Thiadiazole tetranuclear copper complex with antitumor and antibacterial activity, and preparation method and application thereof
The thiadiazole tetranuclear copper complex [Cu4(C3H3S2N2)2(C3H4S2N2)4] was synthesized by solvothermal reaction, which solved the problems of side effects of platinum drugs and easy oxidation of monovalent copper complexes, and achieved low-toxicity antitumor and antibacterial effects.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- XI AN JIAOTONG UNIV
- Filing Date
- 2025-12-10
- Publication Date
- 2026-07-03
AI Technical Summary
Existing platinum-based anticancer drugs have serious side effects, and monovalent copper complexes are easily oxidized in air, which limits their synthesis and application.
The thiadiazole tetranuclear copper complex [Cu4(C3H3S2N2)2(C3H4S2N2)4] was synthesized by solvothermal reaction. Using CuX, thiadiazole and triethylamine as raw materials, the reaction was carried out at a specific temperature and solvent to form a copper complex with antitumor and antibacterial activities.
The prepared copper complexes exhibited good cytotoxicity and antibacterial properties, showing significant cytotoxic effects on tumor cells and effectively inhibiting Gram-positive and Gram-negative bacteria, providing a potential drug option for low-toxicity anticancer and antibacterial treatments.
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Figure CN121494873B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of anticancer chemical drug technology, specifically to thiadiazole tetranuclear copper complexes with antitumor and antibacterial activities, their preparation methods, and applications. Background Technology
[0002] Metal complexes, due to their unique metal-centric coordination modes, have become an indispensable class of compounds in the development of clinical anticancer drugs. Since the advent of cisplatin and related platinum-based drugs, the development of metal-based anticancer drugs has become a research hotspot. However, the serious side effects caused by existing platinum-based complexes have prompted researchers to dedicate themselves to designing and developing novel transition metal anticancer drugs with low toxicity. Transition metal complexes, with their rich coordination geometries and coordination numbers, diverse redox states and electronic properties, unique thermodynamic and kinetic behaviors, and the inherent properties of the metal cations and ligands themselves, show significant application potential in the development of novel metal-based anticancer drugs.
[0003] Copper, a transition metal, is an essential trace element for human survival. It can undergo Fenton-like reactions, catalyzing the production of reactive oxygen species (ROS), affecting the redox balance within tumor cells, and inducing cell death through various mechanisms, such as activating stress pathways, arresting the cell cycle, inhibiting angiogenesis, promoting copper proliferation and apoptosis, and ferroptosis. Copper also possesses anti-inflammatory and antibacterial properties. Due to their broad catalytic activity and electrochemical reactivity, copper complexes have become a promising option for cancer treatment. Monovalent copper complexes are typically synthesized under inert gas protection and are readily oxidized to divalent copper in air, which limits the synthesis and application of cuprous complexes. Summary of the Invention
[0004] To overcome the shortcomings of existing technologies, a thiadiazole tetranuclear copper complex with antitumor and antibacterial activities and its preparation method are provided.
[0005] To achieve the above objectives, the present invention adopts the following technical solution:
[0006] In a first aspect, the present invention provides a thiadiazole tetranuclear copper complex with antitumor and antibacterial activities, wherein the molecular formula of the structure is [Cu4(C3H3S2N2)2(C3H4S2N2)4], where C3H4S2N2 is thiadiazole, also known as 2-methyl-5-mercapto-1,3,4-thiadiazole in Chinese and English.
[0007] English aliases: 1,3,4-Thiadiazole-2(3H)-thione, 5-methyl-; 5-Methyl-1,3,4-thiadiazole-2-thiol; 2-METHYL-5-MERCAPTO THIADIAZOLE (MMTD); MMTD; The structure of the complex is as follows:
[0008] .
