Compound containing quinolinone skeleton and application thereof

Abstract

The invention discloses a compound containing a quinolinone skeleton and application of the compound, the compound containing the quinolinone skeleton has a structure as shown in a formula (I), and the compound can be used for preparing an MAGL inhibitor and a medicine for preventing and / or treating MAGL-related diseases. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of medicinal chemistry, and particularly to a compound containing a quinolinone skeleton and its application. Background Art

[0002] The endocannabinoid system (ECS) consists of cannabinoid receptors (CB1, CB2), endocannabinoid ligands (arachidonoylethanolamide AEA, 2-arachidonoylglycerol 2-AG), and the hydrolases of cannabinoid ligands. CB receptors are the main targets of tetrahydrocannabinol (THC) in plant cannabis, and have the functions of inhibiting adenylate cyclase activity, activating potassium ion channels, and inhibiting voltage-gated calcium channels. Endocannabinoid 2-AG is a full agonist of CB1 and CB2 receptors, and is synthesized "on demand" from phospholipid precursors through a Ca 2+ -dependent mechanism: Ca 2+ influx activates phospholipase C (PLC), hydrolyzes phosphatidylinositol (PI) into diacylglycerol (DAG), and then 2-AG is generated from DAG by diacylglycerol lipase (DAGL). When 2-AG activates CB receptors on target cells, it will be absorbed and degraded, thus terminating the signal transduction pathway. Monoacylglycerol lipase (MAGL) is responsible for 85% of 2-AG hydrolysis, generating arachidonic acid (AA) and glycerol. Downstream AA is metabolized by downstream hydrolases to generate inflammatory mediators such as prostaglandins.

[0003] MAGL is responsible for hydrolyzing 2-AG to generate AA and glycerol, and participates in the signal transduction of the endocannabinoid system. MAGL is highly expressed in the brain, adipose tissue, liver, and intestine. In the brain, MAGL is expressed in the hippocampus, amygdala, and cerebellum. Endocannabinoid signals play important roles in these tissue sites. Therefore, inhibiting MAGL is promising as a target for central nervous system diseases, pain, or liver diseases. Summary of the Invention

[0004] The purpose of the present invention is to provide a compound with MAGL inhibitory activity.

[0005] The compound with MAGL inhibitory activity provided by the present invention has the structure shown in formula (I):

[0006]

[0007] Wherein, R 1 is selected from aryl or heteroaryl;

[0008] R 2 is selected from substituted or aryl or heteroaryl substituted by any number of R 2A ;

[0009] R 2ASelected from -OH, -SH, -CN, halogen, nitro, carboxyl, C 1-8 alkyl, C 1-8 alkoxy, C 1-4 haloalkyl.

[0010] In some embodiments, R 1 , R 2 , the aryl groups are each independently C 6 -C 10 aryl groups, specifically each independently such as phenyl, naphthyl, and more specifically phenyl.

[0011] In some embodiments, R 1 is a heteroaryl group, and R 2 is a substituted or optionally substituted by any number of R 2A aryl group.

[0012] In some embodiments, R 1 , R 2 , the heteroaryl group is a 5- to 10-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1, 2, 3, or 4".

[0013] In some embodiments, R 1 , R 2 , the heteroaryl group is a 5- to 10-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1, 2, or 3".

[0014] In some embodiments, R 1 , R 2 , the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1, 2, or 3".

[0015] In some embodiments, R 1 , R 2 , the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1 or 2".

[0016] In some embodiments, R 1 , R 2 , the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N and S, and the number of heteroatoms is 1 or 2".

[0017] In some embodiments, R 1 , R 2 , the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N and S, and the number of heteroatoms is 1".

[0018] In some embodiments, R 1 , R 2Among them, the heteroaryl group is furyl, thienyl, pyrrolyl, oxazolyl, isoxazolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, triazolyl, 1,3,4-oxadiazolyl, 1,3,4-thiadiazolyl, 1,2,4-oxadiazolyl, 1,2,4-thiadiazolyl, pyridyl, pyrimidinyl, pyridazinyl, benzofuranyl, indolyl, quinolinyl, isoquinolinyl, indazolyl, benzoxazolyl, benzothiazolyl, purinyl, oxazolo-pyridyl.

[0019] In some embodiments, R 1 , R 2 Among them, the heteroaryl group is pyridyl.

[0020] In some embodiments, R 1 is pyridyl, and R 2 is a phenyl group which is substituted or substituted by any number of R 2A .

[0021] In some embodiments, R 2A is one or more, for example, the number is 1, 2, 3 or 4, more specifically 1 or 2. In some instances, each of R 2A is independently selected from -OH, -SH, -CN, cyano group, halogen, nitro group, carboxyl group, methyl group, ethyl group, n-propyl group, isopropyl group, methoxy group, ethoxy group, -CF 3 , CHF 2 or CH 2 F.

[0022] The present disclosure further provides an isotope substitute of the above compound or its pharmaceutically acceptable salt. In some embodiments, the isotope substitute is a deuterated compound.

[0023] In some specific instances, the present invention provides specific compounds having MAGL inhibitory activity, and the structures are as follows:

[0024]

[0025] The present disclosure provides a preparation method of a compound represented by formula (I). Using commercially available 4-Boc-1-(5-bromo-2-pyridyl)piperazine as a raw material, compound 3 obtained by performing a Suzuki coupling reaction with a substituted phenylboronic acid is deprotected to prepare compound 4, and then compound 4 is subjected to a condensation reaction to prepare the compound of formula (I).

[0026]

[0027] The present disclosure also provides a pharmaceutical composition, which comprises the above compound represented by formula (I) or its pharmaceutically acceptable salt or the above isotope substitute, and a pharmaceutically acceptable excipient.

[0028] In some embodiments, the compound represented by formula (I) or a pharmaceutically acceptable salt thereof or the above-mentioned isotope-substituted compound is in a therapeutically effective amount.

[0029] In some embodiments, based on the total weight of the composition, the pharmaceutical composition contains 0.01-99.99% of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned isotope-substituted compound. In some embodiments, the pharmaceutical composition contains 0.1-99.9% of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned isotope-substituted compound. In some embodiments, the pharmaceutical composition contains 0.5%-99.5% of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned isotope-substituted compound. In some embodiments, the pharmaceutical composition contains 1%-99% of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned isotope-substituted compound. In some embodiments, the pharmaceutical composition contains 2%-98% of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned isotope-substituted compound.

[0030] In some embodiments, based on the total weight of the composition, the pharmaceutical composition contains 0.01%-99.99% of a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition contains 0.1%-99.9% of a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition contains 0.5%-99.5% of a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition contains 1%-99% of a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition contains 2%-98% of a pharmaceutically acceptable excipient.

[0031] The present disclosure also provides the use of the compound represented by formula (I) or a pharmaceutically acceptable salt thereof, the above-mentioned isotope-substituted compound or the above-mentioned pharmaceutical composition in the preparation of a MAGL inhibitor.

[0032] The present disclosure also provides the use of the compound represented by formula (I) or a pharmaceutically acceptable salt thereof, the above-mentioned isotope-substituted compound or the above-mentioned pharmaceutical composition in the preparation of a drug for preventing and / or treating MAGL-related diseases.

[0033] In some embodiments, the MAGL-related diseases are central nervous system diseases, metabolic disorders and inflammatory diseases; in some examples, the MAGL-related diseases are depression, anxiety, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, multiple sclerosis, neuropathic pain, inflammatory pain, cancer pain, epilepsy, cancer, fatty liver, non-alcoholic steatohepatitis, liver fibrosis, cholestasis, inflammatory bowel disease.

[0034] Term Definition

[0035] On the other hand, in cases where the present disclosure does not define a specific configuration, the compounds of the present disclosure can exist in specific geometric or stereoisomeric forms. The present disclosure contemplates all such compounds, including cis and trans isomers, (-)- and (+)- enantiomers, (R)- and (S)- enantiomers, diastereoisomers, (D)-isomers, (L)-isomers, and their racemic mixtures and other mixtures, such as enantiomer- or diastereoisomer-enriched mixtures, all of which mixtures are within the scope of the present disclosure. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All such isomers and their mixtures are included within the scope of the present disclosure.

