A copper-based nano coordination polymer and a preparation method and application thereof

Abstract

The application belongs to the technical field of new nano materials and tumor treatment, and relates to a copper-based nano coordination polymer as well as a preparation method and application thereof. The raw materials of the copper-based nano coordination polymer include copper ions, perfluorocarbon and APR-246. 16 When the concentration of CuF 16 When the concentration of CuF @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 by the material can restore the P53 function, inhibit glycolysis, promote TCA cycle, and further sensitize copper death and promote the occurrence of copper death.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of new nanomaterials and tumor treatment, and relates to a copper-based nano coordination polymer as well as a preparation method and application thereof. BACKGROUND

[0002] Copper participates in various physiological processes in the body and plays an important role in the occurrence, development and treatment of tumors. Studies have shown that cancer cells have a higher demand for copper than normal cells, and 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.) is increased, which indicates that copper may be involved in the processes of proliferation, angiogenesis and metastasis of cancer cells. Therefore, regulating copper in the human body has become a new target for cancer treatment, and copper death is a copper-dependent regulatory cell death mode that cannot be blocked by cell signal transduction inhibitors and has shown great application prospects in tumor treatment. Inducing copper death in cancer cells to achieve anti-tumor purposes provides a new idea for tumor treatment. In March 2022, Copper induces cell death by targeting lipoylated TCA cycle proteins was published in Science, which first proposed the concept of copper death and elucidated the mechanism of copper death, i.e., copper ion overload will lead to cell death: imbalance of cellular copper homeostasis, copper ions will directly bind to esterified proteins in the tricarboxylic acid cycle, and FDX1 / LIAS-mediated LA pathway (lipoic acid pathway) will induce oligomerization of DLAT (pyruvate dehydrogenase complex key component) modified by lipoic acid, thereby disrupting the tricarboxylic acid cycle metabolism, triggering cell protein toxicity stress, and ultimately leading to cell death. The study shows that copper death is directly related to the tricarboxylic acid cycle metabolic pathway. However, the metabolic mode of cancer cells will affect the efficacy of copper death, and most cancer cells will mainly use anaerobic glycolysis to meet their energy needs for rapid growth, which limits the TCA cycle in mitochondrial respiration, and thus is not conducive to the occurrence of copper death. However, the current related research has not solved this problem, for example, Shen et al. (Lu X, Chen X, Lin C, et al. Elesclomol loaded copper oxide nanoplatform triggers cuproptosis to enhance antitumor immunotherapy [J]. Adv Sci, 2024, 11(18): e2309984.) constructed an intelligent copper death-inducing nano system ES@CuO by encapsulating copper oxide nanoparticles in a copper ion carrier. ES@CuO can release Cu 2+, which can induce copper death. Although this study can achieve 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 undergoes biodegradation, releasing Cu 2+ which in situ chelates with DSF to form bis(diethyldithiocarbamate)-copper (CuET) with high cytotoxicity. Although this study improves the toxicity of copper nanoplatforms, it still does not address the problem of copper death inhibition from the perspective of cancer cell metabolism. SUMMARY

[0003] In view of the existing problems in cancer treatment by inducing copper death in cancer cells, the present application sensitizes copper death by promoting the development of cancer cell metabolism towards oxidative phosphorylation, which is conducive to the occurrence of copper death.

[0004] P53 is a tumor suppressor gene that can inhibit cell carcinogenesis by regulating metabolism. For cancer cells, inhibiting glycolysis and driving the metabolic mode towards oxidative phosphorylation can promote the occurrence of copper death. Among all malignant tumors, more than 50% will have mutations in this gene. In view of the P53 mutation, APR-246 is selected as a P53 reactivator. APR-246 is a quinuclidone derivative that mainly targets mutant p53, restores the wild-type conformation and function of mutant p53, and induces tumor cell death, which can achieve the effect of sensitizing copper death.

[0005] The technical solutions of the present application are as follows:

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

[0007] Perfluorocarbons, a group of chemically inert, biocompatible synthetic molecules, have been extensively studied or used in clinical practice for various purposes, including artificial blood replacement, organ preservation, ultrasound imaging and fluoromagnetic imaging.

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

[0009] According to the embodiment of the present application, the perfluorocarbon is perfluorodecanoic acid.

[0010] As one of perfluorocarbons, perfluorodecanoic acid has an electron-donating group, which can realize coordination with copper ions.

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

[0012] According to the embodiment of the present application, the copper ions are derived from copper acetate or acetic acid copper.

[0013] In the second aspect of the present application, a preparation method of the above-mentioned copper-based nano coordination polymer is provided, which comprises the following steps: dissolving perfluorocarbon and APR-246 in a solvent and stirring, then adding copper ions and continuing to stir, dialysis, and freeze-drying to obtain the copper-based nano coordination polymer.

