CN107714646A - Amphipathic nature polyalcohol micella of tumor extracellular matrix and preparation method thereof can be penetrated - Google Patents

Abstract

The invention relates to an amphiphilic polymer micelle capable of penetrating tumor extracellular matrix: comprising antitumor drugs, amphiphilic polymers and collagenase, the amphiphilic polymer wraps the antitumor drugs in Internally, the amphiphilic polymer is externally linked to the collagenase by a covalent bond. The present invention also provides a preparation method thereof: react the amphiphilic polymer and collagenase under the action of a cross-linking agent in a solvent at 20-26°C, and remove the first organic solvent; The hydrophilic polymer and the anti-tumor drug are dissolved in the second organic solvent to obtain a drug solution; the drug solution is added to pure water to remove the second organic solvent to form amphiphilic polymer micelles that can penetrate the tumor extracellular matrix . The collagenase on the surface of the micelles of the present invention can penetrate the extracellular matrix, so that the antitumor drugs can better enter the tumor site, increase the amount of drugs in the tumor cells, and thus cause the apoptosis of the tumor cells.

Description

Amphiphilic polymer micelles capable of penetrating tumor extracellular matrix and preparation method thereof

technical field

The invention relates to the field of pharmaceutical preparations, in particular to an amphiphilic polymer micelle capable of penetrating tumor extracellular matrix and a preparation method thereof.

Background technique

In recent years, the incidence of cancer is increasing year by year, seriously threatening human life and health. Although the methods and technologies of tumor treatment have been continuously improved, and the survival rate of patients has been greatly improved, most of the patients are already in the advanced stage when the disease occurs, and they cannot receive surgical treatment. They can only be treated conservatively with anticancer drugs. Chemical drugs usually have poor water solubility, obvious side effects such as cardiotoxicity and bone marrow suppression, and direct application will cause relatively large damage to normal cells of patients.

The micelles with the PEG hydrophilic block can avoid the non-specific recognition and phagocytosis of the drug-loaded system in the reticuloendothelial system, and then enter and transport the drug to the liver, spleen and other organs. Many patents and literature reports have shown that modifying the targeting molecule at the end of PEG can make the drug targeted. The biofunctionalization of PLC-PEG-B nanoparticles with transferrin as a targeting factor can significantly improve the specific targeting ability of PLA-PEG nanoparticles in brain tumors. Although the targeting carrier has strong tumor-specific binding ability, it has not fundamentally solved a series of barriers in the tumor microenvironment.

The extracellular matrix (ECM) is a three-dimensional macromolecular network composed of collagen, fibronectin, glycoproteins, proteoglycans, and glycosaminoglycans. During tumorigenesis, the ECM undergoes significant changes, leading to increased formation of fibrotic matrix, increased stiffness, excessive deposition of ECM components, and abnormal ECM remodeling, so ECM components can significantly affect drug delivery.

After chemical drugs are made into nanoparticles, the curative effect can be improved and the side effects can be reduced. Currently, a major challenge in applying nanomedicines to treat cancer is their difficulty in penetrating tumor extracellular matrix.

Contents of the invention

In order to solve the above-mentioned technical problems, the object of the present invention is to provide a kind of amphiphilic polymer micelle that can penetrate tumor extracellular matrix (ECM). The ability to penetrate and increase the entry of chemical drugs into tumor cells, thereby causing apoptosis of tumor cells.

The present invention provides an amphiphilic polymer micelle capable of penetrating tumor extracellular matrix (ECM): it includes antitumor drugs, amphiphilic polymers and collagenase, and the amphiphilic polymer binds the antitumor drugs Wrapped inside, the amphiphilic polymer exterior is covalently linked to collagenase.

Further, the antineoplastic drug is one or more of docetaxel, doxorubicin and paclitaxel.

Further, the amphiphilic polymer is one or more of PCL-PEG copolymer, PLA-PEG copolymer and PLGA-PEG copolymer. Specifically, the amphiphilic polymer is PCL-PEG-COOH or PLA-PEG-COOH, PLGA-PEG-COOH. Wherein, PCL is polycaprolactone, PEG is polyethylene glycol, PLA is polylactic acid, and PLGA is polylactic acid-glycolic acid copolymer.

Further, the molar ratio of the collagenase to the amphiphilic polymer is 1:100-1000. Preferably, the molar ratio of collagenase to amphiphilic polymer is 1:100.

