse spinal cord tissue showed that: compared with the placebo group, HB-MOG-1 and HB- Spinal cord infiltrating inflammatory cells decreased and tissue vacuoles decreased after MOG-2 administration, while inflammatory cells infiltrated in MOG polypeptide treatment group.

Detailed ways

The preferred embodiments of the present invention will be described in detail below with reference to the accompanying drawings.

The materials used in the following examples of the present invention are as follows: 2-hydroxyethyl acrylate (97%, Alfa Aesar), poly(ethylene glycol) methyl ether acrylate (480Da), poly(ethylene glycol) ester (258da), Octadecyl acrylate (ODA, 97%, ) and dichloromethane (DCM) were prepared by pre-drying activated 3A molecular sieves for 72 h. Azobisisobutyronitrile (AIBN) was purified by recrystallization in methanol. MOG polypeptide (amino acid sequence: CMEVGWYRSPFSRVVHLYRNGK (SEQ ID NO. 1)) and OVA polypeptide (amino acid sequence: CISQAVHAAHAEINEAGR (SEQ ID NO. 2)) were synthesized by China Peptide Biochemical Co., Ltd.

The present invention develops a functional amphiphilic hyperbranched polymer, and the hydrophobicity of the polymer is adjusted by controlling the ratio of the hydrophobic octadecyl acrylate side chains with different grafting degrees to obtain the amphiphilic hyperbranched polymer 1 and two Affinity hyperbranched polymer 2. The two polymers are attached with MOG polypeptide and OVA polypeptide respectively. Two kinds of polymer cross-linked MOG polypeptides with different hydrophobicity are mainly used in the development and research of drug therapy for multiple sclerosis. Two polymers with different hydrophobicity cross-linked OVA polypeptides are mainly used in basic research on polymers as potential immune adjuvants. The experimental results show that both amphiphilic hyperbranched polymer 1 and amphiphilic hyperbranched polymer 2 can effectively promote the phagocytosis and presentation of antigenic peptides by antigen-presenting cells BMDC, and can effectively activate naive T lymphocytes and induce activated T lymphocytes. Apoptosis of antigen-specific pathogenic T lymphocytes. And its ability to induce antigen presentation, activation and apoptosis were significantly higher than soluble free peptides. Two MOG polypeptide-loaded polymers showed significant therapeutic effects on experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, and promoted motor function in hindlimb paralyzed mice after one treatment cycle effective recovery. In contrast, equivalent doses of soluble free peptides had limited therapeutic effects on the EAE model. Therefore, our experimental results show that the functional amphiphilic hyperbranched polymers designed in this study can be used as effective antigen polypeptide carriers, greatly improve the antigen presentation ability, and induce antigen-specific pathogenic T T in vitro and in vivo. The pathway of cell apoptosis induces immune tolerance and then treats autoimmune diseases. The two hyperbranched polymers with different hydrophobicity designed in this study can be used as antigen carriers in the development of antigen/polypeptide drugs for the treatment of autoimmune diseases by inducing immune tolerance.

Embodiment 1, the synthetic method of superculture polymer

The synthetic method of hyperbranched polymer, synthetic route as shown in Figure 1, comprises the steps:

(1) Synthesis of S-Dodecyl-S'-(a,a'-dimethyl-a"-acetic acid)trithiocarbonate (DMATC): see the synthesis method using novel carbonate carboxyl end-capping as three efficient RAFT agents. polymers (Macromolecules, 2002, 35(18), pp 6754-6756).

(2) Synthesis of DMATC-acrylate: DMATC chain transfer agent and acrylate cross-linking through the esterification of DMATC and hydroxyethyl acrylate catalyzed by DMAP/EDC, the concrete method is as follows: add 1.46g DMATC ( dodecyl trithiocarbonate), 1.15 g EDC hydrochloride (C ₈ H ₁₇ N ₃ ·HCl; 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride) , 0.073g DMAP (4-dimethylaminopyridine), the flask was pre-dried in vacuum at room temperature (18-25°C) overnight; then 1.4g of hydroxyethyl acrylate was added, the flask was protected by nitrogen, reacted for 20min, and then 15mL of anhydrous was added with a syringe Dichloromethane (DCM), the solution was shaken at 40°C for 48h, the DCM solution was washed with brine 6 times in a separating funnel, the organic layer was dried over anhydrous sodium sulfate and collected, DCM was removed by rotary evaporation, and the final product was dried under vacuum, And recovered as yellow oil to obtain DMATC-acrylate.

