A fluorophenyl piperidinone compound and derivatives thereof and pharmaceutical uses thereof

Abstract

The application discloses a fluorophenyl piperidone compound and a derivative and a pharmaceutical application thereof, which can specifically activate a TGR5 receptor, pass through a blood-brain barrier, and be used for preventing or treating central nervous diseases such as chronic pain, anxiety, depression and the like in which the TGR5 receptor is involved.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of medicine, and relates to a fluorophenyl piperidinone compound and derivatives thereof and a pharmaceutical use thereof, in particular to a new type of TGR5 agonist of fluorophenyl piperidinone and the use of the compound in preventing or treating central nervous system diseases such as chronic pain, anxiety, depression and the like which are regulated by TGR5 receptor. BACKGROUND

[0002] Pain perception is one of the most important functions of the central nervous system, and pain lasting more than 6 months is chronic pain. Chronic pain not only manifests as hyperalgesia and allodynia, but also often accompanies mental diseases such as anxiety, depression, cognitive impairment, sleep disorders and the like. Chronic pain seriously troubles about 20% of the global population, leading to loss of working capacity, drug abuse, suicide and the like, and brings great pain and heavy burden to patients and families. Classical analgesic drugs include opioids and non-steroidal anti-inflammatory drugs, and long-term use of opioids can cause serious side effects such as body tolerance and addiction; and non-steroidal anti-inflammatory drugs are also limited in clinical application due to various adverse reactions. Therefore, exploring novel therapeutic targets has been a hot spot in the field of international pain research.

[0003] TGR5 is a bile acid receptor located on the cell membrane, also known as G protein-coupled bile acid receptor 1 (GPBAR1), and is mainly distributed in organs such as gallbladder, intestine, liver, fat, muscle, placenta, brain and the like. As a G protein-coupled receptor, TGR5, after binding with bile acid, couples with Gαs subunit, activates adenylyl cyclase, and increases the intracellular cyclic adenosine monophosphate (cAMP) level, thereby regulating the development of diseases related to metabolism, inflammation and digestive system. More and more studies have shown that TGR5 is widely expressed in neurons, astrocytes and microglia in the central nervous system, participates in the occurrence of diseases such as anxiety, depression, cognitive impairment and neuropathic pain, and plays an important role in central nervous system diseases. Our research group has found that TGR5 is expressed in the anterior cingulate cortex, a key brain area for perceiving pain, and activation of TGR5 can significantly relieve pain response in mice and improve depression-like behavior caused by pain. Therefore, TGR5 can be used as an effective target for treating chronic pain and mental diseases co-occurring with chronic pain.

[0004] At present, major pharmaceutical companies have actively invested in the research and development of TGR5 related small molecules, and have found various TGR5 small molecule agonists with different chemical structures and good activity, such as INT-777 and SB756050. However, the performance of INT-777 and SB756050 in clinical experiments is not satisfactory, which may be due to the low intestinal absorption and blood-brain barrier permeability of these drugs. Therefore, it has great research value to develop new TGR5 agonists for preventing or treating central nervous system diseases such as chronic pain.

[0005] In the previous work, the present inventors, by means of computer-aided drug design supercomputing platform, after virtual screening of 1.5 million bioactive small molecules, found that fluorophenyl piperidone compounds have strong TGR5 agonistic activity. The molecules are first discovered to have TGR5 agonistic activity, can relieve chronic pain, anxiety, depression and other central nervous system diseases, and have great research value. SUMMARY

[0006] The object of the present application is to provide a fluorophenyl piperidone compound and its derivatives and its pharmaceutical use, the use of the compound in preventing or treating chronic pain and other central nervous system diseases regulated by TGR5 receptor.

[0007] In order to achieve the above object, the technical scheme adopted by the present application is:

[0008] A fluorophenyl piperidone compound, the structural formula is as follows:

[0009]

[0010] Among them:

[0011] X is H or hydrogen, hydroxyl, fluorine, chlorine or C1-C5 alkyl;

[0012] X1 is carbon or nitrogen;

[0013] X2 is carbon, oxygen or sulfur;

[0014] R1 is methyl, ethyl, propyl, isopropyl, cyclopropyl, methoxy, ethoxy, propoxy;

[0015] R2 is a substituted or unsubstituted 5-7 membered heteroaromatic ring, a substituted or unsubstituted 5-7 membered saturated heterocyclic ring; or R2 and C at position 2 form an unsubstituted or substituted C2-C6 cycloalkyl;

[0016] Individually represent R configuration, S configuration or racemic.

