A spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix and its application

Abstract

The present invention discloses a spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix and its application. The delivery system includes: i) nanoparticles loaded with drugs using a metal polyphenol network modified with polyvinylpyrrolidone as a carrier; and ii) a metal ion chelator; the nanoparticles are used for targeted tumor delivery of drugs; the metal ion chelator is used to disassemble the nanoparticles when they reach the tumor extracellular matrix to release the drugs, thereby degrading the tumor extracellular matrix, enhancing blood perfusion, relieving tumor hypoxia, promoting drug penetration and T cell infiltration to reshape the tumor microenvironment. The present invention provides a novel delivery strategy for drugs targeting the tumor extracellular matrix, effectively overcoming the problem of insufficient treatment such as PDT caused by the TME. It provides new ideas for drug delivery targeting the tumor extracellular matrix and the treatment of various solid tumors.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix and its applications. Background Art

[0002] Malignant tumors have become one of the major diseases seriously endangering human health. In recent years, more and more tumor-targeted nano-drugs have been developed to effectively accumulate in solid tumors through the enhanced permeability and retention (EPR) effect. At the same time, based on this highly efficient delivery strategy, nano-formulations combining multiple therapies have also been widely developed, such as photodynamic therapy (PDT), sonodynamic therapy (SDT), chemodynamic therapy (CDT), chemotherapy (CT), photothermal therapy (PTT), immunotherapy, etc. Among them, PDT is a newly emerging anti-cancer treatment method. Its treatment mechanism is that under local laser irradiation, the photosensitizer can convert molecular oxygen in the tumor into cytotoxic reactive oxygen species (ROS) to oxidize and kill tumor cells, with characteristics such as non-invasiveness, controllability, and high efficiency, and has been proven to have good application prospects.

[0003] However, the immunosuppressive tumor microenvironment (TME) represented by low oxygen, low pH, and high content of reducing GSH severely restricts the application of most treatment strategies, such as oxygen-dependent PDT, SDT, CDT, radiotherapy, etc.; and hypoxia and low pH lead to the deprivation of the functions of T cells and NK cells, and the accumulation of myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs), thus resulting in immune escape.

[0004] Moreover, the tumor physical microenvironment such as highly dense extracellular matrix (ECM), abnormal tumor vascular and lymphatic systems, and elevated tumor interstitial fluid pressure (IFP) essentially hinders the effective delivery and penetration of nanoparticles (NPs) into deeper regions of the tumor. For example, tumor-associated fibroblasts (CAFs) preferentially located near blood vessels have a strong binding site barrier (BSB) effect on drugs and nanoparticles attempting to enter the tumor core site. They will also preferentially capture most of the nanoparticles and drugs that were originally intended to be delivered to tumor cells, resulting in very few nanoparticles or drugs reaching the tumor core. Although many targeted strategies show great potential in combating the TME at the biochemical level, the rigid ECM physical barrier is always the key problem hindering the maximum exertion of the above various treatment strategies.

[0005] At present, there are few reports on drug delivery to the extracellular matrix. One of the more important reasons is that there are still many difficulties in drug delivery to the extracellular matrix. For example, considering the important role of concentrated ECM in TME, hyaluronic acid (HA) is the main component of over-abundant tumor ECM, which plays a vital role in obstructing blood supply, maintaining the rigid structure of ECM, and producing hypoxia. Using hyaluronidase (HAase) to directly degrade HA seems to be an attractive method to alleviate tumor hypoxia. However, how to maintain the good biological activity of HAase from preparation to entry into tumor tissue, reduce immunogenicity, and how to accurately release it in ECM to prevent nonspecific degradation of normal human tissues and prevent efficient delivery strategies by cells are extremely challenging. Moreover, among many nanocarriers, the response release of most smart responsive nanocarriers to TME is limited and time-consuming, and they are often taken up or cleared by cells too early, which is not conducive to the rapid and effective release of drugs in the extracellular matrix, which also increases the difficulty of nanodrug delivery with targets in the extracellular matrix of tumor cells. Summary of the invention

[0006] In view of the shortcomings of the prior art, the present invention provides a spatiotemporal controllable drug delivery system targeting tumor cell extracellular matrix and its application. The delivery system adopts a metal polyphenol network modified with polyvinyl pyrrolidone to load drugs, and cooperates with a metal ion chelator to achieve targeted tumor delivery and spatiotemporal specific disassembly of drugs acting on tumor cell extracellular matrix, thereby providing a new delivery strategy for tumor cell extracellular matrix drugs, effectively overcoming the problem of insufficient treatment such as PDT caused by TME.

