A novel aporphine compound, its preparation method and application

Abstract

The present invention discloses a novel aporphine compound, its preparation method and applications, and its structural formula is shown as follows. The name of the novel aporphine compound is 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 ). In the present invention: ① By using the computer-aided drug design method in the forefront field of pharmacy, this compound is designed according to the binding requirements of the drug action target; ② This compound is synthesized from 3-methoxyphenethylamine and 2-bromo-5-hydroxybenzaldehyde, and its preparation method is simple; ③ This compound is an aryl hydrocarbon receptor (AhR) agonist and an inducer for the generation of regulatory T cells (Treg cells), and can be used for the prevention and treatment of immune inflammation and related diseases. ④ This compound shows good plasma protein binding rate and bioavailability, and has good prospects for drug development. #imgabs0#

Description

Technical Field

[0001] The present invention relates to a compound and a preparation method and application thereof, in particular to a novel aporpheniride compound and a preparation method and application thereof. Background Art

[0002] The aryl hydrocarbon receptor (AhR) is a ligand-dependent nuclear transcription factor that participates in the body's response to changes in the external environment in multicellular organisms. In the past, its ligands were believed to be environmental carcinogens, such as 2,3,7,8-tetrachlorodibenzodioxin and related halogenated aromatic hydrocarbons and polycyclic aromatic hydrocarbons. It has received widespread attention for its role in reducing the toxicity of environmental pollutants. It was subsequently discovered that AhR is a key regulator of pathological processes such as immune imbalance, lipid metabolism disorders, liver damage, and thymic degeneration. The AhR pathway is involved in inflammatory responses and can regulate the secretion of inflammatory cytokines and the activation of inflammatory-related pathways. AhR can also participate in the regulation of T cell differentiation and function. Regulatory T cells (Treg) are a type of suppressive CD4 + CD25 + T cells play an important regulatory role in maintaining the body's immune tolerance and immune microenvironment homeostasis. Their insufficient number or abnormal function will lead to the occurrence of immune diseases. Promoting Treg cell differentiation can effectively improve autoimmune diseases such as type 1 diabetes, rheumatoid arthritis, systemic lupus erythematosus, etc. However, the current treatment method has the following problems: 1. When the in vitro expanded Treg cells are infused or adoptively transferred back into the patient's body, it is expensive and difficult to obtain efficient and functionally stable Treg cells. 2. The use of low doses of interleukin 2 (IL-2), rapamycin, etc. in vivo to induce Treg cell formation is highly adaptable, but its safety remains to be studied.

[0003] Norisoboldine (NOR) is an aporphyrin alkaloid isolated from the dried root tubers of Linderae serrata, with pharmacological activities such as anti-inflammatory, analgesic, and immunomodulatory. NOR (10, 30 μM) can significantly promote Treg cell differentiation and Foxp3 mRNA expression. Oral administration of NOR (20, 40 mg / kg) can induce intestinal Treg cell formation by activating aryl hydrocarbon receptors and restore the dynamic balance of systemic Th17 / Treg cells, thereby exerting anti-arthritis and anti-colitis effects. However, pharmacokinetic studies have shown that after oral administration of NOR (60 mg / kg) to rats, its half-life (t 1 / 2 ) was 30.20±11.04min, and the highest blood concentration (C max ) is 0.44 μM, and the absolute bioavailability is only 2.77%. Summary of the invention

[0004] OBJECT OF THE INVENTION: The object of the present invention is to provide a new aporphine alkaloid, an AhR agonist and a Treg cell generation inducer that are highly efficient and have excellent druggability prospects, which can be used to treat or alleviate immunopathological disorders and diseases.

[0005] TECHNICAL SOLUTION: The novel aporphine compound or a pharmaceutically acceptable salt thereof according to the present invention has the following chemical structural formula:

[0006]

[0007] The preparation method of the novel aporphine compound or a pharmaceutically acceptable salt thereof includes the following steps:

[0008]

[0009] The preparation method of the novel aporphine compound or a pharmaceutically acceptable salt thereof includes the following steps:

[0010] (1) 3-Methoxyphenethylamine and benzyl chloroformate are stirred and reacted in an aqueous sodium carbonate solution. After the reaction is completed, it is extracted with ethyl acetate, and the ethyl acetate layer is evaporated under reduced pressure to obtain N-benzyloxycarbonyl-2-(3-methoxyphenyl)ethylamine;

[0011] (2) 2-Bromo-5-hydroxybenzaldehyde, potassium carbonate, and benzyl bromide are stirred and reacted in N,N-dimethylformamide. After the reaction is completed, water is added to quench the reaction, and it is extracted with ethyl acetate. The ethyl acetate layers are combined, washed with saturated sodium chloride solution, then anhydrous sodium sulfate is added for drying, filtered, and the solvent is recovered to obtain 2-bromo-5-benzyloxybenzaldehyde;

