Light-responsive controllable enhanced metal-dependent cell death synergistic radionuclide therapy nanoparticle, and preparation method and application thereof

Abstract

The present application relates to the technical field of biological medicine, and specifically discloses a kind of light response controllable enhancement metal-dependent cell death synergistic nuclide treatment nanoparticle and its preparation method and application, the nanoparticle provided by the present application has the following characteristics: light response precision release mechanism, radioactivity amplification mechanism, diagnosis and treatment integrated mechanism and synergistic amplification mechanism, through metal ion release and nuclide internal irradiation synergy, form multilevel mitochondrial protein steady-state destruction and oxidative stress amplification, thereby efficiently inducing metal-dependent cell death, and activating antitumor immune response, solve the problems such as uncontrollable metal ion release, high risk of systemic toxicity, low efficiency of metal death induction, limited efficacy of radionuclide therapy and lack of diagnosis and treatment integrated regulation mechanism in the prior art.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to a photoresponsive nanoparticle that controls the enhancement of metal-dependent cell death in conjunction with radionuclide therapy, its preparation method, and its application. Background Technology

[0002] In recent years, radionuclide therapy has attracted widespread attention due to its ability to precisely irradiate tumor tissues. Compared with traditional radiotherapy, radionuclides can achieve sustained radiation damage by targeting and accumulating at the lesion site, thereby inducing tumor cell death. However, current radionuclide therapies still face limitations in efficacy. On the one hand, radionuclides mainly exert their effects by inducing DNA damage, oxidative stress, and cell cycle arrest, while some tumor cells possess strong DNA damage repair and metabolic adaptation capabilities, easily developing treatment tolerance. On the other hand, while increasing the radionuclide dose can enhance efficacy, it may also increase the risk of radiation damage to normal tissues. Therefore, developing new strategies that can improve the sensitivity of radionuclide therapy and achieve synergistic effects is of great significance.

[0003] It is noteworthy that radioactive metal nuclides can not only induce DNA damage using radiation, but their metal ions can also participate in the cell death process, and the related mechanisms have received increasing attention in recent years. Unlike traditional programmed cell death forms such as apoptosis, necroptosis, pyroptosis, and ferroptosis, metal-dependent cell death (hereinafter referred to as "metal death") exhibits unique molecular mechanisms and metabolic dependence characteristics. For example, copper death (cuproptosis) has been shown to be closely related to the abnormal binding of liponylated proteins in the mitochondrial tricarboxylic acid cycle (TCA). Excess copper ions can interact abnormally with liponylated enzymes, inducing protein aggregation and disrupting mitochondrial protein homeostasis, thereby leading to cell death. Furthermore, iron-dependent lipid peroxidation-induced ferroptosis and other transition metal-mediated oxidative stress imbalances further reveal the crucial role of metal homeostasis regulation in cell fate determination. Simultaneously, ionizing radiation from radionuclides can induce DNA damage and oxidative stress responses, which, combined with metal homeostasis imbalances affecting mitochondrial homeostasis, are expected to jointly promote cell damage. However, in existing technologies, radioactive metals are mostly used as tracers or radiotherapy carriers, and their synergistic regulation with metal-dependent cell death mechanisms has not yet been systematically integrated.

[0004] However, the rational integration of radionuclide therapy and metal-dependent death using existing technologies still faces many limitations. First, effective induction of metal death typically relies on a significant increase in intracellular metal ions. However, metal ions have a wide range of physiological functions in vivo, and non-selective increases can lead to severe systemic metal toxicity and damage to normal tissues, accompanied by the irradiation effects of metal nuclides. Current technologies typically employ metal ion carriers or ion transporters to promote metal entry into cells, or deliver metal salts via nanomaterials to enhance tumor accumulation. However, these methods mostly rely on passive targeting or release triggered by acidic conditions in the tumor microenvironment, lacking active regulatory mechanisms mediated by exogenous stimuli. This makes precise spatiotemporal regulation difficult to achieve, and carries the risk of premature leakage of metal and nuclide metal ions and off-target effects.

[0005] Metal-organic frameworks (MOFs) are widely used in drug delivery and imaging due to their high specific surface area, tunable pore structure, and excellent drug loading capacity. Copper-based MOFs can simultaneously serve as reservoirs and carriers for both radioactive metal isotopes and metals, showing potential advantages in radionuclide therapy and metal death induction. However, existing MOF systems mostly rely on pH or reducing environment responses for structural degradation, and their release process is mainly influenced by the tumor microenvironment, lacking precise external control capabilities and making it difficult to achieve precise metal release within specific regions. Radioactive metal isotopes exhibit unique advantages in the field of integrated tumor diagnosis and treatment. In particular, some radioactive isotopes possess both therapeutic and molecular imaging functions, showing significant application prospects in this field. For example, copper-64 (… 64 Cu can exert therapeutic effects by producing ionizing radiation through decay, and it can also be used in positron emission tomography (PET). Lutetium-177 ( 177 Lu combines the functions of beta particle therapy and single-photon emission computed tomography (SPECT), thus it is expected to non-invasively trace the dynamic distribution of metal elements in vivo, so as to further guide the precise controlled release of metals.

[0006] Furthermore, metal death is significantly dependent on cellular metabolic state. Cells with active mitochondrial respiration and high levels of lipoylated protein expression are more prone to metal-dependent protein homeostasis imbalance. However, existing metal delivery systems typically only focus on increasing metal concentration, lacking active regulation of tumor metabolic state, resulting in some metabolically plastic tumors being insensitive to metal death induction.

[0007] It is evident that the existing technology still has shortcomings in the following aspects: (1) lacks a metal release system that can be spatially precisely controlled by external stimuli; (2) it is difficult to achieve local high-concentration metal accumulation while ensuring safety; (3) it fails to effectively combine metabolic reprogramming strategies to enhance metal death sensitivity; (4) lacks a system scheme that integrates internal irradiation of radioactive metals, metal death induction and imaging monitoring on the same platform.

[0008] Therefore, developing a metal death induction platform that can achieve synergistic enhancement of exogenous controllable metal release, metabolic state regulation, and internal radiation irradiation is of great significance for improving the selectivity and efficacy of tumor treatment. Summary of the Invention

[0009] To address the shortcomings of existing technologies, the present invention aims to provide a photoresponsive nanoparticle that enhances metal-dependent cell death in conjunction with radionuclide therapy, along with its preparation method and application. This addresses issues in existing technologies such as uncontrollable metal ion release, high systemic toxicity risk, low metal death induction efficiency, limited therapeutic efficacy of radionuclide therapy, and lack of integrated diagnostic and therapeutic control mechanisms.

