阅读说明：本技术 一种人卵巢癌细胞抑制剂、制备方法及其应用 (Human ovarian cancer cell inhibitor, preparation method and application thereof ) 是由 杨燕 宾贻东 廖志强 农基健 于 2020-12-24 设计创作，主要内容包括：本发明公开了一种人卵巢癌细胞抑制剂及其制备方法、应用,属于医药领域。其化学名称为六氟磷酸化·2,12,13-三甲基吡啶并[3,2-a]吡啶并[1’,2’:1,2]咪唑并[4,5-c]吩嗪·二-苯并喹啉合铱(Ⅲ)。其制备方法步骤包括:8-羟基喹啉与盐酸、NaClO-3反应；加入无水碳酸钾；加入2-氨基-5-甲基吡啶；加入4,5-二甲基邻苯二胺；得化合物3a；7,8-苯并喹啉和水合三氯化铱反应得化合物4；化合物3a与化合物4反应,得人卵巢癌细胞抑制剂。本发明对人卵巢癌耐药株SK-OV-3/DDP的体外抗肿瘤活性大于8-羟基喹啉衍生物配体和金属基抗癌药物顺铂；对正常细胞HL-7702的毒性很小,体现出靶向抑制人卵巢癌增殖的良好效果。(The invention discloses a human ovarian cancer cell inhibitor, a preparation method and application thereof, and belongs to the field of medicines. Its chemical name is hexafluorophosphorylation.2, 12, 13-trimethylpyrido [3,2-a]Pyrido [1',2':1,2]Imidazo [4, 5-c)]Phenazine bis-benzoquinolinato iridium (III). The preparation method comprises the steps of 8-hydroxyquinoline, hydrochloric acid and NaClO 3 Carrying out reaction; adding anhydrous potassium carbonate; adding 2-amino-5-methylpyridine; adding 4, 5-dimethyl o-phenylenediamine; to obtain a compound 3 a; reacting 7, 8-benzoquinoline with iridium trichloride hydrate to obtain a compound 4; reacting the compound 3a with the compound 4 to obtain the human ovarian cancer cell inhibitor. The in vitro anti-tumor activity of the invention on human ovarian cancer drug-resistant strain SK-OV-3/DDP is greater than that of 8-hydroxyquinoline derivative ligand and metal-based anti-cancer drug cisplatin; toxicity to Normal cell HL-7702Has small sex, and shows good effect of targeted inhibition of human ovarian cancer proliferation.)

1. A human ovarian cancer cell inhibitor, which is hexafluorophosphorylation 2,12, 13-trimethylpyrido [3,2-a ] pyrido [1',2':1,2] imidazo [4,5-c ] phenazine bis-benzoquinolinyliridium (III), and has the following structural formula:

2. the method for preparing the human ovarian cancer cell inhibitor of claim 1, which comprises the following steps:

(1) adding 8-hydroxyquinoline and hydrochloric acid into a container, adding NaClO after the 8-hydroxyquinoline is dissolved₃To obtain a compound 1;

(2) adding the compound 1, anhydrous potassium carbonate and ethanol into a container, stirring to dissolve the compound 1, adding 2-amino-5-methylpyridine, and refluxing a reaction mixture to obtain a compound 2;

(3) adding the compound 2, absolute ethyl alcohol and acetic acid into a container, stirring to dissolve the compound 2, adding 4, 5-dimethyl o-phenylenediamine into the container, and performing reflux reaction to obtain a compound 3 a;

(4) adding 7, 8-benzoquinoline and iridium trichloride hydrate into a container, and then sequentially adding ethylene glycol ethyl ether and water; introducing nitrogen, reacting and refluxing to obtain a compound 4;

(5) adding the compound 4 and the compound 3a into a container, adding ethylene glycol, carrying out reflux reaction under the protection of argon, and adding a saturated ammonium hexafluorophosphate solution after the reaction is finished to obtain a precipitate; the precipitate is hexafluorophosphorylation 2,12, 13-trimethylpyrido [3,2-a ] pyrido [1',2':1,2] imidazo [4,5-c ] phenazine bis-benzoquinolinylidium (III).

