Method for preparing irak-4 degrader

Abstract

Provided are a method for preparing a compound represented by formula (I) and an intermediate thereof. The method has a high yield, a short reaction cycle, good atom economy and low production costs, is friendly to environment, involves a simple post-treatment, and has industrial application prospects.

Description

A method for preparing an IRAK-4 degrading agent

[0001] Cross-reference to related applications

[0002] This disclosure claims the benefit and priority of Chinese Patent Application No. 202411177985.5, filed with the China National Intellectual Property Administration on August 26, 2024, and Chinese Patent Application No. 202511165564.5, filed with the China National Intellectual Property Administration on August 19, 2025, the entire contents of which are incorporated herein by reference. Technical Field

[0003] This disclosure relates to the field of medicinal chemistry, specifically to a method for preparing an IRAK-4 degrading agent. Background Technology

[0004] Proteolysis-targeting chimaeras (PROTACs) technology is an emerging drug development technology that has emerged in recent years. Unlike the mechanism of action of traditional small-molecule inhibitors, PROTACs work by delivering these proteins into the proteasome for complete degradation, thus breaking the traditional definition of drugs. Compared with traditional small-molecule inhibitors, they have several advantages, such as making the target "drugable" instead of "undruggable," not relying on "site-driven" mechanisms, and overcoming the problem of drug resistance.

[0005] Interleukin-1 receptor-associated kinase 4 (IRAK-4) is a member of the intracellular serine-threonine kinase family, the IRAK family. It mediates Toll-like receptor (TLR) and IL-1 receptor (IL-1R) activation-mediated inflammatory signaling pathways, playing a crucial role in innate immune signaling. IRAK4 can be activated by recruitment by MyD88 to form Myddosomes, thereby activating downstream IRAK1 / 2, leading to subsequent hyperphosphorylation, dissociation from the complex, and binding to TRAF6. The activated TRAF6 complex triggers downstream signaling pathways to produce pro-inflammatory cytokines (NF-κB, CREB, AP-1, IRF, etc.). Inhibition of IRAK4 may play a significant role in the pathogenesis and progression of inflammatory diseases. Currently, inhibitors targeting IRAK4 are not approved. Therefore, knocking out IRAK4 using PROTAC technology may be another strategy for treating IRAK4-related diseases.

[0006] PCT / CN2024 / 079646 describes an IRAK4 degrading agent with the structure shown in formula (I) below. Its chemical name is 5-((1R,4R)-2-oxa-5-azabicyclo[2.2.1]heptane-5-yl)-N-(3-(difluoromethyl)-1-(7-(4-(1-(2,6-dioxopiperidin-3-yl)-3-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-4-yl)but-3-yn-1-yl)-7-azaspiro[3.5]nonane-2-yl)-1H-pyrazole-4-yl)pyrazole[1,5-a]pyrimidine-3-amide. This compound exhibits good in vitro and in vivo efficacy and is expected to be developed into a clinical drug.

[0007] Therefore, developing a synthetic route for this compound that has one or more of the following characteristics—high yield, short reaction cycle, good atom economy, low production cost, environmental friendliness, and suitability for industrial scale-up—is of great significance for promoting the early market launch of this product. Summary of the Invention

[0008] This disclosure provides two methods for preparing compounds and intermediates as shown in formula (I) below. These methods have one or more of the following advantages: high yield, short reaction cycle, good atom economy, low production cost, environmental friendliness, and simple post-processing, thus having prospects for industrial application.

[0009] In a first aspect, this disclosure provides a method for preparing a compound of formula (I), its salt, and its stereoisomers, by condensation of a compound of formula (II) or its salt with a compound of formula (III), its salt, or its stereoisomers.

[0010] Y is selected from OH and halogens.

[0011] In another embodiment of this disclosure, the compound of formula (III), its salt, or its stereoisomer is prepared by reduction of the compound of formula (IV), its salt, or its stereoisomer.

[0012] In another embodiment of this disclosure, the compound of formula (IV), its salt, or its stereoisomer is prepared (e.g., by reaction) from the compound of formula (VI), its salt, or its stereoisomer, and the compound of formula (V) or its salt.

[0013] Where X is a leaving group.

[0014] In another embodiment of this disclosure, Y is selected from halogens; preferably, Y is chlorine.

[0015] In another embodiment of this disclosure, X is selected from halogens,

[0016] R 1 Selected from 1-2 Rs 1a Replacement C 1-4 Alkyl, phenyl;

[0017] R 1a Selected from halogens, -NO2, C 1-4 alkyl.

[0018] In another embodiment of this disclosure, X is selected from halogens or CH3SO3-.

[0019] In another embodiment of this disclosure, X is selected from Br or CH3SO3-.

[0020] In another embodiment of this disclosure, when Y in compound (II) is OH, compound (II) or its salt is prepared (e.g., condensed) with compound (III), its salt or its stereoisomer under the conditions of basic catalyst A1, reaction solvent B1 and condensing agent C1 to obtain compound (I), its salt or its stereoisomer; or, when Y in compound (II) is halogen, compound (II) or its salt is prepared (e.g., condensed) with compound (III), its salt or its stereoisomer under the conditions of basic catalyst A1' and reaction solvent B1' to obtain compound (I), its salt or its stereoisomer.

[0021] In another embodiment of this disclosure, the alkaline catalysts A1 and A1' are independently selected from organic or inorganic bases, preferably one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate.

[0022] In another embodiment of this disclosure, the reaction solvents B1 and B1' are independently selected from one or more of tetrahydrofuran, N,N-dimethylformamide, N-methylpyrrolidone, acetonitrile, chloroform, dichloromethane, and toluene.

[0023] In another embodiment of this disclosure, the condensing agent C1 is selected from 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU), and N,N,N',N'-tetramethyl-O-(benzotriazole-1-yl)thiourea hexafluorophosphate. One or more of the following: salt (HBTU), O-benzotriazole-N,N,N',N'-tetramethylurea tetrafluoroborate (TBTU), Carter's condensing agent (BOP), benzotriazole-1-yl-oxytripyrrolylphosphine hexafluorophosphate (PyBOP), (7-azabenzotriazole-1-oxy)tripyrrolylphosphine hexafluorophosphate (PyAOP), carbonylimidazolium (CDI), 1-hydroxybenzotriazole (HOBt), and mixed acid anhydrides.

[0024] In another embodiment of this disclosure, for the compound of formula (II) in which Y is a halogen, it can be obtained by reacting a carboxylic acid structure (i.e., Y is OH) with a halogenated reagent D1, and after the reaction is completed, it can be directly reacted with formula (III) without separation; preferably, the halogenated reagent D1 is selected from one or more of sulfoxides, phosphorus halides, trihalomycin, oxalyl chloride and phosphorus oxychloride; preferably, the halogenated reagent D1 is a sulfoxide, preferably sulfoxide chloride.

[0025] In another embodiment of this disclosure, the alkaline catalyst A1 is selected from organic bases, preferably N,N-diisopropylethylamine; the reaction solvent B1 is selected from N,N-dimethylformamide; and the condensing agent C1 is selected from 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate, 1-hydroxybenzotriazole, or a combination thereof.

[0026] In another embodiment of this disclosure, the alkaline catalyst A1' is selected from organic bases, preferably triethylamine; the reaction solvent B1' is selected from tetrahydrofuran, N-methylpyrrolidone, or a combination thereof.

[0027] In another embodiment of this disclosure, the reaction conditions between the compound of formula (II) or its salt and the compound of formula (III), its salt or its stereoisomer include one or more of the following:

[0028] (a) The molar ratio of the compound of formula (II) or its salt to the compound of formula (III), its salt or its stereoisomer is 1 to 2:1, preferably 1 to 1.5:1, for example 1.1:1, 1.2:1, 1.3:1 or 1.4:1;

[0029] (b) The molar ratio of the compound of formula (III), its salt or stereoisomer to each condensing agent C1 is 1:1 to 1.5, for example 1:1.1, 1:1.2, 1:1.3 or 1:1.4;

[0030] (c) The molar ratio of the compound of formula (III), its salt or its stereoisomer to the basic catalyst A1 is 1:1 to 5, preferably 1:2 to 4, and preferably 1:3;

[0031] (d) The reaction time is 1 to 10 hours, preferably 2 to 5 hours, for example 2.5, 3, 3.5, 4 or 4.5 hours;

[0032] (e) The reaction temperature is 0 to 100°C, preferably 20 to 50°C, for example 25, 30, 35, 40 or 45°C.

[0033] In another embodiment of this disclosure, the reaction conditions between the compound of formula (II) or its salt and the compound of formula (III), its salt or its stereoisomer include one or more of the following:

[0034] (a) The molar ratio of the compound of formula (II) or its salt to the compound of formula (III), its salt or its stereoisomer is 1 to 2:1, preferably 1 to 1.5:1, for example 1.1:1, 1.2:1, 1.3:1 or 1.4:1;

[0035] (b) The molar ratio of the compound of formula (II) or its salt or its stereoisomer to the halogenated reagent D1 is 1:1 to 3, preferably 1:2 to 2.5, for example 1:2.1, 1:2.2, 1:2.3 or 1:2.4;

[0036] (c) The molar ratio of the compound of formula (III), its salt or stereoisomer to the basic catalyst A1' is 1:1 to 5, preferably 1:1 to 1.5, for example 1:1.1, 1:1.2, 1:1.3 or 1:1.4;

[0037] (d) The reaction time is 1 to 10 hours, preferably 2 to 5 hours, for example 2.5, 3, 3.5, 4 or 4.5 hours;

[0038] (e) The reaction temperature is 0 to 100°C, preferably 20 to 50°C, for example 25, 30, 35, 40 or 45°C.

[0039] In another embodiment of this disclosure, a compound of formula (IV), its salt, or its stereoisomer is prepared under the conditions of reducing agent A2, reaction solvent B2, and acidic reagent E2 to obtain a compound of formula (III), its salt, or its stereoisomer.

[0040] In another embodiment of this disclosure, the reducing agent A2 is selected from one or more of iron powder, zinc powder, and sodium thiosulfate.

[0041] In another embodiment of this disclosure, the reaction solvent B2 is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, acetonitrile, ethanol, and dioxane.

[0042] In another embodiment of this disclosure, the acidic reagent E2 is selected from one or more of ammonium chloride, hydrochloric acid, and acetic acid.

[0043] In another embodiment of this disclosure, the reducing agent A2 is zinc powder; the reaction solvent B2 is selected from tetrahydrofuran, acetonitrile, or a combination thereof; and the acidic reagent E2 is selected from ammonium chloride.

[0044] In another embodiment of this disclosure, the reaction conditions for the reduction preparation of compound (III), its salt, or its stereoisomer of formula (IV) to obtain compound (III), its salt, or its stereoisomer of formula (IV) include one or more of the following:

[0045] (a) The molar ratio of formula (IV) to reducing agent A2 is 1:1 to 20; preferably 1:8 to 12; preferably 1:10;

[0046] (b) The reaction time is 1 to 10 hours, preferably 2 to 5 hours, for example 2.5, 3, 3.5, 4 or 4.5 hours;

[0047] (c) The reaction temperature is 0 to 100°C, preferably 0 to 50°C, and more preferably 0 to 10°C, for example 1, 2, 3, 4, 5, 6, 7, 8 or 9°C.

