Drug-loaded polymer vesicle with asymmetric membrane structure, preparation method and application in preparation of drug for treating acute myeloid leukemia

Abstract

The invention discloses a drug-loaded polymer vesicle with an asymmetric membrane structure, a preparation method and application in preparation of a drug for treating acute myeloid leukemia. An amphiphilic triblock polymer with polyaspartic acid PAsp, a targeted amphiphilic block polymer and a small molecule drug are assembled together to prepare the targeted small molecule drug-loaded polymer vesicle with the asymmetric membrane structure. The drug-loaded polymer vesicle disclosed by the invention has many unique advantages, including small size, simple and controllable preparation, reversible crosslinking, in-vivo stability, targeted delivery, high intracellular drug enrichment concentration, sensitive reduction, efficient killing of tumor cells, significant tumor growth inhibition effect and the like, and especially the drug-loaded vesicle disclosed by the invention has effective inhibition on both acute myeloid leukemia cell lines and patient cells. Therefore, the polymer vesicleis expected to become a simple and multifunctional nano platform for efficient and specific targeted delivery of drugs to tumor cells.

Description

technical field

[0001] The invention belongs to the technical field of polymer nano-medicines, and in particular relates to reversible cross-linking and degradable polymer vesicles loaded with small molecule drugs, a preparation method thereof, and an application in targeted therapy of acute myeloid leukemia. Background technique

[0002] Acute myeloid leukemia is a common blood disease, accounting for about 1 / 3 of all leukemias, and its typical feature is abnormal proliferation and differentiation of myeloid cells. If not detected and treated in time, acute myeloid leukemia can induce severe symptoms of acute bone marrow failure, leading to death within weeks or months. In 2018, there were 437,033 new cases and 309,006 deaths worldwide. Therefore, the treatment of leukemia is facing a very severe situation. In the past 30 years, the standard treatment for acute myeloid leukemia has not made great progress. It has always been a combination treatment of cytarabine and dauno...

Examples

Embodiment 1

[0065] Example 1 Synthesis of amphiphilic triblock polymer

[0066] Firstly, amphiphilic block polymers PEG-P(TMC-DTC), PEG-P(CL-DTC) and PEG-P(LA-DTC) were synthesized by ring-opening polymerization. Then, after the terminal hydroxyl groups of the above three amphiphilic block polymers were activated by p-nitrophenyl chloroformate (p-NPC), the amphiphilic triblock polymer was prepared by reacting with PAsp. Specifically, taking the synthesis of PEG-P(TMC-DTC)-PAsp as an example, the synthesis route is as follows:

[0067]

[0068] Wherein, in step (i), the reaction conditions are anhydrous dichloromethane (DCM), pyridine, 25 ºC, 24 hours; in step (ii), the reaction conditions are anhydrous dimethyl sulfoxide (DMSO), PAsp , triethylamine, 30 ºC, 48 hours.

[0069] Concrete synthetic steps are as follows:

[0070]The synthesis of PEG-P(TMC-DTC)-PAsp is divided into two steps, that is, after the terminal hydroxyl group of PEG-P(TMC-DTC) (5.0-(15.0-2.0) kg / mol) is activated...

Embodiment 2

[0071] Example 2 Synthesis of targeted amphiphilic block polymers

[0072] The preparation of targeting amphiphilic block polymer is divided into two steps. Firstly, functionalized amphiphilic block polymers with Mal functional groups and NHS functional groups were synthesized, and then targeted amphiphilic block polymers were obtained by reacting targeting polypeptides with functionalized amphiphilic block polymers. Specifically, take A6-PEG-P (TMC-DTC) as an example. First, Mal-PEG-P(TMC-DTC) (7.5-(14.9-2.1) kg / mol) was synthesized by ring-opening polymerization, and then the Michael addition reaction of the sulfhydryl group of A6 with Mal-PEG-P(TMC-DTC) A6-PEG-P(TMC-DTC) was obtained. Under nitrogen atmosphere, 1 mL of Mal-PEG-P(TMC-DTC) (100 mg, 4.1 µmol) in anhydrous DMSO was added dropwise to 2 mL of A6 (7.47 mg, 8.2 µmol) solution at room temperature for 48 hours. After the reaction, the reaction solution was first dialyzed with DMSO for 36 hours (replace the medium...

Embodiment 3

[0074] Example 3 Preparation of non-targeted drug-loaded polymersomes

[0075] The non-targeted drug-loaded polymersomes were prepared by a solvent displacement method, and encapsulated by the electrostatic interaction between the drug and the PAsp segment in the amphiphilic triblock polymer. Specifically, the amphiphilic triblock polymer is PEG-P(TMC-DTC)-PAsp as an example. Dissolve PEG-P(TMC-DTC)-PAsp in DMSO (40 mg / mL), and inject 100 µL into 900 µL HEPES (pH 6.8, 10 mM) containing small molecule drugs, at 300 rpm After stirring for 3 minutes, the mixture was incubated at 37°C for 8 hours. The non-targeted drug-loaded polymersome cPS-VCR was obtained by dialysis with HEPES (pH 7.4, 10 mM) for 8 hours.