[0014] According to the embodiment of the present application, the solvent is DMF;

[0015] According to the embodiment of the present application, the stirring is carried out at room temperature, and the time is more than 12 h.

[0016] According to the embodiment of the present application, the copper ions are dissolved with the solvent before being added.

[0017] In the third aspect of the present application, the above-mentioned copper-based nano coordination polymer or the copper-based nano coordination polymer prepared by the above-mentioned preparation method is applied to the preparation of an antitumor drug.

[0018] The principle of the present application is that when the material CuF 16 @246 is enriched in the tumor site, it is endocytosed into lysosomes to be lysed, the released copper ions enter mitochondria, enter the tricarboxylic acid cycle, and ultimately trigger the oligomerization of DLAT and the disappearance of Fe-S cluster proteins, thereby triggering copper death; at the same time, the loaded small molecule drug APR-246 is released, so that the mutant P53 is reactivated to wild-type P53, the function of the P53 gene is restored, glycolysis is inhibited, the metabolic mode is driven to oxidative phosphorylation, and copper death is sensitized; in addition, perfluorocarbon is used as an organic ligand of copper ions, which has low toxicity and can produce a strong nuclear magnetic signal, thereby realizing in vivo visualization. BRIEF DESCRIPTION OF DRAWINGS

[0019] Figure 1Transmission electron microscopy image of CuF16;

[0020] Figure 2 Scanning electron microscopy image of CuF16@246;

[0021] Figure 3 Transmission electron microscopy image of CuF16@246;

[0022] Figure 4 Scanning electron microscopy image of CuF16@246;

[0023] Figure 5 Particle size distribution of CuF16 and CuF16@246;

[0024] Figure 6 Infrared spectrum of perfluorodecanedioic acid (F-COOH), CuF16 and CuF16@246;

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

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

[0027] Figure 9 Cytotoxicity of perfluorodecanedioic acid, APR-246, CuF16 and CuF16@246.

[0028] Advantages of the present application

[0029] CuF 16 @246 has a significant cell killing effect, when the concentration of CuF 16 @246 is 200 μg / ml, the survival rate of cancer cells is as low as about 7%, which shows that the drug APR-246 loaded by the material can restore P53 function, inhibit glycolysis, promote TCA cycle, and further sensitize copper death and promote the occurrence of copper death. DETAILED DESCRIPTION

[0030] Preparation and analysis of materials in Example 1

[0031] 1. Synthesis of materials

[0032] (1) Synthesis of CuF 16

[0033] ​Cu(CH3COO)2-H2O (30.5 mg) was dissolved in 20 mL of DMF and stirred at room temperature for 12 h. Then perfluorodecanoic acid (25.0 mg) was added to the above solution in DMF (10 mL) and stirred at room temperature for 12 h. The resulting material was dialyzed in a 3000D dialysis bag and the product CuF 16 .

[0034] (2) CuF 16 @246

[0035] Perfluorodecanoic 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. Then Cu(CH3COO)2-H2O (30.5 mg) was added to the above solution in DMF (10 mL) and stirred at room temperature for 12 h. The resulting material was dialyzed in a 3000D dialysis bag and the product CuF 16 @246

[0036] 2 Characterization of the material

[0037] (1) TEM test

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

[0039] (2) SEM test

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

[0041] (3) Infrared spectroscopy test

[0042] Perfluorodecanoic acid, CuF 16 @246 and CuF 16 @246 were tested by infrared spectroscopy.

[0043] (4) DLS diameter test

[0044] The particle size of CuF 16 was characterized by laser particle size analyzer. 16 @246 The particle size of CuF

[0045] (5) XRD test

[0046] CuF 16 was characterized by XRD. 16 @246 After CuF 16 was dried sufficiently, the CuF 16 powder was loaded onto the sample stage, the scanning range (2°-50°) and speed were set after adjusting the appropriate position and angle, and the X-ray irradiation sample was started for measurement.

[0047] (6) TGA test

[0048] CuF 16 was characterized by TGA. 16 @246 After CuF 16 was dried sufficiently, the CuF 16 powder was subjected to TGA test.

[0049] 3 Result analysis

[0050] At room temperature, CuF 2+ was successfully synthesized by coordinating Cu 16 with perfluorodecanoic acid. From the TEM Figure 1 and SEM Figure 2 images, it was observed that the synthesized CuF 16 had a spherical structure, with a particle diameter of 100±10 nm, uniform size and monodispersity. Figure 6 The infrared spectra of perfluorodecanoic acid, CuF 16 and CuF 16 @246 were compared. Compared with perfluorodecanoic acid, in the spectrum of CuF 16 , the asymmetric stretching peak of Cu-O bond appeared at 1400 cm -1 , and the carboxyl peak at 3512 cm -1 disappeared, thereby verifying the formation of C-O-Cu coordination bond and indicating that the Cu 16 in CuF 2+ successfully coordinated with the carboxyl group in perfluorodecanoic acid. According to Figure 7 , according to the simulated single crystal XRD pattern (Simulated), the main peaks were located at 5.239°, 10.489°, 15.747° and 16.071°, which were consistent with the peak positions of the experimental group material CuF 16 , indicating that it had good phase purity, and indicating that CuF 16 was successfully synthesized.