[0036] In addition, the compounds and intermediates of the present disclosure can also exist in different tautomeric forms, and all such forms are included within the scope of the present disclosure. The term "tautomer" or "tautomeric form" refers to structural isomers of different energies that can interconvert via a low energy barrier.

[0037] The compounds of the present disclosure can be asymmetric, for example, having one or more stereoisomers. Unless otherwise specified, all stereoisomers are included, such as enantiomers and diastereoisomers. Compounds of the present disclosure containing asymmetric carbon atoms can be isolated in optically pure form or in racemic form. The optically pure form can be resolved from the racemic mixture or synthesized by using chiral starting materials or chiral reagents.

[0038] The present disclosure also includes isotopically labeled compounds of the present disclosure that are the same as those described herein, but in which one or more atoms are replaced by atoms having an atomic weight or mass number different from the atomic weight or mass number typically found in nature. Examples of isotopes that can be incorporated into the compounds of the present disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, such as 2 H, 3 H, 11 C, 13 C, 14 C, 13 N, 15 N, 15 O, 17 O, 18 O, 31 P, 32 P, 35 S, 18 F, 123 I, 125 I and 36 Cl, etc.

[0039] Unless otherwise specified, when a position is specifically designated as deuterium (D), that position is understood to have a deuterium abundance greater than the natural abundance of deuterium (which is 0.015%) by at least 1000-fold (i.e., at least 10% deuterium incorporation). The deuterium in the exemplified compounds can have an abundance greater than the natural abundance of deuterium by at least 1000-fold, at least 2000-fold, at least 3000-fold, at least 4000-fold, at least 5000-fold, at least 6000-fold, or a higher abundance of deuterium. The present disclosure also includes various deuterated forms of the compound of formula I. Each available hydrogen atom attached to a carbon atom can be independently replaced by a deuterium atom. Those skilled in the art can refer to relevant literature to synthesize the deuterated forms of the compound of formula I. Commercially available deuterated starting materials can be used in the preparation of the deuterated forms of the compound of formula I, or they can be synthesized using conventional techniques with deuterating reagents, including but not limited to deuterated borane, borane-d3 tetrahydrofuran solution, lithium aluminum deuteride, iodoethane-d1, iodomethane-d3, etc.

[0040] The term "alkyl" refers to a saturated aliphatic hydrocarbon group, which is a straight-chain or branched-chain group containing 1 to 8 carbon atoms, preferably an alkyl group containing 1 to 6 carbon atoms, and more preferably an alkyl group containing 1 to 4 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, etc.

[0041] The term "alkoxy" refers to -O-(alkyl), where alkyl is defined as above. Non-limiting examples of alkoxy include: methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, or tert-butoxy.

[0042] The term "aryl" refers to a 6- to 14-membered fully carbon monocyclic or fused polycyclic (i.e., rings sharing adjacent carbon atom pairs) group having a conjugated π-electron system, preferably 6 to 12 members, such as phenyl and naphthyl.

[0043] The term "heteroaryl" refers to a heteroaromatic system containing 1 to 4 heteroatoms and 5 to 14 ring atoms, where the heteroatoms are selected from oxygen, sulfur, and nitrogen. The heteroaryl is preferably 5 to 12 membered, more preferably 5 or 6 membered. For example, non-limiting examples thereof include: imidazolyl, furyl, thienyl, thiazolyl, pyrazolyl, oxazolyl, isoxazolyl, pyrrolyl, tetrazolyl, pyridyl, pyrimidinyl, thiadiazole, pyrazinyl, triazolyl, indazolyl, benzimidazolyl, etc.

[0044] The term "hydroxy" refers to -OH.

[0045] The term "halogen" refers to fluorine, chlorine, bromine, or iodine.

[0046] The term "haloalkyl" refers to an alkyl group substituted by halogen, where the alkyl group is as defined above.

[0047] The term "haloalkoxy" refers to an alkoxy group substituted by halogen, where the alkoxy group is as defined above.

[0048] The term "cyano" refers to -CN.

[0049] The term "nitro" refers to -NO 2 .

[0050] The term "amino" refers to -NH 2 .

[0051] The term "carboxy" refers to -C(O)OH.

[0052] The term "substituted" means that one or more hydrogen atoms in the group, preferably up to 5, more preferably 1 to 3 hydrogen atoms, are independently replaced by the corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and those skilled in the art can determine the possible or impossible substitutions without much effort (by experiment or theory).

[0053] "Substituted by one or more..." means that it can be substituted by a single or multiple substituents. When substituted by multiple substituents, it can be a plurality of the same substituents or a combination of one or a plurality of different substituents.

[0054] The term "each independently selected from" means that they can be the same or different and are selected from the listed groups.

[0055] In the chemical structure of the compounds described in the present disclosure, the bond represents an unspecified configuration, that is, if there are chiral isomers in the chemical structure, the bond can be or simultaneously contain Two configurations. Although all of the above structural formulas are drawn in certain isomeric forms for simplicity, the present disclosure may include all isomers, such as tautomers, rotamers, geometric isomers, diastereomers, racemates, and enantiomers. In the chemical structure of the compounds described in the present disclosure, the bond is not specified in configuration, that is, the configuration of the bond can be E-type or Z-type, or include both E and Z configurations simultaneously.

[0056] In the present disclosure, the terms "comprising" and "including" can be replaced with "consisting of".

[0057] The term "composition" refers to a mixture of a drug containing one or more of the compounds described herein or their physiologically pharmaceutically acceptable salts or precursors and other chemical components, as well as other components such as physiologically pharmaceutically acceptable carriers and excipients. The purpose of the composition is to facilitate administration to an organism, facilitate absorption of the active ingredient and thereby exert biological activity.

[0058] The term "pharmaceutically acceptable excipient" or "pharmaceutically acceptable excipient" includes, but is not limited to, any adjuvant, carrier, excipient, glidant, sweetening agent, diluent, preservative, dye / colorant, flavoring agent, surfactant, wetting agent, dispersing agent, suspending agent, stabilizing agent, isotonic agent, solvent, or emulsifying agent that has been approved by the US Food and Drug Administration for use in humans or domestic animals.

[0059] Unless otherwise specified, the "compounds" of the present disclosure can independently exist in the form of salts, mixed salts, or non-salts (such as free acids or free bases). When in the form of salts or mixed salts, they can be pharmaceutically acceptable salts or medicinally acceptable salts.

[0060] The terms "pharmaceutically acceptable salt" and "medicinally acceptable salt" can be used interchangeably and refer to pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.

[0061] "Pharmaceutically acceptable acid addition salts" refer to salts formed with inorganic acids or organic acids that can retain the biological effectiveness of the free base without other side effects. These salts can be prepared by methods known in the art.

[0062] "Pharmaceutically acceptable base addition salts" refer to salts formed with inorganic bases or organic bases that can maintain the biological effectiveness of the free acid without other side effects. These salts can be prepared by methods known in the art.

[0063] The compounds described in the present invention have good inhibitory activity against MAGL and have good therapeutic effects on the central nervous system or pain. BRIEF DESCRIPTION OF THE DRAWINGS

[0064] Figure 1 Oil red O staining results of cells in the blank (C), model (M), IV-23-5 μM, and IV-23-10 μM groups.

[0065] Figure 2 Time of mice in the blank, model, JZL184, and IV-23 (10 mg / kg) groups staying on the rotarod.

[0066] Figure 3 Determination results of ALT and ALP contents in mice of the Control, DDC, and IV-26 groups.

[0067] Figure 4 HE staining results of mice in the DDC and IV-26 groups. Detailed implementation manners

[0068] The following further describes the present disclosure in combination with embodiments, but these embodiments do not limit the scope of the present disclosure. For the experimental methods without specific conditions noted in the embodiments of the present disclosure, they are usually carried out under conventional conditions or according to the conditions recommended by the raw material or commodity manufacturers. For reagents without specific sources noted, such reagents can be obtained from any supplier of molecular biology reagents in the quality / purity for molecular biology applications.

[0069] Unless otherwise specified, the reagents used in the following embodiments are all commercially available products.

[0070] In the following preparation methods, the raw material compounds and reagents used in each step and the obtained compounds can each be in the form of salts. Examples of such salts include salts similar to the salts of the compounds of the present invention, etc.

