A palladium complex containing quinolinyl triazolophthalazine ligand, and a preparation method and application thereof

Abstract

The present application belongs to the technical field of metal complexes, and particularly relates to a palladium complex containing a quinoline-based triazolophthalazine ligand, a preparation method and application thereof. The complex structure formula is [PdLCl2], wherein L is 3-(quinoline-2-yl)-[1,2,4]triazole[3,4-a]phthalazine. The preparation method is as follows: PdCl2 is mixed with NaCl and dissolved in a methanol solvent to obtain a Na2PdCl4 solution; 3-(quinoline-2-yl)-[1,2,4]triazole[3,4-a]phthalazine is dissolved in a methanol solvent, the Na2PdCl4 solution is added dropwise, and the complex is obtained through a reflux reaction. The complex can effectively inhibit the proliferation of breast cancer cells, cervical cancer cells and ovarian cancer cells, and can also efficiently induce the apoptosis of the four kinds of tumor cells. Therefore, the complex is expected to become a high-efficiency broad-spectrum potential anti-tumor drug for common cancers in women.

Description

Technical Field

[0001] The present invention belongs to the technical field of metal complexes, and in particular relates to a palladium complex containing a quinolyl triazolophthalazine ligand, a preparation method and an application thereof. Background Art

[0002] Cancer, a type of malignant tumor, is one of the leading causes of death worldwide. Cancer treatment consumes significant medical resources, and the morbidity and mortality rates associated with cancer represent a significant societal burden. Consequently, cancer has become a serious global economic and social problem. Current clinical treatments for tumors include surgical resection, chemotherapy, radiotherapy, targeted therapy, and immunotherapy. In chemotherapy, chemotherapeutic drugs are used to kill tumor cells, and chemotherapy agents can also be combined with other drugs to control tumor growth and spread.

[0003] Platinum drugs are one of the most widely used anticancer drugs in the world, but they have disadvantages such as toxic side effects and drug resistance. Therefore, it is necessary to design new anticancer drugs. 8 Due to their similar electronic configurations and nearly identical dimensions, palladium(II) complexes exhibit similar chemical properties and form square planar metal complexes. Despite this similarity, several studies have shown that palladium(II) complexes exhibit distinct biological effects from platinum(II) complexes. First, some palladium complexes exhibit activity against both cisplatin-sensitive and -resistant cell lines. Second, some palladium complexes have mechanisms of action distinct from those of platinum complexes. Third, some palladium complexes exhibit no mutagenic potential in the Ames test, whereas their platinum analogs increase spontaneous mutation rates. These findings have spurred the development of new bioactive palladium complexes. Recently, palladium complexes have been reported to exhibit significant anticancer activity and lower toxicity compared to some clinically used chemotherapeutic drugs. Quinoline and its derivatives possess diverse biological activities, such as antibacterial, antiviral, anti-inflammatory, and antimalarial activities, and have been reported to exhibit significant anticancer activity. Therefore, the construction of quinoline-based palladium complexes is crucial to addressing the current limitations of platinum-based drugs in anticancer applications. Summary of the Invention

[0004] The purpose of the present invention is to provide a palladium complex containing a quinolyl triazolophthalazine ligand, a preparation method and application thereof, so as to construct a quinoline palladium complex to solve the problem of limited application of current platinum drugs in anticancer treatment.

[0005] To achieve the above object, the technical solution of the present invention is as follows:

[0006] A palladium complex containing a quinolinyl triazolophthalazine ligand has a simplified structural formula of [PdLCl2], wherein L is 3-(quinolin-2-yl)-[1,2,4]triazolo[3,4-a]phthalazine; the structural formula is:

[0007]

[0008] The palladium complex has a planar tetragonal coordination structure. The complex molecule has relatively strong aromaticity and good planarity as a whole, but the coordination structure is slightly distorted.

[0009] A method for preparing the palladium complex containing a quinolyltriazolophthalazine ligand as described above comprises the following steps:

[0010] Step 1: PdCl2 and NaCl are mixed and dissolved in methanol solvent to obtain Na2PdCl4 solution;

[0011] Step 2: dissolving 3-(quinolin-2-yl)-[1,2,4]triazolo[3,4-a]phthalazine in methanol solvent, adding Na2PdCl4 solution dropwise, and performing reflux reaction to obtain the palladium complex.

[0012] Furthermore, in step 1, the molar ratio of PdCl2 to NaCl is 1:2.

