CN114940692A - Compound with anti-lung cancer effect and preparation method and application thereof - Google Patents

Compound with anti-lung cancer effect and preparation method and application thereof Download PDF

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CN114940692A
CN114940692A CN202210688316.9A CN202210688316A CN114940692A CN 114940692 A CN114940692 A CN 114940692A CN 202210688316 A CN202210688316 A CN 202210688316A CN 114940692 A CN114940692 A CN 114940692A
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方波
胡春生
赫佳馨
秦媛媛
谭娜
周新贵
胡馨丹
孟江平
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Chongqing University of Arts and Sciences
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Abstract

The invention discloses a compound with anti-lung cancer effect, a preparation method and application thereof, the compound introduces a ligand containing multiple bonds (double bonds or triple bonds) into cisplatin to form a prodrug, the drug absorption of the cisplatin is improved, the synergistic effect of the cisplatin and the multiple bond ligand is exerted, the effect of treating lung cancer by drugs is enhanced, and the inhibition capability of part of the compound on cisplatin drug-resistant A549 cells is obviously higher than that of the cisplatin.

Description

Compound with anti-lung cancer effect and preparation method and application thereof
Technical Field
The invention belongs to the technical field of medicinal chemistry, and particularly relates to a compound with an anti-lung cancer effect, and a preparation method and application thereof.
Background
The lung cancer is the cancer with the second highest incidence worldwide, and has great threat to human health, and the platinum drugs are very important anti-lung cancer drugs. It is known that platinum drugs exhibit completely different action mechanism and kinetics from conventional organic drugs, i.e., cisplatin forms cross-links with DNA in cell nuclei or causes internal DNA cross-links to inhibit cell division during cell division, thereby achieving anti-tumor effect. Therefore, cisplatin was approved by FDA in 1978 for clinical treatment of various cancers, and it has a very good therapeutic effect on lung cancer, bladder cancer, testicular cancer, early ovarian cancer, head and neck cancer, and breast cancer. However, with the wide application of platinum drugs, the drug resistance and the systemic toxic and side effects of the platinum drugs are not negligible. Moreover, the existing platinum drugs can not well distinguish cancer cells from normal cells, so that the bioavailability is low, and a series of systemic toxicity is induced. Therefore, the development of new platinum drugs is the focus of research today.
In recent years, tetravalent platinum [ pt (iv) -based prodrugs, which are formed by introducing axial ligands on the basis of bivalent platinum [ pt (ii) -based drugs by virtue of their excellent antitumor efficacy, have received much attention, and generally show better antitumor potential than the corresponding bivalent platinum-based drugs. In addition, irreversible inhibitors against EGFR and BTK, which have had little success over the last two decades, have centered on the fact that the structural unsaturation (double and triple bonds) can react with the cysteine of the target protein to form irreversible covalent adducts.
Disclosure of Invention
Aiming at the prior art, the invention provides a compound with an anti-lung cancer effect, a preparation method and application thereof, and aims to solve the problems of drug resistance, systemic toxic and side effects and the like of the existing platinum drugs.
In order to achieve the purpose, the invention adopts the technical scheme that: a compound with anti-lung cancer effect has a structural formula
Figure BDA0003700581840000021
Wherein R is 1 And R 2 Are all electron withdrawing groups.
On the basis of the technical scheme, the invention can be further improved as follows.
Further, R 1 And R 2 Are respectively and independently
Figure BDA0003700581840000022
Further, R 1 And R 2 The same is true.
The invention also provides a preparation method of the compound with the anti-lung cancer effect, which comprises the following steps:
(1) dissolving a compound shown as a formula I in an organic solvent, adding alkali, stirring and reacting for 0.4-0.6 h at room temperature, dropwise adding 1-bromo-2-butyne or 3-bromopropylene, stirring overnight at room temperature, and then sequentially extracting, washing, drying and purifying to obtain a compound shown as a formula II;
Figure BDA0003700581840000023
wherein R is 3 Is composed of
Figure BDA0003700581840000024
(2) Dissolving a compound shown as a formula II in an organic solvent, adding trifluoroacetic acid, stirring at room temperature for reacting for 3-5 hours, and then sequentially carrying out concentration, alkalization, liquid separation, washing and purification treatment to obtain a compound shown as a formula III;
Figure BDA0003700581840000025
(3) dissolving a compound shown as a formula III in an organic solvent, adding cyclopentanedioic anhydride, stirring overnight at room temperature, and then sequentially concentrating, filtering, washing and drying to obtain a compound shown as a formula IV;
Figure BDA0003700581840000031
(4) dissolving a compound shown as a formula IV in an organic solvent, adding 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate and triethylamine, stirring and reacting for 0.4-0.6 h at room temperature, adding dihydroxydichlorodiammineplatinum, stirring and reacting for two days at a dark room temperature, concentrating and purifying to obtain the compound.
Further, in the step (1), the molar ratio of the compound shown in the formula I, alkali, 1-bromo-2-butyne or 3-bromopropylene is 9-11: 12-14: 11-13; the purification in the step (1) is column chromatography, and the volume ratio of the used reagents is 1: 6-20 parts of a mixed solution of ethyl acetate and n-hexane.
