Gold (I) complex containing alkyne ligand, preparation method of gold (I) complex and application of gold (I) complex in preparation of antitumor drugs

Abstract

The invention discloses a gold (I) complex containing an alkyne ligand, a preparation method of the gold (I) complex and application of the gold (I) complex in preparation of antitumor drugs, the gold (I) complex has a structure as shown in a general formula I, a phosphorus ligand is selected from diphenyl-2-pyridine phosphine or tricyclohexylphosphine, and Rx is an organic group selected from a specific alkyne ligand; according to the preparation method, chloroauric acid is taken as an initial raw material, sequentially reacts with dimethyl sulfide and an organic phosphine ligand, and finally is coupled with an alkyne ligand under an alkaline condition, so that the steps are simple and convenient, the conditions are mild, and the yield is high; in-vitro activity experiments show that the complex shows remarkable proliferation inhibition activity on ovarian cancer, malignant melanoma, non-small cell lung cancer and cisplatin-resistant cell strains thereof, the effect of the complex is superior to that of cisplatin and auronafine, and a candidate compound is provided for developing a novel targeted anti-tumor drug for overcoming the drug resistance of platinum drugs.

Description

Technical Field

[0001] This application relates to the field of metal complex drugs and antitumor drugs, and more specifically, to gold (I) complexes containing alkyne-based ligands, their preparation methods, and their application in the preparation of antitumor drugs. Background Technology

[0002] Cancer is one of the major public health challenges worldwide, causing nearly ten million deaths each year. Among the many types of cancer, non-small cell lung cancer, ovarian cancer, and malignant melanoma are particularly difficult to treat due to their highly aggressive nature and tendency to metastasize. Current cancer treatments mainly include surgery, radiotherapy, chemotherapy, targeted therapy, and immunotherapy. Chemotherapy remains crucial for advanced or metastatic tumors.

[0003] Cisplatin, as a first-line chemotherapy drug, is widely used in the treatment of non-small cell lung cancer, ovarian cancer, and malignant melanoma, possessing advantages such as a broad anti-cancer spectrum, effectiveness against hypoxic cells, and potent action. Its mechanism of action primarily involves hydrolysis into a cationic hydrated form after entering the body, similar to a bifunctional alkylating agent, which cross-links with DNA bases, causing DNA damage and thus inhibiting DNA replication and transcription. At high concentrations, it can also affect RNA and protein synthesis. However, the long-term use of cisplatin is severely limited by two factors: firstly, its significant systemic toxicity, such as nephrotoxicity, neurotoxicity, and ototoxicity, and poor targeting leading to significant damage to normal tissues; secondly, tumor cells easily develop resistance. Resistance mechanisms involve enhanced DNA damage repair capabilities and elevated levels of detoxification molecules such as glutathione within cells, leading to decreased drug sensitivity and worsened patient prognosis. Therefore, developing novel metalloid antitumor drugs with higher selectivity, lower toxicity, and the ability to overcome resistance is of great significance.

[0004] In recent years, gold complexes have attracted widespread attention due to their unique antitumor mechanisms. Inspired by research on the antirheumatic drug aurinofen, gold complexes mainly exert their anticancer effects by targeting thioredoxin reductase (TrxR). TrxR is overexpressed in various malignant tumors and is closely related to tumor proliferation, apoptosis resistance, and drug resistance. Unlike platinum drugs, which mainly act on DNA, gold complexes selectively kill tumor cells by inhibiting TrxR activity and disrupting the intracellular redox balance, providing a theoretical possibility for overcoming platinum resistance. Although many gold complexes have shown antitumor potential in vitro, their practical application still faces many challenges, including insufficient inhibitory activity against cisplatin-resistant tumor cells, complex synthetic routes, low yields, harsh reaction conditions, and poor stability of some complexes in physiological environments (such as easy breakage or oxidation of P–Au bonds), leading to increased off-target toxicity. Therefore, developing novel gold(I) complexes that are structurally stable, easy to synthesize, and can effectively target and kill cisplatin-resistant tumor cells remains a key research challenge. Summary of the Invention

[0005] To address the aforementioned problems, this invention provides a novel class of gold(I) complexes containing alkyne-based ligands, their preparation method, and their application in the preparation of antitumor drugs. In the gold(I) complex, the phosphorus ligand is selected from diphenyl-2-pyridinium phosphine or tricyclohexylphosphine, and Rx is an organic group derived from a specific alkyne ligand. This invention utilizes an optimized two-step synthetic method, starting with chloroauric acid, reacting sequentially with dimethyl sulfide and an organophosphine ligand, and finally coupling with the alkyne-based ligand under mild conditions, significantly improving the yield and product purity. In vitro antitumor experiments show that this class of complexes exhibits stronger inhibitory activity against various human tumor cell lines, particularly cisplatin-resistant ovarian cancer, malignant melanoma, and non-small cell lung cancer cell lines, compared to the clinical drugs cisplatin and auronoxine, providing valuable candidate compounds for developing novel targeted therapies to overcome platinum-based drug resistance.