3. The method for preparing the human ovarian cancer cell inhibitor according to claim 2, wherein the step (1) is: adding 8-hydroxyquinoline and hydrochloric acid into a container, heating to 40-45 ℃, adding NaClO after 8-hydroxyquinoline is dissolved₃Continuously stirring and reacting for 1-3h at the temperature of 40-45 ℃; diluting with water after reaction, removing precipitate, extracting the filtrate, mixing the organic phases, washing with water, and rotary evaporating to obtain the solutionPreparing a solid, filtering and precipitating, and recrystallizing the solid to obtain a compound 1;

8-hydroxyquinoline and NaClO₃The molar ratio is 1: 5-6.

4. The method for preparing the human ovarian cancer cell inhibitor according to claim 2, wherein the step (2) is: adding the compound 1 and anhydrous potassium carbonate into a container, adding ethanol, stirring to completely dissolve the compound 1, adding 2-amino-5-methylpyridine for reaction, refluxing the reaction mixture at 80-85 ℃ for 15-20h, cooling, precipitating, filtering, and recrystallizing with o-dichlorobenzene to obtain a compound 2;

the molar ratio of the compound 1 to anhydrous potassium carbonate is 1: 1-2;

the molar ratio of 2-amino-5-methylpyridine to compound 1 is 2-3: 1.

5. The method for preparing the human ovarian cancer cell inhibitor according to claim 2, wherein the step (3) is: adding the compound 2 into a container, adding absolute ethyl alcohol and acetic acid, stirring and dissolving, then adding 4, 5-dimethyl o-phenylenediamine into the container, carrying out reflux reaction at the temperature of 80-90 ℃, cooling to separate out a precipitate, filtering, and recrystallizing the precipitate to obtain a compound 3 a;

the molar ratio of the compound 2 to the 4, 5-dimethyl o-phenylenediamine is 1: 1-2.

6. The method for preparing the human ovarian cancer cell inhibitor according to claim 2, wherein the step (4) is: putting 7, 8-benzoquinoline and iridium trichloride hydrate into a container, and then sequentially adding ethylene glycol ethyl ether and water; introducing nitrogen, heating to 120-;

the molar ratio of the 7, 8-benzoquinoline to the iridium trichloride hydrate is 1-2: 1.

7. The method for preparing the human ovarian cancer cell inhibitor according to claim 2, wherein the step (5) is: adding the compound 4 and the compound 3a into a container, adding ethylene glycol, and heating and refluxing for 10-15h under the protection of argon; cooling, adding water for dilution, and adding a saturated ammonium hexafluorophosphate solution to generate a precipitate; filtering, washing the precipitate with water and ether, and drying; dissolving the precipitate with acetonitrile, and separating with neutral alumina column; leaching a mixed solvent of dichloromethane and acetonitrile, and distilling and spin-drying to remove the solvent to obtain the human ovarian cancer cell inhibitor;

the molar ratio of the compound 4 to the compound 3a is 1: 1-2.

8. The method for preparing the human ovarian cancer cell inhibitor of claim 7, wherein the volume ratio of the dichloromethane to the acetonitrile is 3: 1.

9. Use of the human ovarian cancer cell inhibitor of claim 1 in the preparation of an antitumor medicament.

10. Use of the human ovarian cancer cell inhibitor of claim 1 as an active ingredient in the preparation of an external antitumor drug.

Technical Field

The invention relates to the technical field of medicines, and particularly relates to a human ovarian cancer cell inhibitor, a preparation method and application thereof.

Background

At present, about 5.2 thousands of women are diagnosed with ovarian cancer every year in China, about 2.2 thousands of people die of ovarian cancer, and the incidence of ovarian cancer is in a youthful trend in the last 10 years. Ovarian cancer has the characteristics of easy proliferation, high invasiveness and the like, which is the main reason of failure of clinical treatment, so that the inhibition of the proliferation and invasion of ovarian cancer cells is one of the key points and difficulties in the treatment of ovarian cancer.

The 8-hydroxyquinoline derivative is an intermediate of various medicines (such as quiiodoform, chloroiodoquinoline and the like), and has wide pharmacological activities, such as anticancer activity, anti-AIDS activity, antifungal activity, anti-schistosome activity, antioxidant activity, neuroprotective activity and the like. The research on the antitumor activity of 8-hydroxyquinoline is late, and Shaw et al analyze the structure-activity relationship of different types of 8-hydroxyquinoline derivatives and find that 8-hydroxyquinoline is the main anti-tumor pharmacophore. However, the 8-hydroxyquinoline derivative iridium (III) complex is not reported as a human ovarian cancer cell inhibitor.

