CN113527332B - Biological activity-possessed caspovir derivative, and preparation method and application thereof - Google Patents
Biological activity-possessed caspovir derivative, and preparation method and application thereof Download PDFInfo
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Abstract
The invention belongs to the technical field of pharmaceutical chemistry synthesisIn particular to a caspovir derivative with bioactivity and a preparation method and application thereof. The structural formula of the caspovir derivative is as follows:
wherein: r 1 Is methyl or hydrogen; r 2 Is methylene, isopropyl, phenyl, benzyl, a nitrogen-containing heterocycle, a sulfur-containing heterocycle or an oxygen-containing heterocycle; r is 3 Is hydrogen, methyl, ethyl, phenyl, benzyl, a nitrogen-containing heterocycle, a sulfur-containing heterocycle or an oxygen-containing heterocycle. The preparation method comprises the steps of deprotecting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester to aldehyde, cyclizing with aminobutanol, and then carrying out acylation reaction with an amino compound; or firstly reacting with an amino compound and then cyclizing with aminopropanol; and finally reacting with azide by click to obtain the target compound. The derivative of the present invention has a proliferation inhibitory activity on human tumor cells.
Description
Technical Field
The invention belongs to the technical field of chemical synthesis of medicines, and particularly relates to a biological activity-possessing caspovir derivative, and a preparation method and application thereof.
Background
Human immunodeficiency disease (HIV) was first discovered in the united states in 1981 to be a lentivirus (lentivirus) that infects cells of the human immune system, a class of retroviruses. The virus destroys the immunological competence of human body, leads to the immune system to lose resistance, thereby leading to various diseases and cancers to survive in the human body, and finally leading to AIDS, namely acquired immunodeficiency syndrome, and no effective radical therapy exists up to now. Highly Active Antiretroviral Therapy (HAART) is a common treatment for aids using a single basic drug in combination with two nucleoside reverse transcriptase inhibitors. The integrase inhibitor exerts anti-HIV activity by inhibiting integration of double-stranded DNA generated by reverse transcription of HIV into host chromosomes, and can be used as a basic drug.
Caspovir is an integrase inhibitor which is safe and efficient, has a high gene barrier and good pharmacokinetic properties, is developed by the company of stegansmith, u.s.a., and blocks the chain transfer step of integration of retrovirus deoxyribonucleic acid (DNA) by binding with the active site of integrase.
Tumor is a common serious disease endangering human life in the world today, and has become the "second leading killer" of human death. Most chemotherapy drugs on the market at present have more adverse reactions, and many patients cannot tolerate the adverse reactions to 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 the Kabauvir is mainly in the aspect of resisting HIV, and the Kabauvir can not be found to be applied to the treatment of other diseases. The invention carries out structural modification on the carbopol, takes the carbopol as a mother nucleus, designs and synthesizes a series of 1,2, 3-triazole derivatives through click reaction according to the splicing principle of a biological activity substructure, and evaluates the proliferation inhibition activity of a target compound on various human tumor cells by utilizing an MTT method.
Disclosure of Invention
The technical problem to be solved by the invention is to provide a biological active caspovir derivative which has strong inhibition activity on the proliferation of various human tumor cells, and the invention also provides a preparation method and application thereof.
The structural formula of the caspovir derivative with biological activity is as follows:
wherein:
R 1 is methyl or hydrogen;
R 2 is methylene, isopropyl, phenyl, benzyl, a nitrogen-containing heterocycle, a sulfur-containing heterocycle or an oxygen-containing heterocycle;
R 3 is hydrogen, methyl, ethyl, phenyl, benzyl, a nitrogen-containing heterocycle, a sulfur-containing heterocycle or an oxygen-containing heterocycle.
A mixture of one or more of A, B, C or D;
the preparation method of the caspovir derivative with biological activity comprises the following steps:
(1)1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester is deprotected to aldehyde, and cyclized with aminopropanol to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid; then carrying out acylation reaction with an amino compound to obtain an acylation product;
or alternatively
Reacting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester with an amino compound, and cyclizing with aminopropanol to obtain an acylation product;
(2) and (3) reacting the acylation product with azide by click to obtain the target compound.
Wherein the content of the first and second substances,
the method comprises the following steps of:
adding 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester into formic acid, heating to 60-70 ℃ under the protection of argon, and reacting for 2-4 h; concentrating, adding acetonitrile, adding a mixture of aminopropanol and magnesium trifluoromethanesulfonate or aminopropanol, and carrying out reflux reaction for 2-40 h; TLC detects the material to react completely, adds methylene dichloride and water after concentrating, adjusts the pH of the reaction solution to 2-3, separates the lower organic phase, concentrates to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid;
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 aminopropanol is 1: 1-2; the aminopropanol is S-1-amino-2-propanol or 1-amino-2-propanol;
6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid is acylated with aminophenylacetylene to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (X) carboxamide;
wherein X is 2-alkynyl phenylamino, 3-alkynyl phenylamino or propargyl; the aminophenylacetylene is 2-aminophenylacetylene, 3-aminophenylacetylene or propargylamine.
