CN113527334B

CN113527334B - Triazole derivative capable of influencing calcium ion channel of tumor cell and preparation method and application thereof

Info

Publication number: CN113527334B
Application number: CN202110236472.7A
Authority: CN
Inventors: 孙秀伟; 刘晓斐; 姚小军; 王文君; 王育伟
Original assignee: Jinan Asia Pharma Tech Co ltd
Current assignee: Jinan Asia Pharma Tech Co ltd
Priority date: 2020-04-13
Filing date: 2021-03-03
Publication date: 2022-04-29
Anticipated expiration: 2041-03-03
Also published as: CN113527334A

Abstract

The invention relates to the technical field of pharmaceutical chemistry synthesis, in particular to a triazole derivative capable of influencing a tumor cell calcium ion channel, and a preparation method and application thereof. According to the invention, dolutegravir is taken as a lead compound, a benzyl group in a dolutegravir structure is changed into a 1,2, 3-triazole structure through a click reaction by utilizing a medicine splicing principle, so that a novel compound is obtained, the growth of tumor cells can be inhibited by influencing a calcium ion channel in the tumor cells, and the application value is obvious.

Description

Triazole derivative capable of influencing calcium ion channel of tumor cell and preparation method and application thereof

Technical Field

The invention relates to the technical field of pharmaceutical chemistry synthesis, in particular to a triazole derivative capable of influencing a tumor cell calcium ion channel, and a preparation method and application thereof.

Background

Dolutegravir is a safe and efficient HIV integrase inhibitor with higher gene barrier and good pharmacokinetic properties, is developed by the company of Kulansu Schken in the United states, and is approved by the food and drug administration FDA to be marketed in 2013. Dolutegravir blocks the strand transfer step of retroviral deoxyribonucleic acid (DNA) integration by binding to the integrase active site. The medicine is only taken once a day, the curative effect of treating patients infected with HIV-1 for the first time is equivalent to the effect of raltegravir taken twice a day, the safety is high, the medicine has strong drug resistance property, and the sales of the dolutegravir in 2020 is expected to reach 26 hundred million dollars as an FDA priority evaluation medicine. The results of preclinical studies show that it is low in toxicity, free of genotoxicity and carcinogenic toxicity, and free of significant fertility toxicity and teratogenic toxicity at doses 27 times greater than clinical doses. Clinical studies show that the therapeutic effect of dolutegravir on initially treated HIV-infected patients and patients who fail current therapy but are not treated by integrase inhibitors is better than that of control drugs; also show good efficacy in adult patients who have failed antiviral treatment and who are resistant to raltegravir and/or eltamivir.

Tumors are a common serious disease endangering human life in the world today, and have become the "second largest killer" leading to human death. Statistically, the number of cancer deaths worldwide will increase to 1315 thousands in 2035 years, but most chemotherapy drugs on the market are relatively adverse, and many patients cannot tolerate the adverse reactions and die. Therefore, the research and development of anti-tumor drugs are very challenging and significant in the life science field at present. Because HIV can reduce the immunity of human bodies and further can cause patients to easily suffer from various tumors, and the current research on dolutegravir mainly aims at resisting HIV, and the dolutegravir can not be found to be applied to the treatment of other diseases.

Chinese patent CN107759607A discloses a triazole diazepine compound with anti-tumor activity and a preparation method thereof, belonging to the technical field of medicine synthesis. The technical essential of this patent is:

the patent has an inhibiting effect on both breast cancer cells MCF-7 and liver cancer cells HepG 2.

Dolutegravir is a HIV integrase inhibitor with a diketonic acid structure and is used for inhibiting HIV integrase through carbonyl and Mg in HIV integrase targets²⁺Interaction mediation to inhibit HIV integrationThe activity of the synthase. Dolutegravir has no inhibition effect on tumor cells, and the inhibition rates IC50 on lung cancer cells H460 and H1975 are both above 100uM, so that dolutegravir is considered to be ineffective; therefore, a diketonic acid structure in the dolutegravir structure can be reserved, and a 1,2, 3-triazole structure is introduced, so that the dolutegravir has anti-tumor activity.

Disclosure of Invention

The invention aims to provide a triazole derivative capable of influencing a calcium ion channel of a tumor cell, which takes dolutegravir as a lead compound, utilizes a drug splicing principle, changes a benzyl group in a dolutegravir structure into a 1,2, 3-triazole structure through a click reaction to obtain a new compound, can inhibit the growth of the tumor cell by influencing the calcium ion channel in the tumor cell, and has a remarkable application value; the invention also provides a preparation method and application thereof.

The technical scheme adopted by the invention for solving the technical problems is as follows:

the triazole derivative with the effect on the calcium ion channel of the tumor cells has the following structural formula:

the preparation method of the triazole derivative with the effect on the calcium ion channel of the tumor cells comprises the following steps:

(1) reacting methyl 4-methoxyacetoacetate with N, N-dimethylformamide dimethyl acetal to obtain methyl (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate; performing cyclization reaction on (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyric acid methyl ester and dimethyl oxalate, and hydrolyzing after oxygen-nitrogen conversion to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester; or

4-methoxy methyl acetoacetate and N, N-dimethyl formamide dimethyl acetal react, directly carry out cyclization reaction with dimethyl oxalate, and hydrolyze after oxygen nitrogen conversion to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester;

(2) condensing 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester with m-aminophenylacetylene to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynyl phenylamino) formamide;

(3) d (+) alpha-methylbenzylamine reacts with ethyl acetoacetate to obtain 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate; under the action of sodium borohydride and a catalyst, 1R-3- (1 '-methylbenzylamine) -2-ethyl crotonate is subjected to chiral catalytic hydrogenation to obtain (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate;

(4) under the action of sodium borohydride and zinc chloride or the action of pinacol borane and lanthanum acetate or the action of lithium aluminum hydride, (1R,2R) -3- (1 '-methylbenzylamine) -2-ethyl butyrate formate is obtained to obtain (1R,2R) -3- (1' -methylbenzylamine) -2-butanol;

(5) (1R,2R) -3- (1' -methylbenzylamine) -2-butanol reacts with S-mandelic acid to obtain R-3-aminobutanol-S-mandelate; or (1R,2R) -3- (1' -methylbenzylamine) -2-butanol reacts with acetic acid to obtain R-3-aminobutanol-acetate;

(6)1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynylphenylamino) carboxamide reacts with R-3-aminobutanol-S-mandelate or R-3-aminobutanol-acetate to give (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide;

or

Reacting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester with 3-aminobutanol under the action of a catalyst to obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid; then carrying out acylation reaction with m-aminophenylacetylene to obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxyl-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynyl phenylamino) formamide;

(7) (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynyl phenylamino) formamide and 2-trifluoromethyl phenyl azide take a click reaction to obtain the triazole derivative influencing the calcium ion channel of the tumor cells.

Wherein:

the method comprises the following steps of: under the protection of nitrogen and at the temperature of 15-25 ℃, adding methyl 4-methoxyacetoacetate into tetrahydrofuran, dropwise adding N, N-dimethylformamide dimethyl acetal, and stirring for reaction after dropwise adding; then heating to 25-35 ℃, and concentrating after the reaction is finished to obtain (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxomethyl butyrate;

adding (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxomethyl butyrate into methanol, stirring at room temperature under the protection of nitrogen until the methyl butyrate is completely dissolved, adding dimethyl oxalate, then cooling to 10-12 ℃, adding metal alkoxide, and refluxing; cooling to 0 ℃ after the raw materials completely react, adjusting the pH value to 5-6, and concentrating in vacuum; adding ethyl acetate into the concentrate, separating out an organic phase, and concentrating; then adding methanol and aminoacetaldehyde dimethyl acetal, stirring, cooling to 5 ℃, and adding anhydrous lithium hydroxide for reaction; after the reaction is finished, keeping the reaction temperature at 0-5 ℃, adjusting the pH value of the reaction solution to 1-2, extracting an organic phase, washing the organic phase with saturated saline solution, and concentrating the organic phase to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester;

or the step (1) is as follows:

under the protection of nitrogen and at the temperature of 15-25 ℃, adding methyl 4-methoxyacetoacetate into tetrahydrofuran, dropwise adding N, N-dimethylformamide dimethyl acetal, and stirring for reaction after dropwise adding; then heating to 25-35 ℃, and concentrating after the reaction is finished to obtain (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxomethyl butyrate;