[0048] In another embodiment of this disclosure, a compound of formula (VI), its salt or its stereoisomer is prepared by reacting a compound of formula (V) or its salt with a base catalyst A3 and a reaction solvent B3 to obtain a compound of formula (IV), its salt or its stereoisomer.

[0049] In another embodiment of this disclosure, the alkaline catalyst A3 is selected from one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate.

[0050] In another embodiment of this disclosure, the reaction solvent B3 is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, acetonitrile, and ethyl acetate.

[0051] In another embodiment of this disclosure, the alkaline catalyst A3 is sodium bicarbonate; and the reaction solvent B3 is acetonitrile.

[0052] In another embodiment of this disclosure, the reaction conditions between the compound of formula (VI), its salt, or its stereoisomer with the compound of formula (V) or its salt include one or more of the following:

[0053] (a) The molar ratio of the compound of formula (VI), its salt or its stereoisomer to the compound of formula (V) or its salt is 1:0.2 to 2, preferably 1:0.8 to 1;

[0054] (b) The molar ratio of the compound of formula (VI), its salt or stereoisomer to the base catalyst A3 is 1:1 to 2, preferably 1:1 to 1.5, for example 1:1.1, 1:1.2, 1:1.3 or 1:1.4;

[0055] (c) The reaction time is 5 to 60 hours, preferably 20 to 40 hours, for example 25, 30 or 35 hours;

[0056] (d) The reaction temperature is 0 to 100°C, preferably 40°C to 80°C, preferably 50°C to 80°C, and preferably 55°C to 75°C.

[0057] In another embodiment of this disclosure, a method for synthesizing a compound of formula (I), its salt, or a stereoisomer thereof is provided.

[0058] Wherein, X is selected from halogens or CH3SO3-;

[0059] Y is selected from OH or halogens; preferably, Y is OH.

[0060] The synthesis steps are as follows:

[0061] (1) Compounds of formula (VI) and (V) were reacted in acetonitrile with sodium bicarbonate to give compound (IV);

[0062] (2) Compound (IV) was reduced in tetrahydrofuran by zinc powder to obtain compound (III);

[0063] (3) Compound (II) and compound (III) were prepared by condensation of a basic catalyst, 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate, and 1-hydroxybenzotriazole to obtain compound (I);

[0064] Alternatively, the acyl chloride can be obtained by reacting the compound of formula (II) with thionyl chloride, and then the compound of formula (III) can be prepared under an alkaline catalyst to obtain the compound of formula (I).

[0065] The alkaline catalyst may be selected from N,N-diisopropylethylamine or triethylamine;

[0066] The reaction solvent for this step can be one or more of N,N-dimethylformamide, chloroform, N-methylpyrrolidone, and tetrahydrofuran.

[0067] Secondly, this disclosure provides another method for preparing a compound of formula (I), or a salt thereof, or a stereoisomer thereof, characterized in that it is prepared from a compound of formula (VII) or a salt thereof with a compound of formula (VI), a salt thereof, or a stereoisomer thereof.

[0068] Where X is selected from leaving groups.

[0069] In another embodiment of this disclosure, X is selected from halogens,

[0070] R 1 Selected from 1-2 Rs 1a Replacement C 1-4 Alkyl, phenyl;

[0071] R 1a Selected from halogens, -NO2, C 1-4 alkyl.

[0072] In another embodiment of this disclosure, X is selected from I or CH3SO3-.

[0073] In another embodiment of this disclosure, formula (VII) or its salt is prepared by removing the protecting group from formula (VIII) or its salt.

[0074] PG is an amino protecting group.

[0075] In another embodiment of this disclosure, PG is selected from (a) alkoxycarbonyl amino protecting groups, such as benzyloxycarboxyl (Cbz), tert-butyloxycarbonyl (Boc), methoxycarbonyl (Fmoc), allyloxycarbonyl (Alloc), trimethylsilylethoxycarbonyl (Teoc), methoxycarboxyl (or ethoxycarboxyl), etc.; (b) acyl amino protecting groups, such as phthaloyl (Pht), p-toluenesulfonyl (Tos), trifluoroacetyl (Tfa), o-(p-)nitrobenzenesulfonyl (Ns), tert-valeryl, benzoyl, etc.; (c) alkyl amino protecting groups, such as triphenylmethyl (Trt), 2,4-dimethoxybenzyl (Dmb), p-methoxybenzyl (PMB), benzyl (Bn), etc.; and (d) organosilicon protecting groups, such as trimethylsilyl (TMS), tert-butyldimethylsilyl (TBDMS), triisopropylsilyl (TIPS), diethylisopropylsilyl (DEIPS), etc. Preferably, the amino protecting group is an alkoxycarbonyl group, and more preferably Boc.

[0076] In another embodiment of this disclosure, the compound of formula (VIII) or a salt thereof is prepared by condensation of the compound of formula (II) or a salt thereof with the compound of formula (IX) or a salt thereof;

[0077] in,

[0078] PG is an amino protecting group;

[0079] Y is selected from OH and halogens.

[0080] In another embodiment of this disclosure, Y is selected from halogens.

[0081] In another embodiment of this disclosure, the compound of formula (VI), its salt or its stereoisomer, and the compound of formula (VII), its salt or its stereoisomer, are prepared under alkaline catalyst A4 and reaction solvent B4 to obtain the compound of formula (I), its salt or its stereoisomer.

[0082] In another embodiment of this disclosure, the alkaline catalyst A4 is selected from one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate.

[0083] In another embodiment of this disclosure, the reaction solvent B4 is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, acetonitrile, and ethyl acetate.

[0084] In another embodiment of this disclosure, the alkaline catalyst A4 is selected from potassium bicarbonate; and the reaction solvent B4 is selected from acetonitrile.

[0085] In another embodiment of this disclosure, the reaction conditions between the compound of formula (VI), its salt, or its stereoisomer with the compound of formula (VII) or its salt include one or more of the following:

[0086] (a) The molar ratio of the compound of formula (VI), its salt or its stereoisomer to the compound of formula (VII) or its salt is 1:0.2 to 2, preferably 1:0.8 to 1;

[0087] (b) The molar ratio of the compound of formula (VI), its salt or its stereoisomer to the basic catalyst A4 is 1:1 to 10, preferably 1:4 to 6, and more preferably 1:5;

[0088] (c) The reaction time is 5 to 60 hours, preferably 5 to 20 hours, for example 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18 or 19 hours;

[0089] (d) The reaction temperature is 0 to 100°C, preferably 20 to 60°C, for example 25, 30, 35, 40, 45, 50 or 55°C.

[0090] In another embodiment of this disclosure, the protecting group of formula (VIII) or its salt is removed by catalytic hydrogenolysis, acidolysis, or alkali-catalytic cleavage to prepare formula (VII) or its salt.

[0091] In another embodiment of this disclosure, the acid is selected from organic or inorganic acids, preferably one or more of hydrochloric acid, trifluoroacetic acid, acetic acid, and hydrobromic acid; the base is selected from organic or inorganic bases, preferably one or more of potassium hydroxide, sodium hydroxide, potassium carbonate, triethylamine, piperidine, tetrabutylammonium fluoride, and hydrazine hydrate.

[0092] In another embodiment of this disclosure, formula (VIII) or its salt is prepared in acidic reagent A5 and reaction solvent B5 to obtain formula (VII) or its salt.

[0093] In another embodiment of this disclosure, the acidic reagent A5 is selected from organic acids, preferably trifluoroacetic acid.

[0094] In another embodiment of this disclosure, the reaction solvent B5 is selected from one or more of dichloromethane, trichloromethane, tetrahydrofuran, acetonitrile, 1,4-dioxane, methanol, ethanol, isopropanol, ethyl acetate, isopropyl acetate, N,N-dimethylformamide, N-methylpyrrolidone, acetone, and butanone; preferably dichloromethane.

[0095] In another embodiment of this disclosure, the reaction conditions for preparing formula (VII) or its salt by removing the protecting group include one or more of the following:

[0096] (a) The ratio (mol / ml) of the molar amount of formula (VIII) or its salt to the volume of acidic reagent A5 is 1:1 to 3; preferably 1:2;

[0097] (b) The reaction time is 1 to 10 hours, preferably 1 to 5 hours, for example 1.5, 2, 2.5, 3, 3.5, 4 or 4.5 hours;

[0098] (c) The reaction temperature is 0 to 100°C, preferably 20 to 50°C, for example 25, 30, 35, 40 or 45°C.

[0099] In another embodiment of this disclosure, when Y in compound (II) is OH, compound (IX) or its salt is prepared by reacting compound (II) or its salt with alkaline catalyst A6, reaction solvent B6, and condensing agent C2; or, when Y in compound (II) is halogen, compound (II) or its salt is prepared by reacting compound (IX) or its salt with alkaline catalyst A6' and reaction solvent B6'.

[0100] In another embodiment of this disclosure, the alkaline catalysts A6 and A6' are selected from organic or inorganic bases, preferably one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate.

[0101] In another embodiment of this disclosure, the reaction solvents B6 and B6' are selected from one or more of tetrahydrofuran, N,N-dimethylformamide, N-methylpyrrolidone, acetonitrile, chloroform, dichloromethane, and toluene.

[0102] In another embodiment of this disclosure, the condensing agent C2 is selected from 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU), and N,N,N',N'-tetramethyl-O-(benzotriazole-1-yl)thiourea hexafluorophosphate. One or more of the following: salt (HBTU), O-benzotriazole-N,N,N',N'-tetramethylurea tetrafluoroborate (TBTU), Carter's condensing agent (BOP), benzotriazole-1-yl-oxytripyrrolylphosphine hexafluorophosphate (PyBOP), (7-azabenzotriazole-1-oxy)tripyrrolylphosphine hexafluorophosphate (PyAOP), carbonylimidazolium (CDI), 1-hydroxybenzotriazole (HOBt), and mixed acid anhydrides.

[0103] In another embodiment of this disclosure, when Y in compound (II) is a halogen, it can be obtained by reacting a carboxylic acid structure (i.e., when Y is OH) with a halogenated reagent D2, and after the reaction is completed, it can be directly reacted with compound (IX) without separation; preferably, the halogenated reagent D2 is selected from one or more of sulfoxides, phosphorus halides, trihalomycin, oxalyl chloride and phosphorus oxychloride; preferably, the halogenated reagent D2 is a sulfoxide, preferably sulfoxide chloride.

[0104] In another embodiment of this disclosure, the alkaline catalyst A6 is selected from organic bases, preferably N,N-diisopropylethylamine; the reaction solvent B6 is selected from N,N-dimethylformamide; and the condensing agent C2 is selected from 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole, or a combination thereof.

[0105] In another embodiment of this disclosure, the alkaline catalyst A6' is selected from organic bases, preferably triethylamine; the reaction solvent B6' is selected from tetrahydrofuran, N-methylpyrrolidone, or a combination thereof.

