1H-triazole biphenyl derivative as well as preparation method and medical application thereof

Abstract

The invention discloses a 1H-triazole biphenyl derivative as well as a preparation method and medical application thereof, and particularly relates to the 1H-triazole biphenyl derivative as shown in a general formula (I) and enantiomers, diastereoisomers, tautomers, N-oxides, solvates, physiologically hydrolyzable esters, preparations and pharmaceutically acceptable salts of the 1H-triazole biphenyl derivative. The invention discloses a 1H-triazole biphenyl derivative having an inhibition effect on S1PR1 and a pharmaceutically acceptable salt thereof, pharmacological experiments prove that the compound has a significant inhibition effect on S1PR1, has an effect of relieving pain, and is expected to become a novel analgesic molecule for replacing opioid drugs. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to compounds, their preparation methods and uses, and particularly relates to 1H-triazole biphenyl derivatives, their preparation methods and pharmaceutical uses. Background Art

[0002] S1P is a metabolite of ceramide and is a sphingolipid amphiphile containing a serine polar head group. It is involved in physiological processes such as cell chemotaxis, migration, growth and proliferation, and is also involved in neurogenesis, angiogenesis, cardiac chronotropy and inotropy, endothelial cell permeability, vascular integrity and tone.

[0003] S1P regulates a series of cell processes by binding to five known G protein-coupled sphingosine-1-phosphate receptors. These receptors have different cell-specific expression patterns, are related to different signaling pathways, and are regulated by different mechanisms. S1PR1 is mainly coupled to the Gi protein to inhibit the production of cAMP, S1PR2 and S1PR3 are coupled to Gi, Gq and G12 / 13, and S1PR4 and S1PR5 signal through Gi and G12 / 13. S1PRs are differentially expressed in the immune system. S1PR1 and S1PR4 are expressed by T cells, while B cells express S1PR1, S1PR2, S1PR3 and S1PR4. S1PR1 and S1PR2 are additionally found in macrophages and mast cells, while S1PR1 and S1PR5 are expressed in dendritic cells (DC) and natural killer cells (NK). Additionally, immune cells do not necessarily express all S1PRs simultaneously, as there are different expressions at different stages of cell activation and maturation.

[0004] S1PR1 is the first discovered S1PR subtype. S1PR1 plays a crucial role in controlling lymphocyte egress from the thymus, secondary lymphoid organs, and bone marrow. S1PR1 is also expressed on endothelial cells, where it is involved in dendritic cell recruitment and vascular permeability. S1PR1 plays a key role in the development of the vascular system during embryonic development. S1PR1 binds to Gi / o proteins, activating the ERK and Rac signaling pathways, promoting cell survival, proliferation, and enhancing barrier function. In adult tissues, S1PR1 is highly expressed in the lung, brain, endothelial cells, and immune organs. S1PR1 is also involved in astrocyte proliferation, neuroprotection, heart rate, endothelial integrity, and ischemia-reperfusion injury. When S1PR1 is absent during vascular development, adherens junctions are unstable and the coverage of smooth muscle cells in blood vessels is incomplete. Stimulating S1PR1 can regulate heart rate by interacting with atrial potassium channels, leading to hyperpolarization of cardiomyocytes and reducing the firing of action potentials. In addition, inhibiting S1PR1 increases the penetration of small molecules in the central nervous system. S1PR1 plays a key role in angiogenesis, neurogenesis, immune cell trafficking, endothelial barrier function, and vascular tone. These diverse biological effects are potential therapeutic targets for various diseases and may also produce off-target adverse reactions.

[0005] Currently, there are many studies on S1PR1 selective modulators because they can achieve immunosuppressive effects by regulating lymphocyte migration and can treat immune-related diseases such as MS, IBD, and GVHD. Currently, there is preclinical evidence supporting the potential therapeutic effects of S1PR1 modulators in diseases such as Parkinson's disease, myasthenia gravis, Alzheimer's disease, Huntington's disease, spinal cord injury, cancer, type 1 diabetes, autoimmune myocarditis, dilated cardiomyopathy, and hypertrophic cardiomyopathy.

[0006] In recent years, a number of studies have shown that S1P is an important regulator of the pain pathway, and the ceramide-S1P-S1PR1 pathway plays a key role in peripheral and central pain. S1PR1 antagonists are expected to become a new type of analgesic molecule to replace opioids. However, there are currently few studies on S1PR1 antagonists, only a small number of structural types of antagonists have been reported, and the vast majority of the structures still use a benzene ring as their core linking ring, with a relatively single structural type. And the structure-activity relationship of S1PR1 antagonists has not been systematically elaborated, which limits the development of this type of antagonist. Therefore, finding new structural types of S1PR1 receptor antagonists to improve problems such as opioid addiction has become a new strategy for discovering S1PR1 receptor antagonists with strong activity and good selectivity. Summary of the Invention

[0007] Object of the Invention: The object of the present invention is to provide a class of novel structures and having S1PR 11H-triazole biphenyl derivatives with receptor antagonism and their pharmaceutically acceptable salts. Another object of the present invention is to provide a method for preparing the above-mentioned 1H-triazole biphenyl derivatives. Still another object of the present invention is to provide the application of the above-mentioned 1H-triazole biphenyl derivatives in analgesia.

