Compound containing aniline structure and preparation method and application thereof

Abstract

The invention discloses a compound containing an aniline structure and a preparation method and application of the compound, the structure of the compound is shown as a general formula (1), the compound shows regulation activity on neuronal calcium oscillation, abnormal discharge of primary cortical neurons can be remarkably reduced, VGSCs is selectively inhibited, and the compound has a good application prospect. And neuronal synchronization calcium oscillation induced by an epilepsy inducer 4-AP can be effectively blocked, and a protection effect on neuronal cells is exerted, so that the compound can be applied to prevention and treatment of neurological and mental diseases.

Description

Technical Field

[0001] This invention relates to the field of chemical pharmaceutical technology, and in particular to a compound containing an aniline structure, its preparation method, and its application. Background Technology

[0002] Spontaneous calcium oscillations (SCO) refer to rhythmic fluctuations in intracellular calcium ion concentration. This fundamental signaling mechanism is ubiquitous in various cell types and has been shown to be widely linked to many important human functions. Neuronal SCO is influenced by numerous factors, including neurotransmitter release, neuronal action potential formation, and ion channels (GABA, NMDA, AMPA, and VGSC, etc.), playing a crucial regulatory role in neural development, dendritic activity-dependent growth, and synapse formation. It is closely related to intractable neuropsychiatric disorders such as epilepsy, pain, depression, anxiety, and Alzheimer's disease. The potassium channel blocker 4-aminopyridine (4-AP) enhances neuronal excitability and induces aberrant synchronized calcium oscillations, which can be used to establish models simulating epileptic seizures.

[0003] The development of novel neuropsychiatric drugs is an urgent need in the current pharmaceutical field. Drug development strategies based on the regulatory mechanism of SCOs provide an innovative direction for the development of novel neuropsychiatric drugs, and have significant scientific value and application prospects. Summary of the Invention

[0004] To address the aforementioned problems in existing technologies, this invention provides a compound containing an aniline structure, its preparation method, and its applications. The compound of this invention exhibits regulatory activity on neuronal calcium oscillations, significantly reducing abnormal discharges in primary cortical neurons, selectively inhibiting VGSCs, and effectively blocking neuronal synchronized calcium oscillations induced by the epilepsy inducer 4-AP, thus exerting a protective effect on neurons and being applicable to the prevention and treatment of neuropsychiatric diseases.

[0005] The technical solution of the present invention is as follows:

[0006] The first object of the present invention is to provide a compound containing an aniline structure, the structure of which is shown in general formula (1):

[0007]

[0008] In general formula (1), n ​​represents 1 or 2;

[0009] R1 is represented, either identically or differently, by methyl- and amino-substituted phenyl groups.

[0010] One of them;

[0011] R2 represents hydrogen or amino.

[0012] In one embodiment of the present invention, the structure of the compound containing the aniline structure is shown in general formula (2):

[0013]

[0014] In general formula (2), the expressions are represented in the same or different ways as follows: One of them.

[0015] In one embodiment of the present invention, the specific structure of the compound containing the aniline structure is any one of the following structures:

[0016]

[0017] A second objective of this invention is to provide a method for preparing the above-mentioned compound containing an aniline structure, comprising the following steps:

[0018] The intermediate was obtained by reduction of nitroaniline with tin(II) chloride dihydrate; the target compound was obtained by nucleophilic substitution of the intermediate with amyl chloroformate; or...

[0019] The intermediate was obtained by coupling nitroaniline with bromobenzene; the intermediate was then reduced with tin(II) dihydrate to obtain the target compound.

[0020] In one embodiment of the present invention, a method for preparing a compound containing an aniline structure includes the following steps:

[0021] Nitroaniline and tin(II) chloride dihydrate were refluxed in anhydrous ethanol for 12 hours, cooled to room temperature, and extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to obtain the intermediate.

[0022] At 0°C, the intermediate and NaHCO3 (in anhydrous THF) were added dropwise with amyl chloroformate. After stirring at room temperature for 4 hours, the mixture was extracted with ethyl acetate and water, and the organic layer was purified by silica gel column chromatography to obtain the target compound.

