Metallic polyphenol network modified drug-loaded radioactive microspheres, and preparation method and application thereof

Abstract

The application discloses a metal polyphenol network modified drug-loaded radioactive microsphere and a preparation method and application thereof. A radionuclide is labeled on different microspheres through a precipitation method or an adsorption method, and a metal polyphenol network is modified on the surface of the radioactive microsphere to improve the radionuclide labeling stability and the ability of synchronous drug loading. Subsequently, the drug molecules can be adsorbed on the modified radioactive microsphere to obtain the metal polyphenol network modified drug-loaded radioactive microsphere. The metal polyphenol network modified drug-loaded radioactive microsphere prepared by the application can solve the problems of radionuclide non-target shedding and DNA damage repair caused by nuclear medicine, avoid secondary drug administration, ensure that the drug directly and accurately reaches the tumor target, and further promote the selective internal irradiation treatment based on the radioactive microsphere mediation to play a more effective tumor treatment effect.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of drug-loaded microspheres, in particular to a kind of metal polyphenol network modified drug-loaded radioactive microspheres and its preparation method and application. BACKGROUND

[0002] Radioactive microspheres are mainly used for selective internal radiation therapy (SIRT). As a means of local radiotherapy, SIRT has been widely used for palliative treatment or preoperative down-staging treatment of unresectable primary or metastatic liver cancer. The core of SIRT technology lies in the dual blood supply system of the liver, in which the portal system is mainly responsible for providing blood to normal liver tissue, while the high-density hepatic artery transports more than 85% of blood to tumor tissue. This technology injects microspheres loaded with radionuclides (radioactive microspheres) into the blood supply artery of the tumor, uses the radionuclides loaded on the microspheres to release beta rays, and achieves close-range irradiation of tumor cells, thereby achieving effective treatment. Compared with external radiotherapy, SIRT is expected to cause minimal radiation damage to normal tissues.

[0003] Currently, there are three commercial radioactive microspheres for SIRT of liver cancer, namely 90 Y-glass microspheres (TheraSphere), 90 Y-resin microspheres (SIR-Spheres), and 166 Ho-polylactic acid microspheres (QuiremSpheres). However, 90 The preparation process of Y-glass microspheres is complex, the yield is low, and the production cost is high; 90 The radionuclide of Y-resin microspheres is easy to fall off, which reduces the specific activity and affects the treatment effect, and both 90 Y microspheres cannot be monitored by imaging technology for in vivo distribution and dose evaluation. 166 The weak beta energy of Ho-polylactic acid microspheres may not be conducive to the effective killing of tumor cells, and the short half-life is not conducive to long-distance transportation and storage. Moreover, the beta rays released by the three types of radioactive microspheres not only kill tumor cells but also induce DNA double-strand breaks (DSBs), triggering a series of cellular DNA damage responses (DDRs). These responses will pause the cell cycle and attempt to repair DNA damage, thereby reducing the killing effect of beta rays on tumors to some extent. Therefore, how to improve the treatment effect while reducing damage to normal tissues is a problem that needs to be solved in the current SIRT treatment field. SUMMARY

[0004] To solve the above technical problems, the application provides a metal polyphenol network modified drug-loaded radioactive microsphere and a preparation method and application thereof, a radionuclide is labeled on different microspheres through a precipitation method or an adsorption method, and a metal polyphenol network is modified on the surface of the radioactive microsphere to improve the radionuclide labeling stability and the ability of synchronous drug loading, and subsequently, drug molecules can be adsorbed on the modified radioactive microsphere to obtain the metal polyphenol network modified drug-loaded radioactive microsphere.

[0005] The application is achieved by the following technical solutions:

[0006] The application provides a preparation method of the metal polyphenol network modified drug-loaded radioactive microsphere, including the following steps:

[0007] (1) adding an inorganic salt containing a radionuclide and a precipitant into an inorganic microsphere dispersion liquid, and obtaining radioactive microspheres after labeling is completed; or

[0008] adding an inorganic salt containing a radionuclide into an organic microsphere dispersion liquid, and obtaining radioactive microspheres after labeling is completed;

[0009] (2) mixing the radioactive microspheres obtained in step (1) with a metal salt solution and a polyphenol solution to obtain metal polyphenol network modified radioactive microspheres;

[0010] (3) mixing the metal polyphenol network modified radioactive microspheres obtained in step (2) with a drug solution to obtain the metal polyphenol network modified drug-loaded radioactive microspheres.