It is further preferred that the reaction mixture is,
step (1) is as follows:
stirring and reacting 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid, 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate, N, N-diisopropylethylamine, 3-aminophenylacetylene or 2-aminophenylacetylene, and N, N-dimethylformamide for 11-13h at room temperature; TLC shows that the reaction is finished, the reaction liquid is added into water, solid is separated out, and 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (X) formamide is obtained after suction filtration and drying;
the molar ratio of the 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid to the 3-aminophenylacetylene or the 2-aminophenylacetylene is 1: 1-2;
or step (1) is as follows:
under the protection of nitrogen, 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid is added into methyl tert-butyl ether, adding N, N' -carbonyl diimidazole, heating to 60-80 deg.C, stirring for 2-4h, stopping heating, adding dropwise methyl tert-butyl ether solution containing propargylamine, then reacting for 1-3h at room temperature, adding water, evaporating the solvent under reduced pressure, adding dichloromethane for extraction, and concentrating to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (propargyl) formamide;
the molar ratio of the 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid to propargylamine is 1: 1-3.
Step (1) can also be as follows:
reacting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester, 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate, N, N-diisopropylethylamine, aminophenylacetylene and N, N-dimethylformamide at room temperature under the protection of argon; TLC shows that the reaction is finished, the reaction solution is added into water, solid is separated out, and the 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-diformic acid pyridine-2-methyl formate-5- (X) formamide is dried after suction filtration;
wherein X is 2-alkynyl phenylamino, 3-alkynyl phenylamino or propargyl; the aminophenylacetylene is 2-aminophenylacetylene, 3-aminophenylacetylene or propargylamine; 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 aminophenylacetylene is 1: 1-2;
adding 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-diformic acid pyridine-2-methyl formate-5- (X) formamide into anhydrous formic acid, and heating to 60-70 ℃ under the protection of argon; reacting for 2-4h, vacuum concentrating, adding acetonitrile, stirring for dissolving, adding aminopropanol or a mixture of aminopropanol and calcium chloride or a mixture of aminopropanol and calcium oxide or a mixture of aminopropanol and aluminum sulfate, heating to reflux, reacting for 5-30h, vacuum concentrating, adding dichloromethane, stirring, adding water, separating out a lower organic phase, concentrating, and purifying to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (X) formamide;
wherein X is 2-alkynylanilino, 3-alkynylanilino or propargyl; the molar ratio of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-diformic acid pyridine-2-methyl formate-5- (X) formamide and aminopropanol is 1: 1-2; the aminopropanol is S-1-amino-2-propanol or 1-amino-2-propanol.
Thirdly, the step (2) is carried out according to one of the following steps:
directly carrying out click reaction on the acylation product and azide to obtain a target compound A; demethylating the target compound A to obtain a target compound B; or demethylating the acylation product and then carrying out click reaction with azide to obtain the target compound B.
1. The step (2) is as follows: mixing azide, an acylation product, tert-butyl alcohol, water, tetrahydrofuran, copper sulfate pentahydrate and sodium ascorbate, reacting at 50-70 ℃, monitoring by TLC that raw materials are completely reacted to obtain yellow liquid, adding dichloromethane, filtering, extracting, drying, and evaporating to remove a solvent to obtain a target compound A; reacting the target compound A with anhydrous lithium bromide to obtain a target compound B;
the azide is benzyl azide, 2-bromobenzyl azide, 2-ethylphenyl azide, 3, 5-bis (trifluoromethyl) phenyl azide, 2-fluorophenyl azide, 4-trifluoromethylphenyl azide, 2-methyl-3-nitrophenyl azide, 4-fluorophenyl azide, 3-nitrophenyl azide, 4-ethylphenyl azide, 2-trifluoromethoxyphenyl azide, 4-trifluoromethylbenzyl azide or 2-trifluoromethylphenyl azide.
2. The step (2) can also be as follows: after the acylation product reacts with anhydrous lithium bromide for demethylation, mixing azide, the acylation product after demethylation, tert-butyl alcohol, water, tetrahydrofuran, copper sulfate pentahydrate and sodium ascorbate, reacting at 50-70 ℃, monitoring the complete reaction of raw materials by TLC to obtain yellow liquid, adding dichloromethane, filtering, extracting, drying, and evaporating to remove the solvent to obtain a target compound B;
the azide is benzyl azide, 2-bromobenzyl azide, 2-ethylphenyl azide, 3, 5-bis (trifluoromethyl) phenyl azide, 2-fluorophenyl azide, 4-trifluoromethylphenyl azide, 2-methyl-3-nitrophenyl azide, 4-fluorophenyl azide, 3-nitrophenyl azide, 4-ethylphenyl azide, 2-trifluoromethoxyphenyl azide, 4-trifluoromethylbenzyl azide or 2-trifluoromethylphenyl azide.
The target compound a and the target compound B in the present invention are both target compounds of the present invention.
The application of the caspovir derivative with biological activity is to prepare the caspovir derivative into an anti-cancer medicament.
