Somatostatin receptor 5 antagonist, pharmaceutical composition containing the same, and use thereof

Abstract

The present invention relates to a class of somatostatin receptor 5 antagonists, their pharmaceutical compositions and uses. The somatostatin receptor 5 antagonist of the present invention is a compound having the structure shown in the following general formula I. Pharmacological studies have proven that the compounds of the present invention have good SSTR5 antagonistic activity and can be prepared into drugs for treating diseases mediated by SSTR5.

Description

Technical Field

[0001] The present invention belongs to the field of pharmacy, and particularly relates to a class of somatostatin receptor 5 antagonists, pharmaceutical compositions containing the same, and their pharmaceutical uses. Background Art

[0002] Somatostatin Receptor subtype 5 (SSTR5) is an inhibitory G protein-coupled receptor. In rodents, it is mainly distributed in the pituitary gland, gastrointestinal tract, and islets. In humans, it is highly distributed in the gastrointestinal tract (Regulatory peptides, 2000, 90(1-3): 1-18). Its endogenous ligand is somatostatin (SST), which is mainly divided into SST-14 and SST-28. After SST binds to SSTR5, it can activate SSTR5 and mediate the inhibitory hormone secretion effect. Among them, the activation of SSTR5 in the gastrointestinal tract can inhibit the secretion of gastrointestinal hormones such as GLP-1, GLP-2, GIP, PYY, CCK, etc.; the activation of SSTR5 in pancreatic islet tissue can inhibit the secretion of insulin. (Frontiers in Neuroendocrinology, 34(2013)228–252; Am J Physiol Gastrointest Liver Physiol 279:G983–G989, 2000.); Pharmacological studies have shown that SSTR5 antagonists can antagonize the SSTR5 activation effect mediated by the binding of SSTR5 to endogenous ligands, and then promote the secretion of gastrointestinal hormones such as GLP-1, GLP-2, GIP, PYY, CCK, etc. (Diabetologia, 55(2012)3094-3103), as well as promote insulin secretion and have a positive effect on gallbladder motility. At the same time, in the SSTR5 gene knockout mouse model, compared with wild-type mice, its blood glucose handling ability and insulin resistance effect are significantly improved (Molecular Endocrinology 17(1):93–106).GLP-1 has multiple physiological functions, such as promoting glucose-dependent insulin secretion and inhibiting glucagon secretion, promoting satiety, slowing gastric emptying, and playing a protective role in the liver, kidney, and myocardium. Currently, GLP-1-based drugs have been successfully applied in the fields of type 2 diabetes and obesity; at the same time, they have shown positive efficacy in clinical trials of non-alcoholic fatty liver disease, Alzheimer's disease, and Parkinson's disease; GLP-2 can promote small intestine growth and nutrient absorption, which is crucial for maintaining intestinal homeostasis. GLP-2 analogs have been approved for short bowel syndrome and have shown positive efficacy in animal models of inflammatory bowel disease; GIP mainly acts on the islets to regulate glucose homeostasis in a glucose-dependent manner and has a synergistic effect with GLP-1; PYY can slow gastric emptying and promote satiety and is used in the treatment of obesity; CCK can promote gallbladder contraction and the outflow of bile from the gallbladder (Curr Med Chem. 2019; 26(19): 3407–3423.), and the gallbladder emptying function is related to various gallbladder diseases, (GASTROENTEROLOGY 1996; 111: 765–771; Laboratory Investigation (2015) 95, 124–131;) such as gallstones, cholestasis, and primary sclerosing cholangitis. Therefore, antagonizing SSTR5 is a potential therapy for gallstones, cholestasis, and primary sclerosing cholangitis.

[0003] Further studies have shown that there is a significant synergistic effect between SSTR5 antagonists and receptor agonists that promote gastrointestinal hormone secretion (such as TGR5, GPR40, GPR119, GPR41, GPR43 agonists, etc.) and DPP4 inhibitors that inhibit degradation. The combined use of the three can significantly increase the level of gastrointestinal hormones (Diabetes 2018 Feb; 67(2): 309-320). Therefore, STTR5 antagonists can be used in combination with TGR5 agonists, GPR40 full agonists, GPR119 agonists, GPR41 agonists, GPR43 agonists, and DPP4 inhibitors.

[0004] In summary, the development of a novel class of SSTR5 antagonists is expected to be applied in the treatment of chronic metabolic diseases such as type 2 diabetes, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), inflammatory bowel disease, obesity, gallstones, and cholangitis. At the same time, it can be used in combination with TGR5 agonists, GPR40 modulators, GPR119 agonists, GPR41 agonists, GPR43 agonists, and DPP4 inhibitors for diseases related to the GLP-1 and GIP fields, such as type 2 diabetes, obesity, non-alcoholic fatty liver disease, non-alcoholic hepatic fibrosis, Parkinson's disease, and Alzheimer's disease, and has good clinical application prospects. Summary of the Invention

[0005] One technical object of the present invention is to provide a class of compounds having somatostatin receptor 5 antagonistic effects.

[0006] Another technical object of the present invention is to provide a pharmaceutical composition comprising the said compound.

[0007] Another technical object of the present invention is to provide the use of the said compound or the said pharmaceutical composition in the preparation of a somatostatin receptor 5 antagonist.

[0008] In the first aspect of the present application, there is provided a compound having the structure shown in the following general formula I, or its solvate, hydrate, deuterated compound, stereoisomer, tautomer, pharmaceutically acceptable salt:

[0009]

[0010] Wherein, R1, R2, R3, R4, and R5 are each independently selected from the group consisting of: hydrogen, hydroxy, C1-C6 alkyl, C1-C6 alkoxy, C3-C6 cycloalkyl, halogen, C1-C6 haloalkyl, -OH, -NH2, -N(C1-C3 alkyl)(C1-C3 alkyl), -NH(C1-C3 alkyl), substituted or unsubstituted C6-C 14 aryl; wherein the said substitution means that one or more hydrogen atoms on the aryl are substituted by a group selected from the group consisting of: halogen, C1-C3 haloalkoxy, C1-C3 alkyl, C1-C3 alkoxy, C1-C3 haloalkyl, C3-C6 cycloalkyl, -OH, -NH2, -NH(C1-C3 alkyl), -N(C1-C3 alkyl)(C1-C3 alkyl);

[0011] Alternatively, any two adjacent substituents among R1, R2, R3, R4, and R5 together with the benzene ring form a benzo 5-7 membered N, O or S-containing heterocycle or benzo 5-7 membered carbocycle, and the said heterocycle or carbocycle is unsubstituted or substituted by one or more groups selected from the group consisting of: halogen, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 haloalkoxy, C3-C6 cycloalkyl, -OH, -NH2, -N(C1-C6 alkyl)(C1-C6 alkyl);

[0012] A-G is a structure shown in the following formula III or IV:

[0013]

[0014] In formula III, R8 and R9 are each independently hydrogen, halogen, C1-C3 alkyl,

[0015] C1-C3 alkoxy, C1-C3 haloalkoxy, -OH, -NH2, -NH(C1-C3 alkyl), -N(C1-C3 alkyl)(C1-C3 alkyl); preferably, R6, R7, R8, R9 are each independently selected from hydrogen, halogen, and C1-C3 alkoxy;

[0016] X and Y are each independently CH or N;

[0017] G1 has the following structure from the left side (A) to the right side (benzyl);

[0018]

[0019] In formula IV, R 10 , R 11 , R 13 , R 14 are each independently hydrogen, halogen; R 12 is selected from carboxyl, C1-C3 haloalkoxy; G2 is selected from the following structures from the left side to the right side:

[0020]

[0021] In a specific embodiment, the compound of general formula I is represented by the following general formula IIIa:

[0022]

[0023] In general formula IIIa,

[0024] G1 is selected from where Z is CH2 or C=O;

[0025] R1 and R5 are each independently selected from the group consisting of: hydrogen, C3-C6 cycloalkyl, substituted or unsubstituted phenyl; wherein the substitution means that the hydrogen on the phenyl is substituted by 1, 2, or 3 groups selected from the group consisting of: halogen, C1-C3 haloalkoxy, C1-C3 haloalkyl, C1-C3 alkoxy, C1-C3 alkyl;

[0026] R2 and R4 are each independently selected from the group consisting of: hydrogen, C1-C3 alkoxy, halogen, C1-C3 haloalkyl;

[0027] R3 is selected from hydrogen, hydroxy, halogen, C1-C3 alkyl, C1-C3 haloalkyl, -N(C1-C3 alkyl)(C1-C3 alkyl), substituted or unsubstituted phenyl; wherein the substitution means that the phenyl is substituted by a group selected from the group consisting of: halogen, C1-C3 haloalkoxy, C1-C3 haloalkyl, C1-C3 alkoxy, C1-C3 alkyl;

[0028] R8 and R9 are each independently hydrogen, halogen, C1-C3 alkyl, C1-C3 alkoxy, C1-C3 haloalkoxy,

[0029] X and Y are each independently CH or N.

[0030] In a specific embodiment, in general formula IIIa,

[0031] G1 is selected from wherein Z is CH2 or C=O;

[0032] R1 and R5 are each independently selected from hydrogen, C3-C6 cycloalkyl, phenyl which is unsubstituted or substituted by 1, 2 or 3 halogens;

[0033] R2 and R4 are each independently selected from hydrogen, C1-C3 alkoxy, C1-C3 haloalkyl;

[0034] R3 is selected from hydrogen, hydroxy, halogen, C1-C3 alkyl, -N(C1-C3 alkyl)(C1-C3 alkyl), phenyl which is substituted or unsubstituted by halogen;

[0035] R8 and R9 are each independently hydrogen, halogen, C1-C3 alkoxy,

[0036] X and Y are each independently CH or N. In a specific embodiment, in general formula IIIa,

[0037] G1 is selected from wherein Z is CH2 or C=O;

[0038] R1 and R5 are selected from hydrogen, cyclopropyl, phenyl substituted by 1-3 F;

[0039] R2 and R4 are selected from hydrogen, ethoxy, trifluoromethyl;

[0040] R3 is selected from hydrogen, hydroxy, fluorine, methyl, diethylamino, p-fluorophenyl;

[0041] R8 to R9 are each independently selected from hydrogen, fluorine, methoxy,

[0042] X and Y are each independently CH or N.

[0043] In a specific embodiment, in general formula IIIa,

[0044] G1 is selected from wherein Z is CH2 or C=O;

[0045] R1 and R5 are hydrogen; R2 and R4 are ethoxy; R3 is p-fluorophenyl; R8 and R9 are each independently selected from hydrogen, fluorine, methoxy, X and Y are each independently CH or N.

[0046] In a specific embodiment, in General Formula IIIa,

[0047] One of R1 and R5 is hydrogen and the other is cyclopropyl; one of R2 and R4 is ethoxy and the other is hydrogen; R3 is methyl; R8 and R9 are each independently selected from hydrogen, fluorine, and methoxy, and X and Y are each independently CH or N.

[0048] In another specific embodiment, the compound of General Formula I is represented by the following General Formula IIIa1:

[0049]

[0050] Each substituent in General Formula IIIa1 is defined as above.

[0051] In a specific embodiment, R1, R2, R4, and R5 are each independently hydrogen, halogen, C1-C3 alkoxy, C3-C6 cycloalkyl, C1-C3 alkyl, or phenyl substituted with 1 to 3 halogens;

[0052] R3 is hydrogen, C1-C3 alkyl, or phenyl substituted with 1 to 3 halogens;

[0053] R8 and R9 are each independently hydrogen, fluorine, C1-C3 alkoxy, or C1-C3 alkyl;

[0054] Z is CH2 or C=O;

[0055] X and Y are each independently CH or N.

[0056] In another specific embodiment, the compound of General Formula I is represented by the following General Formula IVa:

[0057]

[0058] In General Formula IVa,

[0059] R1 and R5 are each independently selected from the group consisting of hydrogen, C3-C6 cycloalkyl, and substituted or unsubstituted phenyl; wherein the substitution means that the hydrogen on the phenyl is substituted by 1, 2, or 3 groups selected from the group consisting of halogen, C1-C3 haloalkoxy, C1-C3 haloalkyl, C1-C3 alkoxy, and C1-C3 alkyl;

[0060] R2 and R4 are each independently selected from the group consisting of hydrogen, C1-C3 alkoxy, halogen, and C1-C3 haloalkyl;

[0061] R3 is selected from hydrogen, halogen, C1-C3 alkyl, C1-C3 haloalkyl, and substituted or unsubstituted phenyl; wherein the substitution means that the phenyl is substituted by a group selected from the group consisting of halogen, C1-C3 haloalkoxy, C1-C3 haloalkyl, C1-C3 alkoxy, and C1-C3 alkyl;

[0062] R 10 、R 11 、R 13 、R 14 each independently represents hydrogen, halogen,

[0063] R 12 is selected from carboxyl, C1-C3 haloalkoxy, and G2 is as described above.

[0064] In a specific embodiment, in general formula IVa,

[0065] R1 and R5 are hydrogen; R2 and R4 are ethoxy; R3 is p-fluorophenyl; R 12 is selected from C1-C3 haloalkoxy and carboxyl; R 10 、R 11 、R 13 and R 14 are all hydrogen; G2 is as described above.

[0066] In a specific embodiment, in general formula IVa,

[0067] R1 and R5 are hydrogen; R2 and R4 are ethoxy; R3 is p-fluorophenyl; R 12 is selected from C1-C3 haloalkoxy and carboxyl; R 10 、R 11 、R 13 and R 14 are all hydrogen; G2 is selected from

[0068] In a specific embodiment, in general formula IVa,

[0069] R1 and R5 are hydrogen; R2 and R4 are ethoxy; R3 is p-fluorophenyl; R 12 is trifluoromethoxy or carboxyl, R 10 、R 11 、R 13 and R 14 are all hydrogen; G2 is as described above.

