6-phenyl-2, 2 '-bipyridine platinum (II)-guanine N-heterocyclic carbene complex as well as preparation method and application thereof

Abstract

The invention discloses a preparation method of 6-phenyl-2, 3, 4, 5- The invention discloses a 2, 2 '-bipyridine platinum (II)-guanine N-heterocyclic carbene complex and a preparation method and application thereof, and particularly relates to the complex shown in the specification, the complex has dual inhibitory activity on tumor cells and tumor stem cells at the same time, the problem of platinum drug resistance caused by the tumor stem cells, glutathione and other proteins can be solved, and the drug resistance of the tumor cells and the glutathione can be improved. The compound can be used for preparing drugs for treating various malignant tumors, especially drugs for treating malignant tumors resistant to existing chemotherapeutic drugs. The preparation method is simple and easy to operate. In addition, the 6-phenyl-2, 2 '-bipyridine ligand disclosed by the invention contains carbon anions with relatively strong electron donating ability, and compared with a terpyridine ligand, the 6-phenyl-2, 2'-bipyridine ligand disclosed by the invention can further stabilize a metal ion center, so that the ability of resisting GSH interference is improved, and better stability is shown.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical chemistry, and in particular to a novel 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex, and a preparation method and application thereof. Background Art

[0002] Platinum drugs such as cisplatin are commonly used chemotherapy drugs for the clinical treatment of cancer. The main mechanism by which platinum drugs exert their anti-tumor effects is the formation of platinum-DNA adducts with DNA in the cell nucleus. However, platinum drugs have poor stability and are easily chemically reacted with biological molecules such as glutathione (GSH) containing thiol groups, thereby becoming inactivated and developing drug resistance. In addition, the existence of tumor stem cells is another important factor that causes existing platinum drugs to easily develop drug resistance during the treatment of cancer. Tumor stem cells are a type of cancer cells with stem cell characteristics that have the ability to self-renew and differentiate. The content of tumor stem cells in tumor cells is relatively high, which can lead to drug resistance problems by maintaining a dormant state, increasing DNA repair ability, and blocking apoptosis pathways.

[0003] Guanine is a natural nucleic acid base that participates in the synthesis of nucleic acids, energy metabolism and many other biochemical reactions. In recent years, researchers have developed a series of purine drugs that can effectively treat cancer based on the purine skeleton. At present, it has been reported that the complex can dually inhibit the proliferation activity of tumor cells and tumor stem cells, while the toxicity to normal cells is weak, but there are few reports on platinum compounds that can resist GSH interference. For example, the prior art Chinese patent 2022117262148 discloses a class of terpyridine-platinum (II) complexes containing guanine NHC derivative ligands with excellent inhibitory activity on malignant tumor cells and tumor stem cells. This type of complex improves the problem of poor solubility of guanine, and shows strong inhibitory activity on tumor cells, especially triple-negative breast cancer cells and their stem cells, which are currently difficult to treat, without obvious toxicity to normal cells. However, in order to further improve stability and avoid drug resistance caused by reactions with thiol-containing biomolecules such as GSH, it is necessary to further develop other types of complexes. Summary of the invention

[0004] Purpose of the invention: In view of the poor stability of existing complexes in tumor treatment and the drug resistance of existing platinum drugs caused by tumor stem cells, the present invention aims to provide a new type of 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex with excellent inhibitory activity on the proliferation of various malignant tumor cells and tumor stem cells and stable chemical properties. This type of complex shows strong inhibitory activity on the proliferation of various tumor cells and triple-negative breast cancer stem cells, and has no obvious toxicity to normal cells. In addition, the 6-phenyl-2,2'-bipyridine ligand in this type of complex contains a carbon anion with strong electron donating ability, which can further stabilize the metal ion center compared with the terpyridine ligand, thereby improving the ability to resist GSH interference and showing better stability.

[0005] The invention also provides a preparation method and application of a 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex.

[0006] Technical solution: In order to achieve the above purpose, the 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex described in the general formula (I) of the present invention or its pharmaceutically acceptable salt:

[0007]

[0008] in:

[0009] R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 are each independently selected from hydrogen, halogen, cyano, nitro, trifluoromethyl, hydroxyl, amino, carboxyl, C1-C10 straight-chain alkyl, C1-C10 straight-chain heteroalkyl, C3-C12 branched or cyclic alkyl, C3-C12 branched or cyclic heteroalkyl, C2-C12 alkenyl or alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, C1-C10 alkoxy, substituted or unsubstituted phenoxy, substituted or unsubstituted aromatic hydrocarbonoxy, C1-C10 alkylthio, substituted or unsubstituted phenylthio, substituted or unsubstituted aromatic thio, C1-C10 alkanoyl, C1-C10 substituted alkylamino or C1-C10 substituted alkanoylamino;

[0010] wherein the substituted aryl, substituted heteroaryl, substituted phenoxy, substituted aromatic hydrocarbonoxy, substituted phenylthio, substituted aromatic thio are each independently substituted by 1 to 2 of the following substituents: halogen, hydroxyl, cyano, nitro, 1 to 4 carbon straight or branched alkyl, 1 to 4 carbon straight or branched alkoxy, trifluoromethyl, methylthio, trifluoromethoxy or trifluoromethylthio;

[0011] The C1-C10 straight-chain heteroalkyl, C3-C12 cyclic heteroalkyl, substituted or unsubstituted heteroaryl each independently contains 1 to 4 heteroatoms independently selected from oxygen, nitrogen or sulfur;

[0012] X is O or S;

[0013] A is the anion of the complex;

[0014] b is the charge number of the anion;

[0015] y is the absolute value of 1 / b.

