A new compound for treating neurological diseases

Abstract

The present invention belongs to the field of medical technology, and is a compound having a structure of formula I and a pharmaceutically acceptable salt thereof, wherein R1 and R2 are the same or different and are selected from hydrogen, hydroxyl, C1-6 alkyl, C1-6 alkoxy, halogen atom; R3 is hydrogen, C1-6 alkyl, phenyl, benzyl, phenethyl. The present invention also relates to a method for preparing the above-mentioned compound, and discloses a pharmaceutical composition having the compound or a pharmaceutically acceptable salt thereof as an active ingredient, and their use in preparing a drug for treating ischemic stroke.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and more specifically, relates to a class of compounds with neuron damage protection effects, a preparation method thereof, a pharmaceutical composition containing the compounds, and use thereof as drugs for nervous system diseases. Background Art

[0002] Stroke has become one of the leading causes of death in my country, and ischemic stroke accounts for more than 80% of strokes, with the characteristics of high disability and high mortality. Acute ischemic stroke is a common cerebrovascular disease and one of the main causes of death and disability. It is caused by blockage of cerebral blood vessels, which leads to reduced or interrupted blood flow downstream of the blocked blood vessels, causing local brain tissue ischemia and hypoxia, until the ischemic and hypoxic parts are irreversibly damaged or even the patient dies.

[0003] Its main symptoms include sudden numbness and weakness in the face, arms, or legs (usually on one side of the body); sudden slurred speech, difficulty understanding, vision loss, loss of balance, or difficulty moving; and severe headaches and dizziness without warning.

[0004] The treatment of acute ischemic stroke mainly includes thrombolytic therapy and interventional therapy, as well as symptomatic supportive treatment. Early identification and treatment are the key to improving prognosis.

[0005] The main types of drugs used to treat acute ischemic stroke are:

[0006] Antiplatelet drugs: For most patients with acute ischemic stroke, antiplatelet drugs are used as monotherapy. For example, aspirin can be given orally as soon as possible after the onset of the disease. For those who cannot tolerate aspirin, antiplatelet drugs such as clopidogrel can be used.

[0007] Anticoagulants: They are mainly used for the early treatment of acute cerebral ischemia in patients with ischemic stroke who are in a hypercoagulable state. They include oral anticoagulants such as warfarin, dicoumarol, diphenhydramine, etc., and non-oral anticoagulants such as heparin. Among them, the bleeding risk of low molecular weight heparin is lower than that of ordinary heparin.

[0008] Neuroprotectants: used to prevent and treat the sequelae of ischemic stroke and reduce mortality.

[0009] Thrombolytic drugs: such as tenecteplase. Studies have shown that the effectiveness and safety of intravenous tenecteplase are similar to those of alteplase.

[0010] The drugs currently being studied for neuroprotection mainly include glutamate receptor antagonists, free radical scavengers, calcium channel antagonists, γ-aminobutyric acid (GABA) agonists, cytidine diphosphate choline, inflammatory response inhibitors, estrogen, erythropoietin (EPO), etc. Some experiments have shown that they have certain neuroprotective effects, and their clinical effects need further study. It is urgently needed to develop new drugs as neuroprotective agents for the treatment of acute ischemic stroke. Summary of the invention

[0011] The technical problem to be solved by the present invention is to develop new compounds with neuroprotective effects for preparing drugs for treating acute ischemic stroke and other nervous system diseases.

[0012] To solve the above problems, the present invention provides a compound having a structure of Formula I and a pharmaceutically acceptable salt thereof:

[0013]

[0014] in:

[0015] R1 and R2 are the same or different and are selected from hydrogen, hydroxyl, C1-6 alkyl, C1-6 alkoxy, and halogen;

[0016] R3 is: hydrogen, C 1-6 Alkyl, phenyl, benzyl, phenethyl.