Disclosure of Invention

The invention provides a human ovarian cancer cell inhibitor, a preparation method and application thereof, wherein the inhibitor has higher activity than a clinical medicine cisplatin; the preparation method takes 8-hydroxyquinoline derivatives as active ligands to synthesize the human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]。

The first purpose of the invention is realized by the following technical scheme:

a human ovarian cancer cell inhibitor is named as hexafluorophosphorylation 2,12, 13-trimethyl pyrido [3,2-a ] pyrido [1',2':1,2] imidazo [4,5-c ] phenazine bis-benzoquinolinylium (III), and has the following structural formula:

the second purpose of the invention is realized by the following technical scheme:

the preparation method of the human ovarian cancer cell inhibitor comprises the following steps:

(1) adding 8-hydroxyquinoline and hydrochloric acid into a container, adding NaClO after the 8-hydroxyquinoline is dissolved₃To obtain a compound 1;

(2) adding the compound 1, anhydrous potassium carbonate and ethanol into a container, stirring to dissolve the compound 1, adding 2-amino-5-methylpyridine, and refluxing a reaction mixture to obtain a compound 2;

(3) adding the compound 2, absolute ethyl alcohol and acetic acid into a container, stirring to dissolve the compound 2, adding 4, 5-dimethyl o-phenylenediamine into the container, and performing reflux reaction to obtain a compound 3 a;

(4) adding 7, 8-benzoquinoline and iridium trichloride hydrate into a container, and then sequentially adding ethylene glycol ethyl ether and water; introducing nitrogen, reacting and refluxing to obtain a compound 4;

(5) adding the compound 4 and the compound 3a into a container, adding ethylene glycol, carrying out reflux reaction under the protection of argon, and adding a saturated ammonium hexafluorophosphate solution after the reaction is finished to obtain a precipitate; the precipitate is hexafluorophosphorylation 2,12, 13-trimethylpyrido [3,2-a ] pyrido [1',2':1,2] imidazo [4,5-c ] phenazine bis-benzoquinolinylidium (III).

Preferably, the step (1) is: adding 8-hydroxyquinoline and hydrochloric acid into a container, heating to 40-45 ℃, adding NaClO after 8-hydroxyquinoline is dissolved₃Continuously stirring and reacting for 1-3h at the temperature of 40-45 ℃; and (3) diluting with water after the reaction is finished, removing the precipitate, extracting the filtrate, combining organic phases, washing with water, evaporating the solvent in a rotary manner to obtain a solid, filtering the precipitate, and recrystallizing the solid to obtain the compound 1.

8-hydroxyquinoline and NaClO₃The molar ratio is 1: 5-6; the hydrochloric acid concentration is 35-37% volume fraction.

Preferably, the step (2) is: adding the compound 1 and anhydrous potassium carbonate into a container, adding ethanol, stirring to completely dissolve the compound 1, adding 2-amino-5-methylpyridine for reaction, refluxing the reaction mixture at 80-85 ℃ for 15-20h, cooling, precipitating, filtering, and recrystallizing with o-dichlorobenzene to obtain a compound 2;

the molar ratio of the compound 1 to the anhydrous potassium carbonate is 1: 1-2.

The molar ratio of 2-amino-5-methylpyridine to compound 1 is 2-3: 1.

Preferably, the step (3) is: adding the compound 2 into a container, adding absolute ethyl alcohol and acetic acid, stirring and dissolving, then adding 4, 5-dimethyl o-phenylenediamine into the container, carrying out reflux reaction at the temperature of 80-90 ℃, cooling to separate out a precipitate, filtering, and recrystallizing the precipitate to obtain a compound 3 a.

The molar ratio of the compound 2 to the 4, 5-dimethyl o-phenylenediamine is 1: 1-2.

Preferably, the step (4) is: putting 7, 8-benzoquinoline and iridium trichloride hydrate into a container, and then sequentially adding ethylene glycol ethyl ether and water; introducing nitrogen, heating to 120-phase and 150 ℃ for refluxing for 24-48h, cooling, pouring water into the reaction liquid, stirring to separate out a solid, filtering the solid, washing with water, washing with ethanol, and drying to obtain the compound 4.

The molar ratio of the 7, 8-benzoquinoline to the iridium trichloride hydrate is 1-2: 1.