[0008] Technical solution: The 1H-triazole biphenyl derivative represented by the general formula (I) or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt according to the present invention:

[0009]

[0010] R 1 is selected from H, C 1-6 alkyl, C 3-6 cycloalkyl,

[0011]

[0012] wherein, R 4 is selected from H, OH, C 1-6 alkyl; R 5 is selected from H, C 1-6 alkyl; n = 1, 2;

[0013] R 2 is selected from H, NO 2 、CN、OH、NH 2 、F、Cl、Br、I、C 1-6 alkyl, halo-C 1-6 alkyl, C 1-6 alkoxy, halo-C 1-6 alkoxy;

[0014] R 3 is selected from halogen, halo-C 1-6 alkyl, C 1-6 alkyl.

[0015] The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt,

[0016] R 1 is selected from

[0017]

[0018] The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt,

[0019] R2 Selected from H, NO 2 , CN, OH, NH 2 , F, Cl, Br, I, C 1-4 alkyl group.

[0020] The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt,

[0021] R 3 Selected from Cl, F.

[0022] The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt, and the 1H-triazole biphenyl derivative is selected from the following compounds:

[0023]

[0024]

[0025] The preparation method of the 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt, comprises the following steps:

[0026] (1) Reacting the general formula compound A with an alcohol to obtain the general formula B compound;

[0027] (2) Performing a Click reaction of the general formula compound B with azide to obtain the general formula compound C;

[0028] (3) Connecting the general formula compound C with an amine derivative and performing de-esterification hydrolysis under alkaline conditions to obtain the general formula compound D;

[0029]

[0030] R1′ is an ester formed by R1 and CH 3 OH or CH 3 -(CH 2 ) 1～4 -OH.

[0031] Compound D is the general formula (I) compound, and R 1 , R 2 , R 3 are as described above.

[0032] A pharmaceutical composition, which contains the 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt.

[0033] The use of the 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt, or the pharmaceutical composition as described above in the preparation of an S1PR 1 antagonist drug.

[0034] The use of the 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolysable ester, pharmaceutically acceptable salt or the pharmaceutical composition as described above in the preparation of an analgesic drug.

[0035] For the use as described above, the drug is formulated into different dosage forms by adding pharmaceutically acceptable excipients.

[0036] The pharmaceutically acceptable excipients refer to various conventional excipients required for preparing different dosage forms, such as diluents, binders, disintegrants, glidants, lubricants, flavoring agents, inclusion materials, adsorbent materials, etc., and are prepared into any common oral dosage form by conventional pharmaceutical methods, such as granules, powders, tablets, capsules, pills, oral liquids, decoctions, dripping pills, etc.

[0037] Beneficial effects: Compared with the prior art, the present invention has the following advantages: The present invention discloses a class of 1H-triazole biphenyl derivatives and their pharmaceutically acceptable salts that have an inhibitory effect on the S1PR 1 receptor. Through pharmacological experiments, it is confirmed that these compounds have a significant inhibitory effect on the S1PR 1 receptor and can especially be used as analgesic drugs. Description of the Drawings

[0038] Figure 1 It shows the effect of compound 6 on the thermal pain threshold of mice; wherein, each data represents 8 mice and is expressed as mean±s.e.m. (n = 8); compared with the blank control group, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Detailed Embodiments

[0039] The content of the present invention will be specifically described below through examples. In the present invention, the following examples are for better explaining the present invention and are not used to limit the scope of the present invention.

[0040] Example 1

[0041] Synthesis of target compounds in Examples 1 to 4

[0042] Synthesis of N-(but-3-yn-2-yl)-4-chloro-3-methylaniline (30)

[0043]

[0044] Dissolve but-3-yn-2-ol (0.46 g, 6.62 mmol), DDQ (1.80 g, 7.94 mmol), and triphenylphosphine (2.08 g, 7.94 mmol) in 10 mL of DCM. Add compound 29 (937.38 mg, 6.62 mmol) under an ice bath. After 30 min in the ice bath, stir at room temperature for 10 h. After the reaction is completed as detected by TLC, add saturated saline solution to the reaction solution, extract the aqueous phase with DCM, combine the organic phases, evaporate the organic phases to dryness, and purify by silica gel column chromatography (PE:EA = 50:1) to obtain 0.45 g of a pale yellow oily liquid with a yield of 35%. 1 1H NMR (300 MHz, DMSO-d 6 6) δ (ppm) 7.09 (dd, J = 8.7, 2.9 Hz, 1H), 6.60 (d, J = 3.1 Hz, 1H), 6.55 - 6.44 (m, 1H), 5.97 - 5.88 (m, 1H), 4.16 (t, J = 7.4 Hz, 1H), 3.10 - 3.02 (m, 1H), 2.21 (d, J = 2.9 Hz, 3H), 1.39 (dd, J = 6.9, 3.0 Hz, 3H).

[0045] Synthesis of 4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzoic acid (31)

[0046]

[0047] Dissolve compound 30 (0.45 g, 2.31 mmol), 4-azidobenzoic acid (0.38 g, 2.31 mmol), and CuSO 4 ·5H 2O (5.79 mg, 23.17 μmol) and sodium ascorbate (22.95 mg, 115.85 μmol) were dissolved in a mixed solvent of tert-butanol:water = 1:2, and stirred at room temperature for 10 h. After the reaction was completed as detected by TLC, saturated saline solution was added to the reaction solution, and the aqueous phase was extracted with EA. The organic phases were combined, dried by evaporation, and 0.35 g of the crude product of the compound was obtained, with a yield of 43%. General preparation method 1:

[0048]

[0049] (1) R 6 -NH 2 (0.99 mmol), EDCI-HCl (0.38 g, 1.99 mmol), HOBT (0.27 g, 1.98 mmol), and TEA (0.21 g, 1.98 mmol) were dissolved in DMF. Compound 31 (0.35 g, 0.99 mmol) was added under an ice bath. After stirring for 30 min under the ice bath, the mixture was stirred at room temperature for 10 h. After the reaction was completed as detected by TLC, saturated saline solution was added to the reaction solution, and the aqueous phase was extracted with EA. The organic phases were combined, dried by evaporation, and purified by silica gel column chromatography (PE:EA = 2:1) to obtain a pale yellow oily liquid with a yield of 78% - 86%.