[0006] In a first aspect, the present invention provides a class of gold(I) complexes containing alkyne-based ligands with antitumor activity, characterized in that the general structural formula of the gold(I) complex is shown in general formula I: Ⅰ: In the general formula I, Rx is a group containing an alkyne group.

[0007] In a second aspect, the present invention provides a method for preparing the gold (I) complex according to claim 1, characterized by comprising the following steps: S1: Chloroauric acid and dimethyl sulfide are mixed in a solvent, and the reaction yields the intermediate Au(Me2S)Cl; S2: The intermediate Au(Me2S)Cl obtained in S1 is mixed with an organophosphine ligand in a solvent and reacted under heating conditions to generate an intermediate compound; S3: React the intermediate compound obtained in S2 with an alkyne-based ligand in an alkaline solvent to generate the gold (I) complex.

[0008]

[0009] Preferably, the temperature conditions for the S1 reaction are 20~25℃ and the reaction time is 1~3h.

[0010] Preferably, the organophosphine ligand in S2 is selected from either diphenyl-2-pyridinium phosphine or tricyclohexylphosphine.

[0011] Preferably, the temperature conditions for the S2 reaction are 35~40℃ and the reaction time is 1~3h.

[0012] Preferably, the alkyne-based ligand in S3 is selected from one of acetylenol, 1-phenyl-2-propyn-1-ol, cyclopropynyl acetylene, or 4-acetylenyl anisole.

[0013] Preferably, the temperature conditions for the S3 reaction are 20~25℃ and the reaction time is 10~15h.

[0014] Thirdly, the present invention provides an application of a gold (I) complex containing an alkyne-based ligand, characterized in that it is used in the preparation of antitumor drugs.

[0015] In summary, the present invention has at least one of the following beneficial technical effects: 1. The series of gold (I) complexes provided by this invention, especially complex A1, exhibit extremely strong inhibitory activity against cisplatin-resistant ovarian cancer (A2780 / DDP), malignant melanoma (A375 / DDP), and lung adenocarcinoma (A549 / DDP) cell lines, with a half-maximal inhibitory concentration (IC50) of [value missing]. 50 The IC50 of A1 is significantly lower than that of cisplatin, a first-line clinical drug (e.g., for A2780 / DDP cells). 50 (The concentration of cisplatin was 0.397 μM, while that of cisplatin was 29.238 μM), providing a novel and highly effective candidate compound for overcoming the challenge of platinum-based drug resistance in clinical practice.

[0016] 2. This invention optimizes the synthetic route of gold(I) acetylenic complexes, using commercially available chloroauric acid as the starting material, and obtaining the target product through two key reactions. The entire process is mild and simple to operate, requiring no harsh anhydrous or oxygen-free conditions, and the product yield is generally as high as 85% or more. High-purity crystals can be obtained by recrystallization or gas-phase diffusion, demonstrating good potential for process scale-up.

[0017] 3. The series of gold (I) complexes provided by this invention have a unique mechanism of action, combining good stability and targeting potential, while also having a well-defined structure. Their mechanism of action differs from that of cisplatin, which damages DNA; instead, they primarily target thioredoxin reductase, which is highly expressed in tumor cells. This mechanism gives them better selectivity for tumor cells and theoretically reduces toxic side effects on normal cells. Attached Figure Description

[0018] To more clearly illustrate the technical solutions of the embodiments of this application, the drawings used in the description of the embodiments of this application will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0019] Figure 1 The hydrogen magnetic resonance spectrum of the gold(I) complex A1 provided in the embodiments of this application is shown; Figure 2 The infrared spectrum of gold(I) complex A1 provided in an embodiment of this application is shown; Figure 3 The hydrogen magnetic resonance spectrum of the gold(I) complex A2 provided in the embodiments of this application is shown; Figure 4 The infrared spectrum of gold(I) complex A2 provided in an embodiment of this application is shown; Figure 5 The hydrogen magnetic resonance spectrum of the gold(I) complex A3 provided in the embodiments of this application is shown; Figure 6 The infrared spectrum of gold(I) complex A3 provided in an embodiment of this application is shown; Figure 7 The hydrogen magnetic resonance spectrum of the gold(I) complex A4 provided in the embodiments of this application is shown; Figure 8 The infrared spectrum of gold(I) complex A4 provided in an embodiment of this application is shown; Figure 9 The single-crystal diffraction crystal structure diagram of the gold(I) complex A5 provided in the embodiments of this application is shown; Figure 10 The hydrogen magnetic resonance spectrum of the gold(I) complex A5 provided in the embodiments of this application is shown; Figure 11 The infrared spectrum of gold(I) complex A5 provided in an embodiment of this application is shown. Detailed Implementation

[0020] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. The following description of at least one exemplary embodiment is merely illustrative and is in no way intended to limit the present invention or its application or use. Based on the embodiments of the present invention, any product that is the same as or similar to the present invention, derived by any person under the guidance of the present invention or by combining the features of the present invention with other prior art, falls within the protection scope of the present invention. Furthermore, all other embodiments obtained by those skilled in the art without inventive effort are within the protection scope of the present invention.