Further, the mass ratio of the antitumor drug to the amphiphilic polymer is 0.5-15:100.

Further, the molecular weight of the amphiphilic polymer is 10000-25000 g/mol.

Specifically, in PCL-PEG-COOH, the molecular weight of PCL is 8000g/mol, and the molecular weight of PEG is 2000g/mol; in PLA-PEG-COOH, the molecular weight of PLA is 8000g/mol, and the molecular weight of PEG is 2000g/mol; PLGA -In PEG-COOH, the molecular weight of PLGA is 20000 g/mol, and the molecular weight of PEG is 2000 g/mol.

Further, the collagenase is one or more of type I collagenase, type II collagenase, type IV collagenase and type V collagenase. Preferably, the collagenase is type IV collagenase, and type IV collagenase mainly degrades ECM such as type IV collagen fiber laminin, elastin and nestin, and can digest denatured collagen.

The present invention also provides a method for preparing the above-mentioned amphiphilic polymer micelles that can penetrate tumor extracellular matrix, including the following steps:

(1) React the amphiphilic polymer and the cross-linking agent in the first organic solvent at 20-26°C (room temperature), remove the first organic solvent, and then react with collagenase in pure water at 20-26 Reaction at ℃ (room temperature) to obtain an amphiphilic polymer modified with collagenase;

(2) dissolving the amphiphilic polymer modified with collagenase and the antineoplastic drug in a second organic solvent to obtain a drug solution;

(3) Add the drug solution into pure water, remove the second organic solvent, and form amphiphilic polymer micelles that can penetrate the tumor extracellular matrix.

Further, in step (1), the carbodiimide method is used for the reaction, and the crosslinking agent is N-hydroxysuccinimide (NHS) and 1-ethyl-(3-dimethylaminopropyl) carbon Diimine hydrochloride (EDC).

Further, in step (1), the first organic solvent is removed by rotary evaporation.

Further, in step (1), the first organic solvent is one or more of dimethyl sulfoxide, acetonitrile and chloroform.

Further, in step (2), the second organic solvent is one or more of acetone, methanol, ethanol, dichloromethane and chloroform.

Further, in step (3), the drug solution is added into the pure water under stirring condition, the stirring speed is 400-700rpm, and the stirring time is 4-8h.

In step (3), since the amphiphilic polymer contains hydrophilic and hydrophobic segments, after the drug solution is added to pure water, the hydrophilic ends of the polymer will gather on the outside, and the hydrophobic segments will gather on the inside. The drug is also hydrophobic, so it is encapsulated inside, and after removal of the second organic solvent, an amphiphilic polymer micelle can be formed.

By means of the above solution, the present invention has at least the following advantages:

(1) After the collagenase is modified on the surface of the micelles, the collagen-specific degradation of the extracellular matrix of the tumor can be effectively increased by the collagenase, which can effectively increase the therapeutic effect of the chemical drug delivery into the tumor, and can increase the lethality of the chemotherapeutic drug on tumor cells .

(3) The chemotherapeutic drugs are encapsulated in the micelle core, delivered to the tumor cells, and play a role in killing tumor cells. Simultaneously, the micelle of the present invention has high stability and good physiological compatibility, can increase the stability of chemical medicines, can make the medicines release stably and slowly, improve the bioavailability of medicines, and reduce the toxic and side effects of the body.

(4) The preparation process of the present invention is simple, the encapsulation rate is high, and it is convenient for industrialized production; the physical properties of the prepared amphiphilic polymer micelles with the effect of increasing the penetration of tumor extracellular matrix meet the needs of tumor cell therapy. .

The above description is only an overview of the technical solutions of the present invention. In order to understand the technical means of the present invention more clearly and implement them according to the contents of the description, the preferred embodiments of the present invention and accompanying drawings are described in detail below.

Description of drawings

Fig. 1 is the transmission electron microscope test result of the micelle prepared by the embodiment of the present invention 1, 2, 6 and 7;

Fig. 2 is the in vitro release result of the micelles prepared in Example 4.

detailed description

The specific implementation manners of the present invention will be further described in detail below in conjunction with the accompanying drawings and embodiments. The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

Unless otherwise specified, "DOX" involved in the following examples all represent "doxorubicin"; "DTX" all represent "docetaxel"; "TAXOL" all represent "paclitaxel" and "PEG" all represent "poly "Ethylene glycol"; "Ⅳ collagenase" means "type IV collagenase"; "Ⅰcollagenase" means "type IV collagenase"; "Ⅱ collagenase" means "type II collagenase"; "Ⅴ collagenase" means "type V collagen protease".

