Benzimidazole spleen tyrosine kinase inhibitor as well as preparation method and application thereof

Abstract

The invention relates to the field of antitumor drugs, in particular to a benzimidazole spleen tyrosine kinase inhibitor as well as a preparation method and application thereof. The inhibitor has a molecular structure as shown in a formula 1, in the formula 1, an R0 group is an R1 group containing an amide group, the R1 group is selected from at least one of the following groups, and an R3 group is selected from at least one of the following groups: H, F, Cl and Br; according to the benzimidazole inhibitor, the structure of the benzimidazole inhibitor can be optimized by introducing the R1 group of acylamino and the R2 group of multiple cyclic structures, so that efficient selective inhibition on spleen tyrosine kinase (Syk) is realized.

Description

Technical Field

[0001] The present application relates to the field of anti-tumor drugs, and in particular to a benzimidazole spleen tyrosine kinase inhibitor, a preparation method thereof, and an application thereof. Background Art

[0002] Spleen Tyrosine Kinase (SYK) is a non-receptor intracellular kinase that plays a key role in the B cell receptor (BCR) signaling pathway and immune regulation. SYK is abnormally activated or overexpressed in a variety of malignant tumors (such as diffuse large B-cell lymphoma, acute myeloid leukemia-like tumors, breast cancer tumors, etc.). SYK generally participates in cancer progression by promoting tumor cell proliferation, survival and metastasis. In addition, SYK can also indirectly affect the anti-tumor immune response by regulating the function of immune cells in the tumor microenvironment. Current studies have shown that abnormal expression of SYK is closely related to a variety of pathological processes, such as tumorigenesis, cardiovascular disease, cerebrovascular disease and neurodegenerative diseases. In addition, the SYK signaling pathway shows significant functional disorders in many human diseases. Based on these research findings, it can be clarified that the SYK signaling pathway is an important molecular target for the treatment of autoimmune diseases, malignant tumors and allergic diseases.

[0003] Current inhibitors targeting SYK either have low selectivity, poor pharmacokinetic properties, or high toxicity. These side effects result in poor inhibitory effects of existing SYK inhibitors. Summary of the Invention

[0004] The present application provides a benzimidazole spleen tyrosine kinase inhibitor and its preparation method and application, in order to solve the following technical problem: how to improve the inhibitory effect of spleen tyrosine kinase inhibitor.

[0005] In a first aspect, the present invention provides a benzimidazole spleen tyrosine kinase inhibitor having a molecular structure as shown in Formula 1.

[0006]

[0007] In Formula 1, the R0 group is an R1 group containing an amide group, and the R1 group is selected from at least one of the following:

[0008]

[0009] The R3 group is selected from at least one of the following: H, F, Cl and Br;

[0010] The R2 group is selected from at least one of the following:

[0011] Optionally, the inhibitor has a molecular structure as shown in Formula 2,

[0012]

[0013] Optionally, the inhibitor has a molecular structure as shown in Formula 3,

[0014]

[0015] Optionally, the inhibitor has a molecular structure as shown in Formula 4,

[0016]

[0017] In a second aspect, the present invention provides a method for preparing the inhibitor according to the first aspect, the method comprising:

[0018] Aniline containing an R2 group and 1,1-thiocarbonyldiimidazole are subjected to a first reaction to obtain a first intermediate;

[0019] Conducting a second reaction between 4-nitro-o-phenylenediamine, 1,3-dicyclohexylcarbodiimide and the first intermediate to obtain a second intermediate;

[0020] A third intermediate is obtained by reacting oxalyl chloride, triethylamine, 4-dimethylaminopyridinepyrazole and a carboxylic acid containing an R1 group;

[0021] subjecting tetrahydroxydiboron, 4-4′-bipyridine, the second intermediate and the third intermediate to a fourth reaction to obtain an inhibitor;

[0022] Wherein, the first intermediate has a molecular structure as shown in Formula 5,

[0023]

[0024] The second intermediate has a molecular structure as shown in Formula 6,

[0025]

[0026] The third intermediate has a molecular structure as shown in Formula 7,

[0027]

[0028] Optionally, the temperature of the first reaction is 20° C. to 30° C., and the time of the first reaction is ≥ 4 h; and / or

[0029] The temperature of the second reaction is 80° C. to 85° C., and the time of the second reaction is ≥16 h; and / or

[0030] The temperature of the third reaction is 20° C. to 30° C., and the time of the third reaction is ≥ 2 hours; and / or

[0031] The temperature of the fourth reaction is 100° C. to 105° C., and the time of the fourth reaction is 24 h to 26 h.

[0032] Optionally, the amount n1 of the aniline and the amount n2 of the 1,1-thiocarbonyldiimidazole satisfy: n1:n2=2:1; and / or

[0033] The amount n3 of the 4-nitro-o-phenylenediamine, the amount n4 of the 1,3-dicyclohexylcarbodiimide, and the amount n5 of the first intermediate satisfy: n3:n4:n5=1.6:1.92:1.92; and / or

[0034] The amount n6 of the oxalyl chloride, the amount n7 of the triethylamine, the amount n8 of the 4-dimethylaminopyridinepyrazole and the amount n9 of the carboxylic acid satisfy: n6:n7:n8:n9=7.5:10.0:1.25:5.0.

[0035] Optionally, the amount n10 of the tetrahydroxydiboron, the amount n11 of the 4-4′-bipyridine, the amount n12 of the second intermediate and the amount n13 of the third intermediate satisfy: n10:n11:n12:n13=0.8:0.01:0.2:0.4.

[0036] In a third aspect, an embodiment of the present application provides a pharmaceutical composition for inhibiting spleen tyrosine kinase, wherein the pharmaceutical composition comprises the inhibitor described in the first aspect.