[0021] Specific experimental steps or conditions are not specified in the embodiments; they can be performed according to the conventional experimental steps or conditions described in the prior art. Reagents and other instruments used, unless otherwise specified, are all commercially available conventional reagent products. Furthermore, the accompanying drawings are merely illustrative diagrams of the embodiments of the present invention and are not necessarily drawn to scale. The same reference numerals in the drawings denote the same or similar parts, and therefore, repeated descriptions of them will be omitted. Some block diagrams shown in the drawings are functional entities and do not necessarily correspond to physically or logically independent entities.

[0022] Techniques, methods, and devices known to those skilled in the art may not be discussed in detail, but where appropriate, such techniques, methods, and devices should be considered part of this specification.

[0023] In the description of this invention, it should be understood that the use of terms such as "first" and "second" to define components is merely for the purpose of distinguishing the corresponding components. Unless otherwise stated, the above terms have no special meaning and therefore should not be construed as limiting the scope of protection of this invention.

[0024] Furthermore, the technical features involved in the different embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0025] To enable those skilled in the art to better understand this application, the following examples will be used to provide a detailed description of the gold (I) complex containing alkyne-based ligands provided in this application, its preparation method, and its application in the preparation of antitumor drugs.

[0026] Example Example 1 This embodiment provides a method for preparing a specific compound A1 (acetylenylcyclohexanol·diphenyl-2-pyridinium phosphine alloy (I)), structural characterization data, and its antitumor activity.

[0027] S1: At 25°C, 2 g of chloroauric acid (HAuCl4) was dissolved in 10 mL of a solvent consisting of anhydrous ethanol and deionized water in a 4:1 volume ratio, yielding a clear yellow solution. Dimethyl sulfide (Me2S) was slowly added dropwise to this solution with continuous stirring until the solution color changed from yellow to colorless. The reaction was continued with stirring at room temperature for 2 hours, during which a white solid gradually formed. After the reaction was complete, the resulting suspension was filtered, the white solid was collected, and dried under vacuum to obtain the intermediate Au(Me2S)Cl.

[0028] S2: Dissolve 1 mmol of Au(Me2S)Cl obtained in step S1 in 20 mL of dichloromethane. Add 1.05 mmol of diphenyl-2-pyridinium phosphine ligand to the solution. Place the reaction system in a 37°C water bath and stir under gentle heating for 2 hours. After the reaction is complete, filter the reaction solution to remove insoluble impurities, then cool and allow the filtrate to stand, allowing crystals to precipitate, thus obtaining the purified diphenyl-2-pyridinium phosphine chloride gold intermediate.

[0029] S3: 0.5 mmol of diphenyl-2-pyridinium phosphine chloride was placed in a reaction flask and stirred for 20 minutes to form a suspension. 70 μL of ethynylcyclohexanol (≈0.6 mmol) was added and stirred for 10 minutes to mix thoroughly. Subsequently, a solution of 1 mmol potassium hydroxide dissolved in 0.5 mL of deionized water was added dropwise, and the reaction was continued at room temperature with stirring for 12 hours. After the reaction was completed, the mixture was filtered, and the solid precipitate was collected. The solid was washed with pre-cooled 75% methanol aqueous solution to remove the byproduct potassium chloride and excess ligand. After vacuum drying, the crude product was obtained. The crude product was dissolved in 4 mL of dichloromethane, filtered, and the filtrate was allowed to stand at 4 °C for recrystallization to obtain pure gold(I) complex A1, which was a crystalline solid with a yield of 91.62%.

[0030] The gold(I) complex A1 prepared in this embodiment is crystalline and has the molecular formula: C 25 H 25 AuNOP, with a molecular weight of 583.42, has the following molecular structure:

[0031] Nuclear magnetic resonance hydrogen spectrum such as Figure 1 As shown ( 1 ¹H NMR, 600 MHz, CDCl₃: δ 8.76 (d, J = 4.5 Hz, 1H), 7.97 (t, J = 7.7 Hz, 1H), 7.78–7.64 (m, 5H), 7.40 (tdt, J = 10.9, 7.2, 5.5 Hz, 7H), 1.95 (dd, J = 23.7, 17.4 Hz, 3H), 1.70–1.61 (m, 6H), 1.50 (dt, J = 8.5, 8.1 Hz, 1H), 1.31–1.17 (m, 1H). The chemical shifts and integrated areas of each signal peak are consistent with the target molecule structure.

