A pomalidomide feed additive and its preparation method and application

Abstract

The present invention discloses a pomalidomide feed additive and a preparation method and application thereof, and belongs to the technical field of functional feed additive synthesis. The technical scheme of the present invention is as follows: the pomalidomide feed additive molecule has a structure #imgabs0# wherein n is 1 or 2; R is an alkyl group or a halogen group or a methoxy group or #imgabs1#. The present invention uses 2-nitrobenzoic acid as a starting material, first reacts to obtain 2-(2,6-dioxopiperidin-3-yl)-4-nitroisoindoline-1,3-dione, then reacts to obtain pomalidomide, then introduces an azide group, and finally obtains a target molecule with a novel structure through a click reaction, which has a certain inhibitory effect on urease and can be used as a potential feed additive.

Description

Technical Field

[0001] The invention belongs to the technical field of synthesis of feed additives, and in particular relates to a pomalidomide feed additive and a preparation method and application thereof. Background Art

[0002] Immunomodulators can increase the production of IL-2 by T lymphocytes and reduce the production of proinflammatory cytokines. These effects have important applications in tumor treatment. Pomalidomide, with the chemical name (RS)-4-amino-2-(2,6-dioxopiperidin-3-yl)isoindoline-1,3-dione, is an oral small molecule derivative and a newly launched third-generation immunomodulator. It was developed by Celgene Corporation in the United States. It is a modified and synthesized drug based on the chemical structure of the first-generation IMiD thalidomide. It can enhance the immune response mediated by T cells and natural killer cells, inhibit the production of proinflammatory cytokines in monocytes, and induce tumor cell apoptosis. It has received widespread attention in the treatment of various malignant tumors and immune diseases. It is a very widely used thalidomide derivative. Compared with thalidomide, the amino group connected to the benzene ring of pomalidomide makes its own chemical properties more stable, and it has a stronger immunomodulatory effect than thalidomide. In terms of clinical application, pomalidomide is safer than thalidomide and has fewer adverse reactions. It has almost no teratogenicity and neurotoxicity, and is effective against a variety of blood diseases and solid malignant tumors.

[0003] The 1,2,3-triazole structure is a very important class of nitrogen-containing heterocyclic compounds. It is a five-membered heterocyclic ring constructed by 3 nitrogen atoms and 2 carbon atoms, and its molecular formula is C2N3H3. 1,2,3-triazole has a special planar rigid structure, which gives it a strong ability to embed into DNA. It also has a large dipole moment and can form various non-covalent interactions with different biological targets, such as hydrophobic, hydrogen bonds, van der Waals forces, and dipole-dipole bonds. In addition, the structural characteristics of 1,2,3-triazole allow it to be an electronic equivalent substitute for amides, esters, carboxylic acids, olefin rigid analogs, etc., and therefore has a broad spectrum of biological activity. It is often used as an important molecular building block for the synthesis of active compounds, such as the preparation of antibacterial, antimalarial, antifungal, antiviral, antituberculosis and anticancer active compounds. A variety of clinical drugs and active molecules have been modified with 1,2,3-triazole to enhance their original biological activities or acquire new biological activities. For example, the terminal alkyne was introduced into the core structure of the HIV integrase inhibitor drug dolutegravir, and the compound DTHP was obtained by reacting with 2-trifluoromethylphenyl azide. It has good inhibitory activity against a variety of lung cancer cells, especially H1975 cells, which can cause apoptosis and enhance ROS levels in the cells. The tumor inhibition effect in mice experiments is also very significant. The EGFR inhibitor icotinib has the structural characteristics of the terminal alkyne, and reacted with 3-chlorophenyl azide to obtain compound a7. This compound has good inhibitory effects on both mutant lung cancer cells (PC-9) and wild-type lung cancer cells (H460 and A549), which is better than icotinib. A research group introduced 1,2,3-triazole into the structure of naphthoquinone compounds to obtain compound 7a, which has an inhibitory effect on Caco-2 cells by blocking topoisomerase I and IIα (IC 50 =23.92μM), and showed a certain selectivity; a research group designed indole 2-one derivatives containing 1,2,3-triazole, among which compound 13b had a good inhibitory effect on VEGFR-2 kinase (IC50=26.38nM) and also had inhibitory effects on various tumor cells.

[0004]

[0005] Given the remarkable effects of introducing phenyl-1,2,3-triazole into the drug molecule, in order to find newer and more efficient anti-tumor drugs, we modified pomalidomide and introduced phenyl-1,2,3-triazole groups into its structure, hoping to improve and expand the application of pomalidomide. Summary of the invention

[0006] The technical problem solved by the present invention is to provide a preparation method of a pomalidomide derivative with a novel structure.