[0012] (3) The compound obtained in step (2) is stirred and reacted with methoxymethyltriphenylphosphonium chloride and sodium tert-butoxide in dichloromethane. After the reaction is completed, water is added to quench the reaction, the aqueous layer is discarded, the solvent is recovered, and it is separated by silica gel column chromatography to obtain 2-bromo-5-benzyloxy-(2-styrylmethyl ether);

[0013] (4) The compounds obtained in steps (1) and (3) are stirred and reacted with trifluoroacetic acid in acetonitrile, and filtered to obtain benzyl 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylate;

[0014] (5) The compound obtained in step (4) is refluxed and reacted with palladium acetate, 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl, and cesium carbonate in N,N-dimethylacetamide. After the reaction is completed, it is cooled to room temperature and separated by silica gel column chromatography to obtain benzyl 2-methoxy-6-carboxybenzyl-9-benzyloxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline;

[0015] (6) The compound obtained in step (5) and palladium on carbon are subjected to a stirring reaction in a solution of tetrahydrofuran and methanol under hydrogen protection, and separated by a reversed-phase ODS column to obtain 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline.

[0016] In the preparation method of the novel aporphine compound or a pharmaceutically acceptable salt thereof, in step (1), the molar ratio of 3-methoxyphenethylamine to benzyl chloroformate is 10:1 - 1:10; the concentration of the aqueous sodium carbonate solution is 2 - 12%, and the pH value is 8.0 - 10.0.

[0017] In the preparation method of the novel aporphine compound or a pharmaceutically acceptable salt thereof, in step (2), the molar ratio of 2-bromo-5-hydroxybenzaldehyde to benzyl bromide is 1:1 - 1:3; the molar ratio of 2-bromo-5-hydroxybenzaldehyde to potassium carbonate is 1:1 - 1:5.

[0018] In the preparation method of the novel aporphine compound or a pharmaceutically acceptable salt thereof, in step (3), the molar ratio of 2-bromo-5-benzyloxybenzaldehyde to methoxymethyltriphenylphosphonium chloride is 1:1 - 1:3; the molar ratio of 2-bromo-5-benzyloxybenzaldehyde to sodium tert-butoxide is 1:1 - 1:5.

[0019] In the preparation method of the novel aporphine compound or a pharmaceutically acceptable salt thereof, in step (4), the molar ratio of 2-bromo-5-benzyloxy-(2-styrylmethoxy)benzene to N-benzyloxycarbonyl-2-(3-methoxyphenyl)ethylamine is 1:1 - 1:2; the molar ratio of 2-bromo-5-benzyloxy-(2-styrylmethoxy)benzene to trifluoroacetic acid is 1:1 - 1:5.

[0020] In the preparation method of the novel aporphine compound or a pharmaceutically acceptable salt thereof, in step (5), the molar ratio of 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylic acid phenylmethyl ester to palladium acetate is 1:0.05 - 1:0.2; the molar ratio of palladium acetate to 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl is 1:0.5 - 3; the molar ratio of 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylic acid phenylmethyl ester to cesium carbonate is 1:1 - 1:5.

[0021] Use of the novel aporphine compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating immune diseases.

[0022] A pharmaceutical composition is composed of the novel aporphine compound or its pharmaceutically acceptable salt as the active ingredient and at least one pharmaceutically acceptable carrier.

[0023] Preferably, in the preparation method of the novel aporphine compound or its pharmaceutically acceptable salt, in step (1), the molar ratio of 3-methoxyphenethylamine to benzyl chloroformate is 1:1; the concentration of the sodium carbonate aqueous solution is 10%, and the pH value is 9.0.

[0024] Preferably, in the preparation method of the novel aporphine compound or its pharmaceutically acceptable salt, in step (2), the molar ratio of 2-bromo-5-hydroxybenzaldehyde, potassium carbonate and benzyl bromide is 1:2:1. The reaction condition is an ice bath.

[0025] Preferably, in the preparation method of the novel aporphine compound or its pharmaceutically acceptable salt, in step (3), the molar ratio of 2-bromo-5-benzyloxybenzaldehyde, methoxymethyltriphenylphosphonium chloride and sodium tert-butoxide is 1:1.5:1.5. The reaction condition is an ice bath.

[0026] Preferably, in the preparation method of the novel aporphine compound or its pharmaceutically acceptable salt, in step (4), the molar ratio of 2-bromo-5-benzyloxy-(2-styrylmethyl ether), N-carbobenzyloxy-2-(3-methoxyphenyl)ethylamine and trifluoroacetic acid is 1:1.1:2.