[0023] The molar ratio of nitroaniline to stannous chloride dihydrate is 1:1.5 to 1:4, preferably 1:2;

[0024] The molar ratio of the intermediate to NaHCO3 is 1:1.5 to 1:4, preferably 1:2;

[0025] The molar ratio of the intermediate to amyl chloroformate is 1:1 to 1:2, preferably 1:1.75.

[0026] In one embodiment of the present invention, a method for preparing a compound containing an aniline structure includes the following steps:

[0027] Aniline, bromobenzene, Pd(OAc)2, Cs2CO3 and 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl were dissolved in toluene. Under nitrogen protection, the reaction mixture was heated to 110°C and stirred overnight. After cooling to room temperature, the mixture was filtered through diatomaceous earth and extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to obtain the intermediate.

[0028] The intermediate and tin(II) dihydrate were heated to 80°C and refluxed in ethanol. After the reaction mixture was cooled to room temperature, filtered through diatomaceous earth, and extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to obtain the target compound.

[0029] The molar ratio of aniline to bromobenzene is 0.8:1 to 1:1, preferably 0.9:1;

[0030] The molar ratio of bromobenzene to Pd(OAc)2 is 1:0.05 to 1:0.3, preferably 1:0.1;

[0031] The molar ratio of bromobenzene to Cs2CO3 is 1:2 to 1:5, preferably 1:2.5;

[0032] The molar ratio of bromobenzene to 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl is 1:0.05 to 1:0.3, preferably 1:0.1;

[0033] The molar ratio of the intermediate to stannous chloride dihydrate is 1:2 to 1:10, preferably 1:5.

[0034] A third objective of this invention is to provide an application of the above-mentioned aniline-containing compound for the preparation of drugs for the prevention and treatment of neuropsychiatric diseases.

[0035] In one embodiment of the present invention, the drug is a neuronal function modulator.

[0036] In one embodiment of the present invention, the neuronal function modulator can regulate or protect neuronal function through at least one of the following mechanisms:

[0037] a) It exhibits regulatory activity on spontaneous calcium oscillations in regulatory neurons and can promote the normalization of neuronal function;

[0038] b) It can relieve the abnormal neuronal stimulation caused by the epilepsy inducer 4-aminopyridine, thereby exerting a protective effect on neuronal cells;

[0039] c) It can inhibit voltage-gated sodium channels (VGSCs) associated with epilepsy and pain, demonstrating membrane depolarization of cortical neurons.

[0040] In one embodiment of the present invention, the neuropsychiatric disease is epilepsy, neuropathic pain, depression, Alzheimer's disease, Huntington's disease, Tourette syndrome, demyelinating diseases, schizophrenia, addiction, attention deficit, hyperactivity disorder, or Parkinson's disease.

[0041] In one embodiment of the present invention, the compound containing the aniline structure exists in the form of an organic acid salt or an inorganic acid salt;

[0042] The organic acids are acetic acid, malic acid, maleic acid, citric acid, fumaric acid, tartaric acid, succinic acid, lactic acid, p-toluenesulfonic acid, salicylic acid, or oxalic acid.

[0043] The inorganic acids are hydrochloric acid, sulfuric acid, phosphoric acid, diphosphoric acid, hydrobromic acid, or nitric acid.

[0044] In one embodiment of the present invention, the drug for preventing and treating neuropsychiatric diseases further includes a drug-acceptable carrier, excipient, or excipient.

[0045] The beneficial technical effects of this invention are as follows:

[0046] The compounds of this invention achieve precise regulation of neuronal excitability through specific interactions with neuronal receptors or ion channels, and have the potential to treat neurodegenerative diseases, epilepsy, neuropathic pain, etc. Attached Figure Description

[0047] Figure 1 The hydrogen NMR spectrum of compound 1;

[0048] Figure 2 The carbon NMR spectrum of compound 1;

[0049] Figure 3 The hydrogen NMR spectrum of compound 2;

[0050] Figure 4 The carbon NMR spectrum of compound 2;

[0051] Figure 5 The hydrogen NMR spectrum of compound 3;

[0052] Figure 6 The carbon NMR spectrum of compound 3;

[0053] Figure 7 The hydrogen NMR spectrum of compound 4;

[0054] Figure 8 The carbon NMR spectrum of compound 4;

[0055] Figure 9 The hydrogen NMR spectrum of compound 5;

[0056] Figure 10 This is the carbon NMR spectrum of compound 5. Detailed Implementation

[0057] The present invention will now be described in detail with reference to the accompanying drawings and embodiments.