The invention has the following beneficial effects:
1. according to the invention, the Babyvir is taken as a lead compound, the benzyl in the Babyvir structure is changed into a 1,2, 3-triazole structure by a click reaction by utilizing a medicament splicing principle, and the methoxyl is kept, so that a series of compounds with novel structures are obtained, and the compound has strong proliferation inhibition activity on various human tumor cells and has obvious application value.
2. The invention simultaneously discovers that other chiral compounds and mixtures thereof of the caspovir derivative also have anti-tumor activity, thereby enlarging the application range.
3. In the synthesis process of the key intermediate 3-b, because the acidity is weakened after amidation and is not beneficial to the subsequent cyclization with chiral amino alcohol micromolecules, an acid water absorbent, such as aluminum sulfate, calcium oxide or calcium chloride, is required to be added, and the non-cyclized by-products can be reduced
Can also improve chiral purity.
4. The method for preparing the key intermediate of the caspovir derivative is optimized, and the yield is improved.
Detailed Description
The present invention will be described in further detail with reference to examples.
Compound 1:1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester;
compound 2-a: (3S) -6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid;
compound 2-b: 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid;
compound 2-c: (3S,11aR) -6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid;
compound 3-a: (3S) -6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8- (3-alkynylphenylamino) carboxamide;
compound 3-b: (3S,11aR) -6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8- (3-alkynylphenylamino) carboxamide;
compound 4: 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;
compound 5: (3S,11aR) -6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8- (2-alkynylphenylamino) carboxamide;
compound 6: (3S,11aR) -6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8- (propargyl) carboxamide;
compound 7: (3S,11aR) -6-hydroxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazolo [3,2-a ] pyridine [1,2-d ] pyrazine-8- (3-alkynylphenylamino) carboxamide.
Example 1
Adding the compound 1(5g) into 30mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after about 3h of reaction, vacuum concentrating, adding 50mL of acetonitrile, then adding 1.7g of S-1-amino-2-propanol, heating to reflux, reacting for 3h, and detecting by TLC that the raw materials are completely reacted; vacuum concentrating, adding 50mL of dichloromethane, adding 20mL of water while stirring, adjusting the pH to 2-3 with dilute hydrochloric acid, separating a lower organic phase, extracting an upper aqueous phase with 10mL of dichloromethane for three times, combining the organic phases, and washing with 50mL of saturated salt water for 3 times; then concentrating under vacuum condition to obtain crude product, and finally recrystallizing in methanol for purification to obtain pure compound 2-a (4.1 g); wherein, the A type configuration: the class B configuration is about 1: 1; 1 H NMR(400MHz,DMSO-d 6 ):15.46(d,J＝8.0Hz,1H),8.76(d,J＝4.0Hz,1H),5.53-5.32(m,1H),4.97-4.77(m,1H),4.47-4.35(m,1H),4.20-4.07(m,2H),3.90(d,J＝4.0Hz,3H),3.25-3.03(m,1H),1.36-1.29(m,3H).
example 2
Adding the compound 1(5g) into 30mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; reacting for about 3 hours, performing vacuum concentration, adding 100mL of acetonitrile, then adding 2.4g of 1-amino-2-propanol, heating to reflux, performing vacuum concentration after reacting for 10 hours, adding 100mL of dichloromethane, adding 50mL of water under stirring, adjusting the pH value to 2-3 with dilute hydrochloric acid, separating out a lower organic phase, extracting an upper aqueous phase with 30mL of dichloromethane for three times, combining the organic phases, and washing with 50mL of saturated saline solution for 3 times; then concentrating under vacuum condition to obtain crude product, and finally recrystallizing in methanol for purification to obtain pure compound 2-b (4.3 g); wherein, the A type configuration, the B type configuration, the C type configuration and the D type configuration are about 1:1:1: 1; LC-MS (ESI): 293[ M-H ]] - 。
Example 3
Adding the compound 1(5g) into 30mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after reacting for about 3 hours, vacuum concentrating, adding 50mL of acetonitrile, then adding 1.7g of S-1-amino-2-propanol and 5g of magnesium trifluoromethanesulfonate, heating until refluxing, reacting for 36 hours, and detecting by TLC that the raw materials are completely reacted; adding 50mL of dichloromethane into the concentrated reaction solution, adding 20mL of water while stirring, adjusting the pH to 2-3 with dilute hydrochloric acid, separating a lower organic phase, extracting an upper aqueous phase for three times with 10mL of dichloromethane, combining the organic phases, and washing for 3 times with 50mL of saturated saline solution; then concentrated under vacuum to give crude product, which was finally purified by recrystallization from methanol to give pure compound 2-c (2.1 g).