[0106] In another embodiment of this disclosure, the reaction conditions between the compound of formula (IX) or a salt thereof and the compound of formula (II) or a salt thereof include one or more of the following:

[0107] (a) The molar ratio of the compound of formula (IX) or its salt to the compound of formula (II) or its salt is 1:1 to 1.5, for example 1:1.1, 1:1.2, 1:1.3 or 1:1.4;

[0108] (b) The molar ratio of the compound of formula (IX) or its salt to each condensing agent C2 is 1:1 to 2; preferably 1:1 to 1.5, for example 1:1.1, 1:1.2, 1:1.3 or 1:1.4;

[0109] (c) The molar ratio of the compound of formula (IX) or its salt to the basic catalyst A6 is 1:1 to 5; the molar ratio is 1:2 to 4; preferably 1:2;

[0110] (d) The reaction time is 1 to 30 hours, preferably 15 to 25 hours, for example 16, 17, 18, 19, 20, 21, 22, 23 or 24 hours;

[0111] (e) The reaction temperature is 0 to 100°C, preferably 20 to 50°C, for example 25, 30, 35, 40 or 45°C.

[0112] In another embodiment of this disclosure, a method for synthesizing a compound of formula (I), its salt, or a stereoisomer thereof is provided.

[0113] Wherein, Y is selected from OH or halogen; preferably, Y is OH;

[0114] PG is an amino protecting group;

[0115] The synthesis steps are as follows:

[0116] (1) Compounds of formula (IX) and (II) were catalyzed by 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N,N-diisopropylethylamine in N,N-dimethylformamide to give compound (VIII);

[0117] (2) Formula (VIII) was deprotected in dichloromethane by trifluoroacetic acid catalysis to give compound (VII);

[0118] (3) Formulas (VII) and (VI) are reacted in acetonitrile with potassium bicarbonate to obtain compound (I).

[0119] Thirdly, this disclosure provides a method for preparing p-toluenesulfonate of compound (I), which is prepared by adding p-toluenesulfonic acid to a reaction system of compound (I) and reacting the reaction. Compound (I) is as defined in the preceding scheme.

[0120] In another embodiment of this disclosure, the preparation method of compound (I) p-toluenesulfonate is to obtain it by condensing compound (II) or its salt with compound (III), its salt or its stereoisomer, and then adding p-toluenesulfonic acid to the reaction system to form a salt. The reaction steps of compound (II) or its salt with compound (III), its salt or its stereoisomer are as described in the above scheme.

[0121] In another embodiment of this disclosure, the solvent for the salt formation reaction of the compound of formula (I) toluenesulfonate is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, N-methylpyrrolidone, acetonitrile, chloroform, dichloromethane, toluene, isopropanol, and ethanol.

[0122] In another embodiment of this disclosure, the salt formation reaction conditions of the compound of formula (I) with p-toluenesulfonic acid include one or more of the following:

[0123] (a) The molar ratio of the compound of formula (I) to p-toluenesulfonic acid is 1:1 to 10; preferably 1:2 to 6, for example 1:3, 1:4 or 1:5;

[0124] (b) The reaction time is 1 to 10 hours, preferably 2 to 5 hours, for example 2.5, 3, 3.5, 4 or 4.5 hours;

[0125] (c) The reaction temperature is 0 to 100°C, preferably 20 to 50°C, for example 25, 30, 35, 40 or 45°C.

[0126] In another embodiment of this disclosure, the ratio of compound (I) to p-toluenesulfonate in compound (I) is 1:1.

[0127] In another embodiment of this disclosure, the p-toluenesulfonate of the compound of formula (I) is a hydrate.

[0128] In another embodiment of this disclosure, the molar ratio of p-toluenesulfonate of compound (I) to water is 1:1 to 3, for example 1:2.

[0129] In another embodiment of this disclosure, the water content of the compound (I) p-toluenesulfonate is 1%-5%; preferably, the water content is 1.5%-3.5%; preferably, the water content is no more than 2.5%; preferably, the water content is no more than 2%.

[0130] In another embodiment of this disclosure, the p-toluenesulfonate of compound (I) is a monohydrate.

[0131] Fourthly, this disclosure provides the following compounds, their salts, or stereoisomers thereof.

[0132] Wherein, PG is an amino protecting group, as defined in the previous scheme.

[0133] Fifthly, this disclosure provides a method for preparing a compound of formula (III), its salt, or its stereoisomer, characterized in that the compound of formula (III), its salt, or its stereoisomer is prepared by reduction of a compound of formula (IV), its salt, or its stereoisomer.

[0134] Preferably, the reduction reaction steps of the compound of formula (IV), its salt or its stereoisomer are as described in the preceding scheme;

[0135] Preferably, the compound of formula (IV), its salt, or its stereoisomer is prepared from the compound of formula (VI), its salt, or its stereoisomer, and the compound of formula (V), or its salt.

[0136] Where X is a leaving group;

[0137] Preferably, the reaction steps of the compound of formula (VI), its salt or its stereoisomer with the compound of formula (V) or its salt are as described in the preceding scheme.

[0138] In another embodiment of this disclosure, the salt of the compound of formula (I) or the intermediate described above is preferably a pharmaceutically acceptable salt.

[0139] Definitions and Explanations

[0140] Unless otherwise stated, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase should not be considered uncertain or unclear unless specifically defined, but should be understood in its ordinary sense.

[0141] The term "PG" stands for amino protecting group, and all protecting groups known to those skilled in the art are within the scope of this disclosure. For example, common amino protecting groups include: (1) alkoxycarbonyl amino protecting groups, such as benzyloxycarboxyl (Cbz), tert-butyloxycarbonyl (Boc), methoxycarbonyl (Fmoc), allyloxycarbonyl (Alloc), trimethylsilylethoxycarbonyl (Teoc), methoxycarboxyl (or eth)carboxyl, etc.; (2) acyl amino protecting groups, such as phthaloyl (Pht), p-toluenesulfonyl (Tos), trifluoroacetyl (Tfa), o-(p-)nitrobenzenesulfonyl (Ns), tert-valeryl, benzoyl, etc.; (3) alkyl amino protecting groups, such as triphenylmethyl (Trt), 2,4-dimethoxybenzyl (Dmb), p-methoxybenzyl (PMB), benzyl (Bn), etc.; (4) organosilicon protecting groups, such as trimethylsilyl (TMS), tert-butyldimethylsilyl (TBDMS), triisopropylsilyl (TIPS), diethylisopropylsilyl (DEIPS), etc. Preferably, the amino protecting group is an alkoxycarbonyl group, and more preferably Boc.

[0142] The methods for removing amino protecting groups disclosed herein are techniques well known to those skilled in the art, including but not limited to: catalytic hydrogenolysis, acid hydrolysis, base-catalyzed hydrolysis, or Na / NH3, etc. The acid can be an organic or inorganic acid, such as hydrochloric acid, trifluoroacetic acid, acetic acid, hydrobromic acid, etc.; the base can be an organic or inorganic base, such as potassium hydroxide, sodium hydroxide, potassium carbonate, piperidine, tetrabutylammonium fluoride, hydrazine hydrate, etc. Acid hydrolysis is the preferred deprotection method.

[0143] The term "leaving group" or leaving radical refers to an atom or functional group that detaches from a larger molecule during a chemical reaction. It is a term used in nucleophilic substitution and elimination reactions. Common leaving groups include: -halogen, -OCOR, -OTs, -ONO2, -OH, etc.

[0144] The term "halogen" refers to fluorine, chlorine, bromine, or iodine atoms.

[0145] The term "alkyl" refers to a straight-chain or branched aliphatic saturated hydrocarbon group consisting of carbon and hydrogen. For example, the term "C 1-4 "Alkyl" refers to an alkyl group having 1 to 4 carbon atoms, examples of which include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, etc.

[0146] The term "salt" refers to a derivative obtained by reacting a compound or intermediate structure described in this disclosure with an acid or base. These salts can be prepared during the synthesis, isolation, or purification of the compound or intermediate, or by reacting the purified compound or intermediate in its free form with a suitable acid or base. Salts of intermediate structures can be retained in the product or eliminated during the reaction. The acids include organic and inorganic acids; common organic acids include, but are not limited to, maleic acid, malic acid, formic acid, acetic acid, tartaric acid, p-toluenesulfonic acid, benzoic acid, and methanesulfonic acid; common inorganic acids include, but are not limited to, hydrochloric acid, sulfuric acid, hydrobromic acid, and phosphoric acid. The bases include organic and inorganic bases; common organic bases include, but are not limited to, organic amines (such as triethylamine), alkali metal salts of alcohols (such as sodium methoxide), alkyl lithium metal compounds (such as butyllithium), and amino lithium compounds (such as diisopropylaminolithium); common inorganic bases include sodium hydroxide, potassium hydroxide, sodium carbonate, sodium bicarbonate, potassium carbonate, and potassium bicarbonate.

[0147] The term "pharmaceutically acceptable" refers to compounds, materials, compositions, and / or dosage forms that are suitable for use in human and animal tissues to the extent of reasonable medical judgment without excessive toxicity, irritation, allergic reactions, or other problems or complications, and that are commensurate with a reasonable benefit / risk ratio.

[0148] The term "pharmaceutically acceptable salt" refers to derivatives obtained from the compounds of this disclosure prepared with relatively non-toxic acids or bases. These salts can be prepared during the synthesis, isolation, and purification of the compounds, or by reacting the purified free form of the compounds with suitable acids or bases. When the compounds contain relatively acidic functional groups (e.g., -COOH, -OH, -SO3H, etc.), they react with suitable inorganic or organic cations (bases) to yield base addition salts, including salts formed with alkali metals or alkaline earth metals, ammonium salts formed with amines or their derivatives, and salts formed with amino acids. When the compounds contain relatively basic functional groups (e.g., -NH2, etc.), they react with suitable inorganic or organic anions (acids) to yield acid addition salts, including salts formed with inorganic or organic acids (e.g., carboxylic acids, etc.).

[0149] The term "stereoisomer" refers to isomers resulting from different spatial arrangements of atoms in a molecule. Stereoisomers can be classified into cis-trans isomers and enantiomers, or further divided into enantiomers and diastereomers. Cis-trans isomers are commonly characterized by "Z" and "E" types; stereoisomers are commonly characterized by "R" and "S" types. Mixtures of diastereomers can be separated into individual diastereomers based on their physical / chemical differences using methods well-known in the art, such as chromatography and / or stepwise crystallization. Enantiomers can be separated by chiral column chromatography or by reacting the enantiomer compound with a suitable optically active compound to obtain a mixture of diastereomers, separating each diastereomer, and converting each diastereomer to its corresponding enantiomer. The compounds disclosed herein can be any such isomer, including diastereomers, enantiomers, and mixtures thereof.

[0150] The term "mixed anhydrides" refers to a compound formed by the dehydration of two different organic acids, with the structural formula R1-CO-O-CO-R2, where R1 and R2 are optionally substituted aliphatic or aromatic groups. Here, the aliphatic group can be C... 1-10 Alkyl, C 2- 10 alkenyl, C 2-10 alkynyl group, C 3-10 cycloalkyl, C 3-10 Cycloalkenyl or C 3-10 The cycloalkynyl group can also be a heteroalkyl, heteroalkenyl, heteroynyl, heterocycloalkyl, heterocycloalkenyl, or heterocycloalkynyl group formed by the above-mentioned group containing one or more (e.g., 1, 2, or 3) heteroatoms selected from O, S, N, or P. Here, the aromatic group can be C 6-18 Aryl groups can also be heteroaryl groups formed by the inclusion of one or more (e.g., 1, 2 or 3) heteroatoms selected from O, S, N or P.