Preferably, the step (5) is: adding the compound 4 and the compound 3a into a container, adding ethylene glycol, and heating and refluxing for 10-15h under the protection of argon; cooling, adding water for dilution, and adding a saturated ammonium hexafluorophosphate solution to generate a precipitate; filtering, washing the precipitate with water and ether, and drying; dissolving the precipitate with acetonitrile, and separating with neutral alumina column; leaching a mixed solvent of dichloromethane and acetonitrile, and distilling and spin-drying to remove the solvent to obtain the human ovarian cancer cell inhibitor;

the molar ratio of the compound 4 to the compound 3a is 1: 1-2.

The volume ratio of dichloromethane to acetonitrile was 3: 1.

The third purpose of the invention is realized by applying the human ovarian cancer cell inhibitor in preparing anti-tumor drugs.

The third purpose of the invention can also be realized by the application of the human ovarian cancer cell inhibitor as an effective component in the preparation of external antitumor drugs.

Human ovarian cancer prepared by the inventionCytostatic agents [ Ir (3a) (BQ)₂]Has high antitumor activity on human ovarian cancer drug-resistant strain SK-OV-3/DDP and IC thereof₅₀The value range is 0.002 +/-0.001 mu M, and the in vitro anti-tumor activity of the compound is more than that of 8-hydroxyquinoline derivative ligand and metal-based anti-cancer drug cisplatin; has little toxicity to normal cell HL-7702 (IC)₅₀More than 50 mu M), shows good target inhibition of human ovarian cancer proliferation. Human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]Has potential medicinal value and can be used for preparing antitumor drugs.

Drawings

FIG. 1 shows a human ovarian cancer cell inhibitor [ Ir (3a) (BQ) prepared in example 1 of the present invention₂]Electrospray mass spectrogram of (a);

FIG. 2 shows the human ovarian cancer cell inhibitor [ Ir (3a) (BQ) prepared in example 1 of the present invention₂]The nuclear magnetic resonance hydrogen spectrum of (a);

FIG. 3 shows the human ovarian cancer cell inhibitor [ Ir (3a) (BQ) prepared in example 1 of the present invention₂]Nuclear magnetic resonance carbon spectrum of (a);

FIG. 4 shows the human ovarian cancer cell inhibitor [ Ir (3a) (BQ) prepared in example 1 of the present invention₂]An infrared spectrum of (1);

FIG. 5 shows a human ovarian cancer cell inhibitor [ Ir (3a) (BQ) prepared in the present invention₂]The chemical structural formula of (1);

FIG. 6 is a synthetic route diagram of the present invention.

In FIG. 6, compound 1 is 6, 7-dichloroquinoline-5, 8-dione; the compound 2 is 9-methylpyrido [2',1':2,3]Imidazo [4,5-h]Quinoline-5, 6-dione; the compound 3a is 2,12, 13-trimethylpyrido [3,2-a ]]Pyrido [1',2':1,2]Imidazo [4, 5-c)]A phenazine; compound 4 is iridium (III) chloro-bridged dimer [ (H-BQ)₂Ir(μ-Cl)]₂(ii) a (ii) a Compound [ Ir (3a) (BQ)₂]Is hexafluorophosphorylation 2,12, 13-trimethylpyrido [3,2-a ]]Pyrido [1',2':1,2]Imidazo [4, 5-c)]Phenazine bis-benzoquinolinato iridium (III).

Detailed Description

The technical solution in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. The drawings are only for purposes of illustration and are not intended to be limiting, and are merely schematic and non-limiting. For those skilled in the art to which the invention pertains, several simple deductions or substitutions can be made without departing from the spirit of the invention, and all shall be considered as belonging to the protection scope of the invention.

Example 1

(1) Human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]The preparation and characterization of (1):

weighing a compound 4 and a compound 3a (8-hydroxyquinoline derivative) according to a molar ratio of 1:1.3, putting the mixture into a container, adding ethylene glycol, and heating and refluxing for 13 hours under the protection of argon to obtain a dark red clear solution; cooling to room temperature, adding water for dilution, and then adding a saturated ammonium hexafluorophosphate solution to generate a large amount of red precipitates; filtering, washing with water and ether, and drying; dissolving the dried crude product with acetonitrile, and separating with neutral alumina column; eluting with mixed solvent of V (dichloromethane) and V (acetonitrile) ═ 3:1 to collect yellow component, and vacuum distilling to remove solvent to obtain solid product, i.e. human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]I.e. hexafluorophosphorylation 2,12, 13-trimethylpyrido [3,2-a ]]Pyrido [1',2':1,2]Imidazo [4, 5-c)]Phenazine bis-benzoquinolinyliridium (III) to obtain earthy yellow target product [ Ir (3a) (BQ)₂]。

For the obtained [ Ir (3a) (BQ)₂]And (3) identification:

(1) electrospray mass spectrometry, the spectrum of which is shown in figure 1.