[0011] Further, in step (1), the inorganic microspheres of the inorganic microsphere dispersion liquid can be silica microspheres, and the organic microspheres of the organic microsphere dispersion liquid can be sodium alginate microspheres.

[0012] Further, in step (1), the radionuclide is selected from one or more of 177 Lu, 99 mTc, 32 P, 125 I, 131 I and 255 Ac.

[0013] In the specific embodiment, the 177 Lu radionuclide capable of emitting β particles and low-energy γ rays is selected to label inorganic silica microspheres (SiMS) and organic sodium alginate microspheres (AlgMS) to obtain radioactive microspheres ( 177 Lu-SiMS or 177 Lu-AlgMS), which solves the shortcoming that the traditional radioactive microspheres cannot be used for both treatment and imaging. For the problem of easy falling of the radionuclide, the metal polyphenol network with fast film forming speed and high stability is used to label the radioactive microspheres, and the drug molecules are adsorbed on the modified radioactive microspheres to obtain the metal polyphenol network modified drug-loaded radioactive microspheres.177 Lu-SiMS or 177 The surface modification of Lu-AlgMS or

[0014] Further, in step (1), the concentration of inorganic microspheres in the inorganic microsphere dispersion liquid is 2-6 mg / mL.

[0015] Further, in step (1), the precipitant is K3PO4.

[0016] Further, in step (2), the use amount ratio of the radioactive microspheres, the metal salt solution and the polyphenol solution is 1 mg:(5-40) μL:(5-40) μL.

[0017] Further, in step (2), the volume ratio of the metal salt solution and the polyphenol solution is 1:(1-2).

[0018] Further, in step (2), the metal salt of the metal salt solution is selected from one or more of hafnium salt, iron salt, manganese salt, copper salt, cobalt salt and vanadium salt, preferably hafnium salt, iron salt and manganese salt.

[0019] Among them, hafnium metal element, as a highly potential high-Z (high atomic number) metal, can interact with ionizing radiation significantly, and play a radiosensitizing role; iron metal element can activate ferroptosis pathway to induce cell apoptosis through Fenton reaction; accumulation of manganese metal element can improve the sensitivity of cGAS to double-stranded DNA (dsDNA), thereby activating cGAS-STING pathway and enhancing the effect of immunotherapy. Based on the introduction of the above metal polyphenol network, radionuclides can exhibit great advantages in SIRT application, which can maximize the therapeutic effect while minimizing the dose.

[0020] Further, in step (2), the metal salt of the metal salt solution is HfCl4, FeCl3 or MnCl2.

[0021] Further, in step (2), the concentration of metal salt in the metal salt solution is 5-20 mM.

[0022] Further, in step (2), the polyphenol in the polyphenol solution is selected from one or more of tannic acid, gallic acid, protocatechuic aldehyde, pyrogallol and catechol, preferably tannic acid (TA).

[0023] Further, in step (2), the concentration of polyphenol in the polyphenol solution is 5-20 mM.

[0024] Preferably, in step (2), the concentration of the metal salt in the metal salt solution is the same as the concentration of the polyphenol in the polyphenol solution.

[0025] Further, in step (3), the ratio of the use amount of the metal polyphenol network modified radioactive microspheres to the drug solution is 1 mg:(200-600) μL.

[0026] Further, in step (3), the drug of the drug solution is an ATR inhibitor, doxorubicin hydrochloride (DOX), Evofosfamide (TH-302), Toyocamycin, Atovaquone, a pan-Bcl-2 inhibitor, etc., and the pan-Bcl-2 inhibitor is Sabutoclax.

[0027] Wherein, the ataxia telangiectasia mutated and Rad3-related kinase (ATR) is a key regulator of DNA damage response (DDRs), responsible for sensing replication stress (RS) and sending it to the S and G2 / M checkpoints to facilitate DNA damage repair introduction. The present application takes advantage of the large adsorption capacity and high adsorption efficiency of the metal polyphenol network, and obtains metal polyphenol network modified drug-loaded radioactive microspheres by loading ATR inhibitors (ATRi), to solve the problems of DNA damage repair caused by nuclear drugs, avoid secondary administration, ensure that the drug directly and accurately reaches the tumor target, and further promote the SIRT based on the radioactive microsphere mediated to play a more effective tumor treatment effect.

[0028] Further, in step (3), the concentration of the drug in the drug solution is 10-600 μM.