adding a methanol solution of metal salt into 1, 3-dimethyl-2-imidazolidinone at 0 ℃ under the protection of nitrogen, adding dimethyl oxalate, and stirring; adding methyl (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate, heating to room temperature, adding a methanol solution of a metal salt, and continuing stirring after the addition is finished; adding aminoacetaldehyde dimethyl acetal and diisopropylethylamine, and reacting at room temperature; then concentrating the reaction solution, adding the concentrated reaction solution into ethyl acetate, adjusting the pH value of the reaction solution to be neutral, separating an organic phase, concentrating, and adding the concentrated reaction solution into methanol; adding anhydrous lithium hydroxide under the protection of nitrogen, reacting at 0-5 ℃, adjusting the pH value of the reaction solution to 2-3, extracting an organic phase, washing with saturated saline solution, and concentrating the organic phase to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester;

or the step (1) is as follows:

under the conditions of nitrogen protection and room temperature, dropwise adding N, N-dimethylformamide dimethyl acetal into 4-methoxy methyl acetoacetate for reaction, then adding anhydrous methanol, and stirring until the anhydrous methanol is completely dissolved; adding dimethyl oxalate, then cooling to 10-12 ℃, adding metal alkoxide, and refluxing; TLC monitoring the raw material reaction, cooling to 0 deg.C, adjusting pH to 5-6, concentrating, adding ethyl acetate into the concentrate, and dissolving the concentrate; then adding methanol and aminoacetaldehyde dimethyl acetal, stirring, cooling to 5 ℃, and adding anhydrous lithium hydroxide for reaction; after the reaction is finished, keeping the reaction temperature at 0-5 ℃, adjusting the pH value of the reaction solution to 1-2, extracting an organic phase, washing the organic phase with saturated saline solution, and concentrating the organic phase to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester;

wherein the molar ratio of the methyl 4-methoxyacetoacetate to the N, N-dimethylformamide dimethyl acetal is 1: 1-1.1; the molar ratio of the (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyric acid methyl ester to the oxalic acid dimethyl ester to the metal salt is 1:2-4: 2-3; the molar ratio of the (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyric acid methyl ester to the aminoacetaldehyde dimethyl acetal to the lithium hydroxide is 1:1: 1-4; the metal alkoxide is sodium methoxide, sodium ethoxide or potassium tert-butoxide.

Step 2 is as follows: reacting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester, HATU, DIPEA, m-aminophenylacetylene and N, N-dimethylformamide under the protection of argon at room temperature; adding the reaction solution into water, separating out a solid, performing suction filtration and drying to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-diformic acid pyridine-2-methyl formate-5- (3-alkynyl phenylamino) formamide;

wherein the molar ratio of the 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester to the HATU to the DIPEA to the m-aminophenylacetylene is 1:1:2: 2.

Step three, step (3) is as follows:

adding D (+) alpha-methylbenzylamine and ethyl acetoacetate into tetrahydrofuran, carrying out reflux reaction under the protection of nitrogen, completely reacting raw materials, and carrying out vacuum concentration to obtain 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate;

adding sodium borohydride and anhydrous metal chloride salt into methyl tert-butyl ether, and stirring at room temperature to obtain a solution containing the methyl tert-butyl ether; adding L-proline and 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate into tetrahydrofuran, and stirring at room temperature to obtain a solution containing tetrahydrofuran; then, under the protection of nitrogen, dropwise adding a tetrahydrofuran-containing solution into a methyl tert-butyl ether-containing solution at 0-10 ℃, adding citric acid in batches under the protection of nitrogen, and keeping the temperature and stirring after the addition is finished; after the raw materials are reacted completely, cooling to below-10 ℃, dripping water, then adjusting the pH value of the reaction solution to be neutral, separating an organic phase, washing, drying, performing suction filtration, and evaporating to remove the methyl tert-butyl ether; then adding acetone and n-hexane, dropwise adding a formic acid solution, separating out solids, cooling to 0 ℃, and performing suction filtration to obtain (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate;

wherein the molar ratio of the D (+) alpha-methylbenzylamine to the ethyl acetoacetate is 1: 1-1.1; the anhydrous metal chloride salt is cerium trichloride, calcium chloride or lithium chloride; the molar ratio of the 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate, sodium borohydride, metal chloride salt and L-proline is 1:1.5-2.5:0.5-1: 1; the molar ratio of the 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate to the citric acid is 1: 1;

or the step (3) is as follows:

adding D (+) alpha-methylbenzylamine and ethyl acetoacetate into toluene, carrying out reflux reaction under the protection of nitrogen, completely reacting raw materials, and evaporating toluene and unreacted ethyl acetoacetate to obtain 1R-3- (1' -methylbenzylamine) -2-butenoic acid ethyl ester;

adding sodium borohydride, copper chloride and (R, R) -Quinox P into a mixed solution of tetrahydrofuran and acetonitrile, keeping the temperature between-5 ℃ and 0 ℃, and stirring; under the protection of nitrogen, dropwise adding 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate, adding citric acid in batches, and reacting; TLC monitors the raw material to react completely, then the temperature is reduced to below-10 ℃, water is added dropwise, then the pH value of the reaction solution is adjusted to be neutral, and the organic solvent is removed by evaporation; adding ethyl acetate, separating an organic phase, washing, drying, performing suction filtration, and evaporating to remove ethyl acetate; then adding acetone and n-hexane, dropwise adding a formic acid solution, separating out solids, cooling to 0 ℃, and performing suction filtration to obtain (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate;

wherein the molar ratio of the D (+) alpha-methylbenzylamine to the ethyl acetoacetate is 1: 1-1.1; the molar ratio of the 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate, the sodium borohydride, the copper chloride to the (R, R) -Quinox P is 1:1.5-2.5:0.1-0.5: 0.1; the molar ratio of the 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate to the citric acid is 1:1.

Step four, step (4) is as follows:

adding NaBH into tetrahydrofuran at 0-10 deg.C under nitrogen protection₄And zinc chloride, then dripping a tetrahydrofuran solution dissolved with (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate, controlling the temperature to be 5-10 ℃, and heating to room temperature after dripping is finished; after the raw materials completely react, pouring the reaction solution into ice water, adding methyl tert-butyl ether, separating out an upper organic phase, washing, drying, performing suction filtration, and concentrating the filtrate to obtain (1R,2R) -3- (1' -methylbenzylamine) -2-butanol;

wherein the molar ratio of the (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate to the sodium borohydride to the zinc chloride is 1:4: 2;

or the step (4) is as follows:

adding (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate and pinacolborane into a solvent, adding the solvent containing lanthanum acetate at room temperature under the protection of nitrogen, and heating to 40-60 ℃ for reaction; TLC monitors that the raw materials completely react, reaction liquid is filtered, dichloromethane is added after concentration, washing is carried out, an organic phase is separated, and (1R,2R) -3- (1' -methylbenzylamine) -2-butanol is obtained after concentration;

wherein the molar ratio of the (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate to the pinacol borane is 1: 2-3; the solvent is n-pentane or toluene;

or the step (4) is as follows:

under the conditions of-30 to 0 ℃ and argon protection, adding lithium aluminum hydride into tetrahydrofuran, then dropwise adding a tetrahydrofuran solution dissolved with (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate, and reacting until the raw materials disappear; pouring the reaction solution into ice water, adding methyl tert-butyl ether, separating out an upper organic phase, washing, drying, performing suction filtration, and concentrating the filtrate to obtain (1R,2R) -3- (1' -methylbenzylamine) -2-butanol;

wherein the molar ratio of the (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate to the lithium aluminum hydride is 1: 2-5.

Step five, the step (5) is as follows: adding 25wt.% of ammonium formate aqueous solution into (1R,2R) -3- (1' -methylbenzylamine) -2-butanol and absolute ethanol, adding 10wt.% of Pd/C under the protection of nitrogen, and heating to 55-65 ℃; the raw materials react completely, Pd/C is removed by cooling and suction filtration, and ethanol is evaporated to obtain free R-3-aminobutanol; cooling the reaction temperature to 8-12 ℃, adding isopropanol, adding S-mandelic acid or acetic acid in batches, continuing stirring after the addition is finished, and separating out solids; evaporating to remove isopropanol, taking away residual water in a reaction system, adding isopropanol, heating to reflux and dissolve, cooling to room temperature after dissolving, separating out solids, performing suction filtration, washing a filter cake, and drying to obtain R-3-aminobutanol-S-mandelate or R-3-aminobutanol-S-acetate;

wherein the mass ratio of the (1R,2R) -3- (1 ' -methylbenzylamine) -2-butanol to 25wt.% of the ammonium formate aqueous solution is 1:4-5, the mass ratio of the (1R,2R) -3- (1 ' -methylbenzylamine) -2-butanol to 10wt.% of Pd/C is 5:1-2, and the molar ratio of the (1R,2R) -3- (1 ' -methylbenzylamine) -2-butanol to S-mandelic acid or acetic acid is 1: 1-1.2.