ESI-MS m/z:912.2423[M]⁺Wherein M is a human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]Molecular weight of (2).

(2) The NMR spectrum is shown in FIG. 2.

¹H NMR(500MHz,DMSO-d₆)δ9.40(s,1H),9.08(s,1H),8.81–8.56(m,3H),8.49(d,J＝8.0Hz,1H),8.27–8.17(m,1H),8.13(d,J＝5.0Hz,1H),8.08(d,J＝8.8Hz,1H),7.99(dd,J＝27.8,8.9Hz,3H),7.92–7.83(m,2H),7.70(d,J＝7.9Hz,2H),7.64(d,J＝8.0Hz,1H),7.50(d,J＝9.3Hz,1H),7.41–7.35(m,1H),7.35–7.28(m,2H),6.56(d,J＝7.1Hz,1H),6.47(d,J＝7.1Hz,1H),6.02(d,J＝9.2Hz,1H),2.51(s,3H),2.50(s,3H),2.45(s,3H)。

(3) NMR spectrum of carbon, as shown in FIG. 3.

¹³C NMR(126MHz,DMSO-d₆)δ157.34,156.76,146.99,145.97,144.02,141.75,141.66,141.38,138.24,137.98,136.08,134.25,134.11,130.09,129.84,129.72,129.05,127.70,127.28,127.19,126.96,126.92,125.91,124.77,124.71,123.14,121.04,120.96,113.52,20.41,19.83,18.34.

(4) The infrared spectrum is shown in FIG. 4.

Infrared spectrum of (KBr pellet, cm)^-1)：3426,3032,2929,1616,1531,1446,1437,1390,1202,1099,1015,847,724,556。

Therefore, the obtained yellow target product can be determined to be a human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]The structural formula is shown in figure 5.

Example 2

Human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]The synthetic route of (2) is shown in FIG. 6:

(1) preparation of compound 1: 600mL of concentrated hydrochloric acid (pre-frozen in a refrigerator and 37% by mass concentration) and 14.5g of 8-hydroxyquinoline were added to a 1L round-bottom flask, heated to 40 ℃ and 53g of NaClO were added in portions after the 8-hydroxyquinoline was completely dissolved₃(addition was complete over 60 minutes) and after addition stirring was continued for 2h at 40 ℃. After the reaction was completed, the reaction mixture was diluted to 2L with ice water and CH₂Cl₂(6X 250mL), the organic phases are combined, washed with 3X 200mL of distilled water, the solvent is evaporated off by vacuum rotary evaporation to obtain a yellow solid, the precipitate is filtered, and the solid is recrystallized three times with 40mL of methanol to obtain 6, 7-dichloroquinoline-5, 8-dione, which is recorded as compound 1;

(2) preparation of compound 2: 2.28g (0.01mol) of Compound 1 and 1.363g of anhydrous potassium carbonate (0.01mol) were added to a 50mL round-bottomed flask, 20mL of ethanol was added, stirring was performed to completely dissolve Compound 1, 2.16g (0.02mol) of 2-amino-5-methylpyridine was added in portions, and the reaction mixture was refluxed at 84 ℃ for 15 hours; cooling to room temperature, precipitating, filtering, and recrystallizing with o-dichlorobenzene to obtain compound 2.

(3) Preparation of compound 3 a: weighing 1mmol of compound 2, adding the compound into a 50mL round-bottom flask, adding 5mL of absolute ethyl alcohol and 3mL of acetic acid, stirring for 30min, weighing 1.9mmol of 4, 5-dimethyl o-phenylenediamine, adding the mixture into the flask (dissolving the 4mL of ethanol and adding the solution), refluxing for 10min at 80 ℃, cooling to separate out a golden yellow precipitate (Schiff base substance), filtering, and recrystallizing the solid to obtain a compound 3 a.