In the following examples, in the PCL-PEG-COOH used, the molecular weight of PCL is 8000g/mol, and the molecular weight of PEG is 2000g/mol; in PLA-PEG-COOH, the molecular weight of PLA is 8000g/mol, and the molecular weight of PEG is 2000g/mol; in PLGA-PEG-COOH, the molecular weight of PLGA is 20000g/mol, and the molecular weight of PEG is 2000g/mol.

The preparation of embodiment 1.PCL-PEG-COOH blank micelles

The present embodiment provides the preparation method of the PCL-PEG blank micelles as a control experiment, specifically as follows:

Dissolve 5 mg of PCL-PEG-COOH in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Mix 100 µL of the above polymer solution and 400 µL of acetone again thoroughly. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 2. Preparation of DOX-PCL-PEG-COOH micelles with drug and carrier dosage ratio of 0.5:100

This embodiment provides the preparation method of DOX-PCL-PEG-COOH micelles as a control experiment, specifically as follows:

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 100 mL volumetric flask to obtain a 100 μg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-COOH in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Take 100 μL of polymer solution and 50 μL of doxorubicin solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 3. Preparation of DOX-PCL-PEG-COOH with drug and carrier ratio of 5:100

This embodiment provides the preparation method of DOX-PCL-PEG-COOH micelles as a control experiment, specifically as follows:

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 10 mL volumetric flask to obtain a 1 mg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-COOH in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Take 100 μL of polymer solution and 50 μL of DOX solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 4. Preparation of DOX-PCL-PEG-COOH micelles with drug and carrier ratio of 10:100

This embodiment provides the preparation method of DOX-PCL-PEG-COOH micelles as a control experiment, specifically as follows:

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 10 mL volumetric flask to obtain a 1 mg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-COOH in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Preparation of micellar drug and micellar drug and carrier with a dosage ratio of 10:100: take 100 μL of polymer solution and 100 μL of DOX solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 5. Preparation of DOX-PCL-PEG-COOH micelles with drug and carrier ratio of 15:100

This embodiment provides the preparation method of DOX-PCL-PEG-COOH micelles as a control experiment, specifically as follows:

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 10 mL volumetric flask to obtain a 1 mg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-COOH in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Preparation of micellar drug and carrier with a dosage ratio of 15:100: take 100 μL of polymer solution and 150 μL of DOX solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 6. Preparation of PCL-PEG-IV collagenase blank micelles

This embodiment provides the preparation method of PCL-PEG-IV collagenase blank micelles as a control experiment, specifically as follows:

Accurately weighed NHS (15.3mg, 0.15mmol) and EDC (28.7mg, 0.15mmol) were dissolved in 1mL of dichloromethane and vortexed to dissolve. Take COOH-PEG-PCL (50mg, 0.05mmol) solution and add 5mL of dichloromethane, place it in a magnetic stirrer, stir at 20-26°C for 15min, add NHS and EDC solution at the same time to generate NHS-PEG-PCL, mix Stir for 6-8h, rotary steam to volatilize the complete dichloromethane, add pure water to obtain the product. Add collagen solution (1 mL, 0.5 mg/mL) to the aqueous solution of NHS-PEG-PCL, and stir for 2 h. Membrane dialysis for 24 hours (Mw=3500), changing the water every 6 hours, and freeze-drying for 48 hours to obtain PCL-PEG-Ⅳ collagenase.

Dissolve 5 mg of PCL-PEG-IV collagenase in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Mix 100 µL of polymer solution and 400 µL of acetone well. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 7. Preparation of DOX-PCL-PEG-Ⅳcollagenase micelles with drug and carrier ratio of 0.5:100

The preparation method of functionalized DOX-PCL-PEG-Ⅳcollagenase micelles with the effect of penetrating tumor extracellular matrix of the present invention is as follows, wherein, in the following examples, the carrier refers to the amphiphilic polymer modified with collagenase :

Accurately weighed NHS (15.3mg, 0.15mmol) and EDC (28.7mg, 0.15mmol) were dissolved in 1mL of dichloromethane and vortexed to dissolve. Take PCL-PEG-COOH (50mg, 0.05mmol) solution and add 5mL of dichloromethane, place it in a magnetic stirrer, stir at 20-26°C for 15min, then add NHS and EDC solution at the same time to generate NHS-PEG-PCL, mix Stir for 6-8h, rotary steam to volatilize the complete dichloromethane, add pure water to obtain the product. Add type IV collagenase solution (1 mL, 0.5 mg/mL) to the aqueous solution of NHS-PEG-PCL, and stir for 2 h. The membrane was dialyzed for 24 hours (Mw=3500), the water was changed every 6 hours, and freeze-dried for 48 hours to obtain an amphiphilic polymer modified with collagenase (PCL-PEG-IV collagenase).