[0007] The present invention adopts the following technical solution to solve the above technical problems, a method for preparing a pomalidomide derivative, characterized by the specific steps of:

[0008] 1. Add a certain amount of 2-nitrobenzoic acid into dichlorothionyl, stir for a period of time under nitrogen protection, and then concentrate the reaction system in vacuum. Dissolve the concentrate with dichloromethane, and then slowly drop a dichloromethane solution containing 2-amino-1,5-pentanedioic acid dimethyl ester hydrochloride under stirring. After the addition is complete, continue to stir and react for a period of time, then concentrate the reaction system, add N,N-dimethylformyl, and after complete dissolution, add a certain amount of tert-butyl lithium, barium hydroxide, cesium carbonate and palladium acetate under nitrogen protection, maintain nitrogen atmosphere, transfer to a high-pressure reactor, and protect the autoclave with nitrogen throughout the process. Then stir at room temperature for a period of time and use dichloromethane. The gas in the autoclave is replaced by carbon dioxide, and then carbon dioxide is introduced to make the pressure in the autoclave reach a certain value, and the mixture is slowly heated to a certain temperature. After a period of reaction, the carbon dioxide is removed in vacuo, and the temperature is lowered to a certain temperature. Then ammonia is introduced to make the pressure in the autoclave reach a certain value. The temperature and pressure are maintained and stirred for a period of time, and the mixture is cooled to room temperature. Water and dichloromethane are then added to the reaction system, and the reaction liquid is filtered after stirring to separate the organic phase. The aqueous phase is washed with dichloromethane for several times, and then the organic phase is separated. After concentration, the mixture is separated and purified by silica gel column chromatography to obtain 2-(2,6-dioxopiperidin-3-yl)-4-nitroisoindoline-1,3-dione.

[0009] 2. Add a certain amount of methyl 2-nitrobenzoate and calcium carbonate to a dichloroethane solution, stir, and then slowly drop a certain amount of dichloroethane solution containing a certain amount of iodine chloride. After the addition is complete, stir for a period of time at room temperature, cool and filter the reaction solution, add anhydrous magnesium sulfate, stir for a period of time, filter, add a certain amount of 3-aminopiperidine-2,6-dione hydrochloride, triethylamine and DCC to the filtrate, stir and react for a period of time at room temperature, concentrate, wash the concentrate with ether for several times, then add toluene, add a certain amount of triphenylphosphine and iodine, stir evenly, add palladium acetate and triethylamine, stir for a period of time under nitrogen protection, add anhydrous formic acid, maintain nitrogen atmosphere, heat to 70°C, filter the reaction solution after the reaction is completed, add water, stir, extract with dichloromethane for several times, combine the organic phases, and concentrate to obtain 2-(2,6-dioxopiperidin-3-yl)-4-nitroisoindoline-1,3-dione.

[0010] 3. A certain amount of 2-nitrobenzoic acid is added to dichlorothionyl, and the reaction system is concentrated in vacuum after stirring for a period of time under nitrogen protection. The concentrate is dissolved with dichloromethane, and then a dichloromethane solution containing dimethyl 2-aminoglutaric acid hydrochloride is slowly added dropwise under stirring. After the addition is complete, the stirring reaction is continued for a period of time. A saturated sodium chloride solution is added to the reaction system, and the organic phase is separated after stirring and added to a high-pressure reactor. A certain amount of EDCI, 1-hydroxybenzotriazole and triethylamine are added to the reactor, and the air in the reactor is removed by vacuum, and ammonia is introduced into the reactor to make the pressure in the reactor reach 0.5MPa, and the reactor is heated to 50°C. After the reaction is completed, the reaction solution is filtered, and then the reaction system is adjusted to be neutral with acetic acid, and water is added. The organic phase is separated after stirring and concentrated to obtain N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide.

[0011] 4. In a high-pressure reactor, a certain amount of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, cesium carbonate and palladium acetate are added to N,N-dimethylformamide, the reaction system is protected by nitrogen, and after stirring, an N,N-dimethylformamide solution containing sodium tert-butoxide is quickly added, and then the gas in the reactor is removed by vacuum, and carbon dioxide is introduced to make the pressure in the reactor reach pressure 1, and then slowly heated to 110°C, maintained at the temperature for a period of time, and then cooled to 40°C, and the gas in the reaction system is removed by vacuum again, and hydrogen is introduced into the reactor to make the pressure in the reactor reach pressure 2, and the temperature is maintained and stirred for reaction. After the reaction is completed, the reaction liquid is filtered, and the filtrate is poured into water, and the pH is adjusted to 6 with dilute hydrochloric acid under stirring, and then the reaction system is extracted with dichloromethane for multiple times, and the organic phases are combined, dried, concentrated, and separated by silica gel column chromatography to obtain pomalidomide.