(4) Preparation of compound 4: 0.4301g (2.4mmol) of 7, 8-benzoquinoline, 0.423g (1.2mmol) of iridium trichloride hydrate, 18mL of ethylene glycol ethyl ether and 6mL of deionized water are taken. Adding all the medicines into a 50mL flask in sequence, introducing nitrogen, heating to 120 ℃, refluxing for 24h, naturally cooling to room temperature after the reaction is finished, pouring 200mL of deionized water into the reaction liquid, stirring to separate out a large amount of yellow precipitates, filtering, washing with water and ethanol, and drying in vacuum at 45 ℃ to obtain yellow solids, namely the compound 4.

(5) Human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]The preparation of (1): 0.1mmol of compound 4, 0.2mmol of compound 3a and 15mL of ethylene glycol are added into a 100mL three-neck flask, and heated and refluxed for 10 hours under the protection of argon to obtain a dark red clear solution. After cooling to room temperature and diluting with 40mL of water, a large amount of red precipitate was formed by adding 30mL of saturated ammonium hexafluorophosphate solution. Suction filtration, washing with water and ether for several times and drying. The dried crude product was dissolved and separated by column chromatography on neutral alumina (200 mesh). Eluting with V (dichloromethane): V (acetonitrile) ═ 2:1 mixed solvent to collect yellow component, and vacuum distilling to remove solvent to obtain human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂](ii) a The yield was about 92%.

Example 3

Human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]The synthetic route of (2) is shown in FIG. 6:

(1) preparation of compound 1: 725mL of concentrated hydrochloric acid (pre-frozen in a refrigerator and 35% by mass concentration) and 14.5g of 8-hydroxyquinoline are carefully added to a 1L round-bottom flask, the temperature is raised to 45 ℃, and 64g of NaClO are added in portions after the 8-hydroxyquinoline is completely dissolved₃(addition was complete over 60 minutes) and after addition stirring was continued for 1h at 45 ℃. After the reaction is finished, diluting the reaction product by using ice water to2L, with CH₂Cl₂(6X 250mL), the organic phases are combined, washed with 3X 200mL of distilled water, the solvent is evaporated off by vacuum rotary evaporation to obtain a yellow solid, the precipitate is filtered, and the solid is recrystallized three times with 40mL of methanol to obtain 6, 7-dichloroquinoline-5, 8-dione, which is recorded as compound 1;

(2) preparation of compound 2: 2.28g (0.01mol) of Compound 1 and 2.73g (0.02mol) of anhydrous potassium carbonate were charged into a 50mL round-bottom flask, 25mL of ethanol was added, stirring was carried out to dissolve Compound 1 completely, 3.24g (0.03mol) of 2-amino-5-methylpyridine was added in portions, and the reaction mixture was refluxed at 85 ℃ for 15 hours. Cooling to room temperature, precipitating, filtering, and recrystallizing with o-dichlorobenzene to obtain compound 2.

(3) Synthesis of compound 3 a: weighing 1mmol of compound 2, adding the compound into a 50mL round-bottom flask, adding 14mL of absolute ethanol and 5mL of acetic acid, stirring for 60min, weighing 2.0mmol of 4, 5-dimethyl o-phenylenediamine into the flask (dissolving the 4mL of ethanol and then adding the solution), refluxing for 10min at 80 ℃ (if the solution is not dissolved, continuously refluxing until the solution is dissolved), cooling to separate out a golden yellow precipitate (namely Schiff base), filtering, and recrystallizing the solid to obtain a compound 3 a.

(4) Synthesis of Compound 4: 0.2151g (1.2mmol) of 7, 8-benzoquinoline, 0.423g (1.2mmol) of iridium trichloride hydrate, 15mL of ethylene glycol ethyl ether and 5mL of deionized water are taken. Adding all the medicines into a 50mL flask in sequence, introducing nitrogen, heating to 150 ℃, refluxing for 36h, naturally cooling to room temperature after the reaction is finished, pouring 200mL of deionized water into the reaction liquid, stirring to separate out a large amount of yellow precipitates, filtering, washing with water and ethanol, and drying in vacuum at 45 ℃ to obtain yellow solids, namely the compound 4.