[0050] (2) Then, the product was dissolved in 1 mL of methanol, saturated aqueous KOH solution was added, and after addition, the mixture was stirred at room temperature for reaction. Monitored by TLC, after the reaction was complete, 2N HCl was added to adjust the system to pH = 7, and a pale yellow solid precipitated. Stirring or sonication was continued to make the precipitated solid tend to aggregate. It was filtered by suction, and the filter cake was dried to obtain a pale yellow solid. The total yield of the two steps was 60% - 80%.

[0051] (4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzoyl)-L-alanine ((4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzoyl)-L-alanine, Example 1) synthesis

[0052]

[0053] 0.32 g of a pale yellow solid was obtained according to General Preparation Method 1, with a yield of 11%. Mp. 87 °C (decomposed). 1 HNMR (300 MHz, DMSO-d 6) δ (ppm) 8.94 (d, J = 6.6 Hz, 2H), 8.12 (d, J = 8.4 Hz, 2H), 8.00 (d, J = 8.4 Hz, 2H), 7.57 (s, 1H), 7.23 (d, J = 8.6 Hz, 1H), 7.03 (d, J = 2.7 Hz, 1H), 6.86 (dd, J = 8.7, 2.6 Hz, 1H), 4.89 (q, J = 6.7 Hz, 1H), 4.44 (p, J = 7.3 Hz, 1H), 2.22 (s, 3H), 1.67 (d, J = 6.7 Hz, 3H), 1.41 (d, J = 7.3 Hz, 3H). 13 13C NMR (75 MHz, DMSO-d 6 ) δ (ppm) 174.92, 165.81, 149.59, 141.69, 139.16, 136.80, 134.41, 132.91, 132.35, 132.22, 130.11, 129.66, 129.50, 122.31, 121.01, 120.19, 117.81, 49.35, 49.13, 20.63, 20.58, 17.64. HRMS (ESI-TOF) m / z calc’d for C 21 19 22 1 5 3 3 [M + H] + 428.1445, found 428.1489。

[0054] Synthesis of ((4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzoyl)glycine (Example 2)

[0055]

[0056] A pale yellow solid (0.34 g) was obtained according to General Preparation Method 1, with a yield of 12%. Mp. 78 °C (decomposed). 1 1H NMR (300 MHz, DMSO-d 6) δ (ppm) 9.07 (t, J = 5.9 Hz, 1H), 8.71 (s, 1H), 8.10 (d, J = 8.4 Hz, 2H), 8.00 (d, J = 8.3 Hz, 2H), 7.00 (d, J = 8.5 Hz, 1H), 6.63 (d, J = 2.7 Hz, 1H), 6.50 (dd, J = 8.7, 2.7 Hz, 1H), 5.35 (s, 1H), 4.75 (q, J = 6.6 Hz, 1H), 4.00 (d, J = 5.4 Hz, 2H), 2.14 (s, 3H), 1.56 (d, J = 6.6 Hz, 3H). 13 13C NMR (75 MHz, DMSO-d 6 ) δ (ppm) 174.41, 166.57, 152.25, 146.27, 139.05, 135.91, 134.12, 132.10, 131.97, 129.73, 129.52, 129.39, 129.24, 120.76, 119.77, 116.52, 113.28, 45.98, 33.73, 21.89, 20.45, 17.06. HRMS (ESI-TOF) m / z calc’d for C 20 H 20 ClN 5 O 3 [M + H] + 414.1288, found 414.1339。

[0057] Synthesis of 1-(4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzamido)cyclopropane-1-carboxylic acid (1-(4-(4-(1-((4-chloro-3-methylphenyl)amin0)ethyl)-1H-1,2,3-triazol-1-yl)benzamido)cyclopropane-1-carboxylic acid, Example 3)

[0058]

[0059] A pale yellow solid (0.26 g) was obtained according to General Preparation Method 1, with a yield of 9%. Mp. 92 °C (decomposed). 1 1H NMR (300 MHz, DMSO-d 6) δ (ppm) 9.25 (s, 1H), 8.80 (s, 1H), 8.09 (d, J = 8.4 Hz, 2H), 8.00 (d, J = 8.6 Hz, 2H), 7.60 (d, J = 11.3 Hz, 1H), 7.05 (d, J = 8.5 Hz, 1H), 6.70 (d, J = 2.7 Hz, 1H), 6.56 (dd, J = 8.6, 2.7 Hz, 1H), 4.76 (q, J = 6.6 Hz, 1H), 2.17 (s, 3H), 1.57 (d, J = 6.6 Hz, 3H), 1.43 (q, J = 4.5 Hz, 2H), 1.15 (q, J = 4.6 Hz, 2H). 13 C NMR (75 MHz, DMSO-d 6 ) δ (ppm) 174.41, 166.57, 152.25, 146.27, 139.05, 135.91, 134.12, 132.10, 131.97, 129.73, 129.52, 129.39, 129.24, 120.76, 119.77, 116.52, 113.28, 45.98, 33.73, 21.89, 20.45, 17.06. HRMS (ESI-TOF) m / z calc’d for C 22 H 22 ClN 5 O 3 [M + H] + 440.1445, found 440.1491。