[0016] Preferably, the R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 Each is independently selected from hydrogen, methyl, halogen, cyano, trifluoromethyl, carboxyl, hydroxyl, methoxy, amino, methylamino, dimethylamino, acetamido;

[0017] X is O;

[0018] A is the anion of the complex;

[0019] b is the charge number of the anion;

[0020] y is the absolute value of 1 / b.

[0021] Preferably, the complex is selected from any one of the following:

[0022]

[0023]

[0024] Furthermore, the pharmaceutically acceptable salt is a salt formed by the above-mentioned complex and hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, carbonic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, citric acid, malic acid, tartaric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, succinic acid, fumaric acid, salicylic acid, phenylacetic acid, mandelic acid or ferulic acid.

[0025] The method for preparing the complex or a pharmaceutically acceptable salt thereof of the present invention comprises the following steps:

[0026] After reacting compounds II and III with AgSO3CF3, an acid binding agent is added to carry out coordination reaction to obtain;

[0027] The chemical reaction route is as follows:

[0028]

[0029] Among them, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , X, y, A and b are as defined above.

[0030] The reaction solvent is acetonitrile, methanol, tetrahydrofuran, 1,4-dioxane, or N,N-dimethylformamide; the acid-binding agent is potassium tert-butoxide, sodium tert-butoxide, potassium carbonate, or sodium hydroxide.

[0031] The invention relates to the use of the complex or a pharmaceutically acceptable salt thereof in the preparation of anti-tumor drugs.

[0032] Wherein, the complex or a pharmaceutically acceptable salt thereof is used in the preparation of an anti-tumor drug having a dual inhibitory effect on tumor cells and tumor stem cells.

[0033] Wherein, the tumor is a malignant tumor such as breast cancer or pancreatic cancer.

[0034] The anti-tumor drug composition of the present invention comprises the complex or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

[0035] Wherein, the pharmaceutical composition is preferably a capsule, powder, tablet, granule, pill, injection, syrup, oral solution, inhalant, ointment, suppository or patch.

[0036] Preferably, the above complex and a pharmaceutically acceptable carrier form a pharmaceutical composition. Specifically, the pharmaceutical composition is in the form of a pharmaceutical preparation such as tablets, capsules, syrups, and suspensions, and can be added with common pharmaceutical excipients such as flavors, sweeteners, liquid or solid fillers or diluents.

[0037] The present invention designs a series of 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complexes, which have the dual activity of inhibiting tumor cells and tumor stem cells at the same time, and also overcome the drug resistance problem caused by tumor stem cells. More importantly, the complexes of the present invention have excellent stability and improve the anti-tumor efficacy.

[0038] The 6-phenyl-2,2'-bipyridine of the present invention can form a large π conjugated system and has a strong σ electron-donating ability, and can be combined with platinum to form a platinum complex with a stable structure. Studies have shown that the mechanism of action of the 6-phenyl-2,2'-bipyridine-platinum (II) complex is different from that of platinum drugs such as cisplatin. It mainly acts as a DNA metal intercalator, and is embedded between adjacent DNA base pairs through the π-π interaction between non-covalent ligands, thereby preventing DNA replication and transcription and inducing tumor cell apoptosis.

[0039] The nitrogen heterocyclic carbene (NHC) in the present invention can bind to platinum, stabilize the platinum center, and form an NHC platinum complex. The NHC platinum complex has good stability, few side reactions with other bioactive molecules, and its anti-tumor mechanism is different from that of platinum drugs such as cisplatin, showing great advantages in overcoming the drug resistance problem of platinum drugs such as cisplatin.

[0040] The present invention uses guanine as the NHC skeleton and takes into account the structural advantages of 6-phenyl-2,2'-bipyridine to design and synthesize a new type of 6-phenyl-2,2'-bipyridine platinum (II) complex containing guanine derivative NHC ligands. Compared with platinum drugs such as cisplatin, the stability and anti-tumor activity of the obtained complex are improved, and the problems of platinum drug resistance caused by poor stability and tumor stem cells can be overcome. The obtained complex can not only effectively inhibit the proliferation of human triple-negative breast cancer cells MD-MBA-231 and human triple-negative breast cancer cells MD-MBA-468, and the effect is significantly better than the existing platinum drug cisplatin, but also can effectively inhibit the proliferation of human triple-negative breast cancer cells MD-MBA-231 stem cells. Among them, complex 6 can also inhibit the proliferation of human breast cancer cells MCF-7, human liver cancer cells HepG2 and human prostate cancer cells DU145, but has no obvious proliferation inhibition activity on normal rat kidney cells NRK, and the effect is significantly weaker than the existing platinum drug cisplatin. The complex of the present invention, such as complex 6, can also resist GSH interference.

[0041] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0042] (1) The novel structure of the 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex provided by the present invention improves the physical and chemical properties of the platinum (II) complex by introducing natural nucleic acid guanine base, utilizing the carbon anion of 6-phenyl-2,2'-bipyridine and the guanine nitrogen heterocyclic carbene ligand with strong σ electron donating ability, and can resist the interference of biological molecules such as glutathione (GSH) containing sulfhydryl groups, thereby effectively overcoming the drug resistance caused by stability problems.