Step six, step (6) is as follows:

adding 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynyl phenylamino) formamide into anhydrous formic acid, heating to 60-70 ℃ under the protection of argon, reacting, concentrating, evaporating formic acid, adding acetonitrile, stirring to dissolve, adding R-3-aminobutanol-S-mandelate or R-3-aminobutanol-acetate, heating to reflux, concentrating after the reaction is finished, evaporating solvent, adding dichloromethane and water, separating out an organic phase, washing, and concentrating to obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide;

wherein the molar ratio of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynylphenylamino) formamide, R-3-aminobutanol-S-mandelate or R-3-aminobutanol-acetate is 1: 1-2.

Or the step (6) is as follows:

adding 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester into anhydrous formic acid, heating to reflux under the protection of nitrogen, concentrating to remove formic acid after the reaction is finished, adding acetonitrile, and stirring and dissolving under the protection of nitrogen to obtain an acetonitrile solution; adding 3-aminobutanol, (R, R) -Quinox P and bis (triphenylphosphine) copper nitrate (I) into tetrahydrofuran to obtain a solution containing tetrahydrofuran; under the protection of nitrogen, dropwise adding a solution containing acetonitrile into a solution containing tetrahydrofuran, and after dropwise adding, carrying out reflux reaction; TLC detects the raw material to completely react, the reaction liquid is filtered, the solvent is concentrated and evaporated, dichloromethane and water are added, the pH value of the reaction liquid is adjusted to 1-2, dichloromethane and water are added, the lower organic phase is separated, washed and concentrated to obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid;

adding (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid into N, N-dimethylformamide, and stirring at room temperature under the protection of nitrogen to completely dissolve; adding HATU, controlling the temperature not to exceed 25 ℃, and dropwise adding DIPEA; dissolving m-aminophenylacetylene in N, N-dimethylformamide, adding the N, N-dimethylformamide into a reaction system, and reacting at 25-35 ℃; TLC shows that the reaction is finished, water is added, pulping, suction filtration and drying of a filter cake are carried out to obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxyl-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynyl phenylamino) formamide;

wherein the molar ratio of the 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester, 3-aminobutanol, (R, R) -Quinox P and bis (triphenylphosphine) copper nitrate (I) is 1:1-1.4:0.1: 0.1;

the molar ratio of (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid, HATU, DIPEA and m-aminophenylacetylene is 1:1-1.1:1-1.1: 2.

Seventhly, the step (7) is as follows:

mixing 2-trifluoromethylphenyl azide, (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide, tert-butyl alcohol, water, tetrahydrofuran, copper sulfate pentahydrate and sodium ascorbate, and reacting at 60-80 ℃; the raw materials are reacted completely, dichloromethane is added, and reaction liquid is filtered to obtain yellow liquid; separating out an organic phase, drying, evaporating to remove the solvent to obtain a yellow solid, and recrystallizing with ethanol to obtain the triazole derivative influencing the calcium ion channel of the tumor cells.

The structural formula of the (R, R) -QuinoxP is shown as

The application of the triazole derivative influencing the calcium ion channel of the tumor cells is to prepare the triazole derivative influencing the calcium ion channel of the tumor cells into an anti-cancer medicament.

The invention has the following beneficial effects:

1. the key intermediate 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester is obtained through the one-pot reaction, so that the reaction route is shortened, the reaction yield is improved, and the method has good industrial prospect.

2. The invention designs a route for synthesizing R-3-aminobutanol-salt, adopts a metal salt catalyst, and improves the yield of chiral intermediates in each step.

3. The invention can directly obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxyl-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid with higher chiral purity under the action of a catalyst, and has high industrial production value.

4. According to the invention, dolutegravir is taken as a lead compound, a benzyl group in a dolutegravir structure is changed into a 1,2, 3-triazole structure through a click reaction by utilizing a medicine splicing principle, so that a novel compound is obtained, the growth of tumor cells can be inhibited by influencing a calcium ion channel in the tumor cells, and the application value is obvious.

Drawings

FIG. 1 is a hydrogen spectrum of triazole derivatives having calcium ion channels affecting tumor cells according to the present invention;

FIG. 2 is a hydrogen spectrum of (1R,2R) -3- (1' -methylbenzylamine) -2-butanol in example 16 of the present invention;

FIG. 3 is a hydrogen spectrum of R-3-aminobutanol-acetate in example 19 of the present invention;

FIG. 4 is a hydrogen spectrum of (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide in example 20 of the present invention.

Detailed Description

The present invention is further described below with reference to examples.

Example 1

Under the protection of nitrogen and at the reaction temperature of 15-25 ℃, adding 30g of methyl 4-methoxyacetoacetate into 100mL of tetrahydrofuran, dropwise adding 24g of N, N-dimethylformamide dimethyl acetal, keeping the temperature at room temperature after dropwise adding, stirring for reaction for 1h, then heating to 35 ℃, and concentrating after the reaction is finished to obtain 37g of methyl (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate, wherein the weight ratio of the methyl (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate is LC-MS: 202[ M + H [ ]]⁺。

Example 2

Adding 20g of (Z) -methyl 2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate into 250mL of anhydrous methanol, and stirring the mixture under the room temperature condition and the nitrogen protection until the mixture is completely dissolved; adding 48g of dimethyl oxalate, then cooling to 10 ℃, adding 12g of sodium methoxide, heating to reflux, cooling to 0 ℃ after the raw materials completely react, dropwise adding 2N hydrochloric acid to adjust the pH value of the reaction solution to 5-6, then concentrating in vacuum, adding 150mL of ethyl acetate into the concentrate, concentrating after separating an organic phase, adding 200mL of anhydrous methanol, adding 11g of aminoacetaldehyde dimethyl acetal, stirring for 2.5 hours, cooling to 5 ℃, adding 7.2g of anhydrous lithium hydroxide, and controlling the reaction to be between 0 and 5 ℃; after reacting for 30min, TLC shows that the raw materials completely react, dilute sulfuric acid solution is dripped to adjust the pH value of reaction liquid to 1-2 while keeping the reaction temperature at 0-5 ℃, 100mL of dichloromethane is added, an organic phase is separated, the water phase is extracted for a plurality of times by dichloromethane, the organic phases are combined and washed once by 20mL of saturated saline solution, and the organic phase is concentrated to obtain 19.4g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester;¹H NMR(400MHz,CDCl₃):δ8.42(s,1H),4.54(t,J₁＝4.0Hz,J₂＝4.0Hz,1H),4.14(d,J＝8.0Hz,2H),4.03(s,6H),3.42(s,6H).

example 3

Adding 20g of (Z) -methyl 2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate into 250mL of anhydrous methanol, and stirring the mixture under the room temperature condition and the nitrogen protection until the mixture is completely dissolved; adding 48g of dimethyl oxalate, then cooling to 10 ℃, adding 15g of sodium ethoxide, heating to reflux, cooling to 0 ℃ after the raw materials completely react, dropwise adding 2N hydrochloric acid to adjust the pH value to 5-6, then carrying out vacuum concentration, then adding 150mL of ethyl acetate into the concentrate, separating out an organic phase, concentrating, adding 200mL of anhydrous methanol, adding 11g of aminoacetaldehyde dimethyl acetal, stirring for 2.5 hours, cooling to 5 ℃, adding 7.2g of anhydrous lithium hydroxide, and controlling the reaction to be between 0 and 5 ℃; after 30min of reaction, 2N hydrochloric acid solution is added dropwise to adjust the pH value to 1-2 while keeping the reaction temperature at 0-5 ℃, 100mL of dichloromethane is added, the organic phase is separated, the aqueous phase is extracted for a plurality of times by dichloromethane, the organic phases are combined, washed once by 20mL of saturated saline solution, and the organic phase is concentrated to obtain 16.6g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester.