[0058] Example 1

[0059] Synthetic route

[0060] Synthesis of compound 1b

[0061] A mixture of 3-methyl-5-nitroaniline (152 mg, 1.0 mmol) and tin(II) chloride dihydrate (450 mg, 2.0 mmol) in anhydrous ethanol (5 mL) was refluxed for 12 hours. After cooling to room temperature, it was extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to give compound 1b.

[0062] Synthesis of Compound 1

[0063] Compound 1b (90 mg, 0.75 mmol) and NaHCO3 (126 mg, 1.5 mmol) were mixed in anhydrous THF (5 mL) at 0 °C, and amyl chloroformate (200 mg, 1.3 mmol) was added dropwise. After stirring at room temperature for 4 hours, the mixture was extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to give compound 1. The product was a white solid (55 mg, 16%).

[0064] 1 H NMR(600MHz,DMSO-d6)δ9.47(s,2H),7.48(s,1H),6.90(s,2H),4.04(t,J=6.6Hz,4H) ,2.18(s,3H),1.60(t,J=7.0Hz,4H),1.32(dt,J=7.5,3.9Hz,8H),0.91–0.86(m,6H).

[0065] 13 C NMR (151MHz, DMSO-d6) δ154.0,139.9,138.4,113.7,64.4,40.5,28.7,28.0,22.3,21.9,14.4.

[0066] HRMS(ESI): m / z calcd for C 19 H 31 N₂O₄[M+H] + 351.2282, found 351.2280.

[0067] Example 2

[0068]

[0069] Synthesis of Compound 2

[0070] Compound 2 can be obtained by replacing 3-methyl-5-nitroaniline with 4-methyl-3-nitroaniline, following the same synthetic method as compound 1. The product is a white solid (107 mg, 56%).

[0071] 1 H NMR (600MHz, DMSO-d6) δ9.01(s,2H),7.11(s,3H),4.63(t,J=5.4Hz,2H),4.16(t,J=4.4H z, 4H), 3.63 (t, J = 4.7Hz, 4H), 3.51 (q, J = 5.3Hz, 4H), 3.46 (d, J = 5.3Hz, 4H), 2.04 (s, 3H).

[0072] 13 C NMR (151MHz, DMSO-d6) δ155.0,137.3,128.2,125.7,122.9,72.7,69.2,64.1,60.6,14.0,13.2.

[0073] HRMS(ESI): m / z calcd for C 17 H 27 N₂O₈[M+H] + 387.1767, found 387.1780.

[0074] Example 3

[0075] Synthetic route

[0076]

[0077] Synthesis of compound 3b

[0078] 4-Methyl-3-nitroaniline (1.50 g, 10.0 mmol) and NaHCO3 (1.26 g, 15.0 mmol) were dissolved in anhydrous THF (30 mL), cooled to 0 °C in an ice bath, and amyl chloroformate (1.80 g, 12.0 mmol) was slowly added dropwise. The mixture was then heated to room temperature and stirred for 4 hours. The mixture was extracted with ethyl acetate and water, and the organic layer was purified by silica gel column chromatography to obtain compound 3b.

[0079] Synthesis of compound 3c

[0080] Compound 3b (1.33 g, 5.0 mmol) and tin(II) dihydrate (2.25 g, 10.0 mmol) were refluxed in anhydrous ethanol (20 mL) for 12 hours. After cooling, the pH was adjusted to 9 with saturated NaHCO3 solution. The mixture was filtered through diatomaceous earth and extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to obtain compound 3c.

