Application of charge crosslinking carrier as tumor chemotherapy drug

Abstract

The invention discloses a nano-carrier based on charge crosslinking, a preparation method of the nano-carrier and application of the nano-carrier as a tumor chemotherapy drug. Wherein the nano-carrier is formed by assembling two polymer chains with positive and negative charges through electrostatic interaction, a precursor ligand on a polymer chain B releases an organic ligand under exogenous or endogenous stimulation, and competes for metal ions on a polymer chain A with positive charges to generate a metal complex in situ, and meanwhile, the electrostatic interaction of the nano-carrier is weakened, so that the metal complex is formed. The dissociation of the nano-carrier is caused, a macromolecular chain B containing an organic ligand is released, and the effect of tumor chemotherapy or immunoregulation is achieved. The method is simple to operate, short in time consumption and mild in reaction condition; the material is good in biocompatibility and high in safety degree. Pharmacodynamic experiments in some embodiments show that the nano-carrier can effectively kill the breast cancer and trigger cellular immunogenic death, meanwhile, tumor cell metastasis is inhibited, and a new thought and means are provided for the clinical problem of breast cancer lung metastasis urgently needing to be solved at present.

Description

Technical Field

[0001] The invention belongs to the field of nano materials and their preparation, and specifically relates to a charge cross-linking carrier and a preparation method thereof and application thereof as a tumor treatment drug. Background Art

[0002] Malignant tumors are one of the main diseases that threaten the health of Chinese residents. According to the data from China's death cause monitoring, deaths from malignant tumors account for nearly a quarter of all deaths among residents. As the aging of the population gradually intensifies, the burden of major chronic diseases, including malignant tumors, is on the rise. At present, traditional tumor treatment methods include surgery, chemotherapy, and radiotherapy. Due to the limitations of resection in surgical treatment, immunosuppression caused by radiotherapy, and drug resistance caused by chemotherapy, the effects of these treatment modes are often poor. At present, clinical tumor treatment recommends the use of combined therapy, which improves the efficacy and reduces adverse reactions through multi-mechanism synergy, combination of radiotherapy and chemotherapy, and combination of chemotherapy and immunotherapy.

[0003] Drug carrier design is a key step in the development of drug delivery systems, which directly affects the effectiveness, targeting and safety of drugs. In recent years, many studies have been devoted to the development of drug carriers for efficient tumor treatment. However, the processes of most drug carriers include complex synthesis and preparation processes, high production costs, poor controllability, and weak repeatability. In addition, drug carriers also face "off-target effects". How to ensure the stability of drugs during circulation in the body and how to selectively deliver drugs to tumor cells and release them for efficacy have always been urgent problems to be solved.

[0004] Therefore, it is necessary to develop a simple and efficient drug carrier that can carry multiple drugs. Based on this, a design model of a charge-crosslinked drug carrier is proposed. This type of carrier has a simple process, controllability, and repeatability; it can respond to the microenvironment in the tumor in vivo, release drugs synchronously, avoid side effects caused by metabolic differences between drugs, and improve the therapeutic effect through drug synergy. Summary of the invention

[0005] The purpose of the present invention is to provide a double polymer chain-based charge cross-linked carrier for tumor treatment. The positively charged polymer chain is prepared by the coordination of metal ions with organic ligands on polymer chain A, and is uniformly mixed with the negatively charged polymer chain B loaded with prodrugs through electrostatic interaction.

[0006] The purpose of the present invention is to overcome two major difficulties in the existing treatment model: one is the complexity of drug carrier design. The current design of drug carriers often involves complex synthetic preparation processes, with poor stability and repeatability, and unstable efficacy; the other is the problem of multi-drug delivery for tumor treatment. Due to differences in drug metabolic kinetics and the off-target effects of the drugs themselves, the spatiotemporal distribution of drugs is uneven during combined administration, and the tumor cell selectivity is poor, resulting in poor efficacy and obvious side effects.

[0007] The purpose of the present invention can be achieved by the following technical solutions:

[0008] A metal coordination-mediated charge cross-linked carrier, characterized in that a positively charged polymer chain is prepared by the coordination of metal ions with ligands on polymer chain A, and is uniformly mixed with a negatively charged polymer chain B carrying a prodrug through electrostatic interaction, thereby obtaining a uniform drug carrier that can be dispersed in an aqueous solution.