[0017] The compounds of the present invention having a structure of formula I and pharmaceutically acceptable salts thereof, preferably the compounds of formula I are numbered from top to bottom as follows: I.1, I.2, I.3, I.4:

[0018]

[0019]

[0020]

[0021]

[0022] The compound of formula I of the present invention and its pharmaceutically acceptable salt, the pharmaceutically acceptable salt refers to: the compound and inorganic acid, organic acid salt. In particular, the pharmaceutically acceptable salt of the present invention refers to: hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, phosphate, acetate, propionate, butyrate, lactate, methanesulfonate, p-toluenesulfonate, maleate, benzoate, succinate, tartrate, citrate, fumarate, taurate, gluconate.

[0023] The present invention also relates to a method for preparing a compound of formula I, wherein raw materials II and III react at 0°C to room temperature in the presence of NaH and THF to generate intermediate IV, intermediate IV and intermediate V react at 80-100°C for 8-12h in the presence of toluene, tetrakis(triphenylphosphine)palladium and potassium carbonate, water is added to quench the reaction, and compound I is finally obtained after extraction, washing and purification.

[0024]

[0025]

[0026] Wherein R1, R2, and R3 are as described in claim 1, and M is a methylene group.

[0027] The reaction formula for preparing compound I from intermediate IV and intermediate V is as follows:

[0028]

[0029] The present invention also relates to a pharmaceutical composition for treating a nervous system disease, comprising a therapeutically effective amount of any one of the compounds having a structure of formula I or a pharmaceutically acceptable salt thereof and one or more pharmaceutical carriers, wherein the nervous system disease is acute ischemic stroke.

[0030] In addition, the present invention also relates to the use of the compound having the structure of formula I and its pharmaceutically acceptable salt in preparing a drug for treating a nervous system disease, wherein the nervous system disease is acute ischemic stroke.

[0031] Although the compounds of the present invention can be directly administered without any formulation, the compounds are preferably used in the form of pharmaceutical preparations, and the administration routes may be parenteral routes (eg, intravenous, intramuscular) and oral administration.

[0032] The pharmaceutical composition of the compounds of this invention is prepared as follows: using standard and conventional technology, the compounds of this invention is combined with acceptable solid or liquid carriers in pharmaceutics, and it is arbitrarily combined with acceptable adjuvants and excipients in pharmaceutics to be prepared into microparticles or microspheres. Solid dosage forms include tablets, dispersed particles, capsules, sustained-release tablets, sustained-release pellets, etc. Solid carriers can be at least one material, which can serve as diluents, flavoring agents, solubilizers, lubricants, suspending agents, adhesives, disintegrants and wrappers. Inert solid carriers include magnesium phosphate, magnesium stearate, sucrose, lactose, pectin, propylene glycol, polysorbate 80, dextrin, starch, gelatin, cellulose materials such as methylcellulose, microcrystalline cellulose, low melting point paraffin, polyethylene glycol, mannitol, cocoa butter, etc. Liquid dosage forms include solvents, suspensions such as injections, powders, etc.

[0033] The amount of active ingredient (compound of the present invention) contained in the pharmaceutical composition and unit dosage form can be specifically used according to the patient's condition and the doctor's diagnosis, and the amount or concentration of the compound used can be adjusted within a wide range.

[0034] The compound having the structure of formula I or a pharmaceutically acceptable salt thereof of the present invention has a significant neuroprotective effect and is of great significance for acute ischemic stroke and other nervous system diseases. DETAILED DESCRIPTION

[0035] The following will be referred to in detail in various embodiments of the present invention. Although the present invention is described in conjunction with exemplary embodiments, it should be understood that this specification is not intended to limit the present invention to these exemplary embodiments. On the contrary, the present invention is not only intended to cover these exemplary embodiments, but also to cover various substitutions, modifications, equivalent forms and other embodiments included in the essence and scope of the present invention defined by the appended claims. Unless otherwise specified, the embodiments of the present invention use raw materials that are commercially available or synthesized by conventional methods.

