Copper-based nano coordination polymer as well as preparation method and application thereof

Abstract

The invention belongs to the field of new nano material technology and tumor treatment, and relates to a copper-based nano coordination polymer as well as a preparation method and application thereof. The copper-based nano coordination polymer is prepared from the following raw materials: copper ions, perfluorinated carbon and APR-246. The CuF16 at 246 has a remarkable cell killing effect, and when the concentration of the CuF16 at 246 is 200 mu g / ml, the survival rate of cancer cells is as low as about 7%, which indicates that the drug APR-246 loaded on the material can recover the function of P53, inhibit glycolysis and promote TCA circulation as a P53 reactivator, thereby further sensitizing copper death and promoting the occurrence of copper death.

Description

Technical Field

[0001] The invention belongs to the field of nano new material technology and tumor treatment, and relates to a copper-based nano coordination polymer and a preparation method and application thereof. Background Art

[0002] Copper is involved in many physiological processes in the body and plays an important role in the occurrence, development and treatment of tumors. Studies have shown that compared with normal cells, cancer cells have a higher demand for copper. The copper concentration in the serum or tumor tissue of patients with various cancers (breast cancer, colon cancer, lung cancer, melanoma and pancreatic cancer, etc.) increases, indicating that copper may be involved in the proliferation, angiogenesis and metastasis of cancer cells. Therefore, regulating copper in the human body has become a new target for cancer treatment. Copper death is a copper-dependent regulated cell death method that will not be blocked by inhibitors of cell signal transduction, and shows great application prospects in tumor treatment. By inducing copper death in cancer cells to achieve the purpose of anti-tumor, it provides a new idea for tumor treatment. Copper induces cell death by targeting lipoylatedTCAcycle proteins, published in the journal Science in March 2022, first proposed the concept of copper death and clarified the mechanism of copper death, that is, copper ion overload can lead to cell death: cellular copper homeostasis is unbalanced, copper ions will directly bind to ester-acylated proteins in the tricarboxylic acid cycle, and the FDX1 / LIAS-mediated LApathway (lipoic acid pathway) will induce oligomerization of lipoic acid-modified DLAT (key component of the pyruvate dehydrogenase complex), thereby causing abnormal metabolism of the tricarboxylic acid cycle, triggering cellular protein toxicity stress, and ultimately leading to cell death. This study shows that copper death is directly related to the tricarboxylic acid cycle metabolic pathway. However, the metabolic mode of cancer cells can affect the efficacy of copper death. In order to meet the energy needs of their own rapid growth, most cancer cells mainly metabolize in anaerobic glycolysis, which limits the TCA cycle in mitochondrial respiration, and is therefore not conducive to the occurrence of copper death. However, current related research has not solved this problem. For example, Shen et al. (Lu X, Chen X, Lin C, et al. Elesclomolloaded copper oxide nanoplatform triggers cuproptosis to enhance antitumor immunotherapy [J]. Adv Sci, 2024, 11 (18): e2309984.) constructed an intelligent nanosystem ES@CuO that induces cuproptosis by encapsulating copper oxide nanoparticles in copper ion carriers. ES@CuO can degrade and release Cu after pH response. 2+, triggering copper death. Although this study can achieve the effective delivery of copper ions, it does not solve the problem that copper death is easily inhibited; Xu et al. (Zhou J, Yu Q, Song J, et al. Photothermally triggered copper payload release for cuproptosis-promoted cancer synergistic therapy [J]. Angew Chem Int Ed Engl, 2023, 62 (12): e202213922.) used copper, gold nanorods, silica and disulfiram to construct a photothermally triggered nanoplatform Au@MSN-Cu / PEG / DSF. Under near-infrared laser irradiation, the copper-doped silica framework biodegrades and releases Cu 2+ In situ chelation with DSF generates highly cytotoxic bis(diethyldithiocarbamate)-copper (CuET). Although this study improved the toxicity of the copper nanoplatform, it still did not solve the problem of copper death being inhibited from the perspective of cancer cell metabolism. Summary of the invention

[0003] In order to solve the metabolic problem of cancer treatment by inducing cancer cells to undergo copper death at present, the present application sensitizes copper death by promoting cancer cell metabolism toward oxidative phosphorylation that is conducive to copper death.