[0027] Preferably, in the preparation method of the novel aporphine compound or its pharmaceutically acceptable salt, in step (5), the molar ratio of benzyl 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylate, palladium acetate, 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl and cesium carbonate is 1:0.1:0.2:1.5. The reaction needs to be carried out under nitrogen protection in an oil bath at 130 °C.

[0028] Preferably, in the preparation method of the novel aporphine compound or its pharmaceutically acceptable salt, in step (6), the volume ratio of tetrahydrofuran to methanol is 1:2.

[0029] Use of the novel aporphine compound or its pharmaceutically acceptable salt in the preparation of an aryl hydrocarbon receptor (AhR) agonist and an inducer for the generation of regulatory T cells (Treg cells).

[0030] Use of the novel aporphine compound or its pharmaceutically acceptable salt in the preparation of drugs for colitis, rheumatoid arthritis, psoriasis, and systemic lupus erythematosus.

[0031] Beneficial effects: Compared with the prior art, the present invention has the following advantages: The compounds of the present invention have the therapeutic effect on immune diseases and can activate the AhR-regulated Treg-mediated immune response pathway, preparation methods and applications. At the same time, they have good prospects for drug development, providing a basis for new drug screening. It can significantly reduce the MPO level in the colon tissue of colitis mice, effectively improve the DAI score and intestinal damage, and show an obvious dose-dependence. At the same time, at a low dose (10 mg / kg), it has a better effect than the commonly used drug for colitis, 5-aminosalicylic acid (100 mg / kg). In addition, it also shows a better anti-colitis effect than norisoboldine (10, 20 mg / kg), and the compound of the present invention (10 mg / kg) has a better effect on inducing the formation of Treg cells than norisoboldine (20 mg / kg). In terms of pharmacokinetics, compared with the known norisoboldine, the compounds of the present invention have a more suitable oil-water partition coefficient, can be better absorbed in vivo, show a very high oral bioavailability, and are suitable for treating or alleviating immune diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 is the interaction diagram of 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 ) with AhR protein;

[0033] Figure 2 is the HPLC chromatogram of 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 );

[0034] Figure 3 is the mass spectrum of 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 );

[0035] Figure 4 is the carbon spectrum of 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 );

[0036] Figure 5 is the hydrogen spectrum of 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 );

[0037] Figure 6 is the schematic diagram of the result of 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 ) activating AhR;

[0038] Figure 7 is 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 ) Schematic diagram of the results of promoting Treg cell differentiation;

[0039] Figure 8 is 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 ) Schematic diagram of the results of improving DSS-induced colitis in mice;

[0040] Figure 9 is 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 ) Schematic diagram of the results of improving DSS-induced colitis in mice. Detailed implementation manners

[0041] The technical solutions of the present invention will be further described in detail below with reference to the accompanying drawings and examples.

[0042] The "pharmaceutically acceptable salts" of the compounds of the present invention include the conventional non-toxic salts of the compounds of the present invention formed by reacting with inorganic or organic acids. For example, it includes inorganic acids: hydrobromic acid, hydrochloric acid, sulfuric acid, phosphoric acid, nitric acid, etc., and also includes organic acids: acetic acid, oxalic acid, propionic acid, succinic acid, lactic acid, malic acid, tartaric acid, citric acid, maleic acid, fumaric acid, benzoic acid, phenylacetic acid, salicylic acid, etc.

[0043] The compounds described in the present invention have asymmetric centers, and racemates, racemate mixtures, single enantiomers, and all possible isomers and their mixtures including optical isomers are all included in the present invention.

[0044] Structural design principle of this class of new compounds

[0045] Using the target / ligand molecular docking technology in computer-aided drug design method, based on the minimum binding energy, the molecular structural formula of the new compound with expected immunomodulatory potential is designed, and at the same time, it has good druggability prospects. The structural design method is as follows:

[0046] An AhR homology model is established. Through the conformation with the highest score when the compound is docked with the AhR protein structure, it shows that the lowest energy of the compound binding to AhR is -7.06 ± 0.05 kcal / mol, and the binding pocket to AhR is His285, Phe289, Gly315, His331, Met342, Val344, Phe345, Val357, Gln377. With Met342 Phe345(3.12, ) and Val357 form hydrophobic interactions and form hydrogen bonds with His331, Val344, Val357 and Ser359, with bond lengths as shown. The ClogP value of the compound is 2.596. Figure 1

[0047] The reaction route for synthesizing the compound of the present invention is as follows:

[0048]

[0049] Example 1

[0050] Preparation of the compound 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline

[0051] (I) Synthesis of N-carbobenzyloxy-2-(3-methoxyphenyl)ethylamine

[0052] Using 10% Na 2 CO 3 aqueous solution as the reaction solvent to obtain carbobenzyloxy-protected 3-methoxyphenethylamine.