[0036] Example 1

[0037] p-Iodophenol (500 mg, 2.27 mmol) was dissolved in dry tetrahydrofuran (10 mL), and NaH (60%, dispersed in mineral oil) (60 mg, 2.5 mmol) was added at 0°C. After the bubbles disappeared, MOMCl (190 μL, 2.5 mmol, i.e., chloromethyl methyl ether, commercially available) was added dropwise and reacted at room temperature for 0.5 hours. After the reaction was completed by TLC detection, the reaction solution was poured into a saturated saline solution and extracted with ethyl acetate. The organic phase was washed with saturated saline, dried over anhydrous sodium sulfate, filtered to remove the desiccant, concentrated under reduced pressure, and chromatographed on a silica gel column. The crude product was chromatographed on a silica gel column (petroleum ether: ethyl acetate = 6: 1) to obtain a white solid compound 1 (i.e., intermediate IV mentioned above, 410 mg, yield 68%).

[0038]

[0039] Structural characterization data of the product: 1 H NMR (500 MHz, DMSO- d 6 ): δ 7.60 (d, J = 9.0 Hz, 2H),6.86 (d, J = 9.0 Hz, 2H), 5.17 (s, 2H), 3.36 (s, 3H). 13C NMR (125 MHz, DMSO- d 6 ): δ 157.2, 138.6, 138.6, 119.4, 119.4, 94.3, 85.1, 56.2. HR-ESI-MS m / z : [M+H] + Calcd for C 8 H 9 IO 2 263.9647; Found 264.0625.

[0040] Example 2

[0041] Compound 1 (100 mg, 0.38 mmol) was dissolved in toluene (10 mL), and 3,5-difluorophenylboric acid (90 mg, 0.57 mmol), tetrakistriphenylphosphine palladium (8.8 mg, 0.0076 mmol), and potassium carbonate (131 mg, 0.95 mmol) were added under nitrogen protection, and the mixture was reacted at 100°C for 12 hours. After the reaction was completed by TLC, water (20 mL) was added to quench the reaction, and the mixture was extracted with dichloromethane (3×30 mL). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, the desiccant was filtered out, and the mixture was concentrated under reduced pressure. The crude product was subjected to silica gel column chromatography (petroleum ether: ethyl acetate = 10: 1) to obtain white solid compound 3 (i.e., compound Ⅰ.4 mentioned above, 45 mg, yield 45%).

[0042]

[0043] Structural characterization data of the product: 1H NMR (500 MHz, CDCl3): δ 8.00 (d, J = 2.0 Hz,1H), 7.90 (d, J = 8.0 Hz, 1H), 7.88 (d, J = 8.0 Hz, 1H), 7.85 (d, J = 8.0 Hz,1H), 7.71 (d, J = 8.5 Hz, 2.0Hz, 1H),7.66 (d, J = 9.0 Hz, 2.0Hz, 2H), 7.45(m, 2H), 7.17 (d, J = 9.0 Hz, 2.0Hz, 2H), 5.24 (s, 2H), 3.53 (s, 3H). 13C NMR (125 MHz, CDCl3): δ 157.0, 138.2, 135.0, 133.8, 132.5, 128.6, 128.6, 128.5,128.2, 127.7, 126.4, 125.8, 125.6, 125.3, 116.7, 116.7, 94.6, 56.2. HR-ESI-MSm / z: [M+H]+ Calcd for C18H16O2 264.3240; Found 264.1150.

[0044] Embodiment 3-9:

[0045] Referring to the methods of Examples 1 and 2, the compound of formula I of the present invention was prepared, and the specific contents are shown in Table 1:

[0046]

[0047]

[0048] The structural characterization data of the product obtained in Example 3 are:

[0049] 1H NMR (500 MHz, CDCl3): δ 2.16-2.35 (4H, 2.24 (dddd, J = 14.7, 6.9,4.1, 2.9 Hz), 2.26 (dddd, J = 14.6, 8.4, 6.9, 2.9 Hz)), 5.35 (2H, s), 5.48(1H, ddd, J = 11.1, 5.6, 2.5 Hz), 6.05 (1H, dd, J = 8.6, 8.1 Hz), 6.50 (1H,d, J = 11.1 Hz), 6.87-7.39 (9H, 6.93 (tt, J = 7.9, 1.3 Hz), 6.99(dtd, J =8.2, 1.4, 0.5 Hz), 7.09 (ddd, J = 8.8, 1.2, 0.5 Hz), 7.21 (ddd, J = 8.8, 1.9,0.5 Hz), 7.32 (dddd, J = 8.2, 7.9, 1.4, 0.5 Hz)).13C NMR (125 MHz, CDCl3): δ22.1-22.4 (2C, 22.2 (s), 22.3 (s)), 94.8 (1C, s), 114.3 (2C, s), 116.7 (2C,s), 123.1 (1C, s), 127.1 (1C, s), 127.8(1C, s), 127.9 (1C, s), 129.4 (2C, s),130.0 (2C, s), 135.5 (1C, s), 147.2 (1C, s), 162.3-162.4 (2C, 162.3 (s),162.3 (s)).