[0076] Among them, the theoretical drug loading of the small molecule drug VCR was set at 4.8-9.1 wt.%, and the study found that the particle size of the obtained cPS-VCR was about 30 nm, and the particle size distribution was about 0.1 (Table 1). The drug-loading capaci...

technical field

[0001] The invention belongs to the technical field of polymer nano-medicines, and in particular relates to reversible cross-linking and degradable polymer vesicles loaded with small molecule drugs, a preparation method thereof, and an application in targeted therapy of acute myeloid leukemia. Background technique

[0002] Acute myeloid leukemia is a common blood disease, accounting for about 1 / 3 of all leukemias, and its typical feature is abnormal proliferation and differentiation of myeloid cells. If not detected and treated in time, acute myeloid leukemia can induce severe symptoms of acute bone marrow failure, leading to death within weeks or months. In 2018, there were 437,033 new cases and 309,006 deaths worldwide. Therefore, the treatment of leukemia is facing a very severe situation. In the past 30 years, the standard treatment for acute myeloid leukemia has not made great progress. It has always been a combination treatment of cytarabine and dauno...

Examples

Embodiment 1

[0065] Example 1 Synthesis of amphiphilic triblock polymer

[0066] Firstly, amphiphilic block polymers PEG-P(TMC-DTC), PEG-P(CL-DTC) and PEG-P(LA-DTC) were synthesized by ring-opening polymerization. Then, after the terminal hydroxyl groups of the above three amphiphilic block polymers were activated by p-nitrophenyl chloroformate (p-NPC), the amphiphilic triblock polymer was prepared by reacting with PAsp. Specifically, taking the synthesis of PEG-P(TMC-DTC)-PAsp as an example, the synthesis route is as follows:

[0067]

[0068] Wherein, in step (i), the reaction conditions are anhydrous dichloromethane (DCM), pyridine, 25 ºC, 24 hours; in step (ii), the reaction conditions are anhydrous dimethyl sulfoxide (DMSO), PAsp , triethylamine, 30 ºC, 48 hours.

[0069] Concrete synthetic steps are as follows:

[0070]The synthesis of PEG-P(TMC-DTC)-PAsp is divided into two steps, that is, after the terminal hydroxyl group of PEG-P(TMC-DTC) (5.0-(15.0-2.0) kg / mol) is activated...

Embodiment 2

[0071] Example 2 Synthesis of targeted amphiphilic block polymers

[0072] The preparation of targeting amphiphilic block polymer is divided into two steps. Firstly, functionalized amphiphilic block polymers with Mal functional groups and NHS functional groups were synthesized, and then targeted amphiphilic block polymers were obtained by reacting targeting polypeptides with functionalized amphiphilic block polymers. Specifically, take A6-PEG-P (TMC-DTC) as an example. First, Mal-PEG-P(TMC-DTC) (7.5-(14.9-2.1) kg / mol) was synthesized by ring-opening polymerization, and then the Michael addition reaction of the sulfhydryl group of A6 with Mal-PEG-P(TMC-DTC) A6-PEG-P(TMC-DTC) was obtained. Under nitrogen atmosphere, 1 mL of Mal-PEG-P(TMC-DTC) (100 mg, 4.1 µmol) in anhydrous DMSO was added dropwise to 2 mL of A6 (7.47 mg, 8.2 µmol) solution at room temperature for 48 hours. After the reaction, the reaction solution was first dialyzed with DMSO for 36 hours (replace the medium...

Embodiment 3

[0074] Example 3 Preparation of non-targeted drug-loaded polymersomes

[0075] The non-targeted drug-loaded polymersomes were prepared by a solvent displacement method, and encapsulated by the electrostatic interaction between the drug and the PAsp segment in the amphiphilic triblock polymer. Specifically, the amphiphilic triblock polymer is PEG-P(TMC-DTC)-PAsp as an example. Dissolve PEG-P(TMC-DTC)-PAsp in DMSO (40 mg / mL), and inject 100 µL into 900 µL HEPES (pH 6.8, 10 mM) containing small molecule drugs, at 300 rpm After stirring for 3 minutes, the mixture was incubated at 37°C for 8 hours. The non-targeted drug-loaded polymersome cPS-VCR was obtained by dialysis with HEPES (pH 7.4, 10 mM) for 8 hours.

[0076] Among them, the theoretical drug loading of the small molecule drug VCR was set at 4.8-9.1 wt.%, and the study found that the particle size of the obtained cPS-VCR was about 30 nm, and the particle size distribution was about 0.1 (Table 1). The drug-loading capaci...

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