[0032] Infrared spectroscopy (IR) such as Figure 2 As shown, the oxygen-hydrogen stretching vibration of the hydroxyl group is located at 3454.51 cm⁻¹. -1 At this point, the carbon-oxygen vibration is located at 1099.43 cm⁻¹. -1 The carbon-hydrogen stretching vibration of the benzene ring group is located at 3020.53 cm⁻¹. -1 At this point, the hydrocarbon bending vibration is located at 1481.33 cm. -1 The pyridine hydrocarbon stretching vibration is located at 3053.32 cm⁻¹. -1 At this location, the ring skeleton vibration is located at 1566.2 cm. -1 The antisymmetric stretching vibration of cyclohexyl hydrocarbons is located at 2929.87 cm⁻¹. -1 At this point, the symmetrical stretching vibration is located at 2850.79 cm. -1At this point, the hydrocarbon bending vibration is located at 1431.18 cm. -1 Location; the alkyne group is located at 2123.63 cm. -1 At the test compound, specific vibrations of hydroxyl groups, pyridine groups, benzene ring groups, cyclohexyl groups, and alkyne groups were observed, which were consistent with the overall skeleton of complex A1.

[0033] Example 2 This embodiment provides a method for preparing gold (I) complex A2, namely 1-phenyl-2-propyn-1-ol·diphenyl-2-pyridinium phosphine alloy (I), the specific steps of which are as follows: S1: At room temperature, 2 g of HAuCl4 was dissolved in 10 mL of a solvent consisting of anhydrous ethanol and deionized water in a 4:1 volume ratio to obtain a clear yellow solution. Dimethyl sulfide was slowly added dropwise to this solution with stirring until the solution became colorless. The reaction was continued with stirring at room temperature for 2 hours, resulting in the precipitation of a white solid. After the reaction was complete, the mixture was filtered, and the white solid was collected and dried under vacuum to obtain the intermediate Au(Me2S)Cl.

[0034] S2: Dissolve 1 mmol of Au(Me2S)Cl obtained from S1 in 20 mL of dichloromethane. Add 1.05 mmol of diphenyl-2-pyridinium phosphine ligand to the solution. Heat the reaction system in a 37°C water bath and stir for 2 hours. After the reaction is complete, filter to remove insoluble impurities. Cool the filtrate to crystallize and obtain pure diphenyl-2-pyridinium phosphine chloride gold intermediate.

[0035] S3: At room temperature, 0.5 mmol of the diphenyl-2-pyridinium phosphine chloride gold intermediate obtained in S2 was placed in a vial, and 5 mL of ethanol-water mixed solvent (volume ratio 4:1) was added, and the mixture was stirred for 20 minutes. Then, 67 μl of 1-phenyl-2-propyn-1-ol ligand was added, and the mixture was stirred for 10 minutes to ensure homogeneity. Next, an alkaline solution prepared by dissolving 1 mmol of potassium hydroxide in 0.5 mL of deionized water was added dropwise, and the reaction was continued at room temperature with stirring for 12 hours. After the reaction was complete, the mixture was filtered, and the solid precipitate was collected. The crude product, 1-phenyl-2-propyn-1-ol·diphenyl-2-pyridinium phosphine alloy (I), was obtained by vacuum drying and then vacuum dried. 8 mL of dichloromethane was added to dissolve the 1-phenyl-2-propyn-1-ol·diphenyl-2-pyridinium phosphine alloy (I), and the mixture was filtered to obtain a transparent liquid. The liquid was recrystallized and purified at 4 °C to finally obtain a single-crystal gold (I) complex A2.

[0036] The gold(I) complex A2 prepared in this embodiment has a yield of 89.15%. Its structure was determined by 1H NMR spectroscopy (NMR spectroscopy). 1 Confirmed by 1H NMR and infrared spectroscopy (IR), the molecular formula of A2 is: C 26 H 21AuNOP, with a molecular weight of 591.4, exhibits good antitumor activity, and its structural formula is shown below:

[0037] 1 The H NMR results are as follows Figure 3 As shown 1 ¹H NMR (600 MHz, CDCl₃) δ 8.76 (d, J = 4.6 Hz, 1H), 7.96 (t, J = 7.7 Hz, 1H), 7.78–7.62 (m, 7H), 7.52–7.40 (m, 6H), 7.39–7.31 (m, 3H), 7.27 (d, J = 7.4 Hz, 1H), 5.63 (d, J = 6.0 Hz, 1H), 2.20 (d, J = 6.0 Hz, 1H). The chemical shifts and quantities of hydrogen are consistent with those of the expected compound.