[0151] In this article, the number of carbon atoms mentioned before a group, such as "C", is used to indicate the number of carbon atoms. 1-10 “C” 2-10 “C” 3-10 “C” 6-18 ", includes any subrange within that endpoint range, for example, each containing "C" 1-6 “C” 1-4 “C” 2-6 “C” 2-4 “C” 3-6 “C” 6- 12 “C” 6-10 These subscopes are all considered to be within the scope of this disclosure.

[0152] In this article, "optional substitution" means that the modified group can be unsubstituted or selected from deuterium, halogens, C64 ... 1- 4-alkyl, C 6-12 One or more substituents of aryl or a combination thereof are used for substitution.

[0153] Unless otherwise stated, in this disclosure, the structure of the compound is... This indicates that the atom connecting the bond is a chiral atom, and the compound can be a racemic mixture, isomer, or a mixture of isomers in equal or different amounts. A wedge-shaped solid line bond is used. and wedge-shaped dashed key It represents the absolute configuration of the center of a solid. Attached Figure Description

[0154] Figure 1 shows the concentration levels of inflammatory factors in mouse plasma and peritoneal lavage fluid after oral administration of three different doses of KT-474 (15 mg / kg, 50 mg / kg, and 150 mg / kg) and compound (I) in Test Example 6. Specifically, (a) shows the IL-5 level in mouse plasma, (b) shows the MCP-1 level in mouse plasma, (c) shows the IL-5 level in mouse peritoneal lavage fluid, and (d) shows the MCP-1 level in mouse peritoneal lavage fluid.

[0155] Figure 2 shows the in vivo degradation level of IRAK4 in the spleen of mice after oral administration of three different doses of KT-474 and compound (I) at doses of 15 mg / kg, 50 mg / kg, and 150 mg / kg in Test Example 6.

[0156] Figure 3 shows the TGA-DSC spectrum of p-toluenesulfonate of compound (I). Detailed Implementation

[0157] The compounds disclosed herein can be prepared by a variety of synthetic methods known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthetic methods, and equivalent substitutions known to those skilled in the art. Preferred embodiments include, but are not limited to, the embodiments disclosed herein.

[0158] Summary of experimental instruments:

[0159] The structures of the compounds disclosed herein were determined by nuclear magnetic resonance (NMR) and / or liquid chromatography-mass spectrometry (LC-MS), or ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS). NMR chemical shifts (δ) are given in parts per million (ppm). NMR determinations were performed using a Bruker Neo 400M or Bruker Ascend 400 NMR spectrometer, with solvents including deuterated dimethyl sulfoxide (DMSO-d6), deuterated methanol (CD3OD), and deuterated chloroform (CDCl3), heavy water (D2O), and tetramethylsilane (TMS) as the internal standard.

[0160] The determination was performed using an Agilent 1260-6125B single quadrupole mass spectrometer with a Welch Biomate column (C18, 2.7 μm, 4.6 × 50 mm) or a Waters H-Class SQD2 column with a Welch Ultimate column (XB-C18, 1.8 μm, 2.1 × 50 mm) (ion source: electrospray ionization).

[0161] The determination was performed using ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) with a Waters UPLC H-class SQD mass spectrometer (ion source: electrospray ionization).

[0162] HPLC determinations were performed using a Waters e2695-2998 or Waters ARC and an Agilent 1260 or Agilent Poroshell HPH high-performance liquid chromatograph.

[0163] Thin-layer chromatography (TLC) uses GF254 silica gel plates from Yantai Jiangyou Silica Gel Development Co., Ltd. or GF254 silica gel plates from Rushan Shangbang New Materials Co., Ltd. The TLC plate size is 0.15mm to 0.20mm, and the preparation type is 20×20cm.

[0164] The starting materials used in the embodiments of this disclosure are known and commercially available, or can be synthesized using methods known in the art.

[0165] Unless otherwise specified, all reactions in this disclosure are carried out under continuous magnetic stirring and a dry nitrogen or argon atmosphere, using a dry solvent. Reaction temperatures are expressed in degrees Celsius or °C. Unless otherwise specified, room temperature refers to 25 ± 5 °C.

[0166] Raw material preparation:

[0167] Compound II (where Y is OH) can be prepared by referring to the method for intermediate AEH described in WO2020113233A1.

[0168] Compounds of formula VI can be prepared by referring to the method for intermediate WZ described in WO2019133531A1.

[0169] Formula V is prepared by the following method:

[0170] Preparation steps:

[0171] Compound Va (40.0 g, 1.0 eq, CAS No. 1789048-54-9) and potassium carbonate (84.7 g, 2.5 eq) were added to N,N-dimethylformamide (400 ml). Compound Vb (PG: Boc, CAS No. 1638764-90-5) (94.8 g, 1.1 eq) was added with stirring. The mixture was heated to 100 °C and reacted. After the reaction was complete as detected by TLC, the temperature was lowered to 0–5 °C. The reaction solution was slowly added to a mixed solvent of purified water (2800 ml) and n-heptane (600 ml). The mixture was filtered, and the filter cake was washed with purified water (400 ml). The filter cake was recrystallized from dichloromethane (150 ml) / n-heptane (1500 ml), filtered, and the filter cake was dried to obtain compound Vc (70.0 g, yield 73.9%).

[0172] 1 H NMR (400MHz, DMSO) δ9.14 (s, 1H), δ7.32 (t, 1H, J = 52.8Hz), δ5.06-5.01 (m, 1H), δ3.33-3.31 (m, 2H ), δ3.25-3.22(m,2H), δ2.44-2.39(m,2H), δ2.30-2.25(m,2H), δ1.61-1.54(m,4H), δ1.40(s,9H).

[0173] MS(ESI)m / z:287.14[M-(CH3)3COCO+2H + ].

[0174] Compound Vc (65.0 g) was added to dichloromethane (390 ml), and trifluoroacetic acid (162 ml) was slowly added at 0–5 °C. After the addition was complete, the mixture was stirred at 20–30 °C. After the reaction was complete as detected by TLC, the mixture was concentrated under reduced pressure. Purified water (390 ml) was added to the concentrate, and the mixture was filtered. The filter cake was washed with purified water (130 ml) and dried to obtain compound V (45.3 g, yield 94.0%).

[0175] 1H NMR (400MHz, DMSO) δ9.16 (s, 1H), δ8.72 (s, 1H), δ7.32 (t, 1H, J = 53.2Hz), δ5.08-5.00 (m ,1H), δ3.08-2.97(m,4H), δ2.52-2.43(m,2H), δ2.36-2.30(m,2H), δ1.85-1.79(m,4H).

[0176] MS(ESI) m / z: 287.22 [M+H + ].

[0177] Formula IX is prepared by the following method:

[0178] The compound of formula Vc (PG is Boc) was dissolved in tetrahydrofuran at room temperature, and 10% wet palladium on carbon was added at room temperature. The reaction mixture was then stirred at 30°C under hydrogen balloon protection for 3 hours. LC-MS showed that the starting material reacted completely. The reaction mixture was filtered, and the filter cake was washed with ethyl acetate (100 mL). The combined filtrates were concentrated to give the desired product, compound IX. MS (ESI) M / Z: 357 [M+H + ].

[0179] Example 1: Preparation of compound (I)

[0180] Reaction route:

[0181] Step 1:

[0182] Method 1:

[0183] Compound (V) (10.0 g, 0.8 eq) was added to acetonitrile (100 ml), followed by sodium bicarbonate (3.5 g, 1.2 eq). The mixture was heated to 75 °C, and a solution of compound (VI) (X = methanesulfonate) (17.0 g, 1.0 eq) in acetonitrile (110 ml) was added dropwise. The reaction was maintained at this temperature for 26 hours. After the reaction was complete as detected by TLC, the temperature was lowered to -10 °C, and crystallization was carried out for 2 hours. The mixture was filtered, and the filter cake was washed successively with ammonium chloride solution (150 ml) and purified water (60 ml). The filter cake was dried under vacuum at 50 ± 5 °C to obtain compound (IV) (yield 55.6%).

[0184] 1H NMR (400MHz, DMSO) δ11.13(s,1H),9.13(s,1H),7.45-7.18(m,1H),7.12(m,1H),7.06-6.99(m,2H),5.41-5.37(m,1H),5.03-4.99 (m,1H),4.0-4.00(m,1H),3.68(s,3H),2.89-2.85(m,1H),2.75-2.54(m,5H),2.43-2.24(m,5H),2.12-2.04(m,4H),1.59(d,4H).

[0185] MS(ESI) m / z: 596.32 [M+H + ].

[0186] Method 2:

[0187] Compound (V) (1.0 g, 0.8 eq) and compound (VI) (X = Br) (1.70 g, 1.0 eq) were added to acetonitrile (100 ml), and sodium bicarbonate (0.35 g, 1.2 eq) was added. The mixture was heated to 55 °C and reacted. After the reaction was completed as detected by TLC, the reaction solution was concentrated under reduced pressure. The concentrate was purified by rapid column chromatography (dichloromethane:methanol = 20:1) to obtain compound (IV) (yield 13.6%).

[0188] Step 2:

[0189] Compound (IV) (10 g, 1.0 eq) was added to a mixture of acetonitrile (50 ml) and tetrahydrofuran (100 ml), followed by zinc powder (10.9 g, 10 eq). The temperature was controlled at 0–10 °C, and ammonium chloride solution (110 ml) was added dropwise to the reaction system. After the reaction was complete as detected by TLC, sodium chloride solution (100 ml) was added, and the mixture was filtered through diatomaceous earth. The filtrate was extracted with ethyl acetate (150 ml × 3), and the organic phase was concentrated under reduced pressure to obtain compound (III) (yield 90.3%).

[0190] 1 H NMR (400MHz, DMSO) δ11.13(s,1H),9.13(s,1H),7.19-6.77(m,5H),5.41-5.37(m,1H),4.74-4.69(t,J=8.4Hz,1H),4.08 (s,2H),3.68(s,3H),2.90-2.86(m,1H),2.75-2.54(m,5H),2.43-2.24(m,5H),2.12-2.04(m,3H),1.59(d,J=15.8,4H).

[0191] MS(ESI) m / z: 566.45 [M+H + ].

[0192] Step 3:

[0193] Method 1:

[0194] Compound (II) (5.5 g, 1.2 eq) and N,N-dimethylformamide (0.1 g, 0.08 eq) were added to chloroform (50 ml), and thionyl chloride (6.0 g, 2.8 eq) was added dropwise. The reaction was carried out at room temperature for 2 hours. The reaction was detected by TLC and the mixture was concentrated under reduced pressure. N-methylpyrrolidone (50 ml) was added to the concentrate.

[0195] Compound (III) (10.0 g, 1.0 eq) was added to tetrahydrofuran (50 ml), and a solution of triethylamine (2.3 g, 1.3 eq) and the above acyl chloride (i.e., the above concentrate) in N-methylpyrrolidone was added at 0–10 °C. The reaction was carried out at room temperature for 3 hours, and the reaction was detected by TLC. The solution was added to purified water (250 ml), filtered, and the filter cake was dried at 50 °C to obtain compound (I) (yield 88.7%).