[0010] To achieve the above objectives, the present invention adopts the following technical solution: In a first aspect, the present invention provides a photoresponsive nanoparticle that enhances metal-dependent cell death and synergistically induces radionuclide therapy, the nanoparticle comprising a metal-organic framework material and a photoacid generator loaded on the metal-organic framework material.

[0011] In the technical solution disclosed in this invention, the metal-organic framework material is selected from copper-based metal-organic framework materials or iron-based metal-organic framework materials.

[0012] In the technical solution disclosed in this invention, the metal-organic framework material is doped with radioactive metal isotopes.

[0013] The radioactive metal isotopes are selected from 61 Cu、 62 Cu、 64 Cu、 67 Cu、 68 Ga、 89 Zr、 44 Sc、 47 Sc、 111 In、 177 Lu、 90 Y、 188 Re、 225 One or more of Ac.

[0014] in, 61 Cu、 62 Cu、 64 Cu、 68 Ga、 89 Zr and 44 Sc can be used for PET imaging; 111 In and 177 Lu can be used for SPECT imaging; 67 Cu、 177 Lu、 90 Y、188 Re and 225 Ac can be used in radiotherapy; among which 64 Cu、 67 Cu and 177 Lu has both molecular imaging and therapeutic functions.

[0015] In the technical solution disclosed in this invention, the photoacid generator is selected from cyanine photoacid generators, spiropyran photoacid generators, cyanocyanine photoacid generators, BODIPY photoacid generators and their derivatives.

[0016] In the technical solution disclosed in this invention, the metal-organic framework material is further loaded with metabolic regulatory molecules and / or targeting substances.

[0017] In the technical solution disclosed in this invention, the metabolic regulatory molecule is selected from MDM2 inhibitors, glycolysis inhibitors, mitochondrial function enhancers, chemotherapy drugs, targeted therapy drugs, immunomodulators, or nucleic acid drugs.

[0018] In the technical solution disclosed in this invention, the targeting substance is selected from polypeptides, small molecules, antibodies, aptamers or sugar recognition molecules that can recognize tumor-related receptors, cell surface proteins or microenvironment markers.

[0019] Secondly, the present invention provides a method for preparing the above-mentioned photoresponsively controllable enhanced metal-dependent cell death synergistic radionuclide therapy nanoparticles, comprising the following steps: S1. Preparation of metal-organic framework materials; S2. The photoacid generator is loaded onto the metal-organic framework material to obtain the above-mentioned nanoparticles.

[0020] In the technical solution disclosed in this invention, step S1 further includes: introducing a radioactive metal isotope into the metal-organic framework material by ion exchange or co-doping to obtain a doped metal-organic framework material.

[0021] The technical solution disclosed in this invention also includes: step S3, loading metabolic regulatory molecules and / or targeting substances onto the surface of a metal-organic framework material.

[0022] Thirdly, the present invention also provides the application of the above-mentioned photoresponsively controlled enhanced metal-dependent cell death synergistic radionuclide therapy nanoparticles in the preparation of tumor therapeutic drugs.

[0023] Compared with the prior art, the present invention has the following beneficial effects: The nanoparticles provided by this invention have the following characteristics: (1) Photo-responsive precise release mechanism: Under external light conditions, the photoacid generator releases protons, the local pH drops rapidly, and the MOF structure undergoes acid-sensitive degradation, thereby realizing the spatially confined release of metal ions in the light-illuminated area.

[0024] (2) Metabolic sensitization mechanism: The loaded metabolic regulatory molecules activate specific signaling pathways (such as the MDM2-p53 axis), inhibit glycolysis and enhance mitochondrial respiration, making the cells more sensitive to the imbalance of mitochondrial protein homeostasis induced by metal ions.

[0025] (3) Radioactive amplification mechanism: Doped radioactive metal isotopes release β rays or Auger electrons, which induce DNA damage and oxidative stress, further disrupting mitochondrial homeostasis and lowering the metal death threshold.

[0026] (4) Integrated diagnosis and treatment mechanism: Radioactive metal isotopes can be used for PET imaging to realize real-time distribution monitoring of nanoparticles in the body and assessment of treatment response.

[0027] (5) Synergistic amplification mechanism: The release of metal ions, metabolic reprogramming and internal irradiation of radionuclides work together to form multi-level disruption of mitochondrial protein homeostasis and amplification of oxidative stress, thereby efficiently inducing metal-dependent cell death and activating anti-tumor immune response. Attached Figure Description

[0028] Figure 1 This is a schematic diagram of the construction process of nanoparticles provided in Example 1 of the present invention.

[0029] Figure 2 This is a schematic diagram of the construction process of nanoparticles provided in Embodiment 2 of the present invention.

[0030] Figure 3 These are characterization images of CuM and fCuM@PI prepared in Example 1 of the present invention; where A is the TEM image of CuM, B is the TEM image of fCuM@PI, C is the Zeta potential image, and D is the polydispersity index image.

[0031] Figure 4 The present invention provides a synthetic route and structural characterization diagram of MEH, a photoacid generator of cyanine genus; wherein A is a schematic diagram of the synthetic route of MEH, B is a mass spectrometry characterization diagram of MEH, and C is a chromatographic analysis diagram of MEH.

[0032] Figure 5 MEH, a photoacid molecule provided by this invention 1 H NMR spectrum.

[0033] Figure 6 This is a photograph of a solution undergoing a color change under light-induced illumination.

[0034] Figure 7 The graph shows the test results of the photoinduced acidification process. A represents the pH changes of PAG and fCuM@PI aqueous solutions (1 mL) under light irradiation; B shows the pH change curves of fCuM@PI aqueous solutions with different initial volumes before and after light irradiation; and C represents the photoinduced acidification of Idasanutlin and Cu... 2+ A schematic diagram of the release, and the release of Idasanutlin and Cu by fCuM@PI in the culture medium. 2+ The quantitative detection results, where D represents Cu under different pH conditions and with or without light treatment. 2+ The release curve.

[0035] Figure 8 The graph shows the results of radioactivity stability testing of the nanoparticles prepared in Example 2 of this invention, where A represents the nanoparticles after purification and light treatment. 64 The Radio-iTLC analysis results of Cu-fM@PI are shown in Figure B. 64 Figure showing the in vitro stability evaluation results of Cu-fM@PI in PBS and FBS.

[0036] Figure 9 The figure shows the results of a systematic study at the molecular and cellular functional levels of the nanoparticles prepared in Example 2 of this invention. A shows the binding interaction between Idasanutlin and MDM2 as shown by molecular docking analysis, including two-dimensional and three-dimensional docking models. B shows the detection results of mitochondrial respiratory chain complex I activity after mitochondria were isolated and treated with different concentrations of Idasanutlin. C shows the Western blot analysis results of MDM2, p53, p21, PFKP, and PKM2 under different treatment conditions. D shows the lactate content, acetyl-CoA level, and NAD+ levels under different treatment conditions. + Quantitative analysis results of the / NADH ratio.