[0013] Furthermore, in step 2, the molar ratio of 3-(quinolin-2-yl)-[1,2,4]triazolo[3,4-a]phthalazine to Na2PdCl4 is 1:1.1.

[0014] Furthermore, the temperature of the methanol solvent in step 1 is 40-60°C.

[0015] Furthermore, the temperature of the methanol solvent in step 2 is 65-80°C.

[0016] Furthermore, the temperature of the reflux reaction in step 2 is 65-80° C., and the time is 2-4 hours.

[0017] The invention relates to the use of the aforementioned palladium complex containing a quinolyl triazolophthalazine ligand as a potential anti-breast cancer drug.

[0018] The invention relates to an application of the aforementioned palladium complex containing a quinolyl triazolophthalazine ligand as a potential anti-cervical cancer drug.

[0019] The invention relates to a use of the aforementioned palladium complex containing a quinolyl triazolophthalazine ligand as a potential anti-ovarian cancer drug.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] The palladium complex containing a quinolyltriazolophthalazine ligand of the present invention is obtained under conventional reflux reaction conditions, has a simple preparation process, high yield and purity, and exhibits excellent biological properties. The complex is highly effective in inhibiting the proliferation of breast, cervical, and ovarian cancer cells and inducing apoptosis in these cancers. It is a novel palladium complex containing a quinolyltriazolophthalazine ligand, primarily for the broad-spectrum treatment of common cancers in women. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 The infrared spectrum of the palladium complex containing quinolyl triazolophthalazine ligand and its ligand of the present invention.

[0023] Figure 2 The ultraviolet spectrum of the palladium complex containing quinolyl triazolophthalazine ligand and its ligand of the present invention.

[0024] Figure 3 The present invention contains a palladium complex (a) containing a quinolyl triazolophthalazine ligand and a ligand (b) thereof 1 H NMR spectrum.

[0025] Figure 4 Electrospray mass spectrometry of the palladium complex containing quinolyl triazolophthalazine ligands of the present invention.

[0026] Figure 5 The theoretically calculated optimized structure diagram of the palladium complex containing quinolyl triazolophthalazine ligands of the present invention.

[0027] Figure 6 CCK-8 graph showing that the palladium complex containing quinolyl triazolophthalazine ligand of the present invention inhibits the proliferation of MCF-7 cells.

[0028] Figure 7 CCK-8 graph showing that the palladium complex containing quinolyl triazolophthalazine ligand of the present invention inhibits the proliferation of MDA-MB-231 cells.

[0029] Figure 8 Figure 8 of the CCK-8 assay showing that the palladium complex containing the quinolyl triazolophthalazine ligand of the present invention inhibits the proliferation of Hela cells.

[0030] Figure 9 Figure 8 of the CCK-8 assay showing that the palladium complex containing the quinolyl triazolophthalazine ligand of the present invention inhibits the proliferation of ES-2 cells.

[0031] Figure 10 The data diagram of apoptosis of MCF-7 cells after the palladium complex containing quinolyl triazolophthalazine ligand and its ligand act on the cells.

[0032] Figure 11The data diagram of apoptosis of MDA-MB-231 cells after the palladium complex containing quinolyl triazolophthalazine ligand and its ligand act on the cells.

[0033] Figure 12 The data diagram of the apoptosis of Hela cells after the palladium complex containing quinolyl triazolophthalazine ligand and its ligand act on it.

[0034] Figure 13 The data diagram of apoptosis of ES-2 cells after the palladium complex containing quinolyl triazolophthalazine ligand and its ligand act on the cells. DETAILED DESCRIPTION

[0035] To facilitate understanding of the present invention, the present invention will be described more fully below. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the understanding of the disclosure of the present invention more thorough and comprehensive.

[0036] Unless otherwise defined, all technical terms used hereinafter have the same meanings as those generally understood by those skilled in the art. The technical terms used herein are only for the purpose of describing specific embodiments and are not intended to limit the scope of protection of the present invention.

[0037] Unless otherwise specified, various raw materials, reagents, instruments and equipment used in the present invention can be purchased from the market or prepared by existing methods.

