Preparation and application of germacrone skeleton-based NMDA receptor inhibitor

Abstract

The invention discloses preparation and application of an NMDA receptor inhibitor based on a germacrone skeleton, and relates to the technical field of medicines, in particular to a preparation method of germacrone derivatives shown in general formulas I and II and an effect of the derivatives as neuroprotective agents, and the main application field is cerebral ischemia resistance. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to the preparation of a germacrone derivative and the application of the derivative as a GluN2B-NMDA receptor antagonist. Background Art

[0002] Alzheimer's disease is a heterogeneous disease caused by aging, genetics and environmental factors, and is also a common neurodegenerative disease. Current studies have shown that the pathogenesis of Alzheimer's disease is related to dysfunction of the neurotransmitter system, especially in the cholinergic system, which leads to cognitive impairment and neuropsychiatric disorders in patients. Its common clinical features are progressive memory loss, cognitive dysfunction, abnormal behavioral activities, etc.

[0003] Glutamate is an important excitatory neurotransmitter, and its receptors are important excitatory amino acid receptors in the human body, including ionotropic glutamate receptors and metabotropic glutamate receptors. Ionotropic glutamate receptors can be divided into NMDA receptors, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, and kainate receptors according to different exogenous agonists.

[0004] As an important glutamate receptor, NMDA receptor can regulate the survival of neurons, and plays a very important role in the development of dendrites, axons, synaptic plasticity, the formation of neuronal circuits, and learning and memory activities. In addition, NMDA receptors are also involved in many important physiological processes of the nervous system, and are closely related to a variety of neurodegenerative diseases such as pain, stroke, epilepsy, schizophrenia, depression, and Alzheimer's disease. When NMDA receptors are overactivated, it will cause damage to neuronal cells or even apoptosis, thereby causing functional disorders of the central nervous system, and then triggering the occurrence of the above-mentioned diseases. Therefore, NMDA receptors are considered to be related receptors with great potential for the treatment of degenerative nervous system diseases and are important targets for the treatment of a variety of central nervous system diseases.

[0005] NMDA receptors are usually heterotetramers composed of two NR1 subunits and two NR2 subunits. NMDA receptors have the following characteristics that are significantly different from other glutamate receptors: 1) they are widely distributed in synapses and outside synapses; 2) they have a voltage-dependent Mg 2+ Blocking effect; 3) Ca 2+ It has high permeability; 4) The slow release of glutamate results in a slower kinetic process involving NMDA receptors; 5) The activation of NMDA receptors also requires the participation of glycine or serine in addition to glutamate; 6) The NMDA receptor itself is equipped with a series of regulatory sites, making it extremely sensitive to the extracellular microenvironment.

[0006] Most of the sesquiterpenoid compounds in the volatile oil extracted from plants have anti-inflammatory, anticancer, antiviral, analgesic and antitussive effects. As an important component extracted from turmeric volatile oil, germacron also exhibits anti-tumor, antibacterial and anti-inflammatory pharmacological activities. Different in vitro and in vivo studies have shown that germacron is an effective anticancer compound with significant pharmacological activity against breast cancer, liver cancer, prostate cancer, glioma cells and gastric cancer. It can block the cell cycle and induce apoptosis of cancer cells by regulating different intracellular signal transduction. Germacron can significantly improve the motor dysfunction and spatial learning and memory disorders of mice caused by traumatic brain injury, and can reduce neuronal apoptosis and microglial activation in a dose-related manner. It can also upregulate the expression of antioxidant protein Nrf2 and inhibit the expression of inflammatory response protein p-p65. Studies have also confirmed that germacron can significantly reduce brain water content, infarct volume and neurological deficits in rats with transient middle cerebral artery occlusion / reperfusion injury model, and the MDA content is reduced and the activities of glutathione (GSH), SOD and GSH-Px are increased after germacron treatment. Germacron can reduce Caspase-3 and Bax, and increase the expression of messenger RNA, Bcl-2 and p-Akt. Therefore, the neuroprotective effect of germacron is related to its regulation of neuroinflammation, oxidative stress and anti-apoptosis.

[0007] As a potential drug target and its core role in the central nervous system, the NMDA receptor has always attracted the attention of researchers. It is a ligand-gated, voltage-dependent ion channel that is widely present in the central nervous system. The NMDA receptor can convert specific neuronal activity patterns into long-term changes in synaptic structure and function, thereby controlling memory formation and brain plasticity. As a natural product, germacrone can exhibit multiple and obvious pharmacological effects, but germacrone has a simple structure and produces fewer interactions with target proteins, so further structural optimization is still needed to achieve the optimal neuroprotective effect of germacrone. Therefore, the study of the natural product germacrone and its derivatives has important scientific research significance, which provides a certain basis for promoting the discovery of new and innovative drugs. Summary of the invention

[0008] The purpose of the present invention is to design a new GluN2B-NMDA receptor antagonist capable of interacting with the NMDA receptor cavity based on the binding mode of the traditional selective GluN2B-NMDA receptor antagonist Ifenprodil.