[0037] Figure 10 The figure shows the evaluation results of the synergistic effect of Cu-MOF and Idasanutlin in the combined treatment of 4T1 cells in Example 1 of the present invention. In this figure, A is the concentration-cell survival rate curve after Cu-MOF and Idasanutlin are treated alone in 4T1 cells, and B is the combination index (CI) and synergistic effect analysis results after Cu-MOF and Idasanutlin are administered in a fixed ratio.

[0038] Figure 11The figure shows the functional verification results of Idasanutlin-mediated cell metabolic reprogramming in Example 1 of this invention. A represents the changes in cellular oxygen consumption rate (OCR) under different treatment conditions analyzed by Seahorse XF, including basal respiration, maximum respiratory capacity, and ATP production level. B represents the changes in extracellular acidification rate (ECAR) under different treatment conditions analyzed by Seahorse XF, including glycolysis level, glycolysis capacity, and glycolysis reserve.

[0039] Figure 12 This is a diagram illustrating the effect of the nanoparticles provided by this invention at the cellular level, where A represents the effect of 4T1 cells on... 64 CuCl2, 64 Cu-M@PI and 64 Cellular uptake of Cu-fM@PI, including the proportion of cell membrane-bound portions and the proportion endocytosed into the cell, where B represents different activities. 64 CuCl2, 64 Cu-M@PI and 64 The relative survival rate of 4T1 cells after Cu-fM@PI treatment, C represents Bio-TEM images of 4T1 tumor cells after different treatments (green arrows indicate structurally intact mitochondria, red arrows indicate damaged mitochondria; scale bar is 2 μm), and the corresponding quantitative analysis results of mitochondrial length.

[0040] Figure 13 This is an imaging performance and biodistribution characteristic map of nanoparticles provided by the present invention, where A represents intravenous injection. 64 CuCl2 or 64 A schematic diagram of the process for PET imaging, biodistribution, and autoradiography experiments after Cu-fM@PI administration. B shows representative PET images of tumor-bearing mice at different time points after drug administration. C shows the semi-quantitative analysis results of tumor uptake at different time points. D shows the analysis of the uptake ratio of tumor to muscle tissue. E shows the analysis results of the area under the curve (AUC) of tumor accumulation over time. F shows the in vitro biodistribution analysis results of major organs and tumor tissue.

[0041] Figure 14 The following diagram illustrates the therapeutic effects of the nanoparticles provided by this invention: A is a schematic diagram of the experimental process; B shows representative photographs of tumors removed from each group on day 16 and the corresponding tumor weights; C shows the dynamic monitoring results of tumor volume changes over time in different treatment groups; D shows the tumor growth inhibition rate calculated based on tumor volume; E shows the survival analysis results of each treatment group within 80 days; F shows the immunohistochemical staining results of Ki-67 and the TUNEL (green) immunofluorescence staining results in tumor tissue (scale bar is 100 μm); and G shows the quantitative analysis results of proliferation index and apoptosis index in tumor tissue.

[0042] Figure 15 The diagram shows the results of molecular mechanism analysis of the nanoparticles provided by this invention. In the diagram, A is a heatmap showing the expression levels of representative genes related to copper death, copper homeostasis, glycolysis, mitochondrial respiration, DNA damage response, and immune regulation after standardization and normalization. B is the gene set enrichment analysis results, showing that immune-related signaling pathways were significantly enriched in the tumor of the combined treatment group compared with the control group.

[0043] Figure 16 This is a graph showing the results of flow cytometry analysis of immune cell composition, where A represents the maturation status of dendritic cells (CD11c) in tumor tissue. + CD80 + CD86 + ) and tumor-infiltrating CD8 + and CD4 + T cells (in CD45) + CD3 + Representative flow cytometry plots (gated analysis in cell populations), B represents CD8+ in tumor tissue after different treatments. + and CD4 + T cells in total live cells and CD45 + CD3 + The proportion of cells, CD8 + / CD4 + Ratio, dendritic cell maturation level, CD4 + Regulatory T cells (CD4) in T cells + CD25 + FoxP3 + The quantitative analysis results of the M1 / M2 macrophage ratio and the M1 / M2 macrophage ratio were obtained.

[0044] Figure 17 The images show the results of immunohistochemistry and immunofluorescence.

[0045] Figure 18 The results of the safety evaluation are shown in the figure below. A is a graph showing the change in tumor volume of individual mice in different treatment groups over time. B is a graph showing the change in body weight of mice during treatment. C is a histological evaluation of the major organs of 4T1 tumor-bearing mice, including the H&E staining results of the heart, liver, lungs and kidneys. D is the results of hematological and serum biochemical tests, including white blood cells (WBC), red blood cells (RBC), platelets (PLT), lymphocyte percentage, hemoglobin (HGB), alanine aminotransferase (ALT), aspartate aminotransferase (AST), serum creatinine (CREA), and blood urea nitrogen (BUN).

[0046] Figure 19 A schematic diagram illustrating the overall mechanism of action of the nanoparticles provided by this invention. Detailed Implementation

[0047] The present invention will be further described in detail below through specific preferred embodiments, but the present invention is not limited to the following embodiments.

[0048] It should be noted that, unless otherwise specified, all chemical reagents involved in this invention were purchased through commercial channels.

[0049] This invention provides a method for preparing photoresponsive nanoparticles that controllably enhance metal-dependent cell death and synergistically induce radionuclide therapy, comprising the following steps: S1. Synthesis of Metal-Organic Framework Materials Taking copper-based metal-organic frameworks as an example, copper salts, such as copper nitrate or copper chloride, are selected as metal precursors, and polycarboxylated organic ligands are selected as ligands. The reaction is carried out under solvothermal conditions to form CuM (MOF-199) nanoparticles with stable crystal structures. The solvent is preferably ethanol or dimethylformamide, the reaction temperature is preferably 80-120℃, and the reaction time is preferably 6-24 h. After the reaction, the product is separated by centrifugation, washed with the appropriate solvent, and then dried for later use.

[0050] Meanwhile, in the copper-based metal-organic framework system, this invention, in addition to 64 In addition to Cu, other materials can also be used. 61 Cu、 62 Cu、 67 Isotope doping with copper isotopes such as Cu can impart integrated functionality for PET imaging, radiotherapy, or diagnostics to materials. Furthermore, other methods can be employed as needed. 177 Lu、 90 Y、 89 Zr、 68 Ga、 44 Sc、 47 Sc、 111 In、 188 Re、 225 Radioactive metal nuclides such as Ac can be introduced into metal-organic framework systems through co-doping, ion exchange, or coordination assembly, thereby enabling molecular imaging, radionuclide therapy, or synergistic functional expansion of both.