(5) Human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂]The preparation of (1): 0.1mmol of compound 4, 0.2mmol of compound 3a and 15mL of ethylene glycol are added into a 100mL three-neck flask, and heated and refluxed for 15h under the protection of argon to obtain a dark red clear solution. After cooling to room temperature and diluting with 40mL of water, a large amount of red precipitate was formed by adding 30mL of saturated ammonium hexafluorophosphate solution. Suction filtration, washing with water and ether for several times and drying. The dried crude product was dissolved and separated by column chromatography on neutral alumina (200 mesh). Collecting with mixed solvent of V (dichloromethane) and V (acetonitrile) 2:1Collecting yellow components, and vacuum distilling to remove solvent to obtain human ovarian cancer cell inhibitor [ Ir (3a) (BQ)₂](ii) a The yield was about 90%.

In vitro antitumor Activity test

1. Cell lines and cell cultures

Two human cell strains, namely a human ovarian cancer cisplatin drug-resistant SK-OV-3/DDP cell and a human normal liver HL-7702 cell, are selected for the experiment.

All human cell lines were cultured in RPMI-1640 medium containing 100U/mL penicillin, 10 wt% calf blood, and 100U/mL streptomycin, and placed at 37 deg.C with 5% CO by volume₂Culturing in an incubator.

2. Preparation of test Compounds

Compound 3a, compound 4 and human ovarian cancer cell inhibitor [ Ir (3a) (BQ) are used₂]The purity of the compounds is more than or equal to 95 percent, the DMSO (dimethyl sulfoxide) stock solutions are diluted into a final solution of 20 mu mol/L (the final concentration of DMSO is less than or equal to 1 percent) by using a physiological buffer solution, and the inhibition degree of each compound on the growth of normal cells or selected tumor cells under the concentration is tested.

3. Cell growth inhibition assay (MTT method)

(1) Taking normal cells or tumor cells in a logarithmic growth phase, digesting the cells or tumor cells by trypsin, preparing a cell suspension with the concentration of 5000/mL by using a culture solution containing 10% calf serum, inoculating 190 mu L of the cell suspension into a 96-hole culture plate per hole, and enabling the density of cells to be detected to reach 1000-10000 holes (the edge holes are filled with sterile PBS (phosphate buffered saline);

(2)5％CO₂incubating for 24h at 37 ℃ until a cell monolayer is paved on the bottom of each well, adding 10 mu L of medicine with a certain concentration gradient into each well, and arranging 4 compound wells in each concentration gradient;

(3)5％CO₂incubating at 37 ℃ for 48 hours, and observing under an inverted microscope;

(4) add 10. mu.L of MTT solution (5mg/mL PBS, i.e., 0.5% MTT) to each well and continue culturing for 4 h;

(5) terminating the culture, carefully removing the culture solution in the wells, adding 150 μ L of DMSO into each well to sufficiently dissolve formazan precipitate, mixing uniformly with an oscillator, and measuring the optical density of each well with a microplate reader at a wavelength of 570nm and a reference wavelength of 450 nm;

(6) simultaneously, a zero setting hole (culture medium, MTT, DMSO) and a control hole (cells, culture solution, MTT, a drug dissolving medium with the same concentration, DMSO) are arranged.

(7) The number of living cells was judged from the measured optical density values (OD values), and the larger the OD value, the stronger the cell activity. Using the formula:

calculating the inhibition rate of each compound on the growth of the selected cells, and calculating the IC of each tested compound on each selected cell strain by a Bliss method₅₀Values (in μ M). The results are shown in table 1 below.

TABLE 1 Compound 3a, Compound 4 and Complex [ Ir (3a) (BQ)₂]IC on both cell lines₅₀Value of

IC₅₀The results of the activity screening showed that the inhibitor [ Ir (3a) (BQ)₂]The proliferation inhibition activity of the human ovarian cancer cisplatin resistant SK-OV-3/DDP cell and the human normal liver HL-7702 cell is obviously higher than that of the ligand 3 a. The results of the activity screening experiments show that the inhibitor [ Ir (3a) (BQ)₂]Has high antitumor activity on human ovarian cancer drug-resistant strain SK-OV-3/DDP and IC thereof₅₀The value range is 0.002 +/-0.001 mu M, and the in vitro anti-tumor activity of the compound is far greater than that of 3a ligand and a clinical classical metal-based anti-cancer medicament cisplatin; in addition, the toxicity of the complex 5a-Ir to the normal cell HL-7702 is very small (IC)₅₀More than 50 mu M), shows good target inhibition of the multiplication of human ovarian cancer. In summary, the inhibitor [ Ir (3a) (BQ)₂]Shows good in vivo and in vitro anti-tumor activity and targeting property.