Example 4
Adding 3g of compound 2-a (3g), 3.8g of 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate, 2.5g of N, N-diisopropylethylamine, 2.3g of m-aminophenylacetylene and 50mL of N, N-dimethylformamide into a reaction flask at room temperature, and stirring for reacting for 12 hours; TLC shows that the reaction is finished, the reaction solution is added into 50mL of water, a large amount of solid is separated out, and the compound 3-a (3.6g) is obtained after drying; 1 H NMR(400MHz,DMSO-d 6 ):12.53(s,1H),8.69(s,1H),7.95(s,1H),7.58(s,1H),7.38(s,1H),7.23(s,1H),5.52-5.32(m,1H),4.96-4.77(m,1H),4.44-4.36(m,1H),4.22-4.02(m,2H),3.89(s,3H),3.24-3.03(m,1H),2.89-2.73(m,1H),1.35-1.23(m,3H).
example 5
Adding compound 1(16g), 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate 20g, N, N-diisopropylethylamine 12.5g, m-aminophenylacetylene 12g and N, N-dimethylformamide 500mL into a reaction bottle at room temperature, stirring for reaction at room temperature under the protection of argon, adding the reaction solution into 500mL of water to precipitate a large amount of solid, performing suction filtration, and drying to obtain compound 4(15.7 g).
Example 6
Adding the compound 4(2g) into 10mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after about 3.5h of reaction, after concentration in vacuo, 10mL of acetonitrile was added, stirred to dissolve, S-1-amino-2-propanol (0.55g) was added, heated to reflux, reaction was carried out for 36h, after concentration in vacuo, 200mL of dichloromethane was added, 10mL of water was added with stirring, the lower organic phase was separated, the upper aqueous phase was extracted three times with 50mL of dichloromethane, the organic phases were combined, after concentration, and separation and purification by silica gel column chromatography gave compound 3-b (0.88g), LC-ms (esi): 394[ M + H] + 。
Example 7
Adding the compound 4(2g) into 10mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after about 3.5h of reaction, after vacuum concentration, 10mL of acetonitrile was added, stirred to dissolve, S-1-amino-2-propanol (0.55g) and 2g of calcium chloride were added, after stirring for 10min, heating was carried out to reflux, reaction was carried out for 11h, after vacuum concentration, 200mL of dichloromethane was added, 10mL of water was added with stirring, the lower organic phase was separated, the upper aqueous phase was extracted three times with 50mL of dichloromethane, the organic phases were combined, after concentration, recrystallization in methanol was carried out to purify to obtain compound 3-b (1.63g), LC-ms (esi): 394[ M + H] + 。
Example 8
Adding the compound 4(2g) into 10mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after about 3.5h of reactionAfter vacuum concentration, 10mL of acetonitrile is added, stirring is carried out to dissolve the acetonitrile, 0.55g of S-1-amino-2-propanol and 2g of calcium oxide are added, stirring is carried out for 10min, heating is carried out to reflux, reaction is carried out for 11h, 200mL of dichloromethane is added after vacuum concentration, 10mL of water is added under stirring, a lower organic phase is separated, an upper aqueous phase is extracted by 50mL of dichloromethane for three times, the organic phases are combined, and after concentration, the compound 3-b (1.14g) is obtained through column chromatography separation and purification, and the compound is LC-MS (ESI): 394[ M + H] + 。
Example 9
Adding the compound 4(2g) into 10mL of anhydrous formic acid, and heating to 65 ℃ under the protection of argon and stirring; after reacting for about 3.5 hours, vacuum concentrating, adding 10mL of acetonitrile, stirring to dissolve, adding 0.55g of S-1-amino-2-propanol and 2g of aluminum sulfate, stirring for 10min, heating to reflux, reacting for 7 hours, vacuum concentrating, adding 200mL of dichloromethane, adding 10mL of water under stirring, separating a lower organic phase, extracting an upper aqueous phase with 50mL of dichloromethane for three times, combining the organic phases, and washing for 3 times with 10mL of saturated saline solution; then concentrated under vacuum to give crude product, which was finally isolated and purified by column chromatography to give compound 3-b (1.02g), LC-ms (esi): 394[ M + H] + 。
Example 10
Adding a compound 2-c (3g), 2- (7-azabenzotriazole) -N, N, N ', N ' -tetramethylurea hexafluorophosphate 3.8g, N, N ' -diisopropylethylamine 2.5g, 2-aminophenylacetylene 2.3g and N, N-dimethylformamide 50mL into a reaction bottle at room temperature, and stirring for reacting for 12 h; TLC showed that the reaction was complete, and the reaction solution was added to 500mL of water, and a large amount of solid precipitated, after beating sufficiently for several hours, it was filtered under suction and dried to give Compound 5(3.3 g).