[0051] Furthermore, this invention is not limited to copper-based metal-organic framework materials. Iron salts (such as ferric chloride, ferric nitrate, ferrous chloride, or ferrous sulfate) can be used instead of copper salts as the metal source to prepare iron-based metal-organic framework nanoparticles (FeM), such as MIL-101(Fe), MIL-88(Fe), MIL-53(Fe), or PBAs, using a similar solvothermal method. The reaction temperature, reaction time, ligand type, and solvent system can be appropriately adjusted according to the coordination characteristics of different metal ions. In the iron-based system, further... 89Zr、 68 Ga、 44 Sc、 111 In、 177 Lu、 225 Radioactive metal nuclides such as Ac are used to construct a radioactive Fe-MOF system that combines molecular imaging and radiotherapy functions. Iron ions participate in ferroptosis-related processes, while radioactive nuclides provide continuous internal irradiation; both synergistically enhance tumor treatment efficacy.

[0052] Through the above methods, the present invention is not limited to copper systems, but can also be extended to iron and other transition metal systems to construct a generalized metal death-inducing platform.

[0053] In the technical solutions disclosed in this invention, radioactive metal isotopes can also be introduced into the metal-organic framework through ion exchange or co-doping.

[0054] by 64 Taking Cu as an example, when preparing the metal precursor solution, a predetermined activity of [a specific ingredient] is added. 64 CuCl2 solution, radioactive copper isotopes 64 Cu, along with non-radioactive copper ions, participates in the subsequent construction of the metal-organic framework, thereby obtaining... 64 Cu-doped MOF nanoparticles ( 64 The obtained product was centrifuged and washed to remove unreacted raw materials and free radionuclides before being put into use.

[0055] In a specific embodiment of the present invention, the specific experimental steps are as follows: Trimethylbenzene acid and polyvinylpyrrolidone are dissolved in N,N-dimethylformamide to obtain a trimethylbenzene acid mixture; 64 CuCl2 solution and Cu(NO3)2·3H2O were added to N,N-dimethylformamide and mixed thoroughly to obtain a solution containing... 64 Cu solution, containing 64 The Cu solution was added to the pyromellitic acid mixture and mixed thoroughly. Then, it was transferred to a high-pressure reactor for reaction. After the reaction was completed, the material was centrifuged, washed, and dried to obtain the radioactive copper isotope-doped metal-organic framework material.

[0056] The molar ratio of pyromellitic acid, polyvinylpyrrolidone and Cu(NO3)2·3H2O is 1:1-5:1.8-2.5.

[0057] 64 CuCl2 accounts for 0.001%-1% of the total molar amount of copper source.

[0058] The reaction temperature is 80-120℃, and the reaction time is 6-24h.

[0059] Doping efficiency can be detected by radio-iTLC.

[0060] Under optimal conditions, the doping efficiency is not less than 70%.

[0061] In other embodiments, it may also be doped with 177 Lu、 90 Y、 125 I, 131 I, 188 Re、 225 Radioactive metal isotopes such as Ac are used to meet different imaging or treatment needs.

[0062] S2, Loading of photoacid generator Photoacid generators (PAGs) were dissolved in a suitable solvent, mixed with a MOF dispersion, and incubated at room temperature to allow PAGs to enter the MOF structure via physical adsorption or intercalation. After loading, unbound PAGs were removed by centrifugation and washing. The loading efficiency of the photoacid generators could be quantitatively determined by UV-Vis absorption spectroscopy or high-performance liquid chromatography.

[0063] This step enables the material to release protons and trigger framework degradation under light conditions.

[0064] S3, Loading of metabolic regulatory molecules During the simultaneous loading of photoacid generators, or after the loading of photoacid generators is completed, metabolic regulatory molecules can be introduced into the MOF material structure.

[0065] In this step, the mass ratio of MOF material, acid-producing agent, and metabolic regulatory molecule is 1:0.02-0.1:0.05-0.2.

[0066] In a preferred embodiment, the photoacid generator and the metabolic regulatory molecule can be co-loaded within the metal-organic framework to simplify the process and improve the synergistic release efficiency.

[0067] Specifically, metabolic regulatory molecules (such as the MDM2 inhibitor Idasanutlin) are dissolved in a suitable solvent, mixed with a photoacid generator and a MOF dispersion system, and incubated at room temperature with stirring. This allows the two functional molecules to embed into the MOF structure through pore adsorption or intermolecular interactions. After incubation, unbound components are removed by centrifugation and washing.

[0068] The nanoparticles constructed in the above manner can synergistically trigger the release of metal ions and metabolic regulatory molecules under light stimulation, thus possessing both photoresponsive metal release and metabolic reprogramming regulation functions, providing a basis for the subsequent synergistic induction of metal-dependent cell death.

[0069] S4, Targeted material surface modification To improve the stability of nanoparticles in vivo, prolong their circulation time, and enhance their accumulation capacity at tumor sites, further surface modification with targeted substances can be employed.

[0070] Specifically, the mass ratio of nanoparticles to target material is 1:0.2-1.0.

[0071] The target substances can be selected from, but are not limited to: folic acid (targeting folic acid receptor), transferrin (targeting transferrin receptor), EGFR ligand or antibody fragments, HER2 antibody fragments, and TROP2 antibody fragments targeting tumor cells; and RGD peptides or their derivatives targeting tumor blood vessels (targeting α-cell receptors). v β3 integrin, VEGFR ligand; FAP targeting molecules for tumor stroma or cancer-associated fibroblasts; CD44 ligand for the tumor immune microenvironment; and other peptides, small molecules, antibodies, aptamers or carbohydrate recognition molecules that can recognize tumor-associated receptors, cell surface proteins or microenvironmental markers.

[0072] The target material can be modified onto the MOF surface through covalent coupling, electrostatic adsorption, or other chemical linkages.

[0073] Furthermore, surface coating with polyethylene glycol (PEG) can improve its dispersibility and biocompatibility in body fluid environments, thereby reducing reticuloendothelial system clearance.

[0074] After surface modification, the particle size of the nanoparticles is preferably controlled in the range of 80-250 nm to balance the stability of in vivo circulation and the enhanced penetration and retention effect (EPR effect) of tumor tissue. The modified nanoparticles exhibit good colloidal stability and dispersibility in the serum environment.

