A series of piperidine-substituted benzoic acid compounds and their applications

Abstract

A series of piperidine-substituted benzoic acid compounds and their applications, specifically disclosing compounds represented by formula (I) and pharmaceutically acceptable salts thereof.

Description

[0001] The present invention claims the following priority:

[0002] CN 202011610857.7, application date December 30, 2020;

[0003] CN 202110266011.4, application date March 11, 2021;

[0004] CN 202110751316.4, application date July 2, 2021;

[0005] CN 202111234085.6, application date October 22, 2021. Technical Field

[0006] The present invention relates to a series of piperidine-substituted benzoic acid compounds, and in particular to compounds represented by formula (I) and pharmaceutically acceptable salts thereof. Background Art

[0007] Immune diseases are diseases caused by the loss of balance in immune regulation that affects the body's immune response. The complement system is an important component of the immune system, which includes a group of proteins that usually exist in an inactive state, namely complement. Complement can be activated by substances such as lipopolysaccharide, polysaccharide, peptidoglycan, teichoic acid, and condensed IgA and IgG4 in the alternative activation pathway, mediating immune responses and inflammatory responses. Among them, the activating substance directly activates C3 and then completes the chain reaction of components C5 to C9. Complement factor B, also known as C3 activator precursor, can be cleaved into two fragments, Ba and Bb, by complement factor D. Bb combines with C3b to form the C3 convertase of the alternative pathway to play a role. Complement Factor B acts on the AP pathway. Inhibiting the activity of Factor B can prevent the activation of the API pathway without interfering with the CP and LP pathways, which can avoid the increased risk of infection due to complement system inhibition.

[0008] Currently, there is no small molecule Factor B inhibitor on the market. Novartis' Factor B inhibitor LNP023 is in Phase III clinical research for the treatment of PNH, IgAN, C3G and other diseases. Therefore, there is an urgent need to develop new small molecule inhibitors of the complement system Factor B, increase clinical research and verification, and use them for the treatment of various diseases caused by complement abnormalities. Summary of the invention

[0009] The present invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,

[0010]

[0011] in,

[0012] L is selected from a single bond, NR 4 and O;

[0013] R 1 Selected from fluorine, chlorine, C 1-5 Alkyl, C 3-6 Cycloalkyl, 3-6 membered heterocycloalkyl, -C 1-3 Alkyl-C 1-3 Alkoxy, -C 1-3 Alkyl-C 3-6 Cycloalkyl and -C 1-3 Alkyl-3-6 membered heterocycloalkyl, the C 1-5 Alkyl, C 3-6 Cycloalkyl, 3-6 membered heterocycloalkyl, -C 1-3 Alkyl-C 1-3 Alkoxy, -C 1-3 Alkyl-C 3-6 Cycloalkyl and -C 1-3 alkyl-3-6 membered heterocycloalkyl each independently optionally substituted by 1, 2 or 3 R a replace;

[0014] R 2 , R 3 and R 4 are independently selected from H and C 1-5 Alkyl, the C 1-5 The alkyl group is optionally substituted with 1, 2 or 3 R b replace;

[0015] Each R a and R b are independently selected from H, F, Cl, Br and I;

[0016] The condition is that when R 1 Selected from unsubstituted C 1-5 When alkyl, R 2 and R 3 Not selected from H at the same time.

[0017] In some embodiments of the present invention, the compound or a pharmaceutically acceptable salt thereof is selected from:

[0018]

[0019] in,

[0020] L, R 1 , R 2 and R 3 As defined in the present invention;

[0021] The carbon atom with "*" is a chiral carbon atom, which exists in the form of a single enantiomer (R) or (S) or in the form enriched in one enantiomer.

[0022] In some embodiments of the present invention, the compound or a pharmaceutically acceptable salt thereof is selected from:

[0023]

[0024] Among them, L, R 1 , R 2 and R 3 As defined in the present invention.

[0025] In some embodiments of the present invention, the R 1 Selected from C 3-6 Cycloalkyl, 3-6 membered heterocycloalkyl, -C 1-3 Alkyl-C 1-3 Alkoxy, -C 1-3 Alkyl-C 3-6 Cycloalkyl and -C 1-3 Alkyl-3-6 membered heterocycloalkyl, the C 3-6 Cycloalkyl, 3-6 membered heterocycloalkyl, -C 1-3 Alkyl-C 1-3 Alkoxy, -C 1-3 Alkyl-C 3-6 Cycloalkyl and -C 1-3 alkyl-3-6 membered heterocycloalkyl is each independently optionally substituted by 1, 2 or 3 R a The other variables are as defined in the present invention.

[0026] In some embodiments of the present invention, the R 1 Selected from fluorine, chlorine, CH 3 , CH 2 CH 3 , CH 2 CH 2 CH 3 、CH(CH 3 ) 2 , The CH 3 , CH 2 CH 3 , CH 2 CH 2 CH 3 、CH(CH 3 ) 2 , are independently optionally substituted by 1, 2 or 3 R a The other variables are as defined in the present invention.

[0027] In some embodiments of the present invention, the R 1 Selected from CH 3 CF 3 , CH 2 CH 3 , CH 2 CHF 2 , CH 2 CF 3 , Other variables are as defined in the present invention.

[0028] In some embodiments of the present invention, the R 1 Selected from Other variables are as defined in the present invention.

[0029] In some embodiments of the present invention, the R 1 Selected from C 1-5 Alkyl, R 2 Selected from H and C 1-5 Alkyl, R 3 Selected from C 1-5 Alkyl, and other variables are as defined herein.

[0030] In some embodiments of the present invention, the R 1 Selected from CH 2 CH 3 , R 2 Selected from H and CH 3 , R 3 Selected from CHF 2 and CH 3 , other variables are as defined in the present invention.

[0031] In some embodiments of the present invention, the R 2 Selected from H and CH 3 , the CH 3 Optional 1, 2 or 3 R b The other variables are as defined in the present invention.

[0032] In some embodiments of the present invention, the R 2 Selected from H, CH 3 and CHF 2 , other variables are as defined in the present invention.

[0033] In some embodiments of the present invention, the R 3 Selected from H and CH 3 , other variables are as defined in the present invention.

[0034] In some embodiments of the present invention, the R 4 Selected from H and CH3 , other variables are as defined in the present invention.

[0035] In some embodiments of the present invention, L is selected from a single bond and O, and other variables are as defined herein.

[0036] In some embodiments of the present invention, the structural unit Selected from Other variables are as defined in the present invention.

[0037] In some embodiments of the present invention, the structural unit Selected from Other variables are as defined in the present invention.

[0038] In some embodiments of the present invention, the structural unit Selected from Other variables are as defined in the present invention.

[0039] The present invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,

[0040]

[0041] in,

[0042] L is selected from a single bond, NR 4 and O;

[0043] R 1 Selected from fluorine, chlorine, C 1-5 Alkyl, C 3-6 Cycloalkyl, 3-6 membered heterocycloalkyl, -C 1-3 Alkyl-C 1-3 Alkoxy, -C 1-3 Alkyl-C 3-6 Cycloalkyl and -C 1-3 Alkyl-3-6 membered heterocycloalkyl, the C 1-5 Alkyl, C 3-6 Cycloalkyl, 3-6 membered heterocycloalkyl, -C 1-3 Alkyl-C 1-3 Alkoxy, -C 1-3 Alkyl-C 3-6 Cycloalkyl and -C 1-3 alkyl-3-6 membered heterocycloalkyl is each independently optionally substituted by 1, 2 or 3 R a replace;

[0044] R 2 , R3 and R 4 are independently selected from H and C 1-5 Alkyl, the C 1-5 The alkyl group is optionally substituted with 1, 2 or 3 R b replace;

[0045] Each R a and R b are independently selected from H, F, Cl, Br and I;

[0046] The condition is that when R 1 Selected from C 1-5 When alkyl, R 2 and R 3 Not selected from H at the same time.

[0047] Some other solutions of the present invention are obtained by any combination of the above variables.

[0048] The present invention also provides the following compounds or pharmaceutically acceptable salts thereof:

[0049]

[0050] In some embodiments of the present invention, the compound or a pharmaceutically acceptable salt thereof is selected from

[0051]

[0052]

[0053]

[0054] The present invention also provides the use of the compound or a pharmaceutically acceptable salt thereof in preparing a drug related to complement factor B.

[0055] The present invention also provides the following test method:

[0056] Method 1. PHN Heimann Nephritis Drug Efficacy Test

[0057] 1.1 Experimental purpose: To investigate the ability of the compounds of the present invention to improve the renal function of rats with Heimann nephritis induced by Sheep Anti-Rat Fx1A Serum, including the evaluation of reducing proteinuria levels and improving renal tissue damage.

[0058] 1.2 Experimental animals: Male SD rats, 7-10 weeks old, weighing 200-300 g

[0059] 1.3 Experimental process:

[0060] 1.3.1 Modeling: Rat urine was collected on D-2 days before administration. D1 was the first day of the experiment. The control group (Group 1) was given a single injection of 5 mL / kg Sheep Non-Immune serum through the tail vein; the modeling group and the drug administration group (Groups 2 to 6) were given a single injection of 5 mL / kg Sheep Anti-Rat Fx1A Serum through the tail vein.

[0061] 1.3.2 Administration:

[0062] The administration method was oral gavage, twice a day, with an interval of 8 hours, and the administration volume was 10mL / kg. On D1 day, 1 hour before modeling, animals in Group 1 and Group 2 were given blank solvent 20% PEG 400 / 10% Solutol / 70% water; animals in Group 3 were given LNP023 (60mpk); Groups 4 to 6 were given different concentrations of compound 4B (5mpk, 20mpk and 60mpk); 8 hours after the first administration, each group was administered again, and the dosage and volume were the same as the first time. From D2 to D14 days, animals in each group were given different compounds or solvents for 14 consecutive days (including Day 1) according to the dosage, volume, administration method and frequency on D1.

[0063] 1.3.3 Sample collection:

[0064] (1) Urine collection: Rat urine samples were collected on D-2 days before administration, and 2-4 h and 4-6 h after the first administration on D4, D6, D8, D11, and D14. The samples were transferred into EP tubes and stored in a refrigerator at -80°C to -60°C for the detection of rat urine protein and urine creatinine.

[0065] (2) Kidney tissue collection: All animals were collected on D15 after administration with CO 2 The animals were euthanized by inhalation anesthesia, and bilateral kidneys were collected. The left kidney was cut transversely, and the right kidney was cut longitudinally. The transverse half (left side) plus the longitudinal half (right side) of the kidney were fixed in formalin (placed in the same EP tube), and the remaining transverse half (left side) plus the longitudinal half (right side) of the kidney were embedded in OCT, with the cut surface facing down (placed in the same embedding box). After embedding, the kidneys were stored in a refrigerator at -80℃ to -60℃ as soon as possible for histopathological scoring.

[0066] 1.3.4 Sample analysis: Urine protein and urine creatinine data were read by the analytical instrument HITACHI LST008AS (P). The renal glomerulopathy (characterized by glomerular enlargement, thickening of the basement membrane and Bowman's capsule, and enlarged / rounded podocytes) was scored by histopathological analysis, the tubular degeneration score was determined by histopathological analysis, and the glomerular C3 deposition score was analyzed by IHC or IF staining.

[0067] Technical Effects

[0068] The compounds of the present invention have obvious inhibitory activity on the activation of human serum bypass pathway, can significantly inhibit LPS-stimulated complement activation, reduce urine protein levels, and improve renal function; PK results show that the compounds of the present invention exhibit a longer half-life and a higher drug exposure, have excellent in vivo pharmacokinetic properties, and have better drugability.

[0069] Definition and Description

[0070] Unless otherwise specified, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase should not be considered to be uncertain or unclear in the absence of a special definition, but should be understood according to its ordinary meaning. When a trade name appears in this article, it is intended to refer to its corresponding commercial product or its active ingredient.

[0071] The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0072] The term "pharmaceutically acceptable salt" refers to salts of compounds of the invention, prepared from compounds of the invention having specific substituents with relatively nontoxic acids or bases. When the compounds of the invention contain relatively acidic functional groups, base addition salts can be obtained by contacting such compounds with a sufficient amount of base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine or magnesium salts or similar salts. When the compounds of the invention contain relatively basic functional groups, acid addition salts can be obtained by contacting such compounds with a sufficient amount of acid in a pure solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts, such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, hydrogen sulfate, hydroiodic acid, phosphorous acid, etc.; and organic acid salts, such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid and methanesulfonic acid, and salts of amino acids (such as arginine, etc.), and salts of organic acids such as glucuronic acid. Certain specific compounds of the present invention contain basic and acidic functional groups, and thus can be converted into any base or acid addition salt.