Further, the dosage ratio of the compound shown as the formula II in the step (2) to trifluoroacetic acid is 8 mmol: 3-5 mL; the purification in the step (2) is column chromatography, and the volume ratio of the used reagents is 1: 4-9 parts of a mixed solution of ethyl acetate and n-hexane.
Further, the dosage ratio of the compound shown as the formula II in the step (2) to trifluoroacetic acid is 8 mmol: 3-5 mL; the purification in the step (2) is column chromatography, and the volume ratio of the used reagents is 1: 4-9 parts of a mixed solution of ethyl acetate and n-hexane.
Further, in the step (3), the molar ratio of the compound shown as the formula III to the cyclopentanedioic anhydride is 2-4: 3.
further, in the step (4), the molar ratio of the compound shown as the formula IV, 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate, triethylamine and dihydroxydichlorodiammineplatinum is 0.7-0.8: 0.7-0.8: 0.7-0.8: 0.2 to 0.4; in the step (4), the purification is column chromatography, and the volume ratio of the used reagent is 1: 9-99 parts of a mixed solution of methanol and dichloromethane.
Further, dihydroxydiamminedichloroplatinum is prepared by the following steps: the dihydroxydiamminedichloroplatinum is prepared by the following steps: dropwise adding H into cisplatin aqueous solution at room temperature 2 O 2 And stirring for 3-5 h at 50-70 ℃, filtering, recrystallizing the filtrate at 0-4 ℃, filtering, washing and drying to obtain the product.
The invention also provides application of the compound with the anti-lung cancer effect in preparing an anti-lung cancer medicament.
The invention has the beneficial effects that:
1. the invention introduces a ligand containing multiple bonds (double bonds or triple bonds) into cisplatin to form a prodrug, improves the drug absorption of the cisplatin, simultaneously plays the synergistic effect of the cisplatin and the multiple bond ligand, enhances the effect of the drug on treating lung cancer, and has the inhibition capability of part of compounds on cisplatin-resistant A549 cells which is obviously higher than that of the drug cisplatin.
2. The reason for generating the cisplatin resistance is closely related to the detoxification of glutathione containing cysteine, and the introduction of a multiple bond ligand as an axial ligand of a Pt (IV) prodrug is beneficial to enhancing the effect of the drug on target protein, increasing absorption and reducing side effects and enhancing the treatment effect of platinum drugs on solid tumors, particularly lung cancer.
Drawings
FIG. 1 is a graph showing the results of compounds 16 and 28 inhibiting clonogenic lung cancer cells;
FIG. 2 is a graph showing the results of compounds 16, 28 promoting apoptosis;
FIG. 3 is a graph showing the results of compounds 16 and 28 promoting expression of apoptotic proteins;
FIGS. 4 and 5 are graphs showing the results of compounds 16 and 28 in inhibiting the growth of subcutaneous tumors of lung cancer A549 cells
FIG. 6 is a synthetic scheme of the present invention;
FIGS. 7 and 8 are the hydrogen spectrum and the carbon spectrum of the compound 15 prepared by the invention;
FIGS. 9 and 10 are the hydrogen spectrum and the carbon spectrum of compound 16 prepared by the present invention;
FIGS. 11 and 12 show the hydrogen spectrum and carbon spectrum of compound 17 prepared by the present invention;
FIGS. 13 and 14 show the hydrogen spectrum and carbon spectrum of compound 27 prepared by the present invention;
FIGS. 15 and 16 show the hydrogen and carbon spectra of compound 28 prepared according to the invention;
fig. 17 and 18 show the hydrogen spectrum and carbon spectrum of compound 29 prepared by the present invention.
Detailed Description
The following examples are provided to illustrate specific embodiments of the present invention.
Example 1
A preparation method of a compound with an anti-lung cancer effect comprises the following steps:
(1) to a solution of 10.0mmol 2- (BOC-aminomethyl) phenol in anhydrous N, N-dimethylformamide (15mL) at room temperature was added 13.0mmol Cs 2 CO 3 And stirred for 0.5h, 12.0mmol of 1-bromo-2-butyne was added dropwise and stirred at room temperature overnight, the reaction was monitored by LC-MS, followed by quenching with 30mL of water, extraction with 30mL of ethyl acetate 2 times, followed by combining the organic phases and washing with 40mL of water 2 times, 40mL of brine 1 time, followed by drying over 6g of anhydrous sodium sulfate, followed by purification by column chromatography (ethyl acetate/n-hexane (volume ratio: 1: 12, 1.2L)) to give compound 6 (compound 6 in fig. 6) as a white solid;
compound 6 (tert-butyl 2- (but-2-yn-1-yloxy) phenyl) carbonate), 91.6%; MS (ESI, M/z):262[ M + H] + .