[0060] Synthesis of 1-(4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzoyl)azetidine-3-carboxylic acid (1-(4-(4-(1-((4-chloro-3-methylphenyl)amin0)ethyl)-1H-1,2,3-triazol-1-yl)benzoyl)azetidine-3-carboxylic acid, Example 4)

[0061]

[0062] A pale yellow solid (0.25 g) was obtained according to General Preparation Method 1, with a yield of 9%. Mp. 88 °C (decomposed). 1 HNMR (300 MHz, DMSO-d 6) δ (ppm) 8.72 (s, 1H), 7.96 (d, J = 8.3 Hz, 2H), 7.80 (d, J = 8.4 Hz, 2H), 7.00 (d, J = 8.6 Hz, 1H), 6.63 (d, J = 2.7 Hz, 1H), 6.50 (dd, J = 8.7, 2.7 Hz, 1H), 4.74 (q, J = 6.5 Hz, 1H), 4.49 (t, J = 8.7 Hz, 1H), 4.42 (d, J = 6.8 Hz, 1H), 4.24 (t, J = 9.6 Hz, 1H), 4.12 (dd, J = 10.1, 5.9 Hz, 1H), 3.46 (dq, J = 9.7, 6.9, 6.5 Hz, 1H), 2.14 (s, 3H), 1.55 (d, J = 6.6 Hz, 3H). 13 C NMR (75 MHz, DMSO-d 6 ) δ (ppm) 174.56, 172.57, 168.09, 152.53, 146.94, 138.49, 135.56, 132.63, 129.77, 129.19, 120.20, 119.77, 115.45, 112.15, 55.77, 51.66, 48.89, 45.09, 32.99, 21.90, 21.43, 20.16. HRMS (ESI-TOF) m / z calc’d for C 22 H 22 ClN 5 O 3 [M + H] + 440.1445, found 440.1477。

[0063] Synthesis of the target compound in Example 5

[0064] Synthesis of N-(but-3-yn-2-yl)-4-fluoro-3-methylaniline (33)

[0065]

[0066] Dissolve 3-butyn-2-ol (0.46 g, 6.62 mmol), DDQ (1.80 g, 7.94 mmol), and triphenylphosphine (2.08 g, 7.94 mmol) in 10 mL of DCM. Add compound 32 (0.83 g, 6.62 mmol) under an ice bath. After 30 min in the ice bath, stir at room temperature for 10 h. After detecting the completion of the reaction by TLC, add a saturated sodium chloride solution to the reaction solution, extract the aqueous phase with DCM, combine the organic phases, evaporate the organic phases to dryness, and purify by silica gel column chromatography (PE:EA = 50:1) to obtain 0.35 g of a pale yellow oily liquid with a yield of 30%. 1 HNMR(300MHz, DMSO-d 6 ) δ (ppm) 7.03 (dd, J = 8.7, 2.9 Hz, 1H), 6.61 (d, J = 3.1 Hz, 1H), 6.55 - 6.44 (m, 1H), 5.95 - 5.82 (m, 1H), 4.16 (t, J = 7.4 Hz, 1H), 3.10 - 3.02 (m, 1H), 2.20 (d, J = 2.9 Hz, 3H), 1.33 (dd, J = 6.9, 3.0 Hz, 3H).

[0067] Synthesis of 4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzoic acid (34)

[0068]

[0069] Dissolve compound 33 (0.35 g, 1.98 mmol), 4-azidobenzoic acid (0.32 g, 1.98 mmol), CuSO 4 ·5H 2 O (4.94 mg, 19.80 μmol), and sodium ascorbate (19.61 mg, 99.00 μmol) in a mixed solvent of tert-butanol:water = 1:2. Stir at room temperature for 10 h. After detecting the completion of the reaction by TLC, add a saturated sodium chloride solution to the reaction solution, extract the aqueous phase with EA, combine the organic phases, and evaporate the organic phases to dryness to obtain 0.27 g of the crude product of the compound with a yield of 40%.

[0070] Preparation of the target compound, Example 5:

[0071] (1) Methyl 1-aminocyclopropane-1-carboxylate (0.09 g, 0.79 mmol), EDCI-HCl (0.30 g, 1.58 mmol), HOBT (0.21 g, 1.58 mmol), and TEA (0.16 g, 1.58 mmol) were dissolved in DMF. Compound 34 (0.27 g, 0.79 mmol) was added under an ice bath. After 30 min in the ice bath, the mixture was stirred at room temperature for 10 h. After monitoring the completion of the reaction by TLC, saturated sodium chloride solution was added to the reaction solution. The aqueous phase was extracted with EA, and the organic phases were combined, dried by evaporation, and purified by silica gel column chromatography (PE:EA = 2:1) to obtain 0.24 g of a pale yellow oily liquid with a yield of 70%.

