CN110105402B

CN110105402B - Berberine antitumor platinum (II) complex and synthesis method and application thereof

Info

Publication number: CN110105402B
Application number: CN201910495354.0A
Authority: CN
Inventors: 覃其品; 王振凤; 黄小玲
Original assignee: Yulin Normal University
Current assignee: Yulin Normal University
Priority date: 2019-06-10
Filing date: 2019-06-10
Publication date: 2021-09-07
Anticipated expiration: 2039-06-10
Also published as: CN110105402A

Abstract

The invention relates to a berberine antitumor platinum (II) complex, a synthesis method thereof and application of the complex in preparing antitumor drugs. The synthetic method of the berberine antitumor platinum (II) complex comprises the following steps: (1) dissolving bromo-berberine derivative, potassium halide and dimethyl pyridylamine in a solvent, and reacting at 90 ℃ to obtain a B-TFA ligand; (2) and (2) dissolving the B-TFA ligand prepared in the step (1) and dichloro-bis (dimethyl sulfoxide) platinum (II) in an acetonitrile solution, carrying out coordination reaction for 4-6 hours, and washing and drying a product to obtain the compound. The invention synthesizes the platinum complex by taking the bromo-berberine derivative B-TFA as an active ligand for the first time, shows excellent in-vivo and in-vitro anti-tumor activity and targeting property, has potential medicinal value and is expected to be used for preparing various anti-tumor medicaments. The structural formula of the berberine anti-tumor platinum (II) complex is as follows:

Description

Berberine antitumor platinum (II) complex and synthesis method and application thereof

Technical Field

The invention relates to a platinum high-activity complex and a synthetic method thereof, in particular to a berberine anti-tumor platinum (II) complex and a synthetic method thereof. The invention also relates to the application of the berberine antitumor platinum (II) complex in preparing antitumor drugs.

Background

Since cisplatin was discovered, 4 anticancer drugs of nedaplatin, oxaliplatin, sulplatin and leplatin were listed in succession, but it was still unsatisfactory in improving the therapeutic effect, reducing the toxic and side effects and overcoming the cross-resistance. Scientists and researchers have tried to synthesize a new platinum antitumor complex by performing various structural modifications on platinum, and hopefully, the attempts have led to finding a platinum anticancer drug with higher activity and lower toxicity.

The main chemical component of the fibraurea recisa pierre is fibraurea recisa pierre alkaloid which is mainly distributed in areas such as Guangxi, Yunnan and Guangdong, and is one of the main sources of berberine or berberine. At present, berberine is verified to have the effects of anti-tumor, anti-inflammatory, antibacterial, immunity enhancement, nerve protection, anti-depression activity, blood fat reduction and the like. Hitherto, no related reports exist on the research of synthesizing the metal complex and the action mechanism thereof by taking berberine derivatives as the ligands of the active ingredients of the traditional Chinese medicine. Therefore, designing and searching the berberine anti-tumor rare earth metal complex with high water solubility, high curative effect, low toxic and side effects and strong targeting becomes a problem to be solved at present.

Disclosure of Invention

The first object of the present invention is to provide a berberine anti-tumor platinum (II) complex.

In order to achieve the first object of the present invention, the present invention provides the following technical solutions:

a berberine anti-tumor platinum (II) complex has the following chemical structural formula:

the second purpose of the invention is to provide a synthetic method of a berberine anti-tumor platinum (II) complex.

In order to achieve the second object of the present invention, the present invention provides the following technical solutions:

a synthetic method of a berberine anti-tumor platinum (II) complex comprises the following steps:

(1) dissolving bromo-berberine derivative, potassium halide and dimethyl pyridylamine in a solvent, and reacting at 90 ℃ to obtain a B-TFA ligand;

(2) and (2) dissolving the B-TFA ligand prepared in the step (1) and dichloro-bis (dimethyl sulfoxide) platinum (II) in an acetonitrile solution, carrying out coordination reaction for 4-6 hours, and washing and drying a product to obtain the compound.

Further, the solvent in the step (1) is acetonitrile.

Further, the molar ratio of the bromo-berberine derivative to the potassium halide to the dimethyl pyridylamine is 1:0.8: 1.5-3.2.

Further, the method is characterized in that the reaction time of the step (1) is 12-15 hours.