[0012] 5. In a high-pressure reactor, a certain amount of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, cesium carbonate and palladium acetate are added to N,N-dimethylformamide, the reaction system is protected by nitrogen, and after stirring, a solution of N,N-dimethylformamide containing sodium tert-butoxide is quickly added, and then the gas in the reactor is removed by vacuum, and carbon dioxide is introduced to make the pressure in the reactor reach 1.5 MPa, and then slowly heated to 110°C, and the temperature is maintained for a period of time. After filtering the reaction liquid, the filtrate is poured into water, and the pH is adjusted to 6 with dilute hydrochloric acid under stirring, and then the reaction system is extracted with dichloromethane for several times, the organic phases are combined, concentrated and added to methanol, and then a certain amount of palladium carbon with a content of 10% is added. Under nitrogen protection, hydrazine hydrate is slowly added dropwise, and the reaction temperature is maintained at room temperature during the addition. After the addition is completed, the reaction is continued for 7.5 hours, and the reaction liquid is filtered to remove the catalyst palladium carbon, and concentrated and separated by silica gel column chromatography to obtain pomalidomide.

[0013] 5. In a high-pressure reactor with stirring, a certain amount of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, cesium carbonate and palladium acetate are added to N,N-dimethylformamide, the reaction system is protected by nitrogen, and after stirring, N,N-dimethylformamide solution containing sodium tert-butoxide is quickly added, and then the gas in the reactor is removed by vacuum, and carbon dioxide is introduced to make the pressure in the reactor reach 1.5 MPa, and then slowly heated to 110°C. After maintaining the temperature for a period of time, the reaction liquid is filtered, and the filtrate is poured into water. The pH is adjusted to 6 with dilute hydrochloric acid under stirring, and then the reaction system is extracted with dichloromethane for several times, the organic phases are combined, concentrated, and added to a mixed solution of ethanol and water, and then reduced iron powder is added, acetic acid is added dropwise, and heated to a certain temperature. After the reaction is completed, the reaction liquid is filtered, and the filtrate is evaporated by reduced pressure distillation to evaporate ethanol and water, and acetone and benzene are added to the residue for recrystallization, and the solid is filtered by suction, and the filter cake is dried to obtain pomalidomide.

[0014] 6. Add a certain amount of pomalidomide to hydrochloric acid, stir and add a certain amount of sodium nitrite, maintain the temperature at 0°C, add an aqueous solution of sodium azide, continue stirring for a period of time, add dichloromethane to the reaction system, stir and separate the organic phase, extract the aqueous phase with dichloromethane several times, combine the organic phases, and concentrate to obtain pomalidomide-azide compound.

[0015] 7. Add a certain amount of pomalidomide-azide compound and aminoalkyne compound to a mixed solvent of distilled water, tetrahydrofuran and tert-butanol, then add anhydrous copper sulfate and reagent grade sodium ascorbate, protect with nitrogen, heat to 80°C, reflux reaction until the raw materials react completely, then slowly drop a dichloromethane solution containing a benzoyl chloride compound into the reaction system, stir for a while after the dropwise addition, add a saturated sodium chloride solution, separate the organic phase after stirring, and concentrate the organic phase to obtain the product. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 This is the H NMR spectrum of the target compound prepared in Example 1.

[0017] Figure 2 This is the H NMR spectrum of the target compound prepared in Example 9.

[0018] Figure 3 This is the H NMR spectrum of the target compound prepared in Example 10. DETAILED DESCRIPTION

[0019] The above contents of the present invention are further described in detail below through examples, but this should not be understood as the scope of the above subject matter of the present invention being limited to the following examples, and all technologies implemented based on the above contents of the present invention belong to the scope of the present invention.