[0072]

[0073] (2) Then, the product was dissolved in 1 mL of methanol, saturated aqueous KOH solution was added, and after addition, the reaction was stirred at room temperature. The reaction was monitored by TLC. After completion of the reaction, 2N HCl was added to adjust the system to pH = 7, and a pale yellow solid precipitated. Stirring or sonication was continued to promote the aggregation of the precipitated solid. The solid was filtered by suction, and the filter cake was dried to obtain 0.22 g of a pale yellow solid. The total yield of the two steps was 66%. Synthesis of 1-(4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzamido)cyclopropane-1-carboxylic acid (1-(4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)benzamido)cyclopropane-1-carboxylic acid, Example 5)

[0074]

[0075] A pale yellow solid (0.22 g) was obtained according to the above method with a yield of 8%. Mp. 90 °C (decomposed). 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm) 9.27 (s, 1H), 8.98 (d, J = 2.4 Hz, 1H), 8.11 (dd, J = 8.8, 2.5 Hz, 2H), 8.02 - 7.92 (m, 2H), 7.16 (d, J = 6.5 Hz, 1H), 7.05 (d, J = 7.2 Hz, 2H), 5.01 - 4.88 (m, 1H), 2.14 (s, 3H), 1.77 - 1.68 (m, 3H), 1.42 (q, J = 5.0, 3.5 Hz, 2H), 1.16 (s, 2H). 13 C NMR (75 MHz, DMSO-d 6) δ (pm) 174.34, 166.51, 148.84, 138.84, 134.31, 132.05, 131.92, 129.75, 129.36, 125.52, 125.28, 122.24, 119.88, 116.03, 115.72, 50.03, 33.69, 29.52, 19.98, 17.01, 14.88, 14.84. HRMS(ESI-TOF) m / z calc’d for C 22 H 22 FN 5 O 3 [M + H] + 424.1740, found 424.1773。

[0076] Synthesis of target compounds in Examples 6 - 9

[0077] Synthesis of 4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoic acid (35)

[0078]

[0079] Dissolve compound 30 (0.45 g, 2.31 mmol), 4-azidobenzoic acid (0.41 g, 2.31 mmol), CuSO 4 ·5H 2 O (5.79 mg, 23.17 μmol), and sodium ascorbate (22.95 mg, 115.85 μmol) in a mixed solvent of tert-butanol:water = 1:2, and stir at room temperature for 10 h. After detecting the completion of the reaction by TLC, add saturated brine solution to the reaction solution, extract the aqueous phase with EA, combine the organic phases, and evaporate the organic phases to dryness to obtain 0.35 g of the crude product of the compound, with a yield of 41%.

[0080] General Preparation Method 2:

[0081]

[0082] (1) R 6 -NH 2(0.94 mmol), EDCI-HCl (0.36 g, 1.88 mmol), HOBT (0.25 g, 1.88 mmol), and TEA (0.19 g, 1.88 mmol) were dissolved in DMF. Compound 35 (0.35 g, 0.94 mmol) was added under an ice bath. After 30 min in the ice bath, the mixture was stirred at room temperature for 10 h. After the reaction was completed as detected by TLC, saturated saline solution was added to the reaction solution, and the aqueous phase was extracted with EA. The organic phases were combined, dried by evaporation, and purified by silica gel column chromatography (PE:EA = 2:1) to obtain a pale yellow oily liquid with a yield of 70% - 81%.

[0083] (2) Then, the product was dissolved in 1 mL of methanol, saturated aqueous KOH solution was added, and after addition, the mixture was stirred at room temperature for the reaction. Monitored by TLC, after the reaction was complete, 2N HCl was added to adjust the system to pH = 7, and a solid precipitated. Stirring was continued or ultrasonic treatment was used to make the precipitated solid tend to aggregate. Filtration was carried out by suction, and the filter cake was dried to obtain a pale yellow solid with an overall yield of 57% - 69%.

[0084] Synthesis of ((4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoyl)-L-alanine (Example 6)

[0085]

[0086] 0.29 g of a pale yellow solid was obtained according to the general preparation method II with a yield of 10%. Mp. 132 °C (decomposed). 1 HNMR (300 MHz, DMSO-d 6 ) δ (ppm) 8.82 (s, 1H), 8.73 (d, J = 7.2 Hz, 1H), 7.83 - 7.71 (m, 2H), 7.54 (d, J = 8.2 Hz, 1H), 7.20 (d, J = 8.6 Hz, 1H), 6.97 (d, J = 2.9 Hz, 1H), 6.81 (dd, J = 8.6, 2.7 Hz, 1H), 4.87 (q, J = 6.7 Hz, 1H), 4.41 (p, J = 7.3 Hz, 1H), 2.44 (s, 3H), 2.22 (s, 3H), 1.65 (d, J = 6.7 Hz, 3H), 1.38 (d, J = 7.3 Hz, 3H). 13 C NMR (75 MHz, DMSO-d 6) δ (ppm) 174.59, 168.53, 149.58, 142.13, 138.44, 137.34, 137.07, 136.43, 129.75, 129.41, 125.66, 121.89, 121.69, 120.01, 117.24, 116.79, 49.07, 48.50, 20.49, 20.33, 19.86, 17.28. HRMS(ESI-TOF) m / z calc’d for C 22 H 24 ClN 5 O 3 [M + H] + 442.1601, found 442.1631。

[0087] Synthesis of 1-(4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzamido)cyclopropane-1-carboxylic acid (1-(4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzamido)cyclopropane-1-carboxylic acid, Example 7)

[0088]