[0009]

[0010] The metal ion in the polymer chain A is preferably Cu 2+ , Cu + , Fe 3+ ,Fe 2+ , Zn 2+ ;

[0011] More preferably, Cu 2+ , Cu + , Zn 2+ ;

[0012] In some specific embodiments of the present invention, the metal ion is Cu 2+ .

[0013] The organic ligands in the polymer chain A include but are not limited to molecules containing biological activity; the molecules may be molecules having tumor chemotherapy or immunomodulatory effects, including but not limited to plerixafor (AMD3100), paclitinib, and sotolactis.

[0014] In some specific embodiments of the present invention, the organic ligand of the polymer chain A is preferably plerixafor (AMD3100).

[0015] In some embodiments, the polymer chain A is preferably hyaluronic acid, chitosan, chondroitin sulfate, polylysine; the molecular weight of the polymer is preferably 2k-10w.

[0016] In some embodiments, the prodrug in the negatively charged polymer chain B may be a proligand constituting a metal complex; the proligand constituting the metal complex includes but is not limited to a prodrug molecule of a dithiocarbamic acid derivative;

[0017] In some embodiments, the negatively charged polymer is preferably chondroitin sulfate or sulfated polysaccharide.

[0018] In some embodiments, the prodrug-modified negatively charged polymer chain B is a dithiocarbamate prodrug-modified polymer compound represented by the general formula I:

[0019] General formula Ⅰ:

[0020] in, is a dithiocarbamate group,

[0021] N is the negatively charged polymer chain B mentioned above;

[0022] In some of the embodiments, R1 and R2 are preferably C2-C5 short-chain aliphatic hydrocarbons independently of each other or combined into C3-C8 cycloalkane;

[0023] M is an exogenous / endogenous stimulus response group. In some embodiments of the present invention, preferably, a ROS-sensitive bond such as phenylboronic acid ester, a thioketal bond, a GSH-sensitive group disulfide bond, a pH-sensitive bond such as an imine bond, a light-sensitive group such as o-nitrobenzyl alcohol, an enzyme-specific cleavage chemical bond such as the sequence of the FAP-α enzyme, an X-ray-sensitive bond such as a fluorinated aryl azide;

[0024] In some embodiments of the present invention, the ROS-sensitive bond is more preferably a phenylboronic ester bond, the structure of which is as follows:

[0025]

[0026] L is a linker connecting the prodrug and the negatively charged polymer chain B. In some embodiments of the present invention, it is preferably adipic acid dihydrazide, and the structure is as follows:

[0027]

[0028] The preligand on polymer chain B releases free ligand under exogenous or endogenous stimulation and competes for metal ions on positively charged polymer chain A to generate metal complexes in situ. At the same time, the electrostatic effect of the carrier is weakened, leading to the dissociation of the carrier and releasing the polymer chain B containing the metal ligand, thereby playing a role in tumor chemotherapy or immunomodulation.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] The preparation of this charge-crosslinked carrier is simple, time-saving, and has mild reaction conditions. It can respond to the microenvironment within the tumor in vivo, release drugs synchronously, avoid side effects caused by metabolic differences between drugs, and improve the therapeutic effect through drug synergy.

[0031] In some specific embodiments, the carrier overcomes the problem that dithiocarbamic acid derivative drugs and copper cannot be distributed synchronously at the tumor site to effectively exert anti-tumor effects; secondly, in view of the easy metastasis of tumors, a synergistic strategy of killing tumor cells in situ and activating immunogenic death and inhibiting cell invasion signals is proposed to inhibit tumor cell growth and metastasis. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] The specific embodiments of the present invention are further described in detail below in conjunction with the accompanying drawings, wherein:

[0033] Figure 1 : H NMR spectra of the prodrug BpDTC and the negative control pDTC in Example 1 of the present invention

[0034] Figure 2 : H NMR spectra of carrier polymer chains CBD, CD and HA-AMD in Example 2 of the present invention

[0035] Figure 3 : Dynamic light scattering particle size measurement diagram of Example 3 of the present invention

[0036] Figure 4 : Response and release of the carrier and its individual components to H2O2 in Example 4 of the present invention

[0037] Figure 5 : Cytotoxicity of the carrier of Example 5 of the present invention

[0038] Figure 6 : Example 6 of the present invention vector induced generation of reactive oxygen species

[0039] Figure 7 : Example 7 of the present invention vector induced proteasome accumulation diagram

[0040] Figure 8 : Anti-migration effect diagram of carrier component HA-AMD in Example 8 of the present invention DETAILED DESCRIPTION

[0041] The present invention is further described below in conjunction with the following examples, but the present invention is not limited to the following examples. For example, the concentration and volume of the solution used can be adjusted as required.