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 100 mL volumetric flask to obtain a 100 μg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-IV collagenase in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Take 100 μL of polymer solution and 50 μL of DOX solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water and stir for 4 hours to obtain amphiphilic polymer micelles (DOX-PCL-PEG-IV collagenase micelles) that can penetrate the tumor extracellular matrix after evaporating the organic solvent.

Example 8. Preparation of DOX-PCL-PEG-Ⅳcollagenase micelles with drug and carrier dosage ratio of 5:100

Accurately weighed NHS (15.3mg, 0.15mmol) and EDC (28.7mg, 0.15mmol) were dissolved in 1mL of dichloromethane and vortexed to dissolve. Take PCL-PEG-COOH (50mg, 0.05mmol) solution and add 5mL of dichloromethane, place it in a magnetic stirrer, stir at 20-26°C for 15min, then add NHS and EDC solution at the same time to generate NHS-PEG-PCL, mix Stir for 6-8h, rotary steam to volatilize the complete dichloromethane, add pure water to obtain the product. Add collagen solution (1 mL, 0.5 mg/mL) to the aqueous solution of NHS-PEG-PCL, and stir for 2 h. Membrane dialysis for 24 hours (Mw=3500), changing the water every 6 hours, and freeze-drying for 48 hours to obtain PCL-PEG-Ⅳ collagenase.

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 10 mL volumetric flask to obtain a 1 mg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-IV collagenase in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Take 100 μL of polymer solution and 50 μL of DOX solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 9. Preparation of PCL-PEG-IV collagenase micelles with drug and carrier dosage ratio of 10:100

Accurately weighed NHS (15.3mg, 0.15mmol) and EDC (28.7mg, 0.15mmol) were dissolved in 1mL of dichloromethane and vortexed to dissolve. Take PCL-PEG-COOH (50mg, 0.05mmol) solution and add 5mL of dichloromethane, place it in a magnetic stirrer, stir at 20-26°C for 15min, then add NHS and EDC solution at the same time to generate NHS-PEG-PCL, mix Stir for 6-8h, rotary steam to volatilize the complete dichloromethane, add pure water to obtain the product. Add collagen solution (1 mL, 0.5 mg/mL) to the aqueous solution of NHS-PEG-PCL, and stir for 2 h. Membrane dialysis for 24 hours (Mw=3500), changing the water every 6 hours, and freeze-drying for 48 hours to obtain PCL-PEG-Ⅳ collagenase.

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 10 mL volumetric flask to obtain a 1 mg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-IV collagenase in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Take 100 μL of polymer solution and 100 μL of DOX solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 10. Preparation of PCL-PEG-Ⅳcollagenase micelles with drug and carrier dosage ratio of 15:100

Accurately weighed NHS (15.3mg, 0.15mmol) and EDC (28.7mg, 0.15mmol) were dissolved in 1mL of dichloromethane and vortexed to dissolve. Take PCL-PEG-COOH (50mg, 0.05mmol) solution and add 5mL of dichloromethane, place it in a magnetic stirrer, stir at 20-26°C for 15min, then add NHS and EDC solution at the same time to generate NHS-PEG-PCL, mix Stir for 6-8h, rotary steam to volatilize the complete dichloromethane, add pure water to obtain the product. Add collagen solution (1 mL, 0.5 mg/mL) to the aqueous solution of NHS-PEG-PCL, and stir for 2 h. Membrane dialysis for 24 hours (Mw=3500), changing the water every 6 hours, and freeze-drying for 48 hours to obtain PCL-PEG-Ⅳ collagenase.