[0037] Optionally, the mass of the inhibitor is greater than or equal to 80% of the pharmaceutical composition.

[0038] The above technical solution provided by the embodiment of the present application has the following advantages compared with the prior art:

[0039] The present application provides a benzimidazole spleen tyrosine kinase inhibitor. The inhibitor uses a benzimidazole compound as a skeleton, introduces an amide R1 group, and limits the R1 group to a phenyl group and the R2 group to a variety of cyclic structures such as an NO ring, an NN ring, or a benzene ring. These phenyl groups and cyclic structure groups can improve the functionality of the benzimidazole compound skeleton, enabling the inhibitor to specifically bind to tumor cells, thereby enhancing the inhibitory effect of the spleen tyrosine kinase inhibitor. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the present application and, together with the description, serve to explain the principles of the present application.

[0041] In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the following is a brief introduction to the drawings required for use in the embodiments or the description of the prior art. Obviously, for ordinary technicians in this field, other drawings can be obtained based on these drawings without any creative work.

[0042] Figure 1 A schematic flow chart of a method for preparing an inhibitor provided in an embodiment of the present application;

[0043] Figure 2 A schematic diagram of the principle of a method for preparing an inhibitor provided in an embodiment of the present application;

[0044] Figure 3 A schematic diagram of the molecular structure of a benzimidazole spleen tyrosine kinase inhibitor provided in Examples 1 to 14 of the present application;

[0045] Figure 4 A schematic diagram of the molecular structure of another benzimidazole spleen tyrosine kinase inhibitor provided in Examples 15 to 28 of the present application;

[0046] Figure 5 This is a schematic diagram of the molecular structure of another benzimidazole spleen tyrosine kinase inhibitor provided in Examples 29 to 42 of the present application. DETAILED DESCRIPTION

[0047] To make the purpose, technical solutions, and advantages of the embodiments of this application more clear, the technical solutions in the embodiments of this application will be clearly and completely described below in conjunction with the drawings in the embodiments of this application. Obviously, the described embodiments are part of the embodiments of this application, not all of the embodiments. Based on the embodiments in this application, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of this application.

[0048] Various embodiments of the present application may be presented in the form of a range; it should be understood that the description in the form of a range is only for convenience and brevity and should not be understood as a rigid limitation on the scope of the present application; therefore, the range description should be considered to have specifically disclosed all possible sub-ranges and single numerical values ​​within the range; for example, the range description from 1 to 6 should be considered to have specifically disclosed sub-ranges, such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., as well as single numbers within the range, such as 1, 2, 3, 4, 5 and 6, which apply regardless of the range; in addition, whenever a numerical range is indicated in this document, it is meant to include any cited number (fractional or integer) within the indicated range.

[0049] As used herein, the terms "including," "comprising," and the like mean "including but not limited to." Relational terms such as "first" and "second" are used solely to distinguish one entity or operation from another, and do not necessarily require or imply any actual relationship or order between these entities or operations. "And / or" describes the association relationship between associated objects, indicating that three relationships can exist. For example, "A and / or B" can mean: A exists alone, A and B exist at the same time, and B exists alone; where A and B can be singular or plural. "At least one" means one or more, "plurality" means two or more; "at least one," "at least one of the following," or similar expressions, refers to any combination of these items, including any combination of single or plural items; for example, "at least one of a, b, or c," or "at least one of a, b, and c," can all mean: a, b, c, ab (i.e., a and b), ac, bc, or abc, where a, b, and c can each be single or plural. "Parts" notation, such as parts by weight and parts by mass, indicates the proportional relationship between components. In this article, the parameters described by ratio should be understood as the first term of the proportional formula, in the order in which they are described, and the proportional figures should be understood as the second term. For example, if the mass ratio of substances A, B, and C is 1:2:3, then substances A, B, and C should correspond to the proportional figures in the proportional formula in the order in which they are described, that is, the mass of substance A: the mass of substance B: the mass of substance C = 1:2:3.

[0050] Unless otherwise specified, all raw materials, reagents, instruments and equipment used in this article can be purchased from the market or prepared by existing methods.

[0051] The present invention provides a benzimidazole spleen tyrosine kinase inhibitor having a molecular structure as shown in Formula 1.

[0052]

[0053] In Formula 1, the R0 group is an R1 group containing an amide group, and the R1 group is selected from at least one of the following:

[0054] The R3 group is selected from at least one of the following: H, F, Cl and Br;

[0055] The R2 group is selected from at least one of the following:

[0056] It should be noted that the inhibitor may be a compound having a molecular structure as shown in Formula 1, or a pharmaceutically acceptable salt of such a compound.

[0057] It should be noted that the inhibitor can be prepared into at least one of the following forms: tablets, capsules, aqueous solutions or oily solutions; can also be prepared into at least one of the following forms: suspensions, emulsions, creams, ointments, gels, nasal sprays and suppositories; can also be prepared into at least one of the following forms: finely dispersed powders for inhalation, aerosols for inhalation and sprays for inhalation; can also be prepared into at least one of the following forms: sterile aqueous solutions, oily solutions, suspensions and sterile emulsions for extra-gastrointestinal administration (including intravenous, intramuscular or infusion).

[0058] The inhibitor can also be prepared as a liquid formulation using sterile water or a water-propylene glycol solution as a solvent, and the active ingredient of the inhibitor can also be formulated in a polyethylene glycol aqueous solution. The inhibitor can also be an aqueous solution for oral administration or an aqueous suspension for oral administration; wherein the aqueous solution for oral administration can be prepared by dissolving the active ingredient in water and adding appropriate colorants, flavorings, stabilizers, and thickeners as needed. The aqueous suspension for oral administration can be prepared by dispersing the finely divided active ingredient in water together with a viscous substance. The viscous substance used in this process can be a natural or synthetic gum, resin, methylcellulose, carboxymethylcellulose, or other suspending agents known in the pharmaceutical field.