Further, the reaction temperature of the step (2) is 55-65 ℃.

Further, the volume of the acetonitrile solution in the step (2) is 10-75 mL.

The third purpose of the invention is to provide the application of the berberine antitumor platinum (II) complex in preparing antitumor drugs.

Compared with the prior art, the invention has the beneficial effects that:

1. the invention firstly takes a bromo-berberine derivative B-TFA as an active ligand to synthesize the platinum complex [ Pt (B-TFA) Cl ] Cl (wherein TFA is trifluoroacetic acid), and the synthesis route is simple, the reaction condition is mild, and the yield is high.

2. The invention researches the activity and toxicity experiments of the complex on T-24, SK-OV-3/DDP, A549 tumor cells and normal HL-7702 cells. Experimental results show that the complex has good inhibition effect on human tumor cells, and the IC50 value is 0.1-15.2 mu M; particularly, the compound has better inhibiting effect on human bladder cancer cell T-24, the IC50 values are 0.1 +/-0.1 mu M respectively, and the in vitro anti-tumor activity of the compound is far greater than that of a clinical classical metal-based anti-cancer medicament cisplatin 10.4 +/-1.2 mu M.

3. The berberine anti-tumor platinum (II) complex and the Pt1 complex have little toxicity (IC50 is more than 150 mu M) to normal cells HL-7702, and show good effect of targeted inhibition of proliferation of human bladder cancer cells. In a word, the complex Pt1 shows excellent in-vivo and in-vitro anti-tumor activity and targeting property, has potential medicinal value and is expected to be used for preparing various anti-tumor medicaments.

4. When the berberine antitumor platinum (II) complex is synthesized, the reaction can be carried out only if the mass ratio of the bromo-berberine derivative to the dimethyl pyridylamine is 1: 1.5-3.2; the reaction temperature is 90 ℃, the product is adhered to the bottom and generates carbon chain breakage at the temperature higher than 90 ℃, and the reaction hardly occurs at the temperature lower than 90 ℃.

Drawings

The technical solutions of the present invention are further described in detail with reference to the specific embodiments in the drawings, but the present invention is not limited thereto.

FIG. 1 is an electrospray mass spectrum of B-TFA prepared in example 1 of the present invention;

FIG. 2 is a hydrogen nuclear magnetic resonance spectrum of B-TFA obtained in example 1 of the present invention;

FIG. 3 is a nuclear magnetic resonance carbon spectrum of B-TFA obtained in example 1 of the present invention;

FIG. 4 is a nuclear magnetic resonance fluorine spectrum of B-TFA obtained in example 1 of the present invention;

FIG. 5 is an electrospray mass spectrum of a complex Pt1 prepared in example 1 of the present invention;

FIG. 6 is a NMR spectrum of Pt1 obtained in example 1 of the present invention;

FIG. 7 is a NMR carbon spectrum of Pt1 obtained in example 1 of the present invention;

FIG. 8 shows the nuclear magnetic resonance fluorine spectrum of Pt1 obtained in example 1 of the present invention.

Detailed Description

The raw material bromo-berberine derivative 1 related in the embodiment of the invention is prepared by referring to the existing literature (Huang, Z. -S.; et al. bioorgan. Med. chem.,2008,16, 7582-7591.); in addition, platinum (II) bis (dimethyl sulfoxide) dichloride can be prepared by referring to the existing literature (Al-Allaf, T.A.K.; et Al. Transit. Met. chem.,1998,23:403-406.) which is abbreviated as cis-PtCl₂(DMSO)₂。

Example 1

1.0mol of derivative 1, 0.8mol of potassium iodide and 3.20mol of lutidine are respectively weighed in a high-temperature pressure-resistant tube, 5.0ml of acetonitrile is added, the added solvent is just finished to dissolve the solid, so that the reaction is more complete, and after the reaction is carried out for 12 hours at 90 ℃, yellow solid powder B-TFA ligand is obtained, wherein the yield is 94.0%.

Weighing 1.0mol of ligand B-TFA and 1.0mol of dichloro-bis (dimethyl sulfoxide) platinum (II), placing the ligand B-TFA and 1.0mol of dichloro-bis (dimethyl sulfoxide) platinum (II) into a 100.0mL high-temperature pressure-resistant bottle, adding 25.0mL of acetonitrile solution, carrying out coordination reaction at 60 ℃ for 4.0h, washing with acetonitrile, and drying in vacuum at 45 ℃ to obtain a yellow target product Pt 1. The yield was 90.8%.