[0009] Furthermore, the crystal structure of the complex is monoclinic, with space group C2 / c, and cell parameters: a = 45.33(3) Å, b = 7.869(5) Å, c = 19.582(14) Å, α = 90˚, β = 95.523(9)°, γ = 90˚, Z = 8, and unit cell volume V = 6953(8) Å. 3 The entire structure consists of a thiadiazole ligand and four +1 copper ions. The neutral Cu4 core comprises four Cu(I) ions, four deprotonated mmt⁻ ligands, and two Hmmt ligands. Each deprotonated mmt⁻ ligand bridges an adjacent Cu(I) ion in a 3.21 coordination mode via its thiol S atom and nitrogen atom. Simultaneously, the two Hmmt ligands coordinate with the two Cu(I) ions in a monodentate manner to meet their coordination requirements. The coordination spheres at the centers of Cu1 and Cu2 are defined by a nitrogen atom and two μ2-S atoms derived from the mmt– ligands. In contrast, the Cu3 and Cu4 ions are in an environment exhibiting a distorted tetrahedral coordination geometry; the complex is yellow.
[0010] Secondly, the present invention provides a method for preparing a thiadiazole tetranuclear copper complex with antitumor and antibacterial activities, comprising the following steps:
[0011] Using CuX, thiadiazole, and triethylamine as raw materials, a solvothermal reaction is carried out. During the reaction, triethylamine causes some of the thiadiazole to deprotonate, yielding deprotonated thiadiazole, denoted as mmt⁻ ligand. The undeprotonated thiadiazole is denoted as Hmmt ligand. Each complex is composed of four Cu(I) ions, four mmt⁻ ligands, and two Hmmt ligands. The four mmt⁻ ligands bridge adjacent Cu(I) ions in a 3.21 coordination mode through the sulfur and nitrogen atoms of their thiols. At the same time, the two Hmmt ligands coordinate with the two Cu(I) ions in a monodentate coordination mode to obtain the complex, wherein X is I or Br.
[0012] Furthermore, the molar ratio of CuX, thiadiazole, and triethylamine is 1:1.5 to 2:2, or 1:1.5 to 2.0:2.
[0013] Furthermore, the solvent in the solvothermal reaction is an equal volume mixture of CH3OH and H2O.
[0014] Furthermore, the temperature of the solvothermal reaction is 100~110℃, and the time is 48 h~72 h.
[0015] Thirdly, the present invention provides the application of the thiadiazole tetranuclear copper complex with antitumor and antibacterial activities in the preparation of antitumor drugs.
[0016] Furthermore, the tumor is breast cancer.
[0017] Fourthly, the present invention provides the application of the thiadiazole tetranuclear copper complex with antitumor and antibacterial activities in the preparation of antibacterial agents.
[0018] Furthermore, the bacteria include Gram-positive bacteria and Gram-negative bacteria, wherein the Gram-positive bacteria are Staphylococcus aureus and the Gram-negative bacteria are Escherichia coli.
[0019] Compared with the prior art, the present invention has the following beneficial effects:
[0020] The copper group cluster complexes prepared by the method of this invention exhibited good cytotoxicity (IC50) as confirmed by the MTT assay. 50 =7.57 μM), with corresponding ligand thiadiazole activity (IC50). 50 Compared to (0.87 mM), the addition of copper ions significantly improved the cytotoxicity of the complex. It also exhibits inhibitory effects against Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli, thus serving as a candidate antitumor drug with antibacterial properties. Attached Figure Description
[0021] Figure 1 This is a crystal structure diagram of [Cu4(C3H3S2N2)2(C3H4S2N2)4] in Example 1 of the present invention.
[0022] Figure 2 The X-ray photoelectron spectra of the [Cu4(C3H3S2N2)2(C3H4S2N2)4] complex and the X-ray photoelectron spectra of copper 2p are shown in Example 1 of this invention.
[0023] Figure 3 Thermogravimetric analysis (TGA) of the [Cu4(C3H3S2N2)2(C3H4S2N2)4] complex in Example 1 of this invention.
[0024] Figure 4 This is a high-resolution mass spectrum of the [Cu4(C3H3S2N2)2(C3H4S2N2)4] complex in Example 1 of the present invention.
[0025] Figure 5 The results show the effect of the [Cu4(C3H3S2N2)2(C3H4S2N2)4] complex on the cytotoxicity of MCF-7 cells in Example 1 of this invention.