[0073] Pharmaceutically acceptable salts of the present invention can be synthesized by conventional chemical methods from parent compounds containing acid radicals or bases. Generally, the preparation method of such salts is: in water or an organic solvent or a mixture of the two, these compounds in free acid or base form are reacted with a stoichiometric amount of an appropriate base or acid to prepare.

[0074] The compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, and racemic mixtures and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which are within the scope of the present invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl. All of these isomers and their mixtures are included within the scope of the present invention.

[0075] Unless otherwise indicated, the term "enantiomer" or "optical isomer" refers to stereoisomers that are mirror images of one another.

[0076] Unless otherwise indicated, the term "cis-trans isomers" or "geometric isomers" arises from the inability of a ring to rotate freely about double bonds or single bonds of ring carbon atoms.

[0077] Unless otherwise indicated, the term "diastereomer" refers to stereoisomers that have two or more chiral centers and that are not mirror images of each other.

[0078] Unless otherwise indicated, "(+)" indicates dextrorotatory, "(-)" indicates levorotatory, and "(±)" indicates racemic.

[0079] Unless otherwise specified, the key is a solid wedge. and dotted wedge key To indicate the absolute configuration of a stereocenter, use a straight solid bond. and straight dashed key To indicate the relative configuration of a stereocenter, use a wavy line Indicates a wedge-shaped solid key or dotted wedge key Or use a wavy line Represents a straight solid bond and straight dashed key

[0080] Unless otherwise indicated, the term "tautomer" or "tautomeric form" refers to isomers of different functional groups that are in dynamic equilibrium at room temperature and can rapidly interconvert. If tautomerism is possible (such as in solution), chemical equilibrium of tautomers can be achieved. For example, proton tautomers (also called prototropic tautomers) include interconversions via proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers include interconversions via reorganization of some bonding electrons. A specific example of keto-enol tautomerization is the interconversion between pentane-2,4-dione and 4-hydroxypent-3-en-2-one.

[0081] Unless otherwise indicated, the terms "enriched in one isomer", "isomerically enriched", "enriched in one enantiomer" or "enantiomerically enriched" mean that the content of one isomer or enantiomer is less than 100%, and the content of the isomer or enantiomer is greater than or equal to 60%, or greater than or equal to 70%, or greater than or equal to 80%, or greater than or equal to 90%, or greater than or equal to 95%, or greater than or equal to 96%, or greater than or equal to 97%, or greater than or equal to 98%, or greater than or equal to 99%, or greater than or equal to 99.5%, or greater than or equal to 99.6%, or greater than or equal to 99.7%, or greater than or equal to 99.8%, or greater than or equal to 99.9%.

[0082] Unless otherwise indicated, the term "isomer excess" or "enantiomeric excess" refers to the difference between the relative percentages of two isomers or two enantiomers. For example, if the content of one isomer or enantiomer is 90% and the content of the other isomer or enantiomer is 10%, the isomer or enantiomeric excess (ee value) is 80%.

[0083] The compounds of the invention may contain unnatural proportions of atomic isotopes on one or more of the atoms that make up the compound. For example, the compounds may be labeled with radioactive isotopes, such as tritium ( 3 H), iodine-125( 125 I) or C-14( 14 C). For another example, deuterium can be used to replace hydrogen to form a deuterated drug. The bond between deuterium and carbon is stronger than the bond between ordinary hydrogen and carbon. Compared with undeuterated drugs, deuterated drugs have the advantages of reducing toxic side effects, increasing drug stability, enhancing efficacy, and extending the biological half-life of drugs. All isotopic composition changes of the compounds of the present invention, whether radioactive or not, are included in the scope of the present invention.

[0084] The terms "optional" or "optionally" mean that the subsequently described event or circumstance may but need not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not.

[0085] The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, which may include a variant of deuterium and hydrogen, as long as the valence state of the particular atom is normal and the substituted compound is stable. When the substituent is oxygen (i.e., =O), it means that two hydrogen atoms are replaced. Oxygen substitution does not occur on aromatic groups. The term "optionally substituted" means that it may be substituted or not substituted, and unless otherwise specified, the type and number of the substituents may be arbitrary on the basis of chemical achievable.

[0086] When any variable (e.g., R) occurs more than once in a compound's composition or structure, its definition at each occurrence is independent. Thus, for example, if a group is substituted with 0-2 Rs, the group may be optionally substituted with up to two Rs, and each occurrence of R is an independent choice. In addition, combinations of substituents and / or variants thereof are permitted only if such combinations result in stable compounds.

[0087] When the number of a linking group is 0, such as -(CRR) 0 -, indicating that the connecting group is a single bond.

[0088] When one of the variables is selected from a single bond, it means that the two groups it connects are directly connected. For example, when L in ALZ represents a single bond, it means that the structure is actually AZ.

[0089] When the listed linking group does not indicate its linking direction, its linking direction is arbitrary, for example, The connecting group L is -MW-, in which case -MW- can connect ring A and ring B in the same direction as the reading order from left to right to form You can also connect ring A and ring B in the opposite direction of the reading order from left to right to form Combinations of linkers, substituents, and / or variations thereof are permissible only if such combinations result in stable compounds.

[0090] Unless otherwise specified, when a group has one or more connectable sites, any one or more sites of the group can be connected to other groups through chemical bonds. When the chemical bond connection mode is non-positional and there are H atoms at the connectable sites, when the chemical bonds are connected, the number of H atoms at the site will decrease with the number of connected chemical bonds to become a group with the corresponding valence. The chemical bond connecting the site to other groups can be a straight solid bond. Straight dotted key or wavy line For example, -OCH 3 The straight solid bond in the group indicates that it is connected to other groups through the oxygen atom in the group; The straight dashed bond in the group indicates that the two ends of the nitrogen atom in the group are connected to other groups; The wavy line in the phenyl group indicates that it is connected to other groups through the carbon atoms at positions 1 and 2 in the phenyl group; It means that any connectable site on the piperidine group can be connected to other groups through one chemical bond, including at least These four connection methods, even if the H atom is drawn on -N-, Still includes For groups connected in this way, when one chemical bond is connected, the H at that site will be reduced by one and become a corresponding monovalent piperidine group.

[0091] Unless otherwise specified, the term “C 1-3 "Alkyl" by itself or in combination with other terms refers to a straight or branched chain saturated hydrocarbon group consisting of 1 to 3 carbon atoms. 1-3 Alkyl groups include C 1-2 and C 2-3 Alkyl, etc.; it may be monovalent (such as methyl), divalent (such as methylene) or polyvalent (such as methine). 1-3 Examples of alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.

[0092] Unless otherwise specified, the term “C 1-5 "Alkyl" by itself or in combination with other terms refers to a straight or branched chain saturated hydrocarbon group consisting of 1 to 5 carbon atoms. 1-5 Alkyl groups include C 1-4 , C 1-3 , C 1-2 , C 2-5 , C 2-4 and C 5 Alkyl, etc.; it may be monovalent (such as methyl), divalent (such as methylene) or polyvalent (such as methine). 1-5 Examples of alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), butyl (including n-butyl, isobutyl, s-butyl and t-butyl), pentyl (including n-pentyl, isopentyl and neopentyl), and the like.

[0093] Unless otherwise specified, the term “C 1-3"Alkoxy" by itself or in combination with other terms refers to those alkyl groups containing 1 to 3 carbon atoms which are attached to the rest of the molecule via an oxygen atom. 1-3 Alkoxy includes C 1-2 , C 2-3 , C 3 and C 2 Alkoxy, etc. 1-3 Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy (including n-propoxy and isopropoxy), and the like.

[0094] Unless otherwise specified, “C 3-6 "Cycloalkyl" by itself or in combination with other terms refers to a saturated monocyclic hydrocarbon group consisting of 3 to 6 carbon atoms. 3-6 Cycloalkyl includes C 3-5 , C 4-5 and C 5-6 Cycloalkyl, etc.; it may be monovalent, divalent or polyvalent. 3-6 Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.

[0095] Unless otherwise specified, the term "3-6 membered heterocycloalkyl" by itself or in combination with other terms means a saturated monocyclic cyclic group consisting of 3 to 6 ring atoms, 1, 2, 3 or 4 of which are heteroatoms independently selected from O, S and N, and the rest are carbon atoms, wherein the nitrogen atom is optionally quaternized, and the nitrogen and sulfur heteroatoms are optionally oxidized (i.e., NO and S(O) p , p is 1 or 2). In addition, with respect to the "3-6 membered heterocycloalkyl", a heteroatom may occupy the position at which the heterocycloalkyl is attached to the rest of the molecule. The 3-6 membered heterocycloalkyl includes 4-6 membered, 5-6 membered, 4 membered, 5 membered and 6 membered heterocycloalkyl, etc. Examples of 3-6 membered heterocycloalkyl groups include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothiophenyl (including tetrahydrothiophen-2-yl and tetrahydrothiophen-3-yl, etc.), tetrahydrofuranyl (including tetrahydrofuran-2-yl, etc.), tetrahydropyranyl, piperidinyl (including 1-piperidinyl, 2-piperidinyl and 3-piperidinyl, etc.), piperazinyl (including 1-piperazinyl and 2-piperazinyl, etc.), morpholinyl (including 3-morpholinyl and 4-morpholinyl, etc.), dioxanyl, dithianyl, isoxazolidinyl, isothiazolidinyl, 1,2-oxazinyl, 1,2-thiazinyl or hexahydropyridazinyl, etc.

[0096] Unless otherwise specified, C n-n+m or C n -C n+m Any specific case including n to n+m carbons, such as C 1-12 Including C 1 , C2 , C 3 , C 4 , C 5 , C 6 , C 7 , C 8 , C 9 , C 10 , C 11 , and C 12 , also includes any range from n to n+m, such as C 1-12 Including C 1-3 , C 1-6 , C 1-9 , C 3-6 , C 3-9 , C 3-12 , C 6-9 , C 6-12 , and C 9-12 Etc.; similarly, n-membered to n+m-membered means that the number of atoms in the ring is n to n+m, for example, 3-12-membered ring includes 3-membered ring, 4-membered ring, 5-membered ring, 6-membered ring, 7-membered ring, 8-membered ring, 9-membered ring, 10-membered ring, 11-membered ring, and 12-membered ring, and also includes any range from n to n+m, for example, 3-12-membered ring includes 3-6-membered ring, 3-9-membered ring, 5-6-membered ring, 5-7-membered ring, 6-7-membered ring, 6-8-membered ring, and 6-10-membered ring, etc.

[0097] The compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthesis methods, and equivalent substitutions well known to those skilled in the art. Preferred embodiments include but are not limited to the embodiments of the present invention.

[0098] The solvent used in the present invention is commercially available.

[0099] The present invention uses the following abbreviations: aq represents water; eq represents equivalent; DCM represents dichloromethane; PE represents petroleum ether; DMSO represents dimethyl sulfoxide; EtOAc represents ethyl acetate; EtOH represents ethanol; MeOH represents methanol; DMF represents N,N-dimethylformamide; Cbz represents benzyloxycarbonyl, which is an amine protecting group; BOC represents tert-butyloxycarbonyl, which is an amine protecting group; rt represents room temperature; RT represents retention time; O / N represents overnight; THF represents tetrahydrofuran; Boc represents tert-butyloxycarbonyl, which is an amine protecting group. 2 O represents di-tert-butyl dicarbonate; TFA represents trifluoroacetic acid; HCl represents hydrochloric acid; DIPEA represents diisopropylethylamine; TEA represents triethylamine; NBS represents N-bromosuccinimide; iPrOH represents 2-propanol; mp represents melting point; Pd(PPh 3 ) 4stands for tetrakis(triphenylphosphine)palladium; Pd(dppf)Cl 2 ·CH 2 Cl 2 stands for [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride dichloromethane complex; Pd(dppf)Cl 2 stands for [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride; PPA stands for polyphosphoric acid; NMP stands for N-methylpyrrolidone; TBSCl stands for tert-butyldimethylsilyl chloride; n-BuLi stands for n-butyllithium; TBAF stands for tetrabutylammonium fluoride; psi stands for pounds force per square inch, CO 2 stands for carbon dioxide; DEA stands for diethylamine; PEG300 stands for polyethylene glycol 300; Cremphor EL stands for polyoxyethylene castor oil; PBS stands for phosphate buffered saline.