[0070] In a specific embodiment, the compound of general formula I is selected from one of the following compounds:

[0071]

[0072]

[0073]

[0074]

[0075] In a second aspect of the present application, there is provided a pharmaceutical composition comprising one or more therapeutically effective amounts of a compound of general formula I as described above, or a solvate, hydrate, deuterated compound, stereoisomer, tautomer, pharmaceutically acceptable salt thereof, and optionally a pharmaceutically acceptable excipient.

[0076] In a specific embodiment, the pharmaceutical composition further comprises a DPP4 inhibitor and one or more selected from TGR5 agonists, GPR40 agonists, GPR119 agonists, GPR41 agonists, and GPR43 agonists.

[0077] In a third aspect of the present application, there is provided the use of a compound of general formula I as described above, or a solvate, hydrate, deuterated compound, stereoisomer, tautomer, pharmaceutically acceptable salt thereof, or a pharmaceutical composition as described above in the preparation of a medicament for preventing or treating a disease mediated by SSTR5.

[0078] In a specific embodiment, the SSTR5-mediated diseases include type 2 diabetes, obesity, non-alcoholic fatty liver disease, gallbladder-related diseases, or inflammatory bowel disease.

[0079] In a specific embodiment, the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis; and the gallbladder-related diseases are selected from gallstones, primary sclerosing cholangitis, primary biliary cholangitis, and cholestasis.

[0080] Beneficial effects

[0081] The present invention provides a class of compounds with novel structures. Pharmacological studies have demonstrated that the compounds of the present invention have good SSTR5 antagonistic activity and can be used in the preparation of medicaments for treating diseases mediated by SSTR5. Brief description of the drawings

[0082] Figure 1 Shows the hypoglycemic experimental results of the compound of Example 7 of the present application, where *, represents P < 0.05, and ** represents P < 0.01.

[0083] Figure 2 Shows the gallbladder emptying experimental results of Compound 34 of the present application. Specific embodiments

[0084] Definitions

[0085] Unless otherwise specified, the terms used in the present invention have the following definitions:

[0086] In the present invention, the term "C1-C6" means having 1, 2, 3, 4, 5 or 6 carbon atoms, "C1-C4" means having 1, 2, 3 or 4 carbon atoms, and so on. "3-6 membered" means having 3-6 ring atoms, and so on.

[0087] As used herein, "substituted" means replaced by one or more groups (e.g., 2, 3, 4 or 5 groups). When multiple groups are selected from the same series of candidate substituents, they may be the same or different.

[0088] As used herein, "optionally" means that the defined group can be selected from a series of candidate groups or not selected.

[0089] As used herein, "alkyl" means saturated straight-chain and branched-chain alkyls with a specific number of carbon atoms, and specific examples include but are not limited to methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, tert-pentyl, etc. The "C1-3 alkyl" means saturated straight-chain or branched-chain alkyls with 1, 2 or 3 carbon atoms, and specific examples include but are not limited to methyl, ethyl, n-propyl, isopropyl, etc.

[0090] As used herein, "cycloalkyl" represents a non-aromatic, saturated, cyclic aliphatic hydrocarbon group with a specific number of carbon atoms as ring atoms. Representative examples of "C3-6 cycloalkyl" include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl.

[0091] As used herein, "alkoxy" means all straight-chain or branched-chain alkoxys with a specific number of carbon atoms, and specific examples include but are not limited to methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, etc.

[0092] The term "halogen" means fluorine, chlorine, bromine, iodine.

[0093] In the present invention, substitution on an alkyl or cycloalkyl, if not specified to occur on a specific carbon atom, means that it can occur on any carbon atom where the number of substituents has not reached saturation. When multiple substituents are selected from the same series, they may be the same or different.

[0094] In the present invention, the term "heterocyclic group" means a saturated cyclic group containing at least one ring heteroatom (e.g., N, O or S).

[0095] In the present invention, substitution on a benzene ring, heteroaromatic ring or heterocyclic ring, if not specified to occur on a specific atom, means that it can occur at any position not substituted by other atoms except hydrogen. When multiple substituents are selected from the same series, they may be the same or different.

[0096] "Pharmaceutically acceptable salts" The compounds represented by formula (I) retain the desired biological activity and have minimal toxic side effects. Such pharmaceutically acceptable salts can be obtained directly during the preparation and purification of the compounds, or indirectly by reacting the free acid or free base of the compound with another suitable base or acid.

[0097] The term "solvate" is used herein to describe a molecular complex comprising a compound of the invention and a stoichiometric amount of one or more pharmaceutically acceptable solvent molecules (such as ethanol). When the solvent is water, the term "hydrate" is used.

[0098] Pharmaceutical composition

[0099] When used for treatment, the compounds of the present invention are usually administered in the form of a standard pharmaceutical composition. It contains one or more therapeutically effective amounts of the compound represented by general formula (I), and pharmaceutically acceptable excipients. The pharmaceutically acceptable excipients are pharmaceutically acceptable carriers, excipients or sustained-release agents, etc.

[0100] The compounds and pharmaceutical compositions provided by the present invention can be in various forms, such as tablets, capsules, powders, syrups, solutions, suspensions and aerosols, etc., and can be present in a suitable solid or liquid carrier or diluent. The pharmaceutical composition of the present invention can also be stored in a suitable sterilized device for injection or infusion. The pharmaceutical composition may also contain odorants, flavorants, etc.

[0101] In the present invention, the pharmaceutical composition contains a safe and effective amount (such as 0.1-99.9 parts by weight, preferably 1-90 parts by weight) of the compound represented by general formula (I) or its pharmaceutically acceptable salt; and the balance of pharmaceutically acceptable excipients, where the total weight of the composition is 100 parts by weight. Alternatively, the pharmaceutical composition of the present invention contains 0.1-99.9% by weight, preferably 1-90% by weight of the compound represented by general formula (I) or its pharmaceutically acceptable salt based on the total weight; and the balance of pharmaceutically acceptable excipients, where the total weight of the composition is 100% by weight.

[0102] The preferred ratio of the compound represented by general formula (I) to a pharmaceutically acceptable carrier, excipient or sustained-release agent is that the compound represented by general formula (I) as the active ingredient accounts for more than 60% of the total weight, and the remaining part accounts for 0-40% of the total weight. The amount of the remaining part is preferably 1-20%, and most preferably 1-10%.

[0103] The compound represented by general formula (I) or a pharmaceutical composition comprising the compound represented by general formula (I) can be clinically used in mammals, including humans and animals. The administration routes can include oral administration, nasal inhalation, transdermal absorption, pulmonary administration, or gastrointestinal tract, etc. The preferred administration route is oral administration. It is preferably in unit dosage form, and each dose contains 0.01 mg - 200 mg of the active ingredient, preferably 0.5 mg - 100 mg, and is taken once or in divided doses. Regardless of the administration method, the optimal dose for an individual should be determined according to the specific treatment. Usually, it starts with a small dose and gradually increases the dose until the most suitable dose is found.

[0104] The pharmaceutical composition of the present invention can be administered by oral administration as well as by intravenous, intramuscular, or subcutaneous routes, etc. From the perspective of easy preparation and administration, the preferred pharmaceutical composition is a solid composition, especially tablets and solid-filled or liquid-filled capsules. Oral administration of the pharmaceutical composition is preferred.

[0105] Solid carriers include: starch, lactose, dicalcium phosphate, microcrystalline cellulose, sucrose, kaolin, etc., while liquid carriers include: sterile water, polyethylene glycol, non-ionic surfactants, and edible oils (such as corn oil, peanut oil, and sesame oil), etc., as long as they are suitable for the characteristics of the active ingredient and the specific administration method required. Adjuvants commonly used in the preparation of pharmaceutical compositions can also be advantageously included, such as flavoring agents, pigments, preservatives, and antioxidants such as vitamin E, vitamin C, BHT, and BHA.

[0106] Injectable preparations include, but are not limited to, sterile, injectable, aqueous, oily solutions, suspensions, emulsions, etc. These preparations can also be formulated with suitable parenteral diluents, dispersants, wetting agents, suspending agents, etc. Such injectable preparations can be sterilized by filtration through a bacteria-retaining filter. These preparations can also be formulated with bactericides, which are dissolved or dispersed in the injectable medium or by other methods known in the art.

[0107] Combination therapy

[0108] The compounds of the present invention can be combined with other drugs for the prevention or treatment of SSTR5-mediated diseases.

[0109] The compounds of the present invention can be used in combination with one or more other drugs to treat, prevent or improve diseases for which the compounds of the present invention or other drugs may be effective, and in which these drugs are safer or more effective when used in combination than when any one drug is used alone. The other drugs can be administered simultaneously with, before or after the compound of the present invention by common routes of administration and doses. When the compound of the present invention is used simultaneously with one or more other drugs, a pharmaceutical composition in unit dosage form containing the other drug and the compound of the present invention is preferred. However, combination therapy can also include therapies in which the general formula compounds described herein and one or more other drugs are administered in different overlapping regimens. When used in combination with one or more other active ingredients, the compounds of the present invention and the other drugs can be used at lower doses than when used alone. Wherein the other drugs include but are not limited to TGR5 agonists, GPR119 agonists, GPR40 agonists, PDE4 inhibitors, DPP4 inhibitors, SGLT2 inhibitors, metformin, insulin sensitizers, insulin and its analogs, α-glucosidase inhibitors, sulfonylureas or non-sulfonylurea insulin secretagogues, incretin analogs, etc.

[0110] The present invention will be further illustrated by the following examples. It should be particularly noted that these examples are only used to illustrate the present invention and do not limit the present invention in any way. All parameters and other descriptions in the examples are based on mass unless otherwise stated. The packing used for column chromatography separation is silica gel unless otherwise stated. The experimental methods without specific conditions noted in the following examples are generally carried out under conventional conditions or according to the conditions recommended by the manufacturer.

[0111] Unless otherwise defined, all professional and scientific terms used herein have the same meaning as those familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to those described can be applied to the present invention. The preferred methods and materials described herein are for illustrative purposes only.

[0112] Examples

[0113] Example 1

[0114] 4-(9-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-3-oxo-2,9-diazaspiro[5.5]undecan-2-yl)benzoic acid, trifluoroacetate

[0115]

[0116] Step 1: Preparation of intermediate 2-(4-(methoxycarbonyl)phenyl)-3-oxo-2,9-diazaspiro[5.5]undecane-9-carboxylic acid tert-butyl ester (A1a).

[0117]

[0118] Dissolve tert-butyl 3-oxo-2,9-diazaspiro[5.5]undecane-9-carboxylate (1 eq), methyl 4-bromobenzoate (1.5 eq), anhydrous potassium phosphate (3 eq) and Xantphos ligand (0.2 eq) in 1,4-dioxane solution. After purging with nitrogen, add Pd2(dba)3 (0.1 eq), purge with nitrogen again, and react in a sealed tube at 100 - 110 °C for 3 h. Cool the reaction mixture to room temperature, then filter off the solids in the reaction mixture, and directly load the filtrate onto silica gel for column chromatography purification to obtain the target product. The yield is about 70%.

[0119] MS(ESI): m / z 303.3 [M–Boc+H] +

[0120] 1 H NMR(500MHz,Chloroform-d)δ8.06(d,J=8.5Hz,2H),7.32(d,J=8.6Hz,2H),3.91(s,4H),3.57–3.45(m,4H),3.37(ddd,J=13.9,8.2,3.9Hz,2H),2.60(t,J=7.2Hz,2H),1.86(t,J=7.2Hz,2H),1.59(dtq,J=21.8,8.4,4.2Hz,4H),1.45(s,9H).

[0121] Step 2: Preparation of methyl 4-(3-oxo-2,9-diazaspiro[5.5]undecan-2-yl)benzoate hydrochloride (A1b)

[0122]

[0123] Dissolve the intermediate (A1a) obtained in the previous step in DCM, add an excess of hydrogen chloride / dioxane solution (4N) at room temperature, and react at room temperature for 2 h. After confirming the completion of the reaction by thin-layer chromatography, evaporate the reaction mixture to dryness to obtain the solid target product, which can be directly used in the next step without purification. The yield is 92%

[0124] MS(ESI): m / z 503.3 [M+H] + .

[0125] Step 3: Preparation of intermediate 4-(chloromethyl)-2,6-diethoxy-4′-fluoro-1,1′-biphenyl (A1c)

[0126]

[0127] Preparation method reference: ACS Med.Chem.Lett.2018,9,11,1082–1087

[0128] 1 H NMR(500MHz, chloroform-d) δ 7.39–7.31(m, 2H), 7.12–7.05(m, 2H), 6.68(s, 2H), 4.61(s, 2H), 4.01(q, J = 7.0Hz, 4H), 1.28(t, J = 6.9Hz, 6H).

[0129] MS(ESI): m / z 309.2[M+H] + .

[0130] Step 4: Preparation of methyl 4-(9-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-3-oxo-2,9-diazaspiro[5.5]undecan-2-yl)benzoate (A1d).

[0131]

[0132] Dissolve intermediate 4-(chloromethyl)-2,6-diethoxy-4′-fluoro-1,1′-biphenyl (A1c) (1eq), intermediate methyl 4-(3-oxo-2,9-diazaspiro[5.5]undecan-2-yl)benzoate hydrochloride (A1b) (1eq), and cesium carbonate (1.4eq) in acetonitrile, react at 60 °C for 3 h. After confirming the reaction is complete by thin-layer chromatography, filter off the solid in the reaction solution, directly load the filtrate onto a column, and separate by Flash column chromatography to obtain the target product with a yield of 72%.

[0133] MS(ESI): m / z 575.4[M+H] + .

[0134] 1 H NMR(500MHz, Chloroform-d) δ 8.06(d, J = 8.6Hz, 2H), 7.33(ddd, J = 8.8, 3.9, 1.8Hz, 5H), 7.04(t, J = 8.9Hz, 2H), 6.59(s, 3H), 3.96(q, J = 7.0Hz, 4H), 3.92(s, 3H), 3.52(s, 2H), 3.49(d, J = 3.6Hz, 2H), 2.59(t, J = 7.1Hz, 2H), 2.48(s, 4H), 1.84(t, J = 7.2Hz, 2H), 1.68(s, 4H), 1.23(t, J = 7.0Hz, 6H).