[0196] 1 H NMR (400MHz, d-DMSO) δ11.14(s,1H),9.51(d,J=4.8Hz,1H),8.79-8.76(m,1H),8.41-8.40(d,J=3.6Hz,1H),8.27-8.26(d ,J=5.2Hz,1H),7.39-6.97(m,4H),6.87-6.85(d,J=7.6Hz,0.5H),6.46-6.43(d,J=7.6Hz,0.5H),5.42-5.38(dd,J=12.4,5 .2Hz,1H),5.28(s,0.5H),5.08(s,0.5H),4.97-4.93(t,J=8.0Hz,1H),4.79-4.76(d,J=14.4Hz,1H),3.83-3.81(m,1.5H), 3.75-3.73(m,0.5H),3.68-3.59(m,5H),3.45-3.36(m,1H),2.89-2.85(m,1H),2.73-2.60(m,6H),2.45-2.31(m,5H),2.28 -2.20(m,2H),2.18-1.98(m,3H),1.66-1.61(m,4H).

[0197] MS(ESI) m / z: 808.3 [M+H + ].

[0198] Method 2:

[0199] Compound (II) (200 mg, 1.1 eq), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (350 mg, 1.3 eq), and 1-hydroxybenzotriazole (125 mg, 1.3 eq) were added to N,N-dimethylformamide (4 ml). N,N-diisopropylethylamine (275 mg, 3 eq) was slowly added, and the mixture was stirred for 30 minutes. Compound (III) (400 mg, 1 eq) was then added, and the mixture was reacted at 40 °C for 3 hours. The reaction mixture was slowly added to purified water (16 ml), filtered, and the filter cake was dried at 50 °C to obtain compound (I) (yield 91.2%).

[0200] Example 2: Preparation of compound (I)

[0201] Reaction route:

[0202] Step 1:

[0203] At room temperature, compound (II) (5.39 g, 20.65 mmol) was dissolved in dry N,N-dimethylformamide (70 ml). 1-hydroxybenzotriazole (3.98 g, 29.5 mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (5.65 g, 29.5 mmol), and N,N-diisopropylethylamine (5.08 g, 39.3 mmol) were added sequentially to the reaction solution. Compound (IX) (7 g, 19.66 mmol) was then added to the above reaction solution. The reaction was stirred at 50 °C for 18 hours. After the reaction was complete as detected by TLC, the reaction solution was quenched with water (500 ml), and the mixture was extracted with ethyl acetate (250 ml × 3). The organic phases were combined, washed with saturated brine (300 ml × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting mixture was purified by silica gel column chromatography (eluent: dichloromethane / methanol = 20 / 1) to give 12.1 g of compound (VIII) in 96.9% yield.

[0204] 1H NMR (400MHz, d-DMSO) δ9.50(d,J=4.8Hz,1H),8.78(d,J=7.7Hz,1H),8.41(d,J=4.0Hz,1H),8.26(d,J=5.4Hz,1H),7.15(dd,J=54.5,52.4 Hz,1H),6.87(d,J=7.9Hz,0.5H),6.45(d,J=7.7Hz,0.5H),5.28(s,0.5H),5.08(s,0.5H),5.03-4.90(m,1H),4.77(d,J=15.9Hz,1H),3.82 (d,J=9.2Hz,1.5H),3.74(d,J=7.5Hz,0.5H),3.67-3.54(m,1.5H),3.45(d,J=10.1Hz,0.5H),3.32(s,2H),3.24(d,J=4.9Hz,2H),2.37(t, J=10.2Hz,2H),2.21(t,J=10.3Hz,2H),2.03(t,J=9.8Hz,1H),1.94(d,J=10.2Hz,1H),1.64-1.57(m,2H),1.57-1.49(m,2H),1.40(s,9H).

[0205] MS(ESI)M / Z:599.2[M+H]+.

[0206] Step 2:

[0207] Compound (VIII) (12.0 g, 20.07 mmol) was dissolved in dry dichloromethane (80 ml) at room temperature, and trifluoroacetic acid (40 ml) was added to the reaction solution. The reaction solution was stirred at room temperature for 3 hours. After the reaction was completed as detected by TLC, the reaction solution was concentrated under reduced pressure to give 9.25 g of compound (VII), with a yield of 92.5%.

[0208] 1H NMR (400MHz, d-DMSO) δ12.00(s,3H),9.51(d,J=4.9Hz,1H),8.79(dd,J=7.8,1.6Hz,1H),8.49(s,1.5H),8.44(d,J=4.3Hz,1H),8. 27(d,J=5.3Hz,1H),7.15(td,J=53.6,2.0Hz,1H),6.88(d,J=7.9Hz,0.5H),6.46(d,J=7.8Hz,0.5H),5.28(s,0.5H),5.08(s,.5H) ,4.98(p,J=8.1Hz,1H),4.78(d,J=17.6Hz,1H),3.83(d,J=9.6Hz,1.5H),3.75(d,J=7.6Hz,0.5H),3.68-3.58(m,1.5H),3.46(d,J =10.0Hz,0.5H),3.09(s,2H),3.00(s,2H),2.45(dd,J=11.2,9.6Hz,2H),2.33-2.24(m,2H),2.06-1.93(m,2H),1.88-1.74(m,4H).

[0209] MS(ESI)M / Z:499.2[M+H] + .

[0210] Step 3:

[0211] Compound (VI) (500 mg, 1.0 eq) and compound (VII) (570 mg, 1.0 eq) were dissolved in acetonitrile (30 mL) at room temperature. Potassium bicarbonate (572 mg, 5.0 eq) was added with stirring, and the mixture was heated to 45 °C and stirred overnight. The reaction mixture was cooled to room temperature, and saturated sodium bicarbonate aqueous solution (80 mL) was added. The mixture was then extracted with ethyl acetate (3 × 30 mL). The organic phase was concentrated under reduced pressure, and the concentrate was purified by rapid column chromatography (eluent: dichloromethane / anhydrous methanol = 94 / 6 methanol) to give compound (I) (yield 13%).

[0212] 1H NMR (400MHz, d-DMSO) δ11.14(s,1H),9.51(d,J=4.8Hz,1H),8.79-8.76(m,1H),8.41-8.40(d,J=3.6Hz,1H),8.27-8.26(d ,J=5.2Hz,1H),7.39-6.97(m,4H),6.87-6.85(d,J=7.6Hz,0.5H),6.46-6.43(d,J=7.6Hz,0.5H),5.42-5.38(dd,J=12.4,5 .2Hz,1H),5.28(s,0.5H),5.08(s,0.5H),4.97-4.93(t,J=8.0Hz,1H),4.79-4.76(d,J=14.4Hz,1H),3.83-3.81(m,1.5H), 3.75-3.73(m,0.5H),3.68-3.59(m,5H),3.45-3.36(m,1H),2.89-2.85(m,1H),2.73-2.60(m,6H),2.45-2.31(m,5H),2.28 -2.20(m,2H),2.18-1.98(m,3H),1.66-1.61(m,4H).

[0213] MS(ESI) m / z: 808.3 [M+H + ].

[0214] Example 3: Preparation of p-toluenesulfonate of compound (I)

[0215] Reaction route:

[0216] Compound (II) (5.29 g, 1.15 eq) and N,N-dimethylformamide (0.09 g, 0.07 eq) were added to chloroform (53 ml), and thionyl chloride (6.05 g, 2.88 eq) was added dropwise at 0–10 °C. The reaction was carried out at room temperature for 1 hour. The reaction was completed by TLC. The mixture was concentrated under reduced pressure, and N-methylpyrrolidone (50 ml) was added to the concentrate.

[0217] Compound (III) (10.0 g, 1.0 eq) was added to tetrahydrofuran (50 ml), and a solution of triethylamine (2.33 g, 1.3 eq) and the above acyl chloride (i.e., the above concentrate) in N-methylpyrrolidone was added at 0–10 °C. The reaction was carried out at room temperature for 3 hours, and the reaction was detected by TLC. p-Toluenesulfonic acid (10.84 g, 4.0 eq) was added, and the reaction was stirred at room temperature for 2 hours. Isopropanol (90 ml) was added, and the temperature was lowered to 0–5 °C and stirred for 2 hours. The mixture was filtered, and the filter cake was added to a mixed solvent of acetonitrile (22 ml) and purified water (9 ml) and stirred at 40 °C to dissolve. Acetonitrile (150 ml) was added dropwise, and the mixture was stirred at 0–5 °C to induce crystallization. The mixture was filtered, and the filter cake was dried under vacuum at 60 °C to obtain p-toluenesulfonate of compound (I) with a yield of 63.0% and a purity of 99.2%.

[0218] TGA: Weight loss of 3.44% at 35–166℃; DSC shows two endothermic peaks at 85.36℃ and 248.45℃; Moisture content: 2.6% (KF). See Figure 3 for its TGA-DSC spectrum.

[0219] 1 H NMR(400MHz,d-DMSO)δ11.13(s,1H),9.52(d,J=4.8Hz,1H),9.52(s,1H),8.79-8.77(m,1H),8.45-8.44(d,J=3.6Hz,1H),8 .27-8.26(d,J=5.2Hz,1H),7.50-7.48(m,2H),7.19-7.04(m,6H),6.87-6.14(d,J=7.6Hz,1H),6.46-6.43(d,J=7.6Hz,0.5 H),5.42-5.38(dd,J=12.4,5.2Hz,1H),5.28(s,0.5H),5.08(s,0.5H),4.98-4.93(t,J=8.0Hz,1H),4.79-4.75(d,J=14.4H z,1H),3.83-3.33(m,11H),3.68-3.59(m,5H),3.04-2.83(m,5H),2.66-2.49(m,3H),2.35-2.28(m,6H),2.08-1.80(m,8H).

[0220] Bioactivity testing examples

[0221] The control molecule used in this test example is KT-474, which was prepared according to the preparation method of compound I-417 in patent WO2020113233A1, and its structure is as follows:

[0222] Test Example 1: Degradation of IRAK4 by the compound in SU-DHL-2 and OCI-LY3 cells

[0223] (1) Experimental Objective

[0224] Flow cytometry was used to detect the degradation level of the compound on the IRAK4 protein in the SU-DHL-2 cell line.

[0225] (2) Experimental materials

[0226] Fix Buffer I, BD Phosflow TM 557870

[0227] Perm III buffer, BD Phosflow TM 558050

[0228] LIVE / DEAD TM Fixable Violet Dead Cell Stain Kit, Thermo Fisher, L34966

[0229] Alexa Fluor 647Mouse anti-Human IRAK4 Clone L29-525(RUO), BD Phosflow TM 560315

[0230] (3) Experimental instruments

[0231] Flow cytometer, BD, Model: BD LSRFortessa

[0232] (4) Experimental methods

[0233] Step 1: The SU-DHL-2 cell line was cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 1% penicillin antibiotics.

[0234] Step 2: On Day 1, seed 100 μL of cells into each well of a 96-well plate at a density of 2 x 10⁵. Dissolve the compound in DMSO to a final concentration of 10 mM, then serially dilute it 10 times with culture medium at a 1:5 ratio to achieve a maximum final concentration of 2 μM in the cells. Use wells containing 0.1% DMSO as a positive control. Incubate at 37°C in a 5% CO₂ incubator for 24 hours.

[0235] Step 3: After centrifugation, wash the cells with FACS buffer. Use LIVE / DEAD. TMThe Fixable Blue Dead Cell Stain Kit was used to distinguish between live and dead cells. PE Mouse Anti-Human CD14 was used to stain and label the surface of monocytes, and the cells were incubated at 4°C for 30 minutes.