[0071] The compounds obtained in each step can be directly used in the next reaction in the form of a reaction mixture or a crude product. Alternatively, the compounds obtained in each step can be separated from the reaction mixture and purified according to methods known per se, such as concentration, crystallization, recrystallization, distillation, solvent extraction, fractional distillation, column chromatography, etc. When the raw material compounds and reagents used in each step are commercially available, the commercially available products can also be directly used.

[0072] In the reaction of each step, although the reaction time varies depending on the types of reagents and solvents used, it is usually from 1 minute to 48 hours, preferably from 10 minutes to 12 hours, unless otherwise stated.

[0073] In the reaction of each step, although the reaction temperature varies depending on the types of reagents and solvents used, it is usually from 0 °C to 300 °C, preferably from 78 °C to 150 °C, unless otherwise stated.

[0074] Example 1 Preparation of intermediates

[0075] 1. tert-Butyl 4-(5-(4-(trifluoromethyl)phenyl)pyridin-2-yl)piperazine-1-carboxylate (19b)

[0076]

[0077] 4-Boc-1-(5-bromo-2-pyridyl)piperazine (205 mg), 4-trifluoromethylphenylboronic acid (137 mg), Pd(OAc) 2 (3 mg), Na 2 CO 3 (165 mg) were added to a sealed tube. Under N 2 H 2 O and DMF were added, and the mixture was refluxed for 8 h. After the reaction was completed, heating was stopped, diluted with water, extracted three times with ethyl acetate, washed three times with saturated brine, and the organic phase was dried over anhydrous sodium sulfate. Purification by column chromatography (petroleum ether:ethyl acetate = 16:1) gave 19b, 190 mg of white solid, yield 77%.

[0078] 1 1H NMR (300 MHz, DMSO-d6) δ (ppm): 8.55 (d, J = 2.4 Hz, 1H), 7.97 (dd, J = 8.9, 2.6 Hz, 1H), 7.86 (d, J = 8.2 Hz, 2H), 7.76 (d, J = 8.4 Hz, 2H), 6.97 (d, J = 8.9 Hz, 1H), 3.58 (dd, J = 6.5, 3.7 Hz, 4H), 3.44 (dd, J = 6.1, 3.7 Hz, 4H), 1.43 (s, 9H).

[0079] 2. tert-Butyl 4-(5-(4-chlorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19c)

[0080]

[0081] The synthesis method referred to Intermediate 19a to obtain Intermediate 19c, a white solid, yield 83%.

[0082] 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 8.47 (d, J = 2.4 Hz, 1H), 7.89 (dd, J = 8.9, 2.6 Hz, 1H), 7.66 (d, J = 8.6 Hz, 2H), 7.47 (d, J = 8.6 Hz, 2H), 6.93 (d, J = 8.9 Hz, 1H), 3.55 (dd, J = 6.4, 3.4 Hz, 4H), 3.45 - 3.42 (m, 4H), 1.43 (s, 9H).

[0083] 3. tert-Butyl 4-(5-(3,4-dichlorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19d)

[0084]

[0085] The synthesis method refers to Intermediate 19a to obtain Intermediate 19d, which is a white solid with a yield of 73%.

[0086] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 8.52 (d, J = 2.1 Hz, 1H), 7.99 - 7.88 (m, 2H), 7.66 (s, 2H), 6.94 (d, J = 8.9 Hz, 1H), 3.57 (d, J = 5.2 Hz, 4H), 3.44 (s, 4H), 1.44 (s, 9H).

[0087] 4. tert-Butyl 4-(5-(3-chloro-4-fluorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19e)

[0088]

[0089] The synthesis method refers to Intermediate 19a to obtain Intermediate 19e, which is a white solid with a yield of 70%.

[0090] 1 H NMR (300 MHz, DMSO-d6) δ (ppm): 8.48 (d, J = 2.5 Hz, 1H), 7.91 (dd, J = 8.9, 2.6 Hz, 1H), 7.85 (dd, J = 7.1, 2.3 Hz, 1H), 7.64 (ddd, J = 8.6, 4.7, 2.4 Hz, 1H), 7.46 (t, J = 9.0 Hz, 1H), 6.93 (d, J = 8.9 Hz, 1H), 3.55 (dd, J = 6.5, 3.5 Hz, 4H), 3.44 (d, J = 6.3 Hz, 4H), 1.43 (s, 9H).

[0091] 5. tert-Butyl 4-(5-(4-fluorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19f)

[0092]

[0093] The synthesis method refers to Intermediate 19a to obtain Intermediate 19f, which is a light yellow solid with a yield of 84%.

[0094] 1 H NMR (300 MHz, DMSO-d 6) δ (ppm): 8.44 (d, J = 2.3 Hz, 1H), 7.86 (dd, J = 8.9, 2.5 Hz, 1H), 7.70 - 7.61 (m, 2H), 7.26 (t, J = 8.9 Hz, 2H), 6.93 (d, J = 8.9 Hz, 1H), 3.58 - 3.50 (m, 4H), 3.47 - 3.40 (m, 4H), 1.43 (s, 9H).

[0095] 6. tert-Butyl 4-(5-(3-hydroxyphenyl)pyridin-2-yl)piperazine-1-carboxylate (19 g)

[0096]

[0097] The synthesis method refers to Intermediate 19a to obtain Intermediate 19g, a white solid, with a yield of 73%.

[0098] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 9.51 (s, 1H), 8.39 (d, J = 2.4 Hz, 1H), 7.80 (dd, J = 8.9, 2.5 Hz, 1H), 7.23 (td, J = 7.8, 3.9 Hz, 1H), 7.02 (d, J = 7.8 Hz, 1H), 6.98 - 6.95 (m, 1H), 6.91 (d, J = 8.9 Hz, 1H), 6.71 (dd, J = 8.0, 1.6 Hz, 1H), 3.57 - 3.49 (m, 4H), 3.47 - 3.41 (m, 4H), 1.43 (s, 9H).

[0099] 7. tert-Butyl 4-(5-(4-chloro-2-fluorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19 h)

[0100]

[0101] The synthesis method refers to Intermediate 19a to obtain Intermediate 19h, a white solid, with a yield of 80%.

[0102] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 8.30 (s, 1H), 7.74 (d, J = 8.9 Hz, 1H), 7.57 (d, J = 8.5 Hz, 1H), 7.53 - 7.47 (m, 1H), 7.35 (dd, J = 8.3, 1.7 Hz, 1H), 6.94 (d, J = 8.9 Hz, 1H), 3.55 (dd, J = 6.0, 3.5 Hz, 4H), 3.43 (d, J = 5.1 Hz, 4H), 1.42 (s, 9H).

[0103] 8. tert-Butyl 4-(5-(3,5-difluorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19j)

[0104]

[0105] The synthesis method refers to Intermediate 19a to obtain Intermediate 19j, which is a white solid with a yield of 60%.

[0106] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 8.57 - 8.54 (m, 1H), 7.97 (dd, J = 9.0, 2.6 Hz, 1H), 7.47 - 7.39 (m, 2H), 7.13 (tt, J = 9.4, 2.2 Hz, 1H), 6.93 (d, J = 9.0 Hz, 1H), 3.59 - 3.55 (m, 4H), 3.47 - 3.41 (m, 4H), 1.43 (s, 9H).

[0107] 9. tert-Butyl 4-(5-(3,5-dichlorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19k)

[0108]

[0109] The synthesis method refers to Intermediate 19a to obtain Intermediate 19k, which is a white solid with a yield of 52%.

[0110] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 8.25 (d, J = 2.4 Hz, 1H), 7.78 (d, J = 2.0 Hz, 1H), 7.73 (dd, J = 8.8, 2.5 Hz, 1H), 7.56 (dd, J = 8.3, 2.1 Hz, 1H), 7.50 (d, J = 8.3 Hz, 1H), 6.99 (d, J = 8.9 Hz, 1H), 3.62 (dd, J = 6.6, 3.5 Hz, 4H), 3.50 (dd, J = 6.1, 3.4 Hz, 4H), 1.49 (s, 9H).

[0111] 10. tert-Butyl 4-(5-(3-fluoro-5-chlorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19l)

[0112]

[0113] The synthesis method refers to Intermediate 19a to obtain Intermediate 19l, which is a white solid with a yield of 52%. 11H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.56 (d, J = 2.6 Hz, 1H), 8.04 - 7.91 (m, 2H), 7.76 (d, J = 8.0 Hz, 1H), 7.64 - 7.47 (m, 2H), 7.38 - 7.30 (m, 2H), 7.24 (dd, J = 7.9, 1.0 Hz, 1H), 6.95 (d, J = 9.0 Hz, 1H), 6.58 (dd, J = 9.6, 1.6 Hz, 1H), 3.73 (s, 4H), 3.62 (s, 2H), 3.47 (s, 2H).