(2) Dissolving 8.0mmol of compound 6 in 12mL of dried dichloromethane at room temperature, adding 4mL of trifluoroacetic acid thereto and stirring the reaction at room temperature for 4h, monitoring the reaction by LC-MS, then concentrating the resulting reaction mixture and removing dichloromethane and trifluoroacetic acid, diluting with 30mL of water and 40mL of dichloromethane, then adjusting the pH of the mixture to 8.5 with 1mol/L of sodium hydroxide solution, followed by liquid separation, then washing the resulting organic phase with 30mL of water and 30mL of brine, and then purifying by column chromatography (ethyl acetate/n-hexane (volume ratio: 1: 6, 1.2L)) to obtain a light brown oily product (compound 9 in FIG. 6);
compound 9(2- (but-2-yn-1-yloxy) aniline), 78.4%; MS (ESI, M/z):162[ M + H] + .
(3) Dissolving 3.0mmol of the light brown oily product in 15mL of dried dichloromethane at room temperature, adding 3.0mmol of cyclopentanedioic anhydride thereto and stirring overnight at room temperature, monitoring the reaction by LC-MS, thereafter concentrating the reaction mixture and removing dichloromethane, then adding dichloromethane (5mL) and n-hexane (15mL) to form a suspension, thereafter filtering, washing the cake with 2mL of n-hexane 2 times, followed by vacuum drying to obtain compound 12 (compound 12 in FIG. 6) as a white solid;
compound 12(5- ((2- (but-2-yn-1-yloxy) phenyl) amino) -5-oxopentanoic acid), 808mg, 97.8%, purity 98.7%; 1 H NMR(400MHz,DMSO)δ12.05(s,1H),9.05(s,1H),7.88(d,J＝7.7Hz,1H),7.10-7.04(m,2H),6.91(t,J＝7.5Hz,1H),4.79(s,2H),2.40(t,J＝7.1Hz,2H),2.27(t,J＝7.4Hz,2H),1.83-1.75(m,5H)ppm. 13 C NMR(101MHz,DMSO)δ174.21,170.91,147.99,127.70,124.20,122.71,120.77,112.81,83.83,74.74,56.49,35.17,33.02,20.64,3.20ppm.MS(ESI,m/z):276[M+H] + .
(4) at room temperature, 0.75mmol of compound 12 was dissolved in 2.0mL of dry N, N-dimethylformamide, 0.75mmol of 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU) and 0.75mmol of triethylamine were sequentially added thereto and stirred for 0.5h, then 0.3mmol of dihydroxydiammineplatinum dichloride was added thereto, the reaction flask was wrapped with tinfoil paper in the dark and stirred at room temperature for two days, the reaction was monitored by LC-MS, then N, N-dimethylformamide was removed by evaporation, and the residue was purified by column chromatography (methanol/dichloromethane (volume ratio: 1: 45, 1.0L) to give compound 15 as a white solid (compound 15 in FIG. 6).
15, 295mg, 46.3% of compound, 97.6% of purity. 1 H NMR(400MHz,DMSO)δ8.94(s,2H),7.93(d,J＝7.3Hz,2H),7.10-7.03(m,4H),6.91(t,J＝7.3Hz,2H),6.54(s,6H),4.80(s,4H),2.41(t,J＝6.3Hz,4H),2.29(t,J＝7.1Hz,4H),1.83-1.75(m,10H)ppm. 13 C NMR(101MHz,DMSO)δ180.40,171.19,147.71,127.74,124.04,122.33,120.75,112.73,83.88,74.74,56.47,35.45,35.05,21.73,3.23ppm.MS(ESI,m/z):848[M+H] + .
Wherein, the dihydroxydiammine dichloroplatinum is prepared by the following steps: at room temperature, 6.7mmol of cisplatin was added to 60mL of distilled water, and 100mL of 30% H was added dropwise thereto 2 O 2 Stirring at 60 deg.C for 4 hr, filtering, recrystallizing the filtrate at 2 deg.C, filtering, and adding 3mL H 2 Washing with O for 2 times, and vacuum drying at 60 deg.C. (1.69 g of dihydroxydiamminedichloroplatinum was obtained in a yield of 76%)
Example 2
A preparation method of a compound with an anti-lung cancer effect comprises the following steps:
(1) to a solution of 10.0mmol of 3- (BOC-amino) phenol in anhydrous N, N-dimethylformamide (15mL) at room temperature was added 13.0mmol of Cs 2 CO 3 And stirred for 0.4h, 12.0mmol of 1-bromo-2-butyne was added dropwise and stirred at room temperature overnight, the reaction was monitored by LC-MS, followed by quenching with 30mL of water, extraction with 30mL of ethyl acetate 2 times, then the organic phases were combined and washed with 40mL of water 2 times, 40mL of brine 1 time, dried over 6g of anhydrous sodium sulfate, and then purified by column chromatography (ethyl acetate/n-hexane (volume ratio: 1: 6, 1.2L)) to give compound 7 (compound 7 in fig. 6) as a white solid;
compound 7(3- (but-2-yn-1-yloxy) phenyl) carbonic acid tert-butyl ester 7, 92.8%; MS (ESI, M/z):262[ M + H] + .