[0004] P53 is a tumor suppressor gene that can inhibit cell carcinogenesis by regulating metabolism. For cancer cells, copper death can be promoted by inhibiting glycolysis and driving metabolism toward oxidative phosphorylation. In all malignant tumors, more than 50% will have mutations in this gene. For P53 mutations, APR-246 is selected as a P53 reactivator. APR-246 is a quinuclidine derivative that mainly targets mutant p53, can restore the wild-type conformation and function of mutant p53, and induce tumor cell death, which can achieve the effect of sensitizing copper death.

[0005] The technical solution of this application is as follows:

[0006] In a first aspect, the present application provides a copper-based nano-coordination polymer, characterized in that the raw materials of the copper-based nano-coordination polymer include copper ions and perfluorocarbon.

[0007] Perfluorocarbons, a group of chemically inert synthetic molecules with excellent biocompatibility, have been extensively studied or used in the clinic for a variety of purposes including artificial blood substitutes, organ preservation, ultrasound imaging, and fluorine magnetic imaging.

[0008] According to a specific embodiment of the present invention, the raw material further comprises APR-246; preferably, the added mass range of APR-246 is 2-5 mg.

[0009] According to a specific embodiment of the present invention, the perfluorocarbon is perfluorosebacic acid.

[0010] Perfluorosebacic acid, as a type of perfluorocarbon, has an electron-donating group and can coordinate with copper ions.

[0011] According to a specific embodiment of the present invention, the mass ratio of copper ions: perfluorosebacic acid is 1:(1-6); preferably, the mass ratio of copper ions: perfluorosebacic acid is 1:(2-4); more preferably, the mass ratio of copper ions: perfluorosebacic acid is 1:(2.5-3.2).

[0012] According to a specific embodiment of the present invention, the copper ions are derived from copper acetate or copper acetate.

[0013] In a second aspect of the present application, a method for preparing the above-mentioned copper-based nano-coordination polymer is provided, comprising the following steps: dissolving perfluorocarbon and APR-246 in a solvent and mixing and stirring, then adding copper ions, continuing to stir, dialyze, and freeze-dry to obtain a copper-based nano-coordination polymer.

[0014] According to a specific embodiment of the present invention, the solvent is DMF;

[0015] According to a specific embodiment of the present invention, stirring is carried out at room temperature for more than 12 hours;

[0016] According to a specific embodiment of the present invention, the copper ions are dissolved in a solvent before being added.

[0017] In a third aspect, the present application provides the use of the copper-based nano-coordination polymer or the copper-based nano-coordination polymer prepared by the above-mentioned preparation method in the preparation of anti-tumor drugs.

[0018] The principle of this application is: when the material CuF 16 When @246 accumulates in the tumor site, it will be internalized by cancer cells and enter the lysosome for cleavage. The released copper ions enter the mitochondria and the tricarboxylic acid cycle, eventually causing DLAT oligomerization and the disappearance of Fe-S cluster proteins, thereby inducing copper death. At the same time, the loaded drug small molecule APR-246 is released, which reactivates the mutant P53 and transforms it into wild-type P53, restores the function of the P53 gene, inhibits glycolysis, drives the metabolism towards oxidative phosphorylation, and sensitizes copper death. In addition, perfluorocarbon is used as an organic ligand for copper ions, which has low toxicity and can produce strong nuclear magnetic resonance signals, thereby achieving in vivo visualization. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1This is the transmission electron microscope image of CuF16;

[0020] Figure 2 This is the scanning electron microscope image of CuF16@246;

[0021] Figure 3 This is the transmission electron microscope image of CuF16@246;

[0022] Figure 4 This is the scanning electron microscope image of CuF16@246;

[0023] Figure 5 It is the particle size distribution diagram of CuF16 and CuF16@246;

[0024] Figure 6 The infrared spectra of perfluorosebacic acid (F-COOH for short), CuF16 and CuF16@246;

[0025] Figure 7 Simulated, XRD patterns of CuF16 and CuF16@246;

[0026] Figure 8 Thermogravimetric curves of CuF16 and CuF16@246;

[0027] Fig. 9 The cytotoxicity of perfluorosebacic acid, APR-246, CuF16 and CuF16@246.