[0053]

[0054] At room temperature, dissolve 5 g of Na 2 CO 3 solid in 50 mL of pure water (pH = 9.0), add 30 mmol of 3-methoxyphenethylamine, then add 30 mmol of benzyl chloroformate, stir for 6 h, a yellow oil appears at the bottom of the reaction flask. Remove the yellow oil, add an appropriate amount of pure water and extract with ethyl acetate. Combine the ethyl acetate layers and recover the solvent to obtain the target product with a yield of 90.6%. HRMS (ESI + ): calcd 286.1438 for C 17 H 20 NO 3 + [M + H] + , found 286.1449; HRMS / MS: 91.0541, 147.0805, 225.1275 (CE: 20 eV); UV (MeOH) λ max nm: 200, 220, 275 nm. 1 H NMR (600 MHz, CDCl 3 ), δ H ​: 7.37 (7H, s), 6.84 - 6.67 (3H, m), 5.12 (2H, s), 3.80 (3H, s), 3.48 (2H, s), 2.82 (2H, s).

[0055] (2) Synthesis of 2 - bromo - 5 - benzyloxybenzaldehyde

[0056]

[0057] Under ice - bath conditions, add 45 mmol of 2 - bromo - 5 - hydroxybenzaldehyde to a reaction flask containing 50 mL of DMF. The reaction solution is reddish - brown. Add 90 mmol of K 2 CO 3 . The reaction solution changes from reddish - brown to bright yellow. Then add 45 mmol of benzyl bromide and continue stirring for 24 h. After TLC detection and complete reaction, add pure water to quench the reaction, extract with ethyl acetate, combine the ethyl acetate layers, wash with saturated sodium chloride solution, then add anhydrous sodium sulfate for drying, filter, and recover the solvent to obtain the target product with a yield of 96.2%. HRMS (ESI + ): calcd 291.0015 for C 14 H 12 BrO 2 + [M + H] + , found 291.0021; HRMS (ESI - ): calcd 288.9870 for C 14 H 10 BrO 2 - [M - H] - , found 288.9895; HRMS / MS: 80.9185, 78.9205, 181.0676, 208.7883, 153.0725 (CE: 20 eV); UV (MeOH) λ max nm: 225, 255 nm. 1 1H NMR (600 MHz, CDCl 3 ), δ H : 10.33 (1H, s), 7.59 - 7.33 (8H, m), 5.12 (2H, s).

[0058] (3) Synthesis of 2 - bromo - 5 - benzyloxy - (2 - styrylmethyl ether)

[0059]

[0060] Under ice bath conditions, 40 mmol of 2-bromo-5-benzyloxybenzaldehyde, 60 mmol of methoxymethyltriphenylphosphonium chloride, and 60 mmol of sodium tert-butoxide were added successively. 50 mL of anhydrous dichloromethane was quickly added, and the mixture was stirred for 48 h. After the reaction was completed, the reaction was quenched with pure water. The aqueous layer was discarded, the solvent was recovered, and separation by silica gel column chromatography was carried out. The eluent was petroleum ether: methyl tert-butyl ether 30:1, and the target product was obtained with a yield of 77.1%. HRMS (ESI + ): calcd 319.0328 for C 16 H 16 BrO 2 + [M+H] + , found 319.0325; HRMS / MS: 91.0539, 105.0697, 200.9727 (CE: 20 eV); UV (MeOH) λ max nm: 205, 228 nm. 1 H NMR (600 MHz, CDCl 3 ), δ H : 7.40 (6H, m), 6.98 (1H, t, J = 8.4 Hz), 6.69 (1H, td, J = 8.7 Hz), 6.17 (1H, s), 5.07 (2H, s), 3.77 (3H, d).

[0061] (IV) Benzyl 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylate

[0062]

[0063] In a reaction flask containing 50 mL of acetonitrile, 20 mmol of trifluoroacetic acid, 11 mmol of N-benzyloxycarbonyl-2-(3-methoxyphenyl)ethylamine, and 10 mmol of 2-bromo-5-benzyloxy-(2-styrylmethyl ether) were added successively. The mixture was stirred under ice bath for 5 d, and a large amount of white solid precipitated. The target product was obtained by filtration with a yield of 70.4%. HRMS (ESI - ): calcd 570.1285 for C 31 H 27 BrNO 3 - [M-H] - , found 570.1312; HRMS (ESI + ): calcd 572.1431 for C 31 H 29 BrNO 3 + [M+H] +, found 572.1441; HRMS / MS: 206.0811, 296.1281, 91.0542, 252.1384, 162.0913 (CE: 20 eV); UV(MeOH) λ max nm: 225, 282 nm. 1 H NMR(500 MHz, CDCl 3 ), δ H : 7.35 (11H, m), 7.15 (2H, t, J = 7.4 Hz), 6.76 (2H, dd, J = 13.6, 8.8 Hz), 6.68 (2H, m), 5.56 (1H, m), 4.77 (4H, m), 3.81 (3H, d, J = 6.4 Hz), 3.28 (2H, dd, J = 50.4, 40.8 Hz), 3.02 (2H, dd, J = 13.5, 10.3 Hz), 2.78 (2H, d, J = 16.2 Hz). 13 C NMR(126 MHz, CDCl 3 ), δ C : 158.43, 157.95, 155.44, 138.86, 136.76, 136.45, 135.77, 133.17, 128.77, 128.71, 28.38, 128.19, 128.15, 127.92, 127.60, 118.21, 115.99, 114.99, 113.46, 112.76, 77.41, 77.16, 76.91, 70.35, 67.21, 55.42, 54.18, 43.16, 37.63, 28.96.