(2) Dissolving 8.0mmol of compound 7 in 12mL of dried dichloromethane at room temperature, adding 4mL of trifluoroacetic acid thereto and stirring at room temperature for reaction for 3 hours, monitoring the reaction by LC-MS, then concentrating the resulting reaction mixture and removing dichloromethane and trifluoroacetic acid, diluting with 30mL of water and 40mL of dichloromethane, then adjusting the pH of the mixture to 8 with 1mol/L of sodium hydroxide solution, followed by liquid separation, and then washing the resulting organic phase with 30mL of water and 30mL of brine, and then purifying by column chromatography (ethyl acetate/n-hexane (volume ratio: 1: 4, 1.2L)) to obtain a light brown oily product (compound 10 in FIG. 6);
compound 103- (but-2-yn-1-yloxy) aniline 1083.1%; MS (ESI, M/z):162[ M + H] + .
(3) Dissolving 3.0mmol of the light brown oily product in 15mL of dried dichloromethane at room temperature, adding 3.0mmol of cyclopentanedioic anhydride thereto and stirring overnight at room temperature, monitoring the reaction by LC-MS, concentrating the reaction mixture and removing dichloromethane, adding dichloromethane (5mL) and n-hexane (15mL) to form a suspension, filtering, washing the cake with 2mL of n-hexane 2 times, followed by vacuum drying to obtain compound 13 (compound 13 in FIG. 6) as a white solid;
816mg of the compound 135- ((3- (but-2-yn-1-yloxy) phenyl) amino) -5-oxopentanoic acid, 98.8%; the purity is 96.8 percent. 1 H NMR(400MHz,DMSO)δ12.08(s,1H),9.88(s,1H),7.28-7.14(m,3H),6.65-6.62(m,1H),4.68-4.67(m,2H),2.35-2.24(m,4H),1.83-1.77(m,5H)ppm. 13 C NMR(101MHz,DMSO)δ174.19,170.81,157.68,140.42,129.42,111.90,109.09,105.86,83.46,74.81,55.76,35.44,32.98,20.40,3.20ppm.MS(ESI,m/z):276[M+H] + .
(4) At room temperature, 0.75mmol of compound 13 was dissolved in 2.0mL of dry N, N-dimethylformamide, 0.75mmol of 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU), 0.75mmol of triethylamine were sequentially added thereto and stirred for 0.4h, then 0.3mmol of dihydroxydichlorodiammineplatinum was added thereto, the reaction flask was wrapped with tinfoil paper in the dark and stirred at room temperature for two days, the reaction was monitored by LC-MS, then N, N-dimethylformamide was removed by evaporation, and the residue was purified by column chromatography (methanol/dichloromethane (volume ratio: 1: 9, 1.0L) to give compound 16 as a white solid (compound 16 in FIG. 6).
Compound 16, 358mg, 56.3%, purity 97.2%. 1 H NMR(400MHz,DMSO)δ9.82(s,2H),7.29-7.17(m,6H),6.64-6.55(m,8H),4.68(s,4H),2.35-2.27(m,8H),1.83-1.73(m,10H)ppm. 13 C NMR(101MHz,DMSO)δ180.32,171.14,157.67,140.42,129.40,111.89,109.05,105.83,83.45,74.81,55.76,35.73,35.00,21.54,3.21ppm.MS(ESI,m/z):848[M+H] + .
Wherein, the dihydroxydichlorodiammineplatinum is prepared by the following steps: at room temperature, 6.7mmol of cisplatin was added to 60mL of distilled water, and 100mL of 30% H was added dropwise thereto 2 O 2 Stirring at 50 deg.C for 5 hr, filtering, recrystallizing the filtrate at 0 deg.C, filtering, and adding 3mL H 2 Washing with O for 2 times, and vacuum drying at 50 deg.C.
Example 3
A method for preparing a compound having an anti-lung cancer effect comprises the following steps:
(1) to a solution of 10.0mmol 4- (BOC-amino) phenol in anhydrous N, N-dimethylformamide (15mL) at room temperature was added 13.0mmol Cs 2 CO 3 And stirred for 0.6h, 12.0mmol of 1-bromo-2-butyne was added dropwise and stirred at room temperature overnight, the reaction was monitored by LC-MS and then quenched with 30mL of water,further extracting with 30mL of ethyl acetate for 2 times, then combining the organic phases and washing with 40mL of water for 2 times, 40mL of brine for 1 time, then drying with 6g of anhydrous sodium sulfate, and then purifying by column chromatography (ethyl acetate/n-hexane (volume ratio: 1: 6, 1.2L)) to obtain a white solid compound 8 (compound 8 in FIG. 6);
compound 8(4- (but-2-yn-1-yloxy) phenyl) carbonic acid tert-butyl ester 8, 93.5%; MS (ESI, M/z):262[ M + H] + .
(2) Dissolving 8.0mmol of compound 8 in 12mL of dry dichloromethane at room temperature, adding 4mL of trifluoroacetic acid thereto and stirring at room temperature for reaction for 5 hours, monitoring the reaction by LC-MS, thereafter concentrating the resultant mixture of the reaction and removing dichloromethane and trifluoroacetic acid, further diluting with 30mL of water and 40mL of dichloromethane, thereafter adjusting the pH of the mixture to 9 with 1mol/L of sodium hydroxide solution, followed by liquid separation, further washing the resultant organic phase with 30mL of water and 30mL of brine, and thereafter purifying by column chromatography (ethyl acetate/n-hexane (volume ratio: 1: 4, 1.2L)) to obtain a light brown oily product (compound 11 in FIG. 6);
compound 114- (but-2-yn-1-yloxy) aniline 11, 92.4%; MS (ESI, M/z):162[ M + H] + .