[0007] One aspect of the present invention provides a spatiotemporal controllable drug delivery system targeting the extracellular matrix of tumor cells, comprising: i) nanoparticles loaded with drugs using a metal polyphenol network modified by polyvinyl pyrrolidone as a carrier; and ii) a metal ion chelator; the nanoparticles are used for targeted tumor delivery of drugs; the metal ion chelator is used to disassemble the nanoparticles to release the drugs when the nanoparticles reach the extracellular matrix of tumor cells.

[0008] As a specific embodiment of the present invention, the drug is selected from any one of hyaluronidase, tyrosine kinase inhibitors, and immune checkpoint inhibitors; preferably, the drug is hyaluronidase.

[0009] As a specific embodiment of the present invention, the metal polyphenol network is a porous mesh structure material formed by metal ions and polyphenol compounds through coordination connection; preferably, the metal ions are iron ions; preferably, the polyphenol compounds are pentagalloylglucose.

[0010] As a specific embodiment of the present invention, the preparation method of the nanoparticles includes: preparing a mixed aqueous solution containing ferric chloride, hyaluronidase and polyvinyl pyrrolidone, slowly dropping the mixed aqueous solution into the aqueous solution of pentagalloylglucose, stirring to react, and separating the reaction product from solid and liquid, and collecting the solid part; preferably, in the mixed aqueous solution, the concentration of ferric chloride is 0.16 mg / mL, the concentration of hyaluronidase is 0.6 mg / mL, and the concentration of polyvinyl pyrrolidone is 1 mg / mL; preferably, the concentration of the aqueous solution of pentagalloylglucose is 0.5 mg / mL; preferably, the volume ratio of the mixed aqueous solution to the aqueous solution of pentagalloylglucose is 1:1; preferably, the dropping speed is 1-2 mL / min, and the stirring time is 5-10 min. Preferably, solid-liquid separation can be achieved by centrifugation, the centrifugal speed is 6500-9000 rpm, the time is 2-5 min, and then resuspended with 2-4 mL of deionized water, centrifuged under the same conditions, the supernatant is removed, and the precipitate is collected. Preferably, pentagalloylglucose can be first dissolved in ethanol (30 mM) and then diluted into an aqueous solution.

[0011] As a specific embodiment of the present invention, the mass percentage of the drug in the nanoparticles is 50%-60%.

[0012] As a specific embodiment of the present invention, the molar ratio of the metal ion chelating agent to the metal ions in the carrier is 2 to 3:1; and the chelating efficiency is 95% to 98%.

[0013] As a specific embodiment of the present invention, the metal ion chelator is deferoxamine mesylate.

[0014] Another aspect of the present invention provides the use of the aforementioned spatiotemporally controllable drug delivery system in drug delivery targeting the extracellular matrix of tumor cells.

[0015] Specifically, after the nanoparticles are used for systemic administration, metal ion chelators are used to disassemble the tumor extracellular matrix of the nanoparticles at a specific point when the tumor accumulation reaches a peak, and the drug can be released in a controlled manner. When used for in vivo administration in tumor-bearing mice, nanoparticles (PPFH) loaded with hyaluronidase using a polyvinyl pyrrolidone-modified metal iron ion polyphenol network as a carrier can achieve optimal intratumoral accumulation 24 hours after intravenous injection. At this time, injection of deferoxamine mesylate (DFO) (15-20 mg / kg) can achieve the optimal specific release efficiency. DFO can degrade the Fe in PPFH. 3+ Chelation removal destroys the coordination structure in PPFH and releases Fe 3+ The physical barrier of MPN blocks HAase, rapidly releases the drug, and effectively prevents cellular endocytosis. When PPFH is used for intravenous injection, it is fully resuspended in 2 mL of isotonic solution, preferably 5% glucose solution.