[0075] Example 1 A method for constructing photoresponsive nanoparticles that controllably enhance metal-dependent cell death includes the following steps: S1. Preparation of metal-organic framework material CuM: Weigh out 0.31 g of copper nitrate trihydrate (Cu(NO3)2·3H2O), 0.13 g of trimesic acid (BTC), and 0.10 g of polyvinylpyrrolidone (PVP) for later use. Dissolve BTC and PVP completely in 25 mL of N,N-dimethylformamide (DMF), and dissolve copper nitrate trihydrate completely in 30 mL of DMF. Sonicate each solution for 15 min, then mix and magnetically stir at room temperature for 30 min to obtain a homogeneous blue reaction solution. Transfer the resulting reaction solution to a polytetrafluoroethylene-lined high-pressure reactor and react at 100 °C for 2 h. After the reaction is complete and the mixture cools naturally to room temperature, collect the precipitate by centrifugation (12000 rpm, 10 min), and wash three times sequentially with DMF and anhydrous ethanol to remove unreacted raw materials and impurities. Dry the resulting product at 60 °C to obtain copper-based metal-organic framework nanoparticles CuM (MOF-199).

[0076] Preparation of composite nanomaterials containing S2, Idasanutlin, and photoacid MEH 20 mg of the CuM nanoparticles prepared above were weighed and dispersed in ethanol. Then, 2 mg of the MDM2 inhibitor Idasanutlin and 1 mg of the cyanide-based photoacid generator MEH were added. The mixture was stirred in the dark at room temperature to allow the drug molecules and photoacid molecules to enter the pores of the metal-organic framework and be loaded into the framework structure. After the reaction was complete, the product was collected by centrifugation and washed with ethanol to remove unloaded free drug and photoacid molecules, yielding a composite nanomaterial simultaneously loaded with Idasanutlin and the photoacid generator MEH, namely CuM@PI.

[0077] Preparation of S3, photoacid-responsive copper-based metal-organic framework nanoparticles The obtained CuM@PI (20 mg) was dispersed in 5 mL of PBS buffer, and 10 mg of DSPE-PEG2000-FA was added. The mixture was slowly stirred at room temperature, allowing DSPE-PEG2000-FA to coat the surface of CuM@PI through hydrophobic interactions and surface adsorption, forming a lipid layer with folic acid targeting function. After the reaction was completed, unbound DSPE-PEG2000-FA was removed by centrifugation, and the mixture was redispersed in PBS buffer to obtain folic acid-modified photoacid-responsive copper-based metal-organic framework nanoparticles fCuM@PI.

[0078] Example 2 This embodiment introduces a radioactive copper isotope based on the preparation of the above-mentioned fCuM@PI nanoparticles. 64 Cu is doped.

[0079] A photoresponsive nanoparticle that enhances metal-dependent cell death and synergizes with radionuclide therapy 64 The construction method of Cu-fM@PI includes the following steps: S1. Preparation of radioactive metal isotope-doped metal-organic framework materials: 2 mCi 64 Add approximately 50 μL of CuCl2 solution to 30 mL of DMF solution containing 0.31 g of copper nitrate trihydrate (Cu(NO3)2·3H2O), mix thoroughly, and allow the radioactive copper isotope to settle. 64 Cu and non-radioactive copper ions jointly participate in the subsequent construction process of the metal-organic framework. Then, following the CuM construction method in Example 1, the Cu-containing... 64 A radioactive copper isotope-doped metal-organic framework material was prepared by mixing a copper source solution with a BTC solution and then reacting the mixture with a solvothermal agent. 64 Cu-M ( 64 Cu / Cu MOF-199).

[0080] The obtained radioactive metal-organic framework material was further loaded with Idasanutlin and MEH according to the steps in Example 1, and then modified with DSPE-PEG2000-FA before centrifugation to remove impurities, thus obtaining... 64 Cu-fM@PI.

[0081] A schematic diagram of the construction process of the nanoparticles provided in Embodiment 1 of the present invention is shown below. Figure 1 As shown.

[0082] A schematic diagram of the nanoparticle construction process provided in Embodiment 2 of the present invention is shown below. Figure 2 As shown.

[0083] The characterization diagrams of CuM and fCuM@PI prepared in Example 1 of this invention are as follows: Figure 3 As shown, A is the TEM image of CuM, B is the TEM image of fCuM@PI, C is the Zeta potential plot, and D is the polydispersity index plot. Figure 3As can be seen, both the prepared CuM and fCuM@PI nanoparticles possess regular and uniform nanostructures and good dispersibility. The average particle size of CuM is approximately 86.70 ± 4.52 nm; after drug loading and surface modification, the average particle size of fCuM@PI increases to 99.16 ± 3.22 nm. Simultaneously, the Zeta potential changes from -11.59 ± 3.46 mV to -5.02 ± 1.03 mV, further demonstrating the successful coating of DSPE-PEG2000-FA onto the nanoparticle surface. The PDI values ​​of CuM and fCuM@PI are 0.09 ± 0.03 and 0.11 ± 0.05, respectively, indicating that the obtained nanoparticles possess good uniformity and colloidal stability.

[0084] Example 3 Verification of photoresponsive release performance and controllable release of metal ions To systematically evaluate the photoresponsive release performance and controllable release behavior of the constructed nanoparticles, the structure and photoresponsive characteristics of the photoacid molecule MEH were first characterized.

[0085] The photoacid molecule MEH used in the embodiments of this invention was purchased from Xi'an Ruixi Biotechnology Co., Ltd.

[0086] like Figure 4 As shown, the existing synthetic route for the photoacid molecule MEH is as follows: Figure 4 As shown in Figure A, this invention utilizes mass spectrometry analysis (… Figure 4 (B) and chromatographic analysis ( Figure 4 (C) Confirm the molecular weight and purity of the purchased photosensitive acid molecule MEH; and simultaneously, consider its properties in DMSO-d6. 1 H NMR spectrum ( Figure 5 This further verified the chemical structure and high purity of the target product. The above results indicate that the purchased MEH molecules have a well-defined structure and good purity, and can be used for subsequent experimental research.

[0087] The photoresponse characteristics of MEH were further evaluated. An appropriate amount of MEH was weighed and dissolved in dimethyl sulfoxide (DMSO) to prepare a 10 mM stock solution. Then, an appropriate amount of the stock solution was diluted in ultrapure water to prepare a 100 μM working solution. One mL of the working solution was placed in a transparent quartz cuvette and irradiated at room temperature using a 420 nm LED light source with a light power density of 50 mW / cm². 2 The irradiation time was 5 minutes, and the pH value of the system was monitored in real time using a pH meter. Figure 6As shown, with prolonged illumination, MEH undergoes structural transformation, and the solution color gradually changes from yellow to colorless. Simultaneously, the pH value of the system decreases significantly, indicating that MEH can release protons and form an acidic environment under light stimulation. These results demonstrate that MEH possesses excellent photoresponsive acid-producing capabilities, and can rapidly regulate the local acidic microenvironment under light conditions, providing conditions for subsequent metal-organic framework degradation and the controlled release of copper ions.