(3) Dissolving 3.0mmol of the light brown oily product in 15mL of dried dichloromethane at room temperature, adding 3.0mmol of cyclopentanedioic anhydride thereto and stirring overnight at room temperature, monitoring the reaction by LC-MS, concentrating the reaction mixture and removing dichloromethane, adding dichloromethane (5mL) and n-hexane (15mL) to form a suspension, filtering, washing the filter cake with 2mL of n-hexane 2 times, and drying under vacuum to give compound 14 (compound 14 in FIG. 6) as a white solid;
the compound 145- ((4- (but-2-yn-1-yloxy) phenyl) amino) -5-oxopentanoic acid 14, 802mg, 97.1% purity 99.1%. 1 H NMR(400MHz,DMSO)δ12.07(s,1H),9.76(s,1H),7.49-7.47(m,2H),6.90-6.87(m,2H),4.67(d,J＝2.3Hz,2H),2.32-2.24(m,4H),1.82-1.75(m,5H)ppm. 13 C NMR(101MHz,DMSO)δ174.21,170.27,153.14,132.92,120.55,114.82,83.37,74.93,55.99,35.27,33.02,20.54,3.17ppm.MS(ESI,m/z):276[M+H] + .
(4) At room temperature, 0.75mmol of compound 14 was dissolved in 2.0mL of dry N, N-dimethylformamide, 0.75mmol of 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU) and 0.75mmol of triethylamine were sequentially added thereto and stirred for 0.6h, then 0.3mmol of dihydroxydichlorodiammineplatinum was added thereto, the reaction flask was wrapped with tinfoil paper in the dark and stirred at room temperature for two days, the reaction was monitored by LC-MS, then N, N-dimethylformamide was removed by evaporation, and the residue was purified by column chromatography (methanol/dichloromethane (volume ratio: 1: 9, 1.0L) to give compound 17 as a white solid (compound 17 in FIG. 6).
Compound 17 (377 mg, 59.2%) and purity 98.5%. 1 H NMR(400MHz,DMSO)δ9.68(s,2H),7.49(d,J＝9.0Hz,4H),6.88(d,J＝9.0Hz,4H),6.54(s,6H),4.67(d,J＝2.3Hz,4H),2.29(q,J＝7.4Hz,8H),1.82-1.73(m,10H)ppm. 13 C NMR(101MHz,DMSO)δ180.34,170.58,153.11,132.91,120.53,114.79,83.35,74.92,55.97,35.56,35.04,21.66,3.16ppm.MS(ESI,m/z):848[M+H] + .
Wherein, the dihydroxydiammine dichloroplatinum is prepared by the following steps: at room temperature, 6.7mmol of cisplatin was added to 60mL of distilled water, and 100mL of 30% H was added dropwise thereto 2 O 2 Stirring at 70 deg.C for 3 hr, filtering, recrystallizing the filtrate at 4 deg.C, filtering, and adding 3mL H 2 Washing with O for 2 times, and vacuum drying at 70 deg.C.
Example 4
A preparation method of a compound with an anti-lung cancer effect comprises the following steps:
(1) the procedure in step (1) of example 1 was repeated except for changing 1-bromo-2-butyne to 3-bromopropene in step (1) of example 1 to give compound 18 in FIG. 6;
compound 18 (tert-butyl 2- (allyloxy) phenyl) carbonate, 92.9%; MS (ESI, M/z) 250[ M + H] + .
(2) The same as in step (2) in example 1, compound 21 in FIG. 6 was obtained;
compound 212- (allyloxy) aniline, 75.8%; MS (ESI, M/z) 150[ M + H] + .
(3) The same as in step (3) in example 1, compound 24 in FIG. 6 was obtained;
the compound 245- ((2- (allyloxy) phenyl) amino) -5-oxopentanoic acid, 779mg, 98.1%, purity: 99.3%. 1 H NMR(400MHz,DMSO)δ12.05(s,1H),8.99(s,1H),7.85(d,J＝7.5Hz,1H),7.06–6.87(m,3H),6.11-6.02(m,1H),5.44–5.24(m,2H),4.62(d,J＝4.3Hz,2H),2.41(t,J＝6.9Hz,2H),2.27(t,J＝7.3Hz,2H),1.83-1.76(m,2H)ppm. 13 C NMR(101MHz,DMSO)δ174.17,170.80,148.90,133.69,127.51,124.42,122.80,120.37,117.35,112.66,68.87,35.19,32.98,20.64ppm.MS(ESI,m/z):264[M+H] + .
(4) The same procedure as in step (4) in example 1 gave compound 27 in FIG. 6.
Compound 27, 238mg, 38.5% pure 97.2%. 1 H NMR(400MHz,DMSO)δ8.88(s,2H),7.90(d,J＝7.5Hz,2H),7.05-7.01(m,4H),6.88(t,J＝7.0Hz,2H),6.53(s,6H),6.10-6.04(m,2H),5.44-5.25(m,4H),4.62(d,J＝4.6Hz,4H),2.41(t,J＝6.8Hz,4H),2.29(t,J＝7.3Hz,4H),1.80-1.75(m,4H)ppm. 13 C NMR(101MHz,DMSO)δ180.41,171.11,148.63,133.69,127.57,124.26,122.39,120.37,117.51,112.60,68.91,35.49,35.06,21.74ppm.MS(ESI,m/z):824[M+H] + .