[0009] Another object of the present invention is to provide a method for preparing a germacrone derivative.

[0010] In order to achieve the purpose of the present invention, the following technical scheme is adopted:

[0011] The present invention relates to a germacrone derivative having the following general formula I, II, or a pharmaceutically acceptable salt thereof, or an optically active form thereof:

[0012]

[0013] Among them, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 is any one of the following groups: (1) H atom, (2) hydroxyl group, (3) nitro group, (4) amino group, (5) halogen group, (6) cyano group, (7) halogenated C1-C6 alkyl group, (8) C1-6 alkyl group, C3-8 cycloalkyl group, C2-6 alkenyl group, C2-6 alkynyl group, C1-18 alkoxycarbonyl group, C1-6 alkoxy group, C2-6 alkenyloxy group, C2-6 alkynyloxy group, C3-8 cycloalkenyl group, (9) carbonyl group substituted by various means, any of which The halogens include fluorine, chlorine and bromine.

[0014] Preferably, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 It is any one of the following groups: H atom, hydroxyl, nitro, amino, halogen, cyano, halogenated C1-C6 alkyl, C1-6 alkyl, C3-8 cycloalkyl.

[0015] More preferably, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 It is any one of the following groups: H atom, hydroxyl, halogen, halogenated C1-C6 alkyl, C1-6 alkyl.

[0016] Optically active isomers include R and S configurations.

[0017] Most preferably, the present invention provides the following series of germeracrone derivatives:

[0018]

[0019]

[0020] The pharmaceutically acceptable salt refers to a salt formed by a compound represented by general formula I or II and an acid, wherein the acid is selected from hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, phosphoric acid, nitric acid, formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, methanesulfonic acid, ethanesulfonic acid, toluenesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, trifluoroacetic acid or aspartic acid.

[0021] The synthetic route for preparing the germacrone derivatives represented by general formula I or II of the present invention or their optically active forms or racemates, or diastereoisomer mixtures is as follows: wherein R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 are the corresponding groups at the corresponding positions of compounds 1a to 3c.

[0022] Route 1: Synthesis of 3a~3c

[0023] Using substituted or unsubstituted benzaldehyde as the starting material, a nitration reaction is carried out to obtain intermediate a4. Intermediate a4 is subjected to a reductive amination reaction with L-glutamic acid to obtain intermediate a7. Intermediate a7 is refluxed in ethanol to carry out a cyclization to obtain intermediate a10. Intermediate a10 is reacted with a catalyst FeCl 3 6H 2 O catalyzed secondary cyclization to obtain the key intermediate F1. 6 , R 7 , R 8 , R 9 are the corresponding groups at the corresponding positions of compounds 3a to 3c.

[0024]

[0025] Germacron A1 is reduced to reduce the ketone carbonyl to a hydroxyl group to obtain intermediate A2. Intermediate A2 and intermediate F1 are subjected to ester condensation under the action of a condensing agent to obtain final products 3a to 3c.

[0026]

[0027] Route 2: Synthesis of 1a to 2c

[0028] Substituted benzoic acid and alanine methyl ester salt are subjected to amide condensation under the action of a condensing agent to obtain intermediate D1, and intermediate D1 is demethylated under the action of sodium hydroxide methanol solution to obtain carboxylic acid intermediate E1. Intermediate E1 is subjected to esterification reaction with the reduction product (A2) of germacrone to obtain products 1a~2c. Alanine methyl ester salt includes D type and L type. L type is used to prepare compounds 1a~1c, and D type is used to prepare compounds 2a~2c. R 1 , R 2 , R 3 , R 4 , R 5 are the corresponding groups at the corresponding positions of compounds 1a to 2c.

[0029]

[0030] A pharmaceutical composition comprises a germacrone derivative represented by general formula I or II and one or more of its stereoisomers and pharmaceutically acceptable salts, and a pharmaceutically acceptable carrier.

[0031] The dosage form of the pharmaceutical composition is selected from one or more of the following: tablets, capsules, injections, suppositories, patches, inhalable powder preparations, suspensions, emulsions and ointments.

[0032] Use of the germacrone derivative represented by general formula I or II or its pharmaceutically acceptable salt or its optically active form or the pharmaceutical composition in the preparation of drugs for treating and / or preventing Alzheimer's disease.

[0033] Use of the germacrone derivative represented by general formula I or II or its pharmaceutically acceptable salt or its optically active form or the pharmaceutical composition in the preparation of GluN2B-NMDA receptor antagonist.

[0034] The advantages of the present invention are:

[0035] The present invention synthesizes a series of new germacrone derivatives by 1 H-NMR, 13 C-NMR and ESI-HRMS confirmed the structure of the compound, and evaluated the activity of cell protection. Through pharmacological activity tests, it was found that all germerone derivatives can improve the cell protection rate, especially compound 3a, which has the best activity and is better than the positive control drug Ifenprodil. In addition, the mechanism of action of compound 3a was determined through calcium ion influx experiments on compound 3a. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1The protection rate (%) of the compounds 1a to 3c of the present invention on the PC12 cell injury model;

[0037] Figure 2 The green fluorescence images of PC12 cells at different times in the present invention show the intracellular Ca 2+ Changes in concentration;

[0038] Figure 3 For example Ca of the present invention 2+ Fluorescence intensity curve of each group during the detection time of the influx experiment. DETAILED DESCRIPTION

[0039] The compounds of the present invention and their preparation will be better understood from the following examples, which are intended to illustrate but not to limit the scope of the present invention.