[0236] Step 4: After centrifugation, wash the cells with FACS buffer. Add an equal volume of Fix buffer I to the cultured cells and fix the cells at 37°C for ten minutes.

[0237] Step 5: After centrifugation, wash the cells with FACS buffer. Add 150 μL of Perm III reagent and incubate at 4°C for 30 minutes to lyse the cell membrane. After centrifugation, wash the cells with FACS buffer.

[0238] Step 6: Stain cells with Alexa Fluor 647 Mouse anti-Human IRAK4 Antibody and incubate at 4°C for 1 hour. Perform flow cytometry analysis.

[0239] (5) Data processing

[0240] Streaming data was analyzed using the Flowjo software. The data displayed included data on DC... 50 The (half-maximum degradation concentration) was calculated using Prism 8.0.2 (GraphPad).

[0241] (6) Experimental Results

[0242] The degradation activity of the compound of formula (I) against IRAK4 is shown in Table 1.

[0243] The compound of this formula (I) generally achieves a Dmax of over 50% for both SU-DHL-2 and OCI-LY3 cells, with a maximum of around 90%.

[0244] The results showed that the compound of formula (I) exhibited good degradation activity against the IRAK4 target in SU-DHL-2 and OCI-LY3 cells.

[0245] Table 1. Degradation results of IRAK4 in SU-DHL-2 and OCI-LY3 cells

[0246] Test Example 2: Degradation of IRAK4 in human PBMCs by the compound

[0247] (1) Experimental Objective

[0248] Flow cytometry was used to detect the degradation level of the compound on IRAK4 in human peripheral blood mononuclear cells (hPBMCs).

[0249] (2) Experimental materials

[0250] Fix Buffer I, BD Phosflow TM 557870

[0251] Perm III buffer, BD Phosflow TM 558050

[0252] LIVE / DEAD TM Fixable Blue Dead Cell Stain Kit, for UV excitation, Thermo Fisher, L34962

[0253] PE Mouse Anti-Human CD14, BD Pharmingen TM 555398

[0254] Alexa Fluor 647 Mouse anti-Human IRAK4 Clone L29-525(RUO), BD Phosflow TM 560315

[0255] (3) Experimental instruments

[0256] Flow cytometer, BD, Model: BD LSRFortessa

[0257] (4) Experimental methods

[0258] After thawing, the frozen hPBMCs were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 1% Pen+Strep, and then incubated at 37°C for 1 hour.

[0259] On day 1, 100 μL of cells were seeded in 96-well plates at a rate of 2 x 10⁵ cells per well.

[0260] The compound was dissolved in DMSO to a final concentration of 10 mM, and then diluted 10 times in culture medium at a 1:5 ratio to achieve a maximum final concentration of 2 μM in cells. Cell wells containing 0.1% DMSO were used as a positive control.

[0261] Incubate at 37°C in an incubator containing 5% CO2 for 24 hours.

[0262] After centrifugation, the cells were washed with FACS buffer.

[0263] Use LIVE / DEAD TM The Fixable Blue Dead Cell Stain Kit was used to distinguish between live and dead cells. PE Mouse Anti-Human CD14 was used to stain and label the surface of monocytes, and the cells were incubated at 4°C for 30 minutes.

[0264] After centrifugation, the cells were washed with FACS buffer.

[0265] Add Fix buffer I in an amount equal to the volume of cultured cells and fix the cells at 37°C for ten minutes.

[0266] After centrifugation, the cells were washed with FACS buffer.

[0267] Add 150 μL of Perm III reagent and incubate at 4°C for 30 minutes to permeate the cell membrane.

[0268] After centrifugation, the cells were washed with FACS buffer.

[0269] Cells were stained with Alexa Fluor 647 Mouse anti-Human IRAK4 Antibody and incubated at 4°C for 1 hour.

[0270] Flow cytometry analysis was performed using a flow cytometer.

[0271] (5) Data processing methods

[0272] Streaming data was analyzed using the Flowjo software. The data displayed included data on DC... 50 The calculations were performed using Prism 8.0.2 (GraphPad).

[0273] (6) Experimental Results

[0274] The degradation activity of the compound of formula (I) against IRAK4 is shown in Table 2.

[0275] The results showed that the compound of formula (I) exhibited good degradation activity against IRAK4 in human PBMC cells.

[0276] Table 2. Degradation results of IRAK4 in human PBMC cells

[0277] Test Example 3: Inhibition of Compound on the Induction of Inflammatory Factor Secretion in Human PBMCs

[0278] (1) Experimental Objective

[0279] The effects of PROTAC small molecules on TLR and IL-1-induced IRAK4 downstream inflammatory factors in PBMC cells were detected using the CBA (micro-sample multi-index flow cytometry) method.

[0280] (2) Experimental materials

[0281] (3) Experimental instruments

[0282] Flow cytometer, BD, Model: BD LSRFortessa

[0283] (4) Experimental methods

[0284] Step 1: Human PBMC Recovery

[0285] Step 2: Adjust the PBMC cell density to 1E6 / mL, and seed 150μL of PBMCs with 1640 intact culture medium into a 96-well plate, and incubate at 37℃ for 1 hour.

[0286] Step 3: Inoculate 50 μL of serially diluted small molecule drug (working concentration: 10000 nM, 1:3 dilution) onto a plate in a 37°C incubator for 8 hours.

[0287] Step 4: Add 10 μL of stimulant, LPS final concentration 100 ng / mL, R848 final concentration 1 μg / mL, IL-1β final concentration 100 ng / mL respectively.

[0288] Step 5: CBA assay. The concentrations of IL6 and TNF-α in the cell culture supernatant were determined using Hu IL-6CBA Flex Set A7 and Hu TNF-CBA Flex Set D9.

[0289] Step 6: Prepare standards: 2500 pg / mL, 1250 pg / mL, ... 0.1:2 serial dilutions, 10 concentrations.

[0290] Step 7: Resuspend the mixed capture beads in capture bead diluent (1:50), add cell culture supernatant (50 μL), and incubate at room temperature for 10 minutes.

[0291] Step 8: Add the detection reagent prepared with the detection diluent and incubate at room temperature in the dark for 3 hours. At the end of the incubation, wash once with washing buffer and resuspend for detection.

[0292] (5) Data processing

[0293] IL-6 and TNFα concentrations were calculated using the standard curve method. Data showed that... 50 The calculations were performed using Prism 8.0.2.

[0294] (6) Experimental Results

[0295] The results of the inhibitory effect of the compound of formula (I) on the induction of inflammatory factor secretion in human PBMCs are shown in Tables 3 and 4.

[0296] The results show that the compound of formula (I) can effectively inhibit the secretion of inflammatory factors and has potential value in treating inflammatory diseases.

[0297] Table 3. LPS, R848, and IL-1β induce TNFα secretion in human PBMC cells.

[0298] Table 4. LPS, R848, and IL-1β induce IL-6 secretion in human PBMC cells.

[0299] Test Example 4: Inhibition of hERG potassium channels by the compound

[0300] (1) Experimental Objective

[0301] The effects of the compound on the current of the hERG potassium channel (human Ether-a-go-go Related Gene potassium channel) were tested using a whole-cell electrophysiological manual voltage clamp method.

[0302] (2) Experimental materials

[0303] Sodium chloride (NaCl) Sigma S7653, Potassium chloride (KCl) Sigma P9333, Magnesium chloride (MgCl2) Sigma M1028

[0304] Calcium chloride (CaCl2) Sigma 21115, Glucose Sigma G7528, HEPES Sigma H3375, EGTA Sigma E3889, Sodium hydroxide (NaOH) Sinopharm 10019718, Potassium hydroxide (KOH) Sinopharm 10017018;

[0305] CHO-hERG cell line (Chinese Hamster Ovary): Chinese hamster ovary cells stably expressing the hERG channel.

[0306] Extracellular fluid formulation (mM): 140 NaCl, 5 KCl, 1 CaCl2, 1.25 MgCl2, 10 HEPES and 10 Glucose, adjusted to pH 7.4 with NaOH.

[0307] Intracellular fluid formulation (mM): 140 KCl, 1 MgCl2, 1 CaCl2, 10 EGTA and 10 HEPES, pH adjusted to 7.2 with KOH.

[0308] (3) Experimental instruments

[0309] Patch clamp amplifier (Multiclamp 700B, Axon, USA)

[0310] Digital-to-analog converter (DigiData 1440A, Axon, USA)

[0311] Inverted microscope (IX71, Olympus, Japan)

[0312] Rapid drug delivery system (RSC-200, Bio-Logic, France)

[0313] Micromanipulator (MX7600R, Syskiyou, USA)

[0314] Electrode drawing apparatus (P-97, Sutter, USA)

[0315] Glass electrode (BF150-86-10, Sutter, USA)

[0316] Vibration damping platform and shielding mesh (63-534, TMC, USA)

[0317] Data acquisition and analysis software (pClamp, Axon, USA)

[0318] CO2 incubator (HERAcell 150i, Thermo, USA)

[0319] Biosafety cabinet (MODEL 1384, Thermo, USA)

[0320] Pure water system (Milli Q, Millipore, USA)

[0321] (4) Experimental methods

[0322] (4.1) Cell culture and treatment

[0323] CHO cells stably expressing hERG were cultured in 35 mm diameter cell culture dishes at 37°C in a 5% CO2 incubator. They were passaged every 48 hours at a 1:5 ratio. The culture medium consisted of 90% F12 (Invitrogen), 10% fetal bovine serum (Gibco), 100 g / mL G418 (Invitrogen), and 100 g / mL Hygromycin B (Invitrogen). On the day of the experiment, the cell culture medium was aspirated, the cells were rinsed once with extracellular fluid, and then 0.25% Trypsin-EDTA (Invitrogen) solution was added for digestion at room temperature for 3-5 minutes. The digestion solution was aspirated, the cells were resuspended in extracellular fluid, and then transferred to experimental dishes for electrophysiological recording.

[0324] (4.2) Compound preparation

[0325] On the day of testing, the compound was prepared as a 10 mM stock solution with DMSO, then diluted to 1 mM with DMSO, and finally diluted 1000-fold with extracellular fluid to obtain the final concentration to be tested. For the positive control compound cisapride, 10 μL of a 150 μM cisapride DMSO stock solution was added to 4990 L of extracellular fluid and diluted 500-fold to obtain the final concentration to be tested, 300 nM. The DMSO content in the final test concentration did not exceed 0.2%, as this concentration of DMSO had no effect on hERG potassium channels.

[0326] (4.3) Electrophysiological recording process

[0327] CHO cells stably expressing the hERG potassium channel were used to record hERG potassium channel currents at room temperature using whole-cell voltage-clamp technique. Glass microelectrodes were fabricated from glass electrode blanks (BF150-86-10, Sutter) using a stretching device. The tip resistance after perfusion with electrode fluid was approximately 2-5 MΩ. The glass microelectrodes were inserted into the amplifier probe to connect to the patch-clamp amplifier. Clamping voltage and data recording were controlled and recorded by computer using pClamp software, with a sampling frequency of 10 kHz and a filtering frequency of 2 kHz. After obtaining whole-cell recordings, the cells were clamped at -100 mV. The step voltage to induce hERG potassium current (IhERG) was applied from -100 mV with a 2-second depolarization voltage to +20 mV, then repolarized to -50 mV, held for 1 second, and then returned to -100 mV. This voltage stimulation was applied every 5 seconds. After confirming that the hERG potassium current was stable (1 minute), the drug delivery process began. For each test concentration of the compound, administer for at least 1 minute to reach steady state or for a maximum of 3 minutes, and test at least 2 cells for each concentration (n≥2).