[0042] The CXCL12 (SDF-1) / CXCR4 biological axis consists of the chemokine CXCL12 and its specific receptor CXCR4. Studies have shown that under tumor pathological conditions, this signal axis is highly activated and is associated with tumor growth, invasion, and prognosis. Currently, the only clinical CXCR4 small molecule inhibitor, plerixafor (AMD3100), is used for autologous transplantation in patients with non-Hodgkin's lymphoma and multiple myeloma. In addition, studies have shown that AMD3100 can effectively cut off the CXCL12 / CXCR4 axis and inhibit tumor development and metastasis. At the same time, AMD3100 has a macrocyclic structure that can deliver metal ions to the tumor area to achieve in situ formation of metal complexes.

[0043] Example 1: Preparation of ROS-sensitive diethyldithiocarbamate prodrug BpDTC and its negative control pDTC

[0044] (1) The preparation process of BpDTC compound is as follows:

[0045]

[0046] 1) Preparation of intermediate 2: p-Bromophenylacetic acid (5 g, 1 eq) was dissolved in methanol (50 mL), concentrated sulfuric acid (1.73 mL, 1.5 eq) was dissolved in methanol, and slowly dripped into the above solution, reacted at 65 °C for 2 h, and the reaction was monitored by TLC (petroleum ether: ethyl acetate = 8:1). After the reaction was completed, it was cooled to room temperature, the solvent was dried under reduced pressure, redissolved in ethyl acetate, and washed 3 times with saturated brine. The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and the solvent was removed by spin drying to obtain intermediate 2.

[0047] 2) Preparation of intermediate 3: Intermediate 2 (5.348 g, 1 eq), bis-pinacol borate (11.08 g, 2 eq), [1,1′-bis(diphenylphosphino)ferrocene] dichloropalladium dichloromethane complex (893 mg, 0.05 eq) and potassium acetate (6.42 g, 3 eq) were dissolved in 100 mL of dioxane and reacted at 80 degrees under nitrogen protection. The reaction was monitored by TLC (petroleum ether: ethyl acetate = 10: 1). After the reaction was completed, the solvent was dried under reduced pressure, redissolved with dichloromethane, and the filtrate was filtered. Intermediate 3 was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 20: 1).

[0048] 3) Preparation of intermediate 4: Intermediate 3 (5 g, 1 eq) was dissolved in 10 mL of dichloromethane. Dichlorothionyl (26.5 mL, 20 eq) was slowly added to the reaction system under an ice bath. Two drops of DMF were added to catalyze the reaction. The mixture was refluxed at 50 °C under nitrogen protection and monitored by TLC (petroleum ether: ethyl acetate = 8:1). After the reaction was completed, ice water was added to quench the mixture and dichloromethane was used for multiple extractions. The organic phase was dried over anhydrous sodium sulfate and filtered to obtain intermediate 4.

[0049] 4) Preparation of intermediate 5: Intermediate 4 (4.88 g, 1 eq) was dissolved in 20 mL of acetonitrile, sodium diethyldithiocarbamate (2.83 g, 1.05 eq) was dissolved in 20 mL of acetonitrile and gradually added to the above system, reacted at room temperature, and monitored by TLC (petroleum ether: ethyl acetate = 20: 1). After the reaction was completed, the solvent was dried under reduced pressure, redissolved in ethyl acetate, and washed with water and saturated brine. The organic phase was taken, dried over anhydrous sodium sulfate, and filtered. Intermediate 5 was obtained by purification by silica gel column chromatography (petroleum ether: ethyl acetate = 30: 1).