[0038] Example 1 Preparation method of the complex of the present invention

[0039] NaCl (0.44 mmol, 0.0257 g) and PdCl2 (0.22 mmol, 0.0390 g) were heated to 40°C and dissolved in 10 mL of methanol with stirring to obtain a Na2PdCl4 solution. 3-(Quinolin-2-yl)-[1,2,4]triazolo[3,4-a]phthalazine ligand (0.2 mmol, 0.0598 g) was placed in a 100 mL round-bottom flask. 20 mL of methanol was added and heated to 70°C with stirring to dissolve. The Na2PdCl4 solution was then added dropwise and the mixture was heated under reflux at 70°C with stirring for 3 hours. After the reaction was complete, the mixture was cooled to room temperature, filtered, and washed with methanol to obtain a dark purple solid powder with a yield of 82.5%.

[0040] Example 2 Infrared spectroscopy analysis of the complex of the present invention

[0041] The complex of the present invention and dry potassium bromide solid were mixed in a ratio of 1:100, ground and pressed into tablets, and the density of the tablets at 4000-400 cm -1 Infrared spectrum within the range. The infrared spectrum of the complex and its ligand is shown in Figure 1, Table 1 lists the characteristic stretching vibration absorption peaks of some functional groups.

[0042] Compared with its ligand, the characteristic absorption peaks of the complex of the present invention are shifted to varying degrees. The stretching vibration absorption peak of the NN single bond on the triazole ring in the complex shifts from 1096 cm -1 Redshift to 1116 cm -1 The stretching vibration absorption peak of the C=N double bond on the quinoline ring is 1475 cm -1 Redshift to 1487 cm -1 The stretching vibration absorption peak of the C=N double bond on the triazole ring is 1620 cm -1 Blue shift to 1570 cm -1 , indicating that the triazole ring N and quinoline ring N participate in the coordination of palladium, and there is a stretching vibration absorption peak of Pd-N bond at 750cm -1 , thus proving the synthesis of the complex.

[0043] Table 1 Main infrared spectral data of the complex and its ligand (unit: cm -1 )

[0044]

[0045] Example 3 Ultraviolet spectrum analysis of the complex of the present invention

[0046] DMSO was used to prepare the -2 M ligand and complex stock solution, dilute the solution to 10 -5 M, and then measure the UV spectra of these compounds, such as Figure 2 shown.

[0047] The ligand has an absorption peak near 400nm, which is attributed to the n-π* transition between the lone pair electrons on the heteroatom in the ligand and the planar ring. After the formation of the complex, the absorption peak weakens and shifts. The complex shows a strong absorption peak at 585nm, which may be due to the charge transfer transition between the metal ion and the ligand, indicating that the central metal palladium is coordinated with the heteroatom in the ligand.

[0048] Example 4 Complex of the present invention 1 H NMR spectrum analysis

[0049] DMSO-d 6 As solvent, the ligand and complex were determined 1 H NMR spectrum, see Figure 3The corresponding hydrogen atoms have been assigned in the figure. Comparing the NMR spectra of the complex with those of the corresponding ligands shows that the absorption peaks corresponding to the hydrogen atoms in the ligand and the complex have shifted to varying degrees. This further proves that the palladium metal coordinates with the lone pairs of electrons in the ligand. The addition of palladium metal changes the chemical environment of the hydrogen atoms at various positions, causing the corresponding absorption peaks to shift.

[0050] Example 5 Electrospray mass spectrometry analysis of the complex of the present invention

[0051] Weigh a certain amount of complex and dissolve it in chromatographic grade acetonitrile solvent to prepare a concentration of 1×10 -7 ~1×10 - 5 g / mL solution, and then filtered into a liquid chromatography vial with a 0.22μM filter membrane and tested on the machine. The electrospray ionization mass spectrum of the complex (positive ion) is shown in Figure 4 ,Table 2 attributes the molecular ion peaks in the mass spectra.

[0052] The molecular ion peak with mass-to-charge ratio of 437.24 was assigned to the structure of the complex with one unit positive charge after losing one chloride ion. The main structure of the complex was confirmed by electrospray mass spectrometry.

[0053] Table 2 Electrospray mass spectrometry data of the complexes (positive ions)

[0054]

[0055] Example 6 Theoretical calculation analysis of the complex of the present invention

[0056] The Gaussian 09 program package was used to optimize the structure of the complex at the density functional theory (DFT) B3LYP / 6-31G(d) level. Frequency vibration analysis and wave function stability analysis were performed on the optimized complex structure at the same level. The initial structure of the complex was constructed according to the results inferred from experimental characterization.