[0017] Further, in the compound structural formula, R1 is methyl, ethyl, isopropyl.

[0018] Further, in the compound structural formula, X is hydrogen.

[0019] Further, the pharmaceutically acceptable inorganic acid salt of the compound is hydrochloride, sulfate, hydrobromide or phosphate.

[0020] Further, the pharmaceutically acceptable organic acid salt of the compound is oxalate, maleate, benzoate or fumarate.

[0021] Further, the compound has an asymmetric or chiral center, and can exist in the form of a racemate, R-isomer or S-isomer.

[0022] Further, the compound is applied to preparation of a drug for preventing or treating a central nervous system disease regulated by TGR5.

[0023] Further, the compound can be used alone or mixed with a pharmaceutically acceptable excipient, i.e., an excipient, a diluent, to prepare a tablet, a capsule, a granule or a syrup for oral administration.

[0024] Further, the central nervous system disease is a central nervous disorder disease of chronic pain and chronic pain comorbid.

[0025] Further, the present application also provides a preparation method of the compound L2303, and a synthetic route thereof is as follows:

[0026]

[0027] A derivative structure of a fluorophenyl piperidone compound is as follows:

[0028]

[0029] The present application has the following beneficial effects:

[0030] The present application provides a fluorophenyl piperidine and a derivative thereof, which can effectively stimulate TGR5 receptors, has higher safety compared with existing TGR5 agonists, can penetrate the blood-brain barrier, relieve lipopolysaccharide-induced microglial inflammatory response, inhibit mechanical allodynia and thermal pain of mice after CFA injection, and effectively relieve pain with anxiety-like abnormal behavior. Therefore, the fluorophenyl piperidone compound is a new type of TGR5 small molecule agonist, which has the potential to develop into a new analgesic drug.

[0031] The compound has the use in preparation of a drug for relieving chronic pain and other mental diseases, can significantly reduce the content of pro-inflammatory factors IL-6 and IL-1β in lipopolysaccharide LPS-induced microglial cells and reduce the inflammatory response of neurons in an in vitro experiment; the effect is equivalent to the effect of TGR5 specific agonist INT-777, has no obvious cytotoxicity and can penetrate the blood-brain barrier. In a chronic inflammatory pain model induced by complete Freund's adjuvant (CFA), the compound shows good analgesic effect and has a concentration-dependent effect, which is equivalent to the effect of non-steroidal anti-inflammatory drug celecoxib at a corresponding concentration, and exhibits a strong anti-anxiety behavior effect. BRIEF DESCRIPTION OF DRAWINGS

[0032] Figure 1TGR5 is activated in a dose-dependent manner by compounds L2302 and their derivatives.

[0033] Figure 2 L2302 and its derivatives can alleviate LPS-induced inflammatory responses. Specifically, A. L2302 and its derivatives can reduce the level of LPS-induced pro-inflammatory cytokine IL-6; B. L2302 and its derivatives can reduce the level of LPS-induced pro-inflammatory cytokine IL-1β. n=6-8, **** P < 0.0001, *** P < 0.001 vs control; # P < 0.05 ## P < 0.01INT-777 vs LPS; & P < 0.05 && P < 0.01, L-2302 vs LPS; $ P < 0.05 $$ P < 0.01, L-2303 vs LPS.

[0034] Figure 3 The results are ADMET predictions for compound L2302 and its derivatives.

[0035] Figure 4 L2302 reversed CFA-induced persistent mechanical and thermal pain, as shown in A. Schematic diagram of the experimental process; B and C. L2302 alleviated mechanical pain in the ipsilateral and contralateral paws of CFA-treated mice; D. L2302 reduced paw thickness in CFA-treated mice; E. L2302 alleviated ipsilateral thermal pain behavior in CFA-treated mice. n = 5-6, *p < 0.05, **p < 0.01, ***p < 0.001 vs sham; # p<0.05, ## p<0.01 vs. CFA injection group.