Example 4

Adding 20g of (Z) -methyl 2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate into 250mL of anhydrous methanol, and stirring the mixture under the room temperature condition and the nitrogen protection until the mixture is completely dissolved; stirring at room temperature, adding 48g of dimethyl oxalate, cooling to 10 ℃, adding 25g of potassium tert-butoxide, heating to reflux, cooling to 0 ℃ after the raw materials completely react, dropwise adding 2N hydrochloric acid to adjust the pH value of the reaction solution to 5-6, then adding 150mL of ethyl acetate into the concentrate after vacuum concentration, separating out an organic phase and concentrating; then adding into 300mL of anhydrous methanol, adding 11g of aminoacetaldehyde dimethyl acetal, stirring for 2.5h, cooling to 5 ℃, adding 9.6g of anhydrous lithium hydroxide, and controlling the reaction at 0-5 ℃; after reacting for 40min, keeping the reaction temperature at 0-5 ℃, dropwise adding 2N hydrochloric acid solution to adjust the pH value of the reaction solution to 1-2, concentrating the reaction solution, adding 100mL of dichloromethane, separating out an organic phase, extracting the aqueous phase with dichloromethane for multiple times, combining the organic phases, washing the organic phase with 15mL of saturated saline solution once, and concentrating the organic phase to obtain 25.6g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester.

Example 5

Adding 100mL of methanol solution dissolved with 11g of sodium methoxide into 300mL of 1, 3-dimethyl-2-imidazolidinone in a reaction bottle at the temperature of 0 ℃ under the protection of nitrogen, then adding 48g of dimethyl oxalate, stirring for 10min after the addition is finished, then adding 20g of methyl (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxobutyrate, heating to room temperature, stirring for 1h, then adding 50mL of methanol solution dissolved with 5.5g of sodium methoxide, continuing stirring for 1.5h after the dropwise addition is finished, then adding 11g of aminoacetaldehyde dimethyl acetal and 13g of diisopropylethylamine, stirring for reaction for 3.2h at room temperature, then concentrating the reaction solution, adding 200mL of ethyl acetate after the concentration, adjusting the pH of the reaction solution to be neutral by using diluted hydrochloric acid, then separating an organic phase, washing the organic layer by using 5% sodium bicarbonate aqueous solution, and then concentrating, then adding the mixture into 350mL of methanol, adding 9.6g of anhydrous lithium hydroxide under the protection of nitrogen, and controlling the reaction to be between 0 and 5 ℃; after reacting for 30min, keeping the reaction temperature at 0-5 ℃, dropwise adding glacial acetic acid to adjust the pH value of the reaction solution to 2-3, concentrating the reaction solution, adding 100mL of dichloromethane, separating an organic phase, extracting an aqueous phase with dichloromethane for multiple times, combining the organic phases, washing the organic phase with 20mL of saturated saline solution once, and concentrating the organic phase to obtain 27.9g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester.

Example 6

Under the conditions of nitrogen protection and room temperature, slowly dripping 12g of N, N-dimethylformamide dimethyl acetal into 15g of 4-methoxy methyl acetoacetate, stirring and reacting for 1h after dripping is finished, then adding 250mL of anhydrous methanol, adding 48g of dimethyl oxalate under the conditions of room temperature and nitrogen protection, then cooling to 10 ℃, adding 12g of sodium methoxide in batches, slowly heating to reflux, cooling to 0 ℃ after the raw materials completely react, dripping 2N hydrochloric acid to adjust the pH value of a reaction solution to be 5-6, then carrying out vacuum concentration, then adding 150mL of ethyl acetate into the concentrate, and dissolving the concentrate under the stirring condition; then adding into 300mL of anhydrous methanol, adding 11g of aminoacetaldehyde dimethyl acetal, stirring for 2h, then cooling to 5 ℃, adding 7.2g of anhydrous lithium hydroxide, and controlling the reaction at 0-5 ℃; after reacting for 40min, keeping the reaction temperature at 0-5 ℃, dropwise adding 2N hydrochloric acid solution to adjust the pH value of the reaction solution to 1-2, concentrating the reaction solution, adding 100mL of dichloromethane, separating out an organic phase, extracting the aqueous phase with dichloromethane for multiple times, combining the organic phases, washing with 20mL of saturated saline solution once, and concentrating the organic phase to obtain 29.1g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester.

Example 7

Adding 5g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester, 6g of HATU, 4.1g of DIPEA, 3.7g of m-aminophenylacetylene and 50mL of N, N-dimethylformamide into a reaction bottle at room temperature, and stirring for reaction at room temperature for 12 hours under the protection of argon; the reaction mixture was added to 100mL of water, and a large amount of solid was precipitated, which was filtered and dried to obtain 5.73g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynylphenylamino) carboxamide, LC-MS (ESI): 415[ M + H]⁺。

Example 8

Adding 12g of D (+) alpha-methylbenzylamine and 15g of ethyl acetoacetate into 100mL of toluene in a reaction bottle, heating to reflux under the protection of nitrogen, and completely reacting the raw materials; vacuum concentration to obtain 1R-17g of ethyl 3- (1' -methylbenzylamine) -2-butenoate;¹H NMR(400 MHz,CDCl₃):δ8.97(s,1H),7.33(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),7.24(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),4.66-4.61(m,1H),4.47(s,1H),4.17-4.09(m,2H),1.76(s,3H),1.52(d,J＝4.0Hz,3H),1.27(t,J₁＝4.0Hz,J₂＝4.0Hz,5H).

example 9

Adding 12g of D (+) alpha-methylbenzylamine and 15g of ethyl acetoacetate into 100mL of tetrahydrofuran in a reaction bottle, heating to reflux under the protection of nitrogen, and completely reacting the raw materials; vacuum concentrating to obtain 21g of 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate;¹H NMR(400MHz,CDCl₃):δ8.97(s,1H),7.33(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),7.24(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),4.66-4.61(m,1H),4.47(s,1H),4.17-4.09(m,2H),1.76(s,3H),1.52(d,J＝4.0Hz,3H),1.27(t,J₁＝4.0Hz,J₂＝4.0Hz,5H).

example 10

In a reaction bottle, adding 7.6g of sodium borohydride and 13g of anhydrous cerous chloride into 100mL of methyl tert-butyl ether, and stirring for 30min at room temperature to obtain a solution containing the methyl tert-butyl ether; adding 11g of L-proline and 24g of 1R-3- (1' -methylbenzylamine) -2-butenoic acid ethyl ester into 200mL of tetrahydrofuran at room temperature, and stirring for 20min at room temperature to obtain a solution containing tetrahydrofuran; slowly dripping the obtained tetrahydrofuran-containing solution into a methyl tert-butyl ether-containing solution at 0 ℃ under the protection of nitrogen, keeping the temperature at 0-10 ℃, adding 20g of citric acid in batches under the protection of nitrogen, stirring for 1.5h under heat preservation after dripping, cooling to below-10 ℃ after the raw materials completely react, slowly dripping 100mL of water, keeping the temperatureThe temperature is not more than 0 ℃, saturated sodium hydroxide solution is dripped to adjust the pH value of the reaction solution to be neutral, the reaction solution is filtered, and then an organic phase is separated; the organic phase was washed twice with 20mL of water, once with 10mL of saturated NaCl solution and finally with anhydrous MgSO₄Drying, filtering, concentrating to obtain oily liquid (81% of ethyl [ (1R,2R) -3- (1 '-methylbenzylamine) -2-butyrate) and 19% of ethyl (1R,2S) -3- (1' -methylbenzylamine) -2-butyrate)]Adding the obtained oily liquid into a mixed solution of 70mL of acetone and 140mL of n-hexane, uniformly stirring, slowly dropwise adding a formic acid solution, separating out a large amount of solids, cooling to 0 ℃, stirring for 1h, and performing suction filtration to obtain 19.1g of (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate, wherein the chiral purity reaches 96.4%;¹H NMR(400MHz,CDCl₃):δ7.31(s,4H),7.23-7.22(m,1H),4.13(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),3.88(dd,J₁＝4.0Hz,J₂＝4.0Hz,1H),3.01-2.95(m,1H),2.43(t,J₁＝4.0Hz,J₂＝4.0Hz,1H),2.41-2.34(m,1H),1.58(s,1H),1.32(s,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