[0100] The structure of the compound of the present invention can be confirmed by conventional methods known to those skilled in the art. If the present invention relates to the absolute configuration of the compound, the absolute configuration can be confirmed by conventional technical means in the art. For example, single crystal X-ray diffraction (SXRD) is used to collect diffraction intensity data of the cultured single crystal using a Bruker D8 venture diffractometer, the light source is CuKα radiation, and the scanning mode is φ / ω scanning. After collecting relevant data, the direct method (Shelxs97) is further used to analyze the crystal structure, so that the absolute configuration can be confirmed. BRIEF DESCRIPTION OF THE DRAWINGS

[0101] Figure 1 :Experimental results of LPS-induced complement activation in PD model in mice;

[0102] Figure 2 :The results of in vivo drug efficacy model experiment in rats with passive Heimann nephritis. DETAILED DESCRIPTION

[0103] The present invention is described in detail below by way of examples, but it is not intended to impose any adverse limitations on the present invention. The present invention has been described in detail herein, and specific embodiments thereof are also disclosed therein. It will be apparent to those skilled in the art that various changes and modifications may be made to the specific embodiments of the present invention without departing from the spirit and scope of the present invention.

[0104] Reference Example 1 Preparation of Intermediates M-2 and M-1

[0105]

[0106] first step

[0107] To a solution of compound MA (5.00 g) in acetonitrile (30.0 mL) at 20°C, 4-dimethylaminopyridine (3.79 g) and di-tert-butyl dicarbonate (8.12 g) were added. The reaction solution was stirred at 20°C for 1 hour, diluted with ethyl acetate (100.0 mL), washed with 1N hydrochloric acid (30.0 mL) and saturated brine (50.0 mL*2), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound MB. 1 H NMR (400 MHz, CDCl 3 )δ7.52(d,J=4.0Hz,1H),6.87(d,J=2.4Hz,1H),6.75(d,J=2.4Hz,1H),6.48(d,J=4.0Hz,1H),3.85(s,3H),2.64(s,3H),1.65(s,9H).

[0108] Step 2

[0109] At 20°C, oxalyl chloride (5.25 g) was added to a solution of N-methylformanilide (5.59 g) in dichloromethane (50.0 mL). The reaction solution was stirred for 14 hours at 20°C under a nitrogen atmosphere. The reaction solution was transferred to a constant pressure dropping funnel and slowly added dropwise to a solution of compound MB (9.00 g) in dichloromethane (50.0 mL) at -14°C. The reaction solution was stirred for 3 hours at -14°C under a nitrogen atmosphere. Saturated sodium bicarbonate (50.0 mL) was added to quench the reaction, washed with saturated brine (50 mL*2), the organic phase was dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, the residue was washed with ethanol (20.0 mL), filtered, and the filter cake was dried to obtain compound M-2. 1 HNMR (400MHz, CDCl 3 )δ10.62(s,1H),7.62(d,J=3.6Hz,1H), 7.47(d,J=3.6Hz,1H),6.73(s,1H),3.95(s,3H),2.68(s,3H),1.63(s,9H); LC-MS: m / z=290.1[M+H] + .

[0110] Step 3

[0111] Sodium borohydride (980.7 mg) was added to a methanol (10.0 mL) solution of compound M-2 (3.00 g, 10.37 mmol, 1 eq) at 0°C, and the reaction solution was stirred at 0°C for 2 hours. After the reaction was completed, saturated ammonium chloride solution (200.0 mL) was added to the reaction solution at 0°C. It was then diluted with water (100.0 mL), and the mixture was extracted with ethyl acetate (20.0 mL*2), washed with water (50.0 mL) and saturated brine (50.0 mL), the combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 2.2:1) to obtain compound MC. 1 H NMR (400 MHz, CDCl 3 )δ7.53(d,J=3.60Hz,1H),6.74(s,1H),6.61(d,J=3.60Hz,1H),4.89(s,2H),3.90(s,3H),2.63(s,3H),1.62(s,9H).

[0112] Step 4

[0113] At 20°C, add chloromethylenedimethylammonium chloride (1.41 g) to a solution of compound MC (1.60 g) in methanol (1.2 mL) and dichloromethane (20.0 mL), and stir the reaction mixture for 1 hour. Cool to 0°C, add saturated sodium bicarbonate solution (20.0 mL) to quench, extract with ethyl acetate (30 mL*3), combine the organic phases, wash with saturated brine (20.0 mL*2), and dry with anhydrous sodium sulfate. Filter, and concentrate the filtrate under reduced pressure to obtain compound M-1.

[0114] Example 1 Preparation of Compounds 1A, 1B, 1C and 1D

[0115]

[0116]

[0117] first step

[0118] Compound 1-1 (3.20 g) was dissolved in tetrahydrofuran (30 mL), the reaction solution was placed at -78 ° C, and bis(trimethylsilyl) lithium amide (1M, 10.5 mL) was added under nitrogen protection. The reaction was stirred for 1 hour and then heated to 0 ° C, and compound 1,1,1-trifluoro-N-phenyl-N-((trifluoromethyl)sulfonyl) methanesulfonamide (5.10 g) was added. The reaction solution was heated to 30 ° C and stirred for 12 hours. Water (15 mL) was added to quench, and ethyl acetate (40 mL*3) was used for extraction. The organic phase was washed with saturated brine (40 mL*1), dried over anhydrous sodium sulfate, filtered and concentrated. The residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:0~2:1) to obtain compound 1-2. 1 H NMR (400 MHz, CDCl 3 )δ7.56-7.58(m,2H),7.28-7.38(m,6H),7.17-7.23(m,1H),5.89(br s,1H),5.01-5.19(m,3H),4.22(br s,1H),2.92-2.99(m,1H),2.58-2.73(m,1H),2.20-2.38(m,1H); LC-MS: m / z=467.0[M+H] + .

[0119] Step 2

[0120] At 20°C, 1,1-bis(diphenylphosphino)ferrocenepalladium(II) dichloromethane complex (175.1 mg) and cesium carbonate (1.05 g) were added to a toluene (10 mL) solution of compound 1-2 (0.50 g) and cyclobutylboronic acid (139.3 mg). The reaction solution was heated to 110°C and stirred for 14 hours under a nitrogen atmosphere. The reaction was cooled to room temperature, quenched by adding ice water (20 mL), extracted with ethyl acetate (25 mL*3), the organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:10) to obtain compound 1-3. 1 H NMR (400 MHz, CDCl 3 )δppm 7.30-7.67(m,9H),5.37-5.78(m,2H),5.18-5.24(m,1H),5.06-5.17(m,1H),4.17-4.38(m,1H),2.86-3.01( m,2H),2.16-2.28(m,1H),2.08-2.17(m,2H),1.88-2.02(m,4H),1.71-1.82(m,1H); LC-MS: m / z=373.1[M+H] + .

[0121] Step 3

[0122] Add barium hydroxide (1.16 g) to a mixed solution of isopropanol (2 mL), dioxane (2 mL) and water (4 mL) of compound 1-3 (0.34 g), heat the reaction solution to 100 ° C, and stir for 20 hours. The reaction solution was cooled to room temperature, adjusted to pH <7 with 50% potassium hydrogen sulfate aqueous solution, extracted with ethyl acetate (40 mL * 3), combined organic phases, washed with water (30 mL), saturated brine (25 mL) once, and dried over anhydrous sodium sulfate. Filter, and concentrate the filtrate under reduced pressure to obtain compound 1-4. LC-MS: m / z = 392.1 [M + H] + .

[0123] Step 4

[0124] At 20°C, trimethylsilyldiazomethane (2M, 919μL) was added to a mixed solution of methanol (1.2mL) and toluene (3.6mL) of compound 1-4 (0.36g), and the reaction solution was stirred at 20°C for 1 hour. The reaction solution was cooled to 0°C, quenched by adding acetic acid, diluted with water (40mL), extracted with ethyl acetate (35mL*3), and the organic phases were combined, washed with saturated sodium bicarbonate aqueous solution (25mL), water (25mL), and saturated brine (25mL) in sequence, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:3) to obtain compound 1-5. LC-MS: m / z = 406.1 [M+H] + .

[0125] Step 5

[0126] At 20°C, wet palladium carbon (10%, 0.20 g) and hydrogen chloride-dioxane solution (4M, 0.04 mL) were added to a solution of compound 1-5 (0.24 g) in ethanol (4 mL), and the reaction solution was stirred at 20°C under a hydrogen atmosphere (15 psi) for 1 hour. The reaction solution was filtered and the filtrate was concentrated under reduced pressure to obtain compound 1-6. LC-MS: m / z=274.1[M+H] + .

[0127] Step 6

[0128] Compound M-1 (1.01 g), cesium carbonate (1.64 g) and potassium iodide (334.0 mg) were added to a solution of compound 1-6 (0.55 g) in N,N-dimethylformamide (15 mL) at 20° C. The reaction solution was stirred at 20° C. for 15 hours, slowly poured into water (80 mL), extracted with ethyl acetate (90 mL*3), the organic phases were combined, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1: 1). The crude product was then separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; gradient: acetonitrile%: 57%-87%) to obtain compound 1-7, which was then subjected to chiral separation (chromatographic column: DAICEL CHIRALPAK AD (250mm*30mm, 10μm; mobile phase: A: carbon dioxide, B: [0.1% ammonia water-isopropanol]; gradient: B%: 35%-35%) to obtain compound 1-7A, compound 1-7B, compound 1-7C and compound 1-7D.

[0129] SFC analysis and detection method: Chromatographic column: Chiralpak AD-3 150mm*4.6mm ID, 3μm, mobile phase: A: carbon dioxide, B: isopropanol (0.05% diethylamine), gradient: B%: 5% to 40% gradient flow for 5 minutes, 40% for 5 minutes, 5% for 2.5 minutes, flow rate: 2.5mL / min. Compound 1-7A retention time: 4.238 minutes, ee: 100%; Compound 1-7B retention time: 4.946 minutes, ee: 100%; Compound 1-7C retention time: 5.530 minutes, ee: 99.4%; Compound 1-7D retention time: 5.954min, ee: 100%.

[0130] Two-dimensional NMR identification of compound 1-7C shows that the NOE of the carbon and hydrogen connected to the piperidine ring and the four-membered cyclobutyl group is related to the carbon and hydrogen connected to the benzoic acid, indicating a cis structure. The NOE of the carbon and hydrogen connected to the piperidine ring and the four-membered cyclobutyl group and the carbon and hydrogen connected to the benzoic acid of compound 1-7D is related to the cis structure. LC-MS: m / z = 547.2 [M+H] + .

[0131] Step 7

[0132] Lithium hydroxide monohydrate (3.5 mg) was added to a mixed solution of compound 1-7A (15.0 mg) in methanol (1 mL) and tetrahydrofuran (0.5 mL), and the reaction solution was stirred at 50°C for 18 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 16%-86%) to obtain compound 1A. LC-MS: m / z=433.2[M+H] + .

[0133] Lithium hydroxide monohydrate (4.6 mg) was added to a mixed solution of compound 1-7B (20.0 mg) in methanol (1 mL) and tetrahydrofuran (0.5 mL), and the reaction solution was stirred at 50°C for 18 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 16%-86%) to obtain compound 1B. LC-MS: m / z=433.3[M+H] + .

[0134] Lithium hydroxide monohydrate (23.0 mg) was added to a mixed solution of compound 1-7C (0.10 g) in methanol (1 mL) and tetrahydrofuran (0.5 mL), and the reaction solution was stirred at 50° C. for 15 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 16%-86%) to obtain compound 1C. 1 H NMR (400 MHz, DMSO-d 6 )δppm 10.80(s,1H),7.95(d,J=8.80Hz,2H),7.64(d,J=7.60Hz,2H),7.24(t,J=2.80Hz,1H), 6.65(s,1H),6.45(t,J=2.40Hz,1H),3.70(s,3H),3.54(d,J=12.00Hz,1H),3.10-3.30 (m,2H),2.78-2.81(m,1H),2.50-2.55(m,1H),2.41(s,3H),1.56-2.01(m,8H),1.48-1 .51(m,1H),1.34(m,1H),1.02-1.11(m,1H),0.81-0.93(m,1H); LC-MS: m / z=433.3[M+H]+ .