[0040] Example 1: (S)-1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline-1-carboxylic acid (F1-1)

[0041] Step A: Preparation of 2-nitrobenzaldehyde (a4)

[0042] Under ice bath conditions, potassium nitrate (3.90 g, 0.039 mol) was added to concentrated sulfuric acid (30 mL, d = 1.83 g / mL), the reaction temperature was maintained at 0-3 ° C, benzaldehyde (6.93 g, 0.037 mol) was slowly added dropwise, and the reaction was carried out at a temperature below 5 ° C for 2 hours. After the reaction was completed, the reaction solution was poured into 160 mL of ice-water mixture, and a white solid was precipitated. It was filtered and dried, and 4.89 g of white needle-shaped crystals were obtained by column chromatography (PE: EA = 30: 1), with a yield of 74.63%.

[0043] Step B: Preparation of (S)-2-(2-nitrobenzylamino)-pentanedioic acid (a7)

[0044] Dissolve NaOH (2.38 g, 0.06 mol) in 20 mL of water in an ice bath, and then slowly add L-glutamic acid (4.41 g, 0.03 mol) to the above NaOH solution in batches in an ice bath for later use. Stir and dissolve o-nitrobenzaldehyde (3.71 g, 0.02 mol) in 30 mL of anhydrous ethanol. After it is completely dissolved, slowly add the solution to the above L-glutamic acid reaction solution in an ice bath, and control the dripping time to about 1 hour. After the addition is completed, transfer the reaction solution to room temperature and continue stirring the reaction for 1 hour. Afterwards, move it back to the ice bath to cool down, take NaBH 4(0.76g, 0.02mol) was slowly added to the reaction solution in batches. After the addition was completed, the mixture was moved to room temperature and the reaction was continued to stir for about 3 hours until the reaction solution became clear. After the reaction was completed by TLC monitoring, a small amount of insoluble impurities in the reaction solution were filtered out using diatomaceous earth as a medium, and the filtrate was concentrated to a volume of about 20mL, extracted with ethyl acetate, and the organic layer was discarded. The pH of the aqueous layer was adjusted to 4-5 with 1mol / L hydrochloric acid, and a large amount of white solid was precipitated. The mixture was stirred for 30 minutes at room temperature, filtered, and the filter cake was washed twice with 30mL of water and dried to obtain a white shiny powder solid with a mass of 4.39g and a yield of 78.36%.

[0045] Step C: Preparation of (S)-1-(2-nitrobenzylamino)-5-oxopyrrolidine-2-carboxylic acid (a10).

[0046] Add (S)-2-(2-nitrobenzylamino)-pentanedioic acid (4.39 g, 0.016 mol) to anhydrous ethanol (45 mL), heat to reflux for 2 h, the reaction solution is almost clear, filter while hot to remove a small amount of insoluble matter, cool, precipitate a white solid, filter, collect the filter cake, concentrate the filtrate to about 15 mL, filter, combine the filter cakes, and obtain 4.01 g of white solid, with a yield of 97.23%.

[0047] Step D: Preparation of (S)-1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline-1-carboxylic acid (F1)

[0048] Add (S)-1-(2-nitrobenzylamino)-5-oxopyrrolidine-2-carboxylic acid (4.01 g, 0.015 mol) to anhydrous ethanol (55 mL), heat to dissolve, cool slightly, and add 0.530 g of activated carbon and a catalytic amount of FeCl 3 6H 2 O (0.20 g, 0.75 mmol), after the temperature rises to 70°C, vigorously reflux for 72 h. Filter and concentrate the filtrate to leave about 15 mL. Slowly drop 20 mL of acetone into the mixture under stirring, filter and dry to obtain 1.86 g of white solid with a yield of 56.78%.

[0049] Example 2: (S)-7-Chloro-1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline-1-carboxylic acid (F1-2)

[0050] The preparation method is the same as above.

[0051] Example 3: (S)-7-Bromo-1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline-1-carboxylic acid (F1-3)

[0052] The preparation method is the same as above.

[0053] Example 4: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane 3,7-dien-1-yl-(2S)-2-(4-hydroxybenzamide) propionate (1a)

[0054] Step A: Preparation of (S)-methyl 2-(4-hydroxybenzamide)propionate (D1)

[0055] Take 4-hydroxybenzoic acid (1.93g, 0.014mol) and dissolve it in 10mL ultra-dry DMF. In an ice-water bath, slowly add HATU (8.38g, 0.022mol), drop DIPEA (6.07mL, 0.037mol), after adding, stir and react for 15min in an ice-water bath, then move to room temperature to react for 30min, monitor the reaction by TLC, and keep a sample of the reaction solution. Then add L-alanine methyl ester hydrochloride (2.09g, 0.015mol) in an ice-water bath, stir and react for 15min, move to room temperature to continue the reaction for 2h, and monitor the reaction by TLC. After that, slowly pour the reaction solution into 100mL water, extract with DCM, combine the organic layers, extract the organic layers with saturated brine, retain the organic layers, dry and concentrate, and column chromatography (DCM: MeOH = 50: 1) to obtain 2.90g of white solid, with a yield of 89.05%.