[0059] It should be noted that the present invention provides a benzimidazole spleen tyrosine kinase inhibitor, which achieves efficient and selective inhibition of spleen tyrosine kinase (Syk) through structural optimization. The specific mechanism is as follows:

[0060] 1. Backbone modification strategy: Using benzimidazole as the core, the R1 or R2 groups are linked via amide groups, where R1 uses a phenyl group to enhance the hydrophobic interaction between the inhibitor and the kinase ATP binding domain;

[0061] 2. Ring structure design: R2 introduces an NO ring (e.g., morpholino), an NN ring (e.g., piperazinyl), or a benzene ring to improve the specificity of inhibitor binding to the target through hydrogen bonding networks and π-π stacking interactions;

[0062] 3. Structure-activity relationship: The molecular structure of this inhibitor can precisely occupy the allosteric regulatory site of Syk kinase, achieving high inhibitory activity;

[0063] 4. Tumor targeting: Phenyl groups can adjust the logP value of the compound, which can not only ensure the inhibitor's ability to penetrate the blood-brain barrier, but also avoid the nonspecific distribution of the inhibitor.

[0064] In some optional embodiments, the inhibitor has a molecular structure as shown in Formula 2,

[0065]

[0066]

[0067] In these embodiments, the binding site of the R1 group containing an amide group is clearly defined based on the molecular structure of Formula 1, which can further refine the molecular structure of the inhibitor, so that the inhibitor can improve its inhibitory effect through the phenyl group of the R1 group.

[0068] In some optional embodiments, the inhibitor has a molecular structure as shown in Formula 3,

[0069]

[0070] In these embodiments, the binding site of the R2 group containing an amide group is clarified based on the molecular structure of Formula 1, and the molecular structure of the inhibitor can be further refined, so that the inhibitor can improve its inhibitory effect through various ring structures of the NO ring, NN ring or benzene ring of the R1 group.

[0071] In some optional embodiments, the inhibitor has a molecular structure as shown in Formula 4,

[0072]

[0073] In these embodiments, based on the structure of Formula 1, the specific types of the R1 group and the R2 group and the molecular structure of the inhibitor with the best inhibitory effect can be determined.

[0074] Figure 1 The following is a schematic diagram of a process for preparing an inhibitor provided in an embodiment of the present application;

[0075] Figure 2 The schematic diagram of the principle of the preparation method of an inhibitor provided in the examples of the present application is exemplarily shown;

[0076] Based on a general inventive concept, such as Figure 1 and Figure 2 As shown, the present invention provides a method for preparing the inhibitor, which comprises:

[0077] S1. Aniline containing an R2 group and 1,1-thiocarbonyldiimidazole are reacted to obtain a first intermediate;

[0078] S2. 4-nitro-o-phenylenediamine, 1,3-dicyclohexylcarbodiimide and the first intermediate are subjected to a second reaction to obtain a second intermediate;

[0079] S3. A third reaction is performed with oxalyl chloride, triethylamine, 4-dimethylaminopyridinepyrazole and a carboxylic acid containing an R1 group to obtain a third intermediate;

[0080] S4. subjecting tetrahydroxydiboron, 4-4′-bipyridine, the second intermediate and the third intermediate to a fourth reaction to obtain an inhibitor;

[0081] Wherein, the first intermediate has a molecular structure as shown in Formula 5,

[0082]

[0083] The second intermediate has a molecular structure as shown in Formula 6,

[0084]

[0085] The third intermediate has a molecular structure as shown in Formula 7,

[0086]

[0087] This preparation method is a preparation method for the above-mentioned inhibitor. The specific structure of the inhibitor can refer to the above-mentioned embodiment. Since this preparation method adopts part or all of the technical solutions of the above-mentioned embodiment, it at least has all the beneficial effects brought by the technical solutions of the above-mentioned embodiment, which will not be described one by one here.

[0088] In some optional embodiments, the temperature of the first reaction is 20° C. to 30° C., and the time of the first reaction is ≥ 4 h; and / or

[0089] The temperature of the second reaction is 80° C. to 85° C., and the time of the second reaction is ≥16 h; and / or

[0090] The temperature of the third reaction is 20°C to 30°C, and the time of the third reaction is ≥ 2h; and / or

[0091] The temperature of the fourth reaction is 100° C. to 105° C., and the time of the fourth reaction is 24 h to 26 h.

[0092] In these embodiments, the first reaction at a temperature of 20°C to 30°C and a time of ≥4h can allow aniline containing an R2 group and 1,1-thiocarbonyldiimidazole to fully react to ultimately form a first intermediate with a molecular structure as shown in Formula 5. In addition, the second reaction at a temperature of 80°C to 85°C and a time of ≥16h can allow 4-nitro-o-phenylenediamine, 1,3-dicyclohexylcarbodiimide and the first intermediate to fully react to ultimately form a second intermediate with a molecular structure as shown in Formula 6. In addition, the third reaction at a temperature of 20°C to 30°C and a time of ≥2h can allow oxalyl chloride, triethylamine, 4-dimethylaminopyridinepyrazole and a carboxylic acid containing an R1 group or an R2 group to fully react to ultimately form a third intermediate with a molecular structure as shown in Formula 7. In addition, the fourth reaction with a temperature of 100° C. to 105° C. and a time of 24 h to 26 h can allow tetrahydroxydiboron, 4-4′-bipyridine, the second intermediate and the third intermediate to fully react to finally form an inhibitor product.