[0028] Beneficial Effects of the Invention

[0029] CuF 16 @246 has a significant cell killing effect. 16 When the concentration of @246 was 200 μg / ml, the survival rate of cancer cells was as low as about 7%, indicating that the drug APR-246 loaded by the material, as a P53 reactivator, could restore P53 function, inhibit glycolysis, and promote TCA cycle, thereby further sensitizing copper death and promoting the occurrence of copper death. DETAILED DESCRIPTION

[0030] Example 1 Preparation and Analysis of Materials

[0031] 1. Synthesis of materials

[0032] (1)CuF 16 Synthesis

[0033] Dissolve Cu(CH3COO)2·H2O (30.5 mg) in 20 mL of DMF and stir at room temperature for 12 h. Then add DMF (10 mL) containing perfluorosebacic acid (25.0 mg) to the above solution and stir at room temperature for 12 h. The obtained material is dialyzed with a 3000D dialysis bag and freeze-dried to obtain the product CuF 16 .

[0034] (2)CuF 16 @246's Synthesis

[0035] Perfluorosebacic acid (25.0 mg) and APR-246 (2.0 mg) were dissolved in 10 mL of DMF and stirred at room temperature for 12 h. DMF (10 mL) containing Cu(CH3COO)2·H2O (30.5 mg) was then added to the above solution and stirred at room temperature for 12 h. The obtained material was dialyzed with a 3000D dialysis bag and freeze-dried to obtain the product CuF 16 @246.

[0036] 2 Characterization of materials

[0037] (1) TEM test

[0038] The synthesized CuF 16 CuF 16 @246 was dispersed in DMF and dispersed by ultrasound for 10 min to prepare a uniformly dispersed solution system. 10uL of the above solution was dropped on a copper mesh and evaporated to dryness at room temperature. 16 CuF 16 The morphology and particle size of @246 were observed and analyzed.

[0039] (2) SEM test

[0040] The synthesized CuF 16 CuF 16 @246 was dispersed in DMF and dispersed by ultrasound for 10 min to prepare a uniformly dispersed solution system. 10uL of the above solution was dropped on a silicon wafer and evaporated to dryness at room temperature. 16 CuF 16 The geometric morphology of @246 was observed and analyzed.

[0041] (3) Infrared spectrum test

[0042] The infrared spectrometer was used to analyze the perfluorosebacic acid, CuF 16 and CuF 16 @246 Conduct infrared spectroscopy test.

[0043] (4) DLS diameter test

[0044] The laser particle size analyzer was used to analyze the CuF 16 CuF 16 The particle size of @246 solution was characterized.

[0045] (5) XRD test

[0046] CuF 16 CuF 16 @246 After sufficient drying, the CuF 16 CuF 16 @246 Load the powder onto the sample stage, adjust the appropriate position and angle, set the scanning range (2°~50°) and speed, and start X-ray irradiation of the sample for measurement.

[0047] (6) TGA test

[0048] CuF 16 CuF 16 @246 After sufficient drying, take CuF 16 CuF 16 @246 Powder was subjected to TGA test.

[0049] 3 Results Analysis

[0050] At room temperature, Cu 2+ The copper-based nano-coordination polymer CuF was successfully synthesized by coordination with perfluorosebacic acid. 16 From TEM Figure 1 With SEM Figure 2 The image shows the synthesized CuF 16 It is similar to a spherical structure with a particle diameter of 100±10nm, uniform size and monodisperse. Figure 6 Perfluorosebacic acid, CuF 16 and CuF 16 The infrared spectrum of @246 shows that compared with perfluorosebacic acid, CuF 16 In the spectrum, at 1400 cm -1 The asymmetric stretching peak of Cu-O bond appears at 3512cm -1 The disappearance of the carboxyl peak at , thus verifying the formation of CO-Cu coordination bond, indicating that CuF 16 Cu 2+ The coordination reaction with the carboxyl group in perfluorosebacic acid was successful. Figure 7 According to the simulated single crystal XRD pattern (Simulated), the main peaks are located at 5.239°, 10.489°, 15.747° and 16.071°. The experimental group material CuF 16 The peak position is consistent with that of CuF 16 Successfully synthesized.