example 11

In a reaction bottle, adding 7.6g of sodium borohydride and 6g of anhydrous calcium chloride into 100mL of methyl tert-butyl ether, and stirring for 30min at room temperature to obtain a solution containing the methyl tert-butyl ether; adding 11g of L-proline and 24g of 1R-3- (1' -methylbenzylamine) -2-butenoic acid ethyl ester into 200mL of tetrahydrofuran at room temperature, and stirring for 20min at room temperature to obtain a solution containing tetrahydrofuran; then slowly dripping a tetrahydrofuran-containing solution into a methyl tert-butyl ether-containing solution at 0 ℃ under the protection of nitrogen, keeping the temperature at 0-10 ℃, adding 20g of citric acid in batches under the protection of nitrogen, stirring for 1.5 hours under heat preservation after dripping is finished, cooling to below-10 ℃ after the raw materials completely react, slowly dripping 100mL of water, keeping the temperature not more than 0 ℃, dripping a saturated sodium hydroxide solution to adjust the pH value of a reaction solution to be neutral, filtering the reaction solution, and separating out an organic phase; washing with 20mL of waterThe organic phase was washed twice, once with 10mL of saturated NaCl solution and finally with anhydrous MgSO₄Drying, filtering, concentrating to obtain oily liquid (78% of (1R,2R) -3- (1 '-methylbenzylamine) -2-ethyl butyrate) and 22% of (1R,2S) -3- (1' -methylbenzylamine) -2-ethyl butyrate]Adding the obtained oily liquid into a mixed solution of 70mL of acetone and 140mL of n-hexane, uniformly stirring, slowly dropwise adding a formic acid solution, separating out a large amount of solids, cooling to 0 ℃, stirring for 1h, and performing suction filtration to obtain 16.7g of (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate, wherein the chiral purity reaches 96.9%;¹H NMR(400MHz,CDCl₃):δ7.31(s,4H),7.23～7.22(m,1H),4.13(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),3.88(dd,J₁＝4.0Hz,J₂＝4.0Hz,1H),3.01-2.95(m,1H),2.43(t,J₁＝4.0Hz,J₂＝4.0Hz,1H),2.41-2.34(m,1H),1.58(s,1H),1.32(s,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

example 12

In a reaction bottle, adding 7.6g of sodium borohydride and 4g of anhydrous lithium chloride into 100mL of methyl tert-butyl ether, and stirring for 30min at room temperature to obtain a solution containing the methyl tert-butyl ether; adding 11g of L-proline and 24g of 1R-3- (1' -methylbenzylamine) -2-butenoic acid ethyl ester into 200mL of tetrahydrofuran at room temperature, and stirring for 20min at room temperature to obtain a solution containing tetrahydrofuran; then slowly dripping a tetrahydrofuran-containing solution into a methyl tert-butyl ether-containing solution at 0 ℃ under the protection of nitrogen, keeping the temperature at 0-10 ℃ in the dripping process, adding 20g of citric acid in batches under the protection of nitrogen, keeping the temperature and stirring for 1.5h after dripping is finished, cooling to below-10 ℃ after the raw materials completely react, slowly dripping 100mL of water, keeping the temperature not more than 0 ℃, dripping a saturated sodium hydroxide solution to adjust the pH value of a reaction solution to be neutral, filtering the reaction solution, and separating out an organic phase; the organic phase was washed twice with 20mL of water, once with 10mL of saturated NaCl solution and finally with anhydrous MgSO₄Drying, vacuum filtering, concentrating to obtain oilThe liquid was found to contain 73% of ethyl [ (1R,2R) -3- (1 '-methylbenzylamine) -2-butanoate and 27% of ethyl (1R,2S) -3- (1' -methylbenzylamine) -2-butanoate]Adding the obtained oily liquid into a mixed solution of 70mL of acetone and 140mL of n-hexane, uniformly stirring, slowly dropwise adding a formic acid solution, separating out a large amount of solids, cooling to 0 ℃, stirring for 1h, and performing suction filtration to obtain 14.4g of (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate, wherein the chiral purity reaches 91.5%;¹H NMR(400MHz,CDCl₃):δ7.31(s,4H),7.23-7.22(m,1H),4.13(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),3.88(dd,J₁＝4.0Hz,J₂＝4.0Hz,1H),3.01-2.95(m,1H),2.43(t,J₁＝4.0Hz,J₂＝4.0Hz,1H),2.41-2.34(m,1H),1.58(s,1H),1.32(s,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

example 13

Adding 7.6g of sodium borohydride, 6.7g of copper chloride and 3.3g of (R, R) -Quinoxp into 300mL of a mixed solution of tetrahydrofuran and acetonitrile, stirring, cooling to-5-0 ℃, stirring for 2h, slowly dropwise adding 24g of 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate under the protection of nitrogen, keeping the temperature unchanged, adding 20g of citric acid in batches under the protection of nitrogen, keeping the temperature and stirring for 2h after dropwise adding, and monitoring by TLC that the raw materials are completely reacted; then cooling to below-10 ℃, slowly dropwise adding 120mL of water, keeping the temperature not to exceed-10 ℃, dropwise adding a saturated sodium hydroxide solution to adjust the pH value of the reaction solution to be neutral, filtering the reaction solution, then vacuum-evaporating the organic solvent, adding 200mL of ethyl acetate, and then separating out an organic phase; the organic phase was washed twice with 20mL of water, once with 10mL of saturated sodium chloride solution and finally with anhydrous MgSO₄Drying, vacuum filtering, and concentrating to obtain oily liquid (1R,2R) -3- (1 '-methylbenzylamine) -2-ethyl butyrate 91% and (1R,2S) -3- (1' -methylbenzylamine) -2-ethyl butyrate 9%]Adding the obtained oily liquid into a mixed solution of 70mL of acetone and 140mL of normal hexane, stirring uniformly, slowly dropwise adding a formic acid solution, separating out a large amount of solids, coolingStirring for 1h at 0 ℃, and performing suction filtration to obtain 19.4g of (1R,2R) -3- (1' -methylbenzylamine) -2-ethyl butyrate formate with the chiral purity of 97.3%;¹H NMR(400MHz,CDCl₃):δ7.31(s,4H),7.23-7.22(m,1H),4.13(t,J₁＝4.0Hz,J₂＝8.0Hz,2H),3.88(dd,J₁＝4.0Hz,J₂＝4.0Hz,1H),3.01-2.95(m,1H),2.43(t,J₁＝4.0Hz,J₂＝4.0Hz,1H),2.41-2.34(m,1H),1.58(s,1H),1.32(s,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

example 14

Adding 100mL of tetrahydrofuran into a reaction bottle, adding 14g of sodium borohydride and 26g of zinc chloride under the condition of 0-10 ℃ and under the protection of nitrogen, then slowly dropwise adding 200mL of tetrahydrofuran solution in which 28g of (1R,2R) -3- (1 '-methylbenzylamine) -2-ethyl butyrate formate is dissolved, controlling the internal temperature to be 5-10 ℃, slowly heating to room temperature after dropwise adding, after the raw materials completely react, slowly pouring the reaction solution into 120mL of ice water, adding 100mL of methyl tert-butyl ether, stirring, separating an upper organic phase, washing twice with 20mL of water, washing once with 20mL of saturated sodium chloride solution, drying and suction-filtering 20g of anhydrous magnesium sulfate, concentrating the filtrate to obtain 17.5g of (1R,2R) -3- (1' -methylbenzylamine) -2-butanol (the yield is 95%, chiral purity>99％)。¹H NMR(400MHz,CDCl₃):δ7.33(t,J₁＝4.0Hz,J₂＝4.0Hz,2H),7.25(d,J＝8.0Hz,3H),3.95-3.89(m,2H),3.84-3.81(m,1H),2.96(s,1H),1.80(d,J＝12.0Hz,1H),1.46-1.41(m,1H),1.36(d,J＝4.0Hz,3H),1.10(d,J＝4.0Hz,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

Example 15

In a reaction flask, 28g of (1R,2R) -3- (1' -methylbenzylamine) -2-butyric acid ethyl ester formate and 35g of pinacolborane were added to 250mL of n-pentane under nitrogenUnder the protection of gas, 30mL of n-pentane containing 6.3g of lanthanum acetate is added into the reaction solution at room temperature, the temperature is slowly raised to 40 ℃, the reaction is carried out for 7h, TLC monitors that the raw materials are completely reacted, the reaction solution is filtered, dichloromethane is added after vacuum concentration, then washing is carried out for multiple times by water, an organic phase is separated, and the organic phase is concentrated to obtain 18.1g of (1R,2R) -3- (1' -methylbenzylamine) -2-butanol (with chiral purity)>99％)；¹H NMR(400MHz,CDCl₃):δ7.33(t,J₁＝4.0Hz,J₂＝4.0Hz,2H),7.25(d,J＝8.0Hz,3H),3.95-3.89(m,2H),3.84-3.81(m,1H),2.96(s,1H),1.80(d,J＝12.0Hz,1H),1.46-1.41(m,1H),1.36(d,J＝4.0Hz,3H),1.10(d,J＝4.0Hz,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