[0036] Figure 5 In the open field assay, compound L2302 improved anxiety-like behavior induced by cephalomyelitis (CFA) in mice. Specifically, A. L2302 (10 mg / kg) significantly increased the time spent in the central region by CFA mice; B. L2302 (10 mg / kg) significantly increased the distance traveled in the central region by CFA mice; C. There was no significant difference in the total distance traveled among the groups in the open field assay. n = 5-6 * P<0.05 vs sham; # P<0.05 vs CFA injection group. Detailed Implementation

[0037] The application will be further described in connection with the following examples without being limited to the following examples.

[0038] Example 1. Screening of TGR5 small molecule agonists

[0039] Construction of TGR5 stable cell line: According to the instruction manual, 2 μg of constructed hTGR5 plasmid (pRK-HA-human-TGR5) was added to HEK293 cells for expression using Lipofectamine 2000. The function of the cell line was detected using TGR5 specific agonist INT-777 to confirm the successful construction of the TGR5 stable cell line.

[0040] Screening of TGR5 small molecule agonists: With the aid of computer-aided drug design supercomputing platform, 1.5 million small molecule compound library was subjected to virtual screening, and potential candidate compounds were obtained according to the score value and binding mode. The top 200 candidate compounds with higher activity were selected for TGR5 agonistic activity experiment determination. Using HTRF homogeneous time-resolved fluorescence technology, according to the cAMPGs dynamic kit (PerkinElmer (Cisbio) operation scheme, the TGR5 stably expressed cells were resuspended in a buffer solution containing 1 x stimulation buffer + 500 μM of 3-isobutyl-1-methylxanthine (IBMX). 8000 cells / well / 5 μL cell density was inoculated into a 384-well plate, then 5 μL (50 μM) of the test compound was added, INT-777 was used as a positive control drug, mixed uniformly, and stimulated for 40 min in a 37°C cell incubator. 5 μL of diluted cAMP-d2 and cAMP-Eu was added to the cells, and reacted at room temperature for 1 h. A multifunctional microplate reader was used for detection, with an excitation wavelength of 320 nm and an emission wavelength of 620 nm and 665 nm. The cAMP content was compared by calculating the 620 / 665 nm fluorescence ratio.

[0041] Experimental results: Among the 200 candidate active small molecules, 3 compounds showed TGR5 agonistic activity. Among them, compound L2302 (product of example 1) showed the highest TGR5 activation activity and had stable repeatability (as shown in Figure 1 Table 1). The test results are shown in Table 1, and the structural formula is as follows:

[0042]

[0043] Example 2. Anti-inflammatory effect of L2302 and its derivatives

[0044] Compound L2303 is an optimization based on the structure of L2302, and the structure is as follows:

[0045]

[0046] Specific synthesis steps of compound L2303

[0047] 6-((tert-butyldimethylsilyl)oxy)-3,4-dihydroisoquinolin-1(2H)-one (Compound 1)

[0048]

[0049] Compound 1 (2.5 g, 9.0 mmol) was added in dry anhydrous CH2CI2(40 mL) and refluxed for 3 h. The reaction was cooled and water 100 mL was added and pH was adjusted to 9 using K2CO3. The organic phase was extracted with 3 x 50 mL chloroform and the combined organic phase was washed with 1 x 75 mL brine and dried over Na2S04. Concentration under vacuum gave yellow oil 2.03 g in 70% yield.

[0050] 1 H NMR (600 MHz, DMSO-d6) δ 7.67 (s, 1H), 7.59 (d, J = 7.8 Hz, 1H), 6.70 - 6.89 (m, 2H), 6.63 (d, J = 2.3 Hz, 1H), 3.53 - 3.32 (m, 2H), 3.16 - 2.80 (m, 2H), 0.98 (s, 9H), 0.21 (s, 6H). ESI-HRMS m / z: calcd for C 15 H 23 NO2Si(MH + ), 278.1571, found 278.1564.

[0051] 6-((tert-butyldimethylsilyl)oxy)-3-chloroisoquinoline-2(lH)-carbaldehyde (Compound 2)

[0052]

[0053] Compound 1 (2.5 g, 9.0 mmol) was added in dry anhydrous CH2CI2(40 mL) and refluxed for 3 h. The reaction was cooled and water 100 mL was added and pH was adjusted to 9 using K2CO3. The organic phase was extracted with 3 x 50 mL chloroform and the combined organic phase was washed with 1 x 75 mL brine and dried over Na2S04. Concentration under vacuum gave yellow oil 2.03 g in 70% yield.