[0038] Infrared spectral characteristic peaks such as Figure 4 As shown, the oxygen-hydrogen stretching vibration of the hydroxyl group is located at 3415.93 cm⁻¹. -1 At this point, the carbon-oxygen vibration is located at 1099.43 cm⁻¹. -1 The carbon-hydrogen stretching vibration of the benzene ring group is located at 3020.53 cm⁻¹. -1 At this point, the hydrocarbon bending vibration is located at 1481.33 cm. -1 The pyridine hydrocarbon stretching vibration is located at 3064.89 cm⁻¹. -1 At this location, the ring skeleton vibration is located at 1568.13 cm. -1 Location; the alkyne group is located at 2135.2 cm. -1 The carbon-hydrogen stretching vibration at the 2nd carbon position in the ligand 1-phenyl-2-propyn-1-ol is 2841.15 cm⁻¹. -1 At the test compound, specific vibrations of hydroxyl groups, pyridine ring groups, benzene ring groups, and alkyne groups are present, which are consistent with the overall skeleton of complex A2.

[0039] Example 3 This embodiment provides a method for preparing gold (I) complex A3, namely acetylenecyclohexanol·tricyclohexylphosphine alloy (I), the specific steps of which are as follows: S1: At room temperature, 2 g of HAuCl4 was dissolved in 10 mL of a solvent consisting of anhydrous ethanol and deionized water in a 4:1 volume ratio to obtain a clear yellow solution. Dimethyl sulfide was slowly added dropwise to this solution with stirring until the solution became colorless. The reaction was continued with stirring at room temperature for 2 hours, resulting in the precipitation of a white solid. After the reaction was complete, the mixture was filtered, and the white solid was collected and dried under vacuum to obtain the intermediate Au(Me2S)Cl.

[0040] S2: Dissolve 1 mmol of Au(Me2S)Cl obtained from S1 in 20 mL of dichloromethane. Add 1.05 mmol of tricyclohexylphosphine ligand to the solution. Heat the reaction system in a 37°C water bath and stir for 2 hours. After the reaction is complete, filter to remove insoluble impurities. Cool the filtrate to crystallize and obtain pure tricyclohexylphosphine chlorogold intermediate.

[0041] S3: At room temperature, 0.5 mmol of the tricyclohexylphosphine chloride gold intermediate obtained in S2 was placed in a reaction flask, and 5 mL of ethanol-water mixed solvent (volume ratio 4:1) was added. The mixture was stirred for 20 minutes. Then, 70 μL (≈0.6 mmol) of acetylenol ligand was added, and the mixture was stirred for 10 minutes to ensure homogeneity. Then, an alkaline solution prepared by dissolving 1 mmol of potassium hydroxide in 0.5 mL of deionized water was added dropwise, and the reaction was continued to be stirred at room temperature for 12 hours. After the reaction was completed, the mixture was filtered, and the solid precipitate was collected. After washing with pre-cooled 75% methanol aqueous solution, the precipitate was dried under vacuum to obtain the crude product acetylenol-tricyclohexylphosphine alloy (I). The crude product was dissolved in 4 mL of dichloromethane, filtered to obtain a clear solution, and this solution was recrystallized and purified at 4 °C to finally obtain the gold (I) complex A3.

[0042] The gold(I) complex A3 prepared in this embodiment has the molecular formula C1. 26 H 44 AuOP, with a molecular weight of 600.58 and a yield of 85.91%, has the following structural formula:

[0043] like Figure 5 As shown, 1 1H NMR data (600 MHz, CDCl3, δ): 2.01–1.76 (m, 18H), 1.75–1.58 (m, 9H), 1.43 (dd, J = 16.1, 8.2 Hz, 7H), 1.33–1.16 (m, 10H).

[0044] Infrared spectral characteristic peaks such as Figure 6 As shown: the OH stretching vibration of the hydroxyl group is located at 3462.22 cm⁻¹. -1 At this point, the CO stretching vibration is located at 1093.64 cm⁻¹. -1 The CH asymmetric stretching vibration of the cyclohexyl group is located at 2931.80 cm⁻¹. -1 The symmetrical stretching vibration is located at 2848.86 cm. -1 At this point, the CH bending vibration is located at 1444.68 cm. -1 The characteristic absorption peak of the terminal alkyne group is located at 2115.91 cm⁻¹. -1 Place.

[0045] The above spectral data are in perfect agreement with the expected structure of the target compound acetylenecyclohexanol-tricyclohexylphosphine alloy (I), confirming the successful synthesis of complex A3.

[0046] Example 4 This embodiment provides a method for preparing gold (I) complex A4, namely cyclopropylacetylene·diphenyl-2-pyridinium phosphine alloy (I), the specific steps of which are as follows: S1: At room temperature, 2 g of HAuCl4 was dissolved in 10 mL of a solvent consisting of anhydrous ethanol and deionized water in a 4:1 volume ratio to obtain a clear yellow solution. Dimethyl sulfide was slowly added dropwise to this solution with stirring until the solution became colorless. The reaction was continued with stirring at room temperature for 2 hours, resulting in the precipitation of a white solid. After the reaction was complete, the mixture was filtered, and the white solid was collected and dried under vacuum to obtain the intermediate Au(Me2S)Cl.