Example 16

Adding 28g of (1R,2R) -3- (1 '-methylbenzylamine) -2-ethyl butyrate formate and 35g of pinacol borane into 250mL of toluene in a reaction flask, adding 30mL of toluene solution containing 6.3g of lanthanum acetate into the reaction solution under the condition of room temperature under the protection of nitrogen, slowly heating to 60 ℃, reacting for 4 hours, monitoring the completion of the reaction of raw materials by TLC, filtering the reaction solution, adding dichloromethane after vacuum concentration, washing with water for multiple times, separating out an organic phase, and concentrating the organic phase to obtain 17.4g of (1R,2R) -3- (1' -methylbenzylamine) -2-butanol (with chiral purity)>99％)；¹H NMR(400MHz,CDCl₃):δ7.33(t,J₁＝4.0Hz,J₂＝4.0Hz,2H),7.25(d,J＝8.0Hz,3H),3.95-3.89(m,2H),3.84-3.81(m,1H),2.96(s,1H),1.80(d,J＝12.0Hz,1H),1.46-1.41(m,1H),1.36(d,J＝4.0Hz,3H),1.10(d,J＝4.0Hz,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

Example 17

Adding 200mL of tetrahydrofuran into a reaction bottle, introducing argon for inert gas protection, reducing the temperature to-30 ℃, and adding aluminum hydride20g of lithium, then slowly dropwise adding 100mL of tetrahydrofuran solution dissolved with 28g of (1R,2R) -3- (1 '-methylbenzylamine) -2-ethyl butyrate formate, controlling the internal temperature to be not more than 0 ℃, reacting until the raw materials disappear after dropwise adding, slowly pouring the reaction solution into 100mL of ice water, adding 100mL of methyl tert-butyl ether, stirring uniformly, separating out an upper organic phase, washing twice with 25mL of water, washing once with 25mL of saturated sodium chloride solution, drying 20g of anhydrous magnesium sulfate, carrying out suction filtration, and concentrating the filtrate to obtain 12.8g of (1R,2R) -3- (1' -methylbenzylamine) -2-butanol (with chiral purity)>99％)；¹H NMR(400MHz,CDCl₃):δ7.33(t,J₁＝4.0Hz,J₂＝4.0Hz,2H),7.25(d,J＝8.0Hz,3H),3.95-3.89(m,2H),3.84-3.81(m,1H),2.96(s,1H),1.80(d,J＝12.0Hz,1H),1.46-1.41(m,1H),1.36(d,J＝4.0Hz,3H),1.10(d,J＝4.0Hz,3H),1.31-1.25(m,3H),1.05(d,J＝4.0Hz,3H).

Example 18

Adding 20g of (1R,2R) -3- (1' -methylbenzylamine) -2-butanol and 200mL of absolute ethanol into a reaction bottle, then adding 100g of 25wt.% ammonium formate aqueous solution, adding 4g of Pd/C with the content of 10wt.% under the protection of nitrogen, stirring for 1h at room temperature, slowly heating to 60 ℃ for 3h, keeping the temperature for completely reacting the raw materials, cooling, carrying out suction filtration to remove Pd/C, and carrying out reduced pressure distillation to remove ethanol to obtain free R-3-aminobutanol; cooling the reaction temperature to about 10 ℃, adding 50mL of isopropanol, adding 16.5g of S-mandelic acid in batches, and continuously stirring after adding to gradually separate out solids; and (3) distilling off isopropanol in the reaction liquid under reduced pressure so as to bring away residual water in the reaction system, adding 50mL of isopropanol again, heating to reflux and dissolve, cooling to room temperature after dissolving, separating out a large amount of solids, washing a filter cake with a small amount of isopropanol after suction filtration, and drying the filter cake to obtain 22g of R-3-aminobutanol-S-mandelate (the e.e value is 100%).

Example 19

Adding 20g of (1R,2R) -3- (1' -methylbenzylamine) -2-butanol and 200mL of absolute ethanol into a reaction bottle, then adding 100g of 25wt.% ammonium formate aqueous solution, adding 4g of Pd/C with the content of 10wt.% under the protection of nitrogen, stirring for 2h at room temperature, slowly heating to 60 ℃ for 3h, keeping the temperature for completely reacting the raw materials, cooling, carrying out suction filtration to remove Pd/C, and carrying out reduced pressure distillation to remove ethanol to obtain free R-3-aminobutanol; cooling the reaction temperature to about 10 ℃, adding 50mL of isopropanol, adding 7.2g of acetic acid in batches, continuing stirring after the addition is finished, detecting the pH value of the reaction solution to be 3-4, and gradually precipitating solids in the reaction solution; and (3) distilling off isopropanol in the reaction liquid under reduced pressure so as to bring away residual water in the reaction system, adding 50mL of isopropanol again, heating to reflux and dissolve, cooling to room temperature after dissolving, separating out a large amount of solids, washing a filter cake with a small amount of isopropanol after suction filtration, and drying the filter cake to obtain 12.2g of R-3-aminobutanol-acetate (chiral purity is 100%).

Example 20

Adding 4g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynyl phenylamino) formamide into 20mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after reacting for about 4.5h, concentrating in vacuum and removing formic acid by rotary evaporation, adding 40mL of acetonitrile, stirring to dissolve, adding 2.4g of R-3-aminobutanol-S-mandelate, heating to reflux, and reacting until the raw materials react completely; concentrating, adding 20mL of dichloromethane, adding 10mL of water while stirring, separating a lower organic phase, extracting an upper aqueous phase with 10mL of dichloromethane for three times, combining the organic phases, and washing with 10mL of saturated saline solution for 3 times; then concentrated under vacuum to obtain 3.2g of (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide with chiral purity of 98%.

Example 21

Adding 4g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynyl phenylamino) formamide into 20mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after reacting for about 4.5h, carrying out vacuum concentration to remove formic acid, adding 40mL of acetonitrile, stirring to dissolve, adding 3g of R-3-aminobutanol-acetate, heating to reflux, and reacting until the raw materials react completely; concentrating, adding 20mL of dichloromethane, adding 10mL of water while stirring, separating a lower organic phase, extracting an upper aqueous phase with 10mL of dichloromethane for three times, combining the organic phases, and washing with 10mL of saturated saline solution for 3 times; then concentrated under vacuum to obtain 3.4g of (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide with chiral purity of 99.5%.

Example 22

Adding 4g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-carboxylic acid methyl ester-5- (3-alkynyl phenylamino) formamide into 20mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after reacting for about 4.5h, carrying out vacuum concentration to remove formic acid, adding 40mL of acetonitrile, stirring to dissolve, adding 3.6g of R-3-aminobutanol-S-mandelate and 0.4g of copper chloride, heating to reflux, and reacting until the raw materials react completely; concentrating, adding 20mL of dichloromethane, adding 10mL of water while stirring, separating a lower organic phase, extracting an upper aqueous phase with 10mL of dichloromethane for three times, combining the organic phases, and washing with 10mL of saturated saline solution for 3 times; then concentrated under vacuum to obtain 3.6g of (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide with chiral purity of 99.5%.