The synthetic route of the berberine anti-tumor platinum (II) complex is as follows:

example 2

The difference from example 1 is that ligand B-TFA (1.0mol) and platinum (II) bis (dimethyl sulfoxide) (1.0mol) dichloride are weighed and placed in a 100.0mL high-temperature pressure-resistant bottle, 10.0mL of acetonitrile solution is added, coordination reaction is carried out for 4.0h at 60 ℃, the solution is washed by acetonitrile and dried in vacuum at 45 ℃, and then the yellow target product Pt1 is obtained. The yield was 85.6%.

Example 3

The difference from example 1 is that ligand B-TFA (1.0mol) and platinum (II) bis (dimethyl sulfoxide) (1.0mol) dichloride are weighed and placed in a 100.0mL high-temperature pressure-resistant bottle, 75.0mL of acetonitrile solution is added, coordination reaction is carried out for 4.0h at 60 ℃, the solution is washed by acetonitrile and dried in vacuum at 45 ℃, and then the yellow target product Pt1 is obtained. The yield was 80.0%.

Example 4

The difference from example 1 is that ligand B-TFA (1.0mol) and dichloro-bis (dimethyl sulfoxide) platinum (II) (1.0mol) are weighed and placed in a 100.0mL high-temperature pressure-resistant bottle, 40.0mL of acetonitrile solution is added, coordination reaction is carried out for 4.0h at 60 ℃, the solution is washed by acetonitrile and then dried in vacuum at 45 ℃, and then yellow target product Pt1 is obtained. The yield was 89.1%.

Experimental example 1

To further determine the structure of the ligands and complexes, the structure of the product of example 1 was characterized.

1. Identification of the resulting B-TFA

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

ESI-MS M/z 645.3[ M- (TFA-H) ] +, where M is the molecular weight of Compound B-TFA.

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

¹H NMR(400MHz,CHCl₃-d)δ9.84(s,1H),8.81(d,J＝5.5Hz,2H),8.26-8.20(m,2H),8.20-8.16(m,1H),7.87(dd,J＝4.7,8.3Hz,3H),7.82-7.76(m,1H),7.67(t,J＝6.5Hz,2H),7.36(s,1H),6.85(s,1H),6.11(s,2H),5.01(br t,J＝5.8Hz,2H),4.42(s,4H),4.36(t,J＝6.7Hz,2H),4.06(s,3H),3.29-3.20(m,2H),2.87-2.77(m,2H),1.93-1.82(m,2H),1.65-1.52(m,2H),1.48-1.39(m,2H),1.34-1.29(m,2H),1.23(br s,6H)。

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

¹³C NMR(400MHz,DMSO-d₆)δ158.925-158.561(m,2C)，150.810(s,1C)，150.416-149.847(m,1C)，149.001(s,2C)，147.711(s,1C)，145.275(s,1C)，142.927(s,1C)，137.889-137.473(s,2C)，130.648(s,1C)，126.674(s,1C)，124.990(s,1C)，124.122(s,1C)，123.306(s,1C)，121.687(s,1C)，120.447(s,1C)，120.236(s,1C)，117.399(s,1C)，114.490(s,1C)，108.380(s,1C)，105.426(s,1C)，102.080(s,1C)，74.265(s,1C)，56.981-56.769(m,3C)，55.318(s,1C)，53.926(s,1C)，29.469(s,1C)，28.711–28.390(m,2C)，26.363(s,1C)，25.904(s,1C)，25.226(s,1C)，23.191(s,1C)。

(4) NMR spectrum of fluorine as shown in FIG. 3.

¹⁹F NMR(471MHz,CHCl₃)δ-75.804。

(5) The results of elemental analysis are shown in Table 1.

Table 1 elemental analysis results of Compounds B-TFA and Pt1 in the examples

Thus, the yellow target product was identified as compound B-TFA, having the formula:

2. the obtained complex Pt1 is identified

(1) Electrospray mass spectrometry, the spectrum of which is shown in FIG. 5.

ESI-MS m/z:987.4[M-Cl]⁺Wherein M is the molecular weight of the complex Pt 1.

(2) The NMR spectrum is shown in FIG. 6.