[0054] 1 H NMR (400 MHz, CDC13) δ 9.17 (s, 1H), 7.92 (s, 1H), 7.06 (dd, J = 7.8, 3.2 Hz, 1H), 6.72 - 6.68 (m, 2H), 4.22 (s, 2H), 0.98 (s, 9H), 0.21 (s, 6H). ESI-HRMS m / z: calcd for C 16 H 22 ClNO2Si(MH + ), 324.1181, found 324.1179.

[0055] 6-((tert-butyldimethylsilyl)oxy)-3-(3-fluorophenyl)isoquinoline-2(lH)-carbaldehyde (Compound 3)

[0056]

[0057] Compound 2 (1.8 g, 5.57 mmol), 3-fluorophenylboronic acid (1.16 g, 8.36 mmol) and cesium fluoride (2.54 g, 16.72 mmol) were added to a solution of acetonitrile: water (4: 1, 0.2 M) and flushed with nitrogen for 1 h. After addition of DPPF-dichloropalladium(ll) (203 mg, 0.28 mmol) it was reacted at 80 °C for 16 h. The reaction was cooled, washed with 50 mL ethyl acetate, 3 x 25 mL water and 25 mL brine, dried over Na2S04and the solvent was removed in vacuo. Purification by column chromatography eluting with ethyl acetate / hexane = 1 :4 gave the product as an oil 1.41 g, yield: 66%.

[0058] 1 H NMR (400 MHz, CDC13) δ 9.17 (s, 1H), 7.92 (s, 1H), 7.06 (dd, J = 7.8, 3.2 Hz, 1H), 6.72 - 6.68 (m, 2H), 4.22 (s, 2H), 0.98 (s, 9H), 0.21 (s, 6H). ESI-HRMS m / z: calcd for C 22 H 26 FNO2Si(MH + ), 384.1790, found 384.1783.

[0059] 6-((tert-butyldimethylsilyl)oxy)-3-(3-fluorophenyl)-3,4-dihydroisoquinoline-2(lH)-carbaldehyde (Compound 4)

[0060]

[0061] Pd / C (450 mg) was dissolved in ethyl acetate (0.15 M) under nitrogen protection, after adding compound 3 (1.2 g, 3.13 mmol), hydrogen was filled and stirred at room temperature for 16 h. After the reaction was completed, the oil was filtered and concentrated under vacuum to obtain 1.3 g of oil, with a yield of 89%. 1 H NMR (400 MHz, CDC13) δ 8.07 (s, 1H), 7.33 - 7.29 (m, 1H), 7.13 - 6.96 (m, 3H), 7.08 (d, J = 2.3 Hz, 1H), 7.06 - 6.84 (m, 2H), 5.37 - 5.26 (m, 1H), 4.34 - 4.22 (m, 2H), 3.23 - 3.19 (m, 1H), 2.98 - 2.87 (m, 1H), 0.98 (s, 9H), 0.21 (s, 6H). ESI-HRMS m / z: calcd for C 22 H 28 FNO2Si(MH + ), 386.1946, found 386.1940.

[0062] 6-((tert-butyldimethylsilyl)oxy)-3-(3-fluorophenyl)- tetrahydroisoquinoline (Compound 5)

[0063]

[0064] To compound 4 (1.1 g, 2.87 mmol) was added 13 M hydrochloric acid and heated to reflux for 48 h. The reaction was cooled, neutralized with 5 M sodium hydroxide and extracted with 3 x 100 mL CH2Cl2. The organic phase was washed with 1 x 50 mL water, 1 x 50 mL brine, dried over MgSO4and the solvent was removed under vacuum to obtain an oil. Purification by column chromatography eluting with ethyl acetate / hexane = 1 :6 gave the product as an oil 700 mg, yield: 70%.

[0065] 1 H NMR (400 MHz, CDC13) δ 8.03 (s, 1H), 7.42 - 7.36 (m, 1H), 7.28 - 6.96 (m, 3H), 6.97 (dd, J = 8.4, 2.5 Hz, 1H), 6.74 - 6.69 (m, 2H), 5.26 (t, J = 3.4 Hz, 1H), 5.22 - 5.16 (m, 2H), 3.08 - 2.95 (m, 1H), 2.83 - 2.78 (m, 1H), 0.98 (s, 9H), 0.21 (s, 6H). ESI-HRMS m / z: calcd for C 21 H 28FNOSi(MH + ), 358.1997, found 358.1986.