Example 23

In a reaction bottle, 3g of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester is added into 20mL of anhydrous formic acid and heated to reflux under the protection of nitrogen and stirring; after the reaction is finished, removing formic acid by vacuum concentration, adding 20mL of acetonitrile, and stirring and dissolving under the protection of nitrogen; adding 1.25g (0.014mol) of 3-aminobutanol, 0.33g of (R, R) -Quinox P and 0.65g of copper (I) bis (triphenylphosphine) nitrate into 20mL of tetrahydrofuran, and stirring for 15 min; then under the protection of nitrogen, dropwise adding an acetonitrile solution into tetrahydrofuran solution, heating the system after dropwise adding until reflux and stirring for reaction for 2 hours, and detecting by TLC that the raw materials completely react; filtering the reaction solution, concentrating, adding 20mL of dichloromethane, adding 10mL of water while stirring, adding 2N hydrochloric acid to adjust the pH value of the reaction solution to 1-2, stirring for 10min, separating a lower organic phase, extracting an upper aqueous phase with 10mL of dichloromethane for three times, combining the organic phases, and washing with 10mL of saturated saline solution for 3 times; then concentrating under vacuum to obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5]Pyrazino [2,1-B ] s][1,3]2.62g of oxazine-9-carboxylic acid, chiral purity 99%;¹H NMR(400 MHz,CDCl₃):δ8.43(s,1H),5.30(t,J₁＝4.0Hz,J₂＝4.0Hz,1H),5.02(t,J₁＝4.0Hz,J₂＝8.0Hz,1H),4.41(dd,J₁＝4.0Hz,J₂＝4.0Hz,1H),4.27(dd,J₁＝8.0Hz,J₂＝4.0Hz,1H),4.08(s,3H),4.03-3.99(m,2H),2.25-2.16(m,1H),1.56(d,J＝12.0Hz,1H),1.39(d,J＝8.0Hz,3H).

example 24

Adding 7.0g of (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid and 70mL of N, N-dimethylformamide into a reaction bottle, and stirring at room temperature under the protection of nitrogen to completely dissolve; adding 8.61g of HATU, strictly controlling the system temperature to be not more than 25 ℃, dropwise adding 5.9g of DIPEA, dissolving 5.4g of m-aminophenylacetylene into 10.0mL of N, N-dimethylformamide at the system temperature of not more than 25 ℃, adding the reaction system, and reacting for 12 hours at the temperature of 25-35 ℃; TLC showed the reaction was complete, 500mL of water was added, slurried thoroughly and filtered with suction, and the filter cake was dried to give a brown powdery solid (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide 8.35 g.

Example 25

1g of 2-trifluoromethylphenyl azide, (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9- (3-alkynylphenylamino) carboxamide, 1g of tert-butyl alcohol, 20mL of water, 20mL of tetrahydrofuran, 0.5g of copper sulfate pentahydrate and 1g of sodium ascorbate are sequentially added into a reaction bottle, the raw materials are reacted for 6 hours at 70 ℃, 50mL of dichloromethane is added, the reaction solution is filtered to obtain yellow liquid, an organic phase is separated out, the aqueous phase is extracted twice with 20mL of dichloromethane, the combined organic phase is dried by anhydrous magnesium sulfate, the solvent is evaporated to obtain a yellow solid, recrystallizing with ethanol to obtain 1.3g of a yellowish triazole derivative product with the effect on a tumor cell calcium ion channel. (where the implementation of example 25 and compound nuclear magnetic characterization tests were performed at the university of south china) 1H NMR (400MHz, DMSO-d6):12.62(s,1H),9.09(s,1H),8.72(s,1H),8.23(s,1H),8.08(d, J ═ 8.0Hz,1H),7.98(t, J1 ═ 8.0Hz, J2 ═ 8.0Hz,1H),7.90(t, J1 ═ 8.0Hz, J2 ═ 8.0Hz,1H),7.84(t, J1 ═ 8.0Hz, J2 ═ 12.0Hz,2H),7.68(d, J ═ 8.0Hz,1H),7.49(t, J1 ═ 8.0, J539 ═ 12.0Hz, 8.0Hz, 7.42H, 7.8, 8, 1H, 8H, 1H, 7.8, 8H, 8H, 1H, 8J-0H, 1H, 7.42 (t, 8H, 1H, 8H, 1H, 9H, 7.0H, 8H, 1H, 8H, 7.9H, 8H, 9H, 7.9H, 9H, 8H, 9H, etc.),62H, etc., 3.85(s,3H),2.01-1.94(m,1H),1.52(d, J ═ 12.0Hz,1H),1.29(d, J ═ 8.0Hz, 3H); 13C NMR (101MHz, DMSO-d6) 173.77,161.71,155.15,152.75,146.22,143.09,139.01,134.23,134.06,131.24,130.81,130.35,129.82,129.28,127.52,124.39,120.87,119.54,117.62,116.41,75.80,61.72,60.20,52.36,44.34,29.17,15.67.

According to the invention, dolutegravir is structurally modified, dolutegravir is taken as a mother nucleus, a triazole derivative capable of influencing a tumor cell calcium ion channel is designed and synthesized through click reaction according to a bioactive substructure splicing principle, and the MTT method is utilized to evaluate the proliferation inhibition activity of a target compound on various tumor cells. See example 26 and example 27 for specific steps.

Example 26

Anti-tumor activity test: from CO₂Lung cancer cells A549, H358, H460, H1975, H1650 and PC-9 cell culture dishes are taken out from the incubator and respectively carried out the following operations: aseptic operation is carried out beside an alcohol lamp, a dish cover is opened, culture solution is sucked out of a waste liquid cylinder, the culture solution in a culture dish is washed twice by 2mL PBS, 0.25 wt.% trypsin is used for digestion, digestion is stopped when observation shows that cell gaps are increased and cells are changed into small circle shapes, a liquid transfer gun is used for blowing the bottom of the culture dish to enable the cells to fall off, the obtained cell suspension is transferred into an aseptic centrifuge tube, a centrifuge is set to be 800r/min and 3min for centrifugation, then supernatant in the centrifugal tube is slowly poured, 2-5mL culture solution is added, and cell counting is carried out under an inverted microscope. According to the counting results, the cells were seeded in a 96-well plate at a cell number of 3000 per well. The 96-well plate was placed at 37 ℃ in 5% CO₂Culturing in an incubator for 24 h.

The drug molecules of the invention are formulated to the desired concentration: 20. mu. mol/L, 10. mu. mol/L, 5. mu. mol/L, 2.5. mu. mol/L. From 5% CO₂The 96-well plate is taken out from the incubator, 100 mu L of drug-containing culture medium is added into each well, and 3 multiple wells are simultaneously arranged for each concentration of drug. As a blank well, an equal volume of the corresponding culture medium was added. Placing it at 37 ℃ and 5% CO₂Culturing for 72h in an incubator. Each drug was tested in triplicate with the same batch of cells at different passage numbers. After 72 hours, 10. mu.L of MTT solution (5 mg/mL) was added to each well in the dark, followed byContinuously adding 5% CO₂Culturing for 4h in an incubator, absorbing supernatant by using a pipette gun, adding 100 mu L DMSO into each hole, placing a shaking table for 10min to mix uniformly, measuring the absorbance OD value of the mixture at 490nm by using a microplate reader, and calculating the cell proliferation inhibition rate by the following method: inhibition rate of cell proliferation [ OD ]_Control-OD_{Experiment of}]/OD_ControlX is 100%; the tested target compound has the inhibition rate IC on various lung cancer cells₅₀Respectively, A549(7.46umol/L), H358(3.53umol/L), H460(5.84umol/L), H1975(1.71umol/L), H1650(2.92umol/L) and PC-9(2.97 umol/L). Therefore, the drug molecules have good inhibition effect on mutant and wild lung cancer cells.

Example 27

In order to verify the antitumor activity mechanism of the drug molecule, a calcium ion channel experiment is carried out: from 5% CO₂The lung cancer cell H1975 cell culture dish is taken out from the incubator and respectively subjected to the following operations: aseptic operation is carried out beside an alcohol lamp, a dish cover is opened, culture solution is sucked out of a waste liquid cylinder, the culture solution in a culture bottle is washed twice by 2mL PBS, 0.25 wt.% trypsin is used for digestion, digestion is stopped when observation shows that cell gaps are increased and cells are changed into small circle shapes, a liquid transfer gun is used for blowing the bottom of the culture bottle to enable the cells to fall off, the obtained cell suspension is transferred into an aseptic centrifuge tube, a centrifuge is set to be 800r/min and 3min for centrifugation, then supernatant in the centrifuge tube is slowly poured, 2-5mL culture solution is added, and cell counting is carried out under an inverted microscope. According to the counting result, the obtained product is prepared into 1 × 10 culture solution⁵cells/mL of single cell suspension, then seeded in 6-well plates, and 2mL per well. Placing 6-well plate at 37 deg.C and 5% CO₂Culturing in an incubator for 24 h. Adding 10mmol/L drug molecules on the next day, setting 3 holes at the same time, using the holes as the holes of the blank experiment, adding corresponding culture solution with the same volume, and continuing to culture for 24 hours, wherein each drug is subjected to three experiments by using cells of the same batch and different generations. After 24 hours, the cell pellets are prepared in a sterile centrifuge tube in the same way as the apoptosis treatment method. Adding 5 mu M calcium ion fluorescent probe Fluo-310 into the centrifuge tube0 μ L, gently shaken, and incubated at 37 deg.C with 5% CO₂After incubation in an incubator for 30 minutes and staining, the cells were washed twice with PBS and analyzed by flow cytometry. The results show that the level of calcium ions secreted by the cells after the target compound treatment is significantly increased, with a percentage increase of 58.5%. Thus, the drug molecule of the present invention has a large effect on the intracellular calcium ion secretion level, and after treatment, it increases the calcium ion secretion level.