The preparation method of dihydroxydichlorodiammineplatinum was the same as that in example 1.
Example 5
A preparation method of a compound with an anti-lung cancer effect comprises the following steps:
(1) the same procedures used in step (1) of example 1 were repeated except for changing 1-bromo-2-butyne to 3-bromopropene and 2- (BOC-aminomethyl) phenol to 3- (BOC-amino) phenol in step (1) of example 1 to give compound 19 in FIG. 6;
compound 19(3- (allyloxy) phenyl) tert-butyl carbonate, 96.1%; MS (ESI, M/z) 250[ M + H] + .HRMS(ESI,m/z):[M+Na] + Theoretical value C 14 H 19 NNaO 3 + 272.1257, found 272.1254.
(2) The same as in step (2) in example 1, to give compound 22 in FIG. 6;
compound 223- (allyloxy) aniline 22, 88.2%; MS (ESI, M/z) 150[ M + H] + .HRMS(ESI,m/z):[M+H] + Theoretical value C 9 H 12 NO + 150.0913, found 150.0915.
(3) The same as in step (3) in example 1, compound 25 in FIG. 6 was obtained;
compound 255- ((3- (allyloxy) phenyl) amino) -5-oxopentanoic acid 761mg, 96.3%, purity: 97.2%. 1 H NMR(400MHz,DMSO)δ12.08(s,1H),9.86(s,1H),7.32(s,1H),7.19-7.08(m,2H),6.62-6.60(m,1H),6.07-5.98(m,1H),5.40-5.23(m,2H),4.51-4.50(m,2H),2.35-2.24(m,4H),1.82-1.75(m,2H)ppm. 13 C NMR(101MHz,DMSO)δ174.19,170.81,158.42,140.45,133.72,129.44,117.38,111.50,109.11,105.66,68.07,35.45,32.98,20.40ppm.MS(ESI,m/z):264[M+H] + .
(4) The same procedure as in step (4) in example 1 gave compound 28 in FIG. 6.
Compound 28, 268mg, 43.4%, purity 97.0%. 1 H NMR(400MHz,DMSO)δ9.80(s,2H),7.34(s,2H),7.19-7.10(m,4H),6.62-6.55(m,8H),6.08-5.99(m,2H),5.38(d,J＝17.3Hz,2H),5.25(d,J＝9.7Hz,2H),4.52(s,4H),2.32-2.29(m,8H),1.78-1.75(m,4H)ppm. 13 C NMR(101MHz,DMSO)δ180.33,171.14,158.41,140.45,133.71,129.43,117.38,111.50,109.11,105.62,68.07,35.74,35.00,21.54ppm.MS(ESI,m/z):824[M+H] + .
The preparation method of dihydroxydiamminedichloroplatinum is the same as that of example 1.
Example 6
A preparation method of a compound with an anti-lung cancer effect comprises the following steps:
(1) the same procedures used in step (1) of example 1 were repeated except for changing 1-bromo-2-butyne to 3-bromopropene and 2- (BOC-aminomethyl) phenol to 4- (BOC-amino) phenol in step (1) of example 1 to give compound 20 in FIG. 6;
compound 20(4- (allyloxy) phenyl) tert-butyl carbonate, 93.5%; MS (ESI, M/z) 250[ M + H] + .
(2) The same as in step (2) in example 1, compound 23 in fig. 6 was obtained;
compound 234- (allyloxy) aniline 23, 91.6%; MS (ESI, M/z) 150[ M + H] + .
(3) The same as in step (3) in example 1, compound 26 in fig. 6 is obtained;
compound 265- ((4- (allyloxy) phenyl) amino) -5-oxopentanoic acid, 769mg, 97.3%, purity: 99.8%. 1 H NMR(400MHz,DMSO)δ12.07(s,1H),9.74(s,1H),7.47(d,J＝9.0Hz,2H),6.87(d,J＝9.0Hz,2H),6.07-5.97(m,1H),5.40-5.22(m,2H),4.51(d,J＝5.2Hz,2H),2.32-2.24(m,4H),1.83-1.75(m,2H)ppm. 13 C NMR(101MHz,DMSO)δ174.20,170.21,153.93,133.90,132.61,120.58,117.31,114.63,68.32,35.28,33.02,20.54ppm.MS(ESI,m/z):264[M+H] + .
(4) The same procedure as in step (4) of example 1 gave compound 29 in FIG. 6.
Compound 29, 393mg, 63.5%, purity 99.1%. 1 H NMR(400MHz,DMSO)δ9.67(s,2H),7.48(d,J＝8.9Hz,4H),6.87(d,J＝9.0Hz,4H),6.54(s,6H),6.07-5.97(m,2H),5.40-5.22(m,4H),4.50(d,J＝5.2Hz,4H),2.31-2.26(m,8H),1.80-1.73(m,4H)ppm. 13 C NMR(101MHz,DMSO)δ180.36,170.55,153.91,133.90,132.61,120.58,117.32,114.62,68.32,35.58,35.05,21.68ppm.MS(ESI,m/z):824[M+H] + .