[0093] The temperature of the first reaction can be 20°C, 21°C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C or 30°C.

[0094] The time of the first reaction can be 4 h, 4.5 h, 5.0 h, 5.5 h or 6 h.

[0095] The temperature of the second reaction can be 80°C, 81°C, 82°C, 83°C, 84°C, or 85°C.

[0096] The time of the second reaction can be 16 h, 16.5 h, 17 h, 17.5 h or 18 h.

[0097] The temperature of the third reaction can be 20°C, 21°C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C or 30°C.

[0098] The time of the third reaction can be 2 h, 2.5 h, 3.0 h, 3.5 h or 4 h.

[0099] The temperature of the fourth reaction may be 100°C, 101°C, 102°C, 103°C, 104°C, or 105°C.

[0100] The fourth reaction time can be 24 hours, 24.5 hours, 25 hours, 25.5 hours or 26 hours.

[0101] In some optional embodiments, the amount n1 of the aniline and the amount n2 of the 1,1-thiocarbonyldiimidazole satisfy: n1:n2=2:1; and / or

[0102] The amount n3 of the 4-nitro-o-phenylenediamine, the amount n4 of the 1,3-dicyclohexylcarbodiimide, and the amount n5 of the first intermediate satisfy: n3:n4:n5=1.6:1.92:1.92; and / or

[0103] The amount n6 of the oxalyl chloride, the amount n7 of the triethylamine, the amount n8 of the 4-dimethylaminopyridinepyrazole and the amount n9 of the carboxylic acid satisfy: n6:n7:n8:n9=7.5:10.0:1.25:5.0.

[0104] In these embodiments, the amount ratio of aniline and 1,1-thiocarbonyldiimidazole is 2:1, which allows the first reaction to proceed sufficiently to ultimately form a first intermediate with a molecular structure as shown in Formula 5. In addition, the amount ratio of 4-nitro-o-phenylenediamine, 1,3-dicyclohexylcarbodiimide, and the first intermediate is 1.6:1.92:1.92, which allows the second reaction to proceed sufficiently to ultimately form a second intermediate with a molecular structure as shown in Formula 6. In addition, the amount ratio of oxalyl chloride, triethylamine, 4-dimethylaminopyridinepyrazole, and carboxylic acid is 7.5:10.0:1.25:5.0, which allows the third reaction to proceed sufficiently to ultimately form a third intermediate with a molecular structure as shown in Formula 7.

[0105] In some optional embodiments, the amount n10 of the tetrahydroxydiboron, the amount n11 of the 4-4′-bipyridine, the amount n12 of the second intermediate and the amount n13 of the third intermediate satisfy: n10:n11:n12:n13=0.8:0.01:0.2:0.4.

[0106] In these embodiments, the molar ratio of tetrahydroxydiboron, 4-4′-bipyridine, the second intermediate, and the third intermediate is 0.8:0.01:0.2:0.4, which can enable the fourth reaction to proceed sufficiently to ultimately form the inhibitor product shown in Formula 1.

[0107] Based on a general inventive concept, an embodiment of the present application provides a pharmaceutical composition for inhibiting spleen tyrosine kinase, wherein the pharmaceutical composition includes the inhibitor.

[0108] The pharmaceutical composition is realized based on the above-mentioned inhibitor. The specific structural information of the inhibitor can be referred to the above-mentioned embodiments. Since the pharmaceutical composition adopts part or all of the technical solutions of the above-mentioned embodiments, it has at least all the beneficial effects brought about by the technical solutions of the above-mentioned embodiments, which will not be described one by one here.

[0109] It should be noted that the pharmaceutical composition may be a preparation containing an effective dose of the above-mentioned inhibitor or a pharmaceutically acceptable salt thereof.

[0110] It should be noted that the pharmaceutical composition may be in the form of a unit dose of the inhibitor. In these forms, the pharmaceutical composition is divided into unit doses containing an appropriate amount of the active ingredient. The unit dose form may be a packaged preparation containing a separate amount of the preparation, such as boxed tablets, capsules, and powders in vials or ampoules. The unit dose pharmaceutical composition may also be packaged in the form of capsules, cachets, tablets, or any other packaging that can accommodate an appropriate number of these.

[0111] It should be noted that the active ingredient of the pharmaceutical composition may be only the inhibitor component of the present application, or may be in the form of a combination with other anti-breast cancer compounds. In addition, during the actual application treatment process, the pharmaceutical composition of the present application can be used for combined treatment with other anti-breast cancer drugs. In the actual treatment process, this combined treatment can be achieved by administering various therapeutic components simultaneously, sequentially, or separately. Such pharmaceutical compositions may contain inhibitor components within an effective dose range during the treatment application stage, and may also contain other pharmaceutically active agents within the permitted dose range.

[0112] In some optional embodiments, the mass of the inhibitor is greater than or equal to 80% of the pharmaceutical composition.

[0113] In these embodiments, the mass of the inhibitor is greater than or equal to 80% of the pharmaceutical composition, which can provide sufficient inhibitor for the pharmaceutical composition to ensure the content of the effective component of the pharmaceutical composition.

[0114] The present application is further described below with reference to specific examples. Experimental methods in the following examples where specific conditions are not specified are generally measured in accordance with national / industry standards. If there are no corresponding national / industry standards, the methods are carried out in accordance with commonly used international standards, conventional conditions, or conditions recommended by the manufacturer.