[0029] The present application protects the metal polyphenol network modified drug-loaded radioactive microspheres prepared by the above method.

[0030] The present application also protects the application of the above metal polyphenol network modified drug-loaded radioactive microspheres in the preparation of drugs for tumor diagnosis and treatment.

[0031] The present application has the following beneficial effects:

[0032] The present application constructs a metal polyphenol network modified radioactive microsphere, which not only significantly improves the labeling stability of the radioactive microsphere, but also effectively enhances the radiotherapy effect of radionuclides on tumors by virtue of the introduction of metal. At the same time, thanks to the excellent adsorption capacity and high efficiency of the metal polyphenol network, drugs can be successfully loaded on the metal polyphenol network modified radioactive microsphere, thereby eliminating the tumor locally while inhibiting the DNA self-repair mechanism of tumor cells triggered by radiation damage, thereby preventing tumor recurrence. In addition, the present application also combines radiotherapy and ATRi mediated immune response, which effectively inhibits the metastasis of tumors and the growth of distant metastatic tumors under the synergistic effect, thereby opening up a new way for tumor treatment. BRIEF DESCRIPTION OF DRAWINGS

[0033] Figure 1 Preparation for example 1 177 The flowchart of Lu-SiMS@HTi.

[0034] Figure 2 Preparation for example 1 177 The optical microscope images of Lu-SiMS@HT; wherein, (a) is SiMS, (b) is 177 Lu-SiMS@HT.

[0035] Figure 3 Preparation for example 1 177 The scanning electron microscope (SEM) images of Lu-SiMS@HT; wherein, (a) is SEM image, (b) is Mapping image of Hf element.

[0036] Figure 4 Preparation for example 1-5 177 The actual object images of Lu-SiMS@HT; wherein, from right to left, they are example 2, example 1, example 3, example 4, example 5.

[0037] Figure 5 Preparation for example 1 177 The optical microscope images of Lu-SiMS@HT; wherein, (a) is SiMS, (b) is 177 Lu-SiMS@HT.

[0038] Figure 6 Preparation for example 1 177 The scanning electron microscope (SEM) images of Lu-SiMS@HT; wherein, (a) is SEM image, (b) is Mapping image of Hf element.

[0039] Figure 7 Preparation for example 1 177Lu-SiMS@FT, AlgMS in Example 9 and 177 Optical microscope images of Lu-AlgMS@FT; wherein, (a) is SiMS, (b) is 177 Lu-SiMS@FT, (c) is AlgMS, (d) is 177 Lu-AlgMS@FT.

[0040] Figure 8 SiMS in Example 8 and 177 Lu-SiMS@FT and AlgMS in Example 9 and 177 Scanning electron microscope images of Lu-AlgMS@FT; wherein, (a) is SiMS, (b) is 177 SEM images of Lu-SiMS@FT, (b) is 177 Mapping images of Fe element in Lu-SiMS@FT, (c) is 177 SEM images of Lu-AlgMS@FT, (d) is 177 Mapping images of Fe element in Lu-AlgMS@FT.

[0041] Figure 9 SiMS in Example 10 and 177 Lu-SiMS@MT and AlgMS in Example 11 and 177 Mapping images of Mn element in Lu-AlgMS@MT; wherein, (a) is SiMS, (b) is 177 Lu-SiMS@MT, (b) is 177 Lu-AlgMS@MT.

[0042] Figure 10 SiMS in Example 1 and 177 Lu-SiMS and 177 Results of label stability test of Lu-SiMS@HT in saline and 10% fetal bovine serum (FBS) environments.

[0043] Figure 11 Lu-AlgMS in Example 7 and 177 Lu-AlgMS and 177 Results of label stability test of Lu-AlgMS@HT in 10% fetal bovine serum (FBS) environment.

[0044] Figure 12 Comparison of loading capacity of SiMS and SiMS@HT for ATRi.

[0045] Figure 13 Comparison of loading capacity of AlgMS and AlgMS@HT for ATRi.

[0046] Figure 14 is the SiMS in Example 1, 177 Lu-SiMS, 177 Lu-SiMS@HT and 177 Results of Lu-SiMS@HTi cell viability and apoptosis experiments on RH35 cells.