[0088] Based on this, the fCuM@PI nanoparticles prepared in Example 1 of this invention were dispersed in an aqueous solution and locally irradiated using a 420 nm LED light source (50 mW / cm²). 2 (5 min), and the pH changes of the system were monitored in real time using a pH meter. Figure 7 As shown in Figure A, under illumination, the pH values ​​of both the MEH solution (100 μM) and the fCuM@PI aqueous solution (1 mg / mL) decreased significantly, indicating that illumination can effectively trigger the release of protons from photoacid molecules and induce acidification of the system. Furthermore, as... Figure 7 As shown in Figure B, under different initial volume conditions, the fCuM@PI aqueous solution showed a consistent pH decreasing trend after light irradiation, indicating that the photoinduced acidification process has good repeatability and stability.

[0089] Subsequently, a systematic analysis was conducted on the release behavior of drugs and metal ions under light conditions. For example... Figure 7 As shown in Figure C, illumination can simultaneously trigger Idasanutlin and Cu. 2+ Released from fCuM@PI, its concentration in the culture medium gradually increased over time, indicating that the system possesses effective photocontrolled release capability. Furthermore, as... Figure 7 As shown in Figure D, under different pH environments and light treatment conditions, Cu 2+ The release behavior exhibited significant differences: under illumination, the release rate was significantly higher than that of the unilluminated group; furthermore, release was further accelerated under acidic conditions, suggesting that this nanosystem possesses dual regulatory characteristics of both photoresponsiveness and acid-sensitive response. In contrast, under no-light conditions, the pH change of the system was smaller, and Cu... 2+ The significantly reduced release rate of Idasanutlin indicates that the material's structure remains relatively stable in the absence of external stimuli.

[0090] Further 64 Cu-doped nanoparticles ( 64 The radioactivity stability of Cu-fM@PI was evaluated. Figure 8 As shown in Figure A, after purification 64 Cu-fM@PI exhibits good radiolabeling stability; however, after light treatment, Radio-iTLC analysis results show that free... 64The significantly increased Cu ratio indicates that photoacid-induced local acidification can disrupt the MOF framework structure, thereby accelerating the dissociation and release of radioactive metals. Figure 8 As shown in B, 64 Cu-fM@PI maintained a high radioactivity retention rate after being incubated in PBS and FBS for a certain period of time, further demonstrating that the nanoparticles have good in vitro stability in physiological environments.

[0091] Example 4 In vitro metal death induction and metabolic regulation verification To further elucidate the mechanism of action of the functional molecule Idasanutlin in the nanoparticles of this invention and its regulatory effect on tumor cell metabolism and mitochondrial function, a systematic study at the molecular and cellular functional levels was first conducted.

[0092] like Figure 9 As shown in Figure A, molecular docking analysis revealed that Idasanutlin can form a stable binding with MDM2, and its binding mode exhibits good spatial matching in both two-dimensional and three-dimensional structures, suggesting its potential to block MDM2-p53 interactions. Further verification of its function was conducted at the mitochondrial isolation level. Figure 9 As shown in Figure B, the activity of mitochondrial respiratory chain complex I gradually decreased with increasing Idasanutlin concentration, indicating that it can directly or indirectly affect mitochondrial electron transport function.

[0093] At the protein expression level, 4T1 cells were seeded in 6-well plates, with approximately 2 × 10⁶ cells per well. 5 Cells were cultured overnight at 37°C and 5% CO2 to ensure full cell adhesion. Four treatment groups were then established: Group 1 was the blank control group; Group 2 was treated with Idasanutlin (10 μM); Group 3 was treated with fCuM@PI; and Group 4 was treated with fCuM@PI combined with light irradiation (420 nm light source, 50 mW / cm²). 2 The irradiation time was 5 min, with the final concentration of Idasanutlin in groups 3 and 4 being 10 μM. Cells were collected 6 h after treatment in each group, and the expression levels of relevant proteins were detected by Western blot. Figure 9As shown in Figure C, compared with the blank control group, both the Idasanutlin treatment group and the fCuM@PI-related treatment group showed downregulated MDM2 protein expression, while the expression of p53 and its downstream effector p21 was significantly increased. Among them, the fCuM@PI combined with light treatment group showed the most significant changes. At the same time, the expression levels of the key rate-limiting enzymes of glycolysis, PFKP and PKM2, were significantly reduced, indicating that the nanoparticles constructed in this invention can effectively inhibit MDM2 activity, activate the p53 signaling pathway, and further regulate the glucose metabolism process of tumor cells.

[0094] To further evaluate cellular energy metabolism reprogramming, the Seahorse XF analysis system was used to detect cellular respiration and glycolysis. Specifically, 4T1 cells were seeded in Seahorse XF culture plates, with approximately 1 × 10⁶ cells per well. 4 Cells were cultured overnight at 37°C and 5% CO2. Four treatment groups were then established: Group 1 was the blank control group; Group 2 was the Idasanutlin treatment group (10 μM); Group 3 was the CuM treatment group; and Group 4 was the fCuM@PI combined light treatment group (420 nm light source, light power density of 50 mW / cm²). 2 The irradiation time was 5 min, with the final concentration of Idasanutlin in groups 3 and 4 being 10 μM. Seahorse XF assays were performed 6 h after treatment in each group. In the mitochondrial stress experiment, oligomycin, FCCP, and rotenone / antimycin were added sequentially, and cellular oxygen consumption rate (OCR) was measured, and basal respiration, maximum respiratory capacity, and ATP production levels were calculated. In the glycolytic stress experiment, glucose, oligomycin, and 2-deoxyglucose were added sequentially, and extracellular acidification rate was measured, and glycolytic level, glycolytic capacity, and glycolytic reserve were calculated. Figure 10 As shown in Figure A, compared with the blank control group, the cellular oxygen consumption rate of both the Idasanutlin treatment group and the fCuM@PI combined with light treatment group was significantly increased. Basal respiration, maximum respiratory capacity, and ATP production levels were all significantly increased, with the most significant changes observed in the fCuM@PI combined with light treatment group. This indicates that Idasanutlin can promote mitochondrial oxidative phosphorylation, and the fCuM@PI combined with light treatment can further enhance this effect. Figure 10As shown in Figure B, the extracellular acidification rate detection results indicated that the glycolysis level, glycolytic capacity, and glycolytic reserve were significantly reduced in both the Idasanutlin-treated group and the fCuM@PI combined with light treatment group. The fCuM@PI combined with light treatment group showed the largest decrease, indicating that this system can effectively inhibit glycolytic metabolism in tumor cells and weaken their glycolytic reserve capacity. These results demonstrate that the nanoparticles constructed in this invention can further promote the shift of cellular energy metabolism towards oxidative phosphorylation based on Idasanutlin-mediated metabolic reprogramming, creating favorable metabolic conditions for the subsequent copper death effect.