[0016] On the other hand, the present invention provides the application of the aforementioned spatiotemporally controllable drug delivery system in the preparation of drugs for photodynamic therapy. The drug is a drug for enhancing the treatment of tumors by photodynamic therapy, or a drug for enhancing the improvement of the immunosuppressive tumor microenvironment by photodynamic therapy.

[0017] The beneficial effects of the present invention are as follows:

[0018] 1. The delivery system provided by the present invention consists of two parts: nanoparticles loaded with drugs acting on the tumor extracellular matrix and metal ion chelators. Among them, the nanoparticles have good drug-loading capacity, can encapsulate drug activity, reduce drug immunogenicity, and have good safety; they can target tumor sites and can be specifically disassembled and release drugs by metal ion chelators in the tumor extracellular matrix, degrade the tumor extracellular matrix, enhance blood perfusion, relieve tumor hypoxia, promote drug penetration and T cell infiltration to reshape the tumor microenvironment. Furthermore, it can effectively enhance the intratumoral accumulation and therapeutic effect of subsequent PDT drugs, kill tumors to the greatest extent, and play a bridging role between PDT and immunotherapy. The targeting evaluation and controlled release evaluation carried out on BALB / c female tumor-bearing mice using 4T1 breast cancer as a model confirmed that the delivery strategy of the combination of nanoparticles and metal ion chelators of the present invention has excellent ability to improve the TME and can significantly enhance the PDT efficacy.

[0019] 2. The delivery system provided by the present invention is applicable to the delivery of various drugs acting on the outside of tumor cells, such as HAase, tyrosine kinase inhibitors, immune checkpoint inhibitors, etc., and has general applicability. Description of the Drawings

[0020] Figure 1A : Inhibition of HAase activity determined by DNS method;

[0021] Figure 1B : Transmission electron microscopy (TEM) characterization before and after loading HAase;

[0022] Figure 1C : Infrared wavelength scanning of each group;

[0023] Figure 1D : Particle size change before and after adding DFO;

[0024] Figure 1E : Absorption peak change before and after adding DFO;

[0025] Figure 1F : Absorption intensity change at 600 nm before and after adding DFO;

[0026] Figure 1G : Element content change before and after adding DFO;

[0027] Figure 1H: SDS-PAGE gel electrophoresis;

[0028] Figure 1I : Comparison of different response release systems;

[0029] Figure 1J : Detection by flow cytometry;

[0030] Figure 1K : Determination of HAase activity released after DFO disassembly by DNS method;

[0031] Figure 1L : Comparison of enzyme activities in each group;

[0032] Figure 2A : Hemolytic study;

[0033] Figure 2B : Effects on liver and kidney functions;

[0034] Figure 2C : Tumor targeting of PPFH;

[0035] Figure 2D : Organ distribution of PPFH;

[0036] Figure 3A : Observation of tumor tissue section by scanning electron microscope;

[0037] Figure 3B : HE staining results of tumor tissue sections;

[0038] Figure 3C : Observation by immunofluorescence staining;

[0039] Figure 3D : Detection of tumor tissue hypoxia;

[0040] Figure 3E : Verification of infiltration of immune cells in tumor microenvironment by flow cytometry;

[0041] Figure 4A : Imaging of drug accumulation over time;

[0042] Figure 4B : Ex vivo imaging of tumor tissue after 24 h;

[0043] Figure 4C : Tumor fluorescence quantification results;

[0044] Figure 4D : Permeation of IR780 liposomes;

[0045] Figure 5A : HE staining results;

[0046] Figure 5B : Immunofluorescence staining results;

[0047] Figure 6A : Curves of tumor size in each group changing with time;

[0048] Figure 6B : Tumor imaging in each group;

[0049] Figure 6C : Ex vivo imaging of tumors in each group;

[0050] Figure 6D : Comparison of tumor inhibition rates in each group;

[0051] Figure 7 For histological evaluation of anti-tumor effect;

[0052] Figure 8 For the research results of the improvement of the immune microenvironment by the present invention. Detailed implementation manners

[0053] To make the objectives, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are some but not all of the embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0054] Example 1: PPFH 3000 Synthesis of nanoparticles

[0055] Add 1 ml of PVP (5 mg / mL) aqueous solution, 80 μL of FeCl 3 (10 mg / mL) aqueous solution and 1 ml of HAase (3 mg / mL, enzyme activity 3000 U / mg) aqueous solution into 3 ml of deionized water. After stirring at room temperature for 30 min, it is gradually added dropwise to 5 mL of PGG (0.5 mg / mL) aqueous solution, stirred for 10 min, centrifuged at 7500 rpm for 2 min to remove excess reactants, and redispersed and rinsed once with 4 ml of deionized water and centrifuged again. The precipitate obtained is PPFH 3000 .