[0135] Lithium hydroxide monohydrate (23.0 mg) was added to a mixed solution of compound 1-7D (0.10 g) in methanol (1 mL) and tetrahydrofuran (0.5 mL), and the reaction solution was stirred at 50° C. for 15 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 16%-86%) to obtain compound 1D. 1 H NMR (400 MHz, DMSO-d 6 )δppm 10.80(s,1H),7.95(d,J=8.40Hz,2H),7.64(d,J=7.60Hz,2H),7.24(t,J=2.80Hz,1H ),6.65(s,1H),6.43-6.47(m,1H),3.70(s,3H),3.54(d,J=11.60Hz,1H),3.10-3.30 (m,2H),2.78-2.81(m,1H),2.50-2.55(m,1H),2.41(s,3H),1.54-2.01(m,8H),1.50 (m,1H),1.34(m,1H),1.02-1.11(m,1H),0.81-0.96(m,1H); LC-MS: m / z=433.2[M+H] + .

[0136] Example 2 Preparation of Compounds 2A, 2B, 2C and 2D

[0137]

[0138] first step

[0139] At 0°C, a dichloromethane solution of titanium tetrachloride (1M, 1.2 mL) was dissolved in tetrahydrofuran (1 mL), and a solution of compound 1-1 (0.20 g) and dimethyl maleate (79.0 mg) in tetrahydrofuran (1 mL) was added, and the reaction solution was stirred at 0°C for 1 hour, pyridine (236.6 mg) was added, and the reaction solution was stirred at 20°C for 15 hours. The reaction solution was poured into 10% citric acid to adjust the pH to <7, extracted with ethyl acetate (20 mL*3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:3) to obtain compound 2-2. 1 H NMR (400 MHz, CDCl 3)δppm 7.52(d,J=8.40Hz,2H),7.08-7.32(m,7H),5.23-5.42(m,1H),5.06-5.12(m,1H),4.94-5.05(m,1H),4.01-4.11(m,1H), 3.66-3.72(m,6H),3.29-3.39(m,1H),3.16-3.27(m,1H),2.74-2.90(m,2H),2.54-2.65(m,1H); LC-MS: m / z=449.1[M+H] + .

[0140] Step 2

[0141] Sodium borohydride (16.9 mg) was added to a methanol (2 mL) solution of compound 2-2 (0.20 g) at 0°C. The reaction solution was heated to 25°C and stirred for 3 hours. Ice water (30 mL) was added to quench the mixture, and the mixture was extracted with dichloromethane (30 mL*3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain compound 2-3. LC-MS: m / z=451.4[M+H] + .

[0142] Step 3

[0143] At 25°C. Sodium borohydride (923.7 mg) was added to a solution of compound 2-3 (1.10 g) in ethanol (15 mL). The reaction solution was heated to 70°C and stirred for 2 hours. The reaction solution was cooled to room temperature, quenched with ice water (50 mL), extracted with ethyl acetate (50 mL), the organic phases were combined, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:0) to obtain compound 2-4. LC-MS: m / z = 395.1 [M+H] + .

[0144] Step 4

[0145] At 0°C, sodium hydrogen (60% purity, 60.8 mg) was added to a solution of compound 2-4 (0.50 g) in tetrahydrofuran (6 mL). The reaction solution was stirred at 0°C for 0.5 hours, and methanesulfonyl chloride (0.24 g) was added. The reaction solution was heated to 25°C and stirred for 1.5 hours. Ice water (5 mL) was added to quench, and the mixture was extracted with ethyl acetate (10 mL*3). The organic phases were combined and dried over anhydrous sodium sulfate. The mixture was filtered, the filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 2:1) to obtain compound 2-5. LC-MS: m / z = 473.1 [M+H] + .

[0146] Step 5

[0147] At 0°C, sodium hydrogen (60% purity, 330.1 mg) was added to a solution of compound 2-5 (1.30 g) in tetrahydrofuran (40 mL), and the reaction solution was heated to 20°C and stirred for 1 hour. Then heated to 60°C and stirred for 16 hours. The reaction solution was cooled to room temperature, quenched by adding ice water (60 mL), extracted with ethyl acetate (10 mL*3), the organic phases were combined, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 2:1) to obtain compound 2-6. LC-MS: m / z = 377.1 [M+H] + .

[0148] Step 6

[0149] Add barium hydroxide (1.85 g) to a mixed solution of isopropanol (2 mL), dioxane (2 mL) and water (4 mL) of compound 2-6 (0.55 g), raise the reaction solution to 100 ° C, and stir for 14 hours. The reaction solution was cooled to room temperature, and a 50% potassium hydrogen sulfate aqueous solution was added to adjust the pH <7, and extracted with ethyl acetate (40 mL * 3), and the organic phases were combined, washed with water (40 mL) and saturated brine (45 mL) in turn, and dried over anhydrous sodium sulfate. Filter, and concentrate the filtrate under reduced pressure to obtain compound 2-7 (0.58 g). LC-MS: m / z = 396.1 [M + H] + .

[0150] Step 7

[0151] At 20°C, trimethylsilyldiazomethane (2 M, 1.47 mL) was added to a mixed solution of methanol (1.5 mL) and toluene (4.5 mL) of compound 2-7 (0.58 g), and the reaction solution was stirred at 20°C for 2 hours. The mixture was cooled to 0°C, quenched with acetic acid, diluted with water (40 mL), extracted with ethyl acetate (45 mL*3), combined with organic phases, washed with saturated sodium bicarbonate aqueous solution (45 mL), water (45 mL), saturated brine (50 mL), dried with anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 2:1) to obtain compound 2-8. LC-MS: m / z = 410.1 [M+H] + .

[0152] Step 8

[0153] At 20°C, wet palladium carbon (10% content, 0.2 g) was added to a solution of compound 2-8 (0.40 g) in methanol (5 mL), and the reaction solution was stirred at 20°C in a hydrogen atmosphere (15 psi) for 1 hour. The reaction solution was filtered and the filtrate was concentrated under reduced pressure to obtain compound 2-9. LC-MS: m / z=276.1[M+H] +.

[0154] Step 9

[0155] Compound M-1 (0.65 g), cesium carbonate (769.2 mg) and potassium iodide (156.8 mg) were added to a solution of compound 2-9 (0.26 g) in N,N-dimethylformamide (6 mL) at 20° C. The reaction solution was stirred at 20° C. for 16 hours. The reaction solution was slowly poured into water (50 mL), extracted with ethyl acetate (20 mL * 3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was preliminarily purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:2), and then separated and purified by preparative high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80 * 30mm * 3μm; mobile phase: [water (10mM ammonium bicarbonate) - acetonitrile]; acetonitrile%: 57%-87%) to obtain compound 2-10, which was subjected to chiral separation (column: DAICEL CHIRALPAK AD (250mm * 30mm, 10μm); mobile phase: phase A: carbon dioxide, phase B: [0.1% ammonia water - isopropanol]; B%: 40%-40%) to obtain compounds 2-10A, 2-10B, 2-10C and 2-10D.

[0156] SFC analysis and detection method: Chromatographic column: Chiralpak AD-3 150mm*4.6mm ID, 3μm, mobile phase: A: carbon dioxide, B: isopropanol (0.05% diethylamine), gradient: mobile phase B: 5% to 40% gradient flow for 5 minutes, 40% for 5 minutes, 5% for 2.5 minutes, flow rate: 2.5mL / min. Compound 2-10A retention time: 4.333 minutes, ee: 100%); Compound 2-10B retention time: 4.835min, ee: 99.5%; Compound 2-10C retention time: 5.299min, ee: 99.1%; Compound 2-10D retention time: 5.698min, ee: 98.1%. LC-MS: m / z=549.2[M+H] + .

[0157] Step 10

[0158] Lithium hydroxide monohydrate (18.4 mg) was added to a mixed solution of compound 2-10A (80.0 mg) in methanol (3 mL) and tetrahydrofuran (1.5 mL), and the reaction solution was heated to 50° C. and stirred for 14 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 15%-45%) to obtain compound 2A. 1 H NMR (400MHz, CD 3 OD)δppm 8.12(d,J=8.0Hz,1H),8.12(d,J=8.0Hz,1H),7.61(d,J=8.0Hz,2H),7.31(d,J=2.4Hz,1H),6.77(s,1H),6.33(br s,1H),4.93-4.96(m,2H),4.63(s,1H),4.44-4.80(m,2H),4.22-4.32 (m,1H),4.03-4.09(m,1H),3.77(s,3H),3.67-3.75(m,1H),3.06-3.30 (m,2H),2.51(s,3H),2.42-2.51(m,1H),2.20-2.37(m,1H),1.96-2.09(m,1H),1.72-1.81(m,1H),1.54-1.63(m,1H); LC-MS: m / z=435.2[M+H] + . Lithium hydroxide monohydrate (18.4 mg) was added to a mixed solution of compound 2-10B (80.0 mg) in methanol (3 mL) and tetrahydrofuran (1.5 mL), and the reaction solution was stirred at 50°C for 14 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by HPLC (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 15%-45%) to obtain compound 2B. Two-dimensional nuclear magnetic resonance identified that the carbon-hydrogen bond connected to the four-membered ring oxygen of piperidine was a flat bond, and the carbon-hydrogen bond connected to benzoic acid was an upright bond, and compound 2B was a trans configuration. 1 H NMR (400MHz, CD 3OD)δppm 8.13(d,J=8.4Hz,2H),7.61(d,J=8.4Hz,2H),7.30(d,J=3.2Hz,1H),6.72(s,1H),6.31(d,J=2 .8Hz,1H),4.85-4.91(m,2H),4.36-4.46(m,3H),4.27(d,J=12.8Hz,1H),4.03(d,J=12.8Hz,1 H),3.74(s,3H),3.66-3.72(m,1H),3.16-3.25(m,2H),2.50(s,3H),2.42-2.47(m,1H),2.27- 2.30(m,1H),2.03-2.10(m,1H),1.78-1.81(m,1H),1.59-1.63(m,1H); LC-MS: m / z=435.2[M+H] + .

[0159] Lithium hydroxide monohydrate (4.6 mg) was added to a mixed solution of compound 2-10C (0.02 g) in methanol (1 mL) and tetrahydrofuran (0.5 mL), and the reaction solution was stirred at 50°C for 14 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 15%-45%) to obtain compound 2C. Two-dimensional nuclear magnetic resonance identified that the carbon and hydrogen connected to the four-membered ring oxygen of piperidine and the carbon and hydrogen connected to benzoic acid were both upright bonds, and compound 2C was a cis configuration. 1 H NMR (400MHz, CD 3 OD)δppm 8.13(d,J=8.0Hz,2H),7.61(d,J=8.0Hz,2H),7.31(d,J=3.2Hz,1H),6.75(s,1H),6.32(s,1H) ,4.71-4.81(m,2H),4.47-4.56(m,2H),4.29-4.39(m,2H),3.93-3.958(m,1H),3.42-3.48(m,1 H),3.75(s,3H),3.11-3.21(m,1H),2.79-2.83(m,1H),2.51(s,3H),2.16-2.29(m,1H),1.91- 2.01(m,1H),1.79-1.85(m,1H),1.59-1.74(m,1H),1.39-1.46(m,1H); LC-MS: m / z=435.2[M+H] + .

[0160] Lithium hydroxide monohydrate (4.6 mg) was added to a mixed solution of methanol (1 mL) and tetrahydrofuran (0.5 mL) of compound 2-10D (0.02 g), and the reaction solution was stirred at 50°C for 14 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by high performance liquid chromatography (chromatographic column: Phenomenex Gemini-NX 80*30mm*3μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 15%-45%) to obtain compound 2D. Two-dimensional nuclear magnetic resonance identified that the carbon and hydrogen connected to the four-membered ring oxygen of piperidine and the carbon and hydrogen connected to benzoic acid were both upright bonds, and 2D was a cis configuration. 1 H NMR (400MHz, CD 3 OD)δppm 8.14(d,J=8.0Hz,2H),7.62(d,J=8.0Hz,2H),7.31(d,J=3.2Hz,1H),6.75(s,1H),6.32(s,1H) ,4.71-4.81(m,2H),4.46-4.57(m,2H),4.26-4.45(m,2H),3.98-4.06(m,1H),3.76(s,3H),3. 41-3.49(m,1H),3.14-3.26(m,1H),2.76-2.88(m,1H),2.51(s,3H),2.16-2.29(m,1H),1.95- 2.05(m,1H),1.78-1.87(m,1H),1.56-1.75(m,1H),1.36-1.51(m,1H); LC-MS: m / z=435.2[M+H] + .