[0064] (V) 2-Methoxy-6-(carboxybenzyl)-9-benzyloxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline

[0065]

[0066] To the reaction flask were successively added 5 mmol of 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylic acid benzyl ester, 0.5 mmol of palladium acetate, 1 mmol of 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl, and 7.5 mmol of cesium carbonate. Under nitrogen protection, N,N-dimethylacetamide was quickly added, and the mixture was refluxed with condensation at 130 °C and stirred for 3 h. After the reaction was completed, dichloromethane was added to mix well, pure water was added, and the mixture was extracted several times with dichloromethane. The dichloromethane layers were combined, the solvent was recovered, and the product was separated by silica gel column chromatography using petroleum ether:methyl tert-butyl ether 5:1 as the eluent to obtain the target product with a yield of 47.7%. HRMS(ESI +): calcd 492.2169 for C 32 H 30 NO 4 + [M+H] + , found 492.2167; HRMS / MS: 91.0542, 181.1014, 431.2011, 324.1224 (CE: 20 eV); UV(MeOH) λ max nm: 200, 238, 275, 282, 315 nm. 1 H NMR (500 MHz, CDCl 3 ), δ H : 7.69 (1H, d, J = 8.6 Hz), 7.43 (10H, ddd, J = 28.4, 18.9, 6.9 Hz), 7.28 (1H, s), 7.12 (1H, s), 6.96 (1H, d, J = 8.4 Hz), 6.64 (1H, s), 5.23 (2H, s), 5.12 (2H, s), 4.94 (1H, d, J = 12.6 Hz), 3.87 (3H, s), 3.18 - 2.66 (6H, m). 13 C NMR (126 MHz, CDCl 3 ), δ C : 158.87, 158.78, 137.61, 137.01, 136.95, 135.76, 128.78, 128.15, 128.04, 127.64, 127.05, 125.25, 124.10, 114.99, 114.07, 111.75, 108.30, 70.18, 67.25, 55.50, 51.55, 39.44, 30.81.

[0067] (VI) 2 - Methoxy - 9 - hydroxy - 5,6,6a,7 - tetrahydro - 4H - dibenzo[de,g]quinoline

[0068]

[0069] Add 2 mmol of 2 - methoxy - 6 - (carbobenzyloxycarbonyl)phenyl - 9 - benzyloxy - 5,6,6a,7 - tetrahydro - 4H - dibenzo[de,g]quinoline and 10% palladium - carbon into a reaction flask. Under hydrogen protection, add tetrahydrofuran and methanol, stir at room temperature for 12 h. After the reaction is completed, filter. The filtrate is concentrated to a certain volume to precipitate white powder. After separation by a reverse - phase ODS column, the eluent is 40% methanol - water (containing 0.5% acetic acid), and the yield is 37.3%. HRMS (ESI + ): calcd 268.1332 for C 17H 18 NO 2 + [M+H] + , found 268.1334; HRMS / MS: 251.1067, 236.0833, 221.0959, 203.0852 (CE: 20 eV); UV(MeOH) λ max nm: 215, 282, 303 nm. 1 H NMR(600 MHz, DMSO-d 6 ), δ H : 9.54 (1H, s), 7.59 (1H, d, J = 8.4 Hz), 7.01 (1H, d, J = 1.9 Hz), 6.70 (1H, dd, J = 8.3, 1.9 Hz), 6.66 (1H, s), 6.55 (1H, d, J = 1.5 Hz), 3.76 (3H, s), 3.71 (1H, dd, J = 14.0, 4.5 Hz), 2.83 (2H, dd, J = 17.3, 8.4 Hz), 2.76 - 2.57 (2H, m), 2.54 - 2.48 (2H, m). 13 C NMR(151 MHz, DMSO-d 6 ), δ C : 158.56, 157.56, 137.71, 135.85, 134.48, 127.24, 125.74, 125.46, 115.34, 114.51, 112.13, 106.84, 55.45, 53.25, 43.29, 36.95, 29.61.