(3) Synthesis of hyperbranched polymers (PEG-r-PODA-r-PTTC polytrithiocarbonates):

Method 1: The molar ratio of initiator AIBN (azobisisobutyronitrile): DMATC-acrylate: poly(ethylene glycol) methyl ether acrylate: ODA (octadecyl acrylate) is 1:10:40:20 The reaction is specifically adding 0.011g AIBN, 0.3g RAFT-acrylate, 1.25g poly(ethylene glycol) methyl ether acrylate, 0.422g ODA and 2ml toluene into a 10ml round-bottomed flask and sealing with a rubber septum; the reaction is protected by nitrogen 20min, then stirred in the oil bath at 60°C for 18h, after the reaction was completed, cooled with cold water, in order to completely remove unreacted monomers, the product was stirred in n-hexane for 6h, the solvent was changed every 2h, and finally the hexane was poured out, the final product was The hyperbranched polymer 1 was obtained by vacuum drying and recovery.

Method 2: The molar ratio of the initiator AIBN:DMATC-acrylate:poly(ethylene glycol) methyl ether acrylate:ODA is 1:10:40:10, 0.011g AIBN, 0.3g RAFT-acrylate, 1.25 g poly(ethylene glycol) methyl ether acrylate, 0.211g ODA and 2ml toluene were added to a 10ml round-bottomed flask and sealed with a rubber septum; the reaction was carried out under nitrogen protection for 20min, and then the oil bath was stirred at 60°C for 18h. After the reaction was completed, cooled with cold water , in order to completely remove unreacted monomers, the product was stirred in n-hexane for 6 h, the solvent was replaced every 2 h, and finally the hexane was poured out, and the final product was recovered by vacuum drying to obtain hyperbranched polymer 2.

In this step, the amphiphilic hyperbranched polymer is obtained by adjusting the feed ratio of the hydrophilic monomer poly(ethylene glycol) methyl ether acrylate and the lipophilic ODA, and the obtained hyperbranched polymer is subjected to ¹ H NMR spectroscopy to analyze each compound The hydrophobicity was analyzed by structure and water contact angle, and the results are shown in Figures 2 and 3. The results showed that the water contact angle of hyperbranched polymer 1 was higher than that of hyperbranched polymer 2, indicating that hyperbranched polymer 1 was more lipophilic and hyperbranched polymer 2 was more hydrophilic.

Embodiment 2, hyperbranched polytrithiocarbonate modification

The obtained hyperbranched polytrithiocarbonate was further decomposed into polythiol by butylamine aminolysis reaction under nitrogen protection. In order to prevent the crosslinking of hyperbranched polythiols through thiol oxidation, the thiol groups were converted into thioethers by gradually adding thiol alkenes under nitrogen protection. The specific method is as follows: 0.6 g of hyperbranched polythiocarbonate was dissolved in 1.5 ml DCM (dichloromethane) followed by addition of 0.5 ml butylamine (~20 times), under nitrogen protection. After 30 min, the yellow solution faded to colorless or pale yellow, then nitrogen was bubbled into the solution to remove most of the DCM and butylamine. The product was precipitated by transferring nitrogen degassed hexanes (-10 mL) into the solution. After 15 minutes, a colorless or pale yellow product adhered to the bottom of the bottle, and most of the solvent was removed through a cannula under nitrogen protection. Next, 2 ml of degassed DCM was injected into the reaction to redissolve the resulting polythiol, followed by 0.57 ml of PEGDA (0.63 g, 258 Da) and 0.01 ml of butylamine. After 4 h, the reaction was stopped, the product was precipitated to remove hexane, and the acrylate-containing hyperbranched polysulfide was obtained by vacuum drying.

Example 3. Preparation of hyperbranched polysulfide acrylate cross-linked polypeptide

Both MOG and OVA polypeptides have cysteine at the N-terminus, 80 mg of hyperbranched polymer, 2 mg of peptide (OVA), 0.2 mg of butylamine mixed with 5 ml of water and stirred overnight, and then the solution was put into a dialysis bag (molecular weight cut-off: 3500Da) Dialyzed in deionized water for 2 days, and finally, the product was purified by freeze-drying to obtain hyperbranched polymeric polypeptide 3 (Hyper-branch polymerconjugated MOG-1, HB-OVA-1) and hyperbranched polymeric polypeptide 4 (Hyper-branch polymer conjugatedMOG-1). 2, HB-OVA-2).