[0115] Example 1

[0116] like Figure 1 and Figure 2 As shown, a method for preparing the inhibitor comprises:

[0117] S1. Aniline containing an R2 group and 1,1-thiocarbonyldiimidazole are reacted to obtain a first intermediate; the specific process is as follows:

[0118] 4-Morpholinoaniline (0.5 mmol, 1.0 equiv.) and 1,1-thiocarbonyldiimidazole (1.0 mmol, 2.0 equiv.) were dissolved in 3.0 mL of dichloromethane at room temperature and stirred for a first reaction for 4 hours. After completion of the reaction, the reaction solution was concentrated on a rotary evaporator under reduced pressure to remove the dichloromethane solvent, yielding a black crude product. The black crude product was then purified by silica gel column chromatography (hexane:ethyl acetate = 25 mL:1 mL) to afford the first intermediate 1a (4-morpholinophenyl isothiocyanate) as a white solid in an 88% yield.

[0119] S2. 4-nitro-o-phenylenediamine, 1,3-dicyclohexylcarbodiimide and the first intermediate are subjected to a second reaction to obtain a second intermediate; the specific process is as follows:

[0120] 4-Nitro-o-phenylenediamine (1.6 mmol, 1.0 equiv.) and the first intermediate 1a (1.92 mmol, 1.2 equiv.) were dissolved in 10 mL of acetonitrile, and 1,3-dicyclohexylcarbodiimide (DCC) (1.92 mmol, 1.2 equiv.) was added. The reaction mixture was stirred at 80°C for 16 hours. After the reaction was completed, the reaction solution was cooled to room temperature and then placed in an ice bath to allow the precipitation to completely form. The reaction solution was then filtered, and the solid precipitate was collected. The filter cake was washed with a small amount of dichloromethane to remove unconsumed DCC and the first intermediate 1a. The filter cake was then dried to obtain a crude reaction product. The crude reaction product was purified by silica gel column chromatography (dichloromethane:methanol = 40 mL:1 mL) to obtain the second intermediate 2a as a yellow solid in an 85% yield.

[0121] S3. A third reaction is performed on oxalyl chloride, triethylamine, 4-dimethylaminopyridine pyrazole and a carboxylic acid containing an R1 group to obtain a third intermediate; the specific process is as follows:

[0122] Benzoic acid (5.0 mmol, 1.0 equiv.) was dissolved in dichloromethane at room temperature. Three drops of N,N-dimethylformamide (DMF) were then added dropwise to the dichloromethane and stirred for 10 minutes to form a first reaction system. Oxalyl chloride (7.5 mmol, 1.5 equiv.) was then slowly added dropwise to the reaction system, and a third reaction was carried out with stirring at room temperature for 2 hours. After 2 hours, the reaction was monitored by TLC to confirm the completion of the third reaction. Triethylamine (10 mmol, 2.0 equiv.) and 4-dimethylaminopyridine (1.25 mmol, 0.25 equiv.) were then added to the first reaction system to form a second reaction system. Pyrazole (5.0 mmol, 1.0 equiv.) was then added in small portions to the second reaction system in an ice bath, and the reaction was stirred at 0°C for 2 hours. After the reaction is completed, a saturated NaCl solution is added to the reaction solution of the second reaction system, followed by extraction with 20 mL of dichloromethane solvent. The organic phases are extracted multiple times and combined, followed by addition of anhydrous Na2SO4, and the organic phases are dried, filtered, and concentrated under reduced pressure to obtain a pale yellow oily crude product. The pale yellow oily crude product is purified by silica gel column chromatography (hexane:ethyl acetate = 25 mL:1 mL) to obtain the third intermediate 3a as a colorless oily liquid. The yield of the third intermediate 3a is 78%.

[0123] S4. Tetrahydroxydiboron, 4-4′-bipyridine, the second intermediate and the third intermediate are subjected to a fourth reaction to obtain an inhibitor; the specific process is as follows:

[0124] At room temperature, the second intermediate 2a (0.2 mmol, 1.0 equiv.), tetrahydroxydiboron (0.8 mmol, 4.0 equiv.), and 4-4′-bipyridine (5 mmol%) were dissolved in 3 mL of DMF. After stirring at room temperature for 10 min, the third intermediate 3a (0.4 mmol, 2.0 equiv.) was added, and the temperature was raised to 100°C for a fourth reaction for 24 h. After the reaction was completed, the temperature was cooled to room temperature, quenched by the addition of 9 mL of saturated sodium chloride solution, and extracted with 5 mL of ethyl acetate to obtain an organic phase. The organic phase was washed with saturated sodium chloride solution (3 mL x 3) to completely remove residual DMF solvent. The washed organic phases were then combined, dried over anhydrous NaSO, filtered, and concentrated under reduced pressure to obtain a crude pale yellow solid. The crude pale yellow solid was purified by silica gel column chromatography (dichloromethane:methanol=40 mL:1 mL) to obtain inhibitor 4a as a white solid. The yield of inhibitor 4a was 65%.

[0125] like Figure 3 As shown, the final white solid inhibitor 4a is The NMR results of the inhibitor 4a are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.80(s,1H),10.07(s,1H),9.09(s,1H),8.03– 7.91(m,2H),7.83(s,1H),7.60(d,J=2.3Hz,1H),7.59-7.55(m,2H),7.52( dd,J=8.2,6.2Hz,2H),7.28(dd,J=8.5,2.0Hz,1H),7.21(d,J=8.4Hz,1H), 6.93(d,J=9.1Hz,2H),3.74(dd,J=5.8,3.8Hz,4H),3.06–3.00(m,4H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ165.53,152.07,146.21,135.97,134.05,131.63 ,129.72,128.98,128.77,128.01,119.08,116.69,114.21,66.69,50.01.