[0051] Material CuF after drug loading 16 @246 from TEM Figure 3 With SEM Figure 4 The image shows the synthesized CuF 16 @246 is also a spherical structure, through Figure 5 The particle diameter increases to 130±10nm, and the size is uniform and monodisperse. Figure 6 , compared with perfluorosebacic acid, CuF 16 @246 spectrum at 1400cm -1 The asymmetric stretching peak of Cu-O bond appears at 3512cm -1 The disappearance of the carboxyl peak at , thus confirming the formation of CO-Cu coordination bond; indicating that CuF 16 @Cu in 246 2+ The coordination reaction with the carboxyl group in perfluorosebacic acid was successful. Figure 7 , the material CuF after drug loading 16 @246 crystal structure and CuF 16 Slightly different, the main peaks of the material at 5.239°, 10.489° and 15.747° did not change, indicating that the MOF material did not collapse and crystallized well. However, the main peak at 16.071° disappeared, indicating that the successful loading of the drug changed the original crystal structure, which may be caused by the carbonyl oxygen atom / tertiary amine nitrogen atom in the drug APR-246 competing with water for coordination. Then, the thermogravimetric curve TGA test Figure 8 , according to the simulated CuF 16 Structural calculations show that the mass percentage of water molecules in the material is 7.35209%, indicating that CuF 16 The thermal stability is good before 296℃, and the weight loss of 7.35209% in the first stage is attributed to the departure of coordinated water molecules in the material, which proves that the material has good phase purity. After 296℃, the framework collapses in two stages, indicating that the organic ligands decompose. CuF 16 @246 has good thermal stability in the first stage before 199℃, and the weight loss of 5.30801% is attributed to the departure of coordinated water molecules in the material. Compared with Cu-MOF, the coordinated water is reduced by 2.04399%, which is speculated that 27.8018% of the coordinated water in the material is replaced by drugs. 16 @246 During the skeleton collapse, there were three stages of weightlessness and CuF 16 The later stage of weightlessness has one more stage, proving the successful loading of the drug in the material.

[0052] Example 2

[0053] Study on anti-tumor effect in vitro

[0054] At the cellular level, we investigated the role of CuF 16 CuF 16 @246 has the ability to kill cancer cells. The CCK-8 kit was used to detect the effects of perfluorosebacic acid, APR-246, and CuF 16 and CuF 16 @246 toxicity to cancer cells, the results are as follows Fig. 9 It was found that the survival rate of cancer cells hardly changed with the concentration of perfluorodecanedioic acid, indicating that perfluorodecanedioic acid has low toxicity and high biological safety. 16 As the concentration increases, the survival rate of cancer cells gradually decreases, and at 200 μg / ml, it is as low as about 65%, indicating that the cytotoxicity may be caused by Cu 2+ Caused by copper death; CuF 16 In comparison, CuF 16 @246 has a more significant cell-killing effect. At a concentration of 200 μg / ml, the survival rate of cancer cells is as low as about 7%, indicating that the drug APR-246 loaded in the material acts as a P53 reactivator, which can restore P53 function, inhibit glycolysis, and promote TCA cycle, thereby further sensitizing copper death and promoting the occurrence of copper death.

Claims

1. A copper-based nano-coordination polymer, characterized in that: The raw materials of the copper-based nano coordination polymer include copper ions and perfluorocarbon.

2. The copper-based nano-coordination polymer according to claim 1, characterized in that: The raw materials also include APR-246.

3. The copper-based nano-coordination polymer according to claim 1, characterized in that: The copper ions are derived from organic copper.

4. The copper-based nano-coordination polymer according to any one of claims 1 to 3, characterized in that: The perfluorocarbon is perfluorosebacic acid; And / or, the mass ratio of copper ion:perfluorosebacic acid is 1:(1-6); preferably, the mass ratio of copper ion:perfluorosebacic acid is 1:(2-4); more preferably, the mass ratio of copper ion:perfluorosebacic acid is 1:(2.5-3.2).

5. The copper-based nano-coordination polymer according to any one of claims 1 to 3, characterized in that: The copper ions are derived from copper acetate or copper acetate.

6. A method for preparing the copper-based nano-coordination polymer according to any one of claims 1 to 5, characterized in that: The following steps are involved: Perfluorocarbon and APR-246 are dissolved in a solvent and mixed and stirred, and copper ions are added, and the mixture is stirred continuously, dialyzed, and freeze-dried to obtain a copper-based nano-coordination polymer.

7. The preparation method according to claim 6, characterized in that: The solvent was DMF.

8. The preparation method according to claim 6, characterized in that: The stirring was carried out at room temperature for more than 12 h.

9. The preparation method according to claim 6, characterized in that: The copper ions are dissolved in a solvent before being added.

10. Use of the copper-based nano-coordination polymer described in any one of claims 1 to 5 or the copper-based nano-coordination polymer prepared by the preparation method described in any one of claims 6 to 8 in the preparation of anti-tumor drugs.

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