Example 11
Adding compound 2-c (10.0g) into 150mL of methyl tert-butyl ether under the protection of nitrogen, adding 5.5g of N, N' -carbonyldiimidazole, slowly heating to 70 ℃, stirring for 3h, stopping heating, dropwise adding a solution of propargylamine 5.5g dissolved in 20mL of methyl tert-butyl ether, reacting at room temperature for 2h, adding 100mL of water, evaporating to remove methyl tert-butyl ether under reduced pressure, adding 100mL of dichloromethane for extraction, carrying out layering, extracting an aqueous layer for 2 times by using 100mL of dichloromethane, combining organic phases, and concentrating to obtain compound 6(7.7 g); LC-MS (ESI): 332[ M + H] + 。
Example 12
In a reaction bottle, adding the compound 3-b (40g) and anhydrous lithium bromide (26g) into 300mL of anhydrous tetrahydrofuran, and uniformly stirring at room temperature; slowly heating to reflux, keeping the temperature, stirring for 1.5h, detecting by TLC that reaction raw materials disappear, cooling to room temperature, adding 200mL of water, and stirring for 20 min; adding 400mL of dichloromethane after vacuum concentration, adding 2N hydrochloric acid solution while stirring at room temperature to adjust the pH of the reaction solution to 2-4, extracting, layering, extracting the water phase with 300mL of dichloromethane for three times, combining the organic phases, and concentrating the organic phase to obtain a compound 7(24 g); LC-MS (ESI): 380[ M + H ]] + 。
Example 13
Sequentially adding 0.5g of 2-trifluoromethylphenyl azide, 3-a (0.5g) of a compound, 10mL of tert-butyl alcohol, 10mL of tetrahydrofuran and 10mL of water, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate into a reaction bottle, reacting at 60 ℃, monitoring the completion of the reaction of raw materials by TLC, extracting twice with dichloromethane, drying an organic phase by anhydrous magnesium sulfate, evaporating to remove a solvent, and recrystallizing by methanol to obtain 8-a (0.44g) of a target compound; 1H NMR (400MHz, DMSO-d6) 12.60(d, J ═ 8.0Hz,1H),9.10(s,1H),8.72(s,1H),8.25(s,1H),8.08(d, J ═ 8.0Hz,1H),7.96(t, J1 ═ 8.0Hz, J2 ═ 16.0Hz,1H),7.80-7.70(m,3H),7.69(d, J ═ 8.0Hz,1H),7.50(t, J1 ═ 8.0Hz, J2 ═ 8.0Hz,1H),5.54-5.34(m,1H),4.98-4.79(m,1H),4.46-4.36(m,1H),4.19-4.05(m,2H),3.92 (m, 3.37H), 3.04 (d, 3H), 3.30H, 3H), 3.1H, 1H, 3H, 1H, 3H, 1H, 3.0H, 1H, and 1H.
Example 14
Sequentially adding 0.5g of 2-ethyl phenyl azide, 3-a (0.5g) of a compound, 10mL of tert-butyl alcohol, 10mL of water, 10mL of tetrahydrofuran, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate into a 500mL reaction bottle, reacting at 60 ℃, monitoring the completion of the reaction of raw materials by TLC, extracting twice with dichloromethane, drying an organic phase with anhydrous magnesium sulfate, evaporating to remove a solvent, and recrystallizing with methanol to obtain 8-b (0.41g) of a target compound; 1H NMR (400MHz, DMSO-d6) 12.60(d, J ═ 8.0Hz,1H),9.02(s,1H),8.72(s,1H),8.22(s,1H),7.84(d, J ═ 8.0Hz,1H),7.69(d, J ═ 8.0Hz,1H),7.59-7.45(m,5H),5.54-5.34(m,1H),4.98-4.80(m,1H),4.46-4.36(m,1H),4.19-4.04(m,2H),3.92(d, J ═ 4.0Hz,3H),3.25-3.04(m,1H),2.54(t, J1 ═ 8.0, J35 ═ 8.0, 2H, 1H, 37.30, 7H, 3.42H), 3.8H, 368H, 3.42H, 368H, 3.8H, 3607 (t, 3H).
Example 15
Adding 0.5g of 3, 5-bis (trifluoromethyl) phenyl azide, a compound 3-a (0.5g), 10mL of tert-butyl alcohol, 10mL of water, 10mL of tetrahydrofuran, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate into a 500mL reaction bottle in sequence, reacting at 60 ℃, monitoring the completion of the reaction of raw materials by TLC, extracting twice with dichloromethane, drying an organic phase with anhydrous magnesium sulfate, evaporating the solvent, and recrystallizing with methanol to obtain a target compound 8-c (0.32 g); 1 H NMR(400MHz,DMSO-d 6 ):12.64(s,1H),9.69(s,1H),8.71(s,3H),8.31(s,1H),8.17(s,1H),7.88(s,1H),7.69(s,1H),7.52(s,1H),5.54-5.35(m,1H),4.98-4.79(m,1H),4.45-4.38(m,1H),4.18-4.05(m,1H),3.91(s,3H),3.26-3.05(m,1H),1.37-1.31(m,3H),1.11(s,1H).