[0135] Step 5: Preparation of the end product A1: 4-(9-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-3-oxo-2,9-diazaspiro[5.5]undecan-2-yl)benzoic acid, trifluoroacetate

[0136] Dissolve intermediate A1d (1 eq) in a mixed solvent of 1,4-dioxane and water (volume ratio = 4:1), add lithium hydroxide monohydrate (2 eq), react at 50 °C for 12 hours, confirm the complete reaction by thin-layer chromatography, concentrate and evaporate the reaction solution to dryness, and purify by semi-preparative liquid phase to obtain trifluoroacetate A1e. The yield is 65%.

[0137] The mobile phase for semi-preparative liquid phase is acetonitrile-water (containing 0.1% trifluoroacetic acid), and the elution gradient is: 0 min: 20% acetonitrile - 80% water (containing 0.1% trifluoroacetic acid); 45 min: 75% acetonitrile - 25% water (containing 0.1% trifluoroacetic acid)

[0138] MS(ESI): m / z 561.3 [M+H] + .

[0139] 1 H NMR(500 MHz, DMSO-d6) δ 11.13 (s, 1H), 7.94 (d, J = 8.1 Hz, 2H), 7.39 (d, J = 8.1 Hz, 2H), 7.27 (dd, J = 8.6, 5.8 Hz, 2H), 7.15 (t, J = 8.9 Hz, 2H), 7.02 (s, 2H), 4.22 (s, 2H), 3.97 (q, J = 7.0 Hz, 4H), 3.80–3.57 (m, 2H), 3.20–2.92 (m, 6H), 2.03–1.59 (m, 6H), 1.15 (t, J = 6.9 Hz, 6H).

[0140] Example 2

[0141] 4-(5-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)octahydro-1H-pyrrolo[3,2-c]pyridin-1-yl)benzoic acid, trifluoroacetate

[0142]

[0143] Except that the starting material is replaced with octahydro-6H-pyrrolo[2,3-c]pyridine-6-carboxylic acid (CAS: 1196147-27-9) instead of tert-butyl 3-oxo-2,9-diazaspiro[5.5]undecane-9-carboxylate (CAS: 1251021-18-7), the remaining steps are the same as in Example 1.

[0144] MS(ESI): m / z 519.4 [M+H] + .

[0145] 1 H NMR(500 MHz, DMSO-d6) δ 9.68 (s, 1H), 7.74 (d, J = 8.5 Hz, 2H), 7.30 (dd, J = 8.4, 5.6 Hz, 3H), 7.19 (d, J = 8.8 Hz, 2H), 6.93 (s, 2H), 6.62 (d, J = 8.6 Hz, 2H), 4.34 (d, J = 38.9 Hz, 2H), 4.01 (q, J = 7.1 Hz, 4H), 3.55–3.46 (m, 2H), 3.46–3.41 (m, 2H), 3.28 (d, J = 7.2 Hz, 2H), 2.67–2.61 (m, 1H), 2.59–2.53 (m, 1H), 2.39–2.34 (m, 1H), 1.99 (dt, J = 17.2, 7.0 Hz, 1H), 1.58 (s, 1H), 1.46 (t, J = 7.2 Hz, 1H), 1.23–1.12 (m, 6H).

[0146] Example 3

[0147] 4-(9-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-oxo-3,9-diazaspiro[5.5]undecan-3-yl)benzoic acid, trifluoroacetate

[0148]

[0149] Step 1: Preparation of intermediate 9-(2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-3,9-diazaspiro[5.5]-2-undecanone (A3a)

[0150]

[0151] Dissolve 3,9-diazaspiro[5.5]-2-undecanone (1 eq), intermediate A1c (1 eq) and cesium carbonate (1.4 eq) in acetonitrile, react at 60 °C for 3 h. After confirming the reaction is complete by thin layer chromatography, filter off the solid in the reaction solution, directly mix the filtrate with the sample, and separate by Flash column chromatography to obtain the target product with a yield of 77%.

[0152] MS(ESI): m / z 441.2 [M+H] + .

[0153] 11H NMR (500 MHz, Chloroform-d) δ 7.34 (dd, J = 8.6, 5.8 Hz, 2H), 7.04 (t, J = 8.8 Hz, 2H), 6.60 (s, 2H), 6.15 (s, 1H), 3.96 (q, J = 7.0 Hz, 4H), 3.52 (s, 2H), 3.37–3.31 (m, 2H), 2.64–2.35 (m, 4H), 2.27 (s, 2H), 1.69 (t, J = 6.3 Hz, 2H), 1.57 (s, 4H), 1.24 (t, J = 7.0 Hz, 6H).

[0154] Step 2: Preparation of methyl 4-(9-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-oxo-3,9-diazaspiro[5.5]undecan-3-yl)benzoate (A3b).

[0155]

[0156] Dissolve 9-(2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-3,9-diazaspiro[5.5]undecan-2-one (A3a) (1 eq), methyl 4-bromobenzoate (1.5 eq), anhydrous potassium phosphate (3 eq) and Xantphos ligand (0.2 eq) in 1,4-dioxane solution. After purging with nitrogen, add Pd2(dba)3 (0.1 eq), purge with nitrogen again, and react in a sealed tube at 100 - 110 °C for 3 h. Cool the reaction solution to room temperature, then filter off the solid in the reaction solution, and directly load the filtrate onto silica gel for column chromatography purification to obtain the target product. The yield is 60%.

[0157] MS (ESI): m / z 575.3 [M + H] + .

[0158] 1 1H NMR (500 MHz, Chloroform-d) δ 8.09–8.03 (m, 2H), 7.38–7.31 (m, 4H), 7.05 (t, J = 8.9 Hz, 2H), 6.60 (s, 2H), 3.97 (q, J = 7.0 Hz, 4H), 3.91 (s, 3H), 3.73–3.67 (m, 2H), 3.53 (s, 2H), 2.63–2.53 (m, 2H), 2.53–2.41 (m, 4H), 1.88 (t, J = 6.3 Hz, 2H), 1.70–1.62 (m, 4H), 1.25 (t, J = 7.0 Hz, 6H).

[0159] Step 3: Preparation of the end product A3c: 4-(9-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-oxo-3,9-diazaspiro[5.5]undecan-3-yl)benzoic acid, trifluoroacetate

[0160]

[0161] Dissolve intermediate A3b (1 eq) in a mixed solvent of 1,4-dioxane and water (volume ratio = 4:1), add lithium hydroxide monohydrate (2 eq), react at 50 °C for 12 hours, confirm the complete reaction by thin-layer chromatography, concentrate and evaporate the reaction solution to dryness, and purify by semi-preparative liquid phase to obtain trifluoroacetate A3c. The yield is 68%.

[0162] MS(ESI): m / z 561.3 [M+H] + .

[0163] 1 H NMR(500MHz,DMSO-d6)δ10.06(s,1H),7.95(d,J = 8.2Hz,2H),7.45(d,J = 8.2Hz,2H),7.30(dd,J = 8.5,5.6Hz,2H),7.18(t,J = 8.8Hz,2H),6.88(s,2H),4.32(s,2H),4.00(q,J = 6.9Hz,4H),3.71(s,2H),3.34–3.07(m,6H),2.11–1.65(m,6H),1.18(t,J = 6.9Hz,6H).

[0164] Example 4

[0165] 4-(3-(1-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)piperidin-4-yl)azetidin-1-yl)benzoic acid, trifluoroacetate

[0166]

[0167] Except that the starting material is replaced with tert-butyl 4-(azetidin-3-yl)piperidine-1-carboxylate (CAS: 1314703-47-3) instead of tert-butyl 3-oxo-2,9-diazaspiro[5.5]undecane-9-carboxylate (CAS: 1251021-18-7), the remaining steps are the same as in Example 1.

[0168] MS(ESI): m / z 533.2 [M+H] + .

[0169] 11H NMR (500 MHz, DMSO-d6) δ 7.74 (d, J = 8.3 Hz, 2H), 7.29 (dd, J = 8.3, 5.7 Hz, 2H), 7.15 (t, J = 8.7 Hz, 2H), 6.68 (s, 2H), 6.39 (dd, J = 8.7, 2.4 Hz, 2H), 3.96 (p, J = 7.8, 7.0 Hz, 6H), 3.62 (t, J = 6.7 Hz, 2H), 3.56–3.47 (m, 2H), 2.98–2.84 (m, 2H), 2.11–1.89 (m, 2H), 1.68 (d, J = 12.5 Hz, 2H), 1.48 (dd, J = 17.1, 9.0 Hz, 1H), 1.24 (s, 3H), 1.15 (t, J = 7.0 Hz, 6H).

[0170] Example 5

[0171] 4-(9-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-1-oxo-4,9-diazaspiro[5.5]undec-4-yl)benzoic acid, trifluoroacetate

[0172]

[0173] Except using tert-butyl 1-oxa-4,9-diazaspiro[5.5]undecane-9-carboxylate (CAS: 930785-40-3) instead of tert-butyl 3-oxo-2,9-diazaspiro[5.5]undecane-9-carboxylate (CAS: 1251021-18-7), the preparation method is the same as that of Example 1.

[0174] MS (ESI): m / z 549.2 [M+H] + .

[0175] 1 1H NMR (500 MHz, DMSO-d6) δ 9.97 (s, 1H), 7.77 (d, J = 8.7 Hz, 2H), 7.28 (dd, J = 8.7, 5.8 Hz, 2H), 7.16 (t, J = 8.9 Hz, 2H), 6.97 (d, J = 8.7 Hz, 2H), 6.90 (s, 2H), 4.30 (s, 2H), 3.97 (q, J = 6.9 Hz, 4H), 3.84–3.74 (m, 2H), 3.32–3.23 (m, 4H), 3.17–3.03 (m, 2H), 2.16 (d, J = 14.3 Hz, 2H), 1.77 (td, J = 14.5, 4.1 Hz, 2H), 1.16 (t, J = 7.0 Hz, 6H).

[0176] Example 6

[0177] 4-((2-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-azaspiro[3.3]heptan-6-yl)amino)benzoic acid, trifluoroacetate

[0178]

[0179] Step 1: Preparation of intermediate tert-butyl (2-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-azaspiro[3.3]heptan-6-yl)carbamate (A6a)

[0180]

[0181] Dissolve tert-butyl (2-azaspiro[3.3]heptan-6-yl)carbamate (1 eq), intermediate A1c (1 eq) and cesium carbonate (1.4 eq) in acetonitrile, react at 60 °C for 3 h. After confirming the complete reaction by thin-layer chromatography, filter off the solid in the reaction solution, directly mix the filtrate with the sample, and separate by Flash column to obtain the target product (A6a) with a yield of 81%.

[0182] MS(ESI): m / z 385.3 [M-Boc+H] + .

[0183] 1 1H NMR (500 MHz, Chloroform-d) δ 7.32 (dd, J = 8.7, 5.7 Hz, 2H), 7.03 (t, J = 8.8 Hz, 2H), 6.54 (s, 2H), 3.96 (q, J = 7.0 Hz, 4H), 3.59 (s, 2H), 3.34 (s, 2H), 3.24 (s, 2H), 2.56 (t, J = 9.9 Hz, 2H), 1.97 (td, J = 8.9, 2.9 Hz, 2H), 1.42 (s, 9H), 1.23 (t, J = 7.0 Hz, 6H).

[0184] Step 2: Preparation of intermediate 2-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-azaspiro[3.3]heptan-6-amine hydrochloride (A6b).

[0185]

[0186] Dissolve the intermediate A6a obtained in the previous step in dichloromethane, add an excess of hydrogen chloride / dioxane solution (4N) and react at room temperature for 2 h. After confirming the complete reaction by thin-layer chromatography, evaporate the reaction solution to dryness to obtain the solid target product (A6b), which does not need to be purified and can be directly used in the next step. The yield is 90%.

[0187] MS(ESI): m / z 385.3 [M+H] + .

[0188] Step 3: Preparation of intermediate methyl 4-((2-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-azaspiro[3.3]heptan-6-yl)amino)benzoate (A6c).

[0189]

[0190] Dissolve 2-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-azaspiro[3.3]heptan-6-amine hydrochloride (A6b) (1 eq), methyl 4-bromobenzoate (1.5 eq), anhydrous potassium phosphate (3 eq) and Xantphos ligand (0.2 eq) in 1,4-dioxane solution. After purging with nitrogen, add Pd2(dba)3 (0.1 eq), purge with nitrogen again, and react in a sealed tube at 100 - 110 °C for 3 h. Cool the reaction solution to room temperature, then filter off the solid in the reaction solution, and directly load the filtrate onto a Flash column for purification to obtain the target product (A6c) with a yield of 45%.

[0191] MS(ESI): m / z 519.3 [M+H] + .

[0192] 1 H NMR (500 MHz, Chloroform-d) δ 7.88 (d, J = 8.9 Hz, 2H), 7.37–7.28 (m, 2H), 7.11–6.99 (m, 2H), 6.65–6.59 (m, 4H), 3.97 (q, J = 7.0 Hz, 4H), 3.85 (s, 3H), 3.83 (s, 2H), 3.33 (s, 2H), 2.97 (s, 2H), 1.88 (dt, J = 4.0, 1.8 Hz, 2H), 1.56 (dd, J = 4.4, 1.9 Hz, 2H), 1.25 (t, J = 7.0 Hz, 6H).

[0193] Step 4: Preparation of the final product 4-((2-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2-azaspiro[3.3]heptan-6-yl)amino)benzoic acid (A6d)

[0194]

[0195] Dissolve intermediate A6c (1 eq) in a mixed solvent of 1,4-dioxane and water (volume ratio = 4:1), add lithium hydroxide monohydrate (2 eq), react at 50 °C for 12 hours, confirm the complete reaction by thin-layer chromatography, concentrate the reaction solution to dryness by evaporation, and purify it by semi-preparative liquid phase to obtain A6d with a yield of 66%.