[0066] 6-((tert-butyldimethylsilyl)oxy)-3-(3-fluorophenyl)-3,4-dihydroisoquinoline-2(lH)- 3-methylbutan-l-one (Compound 6)

[0067]

[0068] Compound 5 (600 mg, 1.68 mmol), isovaleric acid (171 mg, 1.68 mmol) and DIEA (326 mg, 2.52 mmol) were dissolved in DMF (20 mL), TBTU (648 mg, 2.01 mmol) was added to the solution. The reaction was stirred at room temperature for 16 h. The reaction was quenched with water, the product was extracted with organic solvent. The organic was dried over MgS04, filtered and concentrated in vacuo to give an oil. Purified by column chromatography eluted with ethyl acetate / hexane = 1 :4 to give 660 mg of oil, yield: 90%.

[0069] 1 H NMR (400 MHz, CDC13) δ 7.43 - 7.36 (m, 1H), 7.26 - 6.98 (m, 3H), 6.94 (dd, J = 7.8, 2.4 Hz, 1H), 6.74 - 6.69 (m, 2H), 5.26 (t, J = 3.4 Hz, 1H), 5.31 - 5.22 (m, 2H), 3.08 - 2.97 (m, 1H), 2.83 - 2.79 (m, 1H), 2.16 - 2.07 (m, 3H), 0.98 - 0.94 (m, 15H), 0.21 (s, 6H). ESI-HRMS m / z: calcd for C 26 H 36 FNO2 Si(MH + ), 442.2572, found 442.2569.

[0070] 1-(3-(3-Fluorophenyl)-6-hydroxy-3,4-dihydroisoquinoline-2(lH)-yl)-3-methylbutan- 1-one (Compound 7)

[0071]

[0072] To a solution of compound 6 (500 mg, 1.13 mmol) in tetrahydrofuran (10 mL) was added TBAF (0.5 mL, 1 M) dropwise with stirring. The solution was stirred at room temperature for 1 h. Concentrated in vacuo to give a mixture. Purified by column chromatography eluted with ethyl acetate / petroleum ether (1 / 100-20 / 100) to give 290 mg of oil, yield 80%.

[0073] 1 H NMR (400 MHz, CDC13) δ 9.29 (s, 1H), 7.43 - 7.36 (m, 1H), 7.11 - 7.09 (m, 3H), 6.97 - 6.94 (m, 1H), 6.77 (s, 1H), 6.62 - 6.59 (m, 1H), 5.26 (t, J = 3.4 Hz, 1H), 4.31 - 4.22 (m, 2H), 3.22 - 2.98 (m, 2H), 2.16 - 2.07 (m, 3H), 0.94 (s, 6H). ESI-HRMS m / z: calcd for C 20 H 22 FNO (MH + ), 328.1707, found 328.1769.

[0074] (2-(hydroxymethyl)oxazol-4-yl)-thiomorpholine-methanone (Compound 8)

[0075]

[0076] Thiomorpholine (5.0 g, 48.5 mmol), 2-(hydroxymethyl)oxazole-4-carboxylic acid (6.9 g, 48.5 mmol) and DIEA (9.4 g, 72.75 mmol) were dissolved in DMF (20 mL) and TBTU (18.7 g, 58.2 mmol) was added to the solution. The reaction was allowed to proceed at room temperature for 16 h. The reaction was quenched with water, extracted, dried over MgS04, filtered and concentrated to an oil. Column chromatography purification with ethyl acetate / hexane = 1 :4 elution gave the product as an oil 9.62 g, yield: 87%.

[0077] 1 H NMR (400 MHz, CDC13) δ 7.69 (s, 1H), 5.39 (s, 1H), 4.79 (s, 2H), 3.33-3.29 (m, 4H), 2.67 - 2.71 (m, 4H). ESI-HRMS m / z: calcd for C9H 12 N2O3S (MH + ), 229.0641, found 229.0638.