[0047] Figure 15 is the SiMS in Example 1, 177 Lu-SiMS, 177 Lu-SiMS@HT and 177 H&E, Ki67, and TUNEL staining data of Lu-SiMS@HTi on animal-level liver tumors. DETAILED DESCRIPTION

[0048] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this invention pertains. The terms used herein in the specification of the present invention are for the purpose of describing specific embodiments only and are not intended to limit the present invention. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0049] The present invention will be further described below with reference to the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it. However, the embodiments are not intended to limit the present invention.

[0050] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the materials and reagents used are all commercially available unless otherwise specified.

[0051] Example 1

[0052] A metal polyphenol network modified drug-loaded radioactive microsphere ( 177 The preparation method of Lu-SiMS@HTi comprises the following steps:

[0053] (1) Disperse 1 mg of SiMS in 0.5 mL of deionized water and add 10 μL of medical 177 LuCl3 solution (about 200 μCi) was shaken in a thermomixer for 5 minutes, and then K3PO4 (1M) buffer solution was added dropwise, and the pH was adjusted to 12 and the shaking was continued for 20 minutes. After the reaction was completed, the microspheres were centrifuged at 2000 rpm for 5 minutes and washed 3 times with deionized water to obtain 177 Lu-SiMS.

[0054] (2) 1mg 177After Lu-SiMS was fully mixed with 10 μL HfCl4 aqueous solution (10 mM) and 10 μL TA (10 mM) aqueous solution (pH = 6.8), the microspheres were washed with deionized water for 3 times to obtain 177 Lu-SiMS@HT.

[0055] (3) 1 mg of 177 After Lu-SiMS@HT was mixed with 300 uL 200 μM ATRi aqueous solution, it was placed for 2 h and washed with deionized water for 3 times to obtain 177 Lu-SiMS@HTi.

[0056] Figure 1 A flowchart for preparing 177 Lu-SiMS@HTi.

[0057] Figure 2 A flowchart for preparing 177 Optical microscope images of SiMS and 177 Lu-SiMS@HT. Figure 3 Scanning electron microscope (SEM) images of 177 Lu-SiMS@HT. Figure 2 It can be seen from Figure 3 that the hafnium tannic acid network is formed on the surface of the silica microspheres, and the Hf element exists.

[0058] Example 2

[0059] A method for preparing 177 Lu-SiMS@HTi, which is basically the same as example 1, the difference is that in step (2), 1 mg of 177 Lu-SiMS was fully mixed with 5 μL HfCl4 aqueous solution (10 mM) and 5 μL TA (10 mM) aqueous solution.

[0060] Example 3

[0061] A method for preparing 177 Lu-SiMS@HTi, which is basically the same as example 1, the difference is that in step (2), 1 mg of 177 Lu-SiMS was fully mixed with 20 μL HfCl4 aqueous solution (10 mM) and 20 μL TA (10 mM) aqueous solution.

[0062] Example 4

[0063] A method for preparing 177The preparation method of Lu-SiMS@HTi is basically the same as that of example 1, except that in step (2), 1 mg of Lu-SiMS is mixed with 30 μL of an aqueous solution of HfCl4(10 mM) and 30 μL of an aqueous solution of TA (10 mM). 177 The Lu-SiMS is mixed with 40 μL of an aqueous solution of HfCl4(10 mM) and 40 μL of an aqueous solution of TA (10 mM).

[0064] Example 5

[0065] A 177 The preparation method of Lu-SiMS@HTi is basically the same as that of example 1, except that in step (2), 1 mg of Lu-SiMS is mixed with 30 μL of an aqueous solution of HfCl4(10 mM) and 30 μL of an aqueous solution of TA (10 mM). 177 The Lu-SiMS is mixed with 40 μL of an aqueous solution of HfCl4(10 mM) and 40 μL of an aqueous solution of TA (10 mM).

[0066] Example 6

[0067] A 177 The preparation method of Lu-SiMS@HTi is basically the same as that of example 1, except that in step (2), 1 mg of Lu-SiMS is mixed with 30 μL of an aqueous solution of HfCl4(10 mM) and 30 μL of an aqueous solution of TA (10 mM). 177 The Lu-SiMS is mixed with 40 μL of an aqueous solution of HfCl4(10 mM) and 40 μL of an aqueous solution of TA (10 mM).