[0089] A pale yellow solid (0.27 g) was obtained according to General Preparation Method 2, with a yield of 9%. Mp. 104 °C (decomposed). 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm) 8.08 (s, 1H), 7.97 (s, 1H), 7.10 - 6.98 (m, 2H), 6.85 (d, J = 8.2 Hz, 1H), 6.32 (d, J = 8.6 Hz, 1H), 5.95 (d, J = 2.7 Hz, 1H), 5.82 (dd, J = 8.7, 2.7 Hz, 1H), 4.06 (q, J = 6.6 Hz, 1H), 1.76 (s, 3H), 1.47 (s, 3H), 0.87 (d, J = 6.6 Hz, 3H), 0.65 (d, J = 6.2 Hz, 2H), 0.38 - 0.29 (m, 2H). 13 13C NMR (75 MHz, DMSO-d 6) δ (ppm) 174.98, 169.06, 152.03, 146.50, 137.70, 136.91, 136.66, 135.16, 128.77, 128.66, 121.08, 119.82, 119.69, 116.46, 115.13, 111.74, 48.47, 44.71, 33.88, 31.33, 29.24, 21.58, 19.71, 19.23, 15.47. HRMS(ESI-TOF) m / z calc’d for C 23 H 24 C1N 5 O 3 [M + H] + 454.1601, found 454.1633.

[0090] (Synthesis of (4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoyl)-D-alanine (Example 8))

[0091]

[0092] A pale yellow solid (0.22 g) was obtained according to General Preparation Method 2, with a yield of 8%. Mp. 104 °C (decomposed). 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm) 8.76 - 8.65 (m, 2H), 7.86 - 7.74 (m, 2H), 7.54 (d, J = 8.2 Hz, 1H), 7.05 (d, J = 8.6 Hz, 1H), 6.65 (d, J = 2.8 Hz, 1H), 6.51 (dd, J = 8.7, 2.8 Hz, 1H), 6.25 - 6.17 (m, 1H), 4.80 - 4.70 (m, 1H), 4.40 (p, J = 7.3 Hz, 1H), 2.45 (s, 3H), 2.19 (s, 3H), 1.57 (d, J = 6.7 Hz, 3H), 1.38 (d, J = 7.3 Hz, 3H). 13 13C NMR (75 MHz, DMSO-d 6) δ (ppm) 174.69, 168.42, 152.56, 147.18, 138.34, 137.43, 136.90, 135.70, 129.40, 121.74, 120.36, 120.23, 117.07, 115.54, 112.31, 48.59, 45.25, 22.12, 20.41, 19.86, 17.40. HRMS(ESI-TOF) m / z calc’d for C 22 H 24 ClN 5 O 3 [M + H] + 442.1601, found 442.1636.

[0093] Synthesis of 1-(4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoyl)azetidine-3-carboxylic acid (1-(4-(4-(1-((4-chloro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoyl)azetidine-3-carboxylic acid, Example 9)

[0094]

[0095] A pale yellow solid (0.31 g) was obtained according to General Preparation Method 2, with a yield of 10%. Mp. 112 °C (decomposed). 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm) 8.67 (s, 1H), 7.83 (d, J = 2.1 Hz, 1H), 7.73 (dd, J = 8.2, 2.2 Hz, 1H), 7.44 (d, J = 8.3 Hz, 1H), 7.01 (d, J = 8.6 Hz, 1H), 6.62 (d, J = 2.7 Hz, 1H), 6.49 (dd, J = 8.7, 2.7 Hz, 1H), 4.73 (q, J = 6.7 Hz, 1H), 4.23 (t, J = 9.5 Hz, 1H), 4.15 - 3.98 (m, 2H), 3.95 (d, J = 8.0 Hz, 1H), 3.51 - 3.37 (m, 1H), 2.38 (s, 3H), 2.16 (s, 3H), 1.55 (d, J = 6.6 Hz, 3H). 13 13C NMR (75 MHz, DMSO-d 6) δ (ppm) 174.22, 169.31, 152.39, 146.96, 137.85, 137.32, 135.51, 133.81, 129.17, 128.51, 121.88, 120.12, 117.19, 115.41, 112.14, 53.68, 50.96, 48.86, 45.13, 32.32, 21.95, 20.16, 19.24. HRMS (ESI-TOF) m / z calc’d for C 23 H 24 ClN 5 O 3 [M + H] + 454.1601, found 454.1636。

[0096] Synthesis of target compounds in Examples 10 - 11

[0097] Synthesis of 4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoic acid (4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoic acid, 36)

[0098]

[0099] Dissolve compound 33 (0.35 g, 1.98 mmol), 4-azido-2-methylbenzoic acid (0.35 g, 1.98 mmol), CuSO 4 ·5H 2 O (4.94 mg, 19.80 μmol), and sodium ascorbate (19.61 mg, 99.00 μmol) in a mixed solvent of tert-butanol:water = 1:2, and stir at room temperature for 10 h. After detecting the completion of the reaction by TLC, add saturated brine solution to the reaction solution, extract the aqueous phase with EA, combine the organic phases, and evaporate the organic phases to dryness to obtain 0.28 g of the crude product of the compound, with a yield of 40%.

[0100] General Preparation Method 3:

[0101]

[0102] (1) R 6 -NH 2(0.79 mmol), EDCI-HCl (0.30 g, 1.58 mmol), HOBT (0.21 g, 1.58 mmol), and TEA (0.16 g, 1.58 mmol) were dissolved in DMF. Compound 36 (0.28 g, 0.79 mmol) was added under an ice bath. After 30 min in the ice bath, the mixture was stirred at room temperature for 10 h. After the reaction was completed as detected by TLC, saturated sodium chloride solution was added to the reaction solution, and the aqueous phase was extracted with EA. The organic phases were combined, dried by evaporation, and purified by silica gel column chromatography (PE:EA = 2:1) to obtain a pale yellow oily liquid with a yield of 70% - 86%.