[0057] According to the possible coordination mode of the ligand, we constructed two isomeric structures with different coordination modes. The first structure is a planar tetracoordinate configuration formed by the coordination of the central palladium with the N of the quinoline ring, the N on the triazole ring, and two chloride ions. The second structure is a tetrahedral configuration formed by the coordination of the central palladium with the N of the quinoline ring, the N on the triazole ring, and two chloride ions. The optimization results show that the tetrahedral configuration in the palladium complex of the second structure is finally optimized to the same planar quadrilateral configuration as the first structural model. In addition, we also tried to build a third structure, which is a tetracoordinate structure formed by the central palladium with the N of the quinoline ring, the N on the naphthalene ring after the C-C single bond in the triazole-naphthyridine ligand rotates, and two chloride ions. However, this structure was not successfully built because of the steric hindrance of the hydrogen atoms on the quinoline ring close to the N of quinoline, and palladium is not easy to coordinate with the N on the naphthalene ring. In summary, Figure 5 The structural model shown is the only stable structure of the complex, in which the angle between the chlorine adjacent to the quinoline ring and the plane formed by the other three coordinating atoms is 3.24°, so the coordination structure is a slightly distorted planar quadrilateral configuration.

[0058] Example 7 Determination of IC50 values ​​of the complexes of the present invention for inhibiting proliferation of breast cancer (MCF-7 and MDA-MB-231) cells, cervical cancer (Hela) cells and ovarian cancer (ES-2) cells

[0059] IC 50 IC refers to the concentration of the inhibitor when the inhibitor inhibits cell proliferation to half of its original number. It is one of the criteria for evaluating the inhibitory ability of the inhibitor. 50 The smaller the value, the stronger the inhibitory ability.

[0060] The principle of CCK-8 method to determine the inhibition of cell proliferation by complexes:

[0061] The CCK-8 reagent contains WST-8, which, in the presence of an electron coupling reagent, is reduced by mitochondrial dehydrogenases to a highly water-soluble orange-yellow formazan product. Within a certain cell population range, the amount of formazan produced is proportional to the number of viable cells; greater cytotoxicity indicates a lighter color. The absorbance at 450nm is measured using a microplate reader and compared to a blank control to further determine the number of viable cells.

[0062] CCK-8 experimental steps: Add 150 μL of sterile 0.85% NaCl solution to each well in the outer circle of the 96-well plate, and add 130 μL of culture medium to each well in the middle. Note: Sodium chloride surrounds the culture medium. When the cells grow to about 90%, digest and centrifuge to make a cell suspension, and count with a hemocytometer. Depending on the culture time, inoculate 50 μL of diluted cell suspension into 130 μL of culture medium in a 96-well plate. After the cells adhere to the wall, add the drug at a final concentration of 0, 1, 5, 10, 20, 30, and 50 μM. After the drug has acted for 24 or 48 hours, aspirate the supernatant, add 100 μL of culture medium containing 10% CCK-8 to each well, culture in an incubator for about 20 to 30 minutes, and measure the absorbance at 450 nm with an enzyme reader. Use origin to process the data, calculate the cell survival rate at each concentration, and use Graphpad fitting to obtain the IC of its inhibitory effect. 50 The inhibitory ability of the complex on cell proliferation was evaluated.

[0063] like Figure 6-9 The experimental results showed that palladium (II) complexes containing quinolyl triazolophthalazine ligands can effectively inhibit the proliferation of MCF-7, MDA-MB-231, Hela and ES-2 cells. When the complexes acted on MCF-7, MDA-MB-231, Hela and ES-2 cells for 48 hours, the IC values ​​for inhibiting tumor cell proliferation were 50 The values ​​were 10.22, 7.91, 10.58 and 5.80 μM, respectively. The inhibitory ability was stronger than or comparable to the inhibitory ability of cisplatin on these four cell lines reported in the literature, showing efficient anti-breast cancer, cervical cancer and ovarian cancer activity.

[0064] Example 8 Detection of the ability of the complex of the present invention to induce apoptosis in MCF-7, MDA-MB-231, Hela and ES-2 cells

[0065] Annexin V-FITC / PI double staining assay to determine the ability of complexes to induce cell apoptosis:

[0066] Phosphatidylserine (PS) in normal cells is distributed on the inner side of the cell membrane, while cells undergoing early apoptosis expose PS on the outer side of the cell membrane. Therefore, PS externalization is a key characteristic of apoptosis. Annexin V is a phospholipid-binding protein that has a high affinity for PS. Annexin V labeled with fluorescein (FITC), i.e., Annexin V-FITC, can be used to detect PS externalization by flow cytometry. Propidium iodide (PI) is a nuclear staining reagent that stains DNA and produces red fluorescence after intercalation into DNA. PI cannot cross intact cell membranes, but it can pass through necrotic cells or cells that have lost their cell membrane integrity in the late stages of apoptosis. Therefore, the combination of Annexin V and PI can be used to distinguish between live cells, necrotic cells, and premature and late apoptotic cells.