[0070] The prepared 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (III 11 ) was detected. The HPLC chromatogram is shown in Figure 2 , the mass spectrum is shown in Figure 3 , the carbon spectrum is shown in Figure 4 , and the hydrogen spectrum is shown in Figure 5 .

[0071] Example 2

[0072] Activation effect on AhR

[0073] Obtained by magnetic bead sorting T cells were seeded in a cell plate pre-coated with anti-CD3, and TGF-β, anti-CD28, IL-2, 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (1, 3 μM) and the positive drug 3,3'-diindolylmethane (DIM, 3 μM) were added. At 37 °C, 5% CO2 Cultured for 72 h under the conditions. The expression of the AhR downstream target gene CYP1A1 was detected by RT-qPCR, and immunofluorescence was used to examine whether it promoted the nuclear translocation of AhR. The results are as Figure 6 shown.

[0074] Effect on Treg differentiation

[0075] Obtained by magnetic bead sorting T cells were seeded in a cell plate pre-coated with anti-CD3, and TGF-β, anti-CD28, IL-2, 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline (1, 3 μM) and the positive drug 3,3'-diindolylmethane (DIM, 3 μM) were added. They were cultured at 37 °C and 5% CO 2 for 72 h under the conditions. Flow cytometry was used to determine the proportion of Treg cells, and RT-qPCR was used to detect the expression of molecules related to Treg differentiation. The results are as Figure 7 shown.

[0076] Plasma protein binding rate detection

[0077] 2 mL of fresh rat plasma anticoagulated with heparin was taken, and III was added 11 to make the final concentration 10, 20, 40 μg / mL. It was shaken on a shaker at 37 °C for 12 h. 200 μL of plasma sample was added to an ultrafiltration tube and centrifuged at 2520 g at room temperature for 30 min. The ultrafiltrate and the sample in the inner ultrafiltration tube were discarded, and it was centrifuged at 1000 g in the reverse direction at room temperature for 5 min to remove the residue in the inner ultrafiltration tube. Then 200 μL of plasma sample was added to the ultrafiltration tube and centrifuged at 2520 g at room temperature for 30 min to prepare the ultrafiltrate. 100 μL of the plasma sample before ultrafiltration and the ultrafiltrate were taken, 100 μL of 0.2 mol / L sodium hydroxide solution was added, and after mixing, it was extracted twice with ethyl acetate, 500 μL each time, vortexed for 3 min, centrifuged at 12000 r / min at 4 °C for 10 min, and 800 μL of the supernatant was taken and evaporated to dryness in a vacuum concentrator at 37 °C. The residue was re-dissolved with 80 μL of the initial mobile phase and centrifuged at 12000 r / min at 4 °C for 10 min, and the supernatant was taken. After verifying that the bioanalytical method was feasible, the concentrations in the ultrafiltrate and plasma were determined, and the plasma protein binding rate (PPB) was calculated. The PPB of the III of this discovery 11 after incubation in plasma for 12 h was 85.94%, showing a high-intensity plasma protein binding ability.

[0078] Bioavailability detection

[0079] Ten healthy male Sprague-Dawley rats (body weight 180 - 200 g) were randomly divided into two groups of 5 rats each. For the gavage group, III 11 was suspended in 0.5% CMC-Na solution, and for the intravenous group, III 11 was prepared with normal saline containing 10% propylene glycol. Blood samples were collected at 5, 10, 20, 30, 60, 90, 120, 240, 480, 720, and 1440 min (n = 5) after gavage administration (30 mg / kg) or intravenous administration (3 mg / kg). The blood was placed in heparin sodium-coated centrifuge tubes and centrifuged at 8000 r / min at 4 °C for 6 min, and the supernatant plasma was taken. After verifying the feasibility of the bioanalytical method, the plasma sample concentration was determined. III 11 The oral bioavailability of the rats was 87.40%.

[0080] The prevention and treatment of immune inflammation and related diseases by the compounds of the present invention can be demonstrated in the following animal models:

[0081] In vivo determination 1: Effect in the colitis model

[0082] Different treatment methods and groups: C57BL / 6J mice were randomly divided into a normal group (Normal), a model group (Model), a NOR group (10, 20 mg / kg), III 11 a group (10, 20 mg / kg), a DIM group (40 mg / kg), and a 5-aminosalicylic acid group (100 mg / kg) according to body weight. Each group of mice had free access to food. The normal group drank pure water freely every day, and the other groups of mice were given an aqueous solution containing 3.5% DSS every day for 7 days, and then pure water for 3 days; the control group and the model group were gavaged with 0.5% CMC-Na every day, and the administration groups were administered drugs every day, with a volume of 10 mL / kg for each.