[0068] Figure 4 The preparation method of Lu-SiMS@HTi is basically the same as that of example 1, except that in step (2), 1 mg of Lu-SiMS is mixed with 30 μL of an aqueous solution of HfCl4(10 mM) and 30 μL of an aqueous solution of TA (10 mM). 177 The Lu-SiMS is mixed with 40 μL of an aqueous solution of HfCl4(10 mM) and 40 μL of an aqueous solution of TA (10 mM). Figure 5 As can be seen from the above table, when the aqueous solution of HfCl4and the aqueous solution of TA are added in excess, the mixed solution becomes turbid, which may be because the excess HfCl4and TA form aggregates, affecting the formation of the metal polyphenol network.

[0069] In addition, the Hf loading rate of Lu-SiMS@HT in examples 1-2 and example 6 is tested by inductively coupled plasma optical emission spectrometer (ICP-OES). 177 The Hf loading rate of Lu-SiMS@HT in example 1 is about 72%-80%. 177 The Hf loading rate of Lu-SiMS@HT in example 2 is about 60%-77%. 177 The Hf loading rate of Lu-SiMS@HT in example 6 is about 44%-60%. 177 The Hf loading rate of Lu-SiMS@HT in example 6 is about 44%-60%, which proves that the ratio of the aqueous solution of HfCl4and the aqueous solution of TA also affects the formation of the metal polyphenol network, especially the Hf loading rate, and the volume ratio of the aqueous solution of HfCl4and the aqueous solution of TA of the same concentration is preferably 1:1.

[0070] Example 7

[0071] A metal polyphenol network modified drug-loaded radioactive microsphere ( 177 The preparation method of Lu-AlgMS@HTi) comprises the following steps:

[0072] (1) Take 100 μL AlgMS dispersion (10 mg / mL) and add 10 μL medical 177 LuCl3 solution (about 200 μCi) was shaken in a constant temperature mixer for 2 h, centrifuged at 2000 rpm for 5 min, and the precipitated microspheres were washed 3 times with deionized water to obtain 177 Lu-AlgMS.

[0073] (2) 100 μL 177 Lu-AlgMS dispersion (10 mg / mL) was thoroughly mixed with 10 μL HfCl4 aqueous solution (10 mM) and 10 μL TA (10 mM) aqueous solution (pH = 6.8), and the microspheres were washed three times with deionized water to obtain 177 Lu-AlgMS@HT.

[0074] (3) 100 μL 177 Lu-AlgMS@HT dispersion (10 mg / mL) was mixed with 300 μL of 200 μM ATRi aqueous solution and allowed to stand for 2 h. The mixture was then washed three times with deionized water to obtain 177 Lu-AlgMS@HTi.

[0075] Figure 5 is the AlgMS in Example 7 and 177 Optical microscope images of Lu-AlgMS@HT; (a) is AlgMS, (b) is 177 Lu-AlgMS@HT. Figure 6 For Example 7 177 Scanning electron microscope (SEM) images of Lu-AlgMS@HT; (a) is the SEM image, and (b) is the Hf element mapping image. Figure 5 and Figure 6 It can be seen from the figure that the hafnium tannic acid network forms a film on the surface of the sodium alginate microspheres, as well as the presence of the Hf element.

[0076] Example 8

[0077] A metal polyphenol network modified drug-loaded radioactive microsphere ( 177 The preparation method of Lu-SiMS@FTD comprises the following steps:

[0078] (1) Disperse 1 mg of SiMS in 0.5 mL of deionized water and add 10 μL of medical 177LuCl3 solution (about 200 μCi) was shaken in a thermomixer for 5 minutes, and then K3PO4 (1M) buffer solution was added dropwise, and the pH was adjusted to 12 and the shaking was continued for 20 minutes. After the reaction was completed, the microspheres were centrifuged at 2000 rpm for 5 minutes and washed 3 times with deionized water to obtain 177 Lu-SiMS.

[0079] (2) 1mg 177 After Lu-SiMS was thoroughly mixed with 10 μL of FeCl3 aqueous solution (10 mM) and 10 μL of TA (10 mM) aqueous solution (pH = 6.8), the microspheres were washed three times with deionized water to obtain 177 Lu-SiMS@FT.

[0080] (3) 1mg 177 Lu-SiMS@FT was mixed with 100uL 200μM DOX aqueous solution and allowed to stand for 2h, then washed three times with deionized water to obtain 177 Lu-SiMS@FTD.

[0081] Example 9

[0082] A metal polyphenol network modified drug-loaded radioactive microsphere ( 177 The preparation method of Lu-AlgMS@FTD comprises the following steps:

[0083] (1) Take 100 μL AlgMS dispersion (10 mg / mL) and add 10 μL medical 177 LuCl3 solution (about 200 μCi) was shaken in a constant temperature mixer for 2 h, centrifuged at 2000 rpm for 5 min, and the precipitated microspheres were washed 3 times with deionized water to obtain 177 Lu-AlgMS.