The preparation method of dihydroxydiamminedichloroplatinum is the same as that of example 1.
Experimental example 1
In fig. 6, compounds 15, 16, 17 and compounds 27, 28, 29 inhibit cell proliferation of lung cancer a549, a549R and HCC 827:
lung cancer cells A549, A549R (cis-platinum resistant cells) and HCC827 were seeded at 3000 cells/well in 96-well plates, and the cells were placed in CO 2 Incubator (37 ℃, 5% CO) 2 ) The culture was carried out overnight. Compounds 15, 16, 17 and compounds 27, 28, 29 were added to the experimental group to final concentrations of 0.2, 0.4, 0.8, 1.6, 3.2, 6.4, 12.5, 25, 50 μ M, respectively, and the control group was added to the corresponding volume of DMSO. After continuing to culture for 48 hours, adding 20 mu L of MTT solution into each well, continuing to incubate for 4 hours, sucking out the culture medium, adding 200 mu L of LDMSO reagent, and shaking the culture medium for 10min at 300 rpm; OD value at 570nm was read using a microplate reader (BioTek staining 5), and the cell growth inhibition ratio (1-OD experiment/OD control). times.100% was measured using a Graphpad Prism 8 meterThe IC50 values were calculated and the results are shown in Table 1.
TABLE 1 IC50 values (μ M) for compounds on lung cancer cells
Compound (I) A549 A549R HCC827
15 ＞50 ＞50 ＞50
16 3.17±0.07 2.01±0.28 1.06±0.04
17 1.98±0.05 3.23±0.91 2.76±0.12
27 ＞50 ＞50 ＞50
28 2.36±0.19 1.52±0.04 1.37±0.05
29 2.03±0.19 2.67±0.38 1.72±0.12
Experimental example 2
Compounds 16, 28 in figure 6 inhibit lung cancer cells a549 and a549R clonogenic:
counting A549 and A549R cells, inoculating into 6-well plate at a ratio of 1000 cells per well, inoculating for 24 hr, adding 2 μ M compound 1 (cisplatin), 13, 25, 16 or 28, standing at 37 deg.C and 5% CO 2 Culturing for about 14 days in an incubator; the supernatant was discarded, the cells fixed with 4% paraformaldehyde, stained with crystal violet for 30min, washed 3 times with PBS, 2min each time. The scanner scans the experimental results and performs statistical analysis. The results are shown in fig. 1, and compared with cisplatin and DMSO groups, compounds 16 and 28 had the ability to significantly inhibit the formation of lung cancer cell clones (× P < 0.01), and ligand compound 13 and compound 25 had no significant inhibitory activity on the formation of lung cancer cell clones.
Experimental example 3
Flow cytometry analysis of compounds 16, 28 in figure 6 promote apoptosis:
a549 and A549R cells in logarithmic growth phase are taken, digested, counted and counted at 2X 10 6 Seed/well in 6-well plates, CO 2 Culturing for 24h in an incubator; adding compounds 16 and 28 to make final concentration 2 μ M, setting 3 multiple wells, and culturing for 24 hr; collecting culture medium containing drug, collecting cells, centrifuging at 1000r/min for 5min, discarding supernatant, and adjusting cell concentration to 10 with Binding buffer 6 Per mL; taking 100 μ L of 10 6 Adding 5 mu L of annexin V-FITC and 5 mu L of PI into the cell suspension with the concentration of one cell/mL, and placing for 15min at room temperature in a dark place; staining for 30min in dark place, detecting apoptosis on the machine, and detecting 10000 cells in each sample. The results are shown in FIG. 2 for compounds 16, 28The number of the induced apoptosis is obviously higher than that of the cisplatin group and the control group.
Experimental example 4
Compounds 16, 28 affect apoptosis-related protein expression:
a549 cells were plated in 6cm dishes with CO 2 Culturing in an incubator overnight; the final concentrations of compounds 16, 28 and cisplatin were adjusted to 2. mu.M, and the cells were collected after further culturing for 48 hours. Adding RIPA lysate (containing protease inhibitor) to break cells, centrifuging at 15000rpm for 30min, and collecting supernatant; protein supernatant is quantified by BCA method, and 30 mu g of total protein is loaded for SDS-PAGE electrophoresis; transferring the protein on the PAGE gel to a PVDF membrane by a wet transfer method, then sealing 5% skimmed milk at room temperature for 2h, and washing for 5 times with TBST, each time for 2 min; bax, clear-PARP antibody (1:1000) was incubated overnight at 4 ℃ and washed 5 times with TBST for 2min each; the dilution ratio of the secondary antibody is 1:20000, the incubation is carried out for 1h at room temperature, and TBST is used for washing for 5 times, each time for 2 min; and detecting the target strip by using an LI-COR Odyessey laser scanner and storing the picture. The results are shown in FIG. 3, and after the compounds 16 and 28 are used for treating cells, the expression of apoptosis proteins Bax and cleaned-PARP is up-regulated, and the apoptosis is promoted.