[0050] 5) Preparation of pDTC: Intermediate 5 (3.2 g, 1 eq) was dissolved in 5 mL tetrahydrofuran, and 5 mL NaOH solution (605 mg, 2 eq) was added. The reaction was allowed to react at room temperature and monitored by TLC (petroleum ether: ethyl acetate = 6:1). After the reaction was completed, the mixture was extracted with ethyl acetate, the aqueous layer was taken, and the pH was adjusted to 5. At this time, a white precipitate was precipitated. The mixture was extracted with dichloromethane several times, dried with anhydrous sodium sulfate, filtered, and dried to obtain the final product BpDTC.

[0051] (2) The preparation process of pDTC compound is as follows:

[0052]

[0053] 1) Preparation of intermediate 6: Methyl bromophenylacetate (1.2 g, 1 eq) was dissolved in 10 mL of acetonitrile, sodium diethyldithiocarbamate (898 mg, 1 eq) was dissolved in 10 mL of acetonitrile and slowly added to the above system, reacted at room temperature, and monitored by TLC (petroleum ether: ethyl acetate = 6:1). After the reaction was completed, the filtrate was filtered, the solvent was dried under reduced pressure, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 20:1) to obtain intermediate 6.

[0054] 2) Preparation of pDTC: Intermediate 6 (1.1 g, 1 eq) was dissolved in 5 mL THF, and 5 mL NaOH solution (443 mg, 3 eq) was added. The reaction was allowed to react at room temperature and monitored by TLC (petroleum ether: ethyl acetate = 6:1). After the reaction was completed, the mixture was extracted with ethyl acetate, the aqueous layer was taken, and the pH was adjusted to 5. At this time, a white precipitate was precipitated, which was extracted with dichloromethane several times. The organic layer was taken, dried with anhydrous sodium sulfate, and filtered to obtain the final product pDTC.

[0055] Example 2: Preparation of prodrug-modified chondroitin sulfate A polymer chains CBD and CD and AMD3100-modified hyaluronic acid polymer chains HA-AMD

[0056] (1) The preparation process of CBD and CD polymer compounds is as follows:

[0057]

[0058] 1) Preparation of CS-ADH: 2g chondroitin sulfate A (CS-A) was dissolved in 20mL of distilled water, stirred to dissolve, 4g 1-ethyl-(3-dimethylaminopropyl) carbodiimide (EDC) and 0.585g N-hydroxysuccinimide (NHS) were added, and the pH was adjusted to 6-7 with hydrochloric acid, and stirred for about 15min to activate the carboxyl group. 8.8g adipic acid dihydrazide (ADH) was added to the system, and NaOH was added to adjust the pH to 6-7, stirred at room temperature for 24h, dialyzed for three days (Mw=3500), and freeze-dried to obtain CS-ADH. The grafting rate was calculated by nuclear magnetic hydrogen spectrum.

[0059] 2) Preparation of CBD: BpDTC (0.6 g, 1 eq) was dissolved in 10 mL DMSO, 4-(4,6-dimethoxytriazine-2-yl)-4-methylmorpholine hydrochloride (DMTMM) (0.232 g, 0.6 eq) was dissolved in 7.5 mL water and added to the above system, activated for 0.5 h. CS-ADH (0.188 g, 0.72 eq) was dissolved in 7.5 mL water and added to the above system, stirred at room temperature for 24 h. After the reaction, dialyzed for three days (Mw=3500), and freeze-dried to obtain CBD. The grafting rate was calculated using H NMR spectroscopy.

[0060] 3) Preparation of CD: pDTC (0.6 g, 1 eq) was dissolved in 10 mL DMSO, 4-(4,6-dimethoxytriazine-2-yl)-4-methylmorpholine hydrochloride (DMTMM) (0.165 g, 0.6 eq) was dissolved in 7.5 mL water and added to the above system, activated for 0.5 h. CS-ADH (0.188 g, 0.72 eq) was dissolved in 7.5 mL water and added to the above system, stirred at room temperature for 24 h. After the reaction was completed, it was dialyzed for three days (Mw=3500) and freeze-dried to obtain CD. The grafting rate was calculated using H NMR spectroscopy.