[0020] Example 1

[0021]

[0022] In a reaction bottle with a stirring device, 1.7 g of 2-nitrobenzoic acid was added to 50 mL of dichlorothionyl, and stirred for 1 h under nitrogen protection, and then the reaction system was vacuum concentrated. The concentrate was dissolved with 100 mL of dichloromethane, and then 50 mL of a dichloromethane solution containing 2.3 g of 2-amino-1,5-pentanedioic acid dimethyl ester hydrochloride was slowly added dropwise at 0 ° C. After the addition was completed, the temperature was raised to room temperature and stirred for 2.5 h. The reaction system was vacuum concentrated, and the concentrate was added to 200 mL of N, N-dimethylformamide and stirred. After complete dissolution, 1.28 g of tert-butyl lithium, 3.4 g of barium hydroxide, 6.5 g of cesium carbonate and 0.45 g of palladium acetate were added under nitrogen protection, and the nitrogen atmosphere was maintained, and the reaction was transferred to a high-pressure reactor. The autoclave was protected by nitrogen throughout the process, and then Stir at room temperature for 30 minutes, replace the gas in the autoclave with carbon dioxide, then introduce carbon dioxide to make the pressure in the autoclave reach 0.2MPa, slowly heat to 120°C, react for 15 hours, remove carbon dioxide in vacuum, cool to 70°C, then introduce ammonia to make the pressure in the autoclave reach 0.15MPa, maintain temperature and pressure and stir to react for 7 hours, cool to room temperature, then add 200mL of water and 100mL of dichloromethane to the reaction system, stir and filter the reaction liquid, separate the organic phase, wash the aqueous phase with 50mL of dichloromethane for several times, then separate the organic phase, dry over anhydrous sodium sulfate, concentrate in vacuum and separate and purify by silica gel column chromatography to obtain 2.57g of 2-(2,6-dioxopiperidin-3-yl)-4-nitroisoindoline-1,3-dione, 1 H NMR (400MHz, DMSO-d6) δ11.15(s,1H),8.32(d,J=8.0Hz,1H),8.20(d,J=4.0Hz,1H),8.08(t,J1=8.0Hz,J2 =4.0Hz,1H),5.16(dd,J1=4.0Hz,J2=4.0Hz,1H),2.89-2.81(m,1H),2.60-2.48(m,2H),2.06-2.01(m,1H).

[0023] Example 2

[0024]

[0025] In a reaction flask with a stirring device, 1.8 g of methyl 2-nitrobenzoate and 3 g of calcium carbonate were added to 100 mL of dichloroethane solution, and then 70 mL of dichloroethane solution containing 1.7 g of iodine chloride was slowly added dropwise after stirring. After the addition was complete, the mixture was stirred for 4.0 h at room temperature, and then the reaction solution was cooled and filtered. 10 g of anhydrous magnesium sulfate was added, and the mixture was stirred for 30 min and filtered. 1.7 g of 3-aminopiperidine-2,6-dione hydrochloride, 1 g of triethylamine, 2 g of DCC reagent, and 50 mL of dichloroethane were added to the filtrate, and the reaction was stirred for 12.5 h at room temperature. The concentrate was washed with 50 mL of ether after vacuum concentration. times, then add 120 mL of toluene to the concentrate and stir to dissolve, then add 2.6 g of triphenylphosphine and 3 g of iodine, stir evenly, then add 0.23 g of palladium acetate and 1 g of triethylamine, stir for 30 min under nitrogen protection, then add 1.5 g of anhydrous formic acid and 20 mL of toluene, maintain nitrogen atmosphere, heat to 70 ° C, react for 5 h, stop the reaction, filter the reaction solution, add 200 mL of water to the filtrate, stir and extract with 80 mL of dichloromethane 5 times, combine the organic phases, and concentrate the organic phases in vacuo to obtain 2.79 g of 2-(2,6-dioxopiperidin-3-yl)-4-nitroisoindoline-1,3-dione, 1 H NMR (400MHz, DMSO-d6) δ11.15(s,1H),8.32(d,J=8.0Hz,1H),8.20(d,J=4.0Hz,1H),8.08(t,J1=8.0Hz,J2 =4.0Hz,1H),5.16(dd,J1=4.0Hz,J2=4.0Hz,1H),2.89-2.81(m,1H),2.60-2.48(m,2H),2.06-2.01(m,1H).

[0026] Example 3

[0027]

[0028] In a reaction bottle with a stirring device, 17 g of 2-nitrobenzoic acid was added to 200 mL of dichlorothionyl, and stirred for 1 h under nitrogen protection. Then, the reaction system was concentrated by heating in vacuum, and the concentrate was dissolved by stirring with 300 mL of dichloromethane. Then, 450 mL of dichloromethane solution containing 23 g of 2-amino-1,5-pentanedioic acid dimethyl ester hydrochloride was slowly added dropwise under stirring. After the addition was complete, the stirring reaction was continued for 2.5 h. 300 mL of saturated sodium chloride solution was added to the reaction system, and the organic phase was separated after stirring. 20 g of anhydrous magnesium sulfate was added, and the mixture was filtered after stirring for 30 min. The filtrate was added to a high-pressure reactor, and EDCI was added to the reactor. 19g and 13.5g of 1-hydroxybenzotriazole and 20g of triethylamine, then add 100mL of N,N-dimethylformamide, remove the air in the reactor by vacuum, introduce ammonia into the reactor to make the pressure in the reactor reach 0.5MPa, heat to 50℃, react for 5h, stop the reaction, filter the reaction liquid, and then adjust the reaction system to neutral with acetic acid, add 300mL of water, separate the organic phase after stirring, and concentrate the organic phase in vacuum to obtain 23.6g of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, MS(ESI + )m / z:278[M+H] + .