[0114] 11. tert-Butyl 4-(5-(4-chlorophenyl)pyridin-2-yl)piperazine-1-carboxylate (19n)

[0115]

[0116] The synthetic method refers to Intermediate 19a to obtain Intermediate 19n, a white solid, with a yield of 83%.

[0117] 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.52 (d, J = 2.4 Hz, 1H), 8.00 - 7.92 (m, 2H), 7.76 (d, J = 8.0 Hz, 1H), 7.71 (t, J = 1.8 Hz, 1H), 7.66 - 7.58 (m, 1H), 7.45 (t, J = 7.8 Hz, 1H), 7.39 - 7.33 (m, 2H), 7.24 (dd, J = 8.0, 1.4 Hz, 1H), 6.96 (d, J = 9.0 Hz, 1H), 6.58 (dd, J = 9.6, 1.7 Hz, 1H), 3.73 (d, J = 7.5 Hz, 4H), 3.59 (s, 2H), 3.46 (s, 2H).

[0118] 12. tert-Butyl 4-(5-(4-nitrophenyl)pyridin-2-yl)piperazine-1-carboxylate (19o)

[0119] The synthetic method refers to Intermediate 19a to obtain Intermediate 19o, a white solid, with a yield of 76%.

[0120] 11H NMR (300 MHz, DMSO-d6) δ (ppm): 8.65 - 8.60 (m, 1H), 8.29 - 8.22 (m, 2H), 8.03 (dd, J = 9.0, 2.6 Hz, 1H), 7.97 - 7.90 (m, 2H), 6.98 (d, J = 9.0 Hz, 1H), 3.61 (dd, J = 6.5, 3.8 Hz, 4H), 3.44 (dd, J = 6.2, 3.9 Hz, 4H), 1.43 (s, 9H).

[0121] 13. tert-Butyl 4-(5-(3-nitrophenyl)pyridin-2-yl)piperazine-1-carboxylate (19p)

[0122] The synthetic method refers to Intermediate 19a to obtain Intermediate 19p, a white solid, with a yield of 70%.

[0123] 1 1H NMR (300 MHz, Chloroform-d) δ (ppm): 8.48 (d, J = 2.3 Hz, 1H), 8.37 (t, J = 1.9 Hz, 1H), 8.19 - 8.11 (m, 1H), 7.88 - 7.82 (m, 1H), 7.80 - 7.74 (m, 1H), 7.59 (t, J = 8.0 Hz, 1H), 6.75 (d, J = 8.9 Hz, 1H), 3.66 - 3.60 (m, 4H), 3.60 - 3.54 (m, 4H), 1.50 (s, 9H).

[0124] 14. tert-Butyl 4-(5-(4-methoxyphenyl)pyridin-2-yl)piperazine-1-carboxylate (19q)

[0125] The synthetic method refers to Intermediate 19a to obtain Intermediate 19q, a white solid, with a yield of 60%.

[0126] 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 8.41 (d, J = 2.4 Hz, 1H), 7.82 (dd, J = 8.8, 2.6 Hz, 1H), 7.59 - 7.50 (m, 2H), 7.04 - 6.96 (m, 2H), 6.91 (d, J = 8.9 Hz, 1H), 3.78 (s, 3H), 3.51 (dd, J = 6.7, 3.2 Hz, 4H), 3.47 - 3.39 (m, 4H), 1.43 (s, 9H).

[0127] 15. tert-Butyl 4-(5-phenylpyridin-2-yl)piperazine-1-carboxylate (19r)

[0128] The synthetic method refers to Intermediate 19a to obtain Intermediate 19r, a white solid, with a yield of 83%.

[0129] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 8.46 (d, J = 2.4 Hz, 1H), 7.88 (dd, J = 8.9, 2.6 Hz, 1H), 7.66 - 7.58 (m, 2H), 7.43 (t, J = 7.6 Hz, 2H), 7.34 - 7.27 (m, 1H), 6.94 (d, J = 8.9 Hz, 1H), 3.54 (dd, J = 6.6, 3.3 Hz, 4H), 3.44 (dd, J = 6.0, 2.7 Hz, 4H), 1.43 (s, 9H).

[0130] Example 2 7-(4-(5-(4-(Trifluoromethyl)phenyl)pyridin-2-yl)piperazin-1-carbonyl)quinolin-2(1H)-one (IV-21)

[0131]

[0132] Dissolve 2-oxo-1,2-dihydroquinoline-7-carboxylic acid (38 mg) in 2 ml of DMF, add HATU (91 mg) and DIPEA (104 μL). After 10 min, add Intermediate 20b (82 mg, 20b is prepared from 19b by deprotection, and the deprotection can be carried out according to the conventional method), and stir at room temperature for 1 h. After monitoring the reaction to completion by TLC, stop the reaction, dilute with water, extract three times with ethyl acetate, wash three times with saturated brine, and dry the organic phase with anhydrous sodium sulfate. Purify by column chromatography (methylene chloride:methanol = 30:1) to obtain the target product IV-21, 34 mg of white solid, with a yield of 31%.

[0133] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.57 (d, J = 2.3 Hz, 1H), 7.98 (t, J = 8.1 Hz, 2H), 7.87 (d, J = 8.2 Hz, 2H), 7.76 (dd, J = 8.2, 3.0 Hz, 3H), 7.35 (s, 1H), 7.25 (d, J = 8.0 Hz, 1H), 6.99 (d, J = 8.9 Hz, 1H), 6.58 (d, J = 9.5 Hz, 1H), 3.74 (d, J = 6.6 Hz, 4H), 3.55 (d, J = 41.5 Hz, 4H).

[0134] Example 3 7-(4-(5-(4-Chlorophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-23)

[0135]

[0136] The synthesis method refers to compound IV-21. Intermediate 20c (20c is prepared by deprotection from 19c, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-23, which is a white solid with a yield of 35%.

[0137] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.49 (d, J = 2.4 Hz, 1H), 7.96 (d, J = 9.6 Hz, 1H), 7.91 (dd, J = 8.9, 2.5 Hz, 1H), 7.75 (d, J = 8.0 Hz, 1H), 7.67 (d, J = 8.6 Hz, 2H), 7.48 (d, J = 8.6 Hz, 2H), 7.35 (s, 1H), 7.24 (dd, J = 8.0, 1.2 Hz, 1H), 6.96 (d, J = 8.9 Hz, 1H), 6.58 (dd, J = 9.6, 1.3 Hz, 1H), 3.72 (d, J = 17.5 Hz, 4H), 3.53 (d, J = 35.5 Hz, 4H).

[0138] Example 4 7-(4-(5-(3,4-Dichlorophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-22)

[0139]

[0140] The synthesis method refers to compound IV-21. Intermediate 20d (20d is prepared by deprotection from 19d, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-22, which is a white solid with a yield of 20%.

[0141] 1 H NMR (300 MHz, DMSO-d 6) δ (ppm): 11.90 (s, 1H), 8.53 (d, J = 2.4 Hz, 1H), 7.98 (s, 1H), 7.96 - 7.90 (m, 2H), 7.76 (d, J = 8.0 Hz, 1H), 7.66 (t, J = 8.7 Hz, 2H), 7.35 (s, 1H), 7.24 (dd, J = 8.0, 1.3 Hz, 1H), 6.96 (d, J = 9.0 Hz, 1H), 6.58 (d, J = 10.0 Hz, 1H), 3.73 (d, J = 7.7 Hz, 4H), 3.65 - 3.41 (m, 4H).

[0142] Example 5 7-(4-(5-(3-Chloro-4-fluorophenyl)pyridin-2-yl)piperazin-1-carbonyl)quinolin-2(1H)-one (IV-27)

[0143]

[0144] The synthesis method refers to Compound IV-21. Intermediate 20e (20e is prepared by deprotection from 19e, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-27, a white solid, with a yield of 58%.