[0126] The mass spectrometer results are as follows: HR-ESI-MS m / z 414.1948 [M+H] +, (calcd for C 24 H 24 N5O2,414.1852).

[0127] Example 2

[0128] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0129] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3a was 78%; the yield of the white solid inhibitor 4a was 63%.

[0130] like Figure 3 As shown, the final white solid inhibitor 4b is The NMR results of the inhibitor 4b are as follows: H spectrum: 1HNMR (400MHz, DMSO-d6): δ10.05(s,1H),9.22(s,1H),7.85(d,J=2.0Hz,1H),7.80–7.72(m,2H),7.56(d,J=8.5Hz,2H),7.43-7.36(m,2H) ,7.29(dd,J=8.4,2.0Hz,1H),7.22(d,J=8.5Hz,1H),6.97-6.91(m,2H),3.74(dd,J=6.0,3.5Hz,4H),3.07-3.02(m,4H),2.40(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6) δ165.18,151.38,137.56,135.41,132.32,131.74,128.17,128.03,124.67,119.17,116.17,113.90,66.17,49.39,20.95.

[0131] The mass spectrometer results are as follows: HR-ESI-MS m / z 428.2124 [M+H] +, (calcd for C 25 H 26 N5O2, 428.2008).

[0132] Example 3

[0133] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0134] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3c was 83%; the yield of the light yellow solid inhibitor 4c was 62%.

[0135] like Figure 3 As shown, the finally obtained light yellow solid inhibitor 4c is The NMR results of the inhibitor 4c are as follows: H spectrum: 1 HNMR(400MHz,DMSO-d6)δ10.18(s,1H),9.20(s,1H),7.87–7.80(m,2H),7.77(dt,J=9.9,2.2Hz,1H),7.63–7.50(m,3H),7.42(td,J =8.6, 2.7Hz, 1H), 7.29 (dd, J = 8.5, 2.0Hz, 1H), 7.24 (d, J = 8.5Hz, 1H), 6.98–6.91 (m, 2H), 3.77–3.71 (m, 4H), 3.06–3.01 (m, 4H). Carbon spectrum: 13C NMR(100MHz,DMSO-d6)δ164.13,163.63,161.20,152.12,138.22,138.15,132.41,130.99,130 .91,124.25,124.22,119.74,118.68,118.47,116.67,114.95,114.72,114.47,66.66,49.86.

[0136] The mass spectrometer results are as follows: HR-ESI-MS m / z 432.1876 [M+H] +, (calcd for C 24 H 23 FN5O2,432.1758).

[0137] Example 4

[0138] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0139] The type of carboxylic acid containing the R1 group was adjusted so that the yield of the third intermediate 3d was 75%; the yield of the light yellow solid inhibitor 4d was 56%.

[0140] like Figure 3 As shown, the finally obtained light yellow solid inhibitor 4d is The NMR results of the inhibitor 4d are as follows: H spectrum: 1 H NMR (400 MHz, DMSO-d6): δ 10.84 (s, 1H), 10.32 (s, 1H), 9.14 (s, 1H), 8.16 (d, J = 8.1 Hz, 2H), 7.96–7.78 (m, 3H), 7.65–7.54 (m, 2H), 7.36–7.17 (m, 2H), 7.01–6.90 (m, 2H), 3.78–3.71 (m, 4H), 3.06–2.97 (m, 4H). 13 C NMR (100MHz, DMSO-d6): δ164.32,152.19,146.22,139.77,134.01,131.69,131.37,131.06 ,128.96,125.83,125.79,125.76,125.72,123.12,119.08,116.67,114.18,66.67,49.98.

[0141] The mass spectrometer results are as follows: HR-ESI-MS m / z 482.1814 [M+H] + ,(calcd for C 25 H 23F3N5O2,482.1726).

[0142] Example 5

[0143] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0144] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3e was 82%; the yield of the white solid inhibitor 4e was 70%.

[0145] like Figure 3 As shown, the final white solid inhibitor 4e is The NMR results of the inhibitor 4e are as follows: H spectrum: 1 HNMR (400MHz, DMSO-d6): δ10.79(s,1H),9.92(s,1H),9.09(s,1H),8.00–7.94(m,2H),7.81(s,1H),7.62–7.55(m,2H),7.26(dd,J=8.5,1.9H z,1H),7.20(d,J=8.4Hz,1H),7.05(d,J=8.8Hz,2H),6.93(d,J=9.0Hz,2H),3.84(s,3H),3.74(dd,J=5.9,3.6Hz,4H),3.05–3.01(m,4H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ164.93,162.10,152.01,146.21,134.04,129.88,128.04,128.00,119.08,116.68,114.27,113.99,66.68,55.86,50.01.

[0146] The mass spectrometer results are as follows: HR-ESI-MS m / z 444.3335 [M+H] + ,(calcd for C 25 H 26 N5O3,444.1957).

[0147] Example 6

[0148] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0149] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3f was 78%; the yield of the white solid inhibitor 4f was 68%.

[0150] like Figure 3 As shown, the final white solid inhibitor 4f is The NMR results of the inhibitor 4e are as follows: H spectrum: 1 H NMR (600MHz, DMSO-d6): δ10.88(s,1H),9.98(s,1H),9.17(s,1H),7.88(s,1H),7.68(dd,J=7.5,1.8Hz,1H),7.61–7.56(m,2H),7.49(ddd,J=8 .9,7.4,1.9Hz,1H),7.25–7.13(m,3H),7.07(td,J=7.4,1.0Hz,1H),6. 96–6.91(m,2H),3.92(s,3H),3.77–3.72(m,4H),3.06–3.00(m,4H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ164.11,156.99,146.14,134.11,132.29,130.29,125.55,120.99,118.94,116.67,112.48,66.68,56.40,50.01.