[0056] Step B: Preparation of (S)-2-(4-hydroxybenzamide)propionic acid (E1)

[0057] At room temperature, the intermediate D1 (2.50 g, 0.011 mol) obtained above was dissolved in a 4 mol / L NaOH methanol solution, stirred for 1 h, and the reaction was monitored by TLC. Afterwards, the reaction solution was concentrated and evaporated to dryness, 15% dilute hydrochloric acid was added to adjust the pH to 3-4, and then extracted with ethyl acetate, the organic layers were combined, the organic layers were extracted with saturated brine, the organic layers were retained, dried and concentrated, and a white solid was obtained with a mass of 2.12 g and a yield of 90.32%.

[0058] Step C: Preparation of (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane-3,7-dien-1-ol (A2)

[0059] Take LiAlH 4 (0.38 g, 0.010 mol) was placed in a 50 mL eggplant-shaped bottle, and 10 mL of tetrahydrofuran (THF) was added to form a suspension. Germacron (2.00 g, 0.0092 mol) was dissolved in 5 mL of THF. Under ice bath conditions, LiAlH 4The THF solution of germacrone was added dropwise to the THF suspension, and the addition time was controlled to be about 2 minutes. After the addition was completed, the mixture was moved to room temperature and stirred for 10 minutes, and the reaction was monitored by TLC. After that, 1 mL of water, 2 mL of 15% NaOH aqueous solution, and 3 mL of water were added to the reaction solution in sequence to quench the reaction, and extracted with petroleum ether, the organic layer was retained, dried over anhydrous sodium sulfate, filtered and evaporated to obtain 1.85 g of a milky white oil, with a yield of 91.72%.

[0060] Step D: Preparation of (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane 3,7-dien-1-yl-(2S)-2-(4-hydroxybenzamide) propionate (1a)

[0061] Take the above intermediate E1 (200 mg, 0.97 mmol) and dissolve it in 10 mL of ultra-dry DCM solution. In an ice-water bath, slowly add EDCI (277 mg, 14.62 mmol) and DMAP (295 mg, 0.78 mmol). Then, stir the reaction in an ice-water bath for 15 min, then add the above intermediate A2 (214 mg, 0.97 mmol) in an ice-water bath, stir the reaction for 15 min, move to room temperature and continue the reaction for 2 h, and monitor the reaction by TLC. After that, extract with saturated brine, retain the organic layer, dry and concentrate, and column chromatography (DCM: MeOH = 50: 1) to obtain 198 mg of white powdery solid, with a yield of 49.87%.

[0062] 1 H NMR(600MHz,Chloroform-d)δ9.68(s,1H),8.70(d,J=7.2Hz,1H),7.97(dd,J=8.6,5.6Hz ,2H),7.31(t,J=8.7Hz,2H),4.79(s,1H),4.52(d,J=12.8Hz,1H),4.42(t,J=7.3Hz,1H),3 .78(d,J=12.5Hz,1H),3.22–2.47(m,2H),2.40(s,1H),2.23(d,J=50.1Hz,2H),2.02(d,J= 6.5Hz,2H),1.87(s,1H),1.90–1.65(m,6H),1.51(d,J=9.5Hz,6H),1.40(d,J=7.3Hz,3H). 13 C NMR (150 MHz, DMSO-d 6)δ170.31,163.57,162.13,160.53,142.83,135.86,131.05,128.16,127.19,126.25,125 .25,122.34,115.40(d,J=21.1Hz),64.54,44.90,34.49,30.07,24.34.ESI-HRMS:calced for C 25 H 33 NO 4 ,[M+H] + ,found 412.1845.

[0063] Example 5: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane 3,7-dien-1-yl-(2S)-2-(4-fluorobenzamide) propionate (1b)

[0064] The 4-hydroxybenzoic acid in step A of Example 4 was replaced by 4-chlorobenzoic acid. The preparation method was the same as that of Example 4, and the yield was 45.82%.

[0065] 1 H NMR(600MHz,Chloroform-d)δ8.70(d,J=7.2Hz,1H),7.97(dd,J=8.6,5.6Hz,2H),7.31 (t,J=8.7Hz,2H),4.79(s,1H),4.52(d,J=12.8Hz,1H),4.42(t,J=7.3Hz,1H),3.78(d,J =12.5Hz,1H),3.22–2.47(m,2H),2.40(s,1H),2.23(d,J=50.1Hz,2H),2.02(d,J=65.0 Hz,2H),1.87(s,1H),1.90–1.65(m,6H),1.51(d,J=94.5Hz,6H),1.40(d,J=7.3Hz,3H). 13 C NMR (150 MHz, DMSO-d 6 )δ170.20,163.68,143.21,131.54,131.09,130.21,129.09,125.65,122.82,64.61,44.74,30.12,24.25.ESI-HRMS:calced for C 25 H 32 ClNO 3 ,[M+H] + ,found 430.0929.