[0145] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.50 (d, J = 2.5 Hz, 1H), 8.00 - 7.91 (m, 2H), 7.87 (dd, J = 7.1, 2.3 Hz, 1H), 7.76 (d, J = 8.0 Hz, 1H), 7.66 (ddd, J = 8.7, 4.6, 2.3 Hz, 1H), 7.47 (t, J = 9.0 Hz, 1H), 7.35 (s, 1H), 7.24 (dd, J = 8.0, 1.4 Hz, 1H), 6.95 (d, J = 9.0 Hz, 1H), 6.58 (dd, J = 9.5, 1.7 Hz, 1H), 3.72 (d, J = 20.1 Hz, 4H), 3.53 (d, J = 39.9 Hz, 4H).

[0146] Example 6 7-(4-(5-(4-Fluorophenyl)pyridin-2-yl)piperazin-1-carbonyl)quinolin-2(1H)-one (IV-24)

[0147]

[0148] The synthesis method refers to Compound IV-21. Intermediate 20f (20f is prepared by deprotection from 19f, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-24, a yellowish-brown solid, with a yield of 41%.

[0149] 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.89 (s, 1H), 8.45 (d, J = 2.4 Hz, 1H), 7.96 (d, J = 9.6 Hz, 1H), 7.88 (dd, J = 8.9, 2.6 Hz, 1H), 7.75 (d, J = 8.0 Hz, 1H), 7.69 - 7.63 (m, 2H), 7.34 (s, 1H), 7.30 - 7.21 (m, 3H), 6.95 (d, J = 8.9 Hz, 1H), 6.57 (dd, J = 9.5, 1.7 Hz, 1H), 3.71 (d, J = 21.9 Hz, 4H), 3.52 (d, J = 30.8 Hz, 4H).

[0150] Example 7 7-(4-(5-(3-Hydroxyphenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-25)

[0151]

[0152] The synthesis method refers to Compound IV-21. Add 20 g of the intermediate (20 g is prepared by deprotection from 19 g, and the deprotection can be carried out according to the conventional method) to obtain the target product IV-25, which is an off-white solid with a yield of 36%.

[0153] 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 9.51 (s, 1H), 8.40 (d, J = 2.3 Hz, 1H), 7.96 (d, J = 9.6 Hz, 1H), 7.82 (dd, J = 8.9, 2.5 Hz, 1H), 7.76 (d, J = 8.0 Hz, 1H), 7.34 (s, 1H), 7.27 - 7.17 (m, 2H), 7.03 (d, J = 7.9 Hz, 1H), 6.99 - 6.96 (m, 1H), 6.94 (d, J = 9.0 Hz, 1H), 6.71 (dd, J = 8.0, 1.6 Hz, 1H), 6.58 (dd, J = 9.5, 1.5 Hz, 1H), 3.71 (d, J = 26.3 Hz, 4H), 3.52 (d, J = 31.3 Hz, 4H).

[0154] Example 8 7-(4-(5-(4-Chloro-2-fluorophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-26)

[0155]

[0156] The synthesis method refers to Compound IV-21. Intermediate 20h (20h is prepared by deprotection of 19h, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-26, which is a white solid with a yield of 33%.

[0157] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.33 (s, 1H), 7.96 (d, J = 9.6 Hz, 1H), 7.77 (t, J = 7.4 Hz, 2H), 7.62 - 7.49 (m, 2H), 7.37 (d, J = 10.7 Hz, 2H), 7.24 (d, J = 8.0 Hz, 1H), 6.97 (d, J = 9.0 Hz, 1H), 6.58 (d, J = 8.8 Hz, 1H), 3.73 (d, J = 11.9 Hz, 4H), 3.53 (d, J = 40.8 Hz, 4H).

[0158] Example 9 7-(4-(5-(3,5-Difluorophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-14)

[0159]

[0160] The synthesis method refers to Compound IV-21. Intermediate 20j (20j is prepared by deprotection of 19j, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-14, which is an off-white solid with a yield of 20%.

[0161] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.57 (d, J = 2.5 Hz, 1H), 8.03 - 7.92 (m, 2H), 7.76 (d, J = 8.0 Hz, 1H), 7.49 - 7.39 (m, 2H), 7.35 (s, 1H), 7.24 (dd, J = 8.0, 1.4 Hz, 1H), 7.14 (tt, J = 9.3, 2.3 Hz, 1H), 6.95 (d, J = 9.0 Hz, 1H), 6.58 (dd, J = 9.6, 1.6 Hz, 1H), 3.73 (s, 4H), 3.54 (d, J = 44.8 Hz, 4H).

[0162] Example 10 7-(4-(5-(3,5-Dichlorophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-15)

[0163]

[0164] The synthesis method refers to Compound IV-21. Intermediate 20k (20k is prepared by deprotection from 19k, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-15, a beige solid, with a yield of 35%.

[0165] 1 H NMR(300MHz,DMSO-d 6 )δ(ppm):11.90(s,1H),8.20(d,J=2.3Hz,1H),7.97(d,J=9.6Hz,1H),7.76(d,J=8.1Hz,1H),7.73(d,J=2.0Hz,1H),7.69(dd,J=8.9,2.5Hz,1H),7.50(dd,J=8.3,2.0Hz,1H),7.45(d,J=8.3Hz,1H),7.35(s,1H),7.24(dd,J=8.0,1.4Hz,1H),6.95(d,J=8.9Hz,1H),6.58(dd,J=9.6,1.7Hz,1H),3.73(d,J=13.2Hz,4H),3.63-3.43(m,4H).

[0166] Example 11 7-(4-(5-(3-chloro-5-fluorophenyl)pyridin-2-yl)piperazin-1-carbonyl)quinolin-2(1H)-one (IV-16)

[0167]

[0168] The synthesis method refers to Compound IV-21. Intermediate 20l (20l is prepared by deprotection from 19l, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-16, a beige solid, with a yield of 30%.

[0169] 1 H NMR(300MHz,DMSO-d 6 )δ(ppm):11.90(s,1H),8.56(d,J=2.6Hz,1H),8.04-7.91(m,2H),7.76(d,J=8.0Hz,1H),7.64-7.47(m,2H),7.38-7.30(m,2H),7.24(dd,J=7.9,1.0Hz,1H),6.95(d,J=9.0Hz,1H),6.58(dd,J=9.6,1.6Hz,1H),3.73(s,4H),3.62(s,2H),3.47(s,2H).

[0170] Example 12 7-(4-(5-(3-chlorophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-19)

[0171]

[0172] The synthesis method refers to Compound IV-21. Intermediate 200n (20n is prepared by deprotection from 19n, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-19, which is a light beige solid with a yield of 38%.

[0173] 1 H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.52 (d, J = 2.4 Hz, 1H), 8.00 - 7.92 (m, 2H), 7.76 (d, J = 8.0 Hz, 1H), 7.71 (t, J = 1.8 Hz, 1H), 7.66 - 7.58 (m, 1H), 7.45 (t, J = 7.8 Hz, 1H), 7.39 - 7.33 (m, 2H), 7.24 (dd, J = 8.0, 1.4 Hz, 1H), 6.96 (d, J = 9.0 Hz, 1H), 6.58 (dd, J = 9.6, 1.7 Hz, 1H), 3.73 (d, J = 7.5 Hz, 4H), 3.59 (s, 2H), 3.46 (s, 2H).

[0174] Example 13 7-(4-(4-(4-(4-nitrophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-41)

[0175] The synthesis method refers to Compound IV-21. Intermediate 20o (20o is prepared by deprotection from 19o, and the deprotection can be carried out according to the conventional method) is added to obtain the target product IV-41, which is a yellow solid with a yield of 46%.

[0176] 1 H NMR (400 MHz, DMSO-d 6) δ (ppm): 11.88 (s, 1H), 8.64 (d, J = 2.5 Hz, 1H), 8.29 - 8.23 (m, 2H), 8.05 (dd, J = 9.0, 2.6 Hz, 1H), 7.95 (ddd, J = 9.5, 5.1, 2.7 Hz, 3H), 7.75 (d, J = 8.0 Hz, 1H), 7.35 (d, J = 1.2 Hz, 1H), 7.24 (dd, J = 8.0, 1.5 Hz, 1H), 7.00 (d, J = 9.0 Hz, 1H), 6.58 (dd, J = 9.6, 1.8 Hz, 1H), 3.75 (s, 4H), 3.56 (d, J = 70.6 Hz, 4H).