[0161] Example 3 Preparation of Compounds 3A, 3B, 3C and 3D

[0162]

[0163]

[0164] first step

[0165] Compound 1-2 (66.0 mg) and cyclopropylboronic acid (24.3 mg) were dissolved in 1,4-dioxane (1 mL), potassium phosphate (96.1 mg) and 1,1'-bis(diphenylphosphino)ferrocenepalladium dichloromethane complex (5.8 mg) were added, and the reaction solution was stirred at 90°C under nitrogen protection for 4.5 hours. The reaction solution was concentrated under reduced pressure, quenched with water (5 mL), extracted with ethyl acetate (10 mL), and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated. The residue was separated and purified by silica gel thin layer chromatography (petroleum ether: ethyl acetate = 2:1) to obtain compound 3-3. 1H NMR (400 MHz, CDCl 3 )δ7.38-7.61(m,2H),7.08-7.33(m,7H),5.44(br s,1H),4.94-5.18(m,2H),3.97-4.24(m,1H),2.75-2.96(m,1H),2.05-2.23(m,1H),1.77-1.80(m,1H) ,1.31-1.44(m,1H),1.11-1.28(m,1H),0.53-0.68(m,2H),0.37-0.52(m,2H); LC-MS: m / z=359.2[M+H] + .

[0166] Step 2

[0167] Compound 3-3 (0.3 g) was dissolved in a mixed solution of isopropanol, 1,4-dioxane and water (2 mL: 2 mL: 5 mL), barium hydroxide (573.6 mg) was added to the reaction solution, and then heated to 100 ° C, and stirred for 12 hours. The reaction solution was cooled to room temperature, 50% potassium hydrogen sulfate aqueous solution was added to adjust the pH to 5-6, and dichloromethane (30 mL x 3) was added for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated to obtain compound 3-4. LC-MS: m / z = 378.0 [M + H] + .

[0168] Step 3

[0169] Compound 3-4 (50.0 mg) was dissolved in a mixed solution of methanol (1.5 mL) and toluene (0.5 mL). (Trimethylsilyl) diazomethane (2M, 199 μL) was added at room temperature, and the reaction solution was stirred for 12 hours. Acetic acid (1 mL) and water (10 mL) were added to quench, and the mixture was extracted with ethyl acetate (10 mL x 3). The organic phase was washed with saturated brine (10 mL x 1), dried over anhydrous sodium sulfate, filtered and concentrated. The residue was separated and purified by silica gel thin layer chromatography (petroleum ether: ethyl acetate = 3:1) to obtain compound 3-5. 1 H NMR (400 MHz, CDCl 3)δ7.95-7.97(m,2H),7.28-7.58(m,7H),5.55(br s,1H),5.03-5.24(m,2H),4.09-4.21(m,1H),3.92(s,3H),2.84-3.05(m,1H),2.11-2.19(m,1H),1.84-1.88(m,1H),1.63(br s,1H),1.37-1.53(m,1H),0.38-0.73(m,4H); LC-MS: m / z=392.2[M+H] + .

[0170] Step 4

[0171] Compound 3-5 (250.0 mg) was dissolved in a mixed solution of methanol (3 mL) and ethyl acetate (3 mL), and barium hydroxide (0.10 g) was added. The reaction solution was heated at 15°C and heated to 40°C. 2 The mixture was stirred for 2 hours under the condition of (15Psi). The reaction solution was directly filtered and concentrated under reduced pressure, and the residue was separated and purified by silica gel thin layer chromatography (petroleum ether: ethyl acetate = 1:1) to obtain compound 3-6. LC-MS: m / z = 259.9 [M+H] + .

[0172] Step 5

[0173] Compound 3-6 (0.20 g) was dissolved in 1,2-dichloroethane (5 mL), compound M-2 (133.9 mg) and sodium triacetyl cyanoborohydride (269.7 mg) were added, and the reaction solution was stirred at 15°C for 20 hours. Water (3 mL) was added to quench, extracted with ethyl acetate (10 mL x 3), the organic phase was washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered and concentrated, and the residue was separated and purified by high performance liquid chromatography (column type: Boston Green ODS 150*30mm*5μm; mobile phase: [water (0.075% trifluoroacetic acid)-acetonitrile]; B (acetonitrile)%: 43%-73%, 9min) to obtain compound 3-7. Compound 3-7 was separated and purified by SFC chiral column chromatography (column shape: DAICEL CHIRALPAK AD (250mm*30mm, 10μm); mobile phase: phase A: carbon dioxide, phase B: [0.1% ammonia water-ethanol]; B%: 35%-35%) to obtain compound 3-7A, compound 3-7B, compound 3-7C, and compound 3-7D.

[0174] SFC detection method: Chromatographic column: Chiralpak AD-3 150mm*4.6mm ID, 3μm, mobile phase: A: carbon dioxide, B: isopropanol (0.05% diethylamine), gradient: B%: 5%~40% gradient flow for 5 minutes, 40% for 5 minutes, 5% for 2.5 minutes, flow rate: 2.5mL / min. Compound 3-7A retention time: 3.781 minutes, ee: 100%; Compound 3-7B retention time: 4.455 minutes, ee: 74.9%; Compound 3-7C retention time: 5.030 minutes, ee: 100%; Compound 3-7D retention time: 5.527min, ee: 97.2%.

[0175] Step 6

[0176] At 20°C, compound 3-7A (10.0 mg) was dissolved in a mixed solution of tetrahydrofuran (0.2 mL) and methanol (0.4 mL), and then an aqueous lithium hydroxide solution (1 M, 0.2 mL) was added, and the reaction was stirred at 20°C for 36 hours. A 50% aqueous potassium hydrogen sulfate solution was added to adjust the pH to 6-7, and the mixture was extracted with ethyl acetate (10 mL), the organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated, and the residue was separated and purified by high performance liquid chromatography (column: Boston Green ODS 150*30mm*5μm; mobile phase: [water (0.075% trifluoroacetic acid)-acetonitrile]; acetonitrile%: 20%-60%) to obtain compound 3A. 1 H NMR (400MHz, CD 3 OD)δ8.24(d,J=8.2Hz,2H),7.76(d,J=8.2Hz,2H),7.35(d,J=3.2Hz,1H),6.78(s,1H),6 .39(d,J=3.0Hz,1H),4.79-4.90(m,1H),4.17-4.48(m,2H),3.77(s,3H),3.42-3.70(m, 2H),2.52(s,3H),2.21-2.39(m,1H),2.02-2.19(m,2H),1.91-1.96(m,1H),1.33-1.55( m,1H),1.03-1.12(m,1H),0.59-0.69(m,2H),0.09-0.19(m,2H); LC-MS: m / z=419.2[M+H] + .

[0177] At 20°C, compound 3-7B (20.0 mg) was dissolved in a mixed solution of tetrahydrofuran (0.3 mL) and methanol (0.6 mL), and then a lithium hydroxide aqueous solution (1 M, 0.3 mL) was added, and the reaction was stirred at 20°C for 36 hours. A 50% potassium hydrogen sulfate aqueous solution was added to adjust the pH to 6-7, and the mixture was concentrated under reduced pressure. The residue was separated and purified by high performance liquid chromatography (column: Boston Green ODS 150*30mm*5μm; mobile phase: [water (0.075% trifluoroacetic acid)-acetonitrile]; acetonitrile%: 20%-60%) to obtain compound 3B. 1 H NMR (400MHz, CD 3 OD)δ8.24(d,J=8.2Hz,2H),7.76(d,J=8.2Hz,2H),7.35(d,J=3.0Hz,1H),6.78(s,1H),6 .39(d,J=3.2Hz,1H),4.79-4.90(m,1H),4.19-4.44(m,2H),3.77(s,3H),3.44-3.71(m, 2H),2.52(s,3H),2.27-2.39(m,1H),2.03-2.17(m,2H),1.88-1.93(m,1H),1.42-1.53( m,1H),1.07-1.12(m,1H),0.62-0.73(m,2H),0.17-0.26(m,2H); LC-MS: m / z=419.2[M+H] + At 20°C, compound 3-7C (18.0 mg) was dissolved in a mixed solution of tetrahydrofuran (0.3 mL) and methanol (0.6 mL), and then lithium hydroxide monohydrate (1 M, 0.3 mL) was added, and the reaction was stirred at 20°C for 36 hours. A 50% aqueous potassium hydrogen sulfate solution was added to adjust the pH to 6-7, and the mixture was concentrated under reduced pressure. The residue was separated and purified by high performance liquid chromatography (column: Boston Green ODS 150*30mm*5μm; mobile phase: [water (0.075% trifluoroacetic acid)-acetonitrile]; acetonitrile%: 20%-60%) to obtain compound 3C. 1 H NMR (400MHz, CD 3OD)δ8.23(d,J=8.0Hz,2H),7.67-7.80(m,2H),7.33(d,J=3.0Hz,1H),6.77(s,1H),6.34(d,J=3.0H z,1H),4.45-4.55(m,1H),4.34(d,J=12.4Hz,1H),4.13(d,J=12.4Hz,1H),3.76(s,3H),3.53-3.66( m,1H),3.21-3.34(m,1H),2.51(s,3H),2.28-2.39(m,1H),1.88-2.04(m,2H),1.63-1.77(m,1H),1 .04-1.21(m,1H),0.55-0.67(m,1H),0.36-0.50(m,2H),0.07-0.28(m,2H); LC-MS: m / z=419.2[M+H] + .

[0178] At 20°C, compound 3-7D (20.0 mg) was dissolved in a mixed solution of tetrahydrofuran (0.3 mL) and methanol (0.6 mL), and then lithium hydroxide monohydrate (1 M, 0.3 mL) was added, and the reaction was stirred at 20°C for 36 hours. A 50% aqueous potassium hydrogen sulfate solution was added to adjust the pH to 6-7, and the mixture was concentrated under reduced pressure. The residue was separated and purified by high performance liquid chromatography (column: Boston Green ODS 150*30mm*5μm; mobile phase: [water (0.075% trifluoroacetic acid)-acetonitrile]; acetonitrile%: 20%-60%) to obtain compound 3D. 1 H NMR (400MHz, CD 3 OD)δ8.24(br s,2H),7.75(br s,2H),7.33(br s,1H),6.76(br s,1H),6.34(br s,1H),4.24-4.58(m,2H),4.09-4.21(m,1H),3.76(br s,3H),3.50-3.66(m,1H),2.59-2.65(m,1H),2.50(br s,3H),2.09-2.30(m,1H),1.86-2.07(m,2H),1.59-1.79(m,1H),1.02-1. 30(m,1H),0.35-0.77(m,3H),0.12-0.51(m,2H); LC-MS: m / z=419.2[M+H] + .

[0179] Example 4 Preparation of Compounds 4A and 4B

[0180]

[0181] first step

[0182] At 25°C, under nitrogen protection, potassium tert-butoxide (1.40 g) was added to a solution of trimethyl sulfoxide iodide (2.50 g) in 1,2-dichloroethane (15.0 mL). The reaction solution was stirred for 0.5 hours, the temperature was reduced to 0°C, and a solution of compound 1-1 (3.50 g) in 1,2-dichloroethane (15.0 mL) was added. The reaction temperature was raised to 25°C and stirred for 12 hours. Water (10.0 mL) was added to quench the mixture, and the mixture was extracted with ethyl acetate (50.0 mL*3). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (petroleum ether / ethyl acetate, petroleum ether ratio: 100% to 35%) to obtain compound 4-2. LC-MS: m / z=349.1[M+H] + .