example 16
Adding 0.5g of 3-trifluoromethylphenyl azide, 0.5g of compound 7(0.5g), 10mL of tert-butyl alcohol, 10mL of tetrahydrofuran and 10mL of water in turn, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate into a reaction flask, reacting at 60 ℃, monitoring the completion of the reaction of raw materials by TLC, extracting twice with dichloromethane, drying an organic phase with anhydrous magnesium sulfate, evaporating the solvent, and recrystallizing with methanol to obtain a target compound 9-a (0.29 g); LC-MS (ESI): 567[ M + H ]] + 。
Example 17
Sequentially adding 0.5g of 3-nitrophenylazide, 0.5g of compound 3-b (0.5g), 10mL of tert-butyl alcohol, 10mL of water, 10mL of tetrahydrofuran, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate into a 500mL reaction bottle, reacting at 60 ℃, monitoring the complete reaction of raw materials by TLC, extracting twice by using dichloromethane, drying an organic phase by anhydrous magnesium sulfate, evaporating to remove a solvent, and recrystallizing by using methanol to obtain a target compound 9-b (0.14 g); LC-MS (ESI): 558[ M + H] + 。
Example 18
Adding 0.5g of 2-trifluoromethyl phenyl azide, 0.5g of compound 5(0.5g), 10mL of tert-butyl alcohol, 10mL of tetrahydrofuran and 10mL of water, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate in turn into a reaction bottle, reacting at the temperature of 60 ℃, monitoring the complete reaction of raw materials by TLC (thin layer chromatography), extracting twice by using dichloromethane, drying an organic phase by anhydrous magnesium sulfate, and evaporating off a solventThen, recrystallization was carried out with methanol to obtain the objective compound 10(0.21 g); LC-MS (ESI): 581[ M + H] + 。
Example 19
Sequentially adding 0.5g of 2-ethyl phenyl azide, 0.5g of compound 6(0.5g), 10mL of tert-butyl alcohol, 10mL of tetrahydrofuran and 10mL of water, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate into a reaction bottle, reacting at the temperature of 60 ℃, monitoring the complete reaction of raw materials by TLC, extracting twice by using dichloromethane, drying an organic phase by anhydrous magnesium sulfate, evaporating the solvent, and recrystallizing by using methanol to obtain a target compound 11(0.29 g); LC-MS (ESI): 479[ M + H ]] + 。
Example 20
Sequentially adding 0.5g of 2-trifluoromethoxyphenyl azide, 0.5g of compound 3-a (0.5g), 10mL of tert-butyl alcohol, 10mL of water, 10mL of tetrahydrofuran, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate into a 500mL reaction bottle, reacting at 60 ℃, monitoring the complete reaction of raw materials by TLC, extracting twice by using dichloromethane, drying an organic phase by using anhydrous magnesium sulfate, evaporating a solvent, and washing by using methanol to obtain a target compound 12(0.411 g); LC-MS (ESI): 597[ M + H] + 。
Example 21
In a 500mL reaction flask, 0.5g of 4-trifluoromethyl benzyl azide, 0.5g of compound 3-a (0.5g), 10mL of tert-butyl alcohol, 10mL of water, 10mL of tetrahydrofuran, 0.25g of copper sulfate pentahydrate and 0.5g of sodium ascorbate are sequentially added, the reaction is carried out at 60 ℃, TLC monitors the complete reaction of raw materials, dichloromethane is used for extraction twice, an organic phase is dried by anhydrous magnesium sulfate, a solvent is evaporated, and then methanol is used for washingTo obtain the target compound 13(0.286 g); LC-MS (ESI): 595[ M + H] + 。
Example 22
From CO 2 Lung cancer cell A549, lung cancer cell H1985 and lung fiber cell CCD19 cell culture bottles are taken out from the incubator and respectively carried out the following operations: aseptic technique 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% 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 centrifuge tube is poured slowly, 2-5 mL 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% CO 2 Culturing in an incubator for 24 h.
Drug molecules were formulated to the desired concentration: 20. mu. mol/L, 10. mu. mol/L, 5. mu. mol/L, 2.5. mu. mol/L. From CO 2 The 96-well plate was removed from the incubator, and 100. mu.L of drug-containing medium was added to each well, with 3 multiple wells 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% CO 2 Culturing for 72h in an incubator. Three experiments were performed with the same batch of cells at different passage numbers for each drug. After 72 hours, 10. mu.L of 5mg/mL MTT solution was added to each well in the dark, and CO addition was continued 2 Culturing for 4h in an incubator, sucking supernatant, adding 100 mu L DMSO into each well, 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 the invention ]/OD Control of X is 100%; the inhibition data IC50 of the target compound 8-a on lung cancer cells A549 and H1975 and lung fibroblast CCD19 cells are respectively 1-3 mu mol/L, 6-8 mu mol/L and 7-9 mu mol/L through detection; the activity of the compounds 8-b and 8-c on A549 is respectively better than 11 and 9-b. We find that the drug molecule has certain inhibition effect on A549 cells and has low toxicity.
Example 23
From CO 2 Tumor cells H460, Huh7, SKOV3, HCT116 and MVF-7 are taken out from the incubator and respectively carried out the following operations: and (3) performing sterile operation beside an alcohol lamp, opening a dish cover, sucking out culture solution into a waste liquid jar, washing cells in the dish for 1 time by using 2mL PBS, digesting by using 0.25% pancreatin, stopping digestion when cell gaps are increased and the cells are changed into a small circle shape, and blowing the bottom of the dish by using a pipette to make the cells fall off. The resulting cell suspension was transferred to a sterile 15mL centrifuge tube at 800r/min for 2min, centrifuged, then the tube was slowly poured over to remove the supernatant, leaving the bottom cells, 3mL of culture medium was added, and the cells were counted under an inverted microscope. According to the counting result, the obtained product is prepared into 1 × 10 culture solution 4 cells/mL of single cell suspension, then seeded into 96-well plates at 100. mu.L per well. The 96-well plate was placed at 37 ℃ in 5% CO 2 Culturing in an incubator for 24 h.