[0177] Example 14 7-(4-(4-(4-(3-Nitrophenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-42)

[0178] The synthesis method refers to Compound IV-21, adding Intermediate 20p (20p is prepared by deprotection from 19p, and the deprotection can be carried out according to the conventional method), to obtain the target product IV-42, a yellow solid, with a yield of 40%.

[0179] 1 H NMR (300 MHz, Chloroform-d) δ (ppm): 12.04 (s, 1H), 8.49 (d, J = 2.4 Hz, 1H), 8.38 (t, J = 1.9 Hz, 1H), 8.17 (ddd, J = 8.2, 2.1, 0.9 Hz, 1H), 7.88 - 7.77 (m, 3H), 7.65 (d, J = 8.1 Hz, 1H), 7.60 (t, J = 8.0 Hz, 1H), 7.52 (s, 1H), 7.30 (d, J = 1.3 Hz, 1H), 6.79 (d, J = 5.7 Hz, 1H), 6.76 (d, J = 6.4 Hz, 1H), 4.02 - 3.73 (m, 4H), 3.72 - 3.52 (m, 4H).

[0180] Example 15 7-(4-(4-(4-(4-Methoxyphenyl)pyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-43)

[0181] The synthesis method refers to Compound IV-21, adding Intermediate 20q (20q is prepared by deprotection from 19q, and the deprotection can be carried out according to the conventional method), to obtain the target product IV-43, an off-white solid, with a yield of 42%.

[0182] 11H NMR (300 MHz, DMSO-d6) δ (ppm): 11.89 (s, 1H), 8.42 (d, J = 2.4 Hz, 1H), 7.96 (d, J = 9.6 Hz, 1H), 7.84 (dd, J = 8.9, 2.5 Hz, 1H), 7.75 (d, J = 8.0 Hz, 1H), 7.55 (d, J = 8.8 Hz, 2H), 7.35 (s, 1H), 7.24 (dd, J = 8.0, 1.3 Hz, 1H), 7.00 (d, J = 8.8 Hz, 2H), 6.93 (d, J = 8.9 Hz, 1H), 6.58 (dd, J = 9.6, 1.4 Hz, 1H), 3.78 (s, 3H), 3.77 - 3.39 (m, 8H).

[0183] Example 16 7-(4-(4-(4-Phenylpyridin-2-yl)piperazine-1-carbonyl)quinolin-2(1H)-one (IV-44)

[0184] The synthetic method refers to Compound IV-21, adding Intermediate 20r (20r is prepared from 19r by deprotection, and the deprotection can be carried out according to the conventional method), to obtain the target product IV-44, off-white solid, with a yield of 46%.

[0185] 1 1H NMR (300 MHz, DMSO-d 6 ) δ (ppm): 11.90 (s, 1H), 8.54 - 8.43 (m, 1H), 7.96 (d, J = 9.5 Hz, 1H), 7.93 - 7.86 (m, 1H), 7.76 (d, J = 8.0 Hz, 1H), 7.63 (d, J = 7.5 Hz, 2H), 7.44 (t, J = 7.4 Hz, 2H), 7.38 - 7.19 (m, 3H), 6.96 (d, J = 8.8 Hz, 1H), 6.58 (d, J = 9.5 Hz, 1H), 3.73 (d, J = 25.4 Hz, 4H), 3.54 (d, J = 35.4 Hz, 4H).

[0186] Biological activity test of the compound

[0187] Example 17 Determination of MAGL inhibitory activity

[0188] 1. Test principle: The MAGL inhibitor screening kit from Cayman provides a simple and efficient method for screening the inhibitory activity of compounds against human MAGL. The principle is that MAGL hydrolyzes an alternative substrate (ethyl 4-nitrophenylacetate instead of 2-AG) to produce a yellow product (4-nitrophenol), which has a maximum absorption at 405 - 412 nm. By measuring the absorbance at this wavelength, the content of the hydrolyzed product 4-nitrophenol is quantified to characterize the enzyme activity.

[0189] 2. Preparation of reagents

[0190] (1) Preparation of buffer: Add 3 mL of concentrated buffer (10×) to 27 mL of pure water for dilution. The diluted buffer (1×) contains 10 mM Tris-HCl, pH 7.2, and 1 mM EDTA, and is used for analyzing and diluting MAGL and the positive drug JZL195. Store it at -20 °C for later use.

[0191] (2) Preparation of human recombinant MAGL: Take 30 μL of MAGL protein and add 570 μL of buffer (1×), then store it for later use.

[0192] (3) Preparation of MAGL substrate: Take 150 μL of substrate and add 450 μL of buffer (1×), then store it for later use.

[0193] (4) Preparation of positive drug and compounds: JZL195 is dissolved in DMSO and buffer in a 1:1 ratio to prepare compound solutions and JZL195 solutions with concentrations of 0.001 μM, 0.01 μM, 0.1 μM, 1 μM, 10 μM, and 100 μM respectively, and store them for later use.

[0194] 3. Experimental operation

[0195] (1) Background wells: Add 160 μL of buffer (1×) and 10 μL of solvent

[0196] (2) 100% initial enzyme activity wells: Add 150 μL of buffer (1×), 10 μL of enzyme, and 10 μL of solvent

[0197] (3) Positive control and drug wells: Add 150 μL of buffer (1×), 10 μL of enzyme, and 10 μL of positive drug or inhibitor. Repeat the experiment 3 times for each of the above wells, as shown in Table 1 specifically.

[0198] Table 1

[0199] Buffer (1×) MAGL Solvent Inhibitor Background 160 μL 10 μL 100% Initial Enzyme Activity 150 μL 10 μL 10 μL Inhibitor 150 μL 10 μL 10 μL

[0200] (4) Mix the substances in the wells and incubate at room temperature for 15 min.

[0201] (5) Add 10 μL of substrate into each well, shake the 96-well plate thoroughly for 10 s, and incubate at room temperature for 10 min. Read the absorbance at 405 nm using a Multiscan GO (Thermo) microplate reader. Calculate the inhibition rate according to the following formula, and then perform nonlinear regression analysis using GraphPad Prism software, using second-order polynomial regression analysis and analyzing the experimental data by applying a mixed-model inhibition fit.

[0202]

[0203] The results are shown in Table 2. It can be seen from Table 2 that the compounds of the present invention have MAGL inhibitory activity.

[0204] Example % Inhibition (10 μM) Example % Inhibition (10 μM) IV-14 90 IV-24 99 IV-15 96 IV-25 85 IV-16 84 IV-26 99 IV-19 83 IV-27 92 IV-20 85 IV-41 92 IV-21 91 IV-42 97 IV-22 92 IV-43 99 IV-23 89 IV-44 99

[0205] Refer to a method similar to this example to determine the IC 50 of the compounds of this application, and the specific method is as follows:

[0206] According to the above method, determine the inhibition rates of the applied compounds at concentrations of 0.001 μM, 0.01 μM, 0.1 μM, 1 μM, 10 μM, and 100 μM. Use second-order polynomial regression analysis and analyze the experimental data by applying a mixed-model inhibition fit to obtain the IC 50 data of the compounds against MAGL inhibition.

[0207] Example 18 Determination of 2-AG Concentration in the Brain

[0208] Use ICR male mice (3 mice / group). Dissolve the test compound IV-23 in 10% DMSO, 10% Tween 80, and 80% physiological saline to prepare a dosing solution. The dose of the test compound is prepared as 10 mg / kg, and the dosing volume is 10 ml / kg. Administer the test compound by gavage. After administering the test compound, separate the brains at 60 min, 120 min, 240 min, and 480 min respectively, and extract the cerebral hemispheres (control separated at 0 min). Freeze the obtained cerebral hemispheres on dry ice, measure the weight of the frozen tissue, weigh about 100 mg of brain tissue, and add 9 times the volume of physiological saline to homogenize the brain tissue. After homogenization, centrifuge the mixture at 15000 rpm for 10 minutes. Aliquot the supernatant, about 50 μL per tube. Determine the 2-AG concentration in the brain using the Elisa method.

[0209] The results of the 2-AG concentration in the brain are shown in Table 3, with the unit ng / g.

[0210] Table 3

[0211]

[0212] As can be seen from Table 3, compound IV-23 was transferred to the brain by oral administration to ICR male mice. Compared with the control group, these compounds significantly increased the concentration of 2-AG in the brain, and showed significant differences at 60min, 120min, and 240min (p<0.05), indicating that IV-23 acts on the MAGL target, causing an increase in the level of 2-AG in the brain.