[0196] MS(ESI): m / z 505.2 [M+H] + .

[0197] 1 H NMR(500 MHz, DMSO-d6) δ 12.16 (s, 1H), 9.24 (s, 2H), 7.74 (d, J = 8.4 Hz, 2H), 7.28 (dd, J = 8.4, 5.6 Hz, 2H), 7.17 (t, J = 8.7 Hz, 2H), 6.93 (s, 2H), 6.70 (d, J = 8.5 Hz, 2H), 4.56 (s, 1H), 4.20 (s, 2H), 3.99 (q, J = 7.0 Hz, 4H), 3.34 (s, 4H), 2.04 (d, J = 4.5 Hz, 2H), 1.50 (d, J = 4.4 Hz, 2H), 1.18 (t, J = 6.9 Hz, 6H).

[0198] Example 7

[0199] 4-(2-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid, trifluoroacetate

[0200]

[0201] Except using tert-butyl 7-oxo-2,6-diazaspiro[3.4]octane-2-carboxylate (CAS: 1234616-51-3) instead of tert-butyl 3-oxo-2,9-diazaspiro[5.5]undecane-9-carboxylate (CAS: 1251021-18-7), the remaining preparation method is the same as that of Example 1.

[0202] MS(ESI): m / z 519.2 [M+H] + .

[0203] 11H NMR (500 MHz, DMSO-d6) δ 11.97 (s, 1H), 7.97 (d, J = 8.5 Hz, 2H), 7.73 (d, J = 8.5 Hz, 2H), 7.28 (dd, J = 8.6, 5.8 Hz, 2H), 7.21–7.09 (m, 2H), 7.03–6.93 (m, 2H), 4.35 (s, 2H), 4.29–4.21 (m, 2H), 4.22–4.07 (m, 4H), 4.01 (q, J = 7.0 Hz, 4H), 3.03 (s, 2H), 1.18 (t, J = 7.0 Hz, 6H).

[0204] Example 8

[0205] 4-(2-(2-Cyclopropyl-5-ethoxy-4-methylbenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid, trifluoroacetate

[0206]

[0207] Step 1: Preparation of intermediate tert-butyl 6-(4-(methoxycarbonyl)phenyl)-7-oxo-2,6-diazaspiro[3.4]octane-2-carboxylate (A8a).

[0208]

[0209] Dissolve tert-butyl 7-oxo-2,6-diazaspiro[3.4]octane-2-carboxylate (1 eq), methyl 4-bromobenzoate (1.5 eq), anhydrous potassium phosphate (3 eq) and Xantphos ligand (0.2 eq) in 1,4-dioxane solution. After purging with nitrogen, add Pd2(dba)3 (0.1 eq), and then purge with nitrogen again. React at 100 - 110 °C in a sealed tube for 3 h. Cool the reaction solution to room temperature, then filter off the solid in the reaction solution, and directly load the filtrate onto a Flash column for purification to obtain the target product (A8a) with a yield of 84%.

[0210] MS (ESI): m / z 261.1 [M - BOC + H] + .

[0211] 1 1H NMR (500 MHz, Chloroform-d) δ 8.04 (d, J = 8.8 Hz, 2H), 7.67 (d, J = 8.8 Hz, 2H), 4.04 (s, 2H), 4.02–3.96 (m, 4H), 3.90 (s, 3H), 2.87 (s, 2H), 1.44 (s, 9H).

[0212] Step 2: Preparation of intermediate methyl 4-(7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoate hydrochloride (A8b).

[0213]

[0214] Dissolve the intermediate A8a obtained in the previous step in dichloromethane, add an excess of hydrogen chloride / dioxane solution (4N), and react at room temperature for 2 h. After confirming the completion of the reaction by thin-layer chromatography, evaporate the reaction solution to dryness to obtain the solid target product (A8b), which can be directly used in the next step without purification. The yield is 93%.

[0215] MS(ESI): m / z 261.1[M+H] +

[0216] Step 3: Preparation of intermediate ethyl 2-cyclopropyl-5-ethoxy-4-methylbenzoate (A8c)

[0217]

[0218] Dissolve ethyl 2-bromo-5-ethoxy-4-methylbenzoate (1 eq), cyclopropylboronic acid (1.5 eq), and cesium carbonate (3 eq) in 1,4-dioxane. After purging with nitrogen, add PdCl2(dppf) (0.1 eq), purge with nitrogen again, and react in a sealed tube at 100 °C for 3 h. Stop the reaction. After the reaction solution cools to room temperature, filter off the solid in the reaction solution, and directly load the filtrate onto a silica gel column for flash column chromatography purification. The target product A8c is obtained with a yield of 85%.

[0219] MS(ESI): m / z 249.1[M+H] + .

[0220] 1 1H NMR(500MHz,Chloroform-d)δ7.25(s,1H),6.81(t,J=0.8Hz,1H),4.37(q,J=7.1Hz,2H),4.04(q,J=7.0Hz,2H),2.59–2.44(m,1H),2.20(s,3H),1.40(m,6H),0.94–0.88(m,2H),0.63–0.58(m,2H).

[0221] Step 4: Preparation of intermediate 2-cyclopropyl-5-ethoxy-4-methylbenzyl alcohol (A8d).

[0222]

[0223] Dissolve the obtained intermediate ethyl 2-cyclopropyl-5-ethoxy-4-methylbenzoate (A8c) (1 eq) in ultra-dry tetrahydrofuran, and add lithium aluminum hydride (1.1 eq) portionwise under an ice bath and react for 2 h. Stop the reaction after confirming complete reaction by thin-layer chromatography, and slowly add dropwise 0.5 M aqueous sodium hydroxide solution to quench. Filter off the solid in the reaction solution with diatomaceous earth, and repeatedly rinse the filter cake with ethyl acetate. After evaporating the filtrate to dryness, re-dissolve it in ethyl acetate and mix with silica gel for sample loading, and purify by Flash column chromatography to obtain the target product (A8d) with a yield of 89%.

[0224] 1 H NMR(500MHz,Chloroform-d)δ6.90(s,1H),6.84(s,1H),4.88(d,J=5.7Hz,2H),4.07(q,J=7.0Hz,2H),2.21(s,3H),1.93(m,1H),1.44(t,J=7.0Hz,3H),0.95–0.88(m,2H),0.67–0.61(m,2H).

[0225] Step 5: Synthesis of intermediate 1-(chloromethyl)-2-cyclopropyl-5-ethoxy-4-methylbenzene (A8e).

[0226]

[0227] Dissolve the obtained intermediate 2-cyclopropyl-5-ethoxy-4-methylbenzyl alcohol (A8d) in dichloromethane, and add an excess of thionyl chloride under an ice bath and react for 3 h. After confirming complete reaction by thin-layer chromatography, evaporate the reaction solution to dryness (add aqueous sodium hydroxide solution to the receiving flask of the rotary evaporator). Then re-dissolve it in dichloromethane, mix with silica gel for sample loading and separate by Flash column chromatography to obtain the target product A8e with a yield of 92%.

[0228] 1 H NMR(500MHz,Chloroform-d)δ6.84(t,J=0.8Hz,1H),6.79(s,1H),4.79(s,2H),4.03(q,J=7.0Hz,2H),2.18(d,J=0.7Hz,3H),1.98(ttd,J=8.5,5.4,0.8Hz,1H),1.41(t,J=6.9Hz,3H),0.96–0.90(m,2H),0.66–0.61(m,2H).

[0229] Step 6: Preparation of intermediate methyl 4-(2-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoate (A8f).

[0230]

[0231] The intermediate methyl 4-(7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoate hydrochloride (A8b) (1 eq), the intermediate 1-(chloromethyl)-2-cyclopropyl-5-ethoxy-4-methylbenzene (A8e) (1 eq), and potassium carbonate (1.5 eq) were dissolved in DMF and reacted overnight at room temperature. Thin layer chromatography confirmed the completion of the reaction. The mixture was extracted three times with an ethyl acetate - water system, and the ethyl acetate layers were combined and washed three times with saturated aqueous sodium chloride solution. The ethyl acetate layer was dried over anhydrous sodium sulfate, loaded onto a sample, and purified by flash column chromatography to obtain the target product A8f with a yield of 70%.

[0232] MS(ESI): m / z 449.2 [M + H] + .

[0233] 1 H NMR(600MHz,Chloroform-d)δ8.04(d,J = 8.9Hz,2H),7.72(d,J = 8.9Hz,2H),6.79(s,2H),4.08(s,2H),4.03(q,J = 7.0Hz,2H),3.91(s,3H),3.78(s,2H),3.37(d,J = 7.4Hz,2H),3.30(d,J = 7.5Hz,2H),2.85(s,2H),2.16(s,3H),1.88(tt,J = 8.4,5.4Hz,1H),1.41(t,J = 7.0Hz,3H),0.90–0.82(m,2H),0.60–0.55(m,2H).

[0234] Step 7: Preparation of the final product intermediate 4-(2-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid.

[0235]

[0236] The intermediate methyl 4-(2-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoate A8f (1 eq) was dissolved in a mixed solvent of methanol, tetrahydrofuran and water (volume ratio = 3:3:1), lithium hydroxide monohydrate (2 eq) was added, and the mixture was reacted at 50 °C for 12 hours. Thin layer chromatography confirmed the completion of the reaction. The reaction solution was concentrated to dryness and purified by semi-preparative liquid chromatography with a yield of 70%.

[0237] MS(ESI): m / z 434.2 [M + H] + .

[0238] 1 1H NMR (600 MHz, DMSO-d6) δ 7.93 (d, J = 8.8 Hz, 2H), 7.77 (d, J = 8.8 Hz, 2H), 6.83 (s, 1H), 6.71 (s, 1H), 4.07 (s, 2H), 3.99 (q, J = 6.9 Hz, 2H), 3.73 (s, 2H), 3.34 (d, J = 6.9 Hz, 2H), 3.30 (d, J = 6.8 Hz, 2H), 2.83 (s, 2H), 2.06 (s, 3H), 1.91 (m, 1H), 1.31 (t, J = 6.9 Hz, 3H), 0.86–0.80 (m, 2H), 0.54–0.50 (m, 2H).

[0239] Example 9

[0240] 4-(2-(2-Cyclopropyl-5-ethoxybenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid, trifluoroacetate

[0241]

[0242] Except for using methyl 2-bromo-5-ethoxybenzoate (CAS: 765944-34-1) instead of ethyl 2-bromo-5-ethoxy-4-methylbenzoate, the remaining synthesis steps are the same as those in Example 8.

[0243] MS (ESI): m / z 421.2 [M+H] + .

[0244] Example 10

[0245] 4-(7-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid, trifluoroacetate

[0246]

[0247] Step 1: Preparation of tert-butyl 2-(4-(methoxycarbonyl)phenyl)-2,7-diazaspiro[3.5]nonane-7-carboxylate (A10a).

[0248]

[0249] Dissolve tert-butyl 2,7-diazaspiro[3.5]nonane-7-carboxylate (1 eq), methyl 4-bromobenzoate (1.5 eq), anhydrous potassium phosphate (3 eq) and Xantphos ligand (0.2 eq) in 1,4-dioxane solution. After purging with nitrogen, add Pd2(dba)3 (0.1 eq), purge with nitrogen again, and react in a sealed tube at 100 - 110 °C for 3 h. Cool the reaction solution to room temperature, then filter off the solid in the reaction solution, directly load the filtrate onto silica gel for column chromatography purification to obtain the target product (A10a).

[0250] MS(ESI): m / z 261.1 [M–Boc+H] +

[0251] 1 1H NMR (500 MHz, Chloroform-d) δ 7.89 (d, J = 8.8 Hz, 2H), 6.36 (d, J = 8.8 Hz, 2H), 3.85 (s, 3H), 3.70 (s, 4H), 3.43–3.38 (m, 4H), 1.81–1.74 (m, 4H), 1.46 (s, 9H).

[0252] Step 2: Preparation of methyl 4-(2,7-diazaspiro[3.5]nonan-2-yl)benzoate trifluoroacetate (A10b).

[0253]

[0254] Dissolve the intermediate A10a obtained in the previous step in dichloromethane, add an excess of trifluoroacetic acid with stirring, stir at room temperature for 30 min, confirm the completion of the reaction by thin-layer chromatography, and evaporate the reaction solution to dryness to obtain the intermediate A10b.

[0255] MS(ESI): m / z 261.1 [M+H] +

[0256] Step 3: Preparation of methyl 4-(7-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoate (A10c)

[0257]

[0258] Dissolve the intermediate A10b (1 eq), the intermediate A1c (1 eq), and cesium carbonate (1.4 eq) in acetonitrile, react at 60 °C for 3 h, filter off the solid in the reaction solution after confirming the completion of the reaction by thin-layer chromatography, directly load the filtrate, and perform Flash column separation to obtain the target product A10c.

[0259] MS(ESI): m / z 532.2 [M+H] + .

[0260] 1 H NMR(500 MHz, Chloroform-d) δ 7.88 (d, J = 8.7 Hz, 2H), 7.34 (dd, J = 8.7, 5.7 Hz, 2H), 7.05 (t, J = 8.8 Hz, 2H), 6.61 (s, 2H), 6.36 (d, J = 8.8 Hz, 2H), 3.97 (q, J = 7.0 Hz, 4H), 3.85 (s, 3H), 3.68 (s, 4H), 3.48 (s, 2H), 2.44 (s, 4H), 1.87 (s, 4H), 1.25 (t, J = 7.0 Hz, 6H).

[0261] Step 4: Preparation of the final product 4-(7-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid

[0262]

[0263] Dissolve intermediate A10c: methyl 4-(7-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoate (1 eq) in a mixed solvent of methanol, tetrahydrofuran and water (volume ratio = 3:3:1), add lithium hydroxide monohydrate (2 eq), react at 50 °C for 12 hours, confirm the complete reaction by thin-layer chromatography, concentrate and evaporate the reaction solution to dryness, and purify by semi-preparative liquid chromatography to obtain the target product.