[0183] Step 2

[0184] Compound 4-2 (1.70 g) was dissolved in dichloromethane (20.0 mL), and then boron trifluoride ether (1.04 g) was added, and the reaction was stirred at 25°C for 12 hours. Water (5.0 mL) was added to quench, and the mixture was extracted with dichloromethane (20.0 mL*3). The organic phase was washed with saturated brine (20.0 mL*1), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:0 to 2:1) to obtain compound 4-3. LC-MS: m / z = 349.1 [M+H] + .

[0185] Step 3

[0186] At 25°C, under nitrogen protection, sodium tert-butoxide (744.8 mg) was added to a tetrahydrofuran (15.0 mL) solution of methyl triphenylphosphonium iodide (3.13 g), and the reaction solution was stirred at 25°C for 1 hour, and a tetrahydrofuran (10.0 mL) solution of compound 4-3 (1.80 g) was added, and the reaction solution was stirred for 12 hours. Water (10.0 mL) was added to quench, and ethyl acetate (20 mL*3) was used for extraction. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated, and the residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:0 to 3:1) to obtain compound 4-4. 1 H NMR (400 MHz, CDCl 3)δ7.57-7.71(m,2H),7.29-7.47(m,7H),5.73-5.79(m,1H),5.63-5.70(m,1H),5.20(br s,2H),4.96-5.06(m,2H),4.14-4.35(m,1H),2.76-2.90(m,1H),2.36-2.39(m,1H),2.04-2. 18(m,1H),1.79-1.83(m,1H),1.64-1.72(m,1H),1.25-1.46(m,1H); LC-MS: m / z=347.1[M+H] + .

[0187] Step 4

[0188] At 20°C, under nitrogen protection, compound 4-4 (1.20 g), zinc copper reagent (4.47 g) and phosphorus oxychloride (584.3 mg) were dissolved in ethylene glycol dimethyl ether (50.0 mL), trichloroacetyl chloride (3.15 g) was added, and the reaction solution was stirred at 20°C for 12 hours. The mixture was concentrated under reduced pressure, and the residue was separated and purified by rapid silica gel column chromatography (ethyl acetate / petroleum ether, ethyl acetate ratio 0-50%) to obtain compound 4-5. LC-MS: m / z=457,459[M+H] + .

[0189] Step 5

[0190] At 20°C, compound 4-5 (1.50 g) was dissolved in dioxane (2.0 mL), and glacial acetic acid (15.75 g) and zinc powder (2.14 g) were added. After the addition was completed, the temperature was raised to 80°C and stirred for 1 hour. The reaction solution was filtered and the filtrate was concentrated under reduced pressure. The residue was separated and purified by rapid silica gel column chromatography (ethyl acetate / petroleum ether, ethyl acetate ratio 0-50%) to obtain compound 4-6. 1 H NMR (400 MHz, CDCl 3 )δ:7.63(d,J=8.4Hz,2H),7.21-7.48(m,7H),5.64(br s,1H),5.19(br s,2H),4.28(br s,1H),3.03-3.20(m,2H),2.64-2.87(m,3H),2.30-2.43(m,1H),2.02-2.19(m,1H),1.67(m,2H),1.21-1.41(m,2H).

[0191] Step 6

[0192] At 15°C, compound 4-6 (1.2 g) was dissolved in dichloromethane (30.0 mL), the temperature was lowered to -15°C, diethylaminosulfur trifluoride (2.49 g) was added, the temperature was slowly raised to 15°C, and stirring was continued for 12 hours. The reaction solution was slowly poured into ice water (100.0 mL) for quenching, the pH was adjusted to 7 with saturated sodium bicarbonate aqueous solution, extracted with dichloromethane (100 mL*3), the organic phases were combined, and concentrated under reduced pressure. The residue was separated and purified by rapid silica gel column chromatography (ethyl acetate / petroleum ether, ethyl acetate ratio 0-50%) to obtain compound 4-7. 1 H NMR (400 MHz, CDCl 3 )δ7.64(d,J=8.4Hz,2H),7.10-7.50(m,7H),5.62(br s,1H),5.19(br s,2H),4.24(br s,1H),2.74-2.77(m,1H),2.52-2.72(m,2H),2.20-2.35(m,1H),2.06-2.22(m,2H),1.78- 1.91(m,1H),1.59-1.68(m,1H),1.50-1.57(m,1H),1.23-1.39(m,1H),1.07-1.21(m,1H).

[0193] Step 7

[0194] At 20°C, compound 4-7 (1.00 g) was dissolved in a mixed solution of isopropanol (5.0 mL), dioxane (5.0 mL) and water (10.0 mL), barium hydroxide (960.1 mg) was added, the temperature was raised to 100°C, and the mixture was stirred for 12 hours. The reaction solution was cooled to room temperature, poured into water (100.0 mL) for quenching, and the pH was adjusted to 5-6 with 1M hydrochloric acid. It was extracted with ethyl acetate (100 mL*3), the organic phases were combined, and the mixture was concentrated under reduced pressure to obtain compound 4-8. LC-MS: m / z=430[M+H] + .

[0195] Step 8

[0196] At 20°C, compound 4-8 (0.98 g) was dissolved in a mixed solution of methanol (5.0 mL) and toluene (15.0 mL), and then trimethylsilyldiazomethane (2M, 2.45 mL) was slowly added, and the reaction was stirred at 20°C for 0.5 hours. Acetic acid (2.0 mL) was slowly added dropwise to quench, and then concentrated under reduced pressure. The concentrate was separated and purified by rapid silica gel column chromatography (ethyl acetate / petroleum ether, ethyl acetate ratio 0-50%) to obtain compound 4-9. The results of two-dimensional nuclear magnetic spectrum (HSQC, COSY, NOE) analysis showed that the CH connected to piperidine and difluorocyclobutane was an upright bond, and there was NOE correlation with the two hydrogens at the α position of the benzene ring, so it was judged that the relative configuration of difluorocyclobutane and methyl benzoate was trans configuration. 1 H NMR (400 MHz, CDCl 3 )δ8.04(br d,J=8.4Hz,2H),7.10-7.50(m,7H),5.64(br s,1H),5.21(s,2H),4.27(br s,1H),3.94(s,3H),2.77-2.92(m,1H),2.51-2.76(m,2H),2.27-2.41(m,1H),2.08-2.26(m,2H), 1.73-1.92(m,1H),1.47-1.69(m,2H),1.30-1.45(m,1H),1.08-1.23(m,1H); LC-MS: m / z=444[M+H] + .

[0197] Step 9

[0198] At 25°C, compound 4-9 (900.0 mg) was dissolved in ethyl acetate (15.0 mL), and then a solution of palladium hydroxide (180.0 mg) in methanol (5.0 mL) was added. The system was replaced with hydrogen three times, and the hydrogen pressure was maintained at 15 psi, and the mixture was stirred for 2 hours. The mixture was filtered and the filtrate was concentrated under reduced pressure to obtain compound 4-10. 1 H NMR (400 MHz, CDCl 3 )δ8.00(d,J=8.4Hz,2H),7.45(d,J=8.4Hz,2H),3.92-3.98(m,1H),3.91(s,3H),2.953-2.96(m,2H),2.64-2.76(m,2H),2.38-2. 52(m,1H),2.05-2.25(m,2H),1.80-1.90(m,1H),1.70-1.79(m,2H),1.61-1.68(m,1H),1.43-1.52(m,1H); LC-MS: m / z=310[M+H] + .

[0199] Step 10

[0200] At 25°C, compound 4-10 (70.0 mg) and compound M-1 (180.0 mg) were dissolved in N,N-dimethylformamide (3.0 mL), and then cesium carbonate (221.2 mg) and potassium iodide (37.6 mg, 226.28 μmol, 1 eq.) were added, and the reaction solution was stirred for 12 hours. The reaction solution was poured into water (100 mL) for quenching, extracted with ethyl acetate (70 mL*3), and the organic phases were combined and concentrated under reduced pressure. The residue was separated and purified by silica gel thin layer chromatography (petroleum ether: ethyl acetate = 3:1) to obtain compound 4-11, and then subjected to chiral separation (chiral column: Phenomenex-Cellulose-2 (250mm*30mm, 10μm)); mobile phase: [A: carbon dioxide, B: ethanol (0.1% ammonia water)]; gradient B%: 20%~20%) to obtain compound 4-11A and compound 4-11B.

[0201] SFC analysis and detection method: Column: Cellulose 2 150mm*4.6mm ID, 5μm, Mobile phase: A: Carbon dioxide, B: Ethanol (0.05% diethylamine), Gradient: B%: 5% to 40% gradient flow for 5 minutes, 40% for 5 minutes, 5% for 2.5 minutes, Flow rate: 2.5mL / min. Retention time of compound 4-11A: 3.267min, retention time of compound 4-11B: 3.514min. LC-MS: m / z=583[M+H] + .

[0202] Step 11

[0203] Compound 4A:

[0204] At 20°C, compound 4-11A (50.0 mg) was dissolved in a mixed solution of tetrahydrofuran (1.0 mL), methanol (2.0 mL) and water (1.0 mL), and then lithium hydroxide (40.0 mg) was added. The reaction solution was heated to 50°C and stirred for 12 hours. The reaction solution was poured into water (20.0 mL), the pH was adjusted to 4-5 with 4 M acetic acid aqueous solution, extracted with ethyl acetate (30 mL*3), the organic phase was collected, washed with saturated brine (20.0 mL), dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was dissolved in a mixed solvent of water (10.0 mL) and acetonitrile (2.0 mL), and freeze-dried to obtain compound 4A (SFC detection method: chromatographic column: Chiralpak AD-3 150 mm*4.6 mm ID, 3 μm, mobile phase: A: carbon dioxide, B: isopropanol (0.05% diethylamine), gradient: B%: 5%-40% gradient flow for 5.5 minutes, 40% maintained for 3 minutes, 5% maintained for 1.5 minutes, flow rate: 2.5 mL / min, retention time: 6.521 minutes, ee: 99.7%). 1 H NMR (400 MHz, DMSO-d 6 )δ:10.80(br s,1H),7.89(d,J=8.4Hz,2H),7.52(br d,J=8.0Hz,2H),7.24(t,J=2.8Hz,1H),6.64(s,1H),6.48(t,J=2.4Hz,1H),3.70(s,3H),3.50-3.60(m,2H),2.67-2.77(m,2H),2 .41(s,3H),2.08-2.25(m,4H),1.71-1.80(m,1H),1.58-1.68(m,1H),1.48-1.58(m,3H),1.34-1.37(m,2H); LC-MS: m / z=469[M+H] + .

[0205] Compound 4B:

[0206] Compound 4-11B (46.0 mg) was dissolved in a mixed solution of tetrahydrofuran (1.0 mL), methanol (2.0 mL) and water (1.0 mL) at 20°C, and then lithium hydroxide (40.0 mg) was added. The reaction solution was heated to 50°C and stirred for 12 hours. The reaction solution was poured into water (20.0 mL), the pH was adjusted to 4-5 with 4 M acetic acid aqueous solution, extracted with ethyl acetate (30 mL*3), the organic phase was collected, washed with saturated brine (20.0 mL), dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was dissolved in a mixed solvent of water (10.0 mL) and acetonitrile (2.0 mL), and freeze-dried to obtain compound 4B (SFC detection method: chromatographic column: Chiralpak AD-3 150 mm*4.6 mm ID, 3 μm, mobile phase: A: carbon dioxide, B: isopropanol (0.05% diethylamine), gradient: B%: 5%-40% gradient flow for 5.5 minutes, 40% maintained for 3 minutes, 5% maintained for 1.5 minutes, flow rate: 2.5 mL / min, retention time: 4.832 minutes, ee: 96.0%). 1 H NMR (400 MHz, DMSO-d 6 )δ:10.80(br s,1H),7.93(d,J=8.0Hz,2H),7.61(br d,J=7.4Hz,2H),7.24(t,J=2.8Hz,1H),6.64(s,1H),6.48(t,J=2.0Hz,1H),3.70(s,3H),3.45 -3.65(m,2H),2.67-2.78(m,2H),2.41(s,3H),2.11-2.28(m,4H),1.71-1.80(m,1H),1.62(br s,1H),1.45-1.58(m,3H),1.30-1.40(m,2H); LC-MS: m / z=469[M+H] + .