[0084] (2) 100 μL 177 Lu-AlgMS dispersion (10 mg / mL) was thoroughly mixed with 10 μL of FeCl3 aqueous solution (10 mM) and 10 μL of TA (10 mM) aqueous solution (pH = 6.8), and the microspheres were washed three times with deionized water to obtain 177 Lu-AlgMS@FT.

[0085] (3) 100 μL 177 Lu-AlgMS@FT dispersion (10 mg / mL) was mixed with 100 μL of 200 μM DOX aqueous solution and allowed to stand for 2 h. The mixture was then washed three times with deionized water to obtain 177 Lu-AlgMS@FTD.

[0086] Figure 7For the SiMS in Example 8 and 177 Lu-SiMS@FT and AlgMS in Example 9 and 177 Optical microscope images of Lu-AlgMS@FT; (a) is SiMS, (b) is 177 Lu-SiMS@FT, (c) is AlgMS, (d) is 177 Lu-AlgMS@FT.

[0087] Figure 8 For Example 8 177 Lu-SiMS@FT and Example 9 177 Scanning electron microscope image of Lu-AlgMS@FT; (a) 177 SEM image of Lu-SiMS@FT, (b) 177 Mapping image of Fe element in Lu-SiMS@FT, (c) 177 SEM image of Lu-AlgMS@FT, (d) 177 Mapping image of Fe element in Lu-AlgMS@FT.

[0088] Example 10

[0089] A metal polyphenol network modified drug-loaded radioactive microsphere ( 177 The preparation method of Lu-SiMS@MTT comprises the following steps:

[0090] (1) Disperse 1 mg of SiMS in 0.5 mL of deionized water and add 10 μL of medical 177 LuCl3 solution (about 200 μCi) was shaken in a thermomixer for 5 minutes, and then K3PO4 (1M) buffer solution was added dropwise, and the pH was adjusted to 12 and the shaking was continued for 20 minutes. After the reaction was completed, the microspheres were centrifuged at 2000 rpm for 5 minutes and washed 3 times with deionized water to obtain 177 Lu-SiMS.

[0091] (2) 1mg 177 After Lu-SiMS was thoroughly mixed with 10 μL of MnCl2 aqueous solution (10 mM) and 10 μL of TA (10 mM) aqueous solution (pH = 6.8), the microspheres were washed three times with deionized water to obtain 177 Lu-SiMS@MT.

[0092] (3) 1mg 177 Lu-SiMS@MT was mixed with 100uL 200μM TH302 aqueous solution and allowed to stand for 2h, then washed three times with deionized water to obtain 177Lu-SiMS@MTT.

[0093] Example 11

[0094] A metal polyphenol network modified drug-loaded radioactive microsphere ( 177 The preparation method of Lu-AlgMS@MTT comprises the following steps:

[0095] (1) Take 100 μL AlgMS dispersion (10 mg / mL) and add 10 μL medical 177 LuCl3 solution (about 200 μCi) was shaken in a constant temperature mixer for 2 h, centrifuged at 2000 rpm for 5 min, and the precipitated microspheres were washed 3 times with deionized water to obtain 177 Lu-AlgMS.

[0096] (2) 100 μL 177 Lu-AlgMS dispersion (10 mg / mL) was thoroughly mixed with 10 μL of MnCl2 aqueous solution (10 mM) and 10 μL of TA (10 mM) aqueous solution (pH = 6.8), and the microspheres were washed three times with deionized water to obtain 177 Lu-AlgMS@MT.

[0097] (3) 100 μL 177 Lu-AlgMS@MT dispersion (10 mg / mL) was mixed with 100 μL of 200 μM TH302 aqueous solution and allowed to stand for 2 h. The mixture was then washed three times with deionized water to obtain 177 Lu-SiMS@MTT.

[0098] Figure 9 For Example 10 177 Lu-SiMS@MT and Example 11 177 Mapping image of Mn element in Lu-AlgMS@MT; (a) 177 Lu-SiMS@MT, (b) 177 Lu-AlgMS@MT.

[0099] Test Example 1

[0100] Test Example 1 177 Lu-SiMS and 177 Lu-SiMS@HT and Example 7 177 Lu-AlgMS and 177 Labeling stability of Lu-AlgMS@HT.