The resulting drug molecules were formulated to the desired concentrations: 0.4, 2.0, 10.0, 20.0 and 50.0 mu mol/L. From CO 2 The 96-well plate was removed from the incubator, the supernatant was removed, and 100. mu.L of the drug-containing medium was added to each well, and 5 wells were provided. As a blank well, an equal volume of the corresponding culture medium was added. Then placing the 96-well plate added with the medicine at 37 ℃ and 5% CO 2 The culture in the incubator is respectively carried out for 72 h. Three experiments were performed with the same batch of cells at different passage numbers for each drug. After 72 hours, the supernatant was removed under dark conditions, mixed solution was prepared by adding 10uL of CCK8 detection solution per 100uL of culture medium, and 100uL of mixed solution was added to each well of 96-well plate cells, including blank control wells. Continuously introducing CO 2 Culturing for 1h in an incubator, measuring the absorbance OD value of the sample at the wavelength of 450nm by using an enzyme-labeling instrument, subtracting the blank OD value from the sample OD value, and calculating and comparing the cell activity.
The inhibitory activity of the compound 8-a obtained in example 13 on the 5 tumor cells was 31.2, 26.6, 12.7, 31.4, 11.5. mu. mol/L, respectively; the inhibitory activity of the compound 8-b obtained in example 14 on these 5 tumor cells was 7.2, 21.2, 24.1, 60.3, 59.8. mu. mol/L, respectively; the inhibitory activity of the compound 8-c obtained in example 15 on these 5 tumor cells was 130.5, 28.7, 74.7, 195.0, 71.8. mu. mol/L, respectively; the inhibitory activity of the compound 12 obtained in example 20 on the 5 tumor cells was 4.7, 6.5, 4.9, 24.1, 27.9. mu. mol/L, respectively; the inhibitory activity of compound 13 obtained in example 21 on these 5 tumor cells was 25.5, 7.2, 5.7, 16.1, 8.9. mu. mol/L, respectively.
While the foregoing embodiments have described the general principles, features and advantages of the present invention, it will be understood by those skilled in the art that the present invention is not limited thereto, and that the foregoing embodiments and descriptions are only illustrative of the principles of the present invention, and various changes and modifications can be made without departing from the scope of the principles of the present invention, and these changes and modifications are within the scope of the present invention.
Claims (10)
1. A biologically active caspovir derivative characterized by the structural formula:
wherein:
R 1 is methyl or hydrogen;
R 2 is methylene, isopropyl, phenyl or benzyl;
R 3 is hydrogen, methyl, ethyl, phenyl or benzyl.
2. Biologically active caspovir derivatives according to claim 1 characterized in that it is a mixture of one or more of A, B, C or D;
3. a process for the preparation of biologically active caspovir derivatives as claimed in claim 1 or 2, comprising the steps of:
(1)1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester is deprotected to aldehyde, and then cyclized with aminopropanol to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid; then carrying out acylation reaction with an amino compound to obtain an acylation product;
or alternatively
Reacting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester with an amino compound, and cyclizing with aminopropanol to obtain an acylation product;
(2) and (3) reacting the acylation product with azide by click to obtain the target compound.
4. The process for the preparation of biologically active caspovir derivatives as claimed in claim 3,
the step (1) is as follows:
adding 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester into formic acid, heating to 60-70 ℃ under the protection of argon, and reacting for 2-4 h; concentrating, adding acetonitrile, adding a mixture of aminopropanol and magnesium trifluoromethanesulfonate or aminopropanol, and carrying out reflux reaction for 2-40 h; TLC detects that the raw materials completely react, dichloromethane and water are added after concentration, the pH value of the reaction solution is adjusted to 2-3, a lower organic phase is separated, and 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid is obtained through concentration;
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 aminopropanol is 1: 1-2; the aminopropanol is S-1-amino-2-propanol or 1-amino-2-propanol;
6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid is acylated with aminophenylacetylene to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (X) carboxamide;
wherein X is 2-alkynyl phenylamino, 3-alkynyl phenylamino or propargyl; the aminophenylacetylene is 2-aminophenylacetylene, 3-aminophenylacetylene or propargylamine.
5. The process for the preparation of biologically active caspovir derivatives as claimed in claim 4,
step (1) is as follows:
stirring and reacting 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid, 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate, N, N-diisopropylethylamine, 3-aminophenylacetylene or 2-aminophenylacetylene and N, N-dimethylformamide for 11-13h at room temperature; TLC shows that the reaction is finished, the reaction liquid is added into water, solid is separated out, and 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (X) formamide is obtained after suction filtration and drying;
the molar ratio of the 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid to the 3-aminophenylacetylene or the 2-aminophenylacetylene is 1: 1-2;
or step (1) is as follows:
under the protection of nitrogen, 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid is added into methyl tert-butyl ether, adding N, N' -carbonyl diimidazole, heating to 60-80 deg.C, stirring for 2-4h, stopping heating, adding dropwise methyl tert-butyl ether solution containing propargylamine, then reacting for 1-3h at room temperature, adding water, evaporating the solvent under reduced pressure, adding dichloromethane for extraction, and concentrating to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (propargyl) formamide;
the molar ratio of the 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydro-oxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8-carboxylic acid to propargylamine is 1: 1-3.