[0056] Example 2: PPFH 300 Synthesis of nanoparticles

[0057] Add 1 ml of PVP (5 mg / mL) aqueous solution, 80 μL of FeCl 3(10 mg / mL) aqueous solution and 1 mL of HAase (3 mg / mL, enzyme activity 300 U / mg) aqueous solution were stirred at room temperature for 30 min and then added dropwise to 5 mL of PGG (0.5 mg / mL) aqueous solution, stirred for 10 min, centrifuged at 7500 rpm for 2 min to remove excess reactants, resuspended and rinsed once with 4 mL of deionized water and centrifuged again to obtain the precipitate, which was PPFH. 300 。

[0058] Example 3: Characterization of PPFH

[0059] The inhibition of HAase activity was determined using the dinitrosalicylic acid method (DNS method). As Figure 1A shown, after HAase was co-incubated with Fe 3+ and PVP in the carrier material, the inhibition of its activity (the reaction concentration of Fe 3+ in Examples 1 and 2 was 160 μg / ml), and the results showed that the separate carrier material, especially Fe 3+ , had a strong inhibitory effect on HAase. It indicated that the carrier material had a strong blocking effect on HAase and could improve the biosafety of the nanoparticles.

[0060] The PPFH nanoparticles obtained in Example 1 or 2 were characterized by transmission electron microscopy (TEM). As Figure 1B shown, the nanoparticles had a typical reticular structure with a size between 200 and 300 nm. Compared with the empty carrier PPF without HAase, PPFH had a deeper and fuller color.

[0061] The PPFH nanoparticles obtained in Example 1 or 2 and PVP and HAase alone were scanned at infrared wavelengths. As Figure 1C shown, it showed effective modification of PVP (characteristic water peak at 3500 nm, characteristic absorption peak of C=O at 1680 nm, and fingerprint spectrum at 1000 - 1500 nm).

[0062] Example 4: In vitro dynamic disassembly study of PPFH

[0063] The disassembly effect of the PPFH nanoparticles obtained in Example 1 after using DFO was verified in vitro. The particle size changes, ultraviolet absorption, and elemental content changes before and after adding or not adding DFO were detected by dynamic light scattering, ultraviolet wavelength scanning, and TEM mapping. As Figure 1D shown, the particle size of PPFH was 220 ± 3 nm, and particle size transformation occurred after adding DFO and destruction, indicating the destruction of its nanostructure; as Figure 1E shown, PPFH had a strong absorption peak of Fe-O bond near 600 nm, and the characteristic peak disappeared after adding DFO. As Figure 1FThe figure shows the curve of the absorption intensity at 600 nm changing with time after adding DFO, indicating that DFO can rapidly destroy its structure and release the drug; as Figure 1G shown, the elements O, N, C, Fe, and S in PPFH. The S element indicates effective encapsulation of HAase, and the Fe element significantly decreases after adding DFO, indicating its effective chelation. SDS-PAGE gel electrophoresis was used to verify the release of HAase. As Figure 1H shown, lane 1 is free HAase at 2 mg / ml, lane 2 is the PPFH precipitate without DFO disruption, lane 3 is the supernatant without DFO disruption, lane 4 is the precipitate after DFO disruption, and lane 5 is the supernatant after DFO disruption. The staining results show that there is no HAase band in lane 3, and an obvious HAase band of the same size as the free group can be seen in lane 5, indicating that DFO can effectively release HAase from PPFH. High-performance liquid chromatography was used to detect the PGG release ability in different response systems. As Figure 1I shown, the content of PGG in the supernatant was detected after incubating the nanoparticles in different response release systems of PBS (pH 5.5), GSH, and PDT for 30 min. It shows that DFO has a more significant disassembly efficiency compared to low pH, while rapid response release is hardly achievable using pH.