[0095] To further evaluate whether there is a synergistic effect between CuM-mediated metal ion release and Idasanutlin-mediated metabolic regulation, 4T1 cells were used as an in vitro model, and cell viability under different treatment conditions was detected by the CCK-8 assay. Figure 11 As shown in Figure A, the concentration-cell viability curves of CuM and Idasanutlin after treatment of 4T1 cells alone were determined. The results showed that both CuM and Idasanutlin inhibited 4T1 cell viability in a concentration-dependent manner, with IC50 values ​​of [missing information]. 50 The values ​​were 90.86 μg / mL and 9.98 μg / mL, respectively. Based on these values, the IC50 values ​​of CuM and Idasanutlin were determined according to their respective values. 50 The drugs were administered in a fixed ratio, and the Combination Index (CI) was calculated using CompuSyn software based on the Chou-Talalay method. Figure 11 As shown in B, at IC values ​​of 0.125×, 0.25×, 0.5×, 1×, and 2× 50 Under fixed-ratio combined drug administration conditions, the cell inhibition rates of the combined treatment groups were 21.58%, 34.63%, 53.19%, 74.36%, and 87.62%, with effect fractions of 0.216, 0.346, 0.532, 0.744, and 0.876, respectively. The CI values ​​calculated using CompuSyn software were 0.86, 0.78, 0.69, 0.61, and 0.58, all less than 1. These results indicate that CuM and Idasanutlin exhibit synergistic effects at low, medium, and high effect levels. Furthermore, as the combined treatment effect intensifies, the CI value further decreases, suggesting that the synergistic effect is more pronounced at higher inhibition levels. These results demonstrate that CuM-mediated metal ion release and Idasanutlin-mediated metabolic regulation can synergistically enhance the killing effect of 4T1 cells and improve the induction efficiency of metal-dependent cell death.

[0096] To further validate the effects of the nanoparticles at the cellular level, 4T1 cells were seeded in 6-well plates, with approximately 2 × 10⁶ cells per well. 5 Cells were cultured overnight at 37 °C with 5% CO2 in complete culture medium to allow for full cell adhesion. After cell adhesion, different radioactive activities (0.0185 MBq, 0.037 MBq, and 0.074 MBq) were added. 64 CuCl2, materials not modified with folic acid 64 Cu-M@PI and folic acid modification 64 Cells were incubated with Cu-fM@PI. After incubation, the supernatant was discarded, the cells were washed twice with PBS, and the cells were collected. Intracellular radioactivity was detected using a gamma counter to evaluate the uptake capacity of different nanoparticles in 4T1 cells. Figure 12 As shown in Figure A, 4T1 cells against 64 CuCl2, 64 Cu-M@PI and 64 Both Cu-fM@PI have a certain uptake capacity, among which 64 The Cu-fM@PI group exhibited a higher rate of endocytosis, suggesting that it possesses good cellular uptake characteristics. Figure 12 As shown in Figure B, under different radioactivity levels (0.37 MBq, 0.74 MBq, and 1.11 MBq), 64 The Cu-fM@PI group showed the most significant inhibitory effect on cell viability, exhibiting a clear dose-dependent effect. Further observation of cell ultrastructural changes using Bio-TEM revealed further evidence. Figure 12 As shown in Figure C, the mitochondrial structure was significantly damaged after treatment, exhibiting characteristics such as shortened length and structural destruction (red arrow), while the mitochondrial structure of the control group remained intact (green arrow). The above results are consistent with the quantitative analysis, indicating that the nanoparticles can effectively induce mitochondrial damage and enhance the metal-dependent cell death effect.

[0097] Example 5 The imaging performance, biodistribution characteristics, and antitumor therapeutic effects of the nanoparticles were further evaluated at the in vivo level. Specifically, the relevant experiments were conducted using a 4T1 subcutaneous tumor-bearing mouse model. 4T1 cells in the logarithmic growth phase were digested, centrifuged, and resuspended in PBS to prepare a cell suspension. Approximately 1 × 10⁶ cells were used. 6 100 μL of 4T1 cells were subcutaneously injected into the right axilla of female BALB / c mice to establish a subcutaneous 4T1 tumor-bearing model. The tumor was allowed to grow to approximately 80–100 mm. 3 Mice were randomly grouped and subsequent experiments were conducted. For PET imaging and biodistribution experiments, mice were injected via tail vein. 64 CuCl2 or 64Cu-fM@PI was administered, and PET imaging was performed at different time points after drug administration. Mice were sacrificed after imaging, and tumor and major organ tissues were collected. Tissue radioactive uptake levels were measured using a gamma counter, and the tumor-targeting ability and in vivo distribution characteristics of the nanoparticles were further evaluated using in vitro autoradiography. For the anti-tumor therapy experiment, mice were randomly divided into PBS group, 64 CuCl2 group, fCuM@PI group, and fCuM@PI combined illumination group (fCuM@PI+L) 64 Cu-fM@PI group and 64 Cu-fM@PI combined illumination group ( 64 Cu-fM@PI+L). Mice in each group were treated with the corresponding formulation via tail vein injection. For the combined light irradiation group, the tumor area was locally irradiated with a light source of wavelength matching the MEH absorption peak after drug administration, at a light power density of 50 mW / cm². 2 The irradiation time was 5 minutes. Tumor volume and mouse weight changes were measured every two days during treatment, and mouse survival was recorded. After treatment, tumor tissue was removed and weighed to evaluate the anti-tumor effects of different treatment regimens. On day 16, tumor tissue and major organs (including heart, liver, spleen, lungs, and kidneys) were collected for further analysis. Histopathological changes were evaluated using H&E staining; tumor cell proliferation and death were evaluated using Ki-67 and TUNEL immunohistochemical staining; and HMGB1 release and CD8+ were detected using immunofluorescence staining. + T cell infiltration and PD-L1 expression; flow cytometry was used to analyze dendritic cells and CD4+ expression in the tumor microenvironment. + T cells, CD8 + The compositional changes of immune cell subsets such as T cells, regulatory T cells, and M1 / M2 macrophages were analyzed. Transcriptome sequencing and related bioinformatics analyses were used to evaluate changes in signaling pathways related to copper death, copper homeostasis regulation, glycolysis, mitochondrial respiration, DNA damage response, and tumor immune regulation, thereby systematically elucidating the anti-tumor mechanism and immunomodulatory effects of the nanoparticles.

[0098] like Figure 13 As shown in Figure A, establish an intravenous injection... 64 CuCl2 or 64 Experimental procedure for Cu-fM@PI followed by PET imaging, biodistribution, and autoradiography analysis. PET imaging results ( Figure 13 (B) shows that 64 Cu-fM@PI showed significant enrichment at the tumor site, which gradually increased over time. Semi-quantitative analysis ( Figure 13 (C) and tumor / muscle ratio ( Figure 13 (D) further confirmed its good tumor targeting and contrast. AUC analysis results ( Figure 13 The results of in vitro biodistribution (E) indicate that it has a high accumulation level in tumor tissues; Figure 13 The distribution characteristics of the nanoparticles in vivo were further verified by the F-test.