[0066] Example 6: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane 3,7-dien-1-yl-(2S)-2-(4-chlorobenzamide) propionate (1c)

[0067] The 4-hydroxybenzoic acid in step A of Example 4 was replaced by 4-fluorobenzoic acid. The preparation method was the same as that of Example 4, and the yield was 40.58%.

[0068] 1 H NMR(600MHz,Chloroform-d)δ8.70(d,J=7.2Hz,1H),7.97(dd,J=8.7,5.7Hz,2H),7.3 1(t,J=8.8Hz,2H),4.79(s,1H),4.52(d,J=12.8Hz,1H),4.42(t,J=7.3Hz,1H),3.78(d ,J=12.5Hz,1H),3.22–2.47(m,2H),2.40(s,1H),2.23(d,J=5.1Hz,2H),2.02(d,J=6.0 Hz,2H),1.87(s,1H),1.90–1.65(m,6H),1.51(d,J=9.5Hz,6H),1.40(d,J=7.3Hz,3H). 13 C NMR (150 MHz, DMSO-d 6 )δ170.08,163.64,158.15,131.54,130.21,128.24,127.40,126.32,124.86, 122.18,114.15,64.71,55.42,44.87,34.50,30.07,24.27.ESI-HRMS:calced for C 25 H 32 FNO 3 ,[M+H] + ,found 414.0805.

[0069] Example 7: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane 3,7-dien-1-yl-(2R)-2-(4-hydroxybenzamide) propionate (2a)

[0070] The synthesis method of compound 2a is the same as that of compound 1a, except that the intermediate L-alanine methyl ester hydrochloride is replaced by the intermediate D-alanine methyl ester hydrochloride to obtain a white powdery solid with a yield of 36.29%.

[0071] 1H NMR(600MHz,Chloroform-d)δ9.68(s,1H),8.70(d,J=7.2Hz,1H),7.97(dd,J=8.6,5.6Hz ,2H),7.31(t,J=8.7Hz,2H),4.79(s,1H),4.52(d,J=12.8Hz,1H),4.42(t,J=7.3Hz,1H),3 .78(d,J=12.5Hz,1H),3.22–2.47(m,2H),2.40(s,1H),2.23(d,J=5.1Hz,2H),2.02(d,J= 6.0Hz,2H),1.87(s,1H),1.90–1.65(m,6H),1.51(d,J=9.5Hz,6H),1.40(d,J=7.3Hz,3H). 13 CNMR (150MHz, DMSO-d 6 )δ172.47,167.81,134.51,133.85,129.75,129.47,128.79,126.51,123.20,116. 15,115.22,71.68,49.56,43.84,39.27,25.86,25.65,20.99,20.27,17.23,16.42. ESI-HRMS:calced for C 25 H 33 NO 4 ,[M+Na] + ,found434.0471.

[0072] Example 8: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane 3,7-dien-1-yl-(2R)-2-(4-chlorobenzamide) propionate (2b)

[0073] The synthesis method of compound 2b is the same as that of compound 1b, except that the intermediate L-alanine methyl ester hydrochloride is replaced by the intermediate D-alanine methyl ester hydrochloride to obtain a white powdery solid with a yield of 48.29%.

[0074] 1H NMR(600MHz,Chloroform-d)δ8.70(d,J=7.2Hz,1H),7.97(dd,J=8.6,5.6Hz,2H),7.3 1(t,J=8.7Hz,2H),4.79(s,1H),4.52(d,J=12.8Hz,1H),4.42(t,J=7.3Hz,1H),3.78(d ,J=12.5Hz,1H),3.22–2.47(m,2H),2.40(s,1H),2.23(d,J=5.1Hz,2H),2.02(d,J=6.5 Hz,2H),1.87(s,1H),1.90–1.65(m,6H),1.51(d,J=9.5Hz,6H),1.40(d,J=7.3Hz,3H). 13 C NMR (150 MHz, DMSO-d 6 )δ170.20,163.68,143.21,131.54,131.09,130.21,129.09,125.65,122.82,64.61,44.74,30.12,24.25.ESI-HRMS:calced for C 25 H 32 ClNO 3 ,[M+H] + ,found 431.0471.

[0075] Example 9: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecane 3,7-dien-1-yl-(2R)-2-(4-fluorobenzamide) propionate (2c)

[0076] The synthesis method of compound 2c is the same as that of compound 1c, except that the intermediate L-alanine methyl ester hydrochloride is replaced by the intermediate D-alanine methyl ester hydrochloride to obtain a white powdery solid with a yield of 47.54%.