[0064] The HAase used in Example 1 was labeled with FITC fluorescence. PPFH-FITC was prepared in the same way. After co-incubating with breast cancer 4T1 cells with or without DFO for 4 h, the FITC fluorescence intensity of the cells was detected by flow cytometry. As Figure 1J shown, after using DFO, the uptake of HAase by the cells is lower, only one-third of that without DFO. It shows the feasibility of the present invention for extracellular delivery.

[0065] The low-activity and high-activity PPFH obtained in Examples 1 and 2 300 and PPFH 3000 nanoparticles were used as controls for each other, and the DNS method was used to determine the activity of HAase released after DFO disassembly. As Figure 1K ~Figure 1l shown, G1-G5 are free standard samples, G6 is the enzyme activity of PPFH 300 without adding DFO, G7 is the enzyme activity of the precipitate of PPFH 300 +DFO, G8 is the enzyme activity of the supernatant of PPFH 300 +DFO, G9 is the enzyme activity of PPFH 3000 without adding DFO, G10 is the enzyme activity of the precipitate of PPFH 3000 +DFO, G11 is the enzyme activity of the supernatant of PPFH 3000 +DFO. The results show that PPFH 300 and PPFH 3000There was no significant difference in enzyme activity, indicating that its activity could be effectively blocked. After adding DFO, PPFH 3000 could release a large amount of active HAase. Comprehensive experiments demonstrated the feasibility, effectiveness, and safety of the PPFH+DFO delivery strategy.

[0066] Example 5: In vivo safety evaluation

[0067] The hemolytic property of PPFH nanoparticles was investigated. As Figure 2A shown, the PPFH nanoparticles in the present invention did not cause hemolytic reactions when used for systemic administration. Further, one week after PPFH+DFO administration, the blood of mice was collected to detect ALT, AST, ALP, UREA, and CREA to evaluate the impact of the drug on liver and kidney functions. The results were as Figure 2B shown. Similar to the blank control mice, all the indicators were within the normal range, indicating that this treatment strategy had no significant impact on liver and kidney functions and had high safety.

[0068] Example 6: In vivo tumor targeting study of PPFH

[0069] PPFH was fluorescently labeled with IR780 and administered to female BALB / c mice bearing 4T1 breast cancer tumors. In vivo imaging was used to observe tumor targeting and organ distribution. As Figure 2C - 2D shown, PPFH had good tumor targeting performance and accumulation behavior, which was enhanced over time and reached a peak at 24 - 36 h. After sacrificing the mice at 36 h, the drug distribution in various organs was observed ex vivo. The results showed that its distribution in the main organs was relatively low, significantly lower than that in the tumor tissue, indicating the good tumor targeting performance of PPFH in the present invention.

[0070] Example 7: Application of PPFH+DFO in ECM degradation and TME improvement

[0071] The nanoparticles obtained in Example 1 or 2 were administered to female BALB / c mice bearing 4T1 breast cancer tumors. PPFH 300 and PPFH 3000 were used as controls for each other to highlight the role of HAase, and the PBS group was used as a blank control. After 24 h, DFO was injected. The mice were sacrificed 24 h after the end of DFO administration, and the tumor tissues were taken to study the degradation of tumor ECM by HAase. As Figure 3A shown, by observing the cross-section of tumor tissues through scanning electron microscopy, the results showed that compared with the PBS group and the PPFH 300 +DFO low-activity group, obvious honeycomb-like pores appeared inside the tumors in the PPFH 3000 +DFO group, indicating that HAase could play a role at the tumor site and the tumor stroma was effectively degraded. HE staining was performed on tumor tissue sections. As Figure 3B shown, compared with the PBS group and PPFH300 Compared with the PPFH+DFO group 3000 The PPFH+DFO group showed a looser cell structure; immunofluorescence staining of α-smooth muscle actin (α-SMA) and collagen in tumor tissue sections was performed to observe whether the degradation effect of HAase disrupted other components in ECM formation and its effect on the expression of hypoxia-inducible factor-1α (HIF-1α). The results were as Figure 3C shown, the PPFH 3000 +DFO group showed lower contents of α-SMA (red fluorescence) and collagen (green fluorescence), as well as lower expression of HIF-1α (red fluorescence) compared with the PBS group and the PPFH 300 +DFO group. Similarly, the hypoxia situation in tumor tissue was detected using the hypoxia probe pimonidazole. As Figure 3D shown, the PBS group and the PPFH 300 +DFO group showed stronger hypoxia (green fluorescence) compared with the PPFH 3000 +DFO group, while the PPFH 3000 +DFO showed obvious improvement in hypoxia. It indicates that the present invention can significantly destroy the tumor ECM structure and improve the tumor hypoxia situation. Hypoxia is an important factor in the immunosuppressive microenvironment. Further study on the improvement of the immunosuppressive tumor microenvironment after ECM degradation. After digesting the above-mentioned tumor tissue into single-cell suspensions, immune cells were fluorescently labeled: cytotoxic T cells (CD3 + , CD8 + ), myeloid-derived suppressor cells MDSC (CD45 + , CD11b + , Gr-1 + ), M2 macrophages (CD45 + , CD11b + , F4 / 80 + ). Flow cytometry was used to verify the infiltration of immune cells in the tumor microenvironment. The results were as Figure 3E shown, the PPFH 3000 +DFO group had enhanced CD8 infiltration, reduced MDSC, and no significant change in M2 macrophages, indicating that the present invention can enhance anti-tumor immunity to a certain extent, improve the TME, and relieve the immunosuppressive microenvironment.