[0103] (2) Then, the product was dissolved in 1 mL of methanol, saturated aqueous KOH solution was added, and after addition, the reaction was stirred at room temperature. Monitored by TLC, after the reaction was complete, 2N HCl was added to adjust the system to pH = 7, and a solid precipitated. Stirring or sonication was continued to make the precipitated solid tend to aggregate. The solid was filtered by suction, and the filter cake was dried to obtain a pale yellow solid with a total yield of 58% - 75% for the two steps.

[0104] Synthesis of 1-(4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzamido)cyclopropane-1-carboxylic acid (1-(4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzamido)cyclopropane-1-carboxylic acid, Example 10)

[0105]

[0106] 0.25 g of a pale yellow solid was obtained according to the general preparation method III with a yield of 9%. Mp. 170 °C (decomposed). 1 HNMR (300 MHz, DMSO-d 6 ) δ (ppm) 8.11 (s, 1H), 7.96 (s, 1H), 7.04 (d, J = 13.2 Hz, 2H), 6.85 (d, J = 8.2 Hz, 1H), 6.10 (t, J = 9.1 Hz, 1H), 5.92 - 5.83 (m, 1H), 5.78 (dt, J = 7.9, 3.6 Hz, 1H), 4.05 (d, J = 6.6 Hz, 1H), 1.76 (s, 3H), 1.39 (s, 3H), 0.86 (d, J = 6.5 Hz, 3H), 0.68 (d, J = 6.4 Hz, 2H), 0.35 (s, 2H). 13 C NMR (75 MHz, DMSO-d 6) δ (ppm) 175.92, 169.98, 155.44, 153.22, 152.38, 144.81, 138.59, 137.72, 137.55, 129.57, 124.85, 124.62, 121.92, 120.53, 117.29, 116.05, 115.70, 115.40, 111.81, 111.72, 46.04, 34.67, 32.19, 22.60, 20.13, 16.44, 15.22, 15.18. HRMS(ESI-TOF) m / z calc’d for C 23 H 24 FN 5 O 3 [M + H] + 438.1897, found 438.1932.

[0107] (4-(4-(1-((4-Fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoyl)-L-alanine ((4-(4-(1-((4-fluoro-3-methylphenyl)amino)ethyl)-1H-1,2,3-triazol-1-yl)-2-methylbenzoyl)-L-alanine, Example 11) synthesis

[0108]

[0109] Prepared according to General Preparation Method 3 to obtain 0.18 g of a pale yellow solid, with a yield of 7%. Mp. 195 °C (decomposed). 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm) 8.56 (d, J = 7.2 Hz, 2H), 7.66 (d, J = 11.4 Hz, 2H), 7.49 (d, J = 8.2 Hz, 1H), 6.72 (t, J = 9.2 Hz, 1H), 6.51 (dd, J = 6.7, 2.8 Hz, 1H), 6.42 (dt, J = 7.7, 3.5 Hz, 1H), 4.67 (q, J = 6.7 Hz, 1H), 2.38 (s, 3H), 2.01 (s, 3H), 1.50 (d, J = 6.6 Hz, 3H), 1.36 (d, J = 7.2 Hz, 3H). 13 13C NMR (75 MHz, DMSO-d 6)δ(ppm) 175.32, 168.80, 155.22, 152.98, 152.16, 144.36, 138.39, 137.36, 136.74, 129.28, 124.67, 124.44, 121.74, 120.23, 117.14, 115.92, 115.41, 115.11, 111.60, 111.51, 49.12, 45.69, 31.81, 29.71, 22.32, 19.76, 17.52, 14.84, 14.80. HRMS(ESI-TOF) m / z calc’d for C 22 H 24 FN 5 0 3 [M + H] + 426.1897, found 426.1931.

[0110] Example 2

[0111] The pharmacological experiments and results of the compounds of general formula I in the present invention are as follows:

[0112] Experimental method:

[0113] The compound to be tested was prepared into a gradient stock solution with DMSO, aliquoted and stored frozen at -20°C. A dilution series of the compound to be tested was prepared and transferred into the experimental plate. The specific steps are as follows: Take a LDV 384-well plate, and add 10 μL of each of the above-prepared compound solutions to the wells in sequence. Dilute the compound to be tested with Bravo. After the dilution program is completed, centrifuge at 1000 rpm for 30 seconds. Transfer the prepared compound solution to the experimental plate Corning using ECHO according to the experimental design.

[0114] Prepare the Stimulation buffer and Detection Reagent required for the experiment: 1x Stimulation buffer: Equilibrate the 5x Stimulation buffer in the kit to room temperature, dilute it with ultrapure water at a ratio of 1:4, and add IBMX to a final concentration of 500 μM before use. Detection Reagent: Equilibrate the Lysis&detection buffer in the kit to room temperature, and dilute cAMP-d2 and Anti-cAMP cryptate at a ratio of 1:20 respectively.

[0115] Prepare cell suspension: Digest human S1P1 cells on the culture dish with 0.25% trypsin, then wash the cells with the culture medium and collect them into a 15 mL centrifuge tube. Centrifuge at 1000 rpm for 5 minutes, and then discard the supernatant. Resuspend the cells with 1×Stimulation buffer, count them on a Countess II FL cell counter, and adjust the cell density to 3.0×105 / mL.