[0043] (2) The 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex of the present invention has the characteristics of dual anti-tumor effects of inhibiting tumor cells and tumor stem cells, while having no obvious toxicity to normal cells. It can effectively overcome the problem of platinum drug resistance caused by tumor stem cells, avoid the toxic side effects caused by multi-drug combination, and improve the anti-tumor efficacy.

[0044] (3) The 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex of the present invention has a simple structure, an ingenious synthetic route design, cheap and readily available raw materials, a safe and environmentally friendly synthetic process, and is easy to mass produce. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 The complex 6 inhibits the proliferation and aggregation of human triple-negative breast cancer stem cells MDA-MB-231CSCs;

[0046] Figure 2 The complex 6 solution is used to detect the complex 6 by mass spectrometry;

[0047] Figure 3 The new complexes were detected by mass spectrometry using the control compound solution. DETAILED DESCRIPTION

[0048] The present invention will be further described below in conjunction with the embodiments.

[0049] Unless otherwise specified, the materials and reagents used in the examples can be obtained from commercial sources.

[0050] Example 1

[0051] Example 1

[0052] Preparation of complex 1

[0053]

[0054] Synthesis of 7-methyl-9-(4-methylbenzyl)guanine iodide

[0055] 9-(4-methylbenzyl)guanine (cas: 101134-15-0) (1.0 g, 3.92 mmol), iodomethane (2.8 g, 19.72 mmol), N,N-dimethylformamide (7.5 mL), react at 65 ° C for 16 hours. Add ammonia water to adjust the pH to 9, and a yellow solid precipitates. The solid is collected and recrystallized from methanol and water to obtain a white solid. Hydrochloric acid and ethyl acetate solution are added to the white solid to adjust the pH to 5, and dried to obtain 0.9 g of a light yellow solid product with a yield of 57.8%.

[0056] 1 H-NMR (400MHz, DMSO-d6): δ (ppm) 11.92 (d, J = 8.1Hz, 1H, N H ),9.29(d,J=4.6Hz,1H,NC H N),7.41(s,2H,N H 2),7.30(d,J=8.1Hz,2H,Ar H ),7.22(d,J=7.9Hz,2H,Ar H ),5.30(s,2H,NC H 2),3.99(s,3H,NC H 3),2.30(s,3H,CC H 3).

[0057] Synthesis of complex 1

[0058] Add 6-(4-trifluoromethylphenyl)-2,2'-bipyridylplatinum chloride (II) (85 mg, 0.16 mmol) (cas: 1242162-54-4) and CH3CN (15 mL) to a 50 mL three-necked flask, heat to reflux, add AgCF3SO3 (61 mg, 0.24 mmol) in dark conditions and continue to react for 2 h. A yellow precipitate appears in the reaction solution, and the filtrate is collected by suction filtration. Add 7-methyl-9-(4-methylbenzyl)guanine iodide (95 mg, 0.24 mmol) and CH3CN (5 mL) to another 50 mL three-necked flask, add AgCF3SO3 (122 mg, 0.48 mmol) in dark conditions at room temperature and continue to react for 30 min. A yellow precipitate appears in the reaction solution, and the filtrate is collected by suction filtration. Combine the two parts of the filtrate, add t-BuOK (30 mg, 0.27 mmol), react at room temperature for 24 hours under nitrogen protection, then heat to reflux and continue to react for 10 hours. Filter the reaction solution with diatomaceous earth as the filter aid, collect the yellow clear filtrate, and evaporate the solvent under reduced pressure. Add 2 mL of CH3CN to the solid, filter out the insoluble matter, add 6 mL of saturated NH4PF6 aqueous solution to the filtrate, precipitate a yellow solid, filter, wash the solid with water, collect the solid, and dry. Add 2 mL of CH3CN to the solid to dissolve it, diffuse and crystallize it with 10 mL of ether to obtain 25 mg of yellow powder solid complex 1, with a yield of 17.2%

[0059] 1 H-NMR (400MHz, Acetonitrile-d3) δ (ppm): 9.37 (s, 1H, NH), 8.24-8.15 (m, 3H, ArH), 8.08 (d, J = 8.0H z,1H,ArH),8.03(d,J=8.1Hz,1H,ArH),7.79(d,J=8.1Hz,1H,ArH),7.71-7.66(m,1H,ArH),7.47-7. 45(m,1H,ArH),7.35(ddd,J=7.2,5.3,2.0Hz,1H,ArH),7.17(d,J=8.0Hz,2H,ArH),6.91-6.73(m,3H ,ArH),5.72(s,2H,NH2),5.52(m,1H,NCH),5.14(m,1H,NCH),4.06(s,3H,NCH3),2.04(s,3H,CCH3).

[0060] 13C-NMR(126MHz, Acetonitrile-d3)δ(ppm):167.98,162.51,157.02,154.63,154.14,152.88,152.65,151.80,141.98,140.26,139.7 6,137.76,133.67,132.59,132.56,128.64,128.60,128.36,125.32,124.11,121.82,120.67,120.55,109.79,48.96,37.09,19.94.

[0061] HR-MS(ESI)m / z[M] + CalcdforC 31 H 25 N7F3OPt,763.1720; Found:763.1709.