[0029] Example 4

[0030]

[0031] In a high-pressure reactor with stirring, add 2.8 g of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, 4 g of cesium carbonate and 0.23 g of palladium acetate to 150 mL of N,N-dimethylformamide. Protect the reaction system with nitrogen. After stirring, quickly add 50 mL of N,N-dimethylformamide solution containing 1.2 g of sodium tert-butoxide. Then, remove the gas in the reactor by vacuum, introduce carbon dioxide, make the pressure in the reactor reach 1.5 MPa, and slowly heat to 110°C. The temperature was maintained for 7 hours, then the temperature was lowered to 40°C, the gas in the reaction system was removed by vacuum again, hydrogen was introduced into the reactor to make the pressure in the reactor reach 2MPa, the temperature was maintained for 10 hours with stirring, the reaction liquid was filtered after the reaction was completed, the filtrate was poured into 1000 mL of water, the pH was adjusted to 6-7 with dilute hydrochloric acid under stirring, and then the reaction system was extracted with 300 mL of dichloromethane for several times, the organic phases were combined, dried, concentrated, and separated by silica gel column chromatography to obtain 2.52 g of pomalidomide.

[0032] Example 5

[0033]

[0034] In a high-pressure reactor with stirring, add 2.8 g of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, 4 g of cesium carbonate and 0.23 g of palladium acetate to 150 mL of N,N-dimethylformamide, protect the reaction system with nitrogen, stir and quickly add 50 mL of N,N-dimethylformamide solution containing 1.2 g of sodium tert-butoxide, then evacuate the gas in the reactor under vacuum, introduce carbon dioxide to make the pressure in the reactor reach 1.5 MPa, slowly heat to 110°C, maintain the temperature for 7 hours, and filter the reaction after the reaction is completed. The filtrate was poured into 300 mL of water, and the pH was adjusted to 6-7 with dilute hydrochloric acid under stirring. The reaction system was then extracted with 100 mL of dichloromethane for several times. The organic phases were combined, concentrated in vacuo, and added to 150 mL of methanol. 0.28 g of palladium carbon with a content of 10% was added. Under nitrogen protection, 5 g of hydrazine hydrate was slowly added dropwise. The reaction temperature was kept at room temperature during the addition. After the addition was completed, the reaction was continued for 7.5 hours. The reaction solution was filtered to remove the catalyst palladium carbon. After the system was concentrated in vacuo, the concentrate was separated by silica gel column chromatography to obtain 2.19 g of pomalidomide.

[0035] Example 6

[0036]

[0037] In a high-pressure reactor with stirring, add 2.8 g of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, 4 g of cesium carbonate and 0.23 g of palladium acetate to 150 mL of N,N-dimethylformamide, protect the reaction system with nitrogen, stir and quickly add 50 mL of N,N-dimethylformamide solution containing 1.2 g of sodium tert-butoxide, then evacuate the gas in the reactor under vacuum, introduce carbon dioxide to make the pressure in the reactor reach 1.5 MPa, slowly heat to 110°C, maintain the temperature for 7 hours, filter the reaction liquid after the reaction is completed, and filter The mixture was poured into 300 mL of water, and the pH was adjusted to 6-7 with dilute hydrochloric acid under stirring. The reaction system was then extracted with 100 mL of dichloromethane for several times. The organic phases were combined, concentrated, and added to a mixture of 100 mL of ethanol and 50 mL of water. 0.6 g of reduced iron powder was added, 5 mL of acetic acid was added dropwise, and the mixture was heated to 100 ° C. After the reaction was completed, the reaction solution was filtered, and the filtrate was distilled under vacuum to remove ethanol and water. 50 mL of acetone and 30 mL of benzene were added to the residue, heated, stirred, cooled, and recrystallized. The solid was filtered, and the filter cake was dried to obtain 2.31 g of pomalidomide.