[0050] HR-ESI-MS m / z: [M+H]+ Calcd for C19H16O2276.1150; Found 275.9047.

[0051] The structural characterization data of the product obtained in Example 4 are:

[0052] 1H NMR (500 MHz, CDCl3): δ 2.16-2.35 (4H, 2.24 (dddd, J = 14.7, 6.9,4.1, 2.9 Hz), 2.26 (dddd, J = 14.6, 8.4, 6.9, 2.9Hz)), 5.10 (2H, s), 5.48(1H, ddd, J = 11.1, 5.6, 2.5 Hz), 6.05 (1H, dd, J = 8.6, 8.1 Hz), 6.50 (1H,d, J = 11.1 Hz), 7.07 (2H, ddd, J = 8.8, 1.1, 0.5 Hz),7.20 (2H, ddd, J = 8.8,1.9, 0.5 Hz).13C NMR (125 MHz, CDCl3): δ 22.1-22.4 (2C, 22.2 (s), 22.3 (s)), 101.0 (1C, s), 114.3 (2C, s), 123.1 (1C, s), 127.1 (1C, s), 127.9 (1C, s),130.0 (2C, s), 135.5 (1C,s), 147.2 (1C, s), 162.3 (1C, s).HR-ESI-MS m / z: [M+H]+Calcd for C13H12O2 200.0837; Found 199.8305.

[0053] The structural characterization data of the product obtained in Example 5 are:

[0054] 1H NMR (500 MHz, CDCl3): δ 2.16-2.35 (4H, 2.24 (dddd, J = 14.7, 6.9,4.1, 2.9 Hz), 2.26 (dddd, J = 14.6, 8.4, 6.9, 2.9Hz)), 3.26 (3H, s), 5.10(2H, s), 5.48 (1H, ddd, J = 11.1, 5.6, 2.5 Hz), 6.05 (1H, dd, J = 8.6, 8.1Hz), 6.50 (1H, d, J = 11.1 Hz), 7.02-7.27 (4H, 7.08 (ddd,J = 8.8, 1.2, 0.5Hz), 7.20 (ddd, J = 8.8, 1.9, 0.5 Hz)).13C NMR (125 MHz, CDCl3): δ 22.1-22.4(2C, 22.2 (s), 22.3 (s)),56.3 (1C, s), 95.2 (1C, s), 114.3 (2C, s), 123.1(1C, s), 127.1 (1C, s), 127.9 (1C, s), 130.0 (2C, s), 135.5 (1C, s), 147.2(1C, s), 162.3 (1C, s).HR-ESI-MSm / z: [M+H]+ Calcd for C14H14O2 214.0994 ;Found213.8536.

[0055] The structural characterization data of the product obtained in Example 6 are:

[0056] 1H NMR (500 MHz, CDCl3): δ 2.31-2.48 (2H, 2.40 (ddd, J = 15.2, 10.0,2.0 Hz), 2.39 (ddd, J = 15.2, 4.0, 3.9 Hz)), 4.34(1H, ddd, J = 10.3, 3.9, 2.0Hz), 4.54-4.64 (2H, 4.59 (s), 4.59 (s)), 5.11-5.21 (2H, 5.16 (s), 5.16 (s)), 5.53 (1H, ddd, J = 11.0, 10.0, 4.0 Hz), 6.05 (1H, d,J = 10.3 Hz), 6.54 (1H,d, J = 11.0 Hz), 7.03-7.27 (4H, 7.09 (ddd, J = 8.8, 1.1, 0.5 Hz), 7.21 (ddd,J = 8.8, 1.9, 0.5 Hz)), 7.28-7.50 (5H, 7.34 (tt, J =7.5, 1.2 Hz), 7.35 (dddd,J = 7.8, 7.5, 1.6, 0.5 Hz), 7.44 (dddd, J = 7.8, 1.2, 0.9, 0.5 Hz)).13C NMR (125 MHz, CDCl3): δ 34.3(1C, s), 68.3 (1C, s), 70.0 (1C, s), 93.3 (1C, s),114.3 (2C, s), 124.9 (1C, s), 127.3 (1C, s), 127.7-127.8 (3C, 127.7 (s),127.8 (s)), 127.9 (1C, s), 128.4(2C, s), 130.0 (2C, s), 135.5 (1C, s), 136.5(1C, s), 147.2 (1C, s), 162.3 (1C, s).HR-ESI-MS m / z: [M+H]+ Calcd forC20H18O3 306.1256; Found305.8790.