[0062] Example 2

[0063] Preparation of complex 2

[0064]

[0065] Synthesis of 7-Methyl-9-octylguanine iodide

[0066] Using 9-octylguanine (cas: 108870-41-3) (1.0 g, 3.80 mmol) and iodomethane (2.7 g, 19.02 mmol) as raw materials, the operation was the same as the synthesis of 7-methyl-9-n-butylguanine iodide to obtain 0.7 g of light yellow solid product with a yield of 45.5%.

[0067] 1 H-NMR (300MHz, DMSO-d6): δ (ppm) 11.87 (s, 1H, N H ),9.27(s,1H,NC H N),7.39(s,2H,N H 2),4.14-4.03(t,J=7.2Hz,2H,NC H 2CH2),3.99(s,3H,NC H 3),1.79(t,J=7.4Hz,2H,NCH2C H 2),1.30-1.23(m,10H,CH2C H 2C H 2C H 2C H 2C H2CH3),0.86(t,J=6.5Hz,3H,CH2C H 3).

[0068] Synthesis of complex 2

[0069] Using 6-phenyl-2,2'-bipyridinium platinum (II) chloride (74 mg, 0.16 mmol) (cas: 128025-96-7), 7-methyl-9-octylguanine iodide (97 mg, 0.24 mmol), AgCF3SO3 (61 mg, 0.24 mmol), AgCF3SO3 (122 mg, 0.48 mmol) and t-BuOK (30 mg, 0.27 mmol) as raw materials, the operation was the same as the synthesis of complex 1 to obtain 21 mg of yellow powdery solid complex 2, with a yield of 15.5%

[0070] 1 H-NMR(400MHz, Acetonitrile-d3)δ(ppm):9.35(s,1H,NH),8.37-8.35(m,1H,ArH),8.30-8.24(m ,2H,ArH),8.15(t,J=8.0Hz,1H,ArH),8.04-8.02(m,1H,ArH),7.94-7.91(m,1H,ArH),7.64-7.59 (m,2H,ArH),7.16(td,J=7.5,1.2Hz,1H,ArH),7.05(td,J=7.4,1.5Hz,1H,ArH),6.73-6.55(m,1H ,ArH),5.65(s,2H,NH2),4.29(t,J=6.9Hz,2H,NCH2),4.09(s,3H,NCH3),1.91-1.84(m,2H,NCH2C H 2),1.24-0.98(m,10H,C H 2C H 2C H 2C H 2C H 2CH3),0.76(t,J=7.0Hz,3H,CH2C H 3).

[0071] 13C-NMR(126MHz, Acetonitrile-d3)δ(ppm):168.35,164.23,157.96,154.48,153.74,152.81,152.59,151.71,147.58,141.76,140.83,139.1 2,136.95,131.64,128.61,125.54,125.09,124.34,119.83,119.50,1 09.89,46.06,36.89,31.43,29.03,28.58,28.54,25.80,22.25,13.35.

[0072] HR-MS(ESI)m / z[M] + CalcdforC 30 H 34 N7OPt,703.2473; Found:703.2462.

[0073] Example 3

[0074] Preparation of complex 3

[0075]

[0076] Using 6-(4-methoxyphenyl)-2,2'-bipyridiniumplatinum(II) chloride (79 mg, 0.16 mmol) (cas: 1242162-46-4), 7-methyl-9-octylguanine iodide (97 mg, 0.24 mmol), AgCF3SO3 (61 mg, 0.24 mmol), AgCF3SO3 (122 mg, 0.48 mmol) and t-BuOK (30 mg, 0.27 mmol) as raw materials, the operation was the same as the synthesis of complex 1 to obtain 19 mg of yellow powdery solid complex 3 with a yield of 13.5%.

[0077] 1H-NMR(300MHz, Acetonitrile-d3)δ(ppm):9.21(s,1H,NH),8.36-8.21(m,3H,ArH),8.0 8(t,J=8.0Hz,1H,ArH),7.94-7.91(m,1H,ArH),7.81-7.78(m,1H,ArH),7.62-7.56(m,2 H,ArH),6.71(dd,J=8.6,2.6Hz,1H,ArH),6.24-6.00(m,1H,ArH),5.63(s,2H,NH2),4.2 9(t,J=7.0Hz,2H,NCH2),4.09(s,3H,NCH3),3.70(s,3H,OCH3),1.93-1.84(m,2H,NCH2C H 2),1.24-0.99(m,10H,C H 2C H 2C H 2C H 2C H 2CH3),0.76(t,J=6.8Hz,3H,CH2C H 3).

[0078] 13 C-NMR(126MHz, Acetonitrile-d3)δ(ppm):168.35,164.09,162.09,158.11,154.20,153.77,152.90,152.56,151.72,141.49,141.19,140.8 2,139.91,128.46,127.30,124.20,122.73,119.17,118.25,109.39,5 4.90,46.05,36.92,31.46,29.04,28.61,28.58,25.84,22.27,13.36.

[0079] HR-MS(ESI)m / z[M] + CalcdforC 31 H 36 N7O2Pt,733.2578; Found:733.2576.