[0151] The mass spectrometer results are as follows: HR-ESI-MS m / z 444.1999 [M+H] + ,(calcd for C 25 H 26 N5O3,444.1957).

[0152] Example 7

[0153] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0154] The type of carboxylic acid containing the R1 group was adjusted so that the yield of the third intermediate 3g was 81%; the yield of the white solid inhibitor 4g was 56%.

[0155] like Figure 3 As shown, the final white solid inhibitor 4g is The nuclear magnetic resonance results of the inhibitor 4g are as follows: Proton spectrum: 1H NMR (400MHz, DMSO-d6): δ10.74(s,1H),9.98(s,1H),9.08(s,1H),7.72(s,1H),7.61–7.55(m,2H),7.40–7.29(m,4H),7.27–7.21(m,1 H),7.17(d,J=8.4Hz,1H),7.07(d,J=8.5Hz,1H),6.94–6.89(m,2H),3.73(dd,J=5.8,3.7Hz,4H),3.63(s,2H),3.05–2.98(m,4H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ168.83,146.14,136.91,134.07,129.52,128.74,126.89,118.98,116.66,112.83,66.67,49.99,43.88.

[0156] The mass spectrometer results are as follows: HR-ESI-MS m / z 428.2080 [M+H] + ,(calcd for C 25 H 26 N5O2, 428.2008).

[0157] Example 8

[0158] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0159] like Figure 3 As shown, by adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3h was 74%; the yield of the light yellow solid inhibitor 4h was 56%.

[0160] The light yellow solid inhibitor 4h obtained is The NMR results of the inhibitor for 4 hours are as follows: 1 HNMR (400 MHz, DMSO-d6): δ10.90 (s, 1H), 10.00 (s, 1H), 9.24–8.98 (m, 1H), 7.71 (d, J=2.0 Hz, 1H), 7.53 (d, J=8.4 Hz, 2H), 7.42–7.34 (m, 2H), 7.22–7.11 (m, 3H), 7.07 (dd, J=8.5, 2.0 Hz, 1H), 6.96–6.89 (m, 2H), 3.77–3.71 (m, 4H), 3.62 (s, 2H), 3.07–2.99 (m, 4H). 13C NMR (100MHz, DMSO-d6): δ168.77,162.75,160.34,152.06,133.00,132.97,132.8 3,131.40,131.32,119.59,116.66,115.53,115.32,113.02,66.66,49.87,42.82.

[0161] The mass spectrometer results are as follows: HR-ESI-MS m / z 444.1960 [M+H] + ,(calcd for C 25 H 25 FN5O2,446.1914).

[0162] Example 9

[0163] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0164] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3i was 68%; the yield of the white solid inhibitor 4i was 49%.

[0165] like Figure 3 As shown, the final white solid inhibitor 4i is The NMR results of the inhibitor 4i are as follows: H spectrum: 1 HNMR (400MHz, DMSO-d6): δ10.89(s,1H),9.96(s,1H),9.20(s,1H),7.73(d,J=2.0Hz,1H),7.59–7.53(m,2H),7.31–7.24(m ,2H),7.17(d,J=8.4Hz,1H),7.09(dd,J=8.4,2.0Hz,1H),6.95–6.86(m,4H),3.73(d,J=7.6Hz,7H),3.05–2.99(m,4H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ169.25,158.45,151.78,146.39,133.67,132.95,1 30.50,128.80,119.38,116.64,114.19,112.96,66.67,55.49,49.92,42.98.

[0166] The mass spectrometer results are as follows: HR-ESI-MS m / z 458.2140 [M+H] + ,(calcd for C 26 H 28 N5O3,458.2114).

[0167] Example 10

[0168] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0169] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3j was 79%; the yield of the white solid inhibitor 4j was 59%.

[0170] like Figure 3 As shown, the final white solid inhibitor 4j is The NMR results of the inhibitor 4j are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.80(s,1H),9.81(s,1H),9.08(s,1H),7.71(d, J=2.0Hz,1H),7.59–7.53(m,2H),7.24(ddt,J=6.5,4.7,1.9Hz,2H),7.16(d ,J=8.4Hz,1H),7.08(dd,J=8.5,2.0Hz,1H),7.00–6.96(m,1H),6.94–6.88( m,3H),3.79(s,3H),3.76–3.72(m,4H),3.62(s,2H),3.05–3.00(m,3H). Carbon spectrum: 13 CNMR (100MHz, DMSO-d6): δ168.85,157.75,151.85,146.26,133.94,132.98,131.11,1 28.36,125.08,120.64,119.16,116.67,112.90,111.22,66.68,55.92,49.99,38.17.

[0171] The mass spectrometer results are as follows: HR-ESI-MS m / z 458.2142 [M+H] + ,(calcd for C 26 H 28 N5O3,458.2114).

[0172] Example 11

[0173] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0174] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3k was 82%; the yield of the light yellow solid inhibitor 4k was 43%.

[0175] like Figure 3As shown, the finally obtained light yellow solid inhibitor 4k is The NMR results of the inhibitor 4k are as follows: H spectrum: 1 HNMR (600MHz, DMSO-d6): δ10.80(s,1H),9.91(s,1H),9.14(s,1H),7.92–7.78(m,1H),7.77–7.71(m,1H),7.61–7.56(m,2H) ,7.23–7.14(m,2H),7.10(dd,J=11.5,2.4Hz,1H),6.95–6.87(m,3H),3.94(s,3H),3.76–3.72(m,4H),3.05–3.01(m,4H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ161.81,157.31,157.21,150.54,144.74,130.73,130. 63,120.63,117.56,115.20,106.17,105.96,99.35,99.09,65.22,55.52,48.51.