[0083] Evaluation of treatment effect and results: Disease Activity Index (DAI) score: The body weight of the mice was recorded daily, and the sum of the percentage of daily body weight loss, fecal traits, and occult blood test scores was divided by 3 as the disease activity index score. The mice were sacrificed on the last day, the colon was taken, its length was measured, its morphological changes were observed, and photographs were taken for recording. After recording, the colon was washed several times with PBS solution, and the colon tissue was stored at -80 °C for later use. The MPO activity of the colon tissue was measured. The proportion of Treg cells in the fresh spleen (LPs) and fresh mesenteric lymph nodes (MLNs) of the colon of colitis mice was measured by flow cytometry, and the expression of Treg differentiation-related molecules in the colon tissue was measured. The results were as follows Figures 8-9As shown. Statistical analysis was performed using GraphPad Prism software. The differences between groups of data were analyzed by One-way ANOVA and Tukey's test. A p < 0.05 indicates a significant difference.

[0084] In vivo assay 2: Role in a rheumatoid arthritis model

[0085] Model establishment: Weighed an appropriate amount of immunogenic chicken type II collagen (CII), dissolved it in 50 mM acetic acid solution, and left it overnight in a 4°C refrigerator to completely dissolve, obtaining a collagen solution with a concentration of 2000 μg / mL. Took an appropriate amount of complete Freund's adjuvant in a mortar, added CII (1:1), and ground it thoroughly to obtain a collagen emulsion. Intradermally injected the collagen emulsion (100 μL / animal) at 2 - 3 injection sites at the base of the mouse tail. The first injection was on day 0 (d0), and the booster immunization was performed on day 21 (d7) in the same manner.

[0086] Evaluation of treatment effect: Observed and recorded the joint lesions of the mouse limbs on d7, d11, d14, d17, d21, d24, and d27 after the primary immunization, and calculated the arthritis index (AI). The scoring method was as follows: 0 = no swelling or redness, 1 = mild swelling of the ankle joint, 2 = moderate redness and swelling from the ankle joint to the toes, 3 = severe swelling from the ankle joint to the toes, 4 = severe redness and stiffness from the ankle joint to the toes. Statistical analysis was performed using GraphPad Prism software. The differences between groups of data were analyzed by One-way ANOVA and Tukey's test. A p < 0.05 indicates a significant difference.

[0087] In vivo assay 3: Role in a psoriasis model

[0088] Model establishment: Shaved the back hair of C57BL / 6 mice, drew a 2×1.5 cm modeling area on the back skin with a marker pen, and continuously applied 62.5 mg of 5% (0.1 g:2 g) imiquimod ointment (Sichuan Mingxin Pharmaceutical Co., Ltd.) to the back skin of the mice for 6 days to induce psoriasis-like skin lesions. The control group was given the same dose of petrolatum ointment.

[0089] Evaluation of treatment effect: Starting from the 1st day after modeling, the erythema, scales, and thickness of the skin lesions were scored daily from 0 to 4, and the scores of the three items were added to obtain the total score. PASI scoring criteria: (1) Erythema: 0 = no erythema, 1 = light red, 2 = red, 3 = dark red, 4 = dark and dull red; (2) Scales: 0 = no scales on the surface, 1 = some skin lesions are covered with scales on the surface, 2 = most skin lesions are covered with scales, 3 = almost all of the skin lesion sites are covered with scales, the scales are layered, 4 = all of the skin lesion sites are covered with scales; (3) Thickness: 0 = the skin lesion is flush with the normal skin, 1 = the skin lesion is slightly higher than the surface of the normal skin, 2 = the skin lesion is moderately elevated, 3 = the skin lesion is thickened and significantly elevated, 4 = the skin lesion is highly thickened and significantly bulged. GraphPad Prism software was used for statistics, and the differences in the data of each group were analyzed by One-way ANOVA and Tukey's test. p < 0.05 indicates a significant difference.

[0090] In vivo assay 4: Role in the systemic lupus erythematosus model

[0091] Model establishment: A systemic lupus erythematosus model was established by intraperitoneal injection of 0.5 mL pristane into mice, and the success of modeling was confirmed by detecting the urinary protein level of the mice.

[0092] Evaluation of treatment effect: Detecting the size of lymph nodes, spleen index, kidney index, observing the pathological changes of the kidneys by H&E staining, etc. GraphPad Prism software was used for statistics, and the differences in the data of each group were analyzed by One-way ANOVA and Tukey's test. p < 0.05 indicates a significant difference.