[0264] MS(ESI): m / z 519.2 [M+H] + .

[0265] 1 H NMR(600 MHz, DMSO-d6) δ 12.12 (s, 1H), 7.76 (d, J = 8.6 Hz, 2H), 7.30 (dd, J = 8.4, 6.0 Hz, 2H), 7.17 (t, J = 8.9 Hz, 2H), 7.07 (s, 2H), 6.42 (d, J = 8.7 Hz, 2H), 4.26 (d, J = 5.0 Hz, 2H), 4.01 (q, J = 6.9 Hz, 4H), 3.78 (s, 2H), 3.69 (s, 2H), 3.29 (d, J = 12.0 Hz, 2H), 2.98 (q, J = 9.8 Hz, 2H), 2.21 - 2.13 (m, 2H), 2.13 - 2.07 (m, 2H), 1.18 (t, J = 6.9 Hz, 6H).

[0266] Example 11

[0267] N-(7-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-7-azaspiro[3.5]nonan-2-yl)-4-(trifluoromethoxy)benzenesulfonamide, trifluoroacetate

[0268]

[0269] Step 1: Preparation of intermediate tert-butyl 2-(4-(trifluoromethoxy)phenyl)sulfonamido)-7-azaspiro[3.5]nonane-7-carboxylate (B11a)

[0270]

[0271] MS(ESI): m / z 365.1 [M–Boc+H] + .

[0272] 4-(Trifluoromethoxy)benzenesulfonyl chloride (1 eq), tert-butyl 2-amino-7-azaspiro[3.5]nonane-7-carboxylate (1 eq) were dissolved in dichloromethane and reacted at room temperature for 2 h. After confirming the completion of the reaction by thin layer chromatography, the reaction solution was directly mixed with samples and purified by flash column chromatography to obtain the target product B11a with a yield of 93%.

[0273] 1 H NMR(500MHz,Chloroform-d)δ7.92–7.88(m,2H),7.37–7.32(m,2H),4.71(d,J = 8.3Hz,1H),3.87–3.77(m,1H),3.29(t,J = 5.7Hz,2H),3.25–3.19(m,2H),2.17(ddt,J = 12.1,7.9,1.7Hz,2H),1.57–1.54(m,2H),1.48(t,J = 5.6Hz,2H),1.43(s,11H).

[0274] Step 2: Preparation of intermediate N-(7-azaspiro[3.5]nonan-2-yl)-4-(trifluoromethoxy)benzenesulfonamide hydrochloride (B11b)

[0275]

[0276] The intermediate B11a obtained in the previous step was dissolved in dichloromethane, and an excess of hydrogen chloride / dioxane solution (4N) was added and reacted at room temperature for 2 h. After confirming the completion of the reaction by thin layer chromatography, the reaction solution was evaporated to dryness to obtain the solid target product (B11b), which did not need to be purified and could be directly used in the next step. The yield was 95%.

[0277] MS(ESI): m / z 365.1 [M+H] +

[0278] Step 3: Preparation of the final product N-(7-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-7-azaspiro[3.5]nonan-2-yl)-4-(trifluoromethoxy)benzenesulfonamide (B11c).

[0279]

[0280] Dissolve intermediate N-(7-azaspiro[3.5]nonan-2-yl)-4-(trifluoromethoxy)benzenesulfonamide hydrochloride B11b (1 eq), intermediate 4-(chloromethyl)-2,6-diethoxy-4′-fluoro-1,1′-biphenyl (A1c) (1 eq), and potassium carbonate (3 eq) in DMF and react at room temperature overnight. Confirm complete reaction by thin-layer chromatography, extract three times with an ethyl acetate-water system, combine the ethyl acetate layers, and wash three times with saturated sodium chloride aqueous solution. After drying the ethyl acetate layer with anhydrous sodium sulfate, purify by preparative liquid chromatography (mobile phase MeCN-H2O system containing 0.1% CF3COOH) to obtain the target product B11c with a yield of 70%.

[0281] MS(ESI): m / z 637.3 [M+H] +

[0282] 1 1H NMR (600 MHz, DMSO-d6) δ 8.22 (d, J = 8.5 Hz, 1H), 7.93 (d, J = 8.8 Hz, 2H), 7.59 (d, J = 8.3 Hz, 2H), 7.31–7.24 (m, 2H), 7.16 (t, J = 8.9 Hz, 2H), 7.04 (s, 2H), 4.20–4.12 (m, 2H), 3.98 (q, J = 7.0 Hz, 4H), 3.71 (h, J = 7.6, 7.1 Hz, 1H), 3.12 (dd, J = 30.4, 11.9 Hz, 2H), 2.82 (q, J = 10.2, 7.3 Hz, 1H), 2.72 (p, J = 10.0 Hz, 1H), 2.18–2.11 (m, 1H), 1.95–1.76 (m, 4H), 1.69–1.53 (m, 3H), 1.16 (t, J = 7.0 Hz, 6H).

[0283] Example 12

[0284] 1-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-4-(1-((4-(trifluoromethoxy)phenyl)sulfonyl)piperidin-4-yl)piperazine, trifluoroacetate

[0285]

[0286] Except using tert-butyl 4-(piperidin-4-yl)piperazine-1-carboxylate (CAS: 205059-24-1) instead of tert-butyl 2-amino-7-azaspiro[3.5]nonane-7-carboxylate (CAS: 1239319-82-4), the remaining synthesis steps are the same as in Example 11.

[0287] MS(ESI): m / z 666.3 [M+H] + .

[0288] 1 H NMR(500MHz, DMSO-d6) δ 7.96–7.87(m, 2H), 7.64(d, J = 8.4Hz, 2H), 7.28(dd, J = 8.6, 5.8Hz, 2H), 7.16(t, J = 8.9Hz, 2H), 6.81(s, 2H), 4.07(s, 2H), 3.97(q, J = 6.9Hz, 4H), 3.77(d, J = 11.6Hz, 2H), 3.41–2.90(m, 9H), 2.36–2.24(m, 2H), 2.06–1.96(m, 2H), 1.69–1.54(m, 2H), 1.16(t, J = 7.0Hz, 6H).

[0289] Example 13

[0290] 1-(1-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)piperidin-4-yl)-4-((4-(trifluoromethoxy)phenyl))sulfonyl)piperazine, trifluoroacetate

[0291]

[0292] Except using tert-butyl 4-piperazinyltetrahydro-1(2H)-pyridinecarboxylate (CAS: 177276-41-4) instead of tert-butyl 2-amino-7-azaspiro[3.5]nonane-7-carboxylate (CAS: 1239319-82-4), the remaining synthesis steps are the same as in Example 11.

[0293] MS(ESI): m / z 666.3 [M+H]+.

[0294] 1H NMR (500 MHz, DMSO-d6) δ 7.91 (d, J = 8.5 Hz, 2H), 7.67 (d, J = 8.4 Hz, 2H), 7.28 (dd, J = 8.7, 5.8 Hz, 2H), 7.17 (t, J = 8.9 Hz, 2H), 6.86 (s, 2H), 4.26 (s, 2H), 3.97 (q, J = 7.0 Hz, 4H), 3.86–3.37 (m, 6H), 3.34–3.06 (m, 4H), 3.06–2.85 (m, 2H), 2.18 (d, J = 13.2 Hz, 2H), 1.88 (d, J = 13.2 Hz, 2H), 1.16 (t, J = 6.9 Hz, 6H).

[0295] Example 14

[0296] 1-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl))-4-(1-((4-(trifluoromethoxy)phenyl)sulfonyl)azetidin-3-yl)piperidine, trifluoroacetate

[0297]

[0298] Except for using tert-butyl 4-(azetidin-3-yl)piperidine-1-carboxylate (CAS: 1314703-47-3) instead of tert-butyl 2-amino-7-azaspiro[3.5]nonane-7-carboxylate (CAS: 1239319-82-4), the remaining synthesis steps are the same as in Example 11.

[0299] MS (ESI): m / z 636.2 [M+H] + .

[0300] 1 1H NMR (500 MHz, DMSO-d6) δ 8.06 (s, 1H), 7.97 (d, J = 8.8 Hz, 2H), 7.83 (dd, J = 8.6, 6.4 Hz, 1H), 7.68 (dt, J = 7.7, 1.1 Hz, 2H), 7.26 (dd, J = 8.7, 5.9 Hz, 2H), 7.19–7.08 (m, 3H), 6.58 (s, 2H), 3.91 (q, J = 7.0 Hz, 4H), 3.80 (t, J = 8.4 Hz, 2H), 3.41 (dd, J = 8.5, 6.4 Hz, 2H), 3.38–3.34 (m, 2H), 2.74 (d, J = 11.0 Hz, 2H), 2.22 (q, J = 7.4 Hz, 1H), 1.71 (s, 2H), 1.41–1.31 (m, 2H), 1.14 (t, J = 6.9 Hz, 6H), 0.94–0.78 (m, 3H).

[0301] Example 15

[0302] 4-((4-(4-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)piperazin-1-yl)piperidin-1-yl)sulfonyl)benzoic acid, trifluoroacetate

[0303]

[0304] The preparation method is the same as that of Example 12, except that 4-(chlorosulfonyl)benzoic acid (CAS: 10130-89-9) is used instead of 4-(trifluoromethoxy)benzenesulfonyl chloride (CAS: 94108-56-2).

[0305] MS(ESI): m / z 626.3 [M+H] + .

[0306] 1 H NMR(500MHz, DMSO-d6) δ 7.67(d, J = 7.9Hz, 2H), 7.35(d, J = 7.9Hz, 2H), 7.32–7.24(m, 2H), 7.17(t, J = 8.9Hz, 2H), 6.81(s, 2H), 4.60(d, J = 26.2Hz, 1H), 4.23–4.01(m, 2H), 3.98(q, J = 6.9Hz, 4H), 3.44(q, J = 7.0Hz, 4H), 3.20–2.89(m, 4H), 2.79(s, 2H), 2.14–1.82(m, 2H), 1.56(s, 2H), 1.17(t, J = 7.0Hz, 6H), 1.05(t, J = 7.0Hz, 2H).

[0307] Example 16

[0308] 1-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-4-((4-(trifluoromethoxy)phenyl)sulfonyl)piperazine

[0309]

[0310] The synthetic procedure is the same as that of Example 12, except that 1-Boc-piperazine (CAS: 57260-71-6) is used instead of tert-butyl 2-amino-7-azaspiro[3.5]nonane-7-carboxylate (CAS: 1239319-82-4).

[0311] MS(ESI): m / z 583.2 [M+H] + .

[0312] 11H NMR (500 MHz, DMSO-d6) δ 7.92 (td, J = 5.8, 2.0 Hz, 2H), 7.69 (ddd, J = 10.9, 6.3, 3.9 Hz, 2H), 7.27 (ddd, J = 8.5, 5.4, 1.9 Hz, 2H), 7.17 (td, J = 8.9, 2.0 Hz, 2H), 6.83–6.78 (m, 2H), 4.31 (s, 2H), 3.96 (qd, J = 6.9, 2.5 Hz, 4H), 3.55 (s, 8H), 1.16 (t, J = 7.0 Hz, 6H).

[0313] Example 17

[0314] 4-((4-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)piperazin-1-yl)sulfonyl)benzoic acid

[0315]

[0316] The preparation method was the same as that of Example 16 except that 4-(chlorosulfonyl)benzoic acid (CAS: 10130-89-9) was used instead of 4-(trifluoromethoxy)benzenesulfonyl chloride (CAS: 94108-56-2).

[0317] MS (ESI): m / z 543.2 [M+H] + .

[0318] 1 1H NMR (500 MHz, DMSO-d6) δ 8.20 (d, J = 8.5 Hz, 2H), 7.89 (d, J = 8.5 Hz, 2H), 7.26 (dd, J = 8.7, 5.8 Hz, 2H), 7.20–7.12 (m, 2H), 6.79 (s, 2H), 4.25 (s, 2H), 3.95 (q, J = 7.0 Hz, 4H), 3.79–3.40 (m, 8H), 1.15 (t, J = 6.9 Hz, 6H).

[0319] Example 18

[0320] 4-(7-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-1-oxo-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid, trifluoroacetate

[0321]

[0322] Except using 2,7-diazaspiro[3.5]nonan-1-one (CAS: 1147422-92-1) instead of 3,9-diazaspiro[5.5]undecan-2-one (CAS: 867006-20-0), the synthesis steps are the same as those in Example 3.

[0323] MS(ESI): m / z 533.2 [M+H] + .

[0324] 1 H NMR(500MHz, DMSO-d6) δ 7.99(dd, J = 8.7, 3.0Hz, 2H), 7.44(d, J = 8.7Hz, 2H, 7.34–7.29(m, 2H), 7.23–7.17(m, 2H), 6.99(s, 1H), 6.88(s, 1H), 4.38(dd, J = 26.2, 5.2Hz, 2H), 4.02(q, J = 7.0Hz, 4H), 3.80(s, 1H), 3.68(s, 1H), 3.56 - 3.47(m, 2H), 3.33–3.24(m, 1H) 3.19–2.09(m, 1H), 2.42(d, J = 14.0Hz, 1H), 2.24(d, J = 14.1Hz, 1H), 2.21–2.04(m, 2H), 1.21(t, J = 7.0, 6H).

[0325] Example 19

[0326] 4-(7-(2-Cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3,5]nonan-2-yl)benzoic acid, trifluoroacetate

[0327]

[0328] Except using intermediate A8e instead of intermediate A1c, the remaining preparation methods are the same as those in Example 10.

[0329] MS(ESI): m / z 435.3 [M+H] + .