[0328] (5) Data processing

[0329] Data analysis and processing were performed using pClamp, GraphPad Prism 8, and Excel software. The degree of inhibition of hERG potassium current (the peak hERG tail current induced at -50mV) by different compound concentrations was calculated using the following formula:

[0330] Inhibition%=[1-(I / Io)]×100%

[0331] Where Inhibition% represents the percentage of inhibition of hERG potassium current by the compound, and I and Io represent the amplitude of hERG potassium current before and after drug administration, respectively.

[0332] (6) Experimental Results

[0333] Table 5. Inhibition results of hERG potassium channels

[0334] Note: @1uM refers to the concentration of the test compound.

[0335] Experimental results show that the compound of formula (I) has no significant inhibitory activity on hERG, indicating that the compound of formula (I) has good safety.

[0336] Test Example 5: In vivo pharmacokinetic determination of the compound

[0337] 1.1 Test Drugs:

[0338] This disclosure pertains to compound (I).

[0339] 1.2 Experimental Animals

[0340] Healthy male Beagle dogs, weighing approximately 8-10 kg, 3 dogs per group.

[0341] 1.3 Experimental Design:

[0342] Three Beagle dogs per group were administered the compound at a dose of 2.5 mg / kg via gavage, in a volume of 5 mL / kg, prepared with 10% DMSO + 50% PEG400 + 40% water (w / v). They were allowed free access to food and water.

[0343] Venous blood samples of 0.5-0.8 mL were collected at 15 min, 30 min, 1 h, 2 h, 4 h, 8 h, and 24 h after oral administration. The samples were placed in K2EDTA tubes and centrifuged at 4 °C to separate the plasma. The concentration of compounds in the plasma was determined by liquid chromatography-tandem mass spectrometry.

[0344] 1.4 Experimental Results:

[0345] The main pharmacokinetic parameters were calculated using WinNonlin. The pharmacokinetic parameters of the canine oral drug are shown in Table 6 below. The experimental results show that the compound of formula (I) has good pharmacokinetic properties.

[0346] Table 6. Pharmacokinetic parameters of some compounds disclosed herein orally in dogs

[0347] Test Example 6: Efficacy evaluation of the compound against IL-33-induced acute inflammation in mice

[0348] (1) Experimental Objective

[0349] IL-33 is a member of the IL-1 cytokine family and a ligand for IL-1 receptor-like 1 or ST2. IL-33 transgenic mice specifically express IL-33 in keratinocytes and spontaneously develop AD (atopic dermatitis)-like eczema, suggesting that the IL-33 pathway plays a crucial role in AD. This study investigated the effects of compounds on inflammation along the IL-1R signaling pathway using an acute inflammatory mouse model induced by intraperitoneal injection of IL-33.

[0350] (2) Experimental materials and methods

[0351] Experimental animals: SPF-grade 6-8 week old male C57BL / 6J mice.

[0352] Compound formulation: 10% DMSO + 50% PEG400 + 40% sterile water for injection. The compound was prepared by dissolving it in DMSO, followed by the addition of PEG400 and sterile water for injection, and adjusting the solution to clear with 6N hydrochloric acid.

[0353] Preparation of IL-33 inducer: Prepare a 1 μg / μL stock solution of recombinant IL-33 powder from mice. When using, add 3960 μL of PBS to 40 μL of the 1 μg / μL IL-33 solution, invert and mix well to obtain 4 mL of IL-33 solution with a concentration of 1 μg / 100 μL. Prepare and use immediately.

[0354] Compound administration and IL-33 inflammation induction: Mice were randomly divided into groups according to body weight and orally administered the compound at final doses of 15 mg / kg, 50 mg / kg, and 150 mg / kg twice daily. Four hours after the last administration of the compound, mice were intraperitoneally injected with IL-33 1 μg / 100 μL PBS to induce inflammation.

[0355] Termination, Sample Collection and Data: Plasma, peritoneal lavage fluid and spleen were collected for subsequent sample indicator testing.

[0356] Eight hours after the last administration, the experiment was terminated. Blood was collected from the heart of anesthetized mice. After anticoagulation with EDTA-K2, the blood samples were centrifuged at 3500 rpm for 15 min, and the supernatant plasma was collected. Euthanized mice were then disinfected by immersion in 75% alcohol for 3 min in a sterile laminar flow hood. 2.5 mL of pre-cooled PBS (4°C) was injected intraperitoneally, and the abdomen was massaged for 5 min. The abdominal skin was then cut open with sterile scissors and forceps to expose the peritoneum. The sternum was lifted at the xiphoid process using sterile forceps, and a 1 mm incision was made in the peritoneum at the xiphoid process. 2 A small incision was made, and a pipette was inserted into the peritoneal cavity to gently blow and collect 2 mL of the lavage fluid. The fluid was then aliquoted into 1.5 mL centrifuge tubes and stored at -80°C. The expression levels of inflammatory factors in mouse plasma and peritoneal lavage fluid were detected by ELISA, and the expression level of IRAK4 protein in mouse spleen was detected by LC-MS.

[0357] (3) Experimental Results and Analysis

[0358] Table 7. Endpoint plasma and tissue drug concentrations

[0359] Table 7 shows that the concentrations of compound (I) in plasma and spleen at a dose of 50 mpk were the same as those of KT-474 at 150 mpk, indicating that compound (I) requires a lower dosage.

[0360] Figure 1 shows the concentration-dependent decrease of inflammatory factors in mouse plasma and peritoneal lavage fluid after three PO doses of compound (I). At comparable plasma drug concentrations, compound (I) exhibited superior anti-inflammatory effects compared to KT474.

[0361] Figure 2 illustrates the in vivo degradation of IRAK4 in mouse spleens after three PO doses of compound (I). At comparable spleen drug concentrations, compound (I) showed 2-3 times better degradation at 50 mpk than KT-474 at 150 mpk.

[0362] Although certain exemplary embodiments and specific examples have been described above, other embodiments and modifications will be apparent from the foregoing description. Technical features described in one embodiment may be combined with technical features described in another embodiment without departing from the inventive concept of this disclosure. Therefore, the inventive concept of this disclosure is not limited to the specifically given embodiments and examples, but is limited to the claims and various equivalents that will be apparent to those skilled in the art.

Claims

1. A method for preparing a compound of formula (I), its salt, or a stereoisomer thereof, comprising condensing a compound of formula (II) or its salt with a compound of formula (III), its salt, or a stereoisomer thereof to obtain the compound of formula (I), its salt, or a stereoisomer thereof. in, Y is selected from OH and halogens.

2. The preparation method according to claim 1, characterized in that, The compound of formula (III), its salt, or its stereoisomer is obtained by reduction of the compound of formula (IV), its salt, or its stereoisomer.

3. The preparation method according to claim 2, characterized in that, The compound of formula (IV), its salt, or its stereoisomer is obtained by reacting the compound of formula (VI), its salt, or its stereoisomer with the compound of formula (V) or its salt. Where X is a leaving group.

4. The preparation method according to any one of claims 1-3, characterized in that, Y is selected from halogens; preferably, Y is chlorine.

5. The preparation method according to claim 3 or 4, characterized in that, X is selected from halogens, R 1 Selected from 1-2 Rs 1a Replacement C 1-4 Alkyl, phenyl; R 1a Selected from halogens, -NO2, C 1-4 alkyl; Preferably, X is selected from halogens or CH3SO3-; Preferably, X is selected from Br or CH3SO3-.

6. The preparation method according to any one of claims 1-5, characterized in that, When Y is OH in the compound of formula (II), the compound of formula (II) or its salt is condensed with the compound of formula (III), its salt or its stereoisomer under the conditions of basic catalyst A1, reaction solvent B1 and condensing agent C1 to obtain the compound of formula (I), its salt or its stereoisomer; or, When Y is a halogen in the compound of formula (II), the compound of formula (II) or its salt is condensed with the compound of formula (III), its salt or its stereoisomer under the conditions of alkaline catalyst A1' and reaction solvent B1' to obtain the compound of formula (I), its salt or its stereoisomer; The alkaline catalysts A1 and A1' are independently selected from organic or inorganic bases, preferably from one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate. The reaction solvents B1 and B1' are independently selected from one or more of tetrahydrofuran, N,N-dimethylformamide, N-methylpyrrolidone, acetonitrile, chloroform, dichloromethane, and toluene; The condensing agent C1 is selected from one or more of the following: 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU), N,N,N',N'-tetramethyl-O-(benzotriazole-1-yl)thiourea hexafluorophosphate (HBTU), O-benzotriazole-N,N,N',N'-tetramethylurea tetrafluoroborate (TBTU), Carter condensing agent (BOP), benzotriazole-1-yl-oxytripyrrolylphosphonium hexafluorophosphate (PyBOP), (7-azabenzotriazole-1-oxy)tripyrrolylphosphonium hexafluorophosphate (PyAOP), carbonylimidazolium (CDI), 1-hydroxybenzotriazole (HOBt), and mixed acid anhydrides.

7. The preparation method according to claim 6, characterized in that, The alkaline catalyst A1 is selected from organic bases, preferably N,N-diisopropylethylamine; the reaction solvent B1 is N,N-dimethylformamide; the condensing agent C1 is selected from 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate, 1-hydroxybenzotriazole, or combinations thereof; or... The alkaline catalyst A1' is selected from organic bases, preferably triethylamine; the reaction solvent B1' is selected from tetrahydrofuran, N-methylpyrrolidone, or a combination thereof.

8. The preparation method according to any one of claims 1-7, characterized in that, The reaction conditions of the compound of formula (II) or its salt with the compound of formula (III), its salt or its stereoisomer include one or more of the following: (a) The molar ratio of the compound of formula (II) or its salt to the compound of formula (III), its salt or its stereoisomer is 1 to 2:1, preferably 1 to 1.5:1; (b) The molar ratio of the compound of formula (III), its salt or stereoisomer or thereof to each of the condensing agents C1 is 1:1 to 1.5; (c) The molar ratio of the compound of formula (III), its salt or its stereoisomer to the basic catalyst A1 is 1:1 to 5, preferably 1:2 to 4, and more preferably 1:

3.

9. The preparation method according to any one of claims 1-7, characterized in that, The reaction conditions of the compound of formula (II) or its salt with the compound of formula (III), its salt or its stereoisomer include one or more of the following: (a) The molar ratio of the compound of formula (II) or its salt to the compound of formula (III), its salt or its stereoisomer is 1 to 2:1, preferably 1 to 1.5:1; (b) The molar ratio of the compound of formula (III), its salt or its stereoisomer to the basic catalyst Al' is 1:1 to 5, preferably 1:1 to 1.

5.

10. The preparation method according to any one of claims 1-9, characterized in that, The compound of formula (IV), its salt, or its stereoisomer is reduced under the conditions of reducing agent A2, reaction solvent B2, and acidic reagent E2 to obtain the compound of formula (III), its salt, or its stereoisomer. The reducing agent A2 is selected from one or more of iron powder, zinc powder, and sodium thiosulfate. The reaction solvent B2 is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, acetonitrile, ethanol, and dioxane; The acidic reagent E2 is selected from one or more of ammonium chloride, hydrochloric acid, and acetic acid.