Claims

1. A novel aporphine compound or a pharmaceutically acceptable salt thereof, characterized in that, the chemical structural formula of the compound is as follows:

2. A method for preparing the novel aporphine compound or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, it comprises the following steps:

3. A method for preparing the novel aporphine compound or a pharmaceutically acceptable salt thereof according to claim 2, characterized in that, it comprises the following steps: (1) Stir and react 3-methoxyphenethylamine and benzyl chloroformate in an aqueous sodium carbonate solution. After the reaction is completed, extract with ethyl acetate, and remove the solvent under reduced pressure from the ethyl acetate layer to obtain N-benzyloxycarbonyl-2-(3-methoxyphenyl)ethylamine; (2) Stir and react 2-bromo-5-hydroxybenzaldehyde, potassium carbonate, and benzyl bromide in N,N-dimethylformamide. After the reaction is completed, add water to quench the reaction, extract with ethyl acetate, combine the ethyl acetate, wash with saturated sodium chloride solution, then add anhydrous sodium sulfate for drying, filter, recover the solvent, and obtain 2-bromo-5-benzyloxybenzaldehyde; (3) Stir and react the compound obtained in step (2) with methoxymethyltriphenylphosphonium chloride and sodium tert-butoxide in dichloromethane. After the reaction is completed, quench the reaction with water, discard the aqueous layer, recover the solvent, and separate by silica gel column chromatography to obtain 2-bromo-5-benzyloxy-(2-styrylmethyl ether); (4) Stir and react the compounds obtained in steps (1) and (3) with trifluoroacetic acid in acetonitrile, and filter to obtain benzyl 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylate; (5) Reflux and react the compound obtained in step (4) with palladium acetate, 2-dicyclohexylphosphino-2′,4′,6′-triisopropylbiphenyl, and cesium carbonate in N,N-dimethylacetamide. After the reaction is completed, cool to room temperature, and separate by silica gel column chromatography to obtain benzyl 2-methoxy-6-carboxybenzyl-9-benzyloxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline; (6) Stir and react the compound obtained in step (5) with palladium on carbon in a solution of tetrahydrofuran and methanol under hydrogen protection, and separate by reverse-phase ODS column to obtain 2-methoxy-9-hydroxy-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline.

4. A method for preparing the novel aporphine compound or a pharmaceutically acceptable salt thereof according to claim 2, characterized in that, in step (1), the molar ratio of 3-methoxyphenethylamine to benzyl chloroformate is 10:1 - 1:10; the concentration of the aqueous sodium carbonate solution is 2 - 12%, and the pH value is 8.0 - 10.

0.

5. A method for preparing the novel aporphine compound or a pharmaceutically acceptable salt thereof according to claim 2, characterized in that, in step (2), the molar ratio of 2-bromo-5-hydroxybenzaldehyde to benzyl bromide is 1:1 - 1:3; the molar ratio of 2-bromo-5-hydroxybenzaldehyde to potassium carbonate is 1:1 - 1:

5.

6. A method for preparing the novel aporphine compound or a pharmaceutically acceptable salt thereof according to claim 2, It is characterized in that in step (3), the molar ratio of 2-bromo-5-benzyloxybenzaldehyde to methoxymethyltriphenylphosphonium chloride is 1:1 - 1:3; the molar ratio of 2-bromo-5-benzyloxybenzaldehyde to sodium tert-butoxide is 1:1 - 1:

5.

7. The preparation method of the novel aporphine compound or its pharmaceutically acceptable salt according to claim 2, it is characterized in that in step (4), the molar ratio of 2-bromo-5-benzyloxy-(2-styrylmethoxybenzene) to N-benzyloxycarbonyl-2-(3-methoxyphenyl)ethylamine is 1:1 - 1:2; the molar ratio of 2-bromo-5-benzyloxy-(2-styrylmethoxybenzene) to trifluoroacetic acid is 1:1 - 1:

5.

8. The preparation method of the novel aporphine compound or its pharmaceutically acceptable salt according to claim 2, it is characterized in that in step (5), the molar ratio of benzyl 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylate to palladium acetate is 1:0.05 - 1:0.2; the molar ratio of palladium acetate to 2-dicyclohexylphosphino-2′,4′,6′-triisopropylbiphenyl is 1:0.5 - 3; the molar ratio of benzyl 1-(2-bromo-5-benzyloxybenzyl)-6-methoxy-3,4-dihydro-2-isoquinolinecarboxylate to cesium carbonate is 1:1 - 1:

5.

9. The use of the novel aporphine compound or its pharmaceutically acceptable salt according to claim 1 in the preparation of drugs for the treatment of colitis, rheumatoid arthritis, psoriasis, and systemic lupus erythematosus.

10. A pharmaceutical composition comprising the novel aporphine compound or its pharmaceutically acceptable salt according to claim 1 as an active ingredient and at least one pharmaceutically acceptable carrier.

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