According to the same method, the difference is that the MOG polypeptide is used to obtain hyperbranched polymeric polypeptide 1 (Hyper-branch polymer conjugated MOG-1, HB-MOG-1) and hyperbranched polymeric polypeptide 2 (Hyper-branch polymerconjugated MOG-2, HB- MOG-2).

Example 4, _Rhodamine B fluorescent agent labeling

Rhodamine B fluorescer labels hyperbranched polymeric MOG polypeptides by amidation of free amines at lysine or arginine sites. 100 mg of hyperbranched polymer polypeptide 1, 2 mg of rhodamine B, NHS and EDC were stirred in water at room temperature for 24 hours in the dark. After the reaction, the solution was put into a dialysis bag (molecular weight cut-off: 3500 Da) and dialyzed in deionized water for 2 days. Finally, the product was purified by freeze drying.

In the same way, hyperbranched polymeric polypeptide 2, hyperbranched polymeric polypeptide 3 and hyperbranched polymeric polypeptide 4 were labeled with Rhodamine B fluorescent agent.

Example 5. Evaluation of therapeutic effect

(1) Preparation of multiple sclerosis mouse model EAE

Mouse EAE is an autoimmune disease model mainly mediated by specifically sensitized CD4+ T cells and characterized by central nervous system mononuclear cell infiltration and demyelination. It is the most classic model of human multiple sclerosis. The experimental animal model, the preparation method is:

a. Preparation of MOG/CFA emulsifier: add 100 mg of tuberculin (Mtb) to 25 ml of Freund's complete adjuvant and mix well (the final concentration of Mtb is 5 mg/ml); the MOG35-55 polypeptide is dissolved in PBS to 2 mg/ml; 1: 1 Mix MOG and CFA/Mtb (the final concentration of MOG is 1 mg/ml); use an ultrasonic homogenizer to emulsify at a frequency (output control 7% duty cycle 50), and operate on ice; after complete emulsification, invert the tube, the emulsion will not Free flow, stored in a refrigerator at 4°C and used within 1 week.

b. Mouse immunization: Prepare 10-week-old C57BL/6 female mice, inject 100 μl MOG/CFA emulsifier subcutaneously at two points on the back of the mouse on day 0, and intraperitoneally inject 100 μl pertussis toxin PTX (2ug/ml) 5 hours later; Day 100 μl PTX was injected intraperitoneally per mouse.

c. The mice were scored on the 10th day after immunization. Double-blind scoring was used in this experiment, that is, all the EAE mice used in the experiment were scored by other researchers who were not involved in the drug treatment of the subject. Scoring was performed on a 5-point scale (Hooke Lab): 1, tail drooping weakness; 2, tail paralysis; 2.5, unilateral hindlimb weakness; 3, unilateral hindlimb paralysis; 3.5, bilateral hindlimb paralysis; 4, complete paralysis of both hindlimbs and front limbs Weakness; mice over 4 points were euthanized.

(2) Evaluation of the therapeutic effect of hyperbranched polymeric MOG polypeptide-1 and hyperbranched polymeric MOG polypeptide-2 on EAE in mice

The peak onset of EAE mice is on the 15th, 17th, and 19th days after immunization, respectively, by tail vein injection of 40 μg (normalized to the mass of MOG polypeptide) hyperbranched polymerized MOG polypeptide-1, hyperbranched polymerized MOG polypeptide-2 or free MOG polypeptide, PBS placebo as a control.

The results showed that the hyperbranched polymer HB-MOG-1 and HB-MOG-2 polypeptides could be rapidly taken up by bone marrow-derived antigen-presenting cells (Fig. 4). Hyperbranched polymers HB-MOG-1 and HB-MOG-2 stimulated the activation of bone marrow-derived antigen-presenting cells (Fig. 5). Hyperbranched polymers HB-MOG-1 and HB-MOG-2 have significant therapeutic effects on EAE disease in mice (Figure 7). The above results show that HB-MOG-1 and HB-MOG-2 can stimulate antigen-presenting cells to present polypeptide antigens and induce apoptosis of activated MOG antigen-specific 2D2 mouse T lymphocytes, suggesting that they may be used for immune tolerance. Research. Treatment experiments with an EAE mouse model of human multiple sclerosis showed that both HB-MOG-1 and HB-MOG-2 could significantly improve the severity of EAE mice, and two intravenous treatments resulted in less hind limb paralysis. The rat regained the ability to walk. The results suggest that HB-MOG-1 and HB-MOG-2 may be used in the development of therapeutic drugs for human multiple sclerosis.