[0330] 11H NMR (500 MHz, DMSO-d6) δ 9.41 (s, 1H), 7.76 (d, J = 8.6 Hz, 2H), 7.07 (s, 1H), 6.83 (s, 1H), 6.41 (d, J = 8.6 Hz, 2H), 4.44 (d, J = 4.8 Hz, 2H), 4.03 (q, J = 6.9 Hz, 2H), 3.79 (s, 2H), 3.68 (s, 2H), 3.40–3.33 (m, 2H), 3.19–3.09 (m, 2H), 2.19–2.12 (m, 2H), 2.12 (s, 3H), 2.09–2.05 (m, 1H), 1.99–1.91 (m, 2H), 1.35 (t, J = 6.9 Hz, 3H), 0.97–0.91 (m, 2H), 0.65–0.60 (m, 2H).

[0331] Example 20

[0332] 4-(7-((4-Ethoxy-2′,3′,4′-trifluoro-5-methyl-[1,1′-biphenyl]-2-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid, trifluoroacetate

[0333]

[0334] Step 1: Preparation of Intermediate 2′-(Chloromethyl)-4′-ethoxy-2,3,4-trifluoro-5′-methyl-1,1′-biphenyl (A21a)

[0335]

[0336] The remaining synthetic steps were the same as those for Intermediate A8e, except that 2,3,4-trifluorobenzeneboronic acid (CAS: 226396-32-3) was used instead of cyclopropylboronic acid (CAS: 411235-57-9).

[0337] MS (ESI): m / z 315.1 [M+H] + .

[0338] 11H NMR (500 MHz, Chloroform-d) δ 7.85 (s, 1H), 7.67 (d, J = 8.0 Hz, 1H), 7.39 (d, J = 8.0 Hz, 1H), 7.17–7.03 (m, 2H), 4.47 (s, 2H). 1H NMR (500 MHz, Chloroform-d) δ 7.08 (s, 1H), 7.07–7.00 (m, 3H), 4.51 (s, 2H), 4.15 (q, J = 7.0 Hz, 2H), 2.27 (s, 3H), 1.49 (t, J = 7.0 Hz, 3H).

[0339] Step 2: Synthesis of the final product 4-(7-((4-Ethoxy-2′,3′,4′-trifluoro-5-methyl-[1,1′-biphenyl]-2-yl)methyl)-2,7-diazaspiro[3,5]nonan-2-yl)benzoic acid

[0340]

[0341] Except for using intermediate A21a instead of intermediate A8e, the remaining synthesis steps are the same as those in Example 20.

[0342] MS (ESI): m / z 525.2 [M+H] + .

[0343] Example 21

[0344] 4-(2-(3-Ethoxy-4-fluorobenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid, trifluoroacetate

[0345]

[0346] Step 1: Synthesis of intermediate methyl 4-(2-(3-ethoxy-4-fluorobenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoate (A22a)

[0347]

[0348] Dissolve intermediate A8b: methyl 4-(7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoate (1 eq) and 3-ethoxy-4-fluorobenzaldehyde (1.1 eq) in ultradry 1,2-dichloroethane and stir at room temperature for 0.5 h. Add sodium triacetoxyborohydride (2 eq) and stir at room temperature overnight. After confirming the reaction is complete by thin layer chromatography, quench with aqueous ammonium chloride solution, extract three times with ethyl acetate, combine the organic layers, dry over anhydrous magnesium sulfate, and purify by flash column chromatography with sample mixing to obtain intermediate A22a.

[0349] MS(ESI): m / z 423.1 [M+H] + .

[0350] Step 2: Preparation of the final product 4-(2-(3-ethoxy-4-fluorobenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid

[0351]

[0352] Dissolve methyl 4-(2-(3-ethoxy-4-fluorobenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoate (1 eq) in a mixed solvent of methanol, tetrahydrofuran and water (volume ratio = 3:3:1), add lithium hydroxide monohydrate (2 eq), react at 50 °C for 12 hours, confirm the completion of the reaction by thin layer chromatography, concentrate and evaporate the reaction solution to dryness, and purify by semi-preparative liquid phase, with a yield of 70%.

[0353] MS(ESI): m / z 399.2 [M+H] +

[0354] Example 22

[0355] 4-(2-(3-Ethoxy-4-methylbenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid, trifluoroacetate

[0356]

[0357] Except for using 3-ethoxy-4-methylbenzaldehyde instead of 3-ethoxy-4-fluorobenzaldehyde, the remaining synthesis steps are the same as those in Example 21.

[0358] MS(ESI): m / z 395.2 [M+H] + .

[0359] Example 23

[0360] 4-(2-(2-Cyclopropyl-5-(trifluoromethyl)benzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid, trifluoroacetate

[0361]

[0362] Step 1: Preparation of intermediate 2-(chloromethyl)-1-cyclopropyl-4-(trifluoromethyl)benzene (A24a)

[0363]

[0364] Except using ethyl 2-bromo-5-(trifluoromethyl)benzoate (CAS: 1214336-55-6) instead of ethyl 2-bromo-5-ethoxy-4-methylbenzoate (CAS: 1350759-94-2), the remaining synthesis steps are the same as those of intermediate A8e.

[0365] 1 H NMR(500MHz,Chloroform-d)δ7.60(d,J=2.0Hz,1H),7.50(dd,J=8.1,2.0Hz,1H),7.12(d,J=8.1Hz,1H),4.82(s,2H),2.13(tt,J=8.5,5.3Hz,1H),1.13–1.05(m,2H),0.80–0.73(m,2H).

[0366] Step 2: Preparation of 4-(2-(2-cyclopropyl-5-(trifluoromethyl)benzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid

[0367]

[0368] Except using intermediate A24a instead of intermediate A8e, the remaining synthesis steps are the same as those of Example 8.

[0369] MS(ESI):m / z 445.2[M+H] + .

[0370] Example 24

[0371] 4-(7-(3-Ethoxy-4-fluorobenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid, trifluoroacetate

[0372]

[0373] Except using intermediate A10b instead of intermediate A8b, the remaining preparation methods are the same as those of Example 21.

[0374] MS(ESI):m / z 399.3[M+H] + .

[0375] 11H NMR (500 MHz, Methanol-d4) δ 7.82 (d, J = 8.7 Hz, 2H), 7.24 (d, J = 7.9 Hz, 1H), 7.17 (dd, J = 11.0, 8.3 Hz, 1H), 7.03 (dq, J = 6.1, 1.9 Hz, 1H), 6.42 (d, J = 8.7 Hz, 2H), 4.26 (s, 2H), 4.12 (q, J = 7.0 Hz, 2H), 3.87–3.63 (m, 4H), 3.50–3.35 (m, 2H), 3.14–2.97 (m, 2H), 2.16 (s, 2H), 2.03 (s, 2H), 1.41 (t, J = 7.0 Hz, 3H).

[0376] Example 25

[0377] 4-(7-(4-(Diethylamino)-3-ethoxybenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid, trifluoroacetate

[0378]

[0379] Step 1: Preparation of intermediate: Methyl 4-(diethylamino)-3-ethoxybenzoate (A26a).

[0380]

[0381] Dissolve methyl 4-amino-3-hydroxybenzoate (1 eq) in DMF, add cesium carbonate (3.5 eq), add iodoethane 4 eq with stirring, and react at 70 °C overnight. Confirm the completion of the reaction by thin layer chromatography, extract 3 times with an ethyl acetate and water system, combine the organic layers and wash 3 times with saturated sodium chloride solution. Dry the organic layer with anhydrous magnesium sulfate, filter, and flash column separate the filtrate after sample mixing to obtain the target product A26a.

[0382] MS (ESI): m / z 252.2 [M+H] + .

[0383] 1 1H NMR (500 MHz, Chloroform-d) δ 7.57 (dd, J = 8.3, 1.5 Hz, 1H), 7.48 (s, 1H), 6.83 (s, 1H), 4.11 (q, J = 7.0 Hz, 2H), 3.86 (s, 3H), 3.29 (q, J = 7.1 Hz, 4H), 1.46 (t, J = 7.0 Hz, 3H), 1.11 (t, J = 7.0 Hz, 6H).

[0384] Step 2: Preparation of intermediate (4-(Diethylamino)-3-ethoxyphenyl)methanol (A26b).

[0385]

[0386] Dissolve the intermediate A21a (1 eq) obtained in the previous step in ultradry tetrahydrofuran, add a 2.4 M solution of lithium aluminum tetrahydride in tetrahydrofuran (1 eq) under stirring at room temperature, and react at room temperature for 1 h. Confirm the completion of the reaction by thin-layer chromatography, quench with 0.5 M aqueous sodium hydroxide solution, filter through diatomaceous earth, repeatedly wash the filter cake with ethyl acetate, evaporate the filtrate to dryness, redissolve and sample with ethyl acetate, and separate by Flash column chromatography to obtain the target product A26b.

[0387] MS(ESI): m / z 224.2 [M+H] + .

[0388] 1 H NMR(500 MHz, Chloroform-d) δ 6.92–6.83 (m, 3H), 4.61 (s, 2H), 4.09 (q, J=6.9 Hz, 2H), 3.16 (q, J=7.0 Hz, 4H), 1.45 (t, J=7.0 Hz, 3H), 1.04 (t, J=7.1 Hz, 6H).

[0389] Step 3: Preparation of intermediate 4-(chloromethyl)-2-ethoxy-N,N-diethylaniline (A26c).

[0390]

[0391] Dissolve the intermediate A26b obtained in the previous step in dichloromethane, add an excess of thionyl chloride under stirring at room temperature, and react at room temperature for 30 min. Confirm the completion of the reaction by thin-layer chromatography, and evaporate the solvent to obtain the intermediate A26c.

[0392] MS(ESI): m / z 242.2 [M+H] + .

[0393] Step 4: Preparation of intermediate methyl 4-(7-(4-(diethylamino)-3-ethoxybenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoate (A26d).

[0394]

[0395] Dissolve intermediate A10b (1 eq), intermediate A26c (1.5 eq), and potassium carbonate (2 eq) in DMF, and stir and react overnight at room temperature. Confirm the completion of the reaction by thin-layer chromatography, extract 3 times with an ethyl acetate / water system, combine the organic layers, and wash 3 times with saturated sodium chloride solution. Dry the organic layer with anhydrous magnesium sulfate, filter, evaporate the filtrate to dryness, and separate by preparative TLC plate to obtain the target product A26d.

[0396] MS(ESI): m / z 466.3 [M+H] + .

[0397] 1 H NMR(500 MHz, Chloroform-d) δ 7.91 (d, J=8.8 Hz, 2H), 6.89 (d, J=8.0 Hz, 1H), 6.88 (s, 1H) 6.81 (d, J=8.0 Hz, 1H), 6.38 (d, J=8.8 Hz, 2H), 4.12 (q, J=7.0 Hz, 2H), 3.88 (s, 3H), 3.69 (s, 4H), 3.46 (s, 2H), 3.19 (q, J=7.0 Hz, 4H), 2.42 (br, 4H), 1.88 (br, 4H), 1.48 (t, J=7.0 Hz, 3H), 1.08 (t, J=7.1 Hz, 6H).

[0398] Step 5: Preparation of the final product 4-(7-(4-(diethylamino)-3-ethoxybenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid

[0399]

[0400] Dissolve intermediate A26d: methyl 4-(7-(4-(diethylamino)-3-ethoxybenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoate (1 eq) in a mixed solvent of methanol, tetrahydrofuran and water (volume ratio = 3:3:1), add lithium hydroxide monohydrate (2 eq), react at 50 °C for 12 hours, confirm the reaction is complete by thin layer chromatography, concentrate and evaporate the reaction solution to dryness, and purify by semi-preparative liquid chromatography with a yield of 70%.

[0401] MS(ESI): m / z 452.3 [M+H] + .

[0402] 1 H NMR(500 MHz, Methanol-d4) δ 7.83 (d, J=8.8 Hz, 2H), 7.68–7.60 (m, 1H), 7.45 (d, J=7.1 Hz, 1H), 7.30 (d, J=8.1 Hz, 1H), 6.44 (d, J=8.8 Hz, 2H), 4.36 (s, 2H), 4.31 (q, J=7.0 Hz, 2H), 3.78 (br, 4H), 3.62 (br, 4H), 2.13 (br, 4H), 1.49 (t, J=7.0 Hz, 3H), 1.08 (t, J=7.2 Hz, 6H).

[0403] Example 26

[0404] 4-(2-(3-Ethoxy-4-hydroxybenzyl)-7-oxo-2,6-diazaspiro[3.4]octan-6-yl)benzoic acid, trifluoroacetate

[0405]

[0406] The remaining synthesis steps are the same as those in Example 21, except that 3-ethoxy-4-hydroxybenzaldehyde is used instead of 3-ethoxy-4-fluorobenzaldehyde.

[0407] MS(ESI): m / z 397.2 [M+H] + .

[0408] 1 H NMR(500MHz, DMSO-d6) δ 7.89 (d, J = 8.5Hz, 2H), 7.59 (d, J = 8.6Hz, 2H), 6.80 (d, J = 1.7Hz, 1H), 6.72 (d, J = 8.0Hz, 1H), 6.65 (dd, J = 8.0, 1.7Hz, 1H), 4.01 (s, 2H), 4.00 (q, J = 6.8Hz, 2H), 3.44 (s, 2H), 3.21 (d, J = 7.1Hz, 2H), 3.16 (d, J = 7.0Hz, 2H), 2.76 (s, 2H), 1.32 (t, J = 7.0Hz, 3H).

[0409] Example 27

[0410] 4-(7-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)-3-fluorobenzoic acid, trifluoroacetate

[0411]

[0412] The remaining synthesis steps are the same as those in Example 10, except that methyl 3-fluoro-4-bromobenzoate is used instead of methyl p-bromobenzoate.

[0413] MS(ESI): m / z 537.3 [M+H] + .