[0051] CuF after drug loading 16 @246From TEM Figure 3 With SEM Figure 4 Image observation of synthesized CuF 16 @246Also for spherical structure, by Figure 5 Particle diameter increased to 130 ± 10 nm, uniform size and monodisperse. According to the infrared spectrum Figure 6 , compared with perfluorosuberic acid, CuF 16 @246In the spectrum at 1400 cm -1 , the asymmetric stretching peak of Cu-O bond, and the disappearance of carboxyl peak at 3512 cm -1 , thus also verified the formation of C-O-Cu coordination bond; CuF 16 @246in Cu 2+ and carboxyl in perfluorosuberic acid successfully coordinated. According to Figure 7 , the crystal structure of CuF 16 @246after drug loading is slightly different from CuF 16 , the main peak of the material at 5.239°, 10.489° and 15.747° did not change, indicating that the MOF material did not collapse the skeleton and crystallized well. But the main peak at 16.071° disappeared, indicating the successful loading of the drug, which changed the original crystal structure, which may be caused by the competition of the carbonyl oxygen atom / tertiary amine nitrogen atom in the drug APR-246 with water. Then, through the thermal gravimetric curve TGA test Figure 8 , according to the structure calculation of CuF 16 , it is found that the mass percentage of water molecules in the material is 7.35209%, indicating that CuF 16 has good thermal stability before 296℃, and the first stage weight loss of 7.35209% is attributed to the removal of coordinated water molecules in the material, which proves that the material has good phase purity. After 296℃, the skeleton collapses in two stages, indicating that the organic ligand decomposes. CuF 16 @246has good thermal stability before 199℃, and the weight loss of 5.30801% is attributed to the removal of coordinated water molecules in the material, which is 2.04399% less than that of Cu-MOF, which is speculated to be the replacement of 27.8018% of the coordinated water in the material by the drug. After 199℃, CuF 16 @246has three stages of weight loss in the process of skeleton collapse, compared with CuF 16 , the later weight loss has an additional stage, which proves the successful loading of the drug in the material.

[0052] Example 2

[0053] In vitro antitumor effect study

[0054] At the cellular level, we explored CuF 16 With CuF 16 The killing effect of APR-246 on cancer cells was investigated using a CCK-8 assay kit. 16 and CuF 16 The toxic effects of @246 on cancer cells were as follows: Figure 9 As shown, it was found that the survival rate of cancer cells remained almost unchanged with variations in perfluorosebaic acid (PFSA) concentration, indicating that PFSA has low toxicity and high biocompatibility. In contrast, CuF... 16 As the concentration increases, the survival rate of cancer cells gradually decreases, reaching as low as 65% at 200 μg / ml, indicating that the cytotoxicity may be due to Cu. 2+ Caused by copper death; related to CuF 16 In comparison, CuF 16 @246 exhibits 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 on the material, as a P53 reactivator, can restore P53 function, inhibit glycolysis, promote TCA cycle, and thus further sensitize copper death and promote its occurrence.

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; The raw materials further include APR-246; The perfluorocarbon is perfluorodecanoic acid; The mass ratio of the copper ions to the perfluorodecanoic acid is 1:(1-6).

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

3. The copper-based nano-coordination polymer according to claim 1, characterized in that, The mass ratio of the copper ions to the perfluorodecanoic acid is 1:(2-4).

4. The copper-based nano-coordination polymer according to claim 3, characterized in that, The mass ratio of the copper ions to the perfluorodecanoic acid is 1:(2.5-3.2).

5. The copper-based nano-coordination polymer according to any one of claims 1-4, wherein, 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 1-5, characterized by, The method comprises the following steps: The perfluorocarbon and the APR-246 are dissolved in a solvent and stirred, then the copper ions are added and the stirring is continued, and the copper-based nano coordination polymer is obtained after dialysis and freeze-drying.

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

8. The preparation method according to claim 6, characterized in that, The stirring is carried out at room temperature and the time is more than 12 hours.

9. The production method according to claim 7 or 8, characterized by, The copper ions are dissolved in the solvent before being added.

10. The copper-based nano coordination polymer of any one of claims 1-5 or prepared by the preparation method of any one of claims 6-9 in the preparation of an antitumor drug.

Images