[0061] (2) The preparation process of HA-AMD polymer compound is as follows:

[0062]

[0063] 1) Synthesis of HA-MA: 3g HA (1w-10w) was dissolved in 50mL water, the reaction solution was placed at 0℃, 5.734g methacrylic anhydride was slowly added, the pH was adjusted to 8-9 with 5M NaOH, and the reaction was continued to be stirred at 0℃ for 8h (the reaction pH was adjusted to be stable at 8-9 during the reaction). After the reaction was completed, the reaction solution was added dropwise to 500mL anhydrous ethanol, stirred for 10min, and then the precipitate was taken and washed three times with anhydrous ethanol (500mL×3), the precipitate was collected by suction filtration and vacuum dried to obtain HA-MA. The grafting rate was calculated by nuclear magnetic hydrogen spectrum.

[0064] 2) Synthesis of HA-AMD: HA-MA (100 mg, 1 eq) was dissolved in a mixture of 0.5 mL DMSO and 2 mL water, triethylamine (15 mg, 1 eq) was added, and plerixafor (AMD3100) (100 mg, 1.5 eq) was added to methanol and slowly dripped into the above system. Under nitrogen protection, the reaction was carried out at room temperature for 24 h in the dark. After the reaction, the mixture was dialyzed for three days (Mw=3500) and freeze-dried to obtain HA-AMD. The grafting rate was calculated by H NMR spectroscopy.

[0065] Example 3: Construction of charge-crosslinked nanocarriers and dynamic light scattering particle size determination

[0066] The mother solution of HA-AMD (containing AMD3100 5 mM, the concentrations mentioned below all refer to AMD3100 concentrations) was prepared with ultrapure water, the mother solution of prodrug-modified polymer chain CBD (containing prodrug BpDTC 1 mM, the concentrations mentioned below all refer to BpDTC concentrations) was prepared with DMSO: ultrapure water at a ratio of 1:1, and the copper chloride solution (20 mM) was prepared with ultrapure water. The copper-coordinated HA-AMD-Cu (1 mM) was obtained by mixing HA-AMD and copper solution. The 1 mM HA-AMD-Cu aqueous solution was dropped into the 1 mM CBD solution at a ratio of 1:8, and the magnetic stirring was continued (650 r / min) for 60 min to allow HA-AMD-Cu to complex with CBD through electrostatic interaction. The nanocarrier (CBD-HM-Cu) was then obtained by ultrasound and extrusion, and stored at 4°C.

[0067] The corresponding negative nanocarrier CD-HM-Cu replaces the above CBD chain with the CD chain, and other methods remain the same.

[0068] Figure 3 The dynamic light scattering particle size results showed that the obtained nanocarriers had a uniform particle size of approximately 230 nm and a PDI of 0.176.

[0069] Example 4: Response of Nanocarriers and Their Individual Components to H2O2 Release

[0070] The sensitivity of the selected nanocarriers and their individual components to different concentrations of H2O2 and the active metal complex Cu(DTC)2 were determined by HPLC. A C8 column was selected, the mobile phase was CH3OH:H2O =90%:10% (0.05% HCOOH, v / v), and the characteristic ultraviolet absorption peak of Cu(DTC)2 was selected at 432 nm.

[0071] The nanocarriers and individual components were mixed with H2O2 (50 mM) or without H2O2, dissolved in a solution containing 5% DMSO and 95% H2O (v / v), and detected after incubation for 2 h.

[0072] Figure 4 It showed that under the condition of 50 mM H2O2 concentration, the nanocarrier group CBD-HM-Cu with response activation ability and the prodrug modified polymer chain CBD-Cu alone had significant Cu(DTC)2 production, while no Cu(DTC)2 production was detected in other experimental groups without response activation ability.

[0073] Example 5: Cytotoxicity of Nanocarriers

[0074] MTT assay: 4T1 cells were seeded in 96-well plates (5 × 104 cells / well) and incubated for 24 h. Then, the nanocarriers CBD-HM-Cu and CD-HM-Cu were treated with 0.625μM, 1.25μM, 2.5μM, 5μM, 10μM, and 20μM (referring to the drug concentration in the polymer chain CBD or CD). After each group was placed in a 37°C cell culture incubator for 24 h, 5mg / mL MTT solution was added to each well, and the culture medium was replaced with 100 μL DMSO after incubation for another 4 hours. The optical density (OD) of each well was measured at 490 nm using a microplate reader.