[0414] 11H NMR (500 MHz, DMSO-d6) δ 9.94 (s, 1H), 7.62 (d, J = 8.2 Hz, 1H), 7.50 (d, J = 13.4 Hz, 1H), 7.34–7.26 (m, 2H), 7.18 (t, J = 8.7 Hz, 2H), 6.87 (s, 2H), 6.53 (t, J = 8.7 Hz, 1H), 4.29 (s, 2H), 3.99 (q, J = 6.8 Hz, 4H), 3.95 (s, 2H), 3.81 (s, 2H), 3.35 (d, J = 11.7 Hz, 2H), 3.04 (q, J = 10.4, 9.9 Hz, 2H), 2.19 (d, J = 13.7 Hz, 2H), 1.93 (t, J = 12.1 Hz, 2H), 1.18 (t, J = 6.8 Hz, 6H). Example 28

[0415] 4-(7-((2,6-Diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)-2-methoxybenzoic acid, trifluoroacetate

[0416]

[0417] Except for using methyl 4-bromo-2-methoxybenzoate instead of methyl 4-bromobenzoate, the remaining synthesis steps are the same as those in Example 10.

[0418] MS (ESI): m / z 549.3 [M+H] + .

[0419] 1 1H NMR (500 MHz, DMSO-d6) δ 7.89 (d, J = 8.5 Hz, 2H), 7.59 (d, J = 8.6 Hz, 2H), 6.80 (d, J = 1.7 Hz, 1H), 6.72 (d, J = 8.0 Hz, 1H), 6.65 (dd, J = 8.0, 1.7 Hz, 1H), 4.01 (s, 2H), 4.00 (q, J = 6.8 Hz, 2H), 3.44 (s, 2H), 3.21 (d, J = 7.1 Hz, 2H), 3.16 (d, J = 7.0 Hz, 2H), 2.76 (s, 2H), 1.32 (t, J = 7.0 Hz, 3H).

[0420] Example 29:

[0421] 5-(7-(2-Cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]non-2-yl)picolinic acid (29)

[0422]

[0423] Step 1: Preparation of tert-butyl 7-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]nonane-2-carboxylate (29a)

[0424]

[0425] Dissolve A8e (1 eq), 2-tert-butoxycarbonyl-2,7-diazaspiro[3.5]nonane (1 eq) and potassium carbonate (2 eq) in acetonitrile, react overnight at room temperature. After confirming the complete reaction by thin-layer chromatography, mix the reaction solution with silica gel and perform column chromatography to obtain intermediate 29a with a yield of 79%. 1 H NMR (500 MHz, Chloroform-d) δ 7.12 (s, 1H), 6.79 (s, 1H), 4.06 (q, J = 6.9 Hz, 2H), 3.92 (s, 2H), 3.64 (s, 4H), 2.83–2.47 (m, 4H), 2.17 (s, 3H), 2.00–1.91 (m, 4H), 1.90–1.83 (m, 1H), 1.44 (s, 9H), 1.40 (t, J = 6.9 Hz, 3H), 0.95–0.81 (m, 2H), 0.64–0.49 (m, 2H).

[0426] Step 2: Preparation of 7-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]nonane (29b)

[0427]

[0428] Dissolve 29a in dichloromethane, add an excess of hydrogen chloride - dioxane solution (4N) at room temperature, stir at room temperature for 2 hours. After confirming the complete reaction by thin-layer chromatography, concentrate the reaction solution under reduced pressure and use it directly for the next step without purification.

[0429] Step 3: Preparation of methyl 5-(7-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]non-2-yl)picolinate (29c)

[0430]

[0431] Dissolve methyl 5-fluoropicolinate (1.05 eq), 29b (1 eq) and potassium carbonate (2 eq) in DMF, react overnight at 90 °C. After confirming the complete reaction by thin-layer chromatography, add ice water to the reaction solution, extract three times with ethyl acetate, combine the organic phases and wash with saturated brine 3 times. Dry the organic phase, mix with silica gel and perform column chromatography to obtain intermediate 29c with a yield of 68%. 1HNMR(500MHz, Chloroform-d) δ 8.76 (d, J = 2.1 Hz, 1H), 7.98 (dd, J = 8.8, 2.3 Hz, 1H), 6.87 (s, 1H), 6.78 (s, 1H), 6.19 (d, J = 8.8 Hz, 1H), 4.03 (q, J = 7.0 Hz, 2H), 3.86 (s, 3H), 3.82 (s, 4H), 3.60 (s, 2H), 2.44 (s, 4H), 2.17 (s, 3H), 2.02–1.95 (m, 1H), 1.84 (t, J = 5.5 Hz, 4H), 1.40 (t, J = 6.9 Hz, 3H), 0.89–0.80 (m, 2H), 0.62–0.53 (m, 2H).

[0432] Step 4: Preparation of 5-(7-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)picolinic acid (29)

[0433]

[0434] Dissolve 29c in a mixed solution of 1,4-dioxane and water, add lithium hydroxide monohydrate, react at 60 °C overnight, confirm the complete reaction by thin-layer chromatography, concentrate the reaction solution, adjust the pH to neutral with dilute hydrochloric acid, and then purify by semi-preparative liquid phase to obtain the final product 29. 1 H NMR(500MHz, Chloroform-d) δ 7.98 (d, J = 8.5 Hz, 1H), 7.69 (s, 1H), 6.87 (s, 1H), 6.78 (s, 1H), 6.75–6.67 (m, 1H), 4.01 (q, J = 7.0 Hz, 2H), 3.74 (s, 4H), 3.64 (s, 2H), 2.53–2.38 (m, 4H), 2.17 (s, 3H), 2.04–1.93 (m, 1H), 1.87 (t, J = 5.4 Hz, 4H), 1.39 (t, J = 6.9 Hz, 3H), 0.86 (td, J = 8.2, 2.9 Hz, 2H), 0.57 (dd, J = 5.6, 1.6 Hz, 2H).

[0435] Example 30:

[0436] 6-(7-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)nicotinic acid (30)

[0437]

[0438] Preparation method:

[0439] Except that methyl 6-fluoronicotinate was used instead of methyl 5-fluoropicolinate, the remaining preparation method was the same as that in Example 29.

[0440] 1 H NMR(500MHz,Chloroform-d)δ8.78(s,1H),8.03–7.95(m,1H),6.91(s,1H),6.75(s,1H),6.10(d,J=8.8Hz,1H),3.95(q,J=7.0Hz,2H),3.81–3.71(m,6H),2.71–2.45(m,4H),2.15(s,3H),1.96–1.90(m,1H),1.33(t,J=6.9Hz,3H),0.85(d,J=8.2Hz,2H),0.55(d,J=5.2Hz,2H).

[0441] Example 31:

[0442] 4-(2-((6-Cyclopropyl-2,4′-difluoro-3-isopropoxy-[1,1′-biphenyl]-4-yl)methyl)-7-oxo-2,6-diaza[3.4]octan-6-yl)benzoic acid (31)

[0443]

[0444] Preparation method:

[0445] Step 1: Preparation of 1-bromo-3-(ethoxymethoxy)-2-fluorobenzene (31a)

[0446]

[0447] Dissolve 3-bromo-2-fluorophenol (1eq), chloromethyl ethyl ether (1.2eq), and DIPEA (1.5eq) in tetrahydrofuran, react overnight at room temperature, confirm the completion of the reaction by thin-layer chromatography, concentrate the reaction solution, redissolve it in ethyl acetate, and wash it 3 times with saturated ammonium chloride aqueous solution. Dry and concentrate the organic phase to obtain intermediate 31a with a yield of 91%. 1 H NMR(500MHz,Chloroform-d)δ7.17(ddd,J=8.6,6.7,1.9Hz,2H),6.93(td,J=8.2,1.8Hz,1H),5.26(s,2H),3.77(q,J=7.1Hz,2H),1.23(t,J=7.1Hz,4H).

[0448] Step 2: Preparation of 3-(ethoxymethoxy)-2,4′-difluoro-1,1′-biphenyl (31b)

[0449]

[0450] Dissolve 31a (1 eq), 4-fluorobenzeneboronic acid (1.5 eq), and potassium carbonate (3 eq) in a mixed solvent of toluene and water (v:v = 3:2). After purging with nitrogen, add Pd2(dba)3 (0.05 eq) and 2-dicyclohexylphosphino-2′,6′-dimethoxybiphenyl (0.1 eq). Subsequently, react at 100 °C for 3 hours under a nitrogen atmosphere. Confirm the completion of the reaction by thin-layer chromatography. Cool the reaction solution to room temperature, extract with ethyl acetate, combine the organic phases, dry, and purify by column chromatography to obtain intermediate 31b in a yield of 81%. 1 H NMR (500 MHz, Chloroform-d) δ 7.50 (ddd, J = 8.8, 5.4, 1.7 Hz, 2H), 7.21 (td, J = 7.9, 1.7 Hz, 1H), 7.15–7.07 (m, 3H), 7.02 (ddd, J = 7.9, 6.6, 1.7 Hz, 1H), 5.30 (s, 2H), 3.81 (q, J = 7.1 Hz, 2H), 1.25 (t, J = 7.1 Hz, 3H).

[0451] Step 3: Preparation of 3-(ethoxymethoxy)-2,4′-difluoro-[1,1′-biphenyl]-4-carbaldehyde (31c);

[0452]

[0453] Dissolve 31b (1 eq) in ultra-dry tetrahydrofuran, stir at -78 °C for 40 minutes, then dropwise add a n-butyllithium - n-hexane solution (1.05 eq). After the addition is complete, continue to stir at -78 °C for 1 hour. Subsequently, slowly dropwise add an N,N-dimethylformamide solution (1.1 eq) and slowly warm up to 0 °C and react for 2 hours. Confirm the completion of the reaction by thin-layer chromatography. Then add an aqueous ammonium chloride solution to quench the reaction, extract with ethyl acetate, combine the organic phases, dry, concentrate the reaction solution, and obtain intermediate 31c by column chromatography in a yield of 86%. 1 H NMR (500 MHz, Chloroform-d) δ 10.41 (d, J = 0.8 Hz, 1H), 7.68 (dd, J = 8.2, 1.4 Hz, 1H), 7.54 (ddd, J = 8.8, 5.3, 1.8 Hz, 2H), 7.28–7.21 (m, 2H), 7.16 (t, J = 8.7 Hz, 2H), 5.35 (s, 2H), 3.86 (q, J = 7.1 Hz, 2H), 1.24 (t, J = 7.1 Hz, 3H).

[0454] Step 4: Preparation of 2,4′-difluoro-3-hydroxy-[1,1′-biphenyl]-4-carbaldehyde (31d);

[0455]

[0456] Dissolve 31c in ethanol, add an excess of concentrated hydrochloric acid, heat to 50 °C and react for 0.5 hours. Then, transfer the reaction solution to room temperature and react for 1 hour. A white solid precipitates from the reaction solution. Filter the reaction solution by suction and wash the filter cake with water to obtain intermediate 31d. 1 H NMR (500 MHz, Chloroform-d) δ 11.11 (s, 1H), 9.94 (d, J = 1.7 Hz, 1H), 7.57 (ddd, J = 8.8, 5.3, 1.8 Hz, 2H), 7.42 (dd, J = 8.2, 1.5 Hz, 1H), 7.17 (t, J = 8.7 Hz, 2H), 7.05 (dd, J = 8.1, 6.2 Hz, 1H).

[0457] Step 5: Preparation of 6-bromo-2,4′-difluoro-3-hydroxy-[1,1′-biphenyl]-4-carbaldehyde (31e);

[0458]

[0459] Dissolve 31d (1 eq) in DMF, add 1,3-dibromo-1,3,5-triazine-2,4,6-trione (1.05 eq), and react at room temperature under nitrogen protection for 6 hours. Confirm the completion of the reaction by thin-layer chromatography. Quench the reaction with water, extract with ethyl acetate, combine the organic phases and wash with saturated brine 3 times. Then, dry, sample on silica gel and perform column chromatography to obtain intermediate 31e with a yield of 80%. 1 H NMR (500 MHz, Chloroform-d) δ 11.11 (s, 1H), 9.94 (d, J = 1.8 Hz, 1H), 7.58 (ddt, J = 6.9, 5.2, 1.7 Hz, 1H), 7.43 (dd, J = 8.1, 1.5 Hz, 1H), 7.22–7.13 (m, 2H), 7.05 (dd, J = 8.1, 6.2 Hz, 1H).

[0460] Step 6: Preparation of 6-bromo-2,4′-difluoro-3-isopropoxy-[1,1′-biphenyl]-4-carbaldehyde (31f);

[0461]

[0462] Dissolve 31e (1 eq) in DMF, add 2-iodopropane (1.2 eq) and potassium carbonate (1.5 eq), and react at 60 °C overnight. Confirm the completion of the reaction by thin-layer chromatography. Quench the reaction with saturated brine, extract with ethyl acetate, combine the organic phases and wash with water 3 times. Finally, dry, concentrate the organic phase and sample on silica gel, and perform column chromatography to obtain intermediate 31f with a yield of 89%. 11H NMR (500 MHz, Chloroform-d) δ 10.37 (s, 1H), 7.93 (d, J = 1.8 Hz, 1H), 7.35–7.28 (m, 2H), 7.18 (td, J = 8.7, 7.0 Hz, 2H), 4.73–4.56 (m, 1H), 1.40 (ddd, J = 9.0, 6.1, 0.8 Hz, 6H).