[0080] Example 4

[0081] Preparation of complex 4

[0082]

[0083] Synthesis of complex 4

[0084] Using 6-(4-trifluoromethylphenyl)-2,2'-bipyridylplatinum(II) chloride (85 mg, 0.16 mmol) (cas1242162-54-4), 7-methyl-9-octylguanine iodide (97 mg, 0.24 mmol), AgCF3SO3 (61 mg, 0.24 mmol), AgCF3SO3 (122 mg, 0.48 mmol) and t-BuOK (30 mg, 0.27 mmol) as raw materials, the operation was the same as the synthesis of complex 1 to obtain 33 mg of yellow powdery solid complex 4, with a yield of 22.5%.

[0085] 1 H-NMR (300MHz, Acetonitrile-d3) δ (ppm): 8.36-8.24 (m, 4H, ArH, NH), 8.16 (t, J = 8.0Hz ,1H,ArH),8.06-8.03(m,1H,ArH),7.96-7.93(m,1H,ArH),7.76(d,J=8.1Hz,1H,ArH),7 .69(ddd,J=7.1,5.3,1.8Hz,1H,ArH),7.48-7.45(m,1H,ArH),7.18-7.01(m,1H,ArH),5 .96(s,2H,NH2),4.21(t,J=7.2Hz,2H,NCH2),4.12(s,3H,NCH3),1.96-1.92(m,2H,NCH2C H 2),1.46-1.33(m,10H,C H 2C H 2C H 2C H 2C H 2CH3),0.94(t,J=6.9Hz,3H,CH2C H 3).

[0086] 13 C-NMR(126MHz, Acetonitrile-d3)δ(ppm):164.17,161.31,160.26,156.96,155.20,151.39,150.08,150.05,141.23,141.05,140.93,134.8 6,129.46,129.43,128.19,124.97,124.21,121.44,120.55,120.49,1 09.13,44.77,35.14,31.51,28.83,28.81,28.67,25.90,22.37,13.38.

[0087] HR-MS(ESI)m / z[M] + CalcdforC 31 H 33 N7F3OPt,771.2346; Found:771.2283.

[0088] Anal.CalcdforC 31 H 33 F9N7OPPt:C40.62,H3.63,N10.70; Found:C40.55,H3.65,N10.39.

[0089] Example 5

[0090] Preparation of complex 5

[0091]

[0092] Synthesis of 9-cyclopropylmethylguanine

[0093] 2-Amino-6-chloro-9-cyclopropylpurine (cas: 151370-27-3) (2.0 g, 8.94 mmol) and 0.5 M hydrochloric acid aqueous solution (98 mL) were added to a 250 mL three-necked flask and heated to reflux for 16 h. A 10% aqueous solution of sodium hydroxide was added to the reaction solution to adjust the pH to 8, and a white solid precipitated. The solid was collected by suction filtration, washed with water, and dried to obtain 1.7 g of a white solid product with a yield of 92.7%. ESI-MS [M+H] + 206.1.

[0094] Synthesis of 7-Methyl-9-cyclopropylmethylguanine iodide

[0095] Using 9-cyclopropylmethylguanine (1.0 g, 4.87 mmol) and iodomethane (3.5 g, 24.66 mmol) as raw materials, the operation was the same as the synthesis of 7-methyl-9-(4-methylbenzyl)guanine iodide to obtain 0.8 g of light yellow solid product with a yield of 47.3%.

[0096] 1 H-NMR (400MHz, DMSO-d6): δ (ppm) 11.81 (s, 1H, N H ),9.32-9.31(m,1H,NC H N),7.31(s,2H,N H 2), 4.02(s,3H,NC H 3),3.97(d,J=7.3Hz,2H,NC H 2),1.33-1.25(m,1H,NCH2C H),0.64-0.60(m,2H,CHC H 2CH2), 0.51-0.47(m,2H,CHCH2C H 2).

[0097] Synthesis of complex 5

[0098] Using 6-(4-trifluoromethylphenyl)-2,2'-bipyridinium platinum (II) chloride (85 mg, 0.16 mmol) (cas1242162-54-4), 7-methyl-9-cyclopropylmethylguanine iodide (83 mg, 0.24 mmol), AgCF3SO3 (61 mg, 0.24 mmol), AgCF3SO3 (122 mg, 0.48 mmol) and t-BuOK (30 mg, 0.27 mmol) as raw materials, the operation was the same as the synthesis of complex 1 to obtain 22 mg of yellow powdery solid complex 5, with a yield of 16.0%

[0099] 1 H-NMR(300MHz, Acetonitrile-d3)δ(ppm):8.34-8.22(m,5H,ArH,NH),8.13(d,J=8.0Hz,1H,ArH),8.04(d,J=8.1Hz,1H,ArH),7.81(d,J=8.2Hz,1H,ArH),7.62(ddd ,J=6.9,5.6,2.2Hz,1H,ArH),7.51-7.45(m,1H,ArH),7.02-6.81(m,1H,ArH),5.6 8(s,2H,NH2),4.27-4.12(m,2H,NCH2),4.11(s,3H,NCH3),1.42-1.34(m,1H,CH2C H CH2), 0.42-0.35(m,4H,C H 2CHC H 2).

[0100] 13 C-NMR(151MHz, Acetonitrile-d3)δ(ppm):166.81,162.54,157.85,154.84,153.91,152.87,152.81,151.71,151.66,142.11,141 .07,140.03,132.76,132.74,128.80,125.41,124.55,121.96,121.93,120.82,120.71,110.13,50.92,36.88,11.08,3.76,3.60.