[0038] Example 7

[0039]

[0040] In a stirred reaction flask, add 3 g of pomalidomide to 100 mL of hydrochloric acid, stir, add 0.9 g of sodium nitrite, stir for 30 min, maintain at 0 ° C, add 50 mL of an aqueous solution containing 0.8 g of sodium azide, continue stirring for 4 h, add 100 mL of dichloromethane to the reaction system, stir, separate the organic phase, extract the aqueous phase with 50 mL of dichloromethane for several times, combine the organic phases, and concentrate to obtain 2.87 g of pomalidomide-azide compound.

[0041] Example 8

[0042]

[0043] In a reaction flask with a stirring device, 3 g of pomalidomide-azide compound and 4.7 g of erlotinib were added to 150 mL of a mixed solvent of distilled water, tetrahydrofuran and tert-butyl alcohol (the volume ratio of the three was 1:1:1), and then 0.19 g of cuprous iodide was added. Under nitrogen protection, the mixture was heated to 80°C and refluxed for 15 h. The reaction of the raw materials was monitored by TLC to be complete. The reaction system was filtered, and then the reaction system was extracted with 50 mL of dichloromethane for several times. The organic phases were combined and dried over anhydrous magnesium sulfate. Finally, the organic phases were concentrated and separated by silica gel column chromatography to obtain 5.91 g of the product. MS (ESI + )m / z:693[M+H] + .

[0044] Example 9

[0045]

[0046] In a reaction flask with a stirring device, 3 g of pomalidomide-azide compound and 2.23 g of N-butynyl-p-methoxybenzamide were added to 100 mL of a mixed solvent of distilled water, tetrahydrofuran and tert-butanol (the volume ratio of the three was 1:1:1), and then 0.19 g of cuprous iodide was added. Under nitrogen protection, the mixture was heated to 80°C and refluxed for 11 hours. The reaction of the raw materials was monitored by TLC to ensure that the reaction was complete. The reaction system was extracted with 50 mL of dichloromethane for several times. The organic phases were combined and dried over anhydrous magnesium sulfate. Finally, the organic phases were concentrated and separated by silica gel column chromatography to obtain 4.07 g of the product. 1H NMR (400MHz, DMSO-d6) δ11.13(s,1H),8.59(s,1H),8.48(t,J1=4.0Hz,J2=8.0Hz,1H),8.1 8(t,J1=4.0Hz,J2=4.0Hz,1H),8.09(d,J=4.0Hz,2H),7.83(d,J=8.0Hz,2H),6.98(d,J=8. 0Hz, 2H), 5.17 (dd, J1=4.0Hz, J2=8.0Hz, 1H), 3.81 (s, 3H), 3.60 (dd, J1=8.0Hz, J2=8.0Hz, 2H), 3.03 (t, J1=8.0Hz, J2=4.0Hz, 2H), 2.93-2.84 (m, 1H), 2.64-2.59 (m, 1H), 2.59 (s, 2H).

[0047] Example 10

[0048]

[0049] In a reaction flask with a stirring device, 3 g of pomalidomide-azide compound and 2.1 g of N-propynyl-p-methoxybenzamide were added to 100 mL of a mixed solvent of distilled water, tetrahydrofuran and tert-butyl alcohol (the volume ratio of the three was 1:1:1), and then 0.19 g of cuprous iodide was added. Under nitrogen protection, the mixture was heated to 80°C and refluxed for 8.5 h. The reaction of the raw materials was monitored by TLC to be complete. The reaction system was extracted with 50 mL of dichloromethane for several times. The organic phases were combined and dried over anhydrous magnesium sulfate. Finally, the organic phases were concentrated and separated by silica gel column chromatography to obtain 4.42 g of the product. 1 H NMR(400MHz,DMSO-d6)δ11.12(s,1H),8.99(t,J=5.6Hz,1H),8.65(s,1H),8.25– 8.14(m,1H),8.08(d,J=4.9Hz,2H),7.88(d,J=8.8Hz,2H),7.00(d,J=8.8Hz,2H), 5.17(dd,J=12.8,5.4Hz,1H),4.64(d,J=5.7Hz,2H),3.81(s,3H),2.89(d,J=36. 3Hz, 1H), 2.61 (d, J = 17.7Hz, 1H), 2.07 (d, J = 17.7Hz, 1H), 1.29 (d, J = 35.5Hz, 1H).