Experimental example 5
Compounds 16, 28 inhibit the growth of subcutaneous tumors of lung cancer a549 cells:
BALB/c-nu female mice aged 4 weeks were housed in SPF-grade animal rooms and inoculated subcutaneously into the upper left forelimb of mice 10 7 A549 cells with tumor size of about 100mm 3 When, mice were divided into: control group, cisplatin 5mg/kg group, compound 165 mg/kg group and compound 285 mg/kg group, and were administered. The administration mode comprises the following steps: administration to the abdominal cavity; administration volume: 100 mu L of the solution; the administration frequency is as follows: 2 days/time; the administration times are as follows: 4 times. And the body weight of the mice was recorded at the same time as the administration. Mice were sacrificed on day 21 after the first dose, tumors were removed for photography and weighed. The experimental results are shown in fig. 4 and 5, the compounds 16 and 28 can inhibit the growth of subcutaneous tumors of A549, have obvious antitumor activity, and have no obvious toxic or side effect as shown by body weight.
While the present invention has been described in detail with reference to the embodiments, it should not be construed as limited to the scope of the patent. Various modifications and changes may be made by those skilled in the art without inventive step within the scope of the appended claims.

Claims (10)

1. A compound with an anti-lung cancer effect is characterized in that the structural formula of the compound is as follows:
Figure FDA0003700581830000011
wherein R is 1 And R 2 Are all electron withdrawing groups.
2. The compound having an anti-lung cancer effect according to claim 1, characterized in that: the R is 1 And R 2 Are respectively and independently
Figure FDA0003700581830000012
3. The compound having an anti-lung cancer effect according to claim 2, characterized in that: the R is 1 And R 2 The same is true.
4. The method for producing a compound having an anti-lung-cancer effect according to any one of claims 1 to 3, comprising the steps of:
(1) dissolving a compound shown as a formula I in an organic solvent, adding alkali, stirring and reacting for 0.4-0.6 h at room temperature, dropwise adding 1-bromo-2-butyne or 3-bromopropylene, stirring overnight at room temperature, and then sequentially extracting, washing, drying and purifying to obtain a compound shown as a formula II;
Figure FDA0003700581830000013
wherein R is 3 Is composed of
Figure FDA0003700581830000014
(2) Dissolving a compound shown as a formula II in an organic solvent, adding trifluoroacetic acid, stirring at room temperature for reacting for 3-5 hours, and then sequentially performing concentration, alkalization, liquid separation, washing and purification treatment to obtain a compound shown as a formula III;
Figure FDA0003700581830000021
(3) dissolving a compound shown as a formula III in an organic solvent, adding cyclopentanedioic anhydride, stirring overnight at room temperature, and then sequentially concentrating, filtering, washing and drying to obtain a compound shown as a formula IV;
Figure FDA0003700581830000022
(4) dissolving a compound shown as a formula IV in an organic solvent, adding 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate and triethylamine, stirring and reacting for 0.4-0.6 h at room temperature, adding dihydroxy dichlorodiammineplatinum, stirring and reacting for two days at room temperature in a dark place, concentrating and purifying to obtain the compound.
5. The method of claim 4, wherein: in the step (1), the molar ratio of the compound shown as the formula I, alkali, 1-bromo-2-butyne or 3-bromopropylene is 9-11: 12-14: 11-13; the purification in the step (1) is column chromatography, and the volume ratio of the used reagents is 1: 6-20 parts of a mixed solution of ethyl acetate and n-hexane.
6. The method of claim 4, wherein: in the step (2), the dosage ratio of the compound shown as the formula II to the trifluoroacetic acid is 8 mmol: 3-5 mL; the purification in the step (2) is column chromatography, and the volume ratio of the used reagents is 1: 4-9 parts of a mixed solution of ethyl acetate and n-hexane.
7. The method of claim 4, wherein: in the step (3), the molar ratio of the compound shown as the formula III to the cyclopentanedioic anhydride is 2-4: 3.
8. the method of manufacturing according to claim 4, characterized in that: in the step (4), the molar ratio of the compound shown as the formula IV, 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate, triethylamine and dihydroxydichlorodiammineplatinum is 0.7-0.8: 0.7-0.8: 0.7-0.8: 0.2 to 0.4; in the step (4), the purification is column chromatography, and the volume ratio of the used reagent is 1: 9-99 parts of a mixed solution of methanol and dichloromethane.
9. The method according to claim 4 or 8, wherein the dihydroxydiamminedichloroplatinum is prepared by the following steps: dropwise adding H into cisplatin aqueous solution at room temperature 2 O 2 And stirring for 3-5 h at 50-70 ℃, filtering, recrystallizing the filtrate at 0-4 ℃, filtering, washing and drying to obtain the product.
10. Use of a compound having an anti-lung cancer effect according to claim 1 for the preparation of an anti-lung cancer medicament.
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CN105622674A (en) * 2016-02-29 2016-06-01 东南大学 Tetravalent platinum complex with bioactive group and preparation method of tetravalent platinum complex
WO2019166033A2 (en) * 2018-03-02 2019-09-06 天津谷堆生物医药科技有限公司 Vitamin c-platinum complex, intermediate thereof, preparation method, pharmaceutical composition and use
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