[0101] HR-MS(ESI)m / z[M] + CalcdforC 27 H 23 N7F3OPt,713.1564; Found:713.1557.

[0102] Example 6

[0103] Preparation of complex 6

[0104]

[0105] Synthesis of 7-methyl-9-benzylguanine iodide

[0106] Using 9-benzylguanine (cas: 14937-72-5) (1.0 g, 4.15 mmol) and iodomethane (2.9 g, 20.43 mmol) as raw materials, the operation was the same as the synthesis of 7-methyl-9-(4-methylbenzyl)guanine iodide in Example 1 to obtain 0.8 g of light yellow solid product with a yield of 50.3%.

[0107] 1 H-NMR (400MHz, DMSO-d6): δ (ppm) 11.91 (m, 1H, N H ),9.32(s,1H,NC H N), 7.44-7.35(m,7H,Ar H ,N H 2),5.37(s,2H,NC H 2),4.01(s,3H,NC H 3).

[0108] Synthesis of complex 6

[0109] Using 6-(4-trifluoromethylphenyl)-2,2'-bipyridinium platinum (II) chloride (85 mg, 0.16 mmol) (cas1242162-54-4), 7-methyl-9-benzylguanine iodide (92 mg, 0.24 mmol), AgCF3SO3 (61 mg, 0.24 mmol), AgCF3SO3 (122 mg, 0.48 mmol) and t-BuOK (30 mg, 0.27 mmol) as raw materials, the operation was the same as the synthesis of complex 1 to obtain 20 mg of yellow powdery solid complex 6, with a yield of 14.0%

[0110] 1H-NMR(300MHz, Acetonitrile-d3)δ(ppm):8.37-8.22(m,4H,ArH,NH),8.15(t,J=8.1Hz,1H,ArH),8.02(d,J=7.8Hz,1H,ArH),7.92(d,J=7.9Hz,1H,ArH),7.7 4(d,J=8.2Hz,1H,ArH),7.70-7.65(m,1H,ArH),7.51-7.44(m,6H,ArH),7.16-6 .99(m,1H,ArH),6.01(s,2H,NH2),5.45-5.35(m,2H,NCH2),4.08(s,3H,NCH3).

[0111] 13 C-NMR(126MHz, Acetonitrile-d3)δ(ppm):164.09,161.48,160.26,156.86,155.13,151.35,150.15,150.03,141.20,141.03,140.9 0,134.88,134.42,129.43,129.40,129.09,128.84,128.50,128.17,124.98,124.21,121.43,120.55,120.47,109.14,47.91,35.29.

[0112] HR-MS(ESI)m / z[M] + CalcdforC 30 H 23 N7F3OPt,749.1564; Found:749.1520.

[0113] Anal.CalcdforC 30 H 23 F9N7OPPt:C40.28,H2.59,N10.96; Found:C39.45,H2.58,N10.52.

[0114] Example 7

[0115] Preparation of complex 7

[0116]

[0117] Synthesis of 6-(4-chlorophenyl)-2,2'-bipyridine

[0118] 1-[2-oxo-2-(2-pyridyl)ethyl]pyridinium iodide (3.60 g, 11.03 mmol) (cas: 26482-00-8), NH4OAc (7.70 g, 99.89 mmol) and CH3COOH (34 mL) were added to a three-necked flask, and the temperature was raised to reflux under nitrogen protection. After adding 3-(dimethylamino)-4'-chlorophenylacetone hydrochloride (2.73 g, 11.00 mmol) (cas: 1798-83-0), the reaction was continued for 3 h. TLC (petroleum ether: ethyl acetate = 4:1) monitored the complete reaction of the raw materials. After the reaction solution was cooled to room temperature, 90 mL of water was added to precipitate a brown solid, which was filtered, washed and dried. The crude product was separated by column chromatography (petroleum ether: ethyl acetate = 50:1 to 30:1) to obtain 1.64 g of a white solid product with a yield of 55.9%, ESI-MS m / z [M+H] + :267.0,mp90~91℃.

[0119] 1 HNMR(300MHz,Chloroform-d)δ(ppm):8.77-8.75(m,1H,ArH),8.66(d,J=8.0Hz,1H,ArH),8.46(d,J=7.8Hz,1H,ArH),8. 15-8.12(m,2H,ArH),7.97-7.90(m,2H,ArH),7.80(d,J=7.8Hz,1H,ArH),7.53-7.49(m,2H,ArH),7.43-7.39(m,1H,ArH).

[0120] Synthesis of 6-(4-chlorophenyl)-2,2'-bipyridylplatinum(II) chloride

[0121] Add 6-(4-chlorophenyl)-2,2'-bipyridine (1.41 g, 5.29 mmol), K2PtCl4 (2.20 g, 5.30 mmol) and CH3COOH (65 mL) into a three-necked flask, and heat to reflux for 14 h under nitrogen protection. The reaction solution was cooled to room temperature, filtered, and the solid was washed with CH3COOH, MeOH, EA, and Et2O (4 mL × 3) to obtain 1.57 g of white solid product with a yield of 59.8%, ESI-MS m / z [M+H] + :495.9,mp>300℃.