[0050] Embodiment 11

[0051] We conducted an in vitro urease inhibition experiment on the three target compounds obtained: yellow cattle of about 12 months old were selected. After feeding for 1 hour, 400 mL of rumen fluid was collected through an artificial rumen fistula using a special rumen fluid collector, and filtered through 4 layers of gauze for later use. After adding the corresponding reagents to each culture tube according to the amount in the table below, 4 drops of liquid paraffin were added, and the tube was gently shaken on a (39.0±0.5)℃ constant temperature water bath oscillator. After culturing for 8 hours, some culture tubes were taken out from each group, and 4 drops of saturated mercuric chloride solution were immediately added and shaken to terminate the reaction. The ammonia nitrogen content of each tube was determined by the Kjeldahl semi-micro-saturated magnesium oxide distillation method. We can find that the inhibitory activity of the three designed compounds on urease becomes more and more significant with the extension of time, and has a very significant inhibitory effect; and through the CCK8 experiment, the IC of the human renal epithelial cell line 293T cells was inhibited. 50 The values ​​were 40.19 μM, >100 μM, and >100 μM, respectively.

[0052]

[0053]

[0054] Inhibition rate (%) = (ammonia content of control group - ammonia content of test group) ÷ ammonia content of control group × 100%

[0055] serial number Time (h) The inhibition rate of the control group Inhibition rate of test group 1 Inhibition rate of test group 2 Example 8 Product 8 0％ 36.03％ 57.16％ Example 9 Product 8 0％ 34.14％ 53.88％ Example 10 Product 8 0％ 28.99％ 49.01％

[0056] The above embodiments describe the basic principles, main features and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are only for illustrating the principles of the present invention. Without departing from the scope of the principles of the present invention, the present invention may have various changes and improvements, and these changes and improvements all fall within the scope of protection of the present invention.

Claims

1. A pomalidomide feed additive, characterized in that: The pomalidomide feed additive is selected from the following compounds: 。 2. The method for preparing a pomalidomide feed additive according to claim 1, characterized in that: The specific preparation process of the pomalidomide feed additive is as follows: a certain amount of pomalidomide-azide compound and aminoalkyne compound are added to a mixed solvent of distilled water, tetrahydrofuran and tert-butyl alcohol, anhydrous copper sulfate and reagent-grade sodium ascorbate are added, nitrogen protection is applied, the mixture is heated to 80°C, refluxed to react until the raw materials react completely, and then a dichloromethane solution containing a benzoyl chloride compound is slowly dripped into the reaction system, after the dripping is completed, a saturated sodium chloride solution is added after stirring for a period of time, an organic phase is separated after stirring, and the organic phase is concentrated to obtain the pomalidomide feed additive; the molar ratio of the azide compound to the aminoalkyne to the benzoyl chloride compound is 1:1:1; the pomalidomide-azide compound is The aminoacetylene compound is erlotinib, N-butynyl-p-methoxybenzamide or N-propynyl-p-methoxybenzamide.

3. The method for preparing a pomalidomide feed additive according to claim 2, characterized in that: The specific preparation process of the pomalidomide-azide compound is as follows: a certain amount of pomalidomide is added to hydrochloric acid, a certain amount of sodium nitrite is added after stirring, the temperature is maintained at 0°C, an aqueous solution containing sodium azide is added, stirring is continued for a period of time, dichloromethane is added to the reaction system, an organic phase is separated after stirring, the aqueous phase is extracted with dichloromethane for multiple times, the organic phases are combined, and the pomalidomide-azide compound is obtained after concentration; the molar ratio of the pomalidomide to sodium nitrite to sodium azide is 1:1.2:1.

2.

4. The method for preparing a pomalidomide feed additive according to claim 3, characterized in that: The specific preparation process of pomalidomide is as follows: in a high-pressure reactor, a certain amount of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, cesium carbonate and palladium acetate are added to N,N-dimethylformamide, the reaction system is protected by nitrogen, and after stirring, an N,N-dimethylformamide solution containing sodium tert-butoxide is quickly added, and then the gas in the reactor is removed by vacuum, and carbon dioxide is introduced to make the pressure in the reactor reach pressure 1, and then the reactor is slowly heated to 110° C., and the temperature is maintained for a period of time, and then the temperature is lowered to 40° C., and the gas in the reaction system is removed by vacuum again, and hydrogen is introduced into the reactor to make the pressure in the reactor reach pressure 2, and the temperature is maintained. The mixture is stirred for reaction at 4000 ℃, and the reaction liquid is filtered after the reaction is completed. The filtrate is poured into water, and the pH value is adjusted to 6 with dilute hydrochloric acid under stirring. Then, the reaction system is extracted with dichloromethane for multiple times, and the organic phases are combined, dried, concentrated, and separated by silica gel column chromatography to obtain pomalidomide; the molar ratio of the N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide to cesium carbonate and palladium acetate is 1:1.2:0.1; the molar ratio of the N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide to sodium tert-butoxide is 1:1-1.1; the pressure 1 is 1.0-1.5 MPa; and the pressure 2 is 1.0-2.0 MPa.