[0026] Figure 6 This is the result of the [Cu4(C3H3S2N2)2(C3H4S2N2)4] complex on the production of ROS in MCF-7 cells in Example 1 of this invention.
[0027] Figure 7 The image shows the expression results of GPX-4 and FDX-1 in MCF-7 cells after treatment with the [Cu4(C3H3S2N2)2(C3H4S2N2)4] complex in Example 1 of this invention.
[0028] Figure 8 The results of the antibacterial activity of the [Cu4(C3H3S2N2)2(C3H4S2N2)4] complex in Example 1 of this invention against Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli are shown. Detailed Implementation
[0029] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments, but this should not be construed as limiting the invention. Unless otherwise specified, the technical means used in the following embodiments are conventional means well known to those skilled in the art, and the materials, reagents, etc. used in the following embodiments are commercially available unless otherwise specified.
[0030] Example 1
[0031] Weigh 1.0 mmol CuBr, 1.5 mmol thiadiazole and 2.0 mmol triethylamine into a beaker, add a mixture of 4.5 ml methanol and 4.5 ml deionized water, mix and pour into a 15 ml polytetrafluoroethylene bottle, react in a temperature-controlled oven at 110 °C for 72 hours, and after 24 hours cool to room temperature to obtain pure yellow block crystals. The molecular structure of the crystals was determined to be [[Cu4(C3H3S2N2)2(C3H4S2N2)4] by single-crystal X-ray diffraction. The crystals were thoroughly washed with ethanol and dried in air at room temperature.
[0032] Example 2
[0033] Weigh 1.0 mmol CuI, 1.5 mmol thiadiazole and 2.0 mmol triethylamine into a beaker, add a mixture of 4.5 ml methanol and 4.5 ml deionized water, mix and pour into a 15 ml polytetrafluoroethylene bottle, react in a temperature-controlled oven at 110 °C for 72 hours, and after 24 hours cool to room temperature to obtain pure yellow block crystals. The molecular structure of the crystals was determined to be [Cu4(C3H3S2N2)2(C3H4S2N2)4] by single-crystal X-ray diffraction. The crystals were thoroughly washed with ethanol and dried in air at room temperature.
[0034] Example 3
[0035] Weigh 1.0 mmol CuI, 1.5 mmol thiadiazole and 2.0 mmol triethylamine into a beaker, add a mixture of 4.5 ml methanol and 4.5 ml deionized water, mix and pour into a 15 ml polytetrafluoroethylene bottle, react in a temperature-controlled oven at 100 °C for 72 hours, and after cooling to room temperature for 24 hours, obtain pure yellow block crystals. The molecular structure of the crystals was determined to be [Cu4(C3H3S2N2)2(C3H4S2N2)4] by single-crystal X-ray diffraction. The crystals were thoroughly washed with ethanol and dried in air at room temperature.
[0036] The complexes of Examples 1-3 (abbreviated as Cu4) were determined by X-ray single-crystal diffraction to be monoclinic, with space group C2 / c, and cell parameters: a=45.33(3) Å, b=7.869(5) Å, c=19.582(14) Å, α=90˚, β=95.523(9)°, γ=90˚, Z=8, and unit cell volume V=6953(8) Å. 3 Crystal structure such as Figure 1 As shown, the entire structure consists of a thiadiazole ligand and four +1 copper ions. The neutral Cu4 core comprises four Cu(I) ions, four deprotonated mmt⁻ ligands, and two Hmmt ligands. Each deprotonated mmt⁻ ligand bridges adjacent Cu(I) ions in a 3.21 coordination mode via its thiol sulfur atom and nitrogen atom. Simultaneously, the two Hmmt ligands coordinate with the two Cu(I) ions in a monodentate manner to meet their coordination requirements. The coordination spheres at the centers of Cu1 and Cu2 are defined by a nitrogen atom and two μ2-S atoms derived from the mmt⁻ ligands. In contrast, the Cu3 and Cu4 ions are in an environment exhibiting a distorted tetrahedral coordination geometry; the complex is yellow. The X-ray photoelectron spectra of the complex and the X-ray photoelectron spectra of copper 2p are shown below. Figure 2 As shown, the mass change at different temperatures is as follows: Figure 3 As shown in Table 1, the crystallographic data of the coordination compounds are presented.