[0057] The structural characterization data of the product obtained in Example 7 are:

[0058] 1H NMR (500 MHz, CDCl3): δ 1.25 (3H, t, J= 7.0 Hz), 2.34-2.50 (2H,2.42 (ddd, J = 15.1, 7.8, 2.0 Hz), 2.42 (ddd, J = 15.1, 4.0, 3.9 Hz)), 3.39-3.51 (2H, 3.45 (q, J = 7.0 Hz), 3.45 (q, J = 7.0Hz)), 4.36 (1H, ddd, J = 8.3,3.9, 2.0 Hz), 4.54-4.64 (2H, 4.59 (s), 4.59 (s)), 5.11-5.21 (2H, 5.16 (s),5.16 (s)), 5.55 (1H, ddd, J = 10.9, 7.8, 4.0 Hz), 6.07(1H, d, J = 8.3 Hz), 6.54 (1H, d, J = 10.9 Hz), 7.03-7.27 (4H, 7.09 (ddd, J = 8.8, 1.1, 0.5 Hz),7.21 (ddd, J = 8.8, 1.9, 0.5 Hz)), 7.28-7.50 (5H, 7.34 (tt,J = 7.5, 1.2 Hz),7.35 (dddd, J = 7.8, 7.5, 1.6, 0.5 Hz), 7.44 (dddd, J = 7.8, 1.2, 0.9, 0.5Hz)).13C NMR (125 MHz, CDCl3): δ 14.2(1C, s), 34.3 (1C, s), 64.6 (1C, s), 70.0 (1C, s), 72.1 (1C, s), 93.3 (1C, s), 114.3 (2C, s), 124.9 (1C, s), 127.3(1C, s), 127.7-127.8 (3C, 127.7 (s), 127.8(s)), 127.9 (1C, s), 128.4 (2C, s), 130.0 (2C, s), 135.5 (1C, s), 136.5 (1C, s), 147.2 (1C, s), 162.3 (1C, s).HR-ESI-MS m / z: [M+H]+ Calcd for C22H22O3 334.1569; Found333.8590.

[0059] The structural characterization data of the product obtained in Example 8 are:

[0060] 1H NMR (500 MHz, CDCl3): δ 0.71-0.83 (6H, 0.77 (q, J = 8.1 Hz), 0.77(q, J = 8.1 Hz)), 1.19-1.31 (6H, 1.25 (t, J = 7.0Hz), 1.25 (t, J = 7.0 Hz),1.25 (t, J = 7.0 Hz), 1.25 (t, J = 7.0 Hz)), 2.50-2.67 (2H, 2.57 (dd, J =13.5, 3.9 Hz), 2.60 (dd, J = 13.5, 2.0 Hz)), 3.40-3.58 (4H, 3.46 (q, J = 7.0Hz), 3.52 (q, J = 7.0 Hz), 3.52 (q, J = 7.0 Hz)), 4.44 (1H, ddd, J = 9.3,3.9, 2.0 Hz), 5.08-5.17 (2H, 5.13 (s), 5.13 (s)), 6.05 (1H, d, J = 9.3 Hz), 6.28 (1H, s), 7.03-7.15 (2H, 7.09 (ddd, J = 8.8, 1.1, 0.5 Hz), 7.09 (ddd, J =8.8, 1.1, 0.5 Hz)), 7.22 (2H, td, J = 4.7, 1.9 Hz).13CNMR (125 MHz, CDCl3): δ14.1-14.3 (2C, 14.2 (s), 14.2 (s)), 20.1 (2C, s), 29.2 (1C, s), 64.3 (1C, s), 64.6 (1C, s), 72.1 (1C, s), 93.3 (1C, s), 114.3 (2C, s), 124.9 (1C, s), 130.0(2C, s), 132.0 (1C, s), 135.5 (1C, s), 139.0 (1C, s), 147.2 (1C, s), 162.3(1C, s).HR-ESI-MSm / z: [M+H]+ Calcd for C17H19ClO3 306.1023;Found306.8840.