Dissolve 10 mg of DOX powder in methanol, ultrasonically dissolve, and dilute to a 10 mL volumetric flask to obtain a 1 mg/mL DOX standard solution. Dissolve 5 mg of PCL-PEG-IV collagenase in 0.5 mL of acetone to obtain a polymer solution of 10 mg/mL. Take 100 μL of polymer solution and 150 μL of DOX solution, add acetone until 500 μL. Under the condition of vigorous stirring, slowly add 5 mL of pure water, stir for 4 h, and evaporate the organic solvent.

Example 11. Particle Size and Potential Determination

The micellar solutions in Examples 1, 2, 6 and 7 were diluted with pure water to a certain concentration, and the particle size distribution and surface potential of the micelles were measured by a particle size analyzer and a Zeta potential analyzer.

Table 1 shows the particle size distribution and surface potential results of Examples 1, 2, 6, and 7. It can be seen from the table that the particle size of the PCL-PEG-COOH blank micelles is about 110nm, and increases to 180nm after drug loading about. The particle size of PCL-PEG-Ⅳcollagenase micelles is about 310nm, and increases to about 360nm after drug loading. At the same time, it can be seen that no matter before or after drug loading, the potential is around 0mV, showing electrical neutrality.

Table 1 The particle size distribution and surface potential test results of different micellar solutions

Example 12. Morphological observation of micelles

The prepared micelles of Examples 1, 2, 6 and 7 were respectively dropped onto a copper grid covered with a support film, dried under an infrared lamp for 2-4 hours, and the morphology of the nanoparticles was observed by transmission electron microscopy and photographed.

Fig. 1 is the test result of above-mentioned embodiment, and graph a, b, c, d represent the micelle of embodiment 1, 2, 6 and 7 respectively. By comparing the micelles in Figures a and b with those in Figures c and d, we can see that the particle size increases after drug loading, but the shape is still regular and round, and the drug doxorubicin enters the interior of the carrier.

Example 13. Micelle Encapsulation Efficiency and Drug Loading Determination

Take the micelle solutions in Examples 3-5 and 8-10, use high-speed centrifugation to measure the encapsulation efficiency and drug loading, put the prepared micelle in a centrifuge tube, and set the temperature at 8000rap/min, 4°C Then, ultracentrifuge for 30min, take the supernatant, and measure the free drug content.

Weigh 10mg of DOX, add pure water, dissolve it by ultrasonic and set the volume to 100mL, obtain a 100μg/mL DOX aqueous solution standard product to make a series of gradient concentrations, prepare a standard curve, and measure the fluorescence value of the sample.

Calculate the encapsulation efficiency and drug loading of micelles according to the following formula:

Drug loading (%) = W _DOX /W _carrier × 100%

Encapsulation efficiency (%) = (W _DOX -W _free )/W _{total DOX} × 100%

Wherein, _{the total DOX} of W is the total amount of medicine, W _free is the quality of the medicine not wrapped in micelles, and W _carrier is the quality of carrier.

Table 2 shows the encapsulation efficiency and drug loading results of Examples 3-5 and 8-10. When the feeding rate was 15%, the solutions of the two carriers all appeared turbid, and then the dialysis method was selected when the feeding rate was 10%. Encapsulation efficiency and drug loading. It can be seen from Table 2 that the encapsulation efficiency of DOX-PCL-PEG-COOH carrier is 90%, and the drug loading capacity is 9%. After linking IV collagenase, the encapsulation efficiency and drug loading capacity are slightly reduced.

Table 2 Encapsulation efficiency and drug loading test results of different micelles

Example 14. In vitro release investigation

The dialysis bag was boiled with pure water at 60°C for 15 minutes, the DOX-PCL-PEG-COOH micellar solution of Example 4 was placed in the dialysis bag, tied tightly with cotton rope, and suspended in 100 mL of PBS containing 40 mL of pH 7.4 Place in a beaker in a constant temperature shaking box at 37±1°C, shake for 48 hours, take 1mL samples at regular intervals of 0.5, 1, 2, 4, 8, 12, 24, 48 and 72 hours, and add an equal volume of release medium at the same temperature PBS. The samples taken were measured for drug release, and the cumulative drug release percentage was calculated.

Fig. 2 is the test result of the prepared micelle of embodiment 4, as can be seen from the figure, along with the increase of release time, the accumulative release degree of medicine rises gradually, and when the time exceeds 72h, release curve tends to be stable, can It can be seen that DOX-PCL-PEG-COOH has obvious sustained-release effect.