[0207] Example 5 Preparation of Compound 4C

[0208]

[0209]

[0210] first step

[0211] At 25°C, compound 5-1 (1.00 kg) and compound 5-2 (919.93 g) were dissolved in 2-methyltetrahydrofuran (10.0 L), and tetraethoxytitanium (4.81 kg) was added in batches. The reaction solution was heated to 80°C under nitrogen protection and stirred for 16 hours. The reaction solution was poured into ice water (15.0 L), filtered, and the filter cake was washed with 2-methyltetrahydrofuran (2.0 L*3). The filtrate was collected, washed with saturated brine (3.0 L), dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove 2-methyltetrahydrofuran (8.0 L). The remaining solution was poured into petroleum ether (10.0 L) at 0°C, stirred for 1 hour, filtered, and the filter cake was washed with petroleum ether (1.0 L*3), and then dried under reduced pressure to obtain compound 5-3. 1 H NMR (400 MHz, CDCl 3 )δ:7.96-7.98(m,2H),7.73-7.75(m,2H),2.81(s,3H),1.34(s,9H).

[0212] Step 2

[0213] At -20°C, under nitrogen protection, lithium diisopropylamide (2M tetrahydrofuran solution, 12.08 mL) was added dropwise to a solution of compound 5-3 (5.00 g) in 2-methyltetrahydrofuran (100.0 mL). After the addition was complete, the reaction solution was stirred at -20°C to -10°C for 0.5 hours, and a solution of compound 5-4 (5.74 g) in 2-methyltetrahydrofuran (25.0 mL) was added, the temperature was slowly raised, and the mixture was stirred at -20 to -10°C for 7 hours. Saturated aqueous ammonium chloride solution (30.00 mL) was added to quench the reaction, and the mixture was diluted with water (200.0 mL) and extracted with ethyl acetate (100.0 mL*2). The organic phases were combined, washed with saturated brine (100.0 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate, ethyl acetate ratio: 0 to 20%) to obtain compound 5-5. LC-MS: m / z = 439.1

[0214] Step 3

[0215] At 0°C, compound 5-5 (5.00 g) was dissolved in tetrahydrofuran (50.0 mL), sodium borohydride (0.62 g) was added, and the reaction was stirred at 0°C for 3 hours. Water (30.0 mL) was added to quench, and the mixture was extracted with ethyl acetate (30.0 mL*2). The organic phase was washed with saturated brine (30.0 mL*1), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate, ethyl acetate ratio: 0-100%) to obtain compound 5-6. LC-MS: m / z=399.2[M+H] + .

[0216] Step 4

[0217] At 25°C, under nitrogen protection, triphenylphosphine (1.18 g) and diisopropyl azodicarboxylate (913.3 mg) were added to a solution of compound 5-6 (600.0 mg) in tetrahydrofuran (15.0 mL). The reaction solution was stirred at 25°C for 18 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 0-50%) to obtain compound 5-7. LC-MS: m / z = 381.1 [M+H] + .

[0218] Step 5

[0219] At 20°C, under nitrogen protection, compound 5-7 (200.0 mg) was dissolved in methanol (1.0 mL) and concentrated H 2 SO 4 (1.0mL). The reaction solution was heated to 80°C and stirred for 3 hours. The reaction solution was cooled to room temperature, diluted with water (50.0mL), adjusted to pH 7-8 with saturated sodium bicarbonate aqueous solution, and extracted with ethyl acetate (50.0mLx2). The organic phases were combined, washed with saturated brine (50.0mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel thin layer chromatography (dichloromethane: methanol, methanol ratio: 0-10%) to obtain compound 5-8. LC-MS: m / z=310.2[M+H] + .

[0220] Step 6

[0221] At 25°C, sodium borohydride acetate (137.0 mg) was added to 1,2-dichloroethane (1.0 mL) of compound 5-8 (50.0 mg) and compound M-2 (51.4 mg). The mixture was stirred at 25°C for 16 hours. Diluted with water (200.0 mL), extracted with ethyl acetate (50.0 mL*2), combined organic phases, washed with water (100.0 mL*2), dried with anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure and separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate, ethyl acetate ratio: 0-15%) to obtain compound 5-9. LC-MS: m / z=583.4[M+H] + .

[0222] Step 7

[0223] At 25°C, compound 5-9 (30.0 mg) was dissolved in tetrahydrofuran (0.5 mL) and methanol (0.5 mL), and lithium hydroxide monohydrate (13.0 mg) was added, and the reaction solution was stirred at 50°C for 16 hours. The reaction solution was cooled to room temperature, quenched by adding acetic acid (0.1 mL), diluted with water (50.0 mL), extracted with ethyl acetate (25.0 mL*2), and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and dried under reduced pressure to obtain compound 4C.

[0224] LC-MS: m / z=469.1[M+H] + .

[0225] Example 6 Preparation of Compound 4D

[0226]

[0227]

[0228] first step

[0229] At 25°C, compound 5-1 (50.00 g) and compound 6-2 (45.92 g) were dissolved in 2-methyltetrahydrofuran (500.0 mL), and tetraethoxytitanium (235.72 g) was added in batches. The reaction solution was heated to 80°C under nitrogen protection and stirred for 16 hours. The reaction solution was poured into ice water (15.0 L), filtered, the filter cake was washed with 2-methyltetrahydrofuran (3.0 L), the filtrate was collected, dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove 2-methyltetrahydrofuran (400.0 mL). The remaining solution was poured into petroleum ether (200.0 mL) at 0°C and stirred for 1 hour, filtered, the filter cake was collected, and dried under reduced pressure to obtain compound 6-3. 1 H NMR (400 MHz, CDCl 3 )δ:7.95-7.97(d,J=8.0Hz,2H),7.72-7.74(d,J=8.0Hz,2H),2.79(s,3H),1.33(s,9H).

[0230] Step 2

[0231] At -20°C, under nitrogen protection, lithium diisopropylamide (2M tetrahydrofuran solution, 60.40 mL) was added dropwise to a solution of compound 6-3 (25.00 g) in 2-methyltetrahydrofuran (500.0 mL). After the addition was complete, the reaction solution was stirred at -20°C to -15°C for 1 hour, a solution of compound 5-4 (35.35 g) in 2-methyltetrahydrofuran (125.0 mL) was added, the temperature was slowly raised, and the mixture was stirred at -20 to -15°C for 4 hours. A saturated aqueous ammonium chloride solution (60.00 mL) was added to quench the reaction, the mixture was diluted with water (300.0 mL), and the mixture was extracted with ethyl acetate (200.0 mL*2). The organic phases were combined, washed with water (200.0 mL*2), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (petroleum ether:ethyl acetate, ethyl acetate ratio: 0-13%) to obtain compound 6-5.

[0232] Step 3

[0233] At 0°C, compound 6-5 (5.20 g) was dissolved in tetrahydrofuran (52.0 mL), sodium borohydride (0.50 g) was added, and the reaction was stirred at 0°C for 1 hour. Saturated ammonium chloride solution (50.0 mL) was added to quench, and water (200.0 mL) was added to dilute, and extracted with ethyl acetate (100.0 mL*2), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate, ethyl acetate ratio: 0-33%) to obtain compound 6-6. LC-MS: m / z=441.1[M+H] + .

[0234] Step 4

[0235] At -15°C, compound 6-6 (3.30 g) was dissolved in tetrahydrofuran (15.0 mL), and lithium tri-sec-butyl borohydride (1M tetrahydrofuran solution, 5.02 mL) was added. The reaction was stirred at -15°C for 4 hours. Saturated ammonium chloride solution (50.0 mL) was added to quench, and water (200.0 mL) was added to dilute, and extracted with ethyl acetate (50.0 mL*3), and the organic phases were combined, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (dichloromethane: methanol, methanol ratio: 0-5%) to obtain compound 6-7. LC-MS: m / z=399.2[M+H] + .

[0236] Step 5

[0237] Triphenylphosphine (5.30 g) and diisopropyl azodicarboxylate (4.08 g) were added to a solution of compound 6-7 (2.30 g) in tetrahydrofuran (46.0 mL) at 25°C. The reaction solution was stirred at 25°C under nitrogen for 16 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate, ethyl acetate ratio: 0-40%) to obtain compound 6-8. LC-MS: m / z=381.1[M+H] + .

[0238] Step 6

[0239] At 20°C, under nitrogen protection, compound 6-8 (500.0 mg) was dissolved in methanol (5.0 mL) and concentrated H 2 SO 4 (5.0mL). The reaction solution was heated to 80°C and stirred for 16 hours. The reaction solution was cooled to room temperature, the pH was adjusted to 7-8 with saturated sodium bicarbonate aqueous solution, and extracted with ethyl acetate (50.0mL*2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by silica gel thin layer chromatography (dichloromethane: methanol = 0-10%) to obtain compound 6-9. LC-MS: m / z = 310.1 [M+H] + .

[0240] Step 7

[0241] At 25°C, sodium borohydride acetate (356.3 mg) was added to 1,2-dichloroethane (3.0 mL) of compound 6-9 (130.0 mg) and compound M-2 (133.7 mg). The mixture was stirred at 25°C for 16 hours. Water (200.0 mL) was added for dilution, and ethyl acetate (50.0 mL*2) was used for extraction. The organic phases were combined, washed with water (100.0 mL*2), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The filtrate was separated and purified by silica gel column chromatography (petroleum ether: ethyl acetate, ethyl acetate ratio: 0-15%) to obtain compound 6-10. LC-MS: m / z=583.4[M+H] + .

[0242] Step 8

[0243] At 25°C, compound 6-10 (30.0 mg) was dissolved in tetrahydrofuran (0.5 mL) and methanol (0.5 mL), and lithium hydroxide monohydrate (13.0 mg) was added. The reaction solution was stirred at 50°C for 16 hours. The reaction solution was cooled to room temperature, quenched by adding acetic acid (0.1 mL), diluted with water (50.0 mL), extracted with ethyl acetate (25.0 mL*2), and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and dried under reduced pressure to obtain compound 4D. LC-MS: m / z=469.1[M+H]+ .

[0244] Biological test data:

[0245] Experimental Example 1 Inhibition of Wieslab complement alternative pathway activation (enzyme activity test)

[0246] The purpose of this experiment is to The complement system alternative pathway kit is used to measure the inhibitory activity of the compounds of the present invention against the complement alternative pathway in human serum.

[0247] Experimental protocol:

[0248] Dilute the serum with diluent (1:23). Add the drug to the diluted serum, 8 concentration gradients, up to 10mM or 50mM, 5-fold gradient dilution. Incubate at room temperature for 15 minutes. Add the compound and serum mixture to the 96-well plate provided by the kit (100μL / well) and activate at 37 degrees for 1 hour. Wash three times with washing buffer. Add the detection antibody provided by the kit (100μL) and incubate at room temperature for 30 minutes. Wash three times with washing buffer. Add the substrate (100μL) and incubate at room temperature for 30 minutes. Detect the absorbance at 405nM with an enzyme reader.

[0249] Experimental results: The results of the inhibitory activity of the test compounds on the complement alternative pathway are shown in Table 1.

[0250] Table 1: In vitro enzyme activity screening test results

[0251] Compound <![CDATA[AP MAC IC 50 (nM)]]> Compound 3A 23.6 Compound 3C 51.7 Compound 4B 40.7 LNP023 38.0*

[0252] Note: *Average of two tests

[0253] Conclusion: The compounds of the present invention have significant inhibitory activity on activation of human serum bypass pathway.

[0254] Experimental Example 2: Pharmacokinetic Study in Mice

[0255] Experimental purpose: To investigate the plasma pharmacokinetics of the compound of the present invention in male C57BL / 6J mice after single intravenous injection and oral administration.

[0256] Experimental animals: male C57BL / 6J mice, 7-9 weeks old, weighing 17-23 g; Supplier: Shanghai Xipu-Bikai Experimental Animal Co., Ltd.

[0257] Experimental procedure: Injection (IV): dose 1 mg / kg (solvent: 10% PG / 5% Solutol / 85% PBS (1× pH 7.4)); Oral administration (PO): dose 10 mg / kg (solvent: 30% PEG300 / 10% Cremphor EL / 60% PBS (1× pH 7.4))

[0258] Sample collection: 0.03 mL of blood samples were collected from saphenous vein puncture at each time point of the experimental animals, and the actual blood collection time was recorded. All blood samples were added to commercial EDTA-K2 anticoagulant tubes with a specification of 1.5 mL (supplier: Jiangsu Kangjian Medical Supplies Co., Ltd.). After blood sample collection, centrifuge at 4°C and 3000g for 10 minutes within half an hour to draw supernatant plasma, quickly put it into dry ice, and store it in a -80°C refrigerator for LC-MS / MS analysis.