[0067] Annexin V-FITC / PI double staining experimental steps: Seed the cells in a 6-well plate and culture them in a cell culture incubator. When the cells grow to 60% to 70%, replace the culture medium and add palladium complexes and ligands at final concentrations of 0, 1, 10, 20, 30, and 50 μM respectively. After 48 hours of drug action, the suspended cells are directly collected, and the adherent cells are digested with EDTA-free trypsin, followed by centrifugation at 2000r for 5 minutes. Discard the supernatant and wash twice with pre-cooled PBS by gentle pipetting. Add an appropriate volume of 1×Binding buffer working solution to the cell pellet to make the cell concentration reach 1×10 6 Resuspend the cells and transfer 100μL of the cell suspension to a 1.5mL EP tube. Add 5μL of Annexin V-FITC and 5-10μL of PI, protected from light. Mix gently and incubate at room temperature for 15 minutes in the dark. After staining and incubation, add 400μL of 1× Binding Buffer to each tube and mix thoroughly. Flow cytometry should ideally be performed within one hour. Three control groups should be set up: normal cells, Annexin V-FITC staining alone, and PI staining alone.

[0068] like Figure 10-13 The experimental results showed that palladium (II) complexes containing quinolyl triazolophthalazine ligands can effectively induce apoptosis of MCF-7, MDA-MB-231, Hela and ES-2 cells. When 20 μM of the complex acted on MCF-7, MDA-MB-231, Hela and ES-2 cells for 48 hours, the total apoptosis rates were 34.10%, 53.46%, 55.24% and 48.03%, respectively, demonstrating the ability to effectively induce apoptosis of breast cancer, cervical cancer and ovarian cancer cells.

[0069] The above description is only for better explanation of the embodiments of the present invention and is not intended to limit the present invention. Any modifications or equivalent substitutions that do not depart from the spirit and scope of the present invention are intended to be within the scope of the present invention.

Claims

1. A palladium complex containing a quinolinyltriazolophthalazine ligand, characterized in that: The structural formula is: 。 2. A method for preparing a palladium complex containing a quinolyltriazolophthalazine ligand according to claim 1, characterized in that: The steps include: Step 1: PdCl2 and NaCl are mixed and dissolved in methanol solvent to obtain Na2PdCl4 solution; Step 2: dissolving 3-(quinolin-2-yl)-[1,2,4]triazolo[3,4-a]phthalazine in methanol solvent, adding Na2PdCl4 solution dropwise, and performing reflux reaction to obtain the palladium complex.

3. The method for preparing a palladium complex containing a quinolyltriazolophthalazine ligand according to claim 2, wherein: The molar ratio of PdCl2 to NaCl in step 1 is 1:

2.

4. The method for preparing a palladium complex containing a quinolyltriazolophthalazine ligand according to claim 2, wherein: In step 2, the molar ratio of 3-(quinolin-2-yl)-[1,2,4]triazolo[3,4-a]phthalazine to Na2PdCl4 is 1:1.

1.

5. The method for preparing a palladium complex containing a quinolyl triazolophthalazine ligand according to claim 2, wherein: The temperature of the methanol solvent in step 1 is 40-60°C.

6. The method for preparing a palladium complex containing a quinolyltriazolophthalazine ligand according to claim 2, wherein: The temperature of the methanol solvent in step 2 is 65-80°C.

7. The method for preparing a palladium complex containing a quinolinyl triazolophthalazine ligand according to claim 2, wherein: The reflux reaction temperature in step 2 is 65-80° C. and the reaction time is 2-4 h.

8. Use of the palladium complex containing a quinolyltriazolophthalazine ligand according to claim 1 in the preparation of a potential anti-breast cancer drug.

9. Use of the palladium complex containing a quinolyltriazolophthalazine ligand according to claim 1 in the preparation of a potential anti-cervical cancer drug.

10. Use of the palladium complex containing a quinolyltriazolophthalazine ligand according to claim 1 in the preparation of a potential anti-ovarian cancer drug.

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