[0037] Table 1 Crystallographic data of the coordination compounds
[0038]
[0039] Example 4
[0040] MCF-7 cells were seeded at a density of 5 × 10³ cells per well in 96-well plates. The plates were incubated at 37°C with 5% CO2 for 24 hours. Then, DMEM complete medium containing different concentrations of Cu4 prepared in Example 1 was added. After 48 hours of further incubation, 20 µL of 5 mg / mL MTT solution was added to each well, and the cells were incubated for another 4 hours. The supernatant was then carefully removed, and 150 µL of dimethyl sulfoxide (DMSO) was added to each well. The plates were gently shaken for 10 minutes. Finally, the absorbance was measured at 490 nm using a Multiskan Sky reader. This experiment was independently repeated three times to ensure reproducibility. The half-maximal inhibitory rate (IC50) was also typically used. 50 The IC50 value is used to measure the strength of cytotoxicity. 50 Defined as: the concentration of drug required to kill half of the cells.
[0041] Table 2 IC50 of the complex on cells 50 value
[0042]
[0043] like Figure 4 , Figure 5 The MTT assay results shown in Table 2 indicate that the copper cluster complex exhibits good toxicity against MCF-7 and 4TI cells.
[0044] Example 5
[0045] Effect of Cu4 on intracellular ROS levels in MCF-7 cells
[0046] To study the production of intracellular reactive oxygen species (ROS), 3 × 10⁻⁶ 5 MCF-7 cells were seeded in 35 mm culture dishes and cultured overnight at 37°C and 5% CO2. Cu4 was then added, and cells were incubated for 12 hours at different concentrations. After incubation, the cells were washed three times with phosphate-buffered saline (PBS), and 1 mL of serum-free DMEM containing DCFH-DA (0.5 μL 10 μmol solution) was added to each well. Cells were then incubated for another 30 minutes. After removing the staining medium, the cells were washed three times with PBS and observed using a fluorescence microscope.
[0047] like Figure 6As shown, the effect of Cu4 on ROS levels in MCF-7 cells was investigated using the DCFHDA probe. DCFH-DA reacts with ROS to generate green fluorescent DCF. The results showed that after treatment with different concentrations of Cu4 for 4 hours, the intensity of green fluorescence in MCF-7 cells increased in a concentration-dependent manner.
[0048] Example 6
[0049] To further investigate the mechanisms of tumor cell death, we explored the expression of key regulators in the apoptosis, ferroptosis, and pachytrophic death pathways, as detailed below.
[0050] MCF-7 cells were used at a rate of 4 × 10⁴ cells per well. 5 Cells were seeded at a density of 1000 g / well in 6-well plates and incubated for 24 hours. After incubation, cells were treated with Cu4 for a specified time. Following treatment, cells were washed twice with PBS, and then lysed by adding an appropriate amount of lysis buffer to each well. The cell mixture was incubated on ice for 20 minutes, then gently pipetted a few times and collected into 1.5 ml EP tubes. The supernatant was obtained by centrifugation at 14,000 × g for 15 minutes at 4 °C, and the total protein concentration was determined using a BCA protein quantification kit. After separation by SDS-PAGE on a 12% gel, the protein was transferred to a PVDF membrane. The membrane was blocked with 5% skim milk powder solution at room temperature for 1 hour, and then incubated overnight at 4 °C with primary antibodies: anti-Caspase3, anti-GPX4, anti-FDX1, and anti-Beta Actin. The membrane was then incubated with secondary antibodies and developed using an ECL detection kit.
[0051] In our study, we assessed GPX4 protein levels in Cu4-treated MCF-7 cells, such as... Figure 7 As shown, we observed that Cu4 significantly reduced the expression of Caspase3, GPX4 and FDX1 in MCF-7 cells.