[0213] Example 19 Pharmacokinetics Test

[0214] 1. Experimental Materials

[0215] (1) Mobile phase solution: Take 500 mL of Wahaha water, add 2.5 mL of formic acid to it to prepare an aqueous solution containing 0.5% formic acid, mix well, and degas by ultrasonication for 20 min.

[0216] (2) Preparation of stock solution: Accurately weigh 2.0 mg of compound IV-23 and dissolve it in 10 mL of water to make a stock solution of 200.0 ug / mL. Accurately weigh 2.0 mg of imipramine hydrochloride and dissolve it in 10 mL of water to make a stock solution of 200.0 ug / mL. Add methanol to dilute to 3.000 ug / mL as the internal standard working solution and store in a refrigerator at 4°C for future use.

[0217] (3) Preparation of standard curve samples: Dilute the IV-23 stock solution with methanol to a series of standard solutions of 30000, 15000, 7500, 3000, 1500, 300.0, and 30.00 ng / mL. Accurately pipette 10uL of the above series of standard solutions, add 50ul of blank SD rat plasma, add 10μL of the internal standard working solution, mix well, and add 230μL of methanol. The final volume is 300μL, and the final concentration of compound IV-23 is 1000, 500.0, 250.0, 100.0, 50.00, 10.00, and 1.000 ng / mL. After vortex mixing for 5 minutes, centrifuge at 12000r / min for 10 minutes, and transfer the supernatant to an autosampler vial for LC-MS / MS analysis to prepare for the drawing of the standard curve.

[0218] (4) HPLC conditions: The chromatographic column was Waters 5C18-MS-Ⅱ (2.0×250 mm, 5 μm); the mobile phase was 0.5% formic acid water (A)-methanol (B); isocratic elution was adopted, the organic phase ratio was 90%, the flow rate was 0.3 mL / min, the column temperature was 40°C, and the injection volume was 10 μL.

[0219] Preparation of the test article: Weigh 0.5 mg of the test article IV-23 and place it in a 10 mL EP tube. Add 0.125 mL of DMSO and 0.25 mL of Tween-80, then add 4.2 mL of normal saline injection. After dissolution, sonicate and shake to mix evenly until the compound is clear, and prepare a 0.1 mg / mL preparation; prepare and use it on the same day. Weigh 5 mg of the test article IV-23 and place it in a 10 mL EP tube. Add 0.125 mL of DMSO and 0.25 mL of Tween-80, then add 4.625 mL of normal saline injection. After dissolution, sonicate and shake to mix evenly until the compound is clear, and prepare a 1 mg / mL PO preparation and a 0.2 mg / mL IV preparation. The administration volume is 10 mL / kg, and the administration doses are 10 mg / kg and 2 mg / kg respectively; prepare and use it on the same day.

[0220] The pharmacokinetic parameters of IV-23 are shown in Table 4:

[0221]

[0222] a T max , time to peak concentration; b C max , maximum plasma concentration; c AUC, area under the plasma concentration-time curve; d Vd, apparent volume of distribution; e t 1 / 2 , drug half-life; f CL, clearance rate; g MRT, mean residence time.

[0223] As can be seen from Table 4, the compound IV-23 has a high plasma exposure and a long half-life under different administration methods of intravenous injection and gavage, indicating that the compound IV-23 has good pharmacokinetic properties.

[0224] Example 20 Determination of the antidepressant effect of the compound on depressive mice

[0225] Use ICR male mice. After 1 week of adaptation, randomly divide 12 mice as blank controls and raise them normally; put the remaining 70 mice into 50 mL centrifuge tubes (with holes at the end of the tubes for mice to breathe), and bind them for 4 - 8 h every day, increasing from 4 h per day to 8.5 h per day, and bind them continuously for 30 days.

[0226] Model determination:

[0227] The tail suspension test was used to determine whether the mice were modeled (evaluating the despair behavior of the mice): The posterior 1 / 3 of the mouse's tail was fixed with tape and suspended on a bracket, with the head 15 cm from the ground. After the mice adapted for 2 min, they were photographed, and the immobile time of the mice within 4 min was counted.

[0228] Grouping and drug administration:

[0229] After modeling, according to the immobile time data in the tail suspension test, the mice were divided into a model group, a positive drug group (fluoxetine, 8 mg / kg), a compound IV-23 low-dose group (4 mg / kg), and a compound IV-23 high-dose group (8 mg / kg), with 6 - 8 mice in each group. Compound preparation: 10% DMSO, 40% PEG 400, 5% Tween 80, 45% normal saline. Administration volume: 10 ml / kg. The drug was administered continuously for 7 days. After 7 days, behavioral tests such as the tail suspension test, open field test, forced swimming test, and sucrose preference test were conducted.

[0230] The results are shown in Table 5. As can be seen from Table 5, ICR depressive mice can significantly improve the depressive-like behavior caused by chronic restraint after intraperitoneal injection of compound IV-23 for 7 consecutive days.

[0231] Table 5

[0232]

[0233] Example 21 Determination of the in vitro anti-nonalcoholic fatty liver effect of the compound

[0234] 1. Instruments and materials

[0235] (1) Cells and reagents: The human hepatocellular carcinoma cell line HepG2 cells were purchased from the Shanghai Institute of Cell Biology, Chinese Academy of Sciences; fetal bovine serum was purchased from Solarbio Science & Technology Co., Ltd.; DMEM high-glucose medium, 0.25% trypsin solution, and PBS solution were purchased from Nanjing KeyGen Biotech Co., Ltd.; DMSO, oleic acid, palmitic acid, and oil red O were purchased from Guangdong Xilong Chemical Co., Ltd.

[0236] (2) Main instruments: American MULTISKAN MK3 fully automatic microplate reader (Thermo Scientific), vortex oscillator (QL-902); centrifuge, BT224 electronic balance (Sartouris, Germany); HB-202 constant temperature water bath (Beijing Zhongxi Yuanda); BX51 upright microscope (Olympus, Japan); other equipment and instruments including centrifuge tubes, pipettes, pipettes, etc.

[0237] 2. Experimental method:

[0238] (1) Cell culture method

[0239] HepG2 cells were inoculated into high-glucose DMEM culture medium containing 10% fetal bovine serum to allow the cells to adhere and grow. They were cultured in an incubator at 37°C with 5% CO 2 and saturated humidity. According to the cell growth situation, they were digested with 0.25% trypsin every 1 - 2 days for subculture.

[0240] (2) Effects of Compound IV-23 on the NASH cell model induced by free fatty acids in HepG2 cells

[0241] HepG2 cells were cultured in 96-well plates. After the cells adhered and grew to 60 - 70%, they were co-stimulated with free fatty acids (at a molar ratio of palmitic acid:oleic acid of 1:2) and different concentrations of the test drug (Compound IV-23) for 24 h. Three parallel wells were set up and divided into a blank group, a model group, and a drug administration group. The blank group was replaced with blank DMEM, and the drug administration group was respectively added with Compound IV-23 at concentrations of 5 μM and 10 μM prepared with DMEM.

[0242] (3) Observation of intracellular lipid droplets by Oil Red O staining

[0243] The steps of Oil Red O staining were as follows: The cells were washed 3 times with PBS solution, fixed with 10% neutral formaldehyde for 30 min, washed 2 times with PBS solution, stained with Oil Red O at 37°C, and incubated at 37°C for 1 h. Then, the staining solution was discarded and the cells were washed twice with PBS. Under the microscope, the neutral fat in the cells could be specifically stained red to observe the lipid accumulation in the cells.

[0244] 3. Experimental results: Observation of the effects of Compound IV-23 on the NASH cell model by Oil Red O staining

[0245] The Oil Red O staining results of Compound IV-23 were as Figure 1 shown. It can be seen that compared with untreated cells, HepG2 cells exposed to free fatty acids (model group) showed higher intracellular lipid accumulation. After intervention with Compound IV-23, the lipid droplet aggregation in HepG2 cells was significantly inhibited; indicating that Compound IV-23 prepared by the present invention can significantly inhibit lipid accumulation induced by free fatty acids in a dose-dependent manner at 5 μM and 10 μM, and has the effect of treating non-alcoholic fatty liver disease.