[0099] Regarding treatment efficacy, such as Figure 14 Figure A shows the overall experimental procedure. Tumor removal results for each group on day 16 ( Figure 14 (B) The tumor volume and weight were significantly reduced in the treatment group. Tumor growth curve ( Figure 14 (C) and tumor growth inhibition rate calculated based on volume ( Figure 14 (D) indicates that the combined treatment group had a better tumor-suppressing effect. Survival analysis ( Figure 14 (E) showed that this treatment strategy significantly prolonged the survival time of tumor-bearing mice. Histological and immunostaining results ( Figure 14 The results showed that Ki-67 expression was decreased while TUNEL positive signal was enhanced, and quantitative analysis (F) showed that Ki-67 expression was decreased while TUNEL positive signal was enhanced. Figure 14 The results from the study (G) further demonstrate that tumor cell proliferation was inhibited and cell death levels were significantly increased.

[0100] At the molecular mechanism level, such as Figure 15 As shown in Figure A, heatmap analysis reveals systemic changes in the expression of genes related to copper death, copper homeostasis, glycolysis, mitochondrial respiration, DNA damage response, and immune regulation. Gene set enrichment analysis results ( Figure 15 (B) This indicates that the combination therapy can significantly enrich multiple immune-related signaling pathways, suggesting its potential to regulate the tumor immune microenvironment.

[0101] Further analysis of immune cell composition using flow cytometry ( Figure 16 (A) Results showed elevated dendritic cell maturation levels, along with increased tumor infiltration of CD8. + Increased T cells. Quantitative analysis ( Figure 16 (B) indicates that CD8 + T cell percentage and CD8 + / CD4 + An increase in the ratio, a decrease in the proportion of regulatory T cells, and an increase in the M1 / M2 macrophage ratio indicate that the tumor immunosuppression status has been improved.

[0102] Immunohistochemical and immunofluorescence results ( Figure 17 Further analysis showed that PD-L1 expression changed, while CD8 expression also changed. + Enhanced T-cell infiltration and increased HMGB1 release indicate that this treatment strategy can induce immunogenic cell death-related processes.

[0103] In terms of safety evaluation, such as Figure 18 As shown, Figure 18A shows the changes in tumor volume per mouse in each treatment group over time, while the body weight of mice in each group remained stable. Figure 18 (B) No obvious damage was observed in the H&E staining of the main organs. Figure 18 C), hematological and serum biochemical indicators ( Figure 18 The results for both D and H were within the normal range, indicating that the treatment system has good biocompatibility.

[0104] like Figure 19 As shown, the nanoparticles provided by this invention, under external light stimulation, trigger proton release through photoresponsive units, causing local instability or dissociation of the MOF structure, thereby achieving the controllable release of functional components such as copper ions. The released metal ions induce mitochondrial stress, disrupt protein homeostasis and redox balance in tumor cells; simultaneously, Idasanutlin regulates tumor metabolic state, increasing cell sensitivity to metal-dependent cell death; and the radioactive metal further provides an internal irradiation enhancement effect and supports in vivo tracing. Therefore, the nanoparticles of this invention can achieve a synergistic effect of photoresponsive metal release, metabolic regulation, and radiotherapy, not only enhancing anti-tumor effects but also inducing tumor immune microenvironment remodeling, thus achieving a synergistic effect of integrated treatment and diagnosis.

[0105] Finally, it should be noted that the above embodiments do not limit the present invention in any way. Those skilled in the art can make modifications and improvements based on the present invention. Therefore, any modifications or improvements made without departing from the spirit of the present invention are within the scope of protection claimed by the present invention.

Claims

1. A photoresponsive nanoparticle that controlslably enhances metal-dependent cell death and synergistically supports radionuclide therapy, characterized in that, The nanoparticles include copper-based MOF-199 metal-organic framework material, cyanin photoacid generator MEH, metabolic regulatory molecules, and targeting substances. The copper-based MOF-199 metal-organic framework material is doped with radioactive metal isotopes. 64 Cu; The metabolic regulatory molecules are selected from MDM2 inhibitors, glycolysis inhibitors, mitochondrial function enhancers, chemotherapy drugs, targeted therapy drugs, immunomodulators, or nucleic acid drugs. The targeting substance is selected from peptides, small molecules, antibodies, aptamers, or carbohydrate recognition molecules that can recognize tumor-associated receptors, cell surface proteins, or microenvironmental markers.

2. The method for preparing nanoparticles as described in claim 1, characterized in that, Includes the following steps: S1. Dissolve trimellitic acid and polyvinylpyrrolidone in N,N-dimethylformamide to obtain a trimellitic acid mixture; 64 CuCl2 solution and Cu(NO3)2·3H2O were added to N,N-dimethylformamide and mixed thoroughly to obtain a solution containing... 64 Cu solution, containing 64 The Cu solution was added to the pyromellitic acid mixture and mixed evenly. Then it was transferred to a high-pressure reactor for reaction. After the reaction was completed, the material was centrifuged, washed and dried to obtain the radioactive copper isotope-doped metal-organic framework material. S2. Disperse the radioactive copper isotope-doped metal-organic framework material in ethanol solvent, then add the cyanin-based photoacid generator MEH and metabolic regulation molecules to it, stir at room temperature in the dark, and then centrifuge and wash to obtain a composite nanomaterial simultaneously loaded with photoacid generator MEH and metabolic regulation molecules. S3. Disperse the composite nanomaterials in PBS buffer, then add the targeting substance, stir at room temperature, and then centrifuge, wash and dry to obtain nanoparticles.

3. The preparation method according to claim 2, characterized in that, In step S1, the molar ratio of pyromellitic acid, polyvinylpyrrolidone, and Cu(NO3)2·3H2O is 1:1-5:1.8-2.

5. 64 CuCl2 accounts for 0.001%-1% of the total molar amount of copper source.

4. The preparation method according to claim 2, characterized in that, In step S1, the reaction temperature is 80-120℃ and the reaction time is 6-24h.

5. The preparation method according to claim 2, characterized in that, In step S2, the mass ratio of the radioactive copper isotope-doped metal-organic framework material, the cyanine photoacid generator MEH, and the metabolic regulatory molecule is 1:0.02-0.1:0.05-0.

2.

6. The preparation method according to claim 2, characterized in that, In step S3, the mass ratio of the composite nanomaterial to the target substance is 1:0.2-1.

0.

7. The use of the nanoparticles as described in claim 1 or the nanoparticles prepared by the preparation method as described in any one of claims 2-6 in the preparation of drugs for treating breast cancer.

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