[0052] Example 7
[0053] The potential of copper ions in cancer therapy is currently being investigated, particularly their ability to target tumor microbes, thereby inhibiting cancer progression and improving the efficacy of breast cancer treatment. Here, we evaluated the antibacterial activity of Cu4 against Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli. Using the plating method, the results are as follows: Figure 8 As shown, we observed that at a concentration of 100 µg / ml, Cu4 reduced Escherichia coli colonies by more than 50%, while at a concentration of 200 µg / ml, it almost completely eliminated the survival of Escherichia coli and Staphylococcus aureus compared to the control group. These results indicate that Cu4 has a significant inhibitory effect on the growth of Escherichia coli and Staphylococcus aureus.
[0054] It should be noted that when numerical ranges are mentioned in the claims of this invention, it should be understood that the two endpoints of each numerical range and any value between the two endpoints can be selected. To avoid redundancy, the present invention describes preferred embodiments.
[0055] Although preferred embodiments of the invention have been described, those skilled in the art, upon learning the basic inventive concept, can make other changes and modifications to these embodiments. Therefore, the appended claims are intended to be interpreted as including both the preferred embodiments and all changes and modifications falling within the scope of the invention.
Claims
1. A thiadiazole tetranuclear copper complex with antitumor and antibacterial activities, characterized in that, The molecular formula of the complex is [Cu4(C3H3S2N2)2(C3H4S2N2)4], where C3H4S2N2 is 2-mercapto-5-methyl-1,3,4-thiadiazole; the structure of the complex is as follows: 。 2. The thiadiazole tetranuclear copper complex with antitumor and antibacterial activity according to claim 1, characterized in that, The complex has a monoclinic crystal system with space group C2 / c and cell parameters of: a = 45.33(3) Å, b = 7.869(5) Å, c = 19.582(14) Å, α = 90˚, β = 95.523(9)°, γ = 90˚, Z = 8, and unit cell volume V = 6953(8) Å. 3 .
3. The method for preparing a thiadiazole tetranuclear copper complex with antitumor and antibacterial activity as described in claim 1, characterized in that, Includes the following steps: Using CuX, thiadiazole, and triethylamine as raw materials, a solvothermal reaction is carried out. During the reaction, triethylamine causes some of the 2-mercapto-5-methyl-1,3,4-thiadiazole to deprotonate, yielding deprotonated 2-mercapto-5-methyl-1,3,4-thiadiazole, denoted as mmt⁻ ligand. The undeprotonated 2-mercapto-5-methyl-1,3,4-thiadiazole is denoted as Hmmt ligand. Each complex is composed of four Cu(I) ions, four mmt⁻ ligands, and two Hmmt ligands. The four mmt⁻ ligands bridge adjacent Cu(I) ions in a 3.21 coordination mode through the sulfur and nitrogen atoms of their thiols. At the same time, the two Hmmt ligands coordinate with the two Cu(I) ions in a monodentate coordination mode, thus obtaining the complex, wherein X is I or Br.
4. The method for preparing a thiadiazole tetranuclear copper complex with antitumor and antibacterial activity according to claim 3, characterized in that, The molar ratio of CuX, thiadiazole and triethylamine is 1:1.5 to 2:
2.
5. The method for preparing a thiadiazole tetranuclear copper complex with antitumor and antibacterial activity according to claim 3, characterized in that, The solvent in the solvothermal reaction is an equal volume mixture of CH3OH and H2O.
6. The method for preparing a thiadiazole tetranuclear copper complex with antitumor and antibacterial activity according to claim 3, characterized in that, The temperature of the solvothermal reaction is 100~110℃, and the time is 48 h~72 h.
7. The use of the thiadiazole tetranuclear copper complex with antitumor and antibacterial activity as described in claim 1 in the preparation of an anti-breast cancer drug.
8. The application of the thiadiazole tetranuclear copper complex with antitumor and antibacterial activity as described in claim 1 in the preparation of antibacterial agents, characterized in that, The antibacterial agent is used to inhibit Gram-positive bacteria Staphylococcus aureus and / or Gram-negative bacteria Escherichia coli.