[0047] S2: Dissolve 1 mmol of Au(Me2S)Cl obtained from S1 in 20 mL of dichloromethane. Add 1.05 mmol of diphenyl-2-pyridinium phosphine to the solution. Place the reaction system in a 37°C water bath and stir for 2 hours. After the reaction is complete, filter to remove insoluble matter, and cool the filtrate to crystallize, obtaining the purified diphenyl-2-pyridinium phosphine chlorogold intermediate.

[0048] S3: At room temperature, 0.5 mmol of the diphenyl-2-pyridinium phosphine chloride obtained in S2 was placed in a reaction flask and stirred for 20 minutes. Then, 50 μL (0.6 mmol) of cyclopropaneacetylene was added, and the mixture was stirred for 10 minutes to ensure thorough mixing. Next, an alkaline solution prepared by dissolving 1 mmol of potassium hydroxide in 0.5 mL of deionized water was added dropwise, and the reaction was continued at room temperature with stirring for 12 hours. After the reaction was complete, the mixture was filtered, and the solid precipitate was collected. The precipitate was washed with pre-cooled 75% methanol aqueous solution and dried under vacuum to obtain the crude product. The crude product was dissolved in 6 mL of dichloromethane, filtered to obtain a clear solution, and the filtrate was recrystallized at 4 °C to obtain pure gold(I) complex A4, with a yield of 85.06%.

[0049] The gold(I) complex A4 prepared in this embodiment has the molecular formula C1. 22 H 19 AuNP, with a molecular weight of 525.34. Its chemical structure is shown below:

[0050] like Figure 7 As shown, the proton nuclear magnetic resonance spectrum (NMR spectrum) 1¹H NMR, 600 MHz, CDCl₃): δ 8.74 (d, J = 4.7 Hz, 1H), 8.01 (t, J = 7.8 Hz, 1H), 7.77–7.64 (m, 5H), 7.42 (tdd, J = 7.2, 4.9, 3.8 Hz, 6H), 7.35–7.29 (m, 1H), 1.39–1.32 (m, 1H), 0.74–0.67 (m, 4H). The chemical shifts, splitting patterns, and integrated areas of the signal peaks in the spectrum are consistent with the target molecule structure.

[0051] like Figure 8 The infrared (IR) spectroscopy analysis shown reveals the following characteristic absorption peak: the CH stretching vibration of the pyridine ring is located at 3070.68 cm⁻¹. -1 The ring skeleton vibration is located at 1566.20 cm. -1 The CH stretching vibration of the benzene ring is located at 3003.17 cm⁻¹. -1 The in-plane bending vibration of CH is located at 1479.40 cm. -1 The characteristic peak of the C≡C stretching vibration of terminal alkynes is located at 2127.48 cm⁻¹. -1 The CH stretching vibration of the cyclopropyl group is located at 2929.87 cm⁻¹. -1 .

[0052] The above spectroscopic data collectively confirm that the synthesized compound is a structurally correct cyclopropyne-diphenyl-2-pyridinium phosphine alloy (I). The above spectroscopic data are in perfect agreement with the expected structure of the target compound cyclopropyne-diphenyl-2-pyridinium phosphine alloy (I), confirming the successful synthesis of complex A4.

[0053] Example 5 This embodiment provides a method for preparing gold (I) complex A5, namely 4-ethynyl anisole·diphenyl-2-pyridinium phosphine alloy (I), the specific steps of which are as follows: S1: At room temperature, 2 g of HAuCl4 was dissolved in 10 mL of a solvent consisting of anhydrous ethanol and deionized water in a 4:1 volume ratio to obtain a clear yellow solution. Dimethyl sulfide was slowly added dropwise to this solution with stirring until the solution became colorless. The reaction was continued with stirring at room temperature for 2 hours, resulting in the precipitation of a white solid. After the reaction was complete, the mixture was filtered, and the white solid was collected and dried under vacuum to obtain the intermediate Au(Me2S)Cl.

[0054] S2: Dissolve 1 mmol of Au(Me2S)Cl obtained in S1 in 20 mL of dichloromethane, and add 1.05 mmol of diphenyl-2-pyridinium phosphine. Place the reaction mixture in a 37°C water bath and stir for 2 hours. After the reaction is complete, filter to remove insoluble matter, and cool the filtrate to crystallize, obtaining the purified diphenyl-2-pyridinium phosphine chloride-gold intermediate.