[0081] Synthesis of compound 3d

[0082] Compound 3c (236 mg, 1.0 mmol) and triethylamine (202 mg, 2.0 mmol) were dissolved in anhydrous dichloromethane (3 mL), cooled to 0 °C in an ice bath, and a dichloromethane solution of triphosgene (100 mg, 0.33 mmol) (1 mL) was slowly added dropwise. The mixture was stirred at 0 °C for 3 hours, and then a dichloromethane solution of 4-((tert-butyldimethylsilyl)oxo)but-1-ol (408 mg, 2.0 mmol) (1 mL) was added. The mixture was heated to room temperature and stirred for 12 hours. The mixture was concentrated and purified by silica gel column chromatography to obtain compound 3d.

[0083] Synthesis of Compound 3

[0084] Compound 3d (310 mg, 0.5 mmol) was dissolved in anhydrous THF (5 mL), and 2.5 mL of TBAF (1.0 M in THF) was slowly added dropwise at room temperature. After stirring for 3 hours, the mixture was extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to give compound 3. The product was a white solid (55 mg, 16%).

[0085] 1 H NMR (600MHz, DMSO-d6) δ9.50 (s, 1H), 8.76 (s, 1H), 7.49 (s, 1H), 7.14 (d, J = 8. 2Hz,1H),7.05(dd,J=8.6,3.4Hz,1H),4.44(s,1H),4.05(dt,J=8.9,4.7Hz,4 H),3.42(t,J=5.0Hz,2H),2.11(d,J=3.3Hz,3H),1.67–1.56(m,4H),1.49(q, J=7.9Hz,2H),1.33(td,J=7.2,3.8Hz,4H),0.88(dq,J=7.6,4.8,3.9Hz,3H).

[0086] 13C NMR(151MHz,DMSO-d6)δ154.85,154.05,137.69,136.94,130.64,126.02,11 5.35,64.61,64.48,60.78,29.33,28.70,28.04,25.89,22.29,17.60,14.37.

[0087] HRMS(ESI): m / z calcd for C 18 H 29 N₂O₅[M+H] + 353.2076, found 353.2072.

[0088] Example 4

[0089] Synthetic route

[0090]

[0091] Synthesis of Compound 4

[0092] Following the synthetic method of compound 3, the starting material was also 4-methyl-3-nitroaniline. First, 4-((tert-butyldimethylsilyl)oxo)but-1-ol was reacted with triphosgene to generate 4b. Then, 4b was reduced to 4c by stannous(II) chloride dihydrate. Finally, 4c was reacted with amyl chloroformate to generate compound 4. The product was a white solid (210 mg, 44%).

[0093] 1 H NMR(600MHz,DMSO-d6)δ9.51(s,1H),8.75(s,1H),7.49(s,1H),7.14(d,J=8.2Hz,1 H),7.04(dd,J=8.3,2.5Hz,1H),4.45(t,J=5.4Hz,1H),4.05(dtd,J=13.7,6.5,3.4 Hz,4H),3.44–3.40(m,2H),2.11(s,3H),1.62(ddd,J=18.8,8.9,4.2Hz,4H),1.50( q, J=7.3, 6.9Hz, 2H), 1.33 (dd, J=8.6, 4.4Hz, 4H), 0.88 (td, J=6.5, 5.7, 3.1Hz, 3H).

[0094] 13C NMR (151MHz, DMSO-d6) δ154.9,154.0,137.7,136.9,130.6,126.0,115.4,64.6,64.5,60.8,29.3,28.8,28.0,25.8,22.3,17.6,14.4.

[0095] HRMS(ESI): m / z calcd for C 18 H 29 N₂O₅[M+H] + 353.2076, found 353.2071.

[0096] Example 5

[0097] Synthetic route

[0098]

[0099] 3-Methyl-4-nitroaniline (137 mg, 0.9 mmol), 3-nitro-4-methylbromobenzene (214 mg, 1.0 mmol), Pd(OAc)₂ (25 mg, 0.1 mmol), Cs₂CO₃ (812 mg, 2.5 mmol), and 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl (50 mg, 0.1 mmol) were dissolved in toluene (10 mL) under nitrogen protection. The reaction mixture was heated to 110 °C and stirred overnight. After cooling to room temperature, the mixture was filtered through diatomaceous earth and extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to give compound 5c.