[0077] 1H NMR(600MHz,Chloroform-d)δ8.70(d,J=7.2Hz,1H),7.97(dd,J=8.7,5.7Hz,2H),7.3 1(t,J=8.8Hz,2H),4.79(s,1H),4.52(d,J=12.8Hz,1H),4.42(t,J=7.3Hz,1H),3.78(d ,J=12.5Hz,1H),3.22–2.47(m,2H),2.40(s,1H),2.23(d,J=5.1Hz,2H),2.02(d,J=6.5 Hz,2H),1.87(s,1H),1.90–1.65(m,6H),1.51(d,J=9.5Hz,6H),1.40(d,J=7.3Hz,3H). 13 C NMR (150 MHz, DMSO-d 6 )δ170.08,163.64,158.15,131.54,130.21,128.24,127.40,126.32,124.86, 122.18,114.15,64.71,55.42,44.87,34.50,30.07,24.27.ESI-HRMS:calced for C 25 H 32 FNO 3 ,[M+H] + ,found 414.1013.

[0078] Example 10: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecan-3,7-dien-1-yl-(S)-1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline-1-carboxylate (3a)

[0079] Take the intermediate F1-1 (0.22g, 1.09mmol) in a 25ml eggplant-shaped bottle, dissolve it in 10mL dichloromethane (99.9%, extradry), then place the reaction bottle in an ice-water bath, weigh the condensation agent EDCI (0.21g, 1.09mmol) and DMAP (0.089g, 0.73mmol) respectively, and slowly add them to the reaction bottle under stirring. After continuing to stir and react for 20min under ice-water bath, add the intermediate A2 (0.20g, 0.91mmol) obtained above, move to room temperature and continue to stir and react for 2h, and monitor the reaction by TLC. After that, the reaction solution is extracted with saturated brine, the organic layer is retained, dried over anhydrous calcium chloride, filtered and concentrated, and purified by column chromatography (PE:EA=10:1) to obtain white needle crystals with a yield of 31.34%.

[0080] 1H NMR(600MHz,Chloroform-d)δ7.30(d,J=7.8Hz,2H),7.17(s,1H),6.99(d,J=7.8Hz,1H),4.86(d,J=8.2Hz,1 H),4.71(d,J=14.3Hz,1H),4.52(d,J=12.8Hz,1H),3.78(d,J=12.5Hz,1H),3.42(s,1H),3.16–2.81(m,1H), 2.58(d,J=6.1Hz,1H),2.46(d,J=5.0Hz,1H),2.40(s,2H),2.36(d,J=8.6Hz,1H),2.23(d,J=5.1Hz,1H),2.0 8(s,2H),1.97(s,2H),1.87(s,1H),1.78(d,J=3.3Hz,1H),1.69(d,J=9.1Hz,3H),1.59(s,3H),1.43(s,4H). 13 C NMR (150 MHz, DMSO-d 6 )δ170.31,163.89,143.36,139.74,131.07,129.22,129.13,128.76,126.57,125.72,64.56,44.79,35.40,30.11,24.34.ESI-HRMS:calced forC 27 H 34 N 2 O 2 ,[M+H] + ,419.0701,found 419.0749.

[0081] Example 11: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecan-3,7-dien-1-yl-(S)-7-chloro-1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline-1-carboxylate (3b)

[0082] The synthesis method of compound 3b is the same as that of compound 3a, except that intermediate F1-1 is replaced by intermediate F1-2. Column chromatography (DCM:MeOH=20:1) is used to obtain a brown-yellow powdery solid with a yield of 35.62%.

[0083] 1H NMR(600MHz,Chloroform-d)δ7.33–7.24(m,2H),6.95(d,J=8.3Hz,1H),4.79(s,1H),4.65(d,J=14.4Hz,1H) ,4.52(d,J=12.8Hz,1H),4.20(d,J=14.4Hz,1H),4.11(t,J=7.2Hz,1H),3.78(d,J=12.5Hz,1H),3.22–2.47(m ,2H),2.40(s,1H),2.56(ddd,J=11.4,6.3,3.3Hz,1H),2.49–2.41(m,1H),2.40(s,1H),2.36(dt,J=17.5,9.0 Hz,1H),2.23(d,J=5.1Hz,2H),2.02(d,J=6.5Hz,2H),1.87(s,1H),1.90–1.65(m,6H),1.51(d,J=9.5Hz,6H). 13 C NMR (150 MHz, DMSO-d 6 )δ169.48,163.96,139.69,129.22,128.41,126.74,123.07,121.42,65.32,44.90,35.36,29.89,24.41.ESI-HRMS:calced for C 27 H 33 C1N 2 O 2 ,[M+Na] + ,476.0925,found 476.0991.

[0084] Example 12: (3E,7E)-3,7-dimethyl-10-isopropylidene-cyclodecan-3,7-dien-1-yl-(S)-7-bromo-1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline-1-carboxylate (3c)

[0085] The synthesis method of compound 3c is the same as that of compound 3a, except that intermediate F1-1 is replaced by intermediate F1-3. The product is purified by column chromatography (DCM:MeOH=20:1) to obtain a brown powdery solid with a yield of 28.54%.