[0101] The test method is: 177 Lu-SiMS and 177Lu-SiMS@HT were placed in 1 mL saline and 10% fetal bovine serum (FBS) respectively, and the supernatant was centrifuged at 0, 2, 4, 6, 24, 48, 72 and 96 h to detect the radioactivity, and the radioactivity of the bottom microspheres was measured to calculate the labeling stability. Similarly, Lu-SiMS@HT was placed in 1 mL 10% fetal bovine serum (FBS), and the supernatant was centrifuged at 0, 2, 4, 6, 24 and 48 h to detect the radioactivity, and the radioactivity of the bottom microspheres was measured to calculate the labeling stability. 177 Lu-AlgMS and 177 Lu-AlgMS@HT was placed in 1 mL 10% fetal bovine serum (FBS), and the supernatant was centrifuged at 0, 2, 4, 6, 24 and 48 h to detect the radioactivity, and the radioactivity of the bottom microspheres was measured to calculate the labeling stability.

[0102] The test results are shown in Figure 10 and Figure 11 , Figure 10 The test results of the labeling stability of Lu-SiMS and 177 Lu-SiMS@HT in saline and 10% fetal bovine serum (FBS) environment are shown in 177 , Figure 11 The test results of the labeling stability of Lu-AlgMS and 177 Lu-AlgMS@HT in 10% fetal bovine serum (FBS) environment are shown in 177 , it can be seen that the labeling stability of 177 Lu can be greatly improved after HT modification.

[0103] Test Example 2

[0104] 1 mg of SiMS and 100 μL of AlgMS dispersion (10 mg / mL) were added to 10 μL of HfCl4 aqueous solution (10 mM) and 10 μL of TA (10 mM) aqueous solution respectively, and after mixing, the SiMS@HT and AlgMS@HT were obtained by washing with deionized water three times. The loading of ATRi by SiMS and SiMS@HT and AlgMS and AlgMS@HT was tested.

[0105] The test method was as follows: 300 μL of 200 μM ATRi aqueous solution was added to SiMS and SiMS@HT and AlgMS and AlgMS@HT respectively, and 100 μL of supernatant was taken at the corresponding time point for enzyme marker detection. According to the standard curve of ATRi concentration and absorbance, the ATRi loading of SiMS and SiMS@HT and AlgMS and AlgMS@HT was calculated.

[0106] The test results are shown in Figure 12 and Figure 13 , Figure 12 The comparison chart of the loading amount of ATRi by SiMS and SiMS@HT is shown inFigure 13 The loading capacity of ATRi of AlgMS and AlgMS@HT were compared, which proved that the metal polyphenol network modified microspheres could improve the loading of ATRi on the microspheres.

[0107] Test Example 3

[0108] SiMS, 177 Lu-SiMS, 177 Lu-SiMS@HT and 177 Lu-SiMS@HTi in Test Example 1,

[0109] The test method is as follows: (1) cell live and dead experiment: RH35 cells were inoculated in a 24-well plate, 6×10 4 cells per well, and cultured for 24 h, and then treated as follows, G1: DMEM, G2: SiMS (200 μg / mL), G3: 177 Lu-SiMS (25 μCi / mL), G4: 177 Lu-SiMS@HT (25 μCi / mL), G5: 177 Lu-SiMS@HTi (25 μCi / mL). After incubation for 24 h, the culture medium was removed, and 250 μL of Calcein AM / PI detection working solution was added. After incubation at 37°C for 30 min in the dark, the staining effect was observed under a fluorescence microscope (Calcein AM is green fluorescence, Ex / Em = 494 / 517 nm; PI is red fluorescence, Ex / Em = 535 / 617 nm).

[0110] (2) Cell apoptosis experiment: RH35 cells were inoculated in a 6-well plate, 20×10 4 cells per well, and cultured for 24 h, and then the cells were treated as follows, G1: DMEM, G2: SiMS (500 μg / mL), G3: 177 Lu-SiMS (80 μCi / mL), G4: 177 Lu-SiMS@HT (80 μCi / mL), G5: 177 Lu-SiMS@HTi (80 μCi / mL). After incubation for 48 h, the cells were collected into an EP tube using trypsin, 2000 r, 5 min, the supernatant was discarded, the cells were collected, and the apoptosis staining working solution was added, mixed, and incubated at room temperature (20-25°C) for 20 min in the dark, and then placed in an ice bath for flow cytometry detection. Annexin V-FITC is green fluorescence, and propidium iodide (PI) is red fluorescence.