(3) Activation of mouse T cells by hyperbranched polymeric OVA polypeptide-1 and hyperbranched polymeric OVA polypeptide-2

Take the spleen cells of OVA antigen TCR transgenic mice, add 20 μg (standardized to the mass of OVA polypeptide) hyperbranched polymerized OVA polypeptide-1, hyperbranched polymerized OVA polypeptide-2, PBS placebo as a control, and then detect the spleen-derived naive T cells. Expression of activating molecules CD44 and CD69. The results are shown in Figure 6. The results showed that the hyperbranched polymers HB-OVA-1 and HB-OVA-2 could induce the activation of OVA antigen-specific DO11.10 mouse T cells through antigen presentation, suggesting their strong immunogenicity, suggesting that the hyperbranched polymers HB -OVA-1 and HB-OVA-2 have potential applications as immune adjuvants.

Finally, it should be noted that the above preferred embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail through the above preferred embodiments, those skilled in the art should Various changes may be made in details without departing from the scope of the invention as defined by the claims.

sequence listing

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Claims (5)

1. Hyperbranched polymeric polypeptide of an amphiphilic hyperbranched polymer for an antigen carrier, characterized in that: the hyperbranched polymeric polypeptide is obtained by decomposing hyperbranched polymer 2 into polythiol through aminolysis by butylamine under the protection of nitrogen, gradually adding thiol alkene under the protection of nitrogen to convert sulfydryl into thioether and polymerizing the thioether and polypeptide, wherein the amino acid sequence of the polypeptide is shown as SEQ ID NO. 2;

the amphiphilic hyperbranched polymer is prepared by firstly carrying out esterification reaction on dodecyl trithiocarbonate and acrylate under the catalysis of 4-dimethylaminopyridine and EDC hydrochloride to synthesize dodecyl trithiocarbonate-acrylate, and then carrying out esterification reaction on an initiator AIBN, the dodecyl trithiocarbonate-acrylate, poly (ethylene glycol) methyl ether acrylate and octadecyl acrylate in a molar ratio of 1: 10: 40: and (3) synthesizing a hyperbranched polymer by virtue of a hyperbranched reaction under the condition of 10, and marking as a hyperbranched polymer 2.

2. The hyperbranched polymeric polypeptide of claim 1, wherein the esterification reaction is carried out under the protection of nitrogen for 20min, followed by adding dichloromethane, shaking at 40 ℃ for 48h, washing dichloromethane in a separating funnel, drying the organic layer, collecting, removing dichloromethane by rotary evaporation, drying the final product under vacuum, and recovering yellow oil.

3. The hyperbranched polymeric polypeptide of claim 1, wherein the hyperbranched reaction is in particular: introducing nitrogen for protection reaction for 20min, stirring at 60 ℃ and carrying out oil bath for 18h, cooling with cold water after the reaction is finished, stirring the product in n-hexane for 6h for completely removing unreacted monomers, replacing the solvent every 2h, finally pouring out hexane, and drying and recovering the final product in vacuum to obtain the product.

4. The method for synthesizing the hyperbranched polymeric polypeptide of any one of claims 1-3, comprising the steps of:

(1) synthesis of dodecyl trithiocarbonate-acrylate: carrying out esterification reaction on dodecyl trithiocarbonate and acrylate under the catalysis of 4-dimethylaminopyridine and EDC hydrochloride to synthesize dodecyl trithiocarbonate-acrylate;

(2) synthesis of hyperbranched polymer 2: the initiator AIBN, dodecyl trithiocarbonate-acrylate, poly (ethylene glycol) methyl ether acrylate and stearyl acrylate are mixed in a molar ratio of 1: 10: 40: synthesizing a hyperbranched polymer for the antigen carrier by virtue of a hyperbranched reaction under the condition of 10, and marking as a hyperbranched polymer 2;

(3) adding the polypeptide with the amino acid sequence shown as SEQ ID NO.2, the hyperbranched polymer 2 and butylamine into water according to the mass ratio of 1:40:0.1, then filling into a dialysis bag with the molecular weight cutoff of 3500Da, dialyzing, and purifying to obtain the hyperbranched polymeric polypeptide.

5. Use of the hyperbranched polymeric polypeptide of claim 1 for the preparation of a medicament for enhancing the immunogenicity of an immunoadjuvant.

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