11. The preparation method according to claim 10, characterized in that, The reducing agent A2 is zinc powder; the reaction solvent B2 is selected from tetrahydrofuran, acetonitrile, or a combination thereof; and the acidic reagent E2 is ammonium chloride.

12. The preparation method according to any one of claims 1-11, characterized in that, The reaction conditions for reducing the compound of formula (IV), its salt, or its stereoisomer to obtain the compound of formula (III), its salt, or its stereoisomer include a molar ratio of formula (IV) to reducing agent A2 of 1:1 to 20; preferably 1:8 to 12; preferably 1:

10.

13. The preparation method according to any one of claims 1-12, characterized in that, The compound of formula (VI), its salt, or its stereoisomer reacts with the compound of formula (V) or its salt under the conditions of alkaline catalyst A3 and reaction solvent B3 to obtain the compound of formula (IV), its salt, or its stereoisomer. The alkaline catalyst A3 is selected from one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate. The reaction solvent B3 is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, acetonitrile, and ethyl acetate.

14. The preparation method according to claim 13, characterized in that, The alkaline catalyst A3 is sodium bicarbonate; the reaction solvent B3 is acetonitrile.

15. The preparation method according to any one of claims 1-14, characterized in that, The reaction conditions between the compound of formula (VI), its salt, or its stereoisomer with the compound of formula (V) or its salt include one or more of the following: (a) The molar ratio of the compound of formula (VI), its salt or its stereoisomer to the compound of formula (V) or its salt is 1:0.2 to 2, preferably 1:0.8 to 1; (b) The molar ratio of the compound of formula (VI), its salt or its stereoisomer to the base catalyst A3 is 1:1 to 2, preferably 1:1 to 1.

5.

16. A method for preparing a compound of formula (I), a salt thereof, or a stereoisomer thereof, comprising reacting a compound of formula (VII) or a salt thereof with a compound of formula (VI), a salt thereof, or a stereoisomer thereof to obtain the compound of formula (I), a salt thereof, or a stereoisomer thereof. in, X is selected from leaving groups.

17. The preparation method according to claim 16, characterized in that, X is selected from halogens, R 1 Selected from 1-2 Rs 1a Replacement C 1-4 Alkyl, phenyl; R 1a Selected from halogens, -NO2, C 1-4 alkyl; Preferably, X is selected from I or CH3SO3-.

18. The preparation method according to claim 16 or 17, characterized in that, Formula (VII) or its salt is obtained by removing the protecting group from formula (VIII) or its salt. PG is an amino protecting group; Preferably, PG is selected from (a) alkoxycarbonyl amino protecting groups, such as benzyloxycarboxyl (Cbz), tert-butyloxycarbonyl (Boc), methoxycarbonyl (Fmoc), allyloxycarbonyl (Alloc), trimethylsilylethoxycarbonyl (Teoc), methoxycarboxyl, or ethoxycarboxyl; (b) acyl amino protecting groups, such as phthaloyl (Pht), p-toluenesulfonyl (Tos), trifluoroacetyl (Tfa), o-(p-)nitrobenzenesulfonyl (Ns), tert-valeryl, benzoyl; (c) alkyl amino protecting groups, such as triphenylmethyl (Trt), 2,4-dimethoxybenzyl (Dmb), p-methoxybenzyl (PMB), benzyl (Bn); and (d) organosilicon protecting groups, such as trimethylsilyl (TMS), tert-butyldimethylsilyl (TBDMS), triisopropylsilyl (TIPS), diethylisopropylsilyl (DEIPS). Preferably, PG is an alkoxycarbonyl amino protecting group, and more preferably tert-butyloxycarbonyl (Boc).

19. The preparation method according to any one of claims 16-18, characterized in that, The compound of formula (VIII) or a salt thereof is obtained by condensation of the compound of formula (II) or a salt thereof with the compound of formula (IX) or a salt thereof; in, PG is an amino protecting group; Y is selected from OH and halogens; Preferably, Y is selected from halogens.

20. The preparation method according to any one of claims 16-19, characterized in that, The compound of formula (VI), its salt, or its stereoisomer reacts with the compound of formula (VII) or its salt under the conditions of alkaline catalyst A4 and reaction solvent B4 to give the compound of formula (I), its pharmaceutically acceptable salt, or its stereoisomer. The alkaline catalyst A4 is selected from one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate. The reaction solvent B4 is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, acetonitrile, and ethyl acetate.

21. The preparation method according to claim 20, characterized in that, The alkaline catalyst A4 is potassium bicarbonate; the reaction solvent B4 is acetonitrile.

22. The preparation method according to any one of claims 16-21, characterized in that, The reaction conditions of the compound of formula (VI), its salt or its stereoisomer with the compound of formula (VII) or its salt include one or more of the following: (a) The molar ratio of the compound of formula (VI), its salt or its stereoisomer to the compound of formula (VII) or its salt is 1:0.2 to 2, preferably 1:0.8 to 1; (b) The molar ratio of the compound of formula (VI), its salt or its stereoisomer to the basic catalyst A4 is 1:1 to 10, preferably 1:4 to 6, and more preferably 1:

5.

23. The preparation method according to any one of claims 16-22, characterized in that, The protecting group of formula (VIII) or its salt is removed by catalytic hydrogenolysis, acidolysis, or base-catalytic cleavage to obtain formula (VII) or its salt. The acid used in the acid hydrolysis is selected from organic or inorganic acids, preferably from one or more of hydrochloric acid, trifluoroacetic acid, acetic acid, and hydrobromic acid; the base used in the base-catalyzed hydrolysis is selected from organic or inorganic bases, preferably from one or more of potassium hydroxide, sodium hydroxide, potassium carbonate, triethylamine, piperidine, tetrabutylammonium fluoride, and hydrazine hydrate.

24. The preparation method according to any one of claims 16-23, characterized in that, Formula (VIII) or its salt is reacted in acidic reagent A5 and reaction solvent B5 to obtain formula (VII) or its salt; The acidic reagent A5 is selected from organic acids, preferably trifluoroacetic acid; The reaction solvent B5 is selected from one or more of dichloromethane, trichloromethane, tetrahydrofuran, acetonitrile, 1,4-dioxane, methanol, ethanol, isopropanol, ethyl acetate, isopropyl acetate, N,N-dimethylformamide, N-methylpyrrolidone, acetone, and butanone; preferably dichloromethane.

25. The preparation method according to any one of claims 16-24, characterized in that, The reaction conditions for obtaining formula (VII) or its salt by removing the protecting group from formula (VIII) or its salt include a ratio (mol / ml) of the molar amount of formula (VIII) or its salt to the volume of the acidic reagent A5 of 1:1 to 3; preferably 1:

2.

26. The preparation method according to any one of claims 16-25, characterized in that, When Y is OH in the compound of formula (II), the compound of formula (IX) or its salt reacts with the compound of formula (II) or its salt under the conditions of alkaline catalyst A6, reaction solvent B6, and condensing agent C2 to obtain the compound of formula (VIII) or its salt. or, When Y is a halogen in the compound of formula (II), the compound of formula (II) or its salt reacts with the compound of formula (IX) or its salt under the conditions of alkaline catalyst A6' and reaction solvent B6' to obtain the compound of formula (VIII) or its salt; The alkaline catalysts A6 and A6' are selected from organic or inorganic bases, preferably from one or more of triethylamine, N,N-diisopropylethylamine, pyridine, sodium carbonate, sodium bicarbonate, and potassium bicarbonate. The reaction solvents B6 and B6' are selected from one or more of tetrahydrofuran, N,N-dimethylformamide, N-methylpyrrolidone, acetonitrile, chloroform, dichloromethane, and toluene; The condensing agent C2 is selected from one or more of the following: 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU), N,N,N',N'-tetramethyl-O-(benzotriazole-1-yl)thiourea hexafluorophosphate (HBTU), O-benzotriazole-N,N,N',N'-tetramethylurea tetrafluoroborate (TBTU), Carter condensing agent (BOP), benzotriazole-1-yl-oxytripyrrolylphosphonium hexafluorophosphate (PyBOP), (7-azabenzotriazole-1-oxy)tripyrrolylphosphonium hexafluorophosphate (PyAOP), carbonylimidazolium (CDI), 1-hydroxybenzotriazole (HOBt), and mixed acid anhydrides.

27. The preparation method according to claim 26, characterized in that, The alkaline catalyst A6 is selected from organic bases, preferably N,N-diisopropylethylamine; the reaction solvent B6 is selected from N,N-dimethylformamide; the condensing agent C2 is selected from 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole, or a combination thereof; or, The alkaline catalyst A6' is selected from organic bases, preferably triethylamine; the reaction solvent B6' is selected from tetrahydrofuran, N-methylpyrrolidone, or a combination thereof.

28. The preparation method according to any one of claims 16-27, characterized in that, The reaction conditions between the compound of formula (IX) or a salt thereof and the compound of formula (II) or a salt thereof include one or more of the following: (a) The molar ratio of the compound of formula (IX) or its salt to the compound of formula (II) or its salt is 1:1 to 1.5; (b) The molar ratio of the compound of formula (IX) or its salt to each of the condensing agents C2 is 1:1 to 2; preferably 1:1 to 1.5; (c) The molar ratio of the compound of formula (IX) or its salt to the basic catalyst A6 is 1:1 to 5; the molar ratio is 1:2 to 4; preferably 1:

2.

29. A method for preparing p-toluenesulfonate of the compound of formula (I) according to any one of claims 1-15, comprising adding p-toluenesulfonic acid to a reaction system of the compound of formula (I) and reacting to obtain the p-toluenesulfonate of the compound of formula (I).

30. The preparation method according to claim 29, characterized in that, The p-toluenesulfonate is prepared by condensing a compound of formula (II) or its salt with a compound of formula (III), its salt or its stereoisomer, and then adding p-toluenesulfonic acid to the reaction system to form a salt, wherein the reaction step of the compound of formula (II) or its salt with the compound of formula (III), its salt or its stereoisomer is as described in any one of claims 1-15.

31. The preparation method according to claim 29 or 30, characterized in that, The solvent for the salt formation reaction is selected from one or more of tetrahydrofuran, N,N-dimethylformamide, N-methylpyrrolidone, acetonitrile, chloroform, dichloromethane, toluene, isopropanol, and ethanol.

32. The preparation method according to any one of claims 29-31, characterized in that, The salt formation reaction conditions of the compound of formula (I) and the p-toluenesulfonic acid include a molar ratio of the compound of formula (I) to the p-toluenesulfonic acid of 1:1 to 10; preferably 1:2 to 6.

33. The preparation method according to any one of claims 29-32, characterized in that, In the p-toluenesulfonate compound of formula (I), the molar ratio of the compound of formula (I) to the p-toluenesulfonic acid is 1:

1.

34. The preparation method according to any one of claims 29-33, characterized in that, The compound of formula (I) is a hydrated p-toluenesulfonate. Preferably, the compound of formula (I) p-toluenesulfonate is a monohydrate.

35. The following compounds, their salts, or their stereoisomers, in, PG is an amino protecting group, as defined in claim 18.

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