[0246] Example 22 Determination of the in vivo anti-Parkinson's disease effect of the compound

[0247] 1. Experimental method: Mice were acclimated for 1 week. Five animals were randomly selected from the normal group without any treatment and were normally fed. For the remaining four groups of mice, on the first day, they were intraperitoneally injected with the solvent or the drug once. On the second day, they were intraperitoneally injected with MPTP (15 mg / kg, injected once every 2 h, for a total of 4 injections). One hour after the second injection of MPTP, the solvent or the drug was injected. On the third day, they were intraperitoneally injected with the solvent or the drug once. Behavioral tests were performed 2 h after drug administration.

[0248] 2. Behavioral experiment - Rotarod test: The instrument was placed 50 cm above the ground. There was a digital timer at the bottom of each partition, and the time was recorded when the animal fell. Mice were trained on the rotarod at a speed of 20 rpm for a fixed time. A rotator was used to evaluate the grip strength of the animals. Mice were trained continuously for 2 days before drug administration. During the behavioral test, a rotarod with a diameter of 7 cm was used to test the grip strength of the animals at a speed of 20 rpm, and the cut-off time was 180 s.

[0249] 3. Experimental results: As Figure 2 shown, the average time of the blank mice on the rotarod was 59.26 s, and that of the model group was 24.1 s, showing a significant difference from the blank group, indicating that the model was successful. The average time of the positive drug JZL184 group was 46.75 s, showing a certain therapeutic effect, but there was no significant difference from the model group. The average time of the IV-23 (10 mg / kg) group was 53.82 s, showing a significant difference from the model group, indicating that IV-23 (10 mg / kg) could significantly improve the motor ability of Parkinson's mice.

[0250] Example 23 Determination of the cholestatic liver injury effect of the compound

[0251] 1. Experimental method:

[0252] (1) Eighteen male C57BL / 6J mice were randomly divided into 3 groups (n = 6) according to body weight after one week of adaptive feeding: Control group, model group (0.1% DDC), and IV-26 administration group (12 mg / kg + 0.1% DDC). The control group was given normal feed, and the other groups were given feed containing 0.1% DDC for 2 weeks. One week after feeding with feed containing 0.1% DDC, the mice in the administration group were intraperitoneally injected with IV-26 (12 mg / kg, 10 mL / kg) every day for one week, and the control group and the 0.1% DDC group were injected with the solvent (10 mL / kg) in the same way for 1 week. One hour after the last drug administration of the mice, blood was collected from the orbital cavity, the mice were sacrificed, the livers were dissected, after being washed with normal saline, the surface moisture was blotted dry, and the liver lobules were carefully separated and fixed in formalin fixative, and the remaining part of the liver was stored at -80 °C.

[0253] (2) After the blood sample was left standing at room temperature for 30 min, it was centrifuged at 3000 rpm / min for 10 minutes at 4 °C. The upper serum sample was collected, and the contents of alanine aminotransferase (ALT / GPT) and alkaline phosphatase (ALP) in the serum were detected according to the kit instructions.

[0254] (3) Histopathological examination of mouse liver tissue: The fixed mouse liver tissue was fixed in formalin solution for 24 hours, dehydrated with alcohol, embedded in paraffin, and cut into sections with a thickness of 5 μm. The prepared sections were stained with hematoxylin and eosin dyes respectively. The sections after HE staining were dehydrated, cleared and mounted, and observed under a microscope to record the histopathological morphology of the liver tissue.

[0255] 2. Experimental results:

[0256] (1) Effects of Ⅳ-26 on biochemical indexes in the serum of mice with DDC-induced cholestatic liver injury

[0257] The levels of ALT and ALP in the serum are important indicators for measuring liver function and can directly reflect the liver damage. The experimental results are as Figure 3 shown. Compared with the Control group, the levels of ALT and ALP in the serum of the DDC group increased significantly, indicating that the livers of the mice in the DDC group were severely damaged and the model of cholestatic liver injury was successfully established. Compared with the DDC group, the levels of ALT and ALP in the serum of the Ⅳ-26 administration group decreased. The results showed that the test drug Ⅳ-26 could significantly improve DDC-induced cholestatic liver injury.

[0258] (2) Effects of Ⅳ-26 on histopathological changes in the liver of mice with DDC-induced cholestatic liver injury

[0259] The pathological changes of the mouse liver were observed under a microscope after HE staining. The results are as Figure 4 shown. In the Control group, the liver cells of the mice were arranged intact and closely, and all structures were normal. In the DDC group, there were obvious inflammatory cell infiltrations, hepatocyte necrosis and morphological changes. Compared with the DDC group, the Ⅳ-26 administration group could improve liver injury, reduce inflammatory cell infiltration and alleviate hepatocyte necrosis. The results showed that the test drug Ⅳ-26 could improve the histopathological changes of DDC-induced cholestatic liver injury.

Claims

1. A compound having the structure shown in formula (I), or a pharmaceutically acceptable salt or isotopically substituted derivative thereof: Wherein, R 1 is selected from aryl or heteroaryl; R 2 is selected from substituted or optionally substituted by any number of R 2A substituted aryl or heteroaryl; R 2A Selected from -OH, -SH, -CN, halogen, nitro, carboxyl, C 1-8 alkyl, C 1-8 alkoxy, C 1-4 haloalkyl.

2. The compound according to claim 1, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof, characterized in that, R 1 and R 2 each of the aryl groups is independently C 6 -C 10 aryl group. Preferably, each of the aryl groups is independently phenyl or naphthyl. Specifically preferably, in R 1 and R 2 , the aryl group is phenyl; in R 1 and R 2 , the heteroaryl group is a 5- to 10-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1, 2, 3, or 4"; preferably, the heteroaryl group is a 5- to 10-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1, 2, or 3"; more preferably, the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1, 2, or 3"; further preferably, the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N, O, and S, and the number of heteroatoms is 1 or 2"; more preferably, the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N and S, and the number of heteroatoms is 1 or 2"; even more preferably, the heteroaryl group is a 5- to 6-membered heteroaryl group in which "the heteroatoms are selected from N and S, and the number of heteroatoms is 1"; more preferably, in R 1 and R 2 , the heteroaryl group is furyl, thienyl, pyrrolyl, oxazolyl, isoxazolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, triazolyl, 1,3,4-oxadiazolyl, 1,3,4-thiadiazolyl, 1,2,4-oxadiazolyl, 1,2,4-thiadiazolyl, pyridyl, pyrimidinyl, pyridazinyl, benzofuranyl, indolyl, quinolinyl, isoquinolinyl, indazolyl, benzoxazolyl, benzothiazolyl, purinyl, oxazolopyridyl; specifically preferably, in R 1 and R 2 , the heteroaryl group is pyridyl.

3. The compound according to claim 1, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof, characterized in that, R 1 is a heteroaryl, R 2 is a substituted or unsubstituted aryl with any number of R 2A Preferably, R 1 is a pyridyl group, and R 2 is a substituted or unsubstituted phenyl group with any number of R 2A substituents.

4. The compound according to claim 1, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof, characterized in that, R 2A The number is 1, 2, 3 or 4, preferably 1 or 2, and more preferably, R 2A each independently selected from -OH, -SH, -CN, cyano, halogen, nitro, carboxyl, methyl, ethyl, n-propyl, isopropyl, methoxy, ethoxy, -CF 3 , CHF 2 or CH 2 F.

5. A compound having the following structure, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof:

6. A method for preparing the compound of formula (I) according to claim 1.

7. A pharmaceutical composition, characterized in that, it comprises the compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof, and a pharmaceutically acceptable excipient; preferably, the pharmaceutical composition contains 0.01 - 99.99% of the compound according to any one of claims 1 to 7, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof.

8. Use of the compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof, in the preparation of a MAGL inhibitor.

9. Use of the compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt or isotopically substituted derivative thereof, in the preparation of a drug for preventing and / or treating MAGL-related diseases.

10. The use according to claim 9, characterized in that, the MAGL-related diseases are central nervous system diseases, metabolic disorders and inflammatory diseases; preferably, the MAGL-related diseases are depression, anxiety, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, multiple sclerosis, neuropathic pain, inflammatory pain, cancer pain, epilepsy, cancer, fatty liver, non-alcoholic steatohepatitis, liver fibrosis, cholestasis, inflammatory bowel disease.

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