[0061] The structural characterization data of the product obtained in Example 9 are:

[0062] 1H NMR (500 MHz, CDCl3): δ 2.57-2.75 (2H, 2.64 (dd, J = 14.9, 2.0Hz), 2.68 (dd, J = 14.9, 3.9 Hz)), 4.45 (1H, ddd, J =11.2, 3.9, 2.0 Hz), 5.30-5.40 (2H, 5.35 (s), 5.35 (s)), 5.96 (1H, d, J = 11.2 Hz), 6.27 (1H, s), 6.87-7.05 (5H, 6.93 (tt, J = 7.9, 1.3 Hz), 6.94 (ddd, J =8.8, 1.3, 0.5 Hz),6.99 (dtd, 13C NMR (125MHz, CDCl3): δ29.2 (1C, s), 68.3 (1C, s), 94.8 (1C, s), 114.3 (2C, s), 116.7 (2C, s), 124.9(1C, s), 127.8 (1C, s), 129.4 (2C, s),130.0 (2C, s), 132.0 (1C, s), 135.5(1C, s), 139.0 (1C, s), 147.2 (1C, s), 162.3-162.4 (2C, 162.3 (s), 162.3(s)).HR-ESI-MS m / z: [M+H]+ Calcd for C19H15ClO3 326.0710 ; Found325.7847.

[0063] The neuroprotective effect of the compound of the present invention is further illustrated by pharmacodynamic experiments.

[0064] 1. Materials and methods:

[0065] 1.1 Experimental Materials

[0066] 1.1.1 Experimental Animals The experimental mice were SPF-grade Sprague Dawley rats purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd. Among them, rats at 16-18 days of gestation were used for neuron culture.

[0067] 1.1.2 Main reagents and instruments DMEM culture medium, Neurobasal culture medium, fetal bovine serum,

[0068] 0.25% trypsin was purchased from Invitrogen. Cell culture plates, cell culture flasks and centrifuge tubes were purchased from Corning. 2,2'-Azobis(2-methylpropionamidine) dihydrochloride (AAPH) and CCK-8 cell activity kit were purchased from Dongren.

[0069] 1.2 Experimental methods

[0070] 1.2.1 Primary neuron culture

[0071] After anesthetizing 16-18 day gestational age rats with 10% chloral hydrate, the rats were placed in a clean bench, and the embryos were separated under sterile conditions and transferred to Neuralbasal culture medium. The cortex was taken, the meninges and blood vessels were removed, and the cells were washed twice with culture medium, chopped up, and the cells were dispersed by blowing. The cells were digested with 0.25% trypsin for 10 minutes, and all tissue blocks were filtered to remove the cells. The cells were suspended in DMEM culture medium containing 10% fetal bovine serum and inoculated in a 24-well culture plate. The culture medium in the culture plate was removed the next day and replaced with neuronal culture medium: Neuralbasal culture medium containing 2% B-27 additive, and the medium was changed every other day. It was used for experiments after 7 days.

[0072] 1.2.2 AAPH-induced cell damage and cell viability detection:

[0073] 2,2'-Azobis(2-methylpropionamidine) dihydrochloride (AAPH) was dissolved in DMEM to prepare 1mM, 5mM, 10mM, 20mM and 40mM solutions, which were added to neurons respectively. CCK-8 solution was also added at the same time. After 4 hours, the absorbance at 495nm wavelength was measured by spectrophotometer to analyze cell viability and observe the effect of AAPH on cell viability.