[0116] Add the cell suspension to the experimental plate containing the compound using a Multidrop combi, 5 μL per well. Add the prepared 5 μM Forskolin + 1 μM S1P (in stimulation buffer) to the entire plate using a Mantis, 5 μL per well. Apply the film, centrifuge at 300 rpm for 30 seconds, and then incubate at 37 °C for 30 minutes. Add 5 μL of the above-diluted cAMP-d2 + Anti-cAMP cryptate detection reagent to each well of the experimental plate. Centrifuge the experimental plate at 300 rpm for 30 seconds, then let it stand at room temperature for 60 minutes, and then read the values on an Envision.

[0117] Table 1. Antagonistic activity of the compound against S1PR 1 receptors at the cellular level:

[0118] Number <![CDATA[IC 50 (μM)]]> 1 3.57±0.71 2 38.14±1.43 3 3.20±0.64 4 0.83±0.08 5 3.39±0.33 6 0.65±0.02 7 3.16±0.63 8 0.91±0.15 9 1.69±0.35 10 1.34±0.41 11 1.42±0.24

[0119] The hot plate method is a commonly used method for screening and detecting analgesic drugs. It evaluates pain sensitivity by measuring the reaction of animals on a thermostatic hot plate. The hot plate method is not only used for screening and detecting analgesic drugs but also for determining whether the drug acts through the central nervous system or the peripheral nervous system. It has a wide range of applications in pharmacological experiments.

[0120] We divided female mice into four groups and intraperitoneally injected the experimental group mice with doses of 6 at 20 mg / kg, 100 mg / kg, and 200 mg / kg, respectively, while the blank control group was injected with an equal volume of normal saline. We placed the female mice on a thermostatic hot plate at 55 ± 0.5 °C. Evaluate pain sensitivity by measuring the reaction time of the mice licking or lifting their hind paws and turning their heads back, and record the time of the mice licking or lifting their hind paws and turning their heads back at 15 min, 30 min, 60 min, 90 min, and 120 min after injection, respectively.

[0121] As Figure 1As shown, compared with the blank control group, at 15min and 30min after injection, the time for mice to lick their hind feet or lift their hind feet and turn their heads was significantly prolonged, indicating that the drug had a significant analgesic effect on mice during this period. After 30min, the drug was gradually metabolized, and at 60min, the time for mice to lick their hind feet or lift their hind feet and turn their heads was significantly shortened, indicating that most of the drug had been metabolized during this period, or it may be due to damage to the mouse's feet that made it more sensitive to thermal stimulation. At the same time, it can be seen that the analgesic effect of compound 6 on mice is dose-dependent, and high doses are more effective than low doses.

Claims

1. A 1H-triazole biphenyl derivative as represented by the general formula (I) or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, or pharmaceutically acceptable salt: R 1 Selected from H, C 1-6 Alkyl, C 3-6 Cycloalkyl, in, R4 is selected from H, OH, C 1-6 Alkyl; R5 is selected from H, C 1-6 Alkyl; n = 1, 2; R2 is selected from H, NO2, CN, OH, NH2, F, Cl, Br, I, C 1-6 Alkyl, halo-C 1-6 Alkyl, C 1-6 Alkoxy, halo-C 1-6 Alkoxy; R3 is selected from halogen, halo-C 1-6 Alkyl, C 1-6 alkyl.

2. The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, pharmaceutically acceptable salt according to claim 1, characterized in that: R 1 Selected from 3. The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, pharmaceutically acceptable salt according to claim 1, characterized in that: R2 is selected from H, NO2, CN, OH, NH2, F, Cl, Br, I, C 1-4 alkyl.

4. The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, pharmaceutically acceptable salt according to claim 1, characterized in that: R3 is selected from Cl, F.

5. The 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, pharmaceutically acceptable salt according to claim 1, characterized in that: The 1H-triazole biphenyl derivative is selected from the following compounds:

6. A method for preparing the 1H-triazole biphenyl derivative or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, or pharmaceutically acceptable salt as claimed in claim 1, characterized in that: The steps include: (1) reacting a compound of the general formula A with an alcohol to obtain a compound of the general formula B; (2) Compound B undergoes a click reaction with azide to obtain compound C; (3) Compound C of the general formula is connected with an amine derivative and deesterified and hydrolyzed under alkaline conditions to obtain compound D of the general formula; R1′ is R1 and CH3OH or CH3-(CH2) 1～4 -OH formed esters; Compound D is a compound of formula (I), R 1 , R 2 , R 3 As described in claim 1.

7. A pharmaceutical composition, characterized in that It contains the 1H-triazole biphenyl derivative according to claim 1 or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, and pharmaceutically acceptable salt.

8. Use of the 1H-triazole biphenyl derivative according to claim 1 or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, pharmaceutically acceptable salt, or the pharmaceutical composition according to claim 7 in the preparation of an S1PR1 antagonist drug.

9. Use of the 1H-triazole biphenyl derivative according to claim 1 or its enantiomer, diastereomer, tautomer, N-oxide, solvate, physiologically hydrolyzable ester, pharmaceutically acceptable salt or the pharmaceutical composition according to claim 7 in the preparation of analgesic drugs.

10. The use according to claim 8 or 9, characterized in that The drug is added with pharmaceutically acceptable excipients to prepare different dosage forms.

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