¹H NMR(500MHz,DMSO-d₆)δ9.72(s,1H)，8.92(s,1H)，8.77(d,J＝5.3Hz,2H)，8.31(t,J＝7.2Hz,2H)，8.23–8.09(m,1H)，7.99(s,1H)，7.85(s,2H),7.75(s,1H)，7.67(t,J＝6.6Hz,2H)，7.08(s,1H)，6.18(s,2H)，5.37(s,2H)，4.94(d,J＝31.1Hz,4H)，4.24(t,J＝6.4Hz,2H)，4.04(s,3H)，3.05(s,2H)，2.52(s,2H)，1.90-1.75(m,2H)，1.51(s,2H)，1.40(t,J＝7.9Hz,2H)，1.31–1.20(m,4H)，1.16(d,J＝14.4Hz,4H)。

(3) NMR spectrum of carbon, as shown in FIG. 7.

¹³C NMR(126MHz,DMSO-d₆)δ166.31,158.44,158.19,150.84,150.23,149.43,148.10,145.73,143.29,141.82,137.85,133.44,131.07,127.07,125.85,123.99,123.82,122.10,120.88,120.70,108.89,105.86,102.57,74.72,68.50,64.81,57.53,55.84,40.56,40.39,40.23,40.06,39.89,39.72,39.56,29.93,29.25,29.10,29.03,27.58,26.86,26.31,25.64。

(4) NMR spectrum of the product shown in FIG. 8.

¹⁹F NMR(471MHz,DMSO-d₆)δ-73.52。

(5) The results of elemental analysis are shown in Table 1.

Thus, the resulting yellow target product was identified as complex Pt1, having the formula:

experimental example 2

In order to fully illustrate the application of the berberine antitumor platinum (II) complex Pt1 in pharmacy, in-vivo and in-vitro antitumor activity experiments are carried out on the berberine antitumor platinum (II) complex.

Experiment on proliferation inhibition activity of platinum (II) complex Pt1 on various human tumor cell lines

1. Cell lines and cell cultures

4 human cell strains such as human ovarian cancer cisplatin-resistant SK-OV-3/DDP cells, human bladder cancer T-24 cells, human bladder cancer A549 cells, human normal liver HL-7702 cells and the like 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

The ligands B-TFA and Pt1 used were all 95% pure, their DMSO stock solutions were diluted with physiological buffer to 20. mu. mol/L final solution (DMSO final concentration. ltoreq.1%) at which the degree of inhibition of growth of normal cells or selected tumor cells by each compound was 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, and enabling the density of cells to be detected to reach 1000-10000 holes (the edge holes are filled with sterile PBS);

(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 method₅₀The value is obtained. The results are shown in table 2 below.

TABLE 2 IC of Compounds on various cell lines₅₀Value (μ M)

Slave IC₅₀According to the result of activity screening, the proliferation inhibition activity of the complex Pt1 on three tested human tumor cell strains SK-OV-3/DDP, T-24 and A549 cells is obviously higher than that of cis-Pt (DMSO) of metal salt₂Cl₂And ligands B-TFA and TFA thereof, which embody the synergistic effect of ligand B-TFA and a platinum central atom. Wherein the complex Pt1 has better inhibiting effect on T-24 cells and IC thereof₅₀The value is 0.1 +/-0.1 mu M, and the in vitro anti-tumor activity of the compound is far greater than that of a clinical classical metal-based anti-cancer medicament cisplatin (10.4 +/-1.2 mu M); furthermore, the Pt1 complex has little toxicity (IC) to HL-7702 which is a normal cell₅₀The value is more than 150 mu M), and the compound shows good effect of targeted inhibition of bladder cancer cell proliferation. In a word, the complex Pt1 shows excellent in-vitro anti-tumor activity and targeting selectivity on tumor cells, has potential medicinal value and is expected to be used for preparing various anti-tumor medicaments.

The above description is part of the embodiments of the present invention, and is not intended to limit the present invention. It will be apparent to those skilled in the art that various modifications and variations can be made without departing from the principles of the invention.

Claims

1. A berberine anti-tumor platinum (II) complex has the following chemical structural formula:

2. the use of the berberine anti-tumor platinum (II) complex of claim 1 in the preparation of an anti-tumor medicament for treating ovarian cancer or bladder cancer.