[0074] 1.2.3 Protective effect of new compounds on neurons in acute ischemic stroke.

[0075] Primary cultured neurons were first induced to be damaged by 40 mM AAPH for 4 h. The culture medium was replaced and new compounds of different concentrations were added. After 24 h of protection, CCK-8 solution was added. After 4 h of action, the absorbance at 495 nm was measured by a spectrophotometer to analyze cell viability and determine the protective effect on neurons.

[0076] 1.2.4 Anti-apoptotic effects of new compounds

[0077] Primary cultured neurons were first induced to be damaged by 40mM AAPH for 4h. The culture medium was replaced and different concentrations of new compounds were added. After protection for 24h, anti-apoptosis detection reagent solution was added, and then the number of apoptotic cells was observed under a fluorescence microscope. The normal cells and apoptotic cells in 10 fields of view were recorded and their percentages were calculated. Thus, the anti-apoptotic effect of the new compound was determined.

[0078] 1.2.5 Statistics and Analysis

[0079] The experimental results were statistically analyzed using SPSS15.0, and the experimental data were expressed as mean ± standard deviation. The comparison between groups was performed using analysis of variance. P < 0.05 was considered statistically significant.

[0080]

[0081]

[0082]

[0083]

[0084]

[0085]

[0086]

[0087]

[0088]

[0089]

[0090]

[0091]

[0092]

[0093]

[0094]

[0095]

[0096] 2. Results:

[0097] 2.1 AAPH can simulate stroke to prepare a neuron injury model. The expression of neuron-specific marker neurofilament (NF) in primary cultured neurons indicated that the cultured cells were neurons. Different concentrations of AAPH could damage neurons, and cell activity decreased with increasing AAPH concentrations. 40 mM AAPH could significantly induce neuron damage.

[0098] 2.2 New compounds can reduce neuronal damage

[0099] After 40 mM AAPH induced neuronal damage, cerebroside-astrocyte conditioned medium was added for 24 hours. Neuronal cell viability was then detected. The results (Table 2-9) showed that the new compound could reverse the neuronal damage caused by AAPH, and the results were statistically significant.

[0100] 2.3 New compounds can reduce cell apoptosis

[0101] Through the anti-apoptotic effect test of 1.2.4 new compounds, the percentage of apoptotic cells was statistically analyzed (Tables 10-17), indicating that the compounds of the present invention have anti-apoptotic effects on cells.

[0102] For the purpose of illustration, specific exemplary embodiments of the present invention are presented above.

[0103] The description is not intended to be exhaustive of the present invention, nor is it intended to limit the present invention to the disclosed exact form. Obviously, many changes and variations can be made according to the above description. These exemplary embodiments are selected and described to explain the specific principles of the present invention and its practical application, so that those skilled in the art can make and use each exemplary embodiment of the present invention, and various substitutions and modifications thereof. In fact, the scope of the present invention is limited by the appended claims and their equivalents.

Claims

1. Use of a compound having a structure of formula I in the preparation of a drug for treating acute ischemic stroke: in: R1, R2 are the same or different and are selected from hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy group, halogen atom; R3 is: hydrogen, C 1-6 Alkyl, phenyl, benzyl, phenethyl.

2. The use according to claim 1, characterized in that: The compound of formula I is: 。 3. The use according to claim 1, characterized in that: The preparation method of the compound of formula I is as follows: Raw materials II and III react at 0°C to room temperature in the presence of NaH and THF to generate intermediate IV. Intermediate IV and intermediate V react at 80-100°C for 8-12h in the presence of toluene, tetrakis(triphenylphosphine)palladium and potassium carbonate, and finally purify to obtain compound I. Wherein R1, R2, and R3 are as described in claim 1, and M is a methylene group.

4. A pharmaceutical composition for treating acute ischemic stroke, comprising a therapeutically effective amount of a compound having a structure of formula I and one or more pharmaceutically acceptable carriers, in: R1, R2 are the same or different and are selected from hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy group, halogen atom; R3 is: hydrogen, C 1-6 Alkyl, phenyl, benzyl, phenethyl.

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