[0100] A mixture of compound 5c (143 mg, 0.5 mmol) and tin(II) dihydrate (812 mg, 2.5 mmol) in ethanol (10 mL) was heated to 80 °C and refluxed. Afterward, the reaction mixture was cooled to room temperature, filtered through diatomaceous earth, and extracted with ethyl acetate and water. The organic layer was purified by silica gel column chromatography to give compound 5. The product was an oil (25 mg, overall yield 9.1%).

[0101] 1 H NMR(600MHz,DMSO-d6)δ6.95(s,1H),6.67(s,1H),6.64(dd,J=8.1,3.8Hz,2H),6.51(d,J=8.3Hz,1H),6 .15(t,J=1.9Hz,1H),5.99(dt,J=8.0,1.9Hz,1H),4.56(s,2H),4.41(s,2H),2.01(s,3H),1.92(s,3H).

[0102] 13 C NMR (151MHz, DMSO-d6) δ147.3,143.4,130.5,112.8,106.3,103.5,17.2.

[0103] HRMS:m / z calcd.for C 14 H 19 N3[M+H] + :228.1500,found 228.1505.

[0104] Test example:

[0105] 1. In vitro detection of the inhibitory activity of the compounds of the present invention on spontaneous calcium oscillations and 4-AP-induced calcium oscillations in mouse neurons.

[0106] I. Experimental Methods

[0107] Primary neuronal culture of the cerebral cortex: C57 suckling mice aged 0-1 days were decapitated and their brains were harvested. The cerebral cortex was separated under a dissecting microscope, the meninges were carefully removed, and the cortex was prepared into a single-cell suspension. The cell suspension was centrifuged and seeded into 96-well plates and cultured in a cell culture incubator at 37°C, 5% CO2, and 95% humidity. The medium was changed every 4 and 7 days, and the plate was analyzed on day 8.

[0108] Intracellular Ca 2+ Concentration determination: Discard the culture medium from the 96-well plate, add 60 μL of dye loading buffer to each well, and incubate for 1 h at 37°C, 5% CO2, and 95% humidity. Then wash 5 times with Lockes' buffer and place the cells into a FLIPR reader for reading. Excitation wavelengths are set to 470-495 nm, and emission wavelengths to 515-575 nm. One reading is taken every 1 second. Record baseline spontaneous Ca2+. 2+ After shaking for 5 minutes, add 25 μL of the compound working solution at a concentration 8 times the final concentration, and monitor [Ca]. 2+ ] i 15 minutes. To test the antiepileptic activity of the compound, 25 μL of the epilepsy eliminator 4-AP (10 μM) was added, and monitoring continued [Ca]. 2 + ] i The fluorescence intensity was measured over a period of 10 minutes. The data presented are F / F0 values, where F is the fluorescence intensity at any time point and F0 is the baseline fluorescence intensity. Two replicates were used for each compound. Intracellular Ca2+ was observed. 2+Horizontal variation. The frequency and amplitude of SCOs within 5 minutes after compound addition were quantified using Origin software (V7.0) and normalized to the frequency and amplitude recorded in the first 5 minutes for each well. Then, all data were normalized again to the blank control, and the EC50 values ​​of the compound's effect on the amplitude and frequency of spontaneous calcium oscillations and 4-AP-induced calcium oscillations were calculated. 50 value.

[0109] II. Experimental Results

[0110] In vitro activity tests on the compounds of this invention all showed varying degrees of inhibitory activity, and their EC50 values ​​were [not specified]. 50 The values ​​are shown in Table 1.

[0111] Table 1

[0112]

[0113] * indicates a dual EC response. 50 / IC 50 Bell-shaped fit of values; N represents EC 50 Value greater than 40 μM