[0463] Step 7: Preparation of methyl 4-(2-((6-cyclopropyl-2,4′-difluoro-3-isopropoxy-[1,1′-biphenyl]-4-yl)methyl)-7-oxo-2,6-diaza[3.4]octan-6-yl)benzoate (31 g);

[0464]

[0465] Dissolve 31f (1 eq) and A8b (1 eq) in ultradry 1,2-dichloroethane, add molecular sieve for dehydration, then add sodium cyanoborohydride (3.5 eq), and react overnight at room temperature. Confirm the completion of the reaction by thin-layer chromatography. Directly sample the reaction solution with silica gel and obtain intermediate 31g by column chromatography, with a yield of 81%. 1 1H NMR (500 MHz, Chloroform-d) δ 8.04 (d, J = 8.9 Hz, 2H), 7.74–7.66 (m, 2H), 7.33 (dd, J = 8.5, 5.5 Hz, 2H), 7.14 (t, J = 8.7 Hz, 2H), 6.72 (d, J = 1.4 Hz, 1H), 4.47 (p, J = 6.1 Hz, 1H), 4.12 (s, 2H), 3.91 (s, 3H), 3.89 (s, 2H), 3.70 (d, J = 8.5 Hz, 2H), 3.56 (d, J = 8.5 Hz, 2H), 2.87 (s, 2H), 1.60 (tt, J = 8.4, 5.3 Hz, 1H), 1.33 (d, J = 6.1 Hz, 6H), 0.83–0.74 (m, 2H), 0.71–0.62 (m, 2H).

[0466] Step 8: Preparation of 4-(2-((6-cyclopropyl-2,4′-difluoro-3-isopropoxy-[1,1′-biphenyl]-4-yl)methyl)-7-oxo-2,6-diaza[3.4]octan-6-yl)benzoic acid (31)

[0467]

[0468] Except using 31g to replace 29c, the remaining synthesis methods are the same as in Example 29. 11H NMR (600 MHz, DMSO-d6) δ 7.89 (d, J = 8.7 Hz, 2H), 7.57 (d, J = 8.8 Hz, 2H), 7.43–7.37 (m, 2H), 7.29 (t, J = 8.9 Hz, 2H), 6.73 (d, J = 1.2 Hz, 1H), 4.32 (p, J = 6.1 Hz, 1H), 4.02 (s, 2H), 3.60 (s, 2H), 3.29 (d, J = 7.0 Hz, 2H), 3.26 (d, J = 6.9 Hz, 2H), 2.78 (s, 2H), 1.55 (tt, J = 8.4, 5.3 Hz, 1H), 1.26 (d, J = 6.1 Hz, 6H), 0.75 (dd, J = 8.4, 2.0 Hz, 2H), 0.60 (dd, J = 5.3, 1.9 Hz, 2H).

[0469] Example 32:

[0470] 4-(7-(2-Cyclopropyl-5-isopropoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)benzoic acid (32)

[0471]

[0472] Except using 2-iodopropane instead of iodoethane, the preparation method is the same as that of Example 19. 1 1H NMR (600 MHz, Methanol-d4) δ 7.82 (d, J = 8.7 Hz, 2H), 6.91 (s, 1H), 6.76 (s, 1H), 6.39 (d, J = 8.8 Hz, 2H), 4.52 (p, J = 6.1 Hz, 1H), 3.63 (s, 2H), 3.62 (s, 4H), 2.59–2.35 (m, 4H), 1.99 (tt, J = 8.4, 5.4 Hz, 1H), 1.84 (t, J = 5.5 Hz, 4H), 1.30 (d, J = 6.0 Hz, 6H), 0.90–0.82 (m, 2H), 0.57–0.48 (m, 2H).

[0473] Example 33:

[0474] 4-(7-((6-Cyclopropyl-2,4′-difluoro-3-isopropoxy-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)-2-fluorobenzoic acid (33)

[0475]

[0476] Step 1: Preparation of tert-butyl 2-(2-fluoro-4-(methoxycarbonyl)phenyl)-2,7-diazaspiro[3.5]nonane-7-carboxylate (33a)

[0477]

[0478] Except that methyl 4-bromo-3-fluorobenzoate is used instead of methyl 4-bromobenzoate, the remaining preparation method is the same as that of A10a. 1 H NMR(500MHz,Chloroform-d)δ7.68(dd,J=8.4,1.9Hz,1H),7.59(dd,J=13.4,1.8Hz,1H),6.36(t,J=8.7Hz,1H),3.86(s,3H),3.84(d,J=2.3Hz,4H),3.46–3.35(m,4H),1.79(dd,J=6.7,4.5Hz,4H),1.47(s,9H).

[0479] Step 2: Preparation of 4-(7-((6-cyclopropyl-2,4′-difluoro-3-isopropoxy-[1,1′-biphenyl]-4-yl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)-2-fluorobenzoic acid (33)

[0480]

[0481] Except that 33a is used instead of A8b, the remaining preparation method is the same as that of Example 31. 1 H NMR(600MHz,DMSO-d6)δ7.54(dd,J=8.0,1.7Hz,1H),7.47(dd,J=13.9,1.7Hz,1H),7.41(dd,J=8.5,5.6Hz,2H),7.29(t,J=8.9Hz,2H),6.81(s,1H),6.39(t,J=8.7Hz,1H),4.35(p,J=6.1Hz,1H),3.66(s,4H),3.45(s,2H),2.36(s,4H),1.76(t,J=5.6Hz,4H),1.56(td,J=8.4,4.2Hz,1H),1.25(d,J=6.2Hz,6H),0.75(dt,J=8.6,3.1Hz,2H),0.59–0.51(m,2H).

[0482] Example 34

[0483] 4-(7-(2-cyclopropyl-5-ethoxy-4-methylbenzyl)-2,7-diazaspiro[3.5]nonan-2-yl)-2-fluorobenzoic acid (34)

[0484]

[0485] Except that 33a is used instead of A10a, the remaining preparation method is the same as that of Example 19.1 1H NMR (500 MHz, DMSO-d6) δ 7.61 (dd, J = 8.4, 1.8 Hz, 1H), 7.49 (dd, J = 13.5, 1.8 Hz, 1H), 7.06 (s, 1H), 6.83 (s, 1H), 6.52 (t, J = 8.8 Hz, 1H), 4.43 (d, J = 5.1 Hz, 2H), 4.03 (q, J = 7.0 Hz, 2H), 3.96 (s, 2H), 3.79 (s, 2H), 3.35 (d, J = 11.9 Hz, 2H), 3.20–3.10 (m, 2H), 2.16 (d, J = 13.9 Hz, 2H), 2.12 (s, 3H), 2.07 (tt, J = 5.4, 3.0 Hz, 1H), 1.93 (dt, J = 13.0, 7.0 Hz, 2H), 1.35 (t, J = 7.0 Hz, 3H), 0.97–0.90 (m, 2H), 0.66–0.60 (m, 2H). (ESI, m / z): 453.3 [M+H] + 。

[0486] Pharmacological experiment

[0487] Experimental Example 1. In vitro test of SSTR5 antagonistic activity by calcium flux assay

[0488] Experimental principle: After the receptor-bound ligand is activated, it can cause the activation of Gα16 protein, which in turn activates phospholipase C (PLC) to produce IP3 and DAG. IP3 can bind to the IP3 receptors on the endoplasmic reticulum and mitochondria in the cell, thereby causing the release of intracellular calcium. Therefore, measuring the change in intracellular calcium can be used as a method to detect the activation state of hSSTR5. Fluo-4 / AM is a calcium fluorescent probe indicator used to measure calcium ions. As a non-polar lipid-soluble compound, after entering the cell, under the action of cell lipolytic enzymes, the AM group dissociates and releases Fluo-4; since Fluo-4 is a polar molecule and is not easily passed through the lipid bilayer membrane, it can retain Fluo-4 in the cell for a long time. Finally, the level of Gα protein activation can be reflected by measuring the intensity of the excited fluorescence. The SSTR5 agonist Somatostatin can activate the SSTR5 receptor, resulting in a significant increase in the calcium flux response. The SSTR5 antagonist to be tested can inhibit the agonist activity of the SSTR5 receptor, reducing the calcium flux response increased by the agonist stimulation of the receptor.

[0489] Experimental procedure:

[0490] (1) Seed the hSSTR5-CHO-Gɑ16 and mSSTR5-CHO-Gɑ16 cell lines stably expressing the hSSTR5 or mSSTR5 receptor at a density of 30,000 cells / well in a 96-well plate and culture overnight in an incubator at 37°C.

[0491] (2) Aspirate the culture medium and add 40 μL / well of freshly prepared dye, and incubate at a constant temperature in an incubator at 37 °C for 45 minutes.

[0492] (3) Aspirate and discard all the dye, wash once with freshly prepared calcium buffer, and then replace it with 50 μL of calcium buffer.

[0493] (4) Dilute and mix the agonist Somatostatin-14 with calcium buffer.

[0494] (5) Detect with a Flextation instrument. Starting from the 15th second, the instrument automatically adds 25 μL of the pre-prepared agonist, and finally reads the fluorescence value at 525 nm. The EC 50 of the agonist is 9.7 nM, and an agonist concentration of 100 nM is selected for the detection of antagonist activity.

[0495] (6) Dilute and mix the positive control antagonist and the test compound with calcium buffer; among them, the positive control antagonist is 4-(8-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-3-oxo-2,8-diazaspiro[4.5]dec-2-yl)benzoic acid, from the article ACS Med.Chem.Lett. 2018, 9, 1082 - 1087, corresponding to compound 10 therein.

[0496] (7) Aspirate and discard all the dye, wash once with freshly prepared calcium buffer, and then replace it with 50 μL of the antagonist diluted in a 10-fold concentration gradient.

[0497] (8) Detect with a Flextation instrument. Starting from the 15th second, the instrument automatically adds 25 μL of the pre-prepared agonist with a concentration of 100 nM, and finally reads the fluorescence value at 525 nm.

[0498] Experimental results

[0499] The in vitro activity test results of the compounds prepared in the above Examples 1 - 28 are shown in Table 1.

[0500] Table 1

[0501]

[0502]

[0503] Method for representing the activity range: A: 0.1 - 50 nM; B: 50 - 200 nM; C: >200 nM; -: not measured

[0504] The above data indicate that this class of compounds has good SSTR5 antagonistic activity. In particular, Examples 2, 7, 10, 12, 14, 18, 19, 27, and 28 exhibit strong SSTR5 antagonistic activity.

[0505] Experimental Example 2. Oral Glucose Tolerance (OGTT) Test of the Compounds of the Present Invention in C57BL / 6 Normal Mice

[0506] Experimental method: C57BL / 6 mice were fasted for 12 h before the experiment. A dosing group and a blank control group were set up, with 8 mice in each group. The mice in the blank control group were orally administered 200 μl of pure water, and the mice in the dosing group were orally administered 200 μl of an aqueous solution containing the compound of Example 7 above / positive control drug (the same as in Experimental Example 1). Glucose (4 g glucose / kg mouse body weight) was orally administered 60 min after dosing. Blood was taken from the tail before dosing and at 0, 15, 30, 60, 90, and 120 min, and blood glucose was measured using an Accu-Chek Advantage II Glucose Monitor (Roche, Indianapolis, IN, USA).

[0507] The results of the oral glucose tolerance (OGTT) test of normal mice after single administration of the compound are as Figure 1 shown.

[0508] Among them, the positive control is 4-(8-((2,6-diethoxy-4′-fluoro-[1,1′-biphenyl]-4-yl)methyl)-3-oxo-2,8-diazaspiro[4.5]dec-2-yl)benzoic acid, from the article ACS Med. Chem. Lett. 2018, 9, 1082 - 1087.

[0509] The above data indicate that this class of compounds has good hypoglycemic effects.

[0510] Experimental Example 3. Gallbladder Emptying Promotion Experiment

[0511] Experimental method: C57BL / 6J mice aged 8 - 10 weeks were fasted for 17 - 18 hours before the experiment and allowed free access to water. Subsequently, the mice were grouped by body weight, with 4 mice in each group. They were respectively intragastrically administered an aqueous solution of the compound prepared in Example 34 (30 mg / kg) or an equal volume of distilled water (control group), and the dosing volume was 10 mL / kg. One hour after dosing, 200 μl of egg yolk was orally administered to the mice. After 15 minutes, the mice were sacrificed by cervical dislocation and dissected. The gallbladder was removed, and the bile was squeezed out and weighed using an analytical balance.

[0512] The experimental results are as Figure 2As shown, it can be seen from the figure that the compound of Example 34 of the present application can significantly promote the gallbladder emptying of mice compared with the control group. Therefore, it can be applied to the prevention and treatment of gallbladder-related diseases such as gallstones, cholestasis, and primary sclerosing cholangitis.

[0513] The above are only specific embodiments of the present invention, but the protection scope of the present invention is not limited thereto. Any person skilled in the art within the technical scope disclosed by the present invention can easily think of changes or substitutions, which should be covered within the protection scope of the present invention. Therefore, the protection scope of the present invention should be subject to the protection scope of the claims.

Claims

1. A compound represented by the following general formula IIIa1, or a pharmaceutically acceptable salt thereof: In general formula IIIa1, R 1, R2, R4, and R5 are each independently hydrogen, halogen, C1-C3 alkoxy, C3-C6 cycloalkyl, C1-C3 alkyl, or phenyl substituted with 1-3 halogens; R3 is hydrogen, halogen, C1-C3 alkyl or phenyl substituted with 1-3 halogens; R8 and R9 are each independently hydrogen, fluorine, C1-C3 alkoxy, C1-C3 alkyl; Z is CH2 or C=O; X and Y are each independently CH or N.

2. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein, The compound of general formula IIIa1 is selected from one of the following compounds:

3. A pharmaceutical composition comprising one or more therapeutically effective amounts of the compound of general formula IIIa1 as described in claim 1 or 2, or a pharmaceutically acceptable salt thereof, and optionally a pharmaceutically acceptable excipient.

4. The pharmaceutical composition according to claim 3, wherein, The pharmaceutical composition further comprises a DPP4 inhibitor and one or more selected from TGR5 agonists, GPR40 agonists, GPR119 agonists, GPR41 agonists and GPR43 agonists.

5. Use of a compound of general formula IIIa1 as claimed in claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition as claimed in claim 3 or 4, in the manufacture of a medicament for the prevention or treatment of a disease mediated by SSTR5, wherein, The SSTR5-mediated diseases are selected from type 2 diabetes and gallbladder-related diseases.

6. The use according to claim 5, wherein The gallbladder-related diseases are selected from gallstones, primary sclerosing cholangitis, primary biliary cholangitis and cholestasis.

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