[0086] 1H NMR(600MHz,Chloroform-d)δ7.32–7.24(m,2H),6.95(d,J=8.4Hz,1H),4.86(d,J=8.2Hz,1H),4.65(d,J=14.4Hz,1H),4.52(d ,J=12.8Hz,1H),4.20(d,J=14.4Hz,1H),4.11(dd,J=7.9,6.4Hz,1H),3.78(d,J=12.5Hz,1H),3.16–2.81(m,1H),3.16–2.81(m ,1H),2.56(ddd,J=10.2,5.9,2.2Hz,1H),2.44(dd,J=10.2,4.5Hz,1H),2.40–2.31(m,3H),2.23(d,J=5.1Hz,1H),2.04(dtd,J =13.0,8.8,8.3,4.4Hz,1H),1.97(s,2H),1.87(s,1H),1.78(d,J=3.3Hz,1H),1.69(d,J=9.1Hz,3H),1.59(s,3H),1.43(s,4H). 13 C NMR (150 MHz, DMSO-d 6 )δ170.30,163.68,143.25,135.87,131.09,129.11,125.68,122.82,115.47,64.57,44.76,40.41,34.49,30.10,24.32.ESI-MS:calced for C 27 H 33 Bn 2 O 2 ,[M+H] + ,498.3,found 498.3.

[0087] Example 13: Pharmacological studies of the compounds of the present invention

[0088] (1) In vitro activity test of compounds

[0089] PC12 cells in logarithmic growth phase were seeded in 96-well plates at a cell density of 5000 cells / well and a volume of 100 μL / well. The cells were cultured for 24 h. The test group was added with compounds at final concentrations of 0.05, 0.5, and 5 μM / L, and the blank group and NMDA injury model group were added with an equal volume of blank culture medium. After 6 h, the culture medium was discarded and replaced with HEPES buffer (1×HMF, Mg 2+-free, CC0074, Leagene, China), the test group and the model group were added with HEPES buffer with a final concentration of 2mM NMDA, and the blank group was added with normal HEPES buffer for 30min. The buffer was discarded and replaced with normal culture medium for 12h. 10μL MTT solution (concentration of 5mg / mL) was added to the cell fluid and incubated at 37°C for 4h. The supernatant was carefully aspirated, discarded, and 150μL / well DMSO was added. After shaking and dissolving, the OD value was measured at 490nm using an ELIX 800, Bio-TEK instruments, Inc., and the cell protection rate was calculated. At the same time, Ifenprodil was used as a positive control.

[0090] Cell protection rate: (1-(OD test-OD model) / (OD blank-OD model))×100%

[0091] Test results such as Figure 1 As shown, it can be seen that germacron has certain neuroprotective activity at low concentrations, but has more obvious cytotoxicity at high concentrations; when the hydroxyl group of germacron is connected to other structural formulas, the neuroprotective activity of the compound is significantly better than that of the control drug Ifenprodil (3a~3c); when H, Br or Cl atoms are introduced to the benzene ring in the 1,2,3,9-tetrahydropyrrolo[2,1-b]quinazoline structure, the activity of the compound is not much different. Among them, when there is no substituent on the benzene ring, the neuroprotective activity of compound 3a is the best, which can reach 29.63% at low concentrations; when the substituent of the hydroxyl group of germacron is a benzoic acid connected to alanine, the neuroprotective activity is increased to a certain extent compared with germacron (1a~2c); from the effect of different R / S configurations of the compound on the activity of the compound, it is found that the configuration of the connecting fragment alanine has little effect on the neuroprotective activity of the compound.

[0092] (2) Ca of compound 3a 2+ Inflow experiment

[0093] PC12 cells in logarithmic growth phase (density 6.0×10 4 After culturing for 24 h in a 3.5 cm confocal plate, the cells were treated with or without 0.05 μM of the test compound (Ifenprodil or compound 3a obtained above) for 6 h. Fresh serum-free DMEM medium containing Fluo-4AM (2.5 μM, S1060, Beyotime, China) was added and incubated for another 0.5 h. The medium was discarded and replaced with HEPES buffer (1×HMF, Mg 2+-free, CC0074, Leagene, China) were used to wash the cells twice. After adding HEPES buffer (1 ml), the cells were scanned with a laser scanning confocal microscope (LSMT-PMT, Observer Z1, Zeiss) for 50 s until the baseline was stable, and NMDA (final concentration of 500 μM) or an equal amount of HEPES buffer was added, and the fluorescence intensity changes were detected by scanning for 10 min.

[0094] The test results are as follows Figure 2 As shown, the cytoplasmic Ca 2+ The concentration was stable during the detection time, while after adding NMDA, the cytoplasmic Ca 2+ The concentration of intracellular Ca increased significantly after pretreatment with Ifenprodil and 3a. 2+ Inflows have significantly decreased. Figure 3 The curve shows that compound 3a exhibits a similar inhibitory effect to Ifenprodil. 2+ The overall effect of Inflow is better than Ifenprodil.