[0114] As shown in Table 1, compounds 1-5 exhibit inhibitory activity against both spontaneous and 4-AP-induced calcium oscillations in neurons. Specifically, compound 1 concentration-dependently reduces the amplitude and frequency of calcium oscillations, with its amplitude IC50 value being [value missing]. 50 The value is 33.56μM, and the frequency IC is... 50 The value was 25.79 μM. Compound 2 concentration-dependently reduced the amplitude and frequency of calcium oscillations, with its amplitude IC50 being 25.79 μM. 50 The value is 31.42μM, and the frequency is IC. 50 The value was 36.54 μM. Compound 3 concentration-dependently reduced the amplitude and frequency of calcium oscillations, with its amplitude IC50 being 36.54 μM. 50 The value is 3.14μM, and the frequency IC is... 50 The value was 1.96 μM. Compound 4 concentration-dependently reduced the amplitude and frequency of calcium oscillations, with its amplitude IC50 being 1.96 μM. 50 Value is 9.00μM, frequency IC 50 The value was 11.55 μM. Compound 5 concentration-dependently reduced the amplitude and frequency of calcium oscillations, with its amplitude IC50 being 11.55 μM. 50 The value is 35.78μM, and the frequency IC is... 50 The value is 39.26 μM.

[0115] Compounds 1, 3, and 4 showed inhibitory activity against both the frequency and amplitude of 4-AP-induced neuronal calcium oscillations. Compound 1 showed inhibitory activity only against the amplitude of 4-AP-induced neuronal calcium oscillations, with an IC50 value of [missing value].50 The value was 37.35 μM. Compound 3 concentration-dependently reduced the amplitude of calcium oscillations, with an amplitude IC50 of 37.35 μM. 50 The value is 6.71 μM, and its effect on frequency exhibits double EC. 50 / IC 50 Bell-shaped fitting of values, EC 50 The value is 10.00 μM, IC 50 The value was 28.51 μM. Compound 4 concentration-dependently reduced the amplitude of calcium oscillations, with an amplitude IC50 of 28.51 μM. 50 The value is 25.01μM, and its frequency IC 50 The value was 31.33 μM. The other compounds had no significant effect on 4-AP-induced neuronal calcium oscillations.

[0116] Activity tests showed that the compound could significantly inhibit spontaneous and 4-AP-induced calcium oscillations in neurons, providing important reference value for the preparation of inhibitors of abnormal neuronal discharges.

Claims

1. A compound containing an aniline structure, characterized in that, The structure of the compound is shown in general formula (1): In general formula (1), n ​​represents 1 or 2; R1 is represented, either identically or differently, by methyl- and amino-substituted phenyl groups. One of them; R2 represents hydrogen or amino.

2. The compound containing an aniline structure according to claim 1, characterized in that, The structure of the compound is shown in general formula (2): In general formula (2), the expressions are represented in the same or different ways as follows: One of them.

3. The compound containing an aniline structure according to claim 1, characterized in that, The specific structure of the compound is any one of the following structures:

4. A method for preparing the compound containing the aniline structure as described in claim 1, characterized in that, Includes the following steps: The intermediate was obtained by reduction of nitroaniline with tin(II) chloride dihydrate; the target compound was obtained by nucleophilic substitution of the intermediate with amyl chloroformate; or... The intermediate was obtained by coupling nitroaniline with bromobenzene; the intermediate was then reduced with tin(II) dihydrate to obtain the target compound.

5. The application of the compound containing the aniline structure as described in claim 1, characterized in that, Used to prepare drugs for the prevention and treatment of neuropsychiatric diseases.

6. The application according to claim 5, characterized in that, The drug is a neuronal function modulator.

7. The application according to claim 5, characterized in that, Neuropsychiatric disorders include epilepsy, neuropathic pain, depression, Alzheimer's disease, Huntington's disease, Tourette syndrome, demyelinating diseases, schizophrenia, addiction, attention deficit hyperactivity disorder, or Parkinson's disease.

8. The application according to claim 5, characterized in that, Compounds containing aniline structures exist in the form of organic acid salts or inorganic acid salts; The organic acids are acetic acid, malic acid, maleic acid, citric acid, fumaric acid, tartaric acid, succinic acid, lactic acid, p-toluenesulfonic acid, salicylic acid, or oxalic acid. The inorganic acids are hydrochloric acid, sulfuric acid, phosphoric acid, diphosphoric acid, hydrobromic acid, or nitric acid.

9. The application according to claim 5, characterized in that, Drugs for the prevention and treatment of neuropsychiatric disorders also include drug-acceptable carriers, excipients, or excipients.

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