[0078] 1-(3-(3-Fluorophenyl)-6-((4-(thiomorpholine-4-carbonyl)oxazol-2-yl)methoxy)-3,4- dihydroisoquinolin-2(lH)-yl)-3-methylbutan-l-one (Compound L2303)

[0079]

[0080] To a solution of compound 7 (200 mg, 0.610 mmol), compound 8 (347 mg, 1.52 mmol) and PPh3 (0.39 mg, 1.52 mmol) in tetrahydrofuran (5 mL) was added DEAD (0.65 mg, 1.52 mmol, 40 wt% in toluene) dropwise at 0-5 °C under nitrogen. The reaction was stirred at room temperature for 1 h and then concentrated. Purification by column chromatography (ethyl acetate:hexane = 1:100-1:1) gave the target compound 200 mg, yield 60%

[0081] 1 H NMR (400 MHz, CDC13) δ 7.69 (s, 1H), 7.36-7.28 (m, 2H), 7.13-7.09 (m, 2H), 6.97-6.92 (m, 2H), 6.79 (d, J = 7.8, 1H), 5.39 (s, 2H), 5.26 (t, J = 3.4 Hz, 1H), 4.51-4.41 (m, 2H), 3.33-3.23 (m, 4H), 2.96-2.87 (m, 2H), 2.71-2.69 (m, 4H), 2.10-2.07 (m, 3H), 0.96-0.87 (m, 6H). ESI-HRMS m / z: calcd for C 29 H 32 FN3O4S (MH + ), 538.2170, found 538.2167.

[0082] Synthesized according to the design of Preparation Example 1. Through in vitro activity detection experiments, it was found that the fluorophenyl piperidone compounds of this type have strong TGR5 agonistic activity and have a significant alleviating effect on the inflammatory response of BV2 microglial cells induced by lipopolysaccharide (LPS).

[0083] TGR5 agonistic activity experiment of L2302 and its derivatives: The stock solution of compounds L2302 and L2303 was diluted 2 times with normal saline, and the activation of L2302 and L2303 on TGR5 receptor was determined according to the experimental protocol of cAMP Gs dynamic kit (PerkinElmer (Cisbio)).

[0084] Experimental results: L2302 and L2303 can activate TGR5 protein in a dose-dependent manner, and the half maximal effective concentration for activating TGR5 is 10.6 μM and 5.43 μM, respectively, and the results are shown in Figure 1 .

[0085] L2302 and its derivatives alleviate LPS-induced BV2 cell inflammatory response: logarithmic growth period of BV2 cells were inoculated in 96-well plates, the cell density was 8000 per well, 100 μL per well. After 24 h, the supernatant was discarded, and different concentrations (0.1, 1.0, 10 μM) of compounds were added to the drug group, 6 parallel holes were set in each group, and the action was 2 h. Add LPS stock solution 100 μL (final concentration 1 μg / ml), and set up blank control group and LPS group, continue to act for 24 h. The supernatant of different test groups of cells was absorbed into the 96-well plate coated with IL-6, IL-1β, and the determination was carried out according to the kit operation steps.

[0086] Experimental results: LPS (1 μg / mL) induced BV2 cell proinflammatory factor IL-6 and IL-1β levels increased significantly, and compounds L-2302 and L-2303 at 0.1 μM, 1.0 μM and 10 μM could significantly reduce the content of IL-6 and IL-1β, which was comparable to the effect of TGR5 specific agonist INT-777. These results show that fluorophenyl piperidone compounds can inhibit LPS-induced inflammatory response of microglial cells, and the detection results are shown in Figure 2 .

[0087] Example 3. Drug evaluation of L2302 and its derivatives

[0088] Experimental method: using the ADME module in Discovery Studio software, input the structure of compound L2302 and L2303, select "Calculate Molecular Properties" in "Small Molecules" module, click ADME descriptors to set parameters, click run, and predict the absorption, distribution and toxicity of the compounds. The data of water solubility, passive intestinal absorption, blood brain barrier penetration, liver toxicity of compounds L2302 and L2303 at 25℃ were obtained.

[0089] Experimental results: compared with INT-777, compounds L2302 and L2303 have good blood brain barrier penetration and gastrointestinal absorption rate, and the ADME prediction results are shown in Figure 3 .

[0090] Example 4. Therapeutic effect of L2302 on pain and anxiety depression

[0091] Experimental animals: C57BL / 6 male mice (about 8 weeks old) were purchased from the experimental animal center of air force medical university. The feeding conditions were 12 h light / 12 h dark cycle, temperature 22-26℃, humidity 55%-60%, free water and food. All animal experiments were approved by the air force medical university experimental animal welfare and ethics committee.