Example 28

Toxicity assay on C8166 cells: 4X 10⁵100 mu L of/mL C8166 cell suspension is mixed with different solutions of the drug to be detected, and 3 repeated holes are arranged. At the same time, a control well containing no drug was placed at 37 ℃ with 5% CO₂Culturing for 3 days, and detecting cytotoxicity by MTT colorimetric method. ELx800 OD value is measured by enzyme-labeling instrument, the measuring wavelength is 595nm, and the reference wavelength is 630 nm. Calculating to obtain CC₅₀The value (50% cytoxic Concentration), i.e. the Concentration of drug that is toxic to 50% of the normal T lymphocyte cell line C8166.

Inhibition assay for HIV-1IIIB Cytopathic (CPE): mix 8X 10⁵mu.L/well of C8166 cells were plated in 96-well cell culture plates containing 100. mu.L/well gradient dilution drug, and 50. mu.L of HIV-1IIIB dilution supernatant, 1300TCID 50/well, was added. 3 repeat wells were provided. And setting a normal cell control hole without the medicine. 3TC is a positive drug control. 37 ℃ and 5% CO₂After 3 days of culture, the formation of syncytia was counted under an inverted microscope (100X). EC (EC)₅₀(50% Effective Concentration) is the Concentration of drug that inhibits syncytia formation by 50%.

Calculating the formula: according to the experimental results, a line drawing is drawn by origin7.5, according to Reed&The Muench method calculates the 50% Effective Concentration (EC) of the sample for inhibiting the virus₅₀) 50% inhibitory cell growth concentration (CC)₅₀) And the Therapeutic index TI value (Therapeutic index) of anti-HIV-1 activity is: TI ═ CC₅₀/EC₅₀. 1. Cell growth survival (%) ═ experimental well OD value/control well OD value × 100; 2. the inhibition rate (%) of HIV-1 cytopathic effect was (number of syncytia in test wells/number of syncytia in control wells) × 100.

The compound obtained in example 25 had CC50 and EC50 of 6.21 and 2.61. mu. mol/L, respectively; TI was 2.38 respectively.

Claims

1. The triazole derivative with the effect on the calcium ion channel of the tumor cells is characterized in that the structural formula of the compound is as follows:

。

2. the preparation method of the triazole derivative capable of affecting the calcium ion channel of the tumor cells according to claim 1, which is characterized by comprising the following steps:

or

3. The preparation method of triazole derivatives having effect on calcium ion channel of tumor cells according to claim 2,

the step (1) is as follows: under the protection of nitrogen and at the temperature of 15-25 ℃, adding methyl 4-methoxyacetoacetate into tetrahydrofuran, dropwise adding N, N-dimethylformamide dimethyl acetal, and stirring for reaction after dropwise adding; then heating to 25-35 ℃, and concentrating after the reaction is finished to obtain (Z) -2- ((2, 2-methylamino) methylene) -4-methoxy-3-oxomethyl butyrate;

or the step (1) is as follows:

under the conditions of nitrogen protection and room temperature, dropwise adding N, N-dimethylformamide dimethyl acetal into 4-methoxy methyl acetoacetate for reaction, then adding anhydrous methanol, and stirring until the anhydrous methanol is completely dissolved; adding dimethyl oxalate, then cooling to 10-12 ℃, adding metal alkoxide, and refluxing; cooling to 0 deg.C after the raw materials react completely, adjusting pH to 5-6, concentrating, adding ethyl acetate into the concentrate, and dissolving the concentrate; then adding methanol and aminoacetaldehyde dimethyl acetal, stirring, cooling to 5 ℃, and adding anhydrous lithium hydroxide for reaction; after the reaction is finished, keeping the reaction temperature at 0-5 ℃, adjusting the pH value of the reaction solution to 1-2, extracting an organic phase, washing the organic phase with saturated saline solution, and concentrating the organic phase to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester;

4. The preparation method of the triazole derivatives having the effect on the calcium ion channel of tumor cells according to claim 2, wherein the step (2) is as follows: reacting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester, HATU, DIPEA, m-aminophenylacetylene and N, N-dimethylformamide under the protection of argon at room temperature; adding the reaction solution into water, separating out a solid, performing suction filtration and drying to obtain 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-diformic acid pyridine-2-methyl formate-5- (3-alkynyl phenylamino) formamide;

5. The preparation method of triazole derivatives having effect on calcium ion channel of tumor cells according to claim 2,

the step (3) is as follows:

or the step (3) is as follows:

adding D (+) alpha-methylbenzylamine and ethyl acetoacetate into toluene, carrying out reflux reaction under the protection of nitrogen, completely reacting raw materials, and carrying out vacuum concentration to obtain 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate;

wherein the molar ratio of the D (+) alpha-methylbenzylamine to the ethyl acetoacetate is 1: 1-1.1; the molar ratio of the 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate, the sodium borohydride, the copper chloride to the (R, R) -Quinox P is 1:1.5-2.5:0.1-0.5: 0.1; the molar ratio of the 1R-3- (1' -methylbenzylamine) -2-ethyl crotonate to the citric acid is 1: 1;

the structural formula of the (R, R) -QuinoxP is shown as

。

6. The preparation method of triazole derivatives having effect on calcium ion channel of tumor cells according to claim 2,

the step (4) is as follows:

or the step (4) is as follows:

7. The preparation method of triazole derivatives having effect on calcium ion channel of tumor cells according to claim 2,

the step (5) is as follows: adding 25wt.% of ammonium formate aqueous solution into (1R,2R) -3- (1' -methylbenzylamine) -2-butanol and absolute ethanol, adding 10wt.% of Pd/C under the protection of nitrogen, and heating to 55-65 ℃; the raw materials react completely, Pd/C is removed by cooling and suction filtration, and ethanol is evaporated to obtain free R-3-aminobutanol; cooling the reaction temperature to 8-12 ℃, adding isopropanol, adding S-mandelic acid or acetic acid in batches, continuing stirring after the addition is finished, and separating out solids; evaporating to remove isopropanol, taking away residual water in a reaction system, adding isopropanol, heating to reflux and dissolve, cooling to room temperature after dissolving, separating out solids, performing suction filtration, washing a filter cake, and drying to obtain R-3-aminobutanol-S-mandelate or R-3-aminobutanol-S-acetate;

8. The preparation method of triazole derivatives having effect on calcium ion channel of tumor cells according to claim 2,

the step (6) is as follows:

9. The preparation method of triazole derivatives having effect on calcium ion channel of tumor cells according to claim 2,

the step (6) is as follows:

adding 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester into anhydrous formic acid, heating to reflux under the protection of nitrogen, concentrating to remove formic acid after the reaction is finished, adding acetonitrile, and stirring and dissolving under the protection of nitrogen to obtain a solution containing the acetonitrile; adding 3-aminobutanol, (R, R) -Quinox P and bis (triphenylphosphine) copper nitrate (I) into tetrahydrofuran to obtain a solution containing tetrahydrofuran; under the protection of nitrogen, dropwise adding a solution containing acetonitrile into a solution containing tetrahydrofuran, and after dropwise adding, carrying out reflux reaction; TLC detects the raw material to completely react, the reaction liquid is filtered, the solvent is concentrated and evaporated, dichloromethane and water are added, the pH value of the reaction liquid is adjusted to 1-2, dichloromethane and water are added, the lower organic phase is separated, washed and concentrated to obtain (4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid;

(4R,12AS) -3,4,6,8,12, 12A-hexahydro-7-methoxy-4-methyl-6, 8-dioxo-2H-pyrido [1',2':4,5] pyrazino [2,1-B ] [1,3] oxazine-9-carboxylic acid, HATU, DIPEA, m-aminophenylacetylene in a molar ratio of 1:1-1.1:1-1.1: 2;

the structural formula of the (R, R) -QuinoxP is shown as

；

The step (7) is as follows:

10. The application of the triazole derivative capable of influencing the calcium ion channel of the tumor cells according to claim 1, which is characterized in that the triazole derivative capable of influencing the calcium ion channel of the tumor cells is used for preparing an anticancer drug.