[0122] 1HNMR(300MHz,DMSO-d6)δ(ppm):8.80-8.79(m,1H,ArH),8.50-8.45(m,1H,ArH),8.35-8.30(m,1H,ArH),8.16-8 .09(m,2H,ArH),7.90-7.88(m,2H,ArH),7.64-7.58(m,1H,ArH),7.42-7.26(m,1H,ArH),7.13-7.11(m,1H,ArH).

[0123] Synthesis of complex 7

[0124] Using 6-(4-chlorophenyl)-2,2'-bipyridiniumplatinum(II) chloride (79 mg, 0.16 mmol), 7-methyl-9-octylguanine iodide (97 mg, 0.24 mmol), AgCF3SO3 (61 mg, 0.24 mmol), AgCF3SO3 (122 mg, 0.48 mmol) and t-BuOK (30 mg, 0.27 mmol) as raw materials, the operation was the same as the synthesis of complex 1 to obtain 19 mg of yellow powdery solid complex 7 with a yield of 13.4%.

[0125] 1 H-NMR(400MHz, Acetonitrile-d3)δ(ppm):9.71(s,1H,NH),8.34-8.22(m,3H,ArH ),8.17(t,J=8.0Hz,1H,ArH),8.06-8.04(m,1H,ArH),7.93-7.90(m,1H,ArH),7.6 5-7.59(m,2H,ArH),7.19(dd,J=8.3,2.1Hz,1H,ArH),6.73-6.56(m,1H,ArH),5.7 8(s,2H,NH2),4.34-4.23(m,2H,NCH2),4.09(s,3H,NCH3),1.91-1.84(m,2H,NCH2C H 2),1.31-0.99(m,10H,C H 2C H 2C H 2C H 2C H 2CH3),0.77(t,J=7.0Hz,3H,CH2C H 3).

[0126] 13C-NMR(126MHz, Acetonitrile-d3)δ(ppm):167.05,163.09,157.94,154.57,153.89,152.93,152.70,151.74,146.36,141.93,141.04,13 6.66,136.07,128.72,126.72,124.96,124.45,120.17,119.81,110.11,46.10,36.88,31.45,29.07,28.61,28.61,25.87,22.26,13.35.

[0127] HR-MS (ESI) m / z [M+H] + CalcdforC 30 H 33 N7ClOPt,737.2083; Found:738.2077.

[0128] Example 8

[0129] Pharmacological experiments

[0130] In this experiment, the CCK8 method was used to test the in vitro tumor cell proliferation inhibition activity of the complex of the present invention and the commonly used platinum anti-tumor drugs in clinical practice.

[0131] (1) Experimental methods

[0132] The cells were plated at 4 × 10 3 The density of cells / well (100μL) was inoculated into a 96-well plate, and the 96-well plate was placed in an incubator at 37°C and 5% CO2 overnight. On the second day, 100μL of fresh culture medium containing 200μM of each compound complex was added to the first column of wells, mixed evenly, and then applied by half-dilution. After a certain period of incubation for 96h, 20μL CCK8 reagent was added to each well. After incubation for another 2h, OD450nm was measured with an enzyme reader. A freshly prepared 10mM solution of the compound complex in N,N-dimethylformamide (DMF) was used as a stock solution, stored at -20°C for standby use, and diluted to 200μM with complete culture medium when used; cisplatin was prepared with culture medium before each use.

[0133] (2) Test results

[0134] The anti-proliferative activity data of the complex of the present invention on human triple-negative breast cancer cells MDA-MB-231, human triple-negative breast cancer cells MDA-MB-468, human triple-negative breast cancer stem cells MDA-MB-231CSCs, human breast cancer cells MCF-7, human liver cancer cells HepG2, human prostate cancer cells DU145 and normal rat kidney cells NRK are shown in Tables 1 and 2, and cisplatin is used as a positive control drug;

[0135] Table 1 Antiproliferative activity data of the complexes on MDA-MB-231, MDA-MB-468 and MDA-MB-231CSCs, with an action time of 96 hours

[0136]

[0137]

[0138] Table 2 Antiproliferative activity data of the complexes on MCF-7, HepG2, DU145 and NRK, the action time is 96 hours

[0139]

[0140] As can be seen from Table 1, the complex has good inhibitory activity on malignant tumor cells: the complex has better proliferation inhibitory activity on human triple-negative breast cancer cells MDA-MB-231, human triple-negative breast cancer cells MDA-MB-468, and human breast cancer stem cells MDA-MB-231 than the positive drug cisplatin; as can be seen from Table 2, complex 6 has good proliferation inhibitory activity on human breast cancer cells MCF-7, human liver cancer cells HepG2, and human prostate cancer cells DU145; complex 6 has no obvious proliferation inhibitory activity on normal rat kidney cells NRK, and its inhibitory activity is significantly weaker than the positive drug cisplatin. The above explanation shows that the complex of the present invention shows strong inhibitory activity on breast cancer, liver cancer and prostate cancer cells, especially triple-negative breast cancer cells and their stem cells, which are currently difficult to treat, and has no obvious toxicity to normal cells, and has very good selectivity.

[0141] Example 9

[0142] Pharmacological experiments

[0143] In this experiment, the complex 6 of the present invention was tested for inhibiting the in vitro proliferation and in vitro clustering of breast cancer stem cells.