[0075] The group without drug addition was used as the control group, and the well without cells was used as the blank group. The cell survival rate was calculated according to formula 2-1:

[0076]

[0077] Figure 5 It was shown that after 24 hours of co-incubation, the cell survival rate of the CBD-HM-Cu carrier group had a significant tumor cell killing effect as the concentration increased. At a concentration of 20μM, the cell survival rate was only 5%, while the CD-HM-Cu group that could not be activated by response still had no killing effect at high concentrations. This shows that CBD-HM-Cu can respond to H2O2 in tumor cells and be activated to generate Cu(DTC)2 to take effect.

[0078] Example 6: Nanocarrier-induced generation of reactive oxygen species

[0079] DCFH-DA was used as a ROS probe to detect intracellular ROS generation by FCM. The specific sample processing method is as follows:

[0080] 4T1 cells were cultured at 5 × 10 4The cells were seeded at a density of 100 μM / well in a 12-well plate and incubated in a 37°C, 5% CO2 incubator for 24 h. After the cells were completely attached to the wall, the original culture liquid was aspirated and 1 mL of DMEM containing drug culture solution was added: Cu(DTC)2 (2.5 μM), CBD (20 μM), CBD+HM (CBD is 20 μM, HM is 2.5 μM), CD-HM-Cu NPs (CD is 20 μM, HM is 2.5 μM, copper is 5 μM), CBD-HM-Cu NPs (CBD is 20 μM, HM is 2.5 μM, copper is 5 μM), and each group was incubated in a 37°C incubator for 7 h. After the incubation was completed, the drug-containing DMEM solution was aspirated and the cells were washed 3 times with PBS. Subsequently, DMEM culture medium containing DCFH-DA probe (10 μM) was added and the cells were incubated in an incubator for 30 min. After the incubation, the DMEM solution containing DCFH-DA was removed, and the cells were washed three times with PBS. The cells were digested with trypsin and collected for FCM detection.

[0081] Figure 6 It was shown that after incubation with 4T1 cells, the Cu(DTC)2 group and the CBD-HM-Cu NPs group showed a significant enhancement in the fluorescence signal of DCFH-DA compared with the CT group, indicating an increase in the level of intracellular ROS.

[0082] Example 7: Nanocarrier-induced proteasome accumulation

[0083] 4T1 cells were seeded in 6-well plates (10 5 cells / well) and incubated for 24 h. Then, the cells were incubated with each group in Example 6 for another 10 hours (Cu(DTC)2 incubation time was 5 h). The cells were washed 3 times with PBS and then collected by centrifugation at 1500 rpm for 5 minutes. The cells were lysed on ice with RIPA lysis buffer for 30 minutes. Proteins in the supernatant were obtained by centrifugation at 12000 rpm for 20 minutes, and protein quantification was performed using a BCA kit. Subsequently, proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and then transferred to a PVDF membrane. After blocking in 5% skim milk at room temperature for 2 hours, the PVDF membrane was incubated with ubiquitin protein and β-Actin primary antibodies at 4°C overnight. Subsequently, the PVDF membrane was incubated with HRP-conjugated secondary antibodies at room temperature for 2 hours, and finally observed by chemiluminescent reagent autoradiography.

[0084] Figure 7 It was shown that Cu(DTC)2 and CBD-HM-Cu NPs could effectively accumulate ubiquitinated proteins.

[0085] Example 8: Anti-migration effect of nanocarrier component HA-AMD

[0086] Three horizontal scratches were made in each well of a 6-well plate in advance, and then 4T1 cells were inoculated into the 6-well plate and incubated to allow the cells to proliferate to a confluency of 90%. Three vertical scratches were made on the cells using a sterile 20 μL pipette tip, and dead cells were washed away with PBS. 2 mL of drug-containing DMEM medium was added to the experimental group: HA-AMD (2.5 μM), AMD (5 μM), and drug-free medium was added to the control well. Photos were taken at 0 h and 24 h after the scratches, and 3 scratch intersections were selected for each well to be photographed, and the empty area in the middle was measured using ImageJ software.

[0087] Mobility calculation method: Mobility 24 h (%) =(Area 0 h -area 24 h ) / area 0 h x100%.

[0088] Figure 8 It was shown that both AMD and HA-AMD could inhibit the migration of tumor cells.

Claims

1. A metal coordination-mediated charge cross-linked carrier, characterized in that: The positively charged polymer chain is prepared by the coordination of metal ions with organic ligands on the polymer chain A, and is uniformly mixed with the negatively charged polymer chain B loaded with the prodrug through electrostatic interaction.