Example 15. Preparation of DOX-PLA-PEG-COOH micelles with drug and carrier ratio of 0.5:100

The amphiphilic polymer in Example 7 was replaced by PLA-PEG-COOH, and other experimental steps were referred to in Example 7 to prepare amphiphilic polymer micelles that could penetrate tumor extracellular matrix.

Example 16. Preparation of DOX-PLGA-PEG-COOH micelles with drug and carrier ratio of 0.5:100

The amphiphilic polymer in Example 7 was replaced with PLGA-PEG-COOH, and other experimental steps were referred to in Example 7 to prepare amphiphilic polymer micelles that could penetrate tumor extracellular matrix.

Example 17. Preparation of DTX-PCL-PEG-I collagenase micelles with drug and carrier ratio of 5:100

In Example 8, the antitumor drug was replaced by -DTX, IV collagenase was replaced by I collagenase, and other experimental procedures were referred to in Example 8, and amphiphilic polymer micelles that could penetrate tumor extracellular matrix were prepared.

Example 18. Preparation of TAXOL-PCL-PEG-Ⅱcollagenase micelles with drug and carrier ratio of 5:100

The anti-tumor drug in Example 8 was replaced by TAXOL, IV collagenase was replaced by II collagenase, and other experimental procedures were referred to in Example 8, and amphiphilic polymer micelles that could penetrate tumor extracellular matrix were prepared.

Example 19. Preparation of TAXOL-PCL-PEG-Ⅴcollagenase micelles with drug and carrier ratio of 5:100

The antitumor drug in Example 8 was replaced with TAXOL, IV collagenase was replaced with V collagenase, and other experimental steps were referred to in Example 8, and amphiphilic polymer micelles that could penetrate tumor extracellular matrix were prepared.

The above is only a preferred embodiment of the present invention, and is not intended to limit the present invention. It should be pointed out that for those of ordinary skill in the art, some improvements can be made without departing from the technical principle of the present invention. and modifications, these improvements and modifications should also be considered as the protection scope of the present invention.

Claims (10)

1. An amphiphilic polymer micelle capable of penetrating tumor extracellular matrix, characterized in that: comprising antineoplastic drugs, amphiphilic polymers and collagenase, said amphiphilic polymers combine said antitumor The drug is wrapped inside, and the amphiphilic polymer is covalently linked to the collagenase on the outside. 2. The amphiphilic polymer micelle capable of penetrating tumor extracellular matrix according to claim 1, characterized in that: the antitumor drug is one or more of docetaxel, doxorubicin and paclitaxel Several kinds. 3. The amphiphilic polymer micelle capable of penetrating tumor extracellular matrix according to claim 1, characterized in that: the amphiphilic polymer is PCL-PEG copolymer, PLA-PEG copolymer and PLGA -One or more of PEG copolymers. 4 . The amphiphilic polymer micelle capable of penetrating tumor extracellular matrix according to claim 1 , wherein the molar ratio of the collagenase to the amphiphilic polymer is 1:100-1000. 5. The amphiphilic polymer micelle capable of penetrating tumor extracellular matrix according to claim 1, characterized in that: said collagenase is type I collagenase, type II collagenase, type IV collagenase and type V collagenase One or more of collagenases. 6. A method for preparing an amphiphilic polymer micelle capable of penetrating tumor extracellular matrix according to any one of claims 1-5, characterized in that it comprises the following steps: (1) react the amphiphilic polymer and the cross-linking agent in the first organic solvent at 20-26° C., remove the first organic solvent, and then react with collagenase in pure water at 20-26° C. React at 26°C to obtain an amphiphilic polymer modified with collagenase; (2) dissolving the amphiphilic polymer modified with collagenase and the antitumor drug in a second organic solvent to obtain a drug solution; (3) Add the drug solution into water, remove the second organic solvent, and form the amphiphilic polymer micelles that can penetrate the tumor extracellular matrix. 7. preparation method according to claim 6 is characterized in that: in step (1), described linking agent is N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl base) carbodiimide hydrochloride. 8. The preparation method according to claim 6, characterized in that: in step (1), the first organic solvent is one or more of dimethyl sulfoxide, acetonitrile and chloroform. 9. The preparation method according to claim 6, characterized in that: in step (2), the second organic solvent is one or more of acetone, methanol, ethanol, methylene chloride and chloroform. 10. The preparation method according to claim 6, characterized in that: in step (3), the drug solution is added into pure water under stirring condition, and the stirring speed is 400-700rpm.