[0144] (1) Experimental methods

[0145] MDA-MB-231CSCs cells were cultured at 1×10 5The density of cells / well (2.5mL) was inoculated into a 6-well plate, and the 6-well plate was placed in an incubator at 37°C and 5% CO2 overnight. The next day, after all the cells were attached to the wall, the complete medium was aspirated, and 6.25μM and 3.12μM complex 6 solutions were prepared with a medium containing 10% serum. The complete medium, 6.25μM complex 6 solution and 3.12μM complex 6 solution were added to the 6-well plate, respectively, and incubated for 96h. The cells were observed and photographed using a fluorescent inverted microscope.

[0146] (2) Test results

[0147] The inhibitory effect of the complex 6 of the present invention on the proliferation and clustering of human triple-negative breast cancer stem cells MDA-MB-231CSCs is shown in Figure 1 , and the blank was used as control; Figure 1 It can be seen that complex 6 can effectively inhibit the in vitro proliferation and in vitro aggregation of human triple-negative breast cancer stem cells MDA-MB-231CSCs.

[0148] Example 10

[0149] Stability test

[0150] In this experiment, the complex of the present invention was co-incubated with GSH in vitro, and the stability of the complex was determined by mass spectrometry detection of the system.

[0151] (1) Experimental methods

[0152] Prepare 400 μL of 0.25 mM DMF / PBS (DMF=100 μL, PBS=300 μL) solution of complex 6 and control compound (structure shown below), add 100 μL of 10 mM GSH solution in PBS, shake well, wherein the pH of PBS used is 7.4, incubate at room temperature for two weeks, and then perform mass spectrometry on the system.

[0153]

[0154] (2) Test results

[0155] The experimental results are shown in Figure 2 After the complex 6 of the present invention was co-incubated with in vitro glutathione for two weeks, complex 6 was detected in the system, but no new complex was detected, indicating that complex 6 has a good ability to resist GSH interference and has good stability. After the control was co-incubated with in vitro glutathione for two weeks, a new complex ( Figure 3 ), indicating that the control reacted chemically with GSH. Therefore, the stability of the complex 6 of the present invention is higher than that of the analogs containing terpyridine ligands.

Claims

1. A 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex of general formula (I) or a pharmaceutically acceptable salt thereof: in: R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 are each independently selected from hydrogen, halogen, cyano, nitro, trifluoromethyl, hydroxyl, amino, carboxyl, C1-C10 straight-chain alkyl, C1-C10 straight-chain heteroalkyl, C3-C12 branched or cyclic alkyl, C3-C12 branched or cyclic heteroalkyl, C2-C12 alkenyl or alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, C1-C10 alkoxy, substituted or unsubstituted phenoxy, substituted or unsubstituted aromatic hydrocarbonoxy, C1-C10 alkylthio, substituted or unsubstituted phenylthio, substituted or unsubstituted aromatic thio, C1-C10 alkanoyl, C1-C10 substituted alkylamino or C1-C10 substituted alkanoylamino; wherein the substituted aryl, substituted heteroaryl, substituted phenoxy, substituted aromatic hydrocarbonoxy, substituted phenylthio, substituted aromatic thio are each independently substituted by 1 to 2 of the following substituents: halogen, hydroxyl, cyano, nitro, 1 to 4 carbon straight or branched alkyl, 1 to 4 carbon straight or branched alkoxy, trifluoromethyl, methylthio, trifluoromethoxy or trifluoromethylthio; The C1-C10 straight-chain heteroalkyl, C3-C12 cyclic heteroalkyl, substituted or unsubstituted heteroaryl each independently contains 1 to 4 heteroatoms independently selected from oxygen, nitrogen or sulfur; X is O or S; A is the anion of the complex; b is the charge number of the anion; y is the absolute value of 1 / b.

2. The 6-phenyl-2,2'-bipyridine platinum (II)-guanine nitrogen heterocyclic carbene complex or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 Each is independently selected from hydrogen, methyl, halogen, cyano, trifluoromethyl, carboxyl, hydroxyl, methoxy, amino, methylamino, dimethylamino, acetamido; X is O; A is the anion of the complex; b is the charge number of the anion; y is the absolute value of 1 / b.

3. The complex according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: The complex is preferably selected from any one of the following:

4. A method for preparing the complex or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The steps include: After reacting compounds II and III with AgSO3CF3, an acid binding agent is added to carry out coordination reaction to obtain; The chemical reaction route is as follows: Among them, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , X, y, A and b are defined the same as in claim 1.

5. The method for preparing the complex or a pharmaceutically acceptable salt thereof according to claim 4, characterized in that: The reaction solvent is acetonitrile, methanol, tetrahydrofuran, 1,4-dioxane, or N,N-dimethylformamide; the acid-binding agent is potassium tert-butoxide, sodium tert-butoxide, potassium carbonate, or sodium hydroxide.

6. Use of the complex according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof in the preparation of an anti-tumor drug.

7. The use according to claim 6, characterized in that: The complex or a pharmaceutically acceptable salt thereof is used in the preparation of an anti-tumor drug having a dual inhibitory effect on tumor cells and tumor stem cells.

8. The use according to claim 6, characterized in that: The tumor is a malignant tumor such as breast cancer or pancreatic cancer.

9. An anti-tumor drug composition, characterized in that: It comprises the complex described in any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

10. The pharmaceutical composition according to claim 9, characterized in that The pharmaceutical composition is preferably a capsule, powder, tablet, granule, pill, injection, syrup, oral solution, inhalant, ointment, suppository or patch.

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