[0055] S3: At room temperature, 0.5 mmol of diphenyl-2-pyridinium phosphine chloride obtained in S2 was placed in a reaction flask, and 5 mL of solvent A was added. The mixture was stirred for 20 minutes. Then, 78 μL of 4-ethynyl anisole (approximately 0.6 mmol) was added, and the mixture was stirred for 10 minutes to ensure thorough mixing. Then, an alkaline solution prepared by dissolving 1 mmol of potassium hydroxide in 0.5 mL of deionized water was added dropwise, and the reaction was continued at room temperature with stirring for 12 hours. After the reaction was complete, the mixture was filtered, and the solid precipitate was collected and dried under vacuum to obtain the crude product. The crude product was dissolved in 6 mL of dichloromethane, filtered to obtain a clear solution, and the filtrate was allowed to stand for 10 minutes. Then, the solution was incubated at 4 °C using the ether vapor-phase diffusion method to finally obtain a single crystal of gold(I) complex A5 suitable for single-crystal diffraction analysis, with a yield of 88.16%.

[0056] The gold (I) complex A5 prepared in this embodiment has the molecular formula C1. 26 H 21 AuNOP, with a molecular weight of 591.37, has crystallographic parameters shown in Table 1, partial bond lengths and bond angles shown in Table 2, and a crystal structure as shown in Table 2. Figure 9 As shown, the chemical structural formula is as follows:

[0057] Table 1. Crystal data of gold(I) complex A5

[0058] Table 2. Bond lengths (Å) and bond angles (Å) of gold(I) complex A5 o )

[0059] like Figure 10 As shown, 1 1H NMR data (600 MHz, CDCl3, δ): 8.77 (d, J = 4.7 Hz, 1H), 8.04 (t, J = 7.8 Hz, 1H), 7.74 (dddd, J = 21.0, 12.6, 6.1, 1.6 Hz, 5H), 7.50–7.41 (m, 8H), 7.37–7.33 (m, 1H), 6.80–6.76 (m, 2H), 3.78 (s, 3H).

[0060] like Figure 11 As shown, the hydrocarbon stretching vibration of pyridine is located at 3051.39 cm⁻¹. -1 At this location, the ring skeleton vibration is located at 1566.20 cm. -1 The carbon-hydrogen stretching vibration of the benzene ring group is located at 3008.95 cm⁻¹. -1At this point, the hydrocarbon bending vibration is located at 1481.33 cm. -1 Location; the alkyne group is located at 2110.12 cm. -1 The asymmetric stretching vibration of the C-H single bond in the methyl group is located at 2947.23 cm⁻¹. -1 At this point, the symmetrical stretching vibration is located at 2893.22 cm. -1 The carbon-oxygen single bond stretching vibration is located at 1170.79 cm⁻¹. -1 Place.

[0061] Example 6 In this embodiment, the in vitro proliferative inhibitory activity of the gold (I) complexes described in this invention (represented by A1-A5 prepared in Examples 1-5) against various tumor cells was detected by the MTT assay, and their half-maximal inhibitory concentrations (IC50) were calculated. 50 The specific method is as follows: Complete culture medium containing 10% fetal bovine serum and 1% penicillin-streptomycin was used. Six tumor cell lines, including human ovarian cancer cells (A2780), human melanoma cells (A375), human non-small cell lung cancer cells (A549), and their corresponding cisplatin-resistant strains (A2780 / DDP, A375 / DDP, A549 / DDP), were rapidly thawed in a 37°C water bath after being taken from liquid nitrogen.

[0062] Transfer the thawed cell suspension to a centrifuge tube, add 2 mL of complete culture medium, and centrifuge at 400×g for 3 minutes. Discard the supernatant, resuspend the cells in 2 mL of fresh complete culture medium, and then seed them into T75 cell culture flasks, adding culture medium to a total volume of 10 mL. Incubate the cells in a 37°C, 5% CO2 incubator for 24 hours, then replace with fresh culture medium.

[0063] When the cells reach 80%–90% confluence, passage them. Discard the old culture medium, wash the cells 1–2 times with PBS, add 2 mL of trypsin digestion solution containing EDTA, and digest at 37°C for about 3 minutes. When 70%–80% of the cells become rounded and detach, add 4 mL of complete culture medium to stop the digestion. Centrifuge the cell suspension at 400×g for 3 minutes, discard the supernatant, resuspend the cells in complete culture medium, and seed them into new T75 culture flasks at a ratio of 1:2 to 1:4. Passage the cells every 2–3 days, maintaining them in the logarithmic growth phase.

[0064] Six cell lines in the logarithmic growth phase were collected, the culture medium was discarded, and the cells were washed twice with PBS. After trypsin digestion and cell counting, the cell density of each cell type was adjusted to 5 × 10⁶ cells / year using complete culture medium. 4Cells / mL. Add 100 μL of cell suspension (approximately 5000 cells / well) to each well of a 96-well plate, and reserve at least 3 wells as blank controls (containing only complete culture medium, without cells). Incubate the 96-well plate in a 37°C, 5% CO2, 95% humidity incubator for 24 hours to allow the cells to adhere fully.