2. The metal coordination-mediated charge cross-linked carrier according to claim 1, characterized in that: The metal ions include but are not limited to Cu 2+ , Cu + , Fe 3+ , Fe 2+ , Zn 2+ , Ca 2+ , Ni 2+ , Mn 2+ , Co 2+ , Mg 2+ ,Hg 2+ ,Pb 2+ .

3. The metal coordination-mediated charge cross-linked carrier according to claim 1, characterized in that: The organic ligand on the polymer chain A has the following structural characteristics: it contains one or more of the coordinating groups such as carboxyl, hydroxyl, N electron donor, N and O electron donor or N, O and S electron donor; the organic ligand containing the coordinating group includes but is not limited to monodentate ligand, multidentate ligand, amphipathic ligand; the organic ligand includes but is not limited to heterocyclic compounds, fatty amines, sulfur compounds, carboxyl compounds, hydroxyl compounds, phosphorus-containing compounds; the organic ligand includes but is not limited to biologically active molecules such as Plerixafor (AMD3100), Abametapir, Doxorubicin, Pacritinib, Sotorasib, Bortezomib.

4. The metal coordination-mediated charge cross-linked carrier according to claim 1, characterized in that: The positively charged polymer chain A includes but is not limited to hyaluronic acid, chitosan, chondroitin sulfate, fucoidan, polylysine, polylactic acid, polyvinyl alcohol, dextran, polyvinyl pyrrolidone, and aliphatic polyamide; the molecular weight of the polymer is 2k-50w.

5. The metal coordination-mediated charge cross-linked carrier according to claim 1, characterized in that: The prodrug-modified negatively charged polymer chain B, the prodrug may be a proligand constituting a metal complex; the proligand constituting a metal complex includes but is not limited to a prodrug molecule of a dithiocarbamic acid derivative; The negatively charged polymer chains include, but are not limited to, chondroitin sulfate, sulfated polysaccharides, polyacrylic acid, polyvinyl sulfonic acid, and polyvinyl phosphoric acid.

6. The prodrug-modified negatively charged polymer chain B according to claim 5, characterized in that: It is a polymer compound modified with a dithiocarbamic acid prodrug as shown in the general formula Ⅰ: General formula Ⅰ: in, is a dithiocarbamate group, N is the negatively charged polymer chain B mentioned above; R1 and R2 can be different or the same C1-C10 alkyl chains, C2-C10 olefin chains or alkyne chains, or can be aliphatic chains substituted or polysubstituted in the middle or at the end by hydroxyl, carboxyl, cyano, nitro, or different halogen atoms (F, Cl, Br, I), or combined into 3-8 membered aliphatic rings, aliphatic aromatic rings, and heterocyclic rings, including single-substituted or polysubstituted heterocyclic rings containing nitrogen, sulfur, oxygen, and boron; M is an exogenous / endogenous stimulus response group, including but not limited to ROS sensitive bonds such as phenylboronic acid ester, thioketal bond, diselenide bond, ditelluride bond, etc., GSH sensitive bonds such as disulfide bonds, pH sensitive bonds such as hydrazone bonds, imine bonds, etc., light sensitive groups such as o-nitrobenzyl alcohol, etc., chemical bonds specifically cleaved by enzymes such as FAP-α enzyme, MMP enzyme corresponding sequences, etc., X-ray sensitive bonds such as fluorinated aromatic azide, etc.; L is a linker connecting the prodrug and the negatively charged polymer chain B, including but not limited to adipic acid dihydrazide, short chain PEG, C2-C 10 Saturated or unsaturated fatty chains are connected by triazoles (formed by click-reactive groups such as azide and alkyne).

7. The metal coordination-mediated charge cross-linked carrier according to claims 1-6, characterized in that: The preligand on polymer chain B releases free ligand under exogenous or endogenous stimulation and competes for metal ions on positively charged polymer chain A to generate metal complexes in situ. At the same time, the electrostatic effect of the carrier is weakened, leading to the dissociation of the carrier and releasing polymer chain B containing organic ligand, thereby playing a role in tumor chemotherapy or immunomodulation.

8. Use of the metal coordination-mediated charge cross-linked carrier according to any one of claims 1 to 7 in a drug for treating and / or preventing cancer.

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