[0092] Experimental method: The mice were randomly divided into 6 groups: control group, CFA group, CFA+1mg / kg L-2302, CFA+5mg / kg L-2302, CFA+10mg / kg L-2302 and CFA+10mg / kg Celebrex positive control group. The mice were injected with 10μL of 50% CFA (CFA: normal saline = 1:1) into the left hind footpad to induce a chronic pain model. L2302 was dissolved in sterile normal saline to prepare different concentrations of 1, 5, and 10mg / kg for the administration groups, which were administered by gavage for 7 consecutive days. The blank control group was given the same volume of sterile normal saline. 60 minutes after administration, the pain threshold of the mice was determined according to the experimental plan

[0093] Mechanical pain threshold test: Von Frey fine fiber filaments in the range of 0.08-2.00g were used to evaluate mechanical sensitivity. The baseline foot pain threshold of the mice was determined before modeling. On the first day, the CFA was injected into the foot according to the grouping, and the control group was treated with normal saline. On the second to eighth day, the mice were administered different doses of L-2302 and Celebrex by gavage. 60 minutes after administration, the Von Frey fiber filament was applied vertically to the center of the mouse's left or right hind paw, and the mouse's mechanical pain was evaluated with the endpoint of causing slight bending. The manifestations of mouse pain included licking the foot, trying to bite the fiber, quickly avoiding, and foot withdrawal, etc. Each mouse was tested twice with an interval of more than 3 minutes, and a total of 10 times. Finally, more than 50% of the positive reaction points were included as the pain threshold.

[0094] Hot pain threshold test: The mice were placed on a constant temperature plate at 55±1℃, and the time required for the mice to lift their feet from the start was recorded as the pain threshold.

[0095] Open field test: The Shanghai Fanbi Intelligent Technology Tracking Master V3.0 acquisition and analysis system was used to place the mice in the open field (40x40x40cm), and the spontaneous activity of the mice within 15 minutes, the residence time in the central and peripheral areas, and the movement distance were recorded and analyzed.

[0096] Experimental results: Injection of CFA can induce obvious mechanical pain and thermal pain in mice, and compound L2302 can reduce CFA-induced pain in a dose- and time-dependent manner. On the seventh day after administration, the analgesic effect reached the highest value. 10mg / kg of compound L2302 can significantly inhibit the mechanical pain and thermal pain behavior of CFA-induced mice, and the analgesic effect is comparable to that of 10mg / kg Celebrex positive control group. The results are shown in Figure 4 In addition, the open field test results show that 10mg / kg of compound L2302 can significantly alleviate the CFA-induced pain and anxiety-like behavior in mice, and the results are shown in Figure 5 .

[0097] Representative compounds were evaluated for their activity to activate TGR5, as shown in Table 1, both compound L2302 and compound L2303 significantly activated TGR5 to produce cAMP. The half maximal effective concentration (EC50) of compound L2302 and L2303 to activate TGR5 was calculated to be 10.7 ± 0.4 μΜ and 5.43 ± 2.3 μΜ (n = 6-8), respectively, by HTRF homogenous time-resolved fluorescence technology.

[0098] Example EC50 (μM) Example 1 10.7 μM Example 2 5.43 μM

Claims

1. A fluorophenylpiperidinone compound, characterized by, The structural formula is as follows (I) or (II): (I) (II).

2. The fluorophenylpiperidinone compound according to claim 1, characterized by, The pharmaceutically acceptable salt of the compound is hydrochloride, sulfate, hydrobromide, phosphate, oxalate, maleate, benzoate or fumarate.

3. Use of the fluoro-phenyl-piperidinone compound of claim 1 in the preparation of a medicament for preventing or treating a central nervous system disease modulated by TGR5.

4. The use of the fluoro-phenylpiperidinone compound according to claim 3 for manufacturing a medicament for preventing or treating a central nervous system disease modulated by TGR5, characterized in that, The compound can be used alone or mixed with a pharmaceutically acceptable excipient.

5. The use of the fluorophenylpiperidone compounds according to claim 3 in the preparation of drugs for the prevention or treatment of TGR5-regulated central nervous system diseases, characterized in that, The central nervous system disease is chronic pain and central nervous system disorder disease comorbid with chronic pain.

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