[0072] Example 8: Study on the effect of PPFH+DFO on drug accumulation

[0073] Further study was conducted to determine whether the loosened ECM could enhance drug penetration and improve drug accumulation. After 24 hours of administering DFO as in Example 7, the model photosensitizer IR780 liposomes were intravenously injected, and in vivo imaging technology was used to observe the accumulation behavior of IR780 liposomes. Figure 4AFor imaging of drug accumulation over time, Figure 4B For ex vivo imaging of tumor tissues after 24 h, Figure 4C For tumor fluorescence quantification results, showing PPFH 3000 + DFO group had an enhancing effect on drug accumulation, significantly higher than the other two groups. For frozen sections of tumor tissues, the penetration of IR780 liposomes was observed, and the results were as Figure 4D shown. Similarly, PPFH 3000 + DFO showed a larger area of IR780 fluorescence. The results indicated that the present invention could enhance drug accumulation in solid tumors to enhance the therapeutic effect.

[0074] Example 9: Evaluation of the photosensitizing effect of PPFH + DFO on PDT

[0075] Further on the basis of Example 8, photodynamic therapy was carried out 12 h after administration of IR780 liposomes (model 808 nm laser, 1 W / cm 2 , 10 min / animal). The mice were sacrificed 24 h after the treatment ended, and the tumors were taken for histological analysis. As Figure 5A - 5B shown, i was the PBS group, ii was the single PDT group, iii was the PPFH 300 + DFO + PDT group, iv was the PPFH 3000 + DFO + PDT group. As Figure 5A shown, the HE staining results showed that compared with the dense cell structure in other groups, group iv was looser, the cell nuclei showed shrinkage, indicating an apoptotic state. Calreticulin (CRT) exposure was used as a marker of immunogenic cell death (ICD), and immunofluorescence staining was performed on it. The results were as Figure 5B shown. Compared with other groups, group iv showed more obvious green fluorescence of CRT, and cell rupture was visible. It was shown that the present invention could significantly enhance the therapeutic effect of PDT.

[0076] Example 10: Evaluation of anti-tumor application

[0077] On the basis of Example 9, the specific anti-tumor effect of the present invention was further studied. It was divided into G1: PBS, G2: PDT, G3: PPFH 300 + DFO + PDT, G4: PPFH 3000 + DFO, G5: PPFH 3000 + PDT, G6: PPFH 3000 + DFO + PDT, a total of 6 groups for anti-tumor treatment. As Figure 6A - 6D shown, compared with the PBS group, the PDT group significantly inhibited tumor growth (the tumor inhibition rate was 63.7%), and one was completely cured. Alone PPFH 3000+DFO treatment only showed a weak tumor suppression effect (tumor inhibition rate: 31.0%), PPFH 300 +DFO + PDT hardly enhanced the anti-tumor effect of PDT, indicating that hyaluronidase played an important role in it, PPFH 3000 +PDT enhanced the therapeutic effect of PDT to a certain extent (tumor inhibition rate: 72.7%, two were completely cured), while PPFH 3000 +DFO + PDT group further enhanced the efficacy of PDT, with a tumor inhibition rate as high as 97.1%, and 4 were completely cured. HE staining and TUNEL staining were performed on tumor tissues to evaluate cell apoptosis. As Figure 7 shown, HE staining showed that G6 exhibited more obvious cell apoptosis morphology compared with other groups, and TUNEL staining also found a large area of positive regions in G6, which was much higher than other groups. It showed that the treatment strategy in the present invention had excellent anti-tumor effects.