Claims

1. A germeracone derivative represented by general formula I or II or a pharmaceutically acceptable salt thereof or an optically active form thereof: in, R1, R2, R3, R4, R5, R6, R7, R8, and R9 are any of the following groups: (1) an H atom, (2) a hydroxyl group, (3) a nitro group, (4) an amino group, (5) a halogen group, (6) a cyano group, (7) a halogenated C1-C6 alkyl group, (8) a C1-6 alkyl group, a C3-8 cycloalkyl group, a C2-6 alkenyl group, a C2-6 alkynyl group, a C1-18 alkoxycarbonyl group, a C1-6 alkoxy group, a C2-6 alkenyloxy group, a C2-6 alkynyloxy group, a C3-8 cycloalkenyl group, (9) (10) substituted carbonyl, which is arbitrarily substituted by one or more substituents selected from H atom, hydroxyl, C1-6 alkyl, amino, C1-6 alkylamino, C1-6 alkoxy and C3-8 cycloalkyl; (11) substituted amino, which is arbitrarily substituted by one or more substituents selected from H atom, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C2-6 alkenylsulfonyl, C1-6 alkylcarbonyl, C2-6 alkenylcarbonyl and C2-6 alkynylcarbonyl; the halogens include fluorine, chlorine and bromine.

2. The germacrone derivative or its pharmaceutically acceptable salt or its optically active form according to claim 1, characterized in that: R1, R2, R3, R4, R5, R6, R7, R8, and R9 are any one of the following groups: H atom, hydroxyl, nitro, amino, halogen, cyano, halogenated C1-C6 alkyl, C1-6 alkyl, or C3-8 cycloalkyl.

3. The germacrone derivative or its pharmaceutically acceptable salt or its optically active form according to claim 2, characterized in that: R1, R2, R3, R4, R5, R6, R7, R8, and R9 are any one of the following groups: H atom, hydroxyl, halogen, halogenated C1-C6 alkyl, and C1-6 alkyl.

4. The germacrone derivative or a pharmaceutically acceptable salt thereof or an optically active form thereof according to claim 1, characterized in that: The pharmaceutically acceptable salt refers to a salt formed by a compound represented by general formula I or II and an acid, wherein the acid is selected from: hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, phosphoric acid, nitric acid, formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, methanesulfonic acid, ethanesulfonic acid, toluenesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, trifluoroacetic acid or aspartic acid; optically active forms, isomers include R and S configurations.

5. The germacrone derivative or a pharmaceutically acceptable salt thereof or an optically active form thereof according to claim 1, characterized in that: Selected from the following compounds:

6. The method for preparing the germacrone derivative or its pharmaceutically acceptable salt or its optically active form according to claim 5, characterized in that: The following steps are involved: Route 1: Synthesis of 3a~3c Using substituted or unsubstituted benzaldehyde as a starting material for nitration reaction to obtain intermediate a4, intermediate a4 and L-glutamic acid undergo reductive amination reaction to obtain intermediate a7, intermediate a7 undergoes cyclization under reflux in ethanol to obtain intermediate a10, intermediate a10 undergoes secondary cyclization under the catalysis of FeCl3·6H2O to obtain key intermediate F1, R6, R7, R8, R9 are corresponding groups at corresponding positions of compounds 3a to 3c; Germacron A1 is reduced to reduce the ketone carbonyl to a hydroxyl group to obtain intermediate A2, and intermediate A2 is subjected to ester condensation with intermediate F1 under the action of a condensing agent to obtain final products 3a to 3c; Route 2: Synthesis of 1a to 2c Substituted benzoic acid and alanine methyl ester salt are subjected to amide condensation under the action of a condensing agent to obtain an intermediate D1, intermediate D1 is demethylated under the action of a sodium hydroxide methanol solution to obtain a carboxylic acid intermediate E1, intermediate E1 is subjected to an esterification reaction with a reduction product of germacrone to obtain products 1a-2c; alanine methyl ester salt includes D type and L type, L type is used to prepare compounds 1a-1c, and D type is used to prepare compounds 2a-2c; R1, R2, R3, R4, and R5 are corresponding groups at corresponding positions of compounds 1a-2c; 7. A pharmaceutical composition, characterized in that It comprises one or more of the germacrone derivatives represented by general formula I or II and stereoisomers thereof, pharmaceutically acceptable salts, and pharmaceutically acceptable carriers.

8. The pharmaceutical composition according to claim 7, characterized in that The dosage form is selected from the group consisting of tablets, capsules, injections, suppositories, patches, inhalable powder preparations, suspensions, emulsions and ointments.

9. Use of the germacrone derivative according to claim 1 or its pharmaceutically acceptable salt or its optically active form or the pharmaceutical composition according to claim 7 in the preparation of a drug for treating and / or preventing Alzheimer's disease.

10. Use of the germacrone derivative according to claim 1 or its pharmaceutically acceptable salt or its optically active form or the pharmaceutical composition according to claim 7 in the preparation of a GluN2B-NMDA receptor antagonist.

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