5. The method for preparing a pomalidomide feed additive according to claim 3, characterized in that: The specific preparation process of pomalidomide is as follows: in a high-pressure reactor, a certain amount of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, cesium carbonate and palladium acetate are added to N,N-dimethylformamide, the reaction system is protected by nitrogen, and after stirring, an N,N-dimethylformamide solution containing sodium tert-butoxide is quickly added, and then the gas in the reactor is removed by vacuum, and carbon dioxide is introduced to make the pressure in the reactor reach 1.5MPa, and then the reaction mixture is slowly heated to 110°C, and the temperature is maintained for a period of time, and then the reaction liquid is filtered, the filtrate is poured into water, and the pH is adjusted to 6 with dilute hydrochloric acid under stirring, and then extracted with dichloromethane. The reaction system is taken for multiple times, the organic phase is combined, and the organic phase is added to methanol after concentration, and then a certain amount of palladium carbon with a content of 10% is added. Under nitrogen protection conditions, hydrazine hydrate is slowly added dropwise, and the reaction temperature is kept at room temperature during the dropping process. After the dropping is completed, the reaction is continued for a period of time, and the catalyst palladium carbon is removed by suction filtration. After concentration, pomalidomide is obtained by separation through silica gel column chromatography; the mass ratio of the N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide to 10% palladium carbon is 10:1; the molar ratio of the N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide to hydrazine hydrate is 1:5-10.

6. The method for preparing a pomalidomide feed additive according to claim 3, characterized in that: In a high-pressure reactor with stirring, a certain amount of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide, cesium carbonate and palladium acetate are added to N,N-dimethylformamide, and the reaction system is protected by nitrogen. After stirring, N,N-dimethylformamide solution containing sodium tert-butoxide is quickly added, and then the gas in the reactor is removed by vacuum, and carbon dioxide is introduced to make the pressure in the reactor reach 1.5MPa. It is slowly heated to 110°C, and the temperature is maintained for a period of time. After that, the reaction liquid is filtered, and the filtrate is poured into water. The pH is adjusted to 6 with dilute hydrochloric acid under stirring, and then dichloromethane is used. The reaction system is extracted for multiple times, the organic phases are combined, concentrated and added to a mixture of ethanol and water, reduced iron powder is added, acetic acid is added dropwise and heated to a certain temperature, the reaction solution is filtered after the reaction is completed, the filtrate is subjected to reduced pressure distillation to evaporate ethanol and water, acetone and benzene are added to the residue for recrystallization, the solid is filtered, and the filter cake is dried to obtain pomalidomide; the volume ratio of ethanol to water is 5:2; the mass ratio of the feeding amount of N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide to reduced iron powder is 1:0.1-0.2; and the certain temperature is 80-100°C.

7. A method for preparing a pomalidomide feed additive according to any one of claims 4 to 6, characterized in that: The specific preparation process of the N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide is as follows: a certain amount of 2-nitrobenzoic acid is added into dichlorothionyl, and the reaction system is concentrated in vacuo after stirring for a period of time under nitrogen protection, the concentrate is dissolved with dichloromethane, and then a dichloromethane solution containing dimethyl 2-aminoglutarate hydrochloride is slowly added dropwise under stirring, and the stirring reaction is continued for a period of time after the addition is complete, a saturated sodium chloride solution is added to the reaction system, and the organic phase is separated after stirring, and added into a high-pressure reactor, and a certain amount of EDCI and 1-hydroxybenzotriazole and tris(III) are added into the reactor. Ethylamine, the air in the reaction kettle is removed by vacuum, ammonia is introduced into the reaction kettle to make the pressure in the kettle reach 0.5MPa, and the reaction is heated to 50°C. After the reaction is completed, the reaction liquid is filtered, and then acetic acid is used to adjust the reaction system to be neutral, water is added, and the organic phase is separated after stirring, and N-3-(2-nitrobenzene)formamide-cyclopentanedicarboxamide is obtained after concentration; the molar ratio of the feeding amount of the 2-nitrobenzoic acid to the 2-aminoglutaric acid dimethyl ester hydrochloride is 1:1-1.1; the molar ratio of the feeding amount of the 2-nitrobenzoic acid to the EDCI, 1-hydroxybenzotriazole and triethylamine is 1:1:1:

2.

8. Use of the pomalidomide feed additive according to claim 1 in the preparation of a product for inhibiting urease activity.

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