[0078] Example 11: Study on improving the immunosuppressive tumor microenvironment

[0079] After the treatment in Example 10, the immune cells in the tumor tissues of mice were stained, including cytotoxic T cells (CD3 + , CD8 + ), myeloid-derived suppressor cells MDSC (CD45 + , CD11b + , Gr-1 + ), M2 macrophages (CD45 + , CD11b + , F4 / 80 + ), and dendritic cells DCs (CD11c + , CD80 + , CD86 + ). The results were as Figure 8 shown. Compared with the PBS group, the PDT group significantly enhanced the infiltration of DCs and CD8 cells. However, at the same time, it also enhanced the infiltration of MDSCs and M2 macrophages. Compared with PDT alone, PPFH 3000 +DFO + PDT group further and significantly enhanced the infiltration of DCs and CD8 cells, and reduced the infiltration of MDSCs and M2 macrophages, and the effect was better than other groups. The results proved that this strategy successfully reshaped the immunosuppressive tumor microenvironment, activated anti-tumor immunity, and established a bridge between PDT and immunotherapy.

[0080] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solutions of the present invention, and these simple modifications all belong to the protection scope of the present invention.

Claims

1. A spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix, characterized in that, the delivery system comprises: i) nanoparticles loaded with drugs using a metal polyphenol network modified with polyvinylpyrrolidone; and ii) a metal ion chelator; the metal polyphenol network is a porous network structure material formed by the coordination connection of ferric ions and pentagalloyl glucose; the metal ion chelator is deferoxamine mesylate; the nanoparticles are used for targeted tumor delivery of drugs; the metal ion chelator is used to disassemble the nanoparticles when they reach the tumor extracellular matrix to release the drugs.

2. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 1, characterized in that, the drug is selected from any one of hyaluronidase, tyrosine kinase inhibitor, and immune checkpoint inhibitor.

3. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 2, characterized in that, the drug is hyaluronidase.

4. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 1, characterized in that, the preparation method of the nanoparticles comprises: preparing a mixed aqueous solution containing ferric chloride, hyaluronidase and polyvinylpyrrolidone, slowly dropping the mixed aqueous solution into the aqueous solution of pentagalloyl glucose, stirring to react, and separating the solid and liquid of the reaction product, and collecting the solid part.

5. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 4, characterized in that, in the mixed aqueous solution, the concentration of ferric chloride is 0.16 mg / mL, the concentration of hyaluronidase is 0.6 mg / mL, and the concentration of polyvinylpyrrolidone is 1 mg / mL.

6. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 4, characterized in that, the concentration of the aqueous solution of pentagalloyl glucose is 0.5 mg / mL.

7. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 4, characterized in that, the volume ratio of the mixed aqueous solution to the aqueous solution of pentagalloyl glucose is 1:

1.

8. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 4, characterized in that, the dropping rate is 1 - 2 mL / min, and the stirring time is 5 - 10 min.

9. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 1, characterized in that, the mass percentage content of the drug in the nanoparticles is 50% - 60%.

10. The spatiotemporally controllable drug delivery system targeting the tumor extracellular matrix according to claim 1, characterized in that, the molar ratio of the metal ion chelator to the metal ions in the carrier is 2 - 3:1; the chelation efficiency is 95% - 98%.

11. Use of the spatiotemporally controllable drug delivery system according to any one of claims 1 to 10 in drug delivery targeting the tumor extracellular matrix.

12. Use of the spatiotemporally controllable drug delivery system according to any one of claims 1 to 10 in the preparation of drugs for photodynamic therapy.

13. According to the application described in claim 12, Characterized in that, the drug is a drug for enhancing the treatment of tumors by photodynamic therapy, or a drug for enhancing photodynamic therapy to improve the immunosuppressive tumor microenvironment.

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