[0065] The stock solutions of the gold(I) complexes (A1-A5), the positive control aurinofen, and cisplatin were removed from -20°C and allowed to warm to room temperature. The stock solutions of each gold compound were serially diluted using complete culture medium, with eight concentration gradients established; eight concentration gradients were also established for cisplatin. The final concentration of dimethyl sulfoxide (DMSO) in all drug diluents did not exceed 0.1% to eliminate the interference of solvent toxicity on the experimental results.

[0066] The experimental setup included: the test drug group, the positive control group (Aurinophene, cisplatin), the negative control group (complete culture medium containing 0.1% DMSO), and the blank control group (complete culture medium only).

[0067] After cells have adhered for 24 hours, carefully aspirate the original culture medium from each well of the 96-well plate. Add 100 μL of the corresponding concentration of the test drug working solution, positive control working solution, negative control solution, or blank control solution to each well sequentially according to the pre-defined groupings. Set up three replicates for each concentration. Return the 96-well plate to the incubator and continue culturing for 72 hours.

[0068] After 72 hours of drug treatment, add 10 μL of MTT solution (5 mg / mL, sterilized through a 0.2 μm filter membrane) to each well. Wrap the 96-well plate with aluminum foil to protect it from light and incubate at 37°C in a 5% CO2 incubator for another 4 hours. After incubation, carefully discard the liquid in each well. Add 150 μL of DMSO to each well and place the 96-well plate on a shaker at 37°C, protected from light, and shake slowly for 10 minutes to fully dissolve the purple formazan crystals. Use a microplate reader to measure the absorbance (OD value) of each well at a wavelength of 490 nm. Calculate the cell inhibition rate at each drug concentration using the following formula: Cell inhibition rate (%) = [1 - (OD-treated group - OD blank group) / (OD control group - OD blank group)] × 100% The above experiments were independently repeated three times. Using the logarithm of drug concentration as the x-axis and cell inhibition rate as the y-axis, dose-response curves were fitted using GraphPad Prism software, and the IC50 of each gold(I) complex against six types of tumor cells was finally calculated. 50 The values ​​are shown in Table 3.

[0069] Table 3 IC50 of various gold (I) complexes against six types of tumor cells 50 Value (μM)

[0070] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on the differences from other embodiments. The same or similar parts between the various embodiments can be referred to each other.

[0071] Although preferred embodiments of the present application 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 the preferred embodiments as well as all changes and modifications falling within the scope of the embodiments of the present application.

[0072] Finally, it should be noted that in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or terminal device that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or terminal device. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or terminal device that includes said element.

[0073] The above provides a detailed description of the gold (I) complex containing alkyne-based ligands, its preparation method, and its application in the preparation of antitumor drugs. Specific examples have been used to illustrate the principles and implementation methods of this application. The above description of the embodiments is only for the purpose of helping to understand the method and core ideas of this application. At the same time, for those skilled in the art, there will be changes in the specific implementation methods and application scope based on the ideas of this application. Therefore, the content of this specification should not be construed as a limitation of this application.

Claims

1. A class of gold (I) complexes containing alkyne-based ligands with antitumor activity, characterized in that, The general structural formula of the gold(I) complex is shown in Formula I: Ⅰ: In the general formula I, Rx is a group containing an alkyne group.

2. A method for preparing the gold (I) complex according to claim 1, characterized in that, Includes the following steps: S1: Chloroauric acid and dimethyl sulfide are mixed in a solvent, and the reaction yields the intermediate Au(Me2S)Cl; S2: The intermediate Au(Me2S)Cl obtained in S1 is mixed with an organophosphine ligand in a solvent and reacted under heating conditions to generate an intermediate compound; S3: React the intermediate compound obtained in S2 with an alkyne-based ligand in an alkaline solvent to generate the gold (I) complex.

3. A method for preparing the gold (I) complex of claim 1 according to claim 2, characterized in that, The temperature conditions for the S1 reaction are 20~25℃, and the reaction time is 1~3h.

4. A method for preparing the gold (I) complex of claim 1 according to claim 2, characterized in that, The organophosphine ligand in S2 is selected from either diphenyl-2-pyridinium phosphine or tricyclohexylphosphine.

5. A method for preparing the gold (I) complex of claim 1 according to claim 2, characterized in that, The temperature conditions for the S2 reaction are 35~40℃, and the reaction time is 1~3h.

6. A method for preparing the gold (I) complex of claim 1 according to claim 2, characterized in that, The alkyne-based ligand in S3 is selected from one of acetylenol, 1-phenyl-2-propyn-1-ol, cyclopropynyl acetylene, or 4-acetylenyl anisole.

7. A method for preparing the gold (I) complex of claim 1 according to claim 2, characterized in that, The temperature conditions for the S3 reaction are 20~25℃, and the reaction time is 10~15h.

8. The application of a gold(I) complex containing an alkyne-based ligand, characterized in that, Used in the preparation of anti-tumor drugs.

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