[0259] Data analysis: WinNonlin TM The non-compartmental model of the pharmacokinetic software Version 6.3 (Pharsight, Mountain View, CA) was used to process plasma concentrations, and the pharmacokinetic parameters Cl were calculated using the linear logarithmic trapezoidal method. , T 1 / 2 ,C max ,AUC 0-last , the results are shown in Table 2.

[0260] Table 2 Comparison of PK results between the compounds of the present invention and reference compounds

[0261]

[0262]

[0263] Note: Cl: apparent clearance; T 1 / 2 : The time required to remove half of the compound; C max : Peak concentration; AUC 0-last : 0-integrated area of ​​concentration within the last sampling time.

[0264] Experimental conclusion: The results of the pharmacokinetic study of the compound of the present invention in mice showed that the half-life was longer than that of the reference compound, and the oral plasma exposure was significantly better than that of the reference compound LNP023. The compound of the present invention has excellent pharmacokinetic properties.

[0265] Experimental Example 3: Pharmacokinetic Study in Rats

[0266] Experimental purpose: To investigate the plasma pharmacokinetics of the compound of the present invention in male Wistar Han rats after single intravenous injection and oral administration.

[0267] Experimental animals: Male Wistar Han rats, 7-9 weeks old, weighing 231.03-243.2 g; Supplier: Beijing Weitonglihua Laboratory Animal Technology Co., Ltd.

[0268] Experimental procedure: Injection (IV): 1 mpk (solvent: 30% PEG300 / 10% Cremphor EL / 60% PBS (1× pH 7.4)); Oral administration (PO): 5 mpk or 30 mpk (solvent: 30% PEG300 / 10% Cremphor EL / 60% PBS (1× pH 7.4))

[0269] Sample collection: About 0.3 mL of blood samples were collected from the jugular vein of the experimental animals at each time point, and the actual blood collection time was recorded. All blood samples were added to a commercial EDTA-K2 anticoagulant tube with a specification of 1.5 mL (supplier: Jiangsu Kangjian Medical Supplies Co., Ltd.). After blood sample collection, centrifuge at 4°C and 3200g for 10 minutes to draw the supernatant plasma, quickly put it into dry ice, and store it in a -80°C refrigerator for LC-MS / MS analysis.

[0270] Data analysis: The non-compartmental model of Phoenix WinNonlin 6.3 pharmacokinetic software was used to process plasma concentrations, and the pharmacokinetic parameters Cl, C were calculated using the linear logarithmic trapezoidal method. max , T 1 / 2 , AUC 0-last , the results are shown in Table 3.

[0271] Table 3 Comparison of PK results of the compounds of the present invention and reference compounds in rats

[0272]

[0273] Note: Cl: apparent clearance; T 1 / 2 : The time required to remove half of the compound; C max : Peak concentration (the table shows dose-normalized data); AUC 0-last : 0-integrated area of ​​concentration within the last sampling time (the table shows dose-normalized data); a: the administration dose is 5 mpk; b: the administration dose is 30 mpk.

[0274] Experimental conclusion: The results of the pharmacokinetic study of the compound of the present invention in rats showed that the half-life was longer than that of the reference compound, and the oral plasma exposure was significantly better than that of the reference compound LNP023. The compound of the present invention has excellent pharmacokinetic properties.

[0275] Experimental Example 4: LPS-induced complement activation in vivo PD model in mice

[0276] Experimental purpose: To investigate the inhibitory effect of the compounds of the present invention on complement activation induced by LPS stimulation in mice.

[0277] Experimental animals: female C57BL / 6J mice, 7-9 weeks old, weighing 17-23 g; Supplier: Shanghai Xipu-Bikai Experimental Animal Co., Ltd.

[0278] Experimental procedure: 100 μg of lipopolysaccharide (LPS) from Salmonella typhimurium (Sigma) dissolved in 100 μL sterile PBS was injected intraperitoneally to induce complement activation in mice. Negative control animals received intraperitoneal injections of 100 μL sterile PBS and were administered alone by gavage (PO). Positive control animals received intraperitoneal LPS and PO drug vehicle (0.5% (w / v) methylcellulose and 0.5% (v / v) Tween 80). Dosing and plasma collection time points are as follows:

[0279]

[0280] Sample collection: 0.3 mL of blood sample was collected from the orbital venous plexus. All blood samples were added to a commercial EDTA-K2 anticoagulant tube with a specification of 1.5 mL (supplier: Jiangsu Kangjian Medical Supplies Co., Ltd.). Within half an hour after blood sample collection, centrifuge at 4°C and 3000g for 10 minutes to draw the supernatant plasma, quickly put it into dry ice, and store it in a refrigerator at -80°C for Western blot analysis of downstream C3d protein levels after complement activation.

[0281] Sample analysis: mouse plasma (5 μL) + Lysis buffer (lysate, 27.5 μL) + Loading buffer (loading buffer, 12.5 μL) + Reducing buffer (reducing buffer, 5 μL) were mixed and incubated at 100°C for 15 min. The loading volume was 5 μL / well, i.e., the plasma loading volume in each well was 0.5 μL.

[0282] Experimental results: In this experiment, Novartis' Factor B inhibitor LNP023 was used as the reference drug, and the test compounds were the reference compound LNP023, compound 3A and compound 3C. All three drugs can significantly inhibit the level of C3d, that is, they can inhibit LPS-induced complement activation, and the inhibitory effects of compound 3A and compound 3C are better than LNP023. Specific experimental results are attached Figure 1 , Data are expressed as: Mean±SEM, n=5; ###, p<0.001, normal group vs. model group, t-test; ****, p<0.0001, drug administration group vs. model group, one-way ANOVA.

[0283] Experimental conclusion: The compounds of the present invention can significantly inhibit complement activation stimulated by LPS, and the inhibitory effect is better than that of LNP023.

[0284] Experimental Example 5: In vivo drug efficacy model in rats with passive Heimann nephritis

[0285] Experimental purpose: To investigate the ability of the compounds of the present invention to improve the renal function of rats with Heimann nephritis induced by Sheep Anti-Rat Fx1A Serum, including the evaluation of reducing proteinuria levels and improving renal tissue damage.

[0286] Experimental animals: male SD rats, 7-10 weeks old, weighing 200-300 g; Supplier: Beijing Weitonglihua Experimental Animal Technology Co., Ltd.

[0287] Experimental process:

[0288] 1. Modeling:

[0289] On D-2 days before administration, the rat urine was collected. D1 was the first day of the experiment, and the animals in the control group (Group 1) were given a single injection of 5 mL / kg of Sheep Non-Immune serum through the tail vein; the animals in the modeling group and the drug administration group (Groups 2 to 6) were given a single injection of 5 mL / kg of Sheep Anti-Rat Fx1A Serum through the tail vein.

[0290] 2. Administration:

[0291] The administration method was oral gavage, twice a day, with an interval of 8 hours, and the administration volume was 10mL / kg. On D1 day, 1 hour before modeling, animals in Group 1 and Group 2 were given blank solvent 20% PEG 400 / 10% Solutol / 70% water; animals in Group 3 were given LNP023 (60mpk); Groups 4 to 6 were given different concentrations of compound 4B (5mpk, 20mpk and 60mpk); 8 hours after the first administration, each group was administered again, and the dosage and volume were the same as the first time. From D2 to D14 days, animals in each group were given different compounds or solvents for 14 consecutive days (including Day 1) according to the dosage, volume, administration method and frequency on D1.

[0292] 3. Sample collection:

[0293] Urine collection: Rat urine samples were collected on D-2 days before administration, and 2-4h and 4-6h after the first administration on D4, D6, D8, D11, and D14 days after administration, and transferred to EP tubes and stored in a refrigerator at -80℃ to -60℃ for rat urine protein and urine creatinine detection.

[0294] Kidney sample collection: All animals were anesthetized with CO on Day 15. 2 The rats were euthanized by inhalation anesthesia, and the kidneys were collected from both sides. The left kidney was cut transversely, and the right kidney was cut longitudinally. The transverse half (left side) and the longitudinal half (right side) of the kidney were fixed in formalin (placed in the same EP tube).

[0295] 4. Sample analysis:

[0296] (1) Rat urine creatinine uCRE is normally loaded after ten-fold dilution (Decrease mode), 10 μl sample + 90 μl saline (0.9% saline). If it exceeds the detection limit, it is diluted 20 times (Increase mode), 5 μl sample + 95 μl saline (0.9% saline). Rat urine total protein uTP is detected at the original multiple. If it exceeds the detection limit, it is first diluted 10 times (Decrease mode), 10 μl sample + 90 μl saline (0.9% saline). If it still exceeds the detection limit after ten-fold dilution, it is diluted 100 times (Increase mode), 2 μl sample + 198 μl saline (0.9% saline); the data is read by the analytical instrument HITACHI LST008AS (P).

[0297] (2) Renal pathology scoring: HE staining was used to analyze the degree of drug damage to rat kidneys (paraffin sections). Scoring criteria: 0 indicates normal, 1 indicates a few cell infiltrations in the mesangium, 2 indicates more cell infiltrations in the mesangium, 3 indicates hyperplasia of several glomerular mesangial cells and infiltration of several mesangial cells, and 4 indicates tubular casts, atrophy, glomerular crescent formation, and sclerosis.

[0298] Experimental results: The urine protein and urine creatinine of rats were detected using Novartis' Factor B inhibitor LNP023 as the reference drug. Both LNP023 and compound 4B were able to significantly inhibit the urine protein level of rats and improve the renal function of rats. The efficacy of compound 4B was better than that of LNP023 at a dose of 60 mpk. See the attached for specific experimental results. Figure 2 , Data are expressed as: Mean±SEM, n=5; ###, p<0.001, normal group vs. model group, t-test; ****, p<0.0001, drug administration group vs. model group, one-way ANOVA.

[0299] The pathological scoring results are shown in Table 4:

[0300] Table 4 Pathological scoring results

[0301]

[0302] Experimental conclusion: According to the pathological scoring results, compared with the model group, the compounds of the present invention in different dose groups can significantly improve the degree of renal lesions in the model animals. The compounds of the present invention can reduce the urine protein level of rats, improve renal function, and have better efficacy than LNP023.

Claims

1. A compound represented by formula (I) or a pharmaceutically acceptable salt thereof, in, L is a single bond; R 1 Selected from C 3-6 Cycloalkyl and -C 1-3 Alkyl-C 3-6 Cycloalkyl, the C 3-6 Cycloalkyl and -C 1-3 Alkyl-C 3-6 The cycloalkyl group is optionally substituted by 1, 2 or 3 R a replace; R 2 and R 3 are independently selected from H and C 1-5 Alkyl, the C 1-5 The alkyl group is optionally substituted with 1, 2 or 3 R b replace; Each R a and R b are independently selected from H, F, Cl, Br and I.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, which is selected from: in, L, R 1 , R 2 and R 3 As defined in claim 1; The carbon atom with "*" is a chiral carbon atom, which exists in the form of a single enantiomer (R) or (S) or in the form enriched in one enantiomer.

3. The compound according to claim 2 or a pharmaceutically acceptable salt thereof, which is selected from: in, L, R 1 , R 2 and R 3 As defined in claim 2.

4. The compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, in, R 1 Selected from Said are independently optionally substituted by 1, 2 or 3 R a replace.

5. The compound according to claim 4 or a pharmaceutically acceptable salt thereof, in, R 1 Selected from 6. The compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, in, R 2 Selected from H and CH 3 , the CH 3 Optional 1, 2 or 3 R b replace.

7. The compound according to claim 6 or a pharmaceutically acceptable salt thereof, in, R 2 Selected from H, CH 3 CF 3 and CHF 2 .

8. The compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, in, R 3 Selected from H and CH 3 .

9. The compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, in, Structural unit Selected from 10. The compound according to claim 9 or a pharmaceutically acceptable salt thereof, in, Structural unit Selected from 11. The following compound or a pharmaceutically acceptable salt thereof, 12. The compound according to claim 11 or a pharmaceutically acceptable salt thereof, which is selected from:

13. Use of the compound according to any one of claims 1 to 12 or a pharmaceutically acceptable salt thereof in the preparation of a medicament related to complement factor B.

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