6. The process for the preparation of biologically active caspovir derivatives as claimed in claim 3,
the step (1) is as follows:
reacting 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-pyridinedicarboxylic acid-2-methyl ester, 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate, N, N-diisopropylethylamine, aminophenylacetylene and N, N-dimethylformamide at room temperature under the protection of argon; TLC shows that the reaction is finished, the reaction solution is added into water, solid is separated out, and the 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-diformic acid pyridine-2-methyl formate-5- (X) formamide is dried after suction filtration;
wherein X is 2-alkynyl phenylamino, 3-alkynyl phenylamino or propargyl; the aminophenylacetylene is 2-aminophenylacetylene, 3-aminophenylacetylene or propargylamine; 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 aminophenylacetylene is 1: 1-2;
adding 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-diformic acid pyridine-2-methyl formate-5- (X) formamide into anhydrous formic acid, and heating to 60-70 ℃ under the protection of argon; reacting for 2-4h, vacuum concentrating, adding acetonitrile, stirring for dissolving, adding aminopropanol or a mixture of aminopropanol and calcium chloride or a mixture of aminopropanol and calcium oxide or a mixture of aminopropanol and aluminum sulfate, heating to reflux, reacting for 5-30h, vacuum concentrating, adding dichloromethane, stirring, adding water, separating out a lower organic phase, concentrating, and purifying to obtain 6-methoxy-3-methyl-5, 7-dioxo-2, 3,5,7,11,11 a-hexahydrooxazole [3,2-a ] pyridine [1,2-d ] pyrazine-8- (X) formamide;
wherein X is 2-alkynylanilino, 3-alkynylanilino or propargyl; the molar ratio of 1- (2, 2-dimethoxyethyl) -1, 4-dihydro-3-methoxy-4-oxo-2, 5-dicarboxylic acid pyridine-2-methyl formate-5- (X) formamide to aminopropanol is 1: 1-2; the aminopropanol is S-1-amino-2-propanol or 1-amino-2-propanol.
7. The process for the preparation of biologically active caspovir derivatives as claimed in claim 3,
the step (2) is carried out according to one of the following steps:
directly carrying out click reaction on the acylation product and azide to obtain a target compound A; demethylating the target compound A to obtain a target compound B; or demethylating the acylation product and then carrying out click reaction with azide to obtain a target compound B.
8. The process for the preparation of biologically active caspovir derivatives as claimed in claim 7,
the step (2) is as follows: mixing azide, an acylation product, tert-butyl alcohol, water, tetrahydrofuran, copper sulfate pentahydrate and sodium ascorbate, reacting at 50-70 ℃, monitoring by TLC that raw materials are completely reacted to obtain yellow liquid, adding dichloromethane, filtering, extracting, drying, and evaporating to remove a solvent to obtain a target compound A; reacting the target compound A with anhydrous lithium bromide to obtain a target compound B;
the azide is benzyl azide, 2-bromobenzyl azide, 2-ethylphenyl azide, 3, 5-bis (trifluoromethyl) phenyl azide, 2-fluorophenyl azide, 4-trifluoromethylphenyl azide, 2-methyl-3-nitrophenyl azide, 4-fluorophenyl azide, 3-nitrophenyl azide, 4-ethylphenyl azide, 2-trifluoromethoxyphenyl azide, 4-trifluoromethylbenzyl azide or 2-trifluoromethylphenyl azide.
9. The process for the preparation of biologically active caspovir derivatives as claimed in claim 7,
the step (2) is as follows: after the acylation product reacts with anhydrous lithium bromide for demethylation, mixing azide, the acylation product after demethylation, tert-butyl alcohol, water, tetrahydrofuran, copper sulfate pentahydrate and sodium ascorbate, reacting at 50-70 ℃, monitoring the complete reaction of raw materials by TLC to obtain yellow liquid, adding dichloromethane, filtering, extracting, drying, and evaporating to remove the solvent to obtain a target compound B;
the azide is benzyl azide, 2-bromobenzyl azide, 2-ethylphenyl azide, 3, 5-bis (trifluoromethyl) phenyl azide, 2-fluorophenyl azide, 4-trifluoromethylphenyl azide, 2-methyl-3-nitrophenyl azide, 4-fluorophenyl azide, 3-nitrophenyl azide, 4-ethylphenyl azide, 2-trifluoromethoxyphenyl azide, 4-trifluoromethylbenzyl azide or 2-trifluoromethylphenyl azide.
10. The application of the caspovir derivative with biological activity is characterized in that the caspovir derivative is used for preparing a medicament for resisting lung cancer, liver cancer, ovarian cancer or colon cancer.
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