[0111] The test results are as follows: Figure 14As shown, cell live / dead and apoptosis test results prove that 177 Lu-SiMS@HTi 177 Lu-SiMS has stronger tumor cell killing effect.

[0112] Test Example 4

[0113] Validation was carried out at the animal level, SiMS in Example 1, 177 Lu-SiMS, 177 Lu-SiMS@HT and 177 Lu-SiMS@HTi were used to treat SD rats with successfully induced orthotopic liver cancer models by the administration mode of transcatheter arterial embolization (TAE), and after treatment, the SD rats were humanely sacrificed, and the liver tumors were taken for hematoxylin-eosin (H&E), proliferating cell nuclear antigen (Ki67) and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining, which showed that tumor cell necrosis and apoptosis increased and proliferation decreased, further verifying at the animal level that 177 Lu-SiMS@HTi has a treatment effect on liver cancer.

[0114] The present application uses a metal polyphenol network to modify the surface of organic and inorganic radioactive microspheres, which can improve the labeling stability of 177 Lu on microspheres, improve the tumor retention effect in the animal body, and the drug-loaded radioactive microspheres modified by the metal polyphenol network can achieve inhibition of DNA damage repair of tumor cells due to the loading of ATRi, enhance the tumor killing effect of radioactive microspheres, and avoid secondary administration through SIRT, realizing one-step radio-chemotherapy combined application for tumors.

[0115] Obviously, the above embodiments of the present application are only examples for clearly illustrating the present application, and are not intended to limit the embodiments of the present application. Those skilled in the art should understand that on the basis of the above description, other different forms of changes or variations can also be made. Here, it is not necessary and impossible to exhaust all the embodiments. Any modification, equivalent replacement and improvement made within the spirit and principles of the present application should be included in the protection scope of the claims of the present application.

Claims

1. A method for preparing drug-loaded radioactive microspheres modified with a metal polyphenol network, characterized in that: The following steps are involved: (1) adding an inorganic salt containing a radioactive nuclide and a precipitant to an inorganic microsphere dispersion to obtain radioactive microspheres after labeling; the inorganic microspheres are inorganic silica microspheres; or Adding an inorganic salt containing a radioactive nuclide to an organic microsphere dispersion to obtain radioactive microspheres after labeling; the organic microspheres are organic sodium alginate microspheres; (2) mixing the radioactive microspheres obtained in step (1) with a metal salt solution and a polyphenol solution to obtain radioactive microspheres modified with a metal polyphenol network; the amount ratio of the radioactive microspheres to the metal salt solution and the polyphenol solution is 1 mg: (5-40) μL: (5-40) μL; (3) The metal polyphenol network-modified radioactive microspheres obtained in step (2) are mixed with a drug solution to obtain the metal polyphenol network-modified drug-loaded radioactive microspheres.

2. The preparation method according to claim 1, characterized in that In step (1), the radionuclide is selected from 177 Lu, 99 mTc, 32 P. 125 I. 131 I and 255 One or more of Ac.

3. The preparation method according to claim 1, characterized in that In step (2), the metal salt of the metal salt solution is selected from one or more of hafnium salts, iron salts, manganese salts, copper salts, cobalt salts and vanadium salts.

4. The preparation method according to claim 1, characterized in that In step (2), the polyphenols in the polyphenol solution are selected from one or more of tannic acid, gallic acid, protocatechuic aldehyde, pyrogallol and catechol.

5. The preparation method according to claim 1, wherein In step (2), the concentration of the metal salt in the metal salt solution is 5-20 mM.

6. The preparation method according to claim 1, wherein In step (3), the dosage ratio of the metal polyphenol network-modified radioactive microspheres to the drug solution is 1 mg: (200-600) μL.

7. The preparation method according to claim 1, characterized in that In step (3), the drug in the drug solution is an ATR inhibitor, doxorubicin hydrochloride, Evofosfamide, toyocamycin, atovaquone or a pan-Bcl-2 inhibitor.

8. A metal polyphenol network-modified drug-loaded radioactive microsphere prepared by the method according to any one of claims 1 to 7.

9. Use of the metal polyphenol network-modified drug-loaded radioactive microspheres according to claim 8 in the preparation of drugs for tumor diagnosis and treatment.

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