[0176] The mass spectrometer results are as follows: HR-ESI-MS m / z 462.1909 [M+H] + ,(calcd for C 25 H 25 FN5O3,462.1863).

[0177] Example 12

[0178] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0179] The type of carboxylic acid containing the R1 group was adjusted so that the yield of the third intermediate 31 was 76%; the yield of the light green solid inhibitor 41 was 43%.

[0180] like Figure 3 As shown, the final light green solid inhibitor 41 is The NMR results of the inhibitor 4l are as follows: H spectrum: 1H NMR (400MHz, DMSO-d6): δ10.75(d,J=23.1Hz,1H),9.92(d,J=23.5Hz,1H),9.09(s,1H),7.74–7.64(m,1H),7.59(d,J=8.4Hz,2H),7.27(d ,J=1.9Hz,1H),7.20(t,J=12.3Hz,1H),7.17–7.09(m,2H),6.93(d,J=8.9Hz,2H),3.95(s,3H),3.77–3.71(m,4H),3.05–3.00(m,4H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ163.21,157.81,151.85,136.41,131.59,120.95,116.67,112.95,66.69,56.98,50.02.

[0181] The mass spectrometer results are as follows: HR-ESI-MS m / z 478.1591 [M+H] + ,(calcd for C 25 H 25 ClN5O3,478.1568).

[0182] Example 13

[0183] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0184] The type of carboxylic acid containing the R1 group was adjusted so that the yield of the third intermediate 3m was 85%; the yield of the light green solid inhibitor 4m was 51%.

[0185] like Figure 3 As shown, the final light green solid inhibitor 4m is The NMR results of the inhibitor 4m are as follows: H spectrum: 1 H NMR(600MHz,DMSO-d6)δ10.78(s,1H),9.95(s,1H),9.12(s,1H),7.85(s,1H),7.62–7.56(m,3H),7.39(d,J=1.8Hz,1H),7 .27(dd,J=8.1,1.8Hz,1H),7.23–7.13(m,2H),6.95–6.90(m,2H),3.94(s,3H),3.76–3.72(m,4H),3.04–3.01(m,4H). Carbon spectrum: 13C NMR (100MHz, DMSO-d6): δ163.35,157.71,146.21,133.97,131.78,125.11,124.93,123.92,119.05,116.66,115.70,104.65,66.68,56.96,49.98.

[0186] The mass spectrometer results are as follows: HR-ESI-MS m / z 522.1071 [M+H] + ,(calcd for C 25 H 25 BrN5O3,522.1063).

[0187] Example 14

[0188] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0189] By adjusting the type of carboxylic acid containing the R1 group, the yield of the third intermediate 3n was 58%; the yield of the white solid inhibitor 4n was 59%.

[0190] like Figure 3 As shown, the final white solid inhibitor 4n is The NMR results of the inhibitor 4n are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.81(s,1H),9.97(s,1H),9.10(s,1H),7.76(s,1H),7.62–7.55(m,2H),7.29(s,2 H),7.23(d,J=1.3Hz,2H),6.96–6.91(m,2H),3.88(s,6H),3.74(d,J=2.4Hz,7H),3.03(t,J=4.8Hz,4H). Carbon spectrum: 13 CNMR (100MHz, DMSO-d6): δ164.91,153.08,152.10,146.26,140.57,133.98,131 .05,119.15,116.68,114.58,105.73,66.68,65.36,60.59,56.59,50.00,15.62.

[0191] The mass spectrometer results are as follows: HR-ESI-MS m / z 504.2243 [M+H] + ,(calcd for C 27 H 30 N5O5,504.2169).

[0192] Example 15

[0193] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0194] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5a was 69%.

[0195] like Figure 4 As shown, the final white solid inhibitor 5a is The NMR results of the inhibitor 5a are as follows: H spectrum: 1 H NMR (400 MHz, DMSO-d6): δ 10.10 (s, 1H), 9.11 (s, 1H), 8.00–7.92 (m, 2H), 7.84 (s, 1H), 7.57 (dd, J = 7.9, 5.6 Hz, 3H), 7.52 (t, J = 7.1 Hz, 2H), 7.31–7.19 (m, 2H), 6.96–6.89 (m, 2H), 3.06 (t, J = 5.0 Hz, 4H), 2.50–2.44 (m, 4H), 2.24 (s, 3H). 13 C NMR (100MHz, DMSO-d6): δ165.06,151.65,145.65,135.49,133.35,132.00,131.18,128.31,127.56,118.54,116.48,113.70,54.67,49.02,45.65.

[0196] The mass spectrometer results are as follows: HR-ESI-MS m / z 427.2389 [M+H] + ,(calcd for C 25 H 27 N6O,427.2168).

[0197] Example 16

[0198] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0199] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5b was 59%.

[0200] like Figure 4 As shown, the final white solid inhibitor 5b is The NMR results of the inhibitor 5b are as follows: H spectrum: 1H NMR (400MHz, DMSO-d6): δ10.87(s,1H),10.10(s,1H),9.16(s,1H),7.88(s,1H),7.84–7.76(m,2H),7.67–7.59(m,2H),7.53–7 .39(m,2H),7.38–7.20(m,2H),7.04–6.95(m,2H),3.14(t,J=5.0Hz,4H),2.63(t,J=4.9Hz,4H),2.46(s,3H),2.36(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ165.63,145.97,138.05,135.96,133.90,132.21,128. 67,128.52,125.16,119.01,117.02,56.50,54.94,49.29,45.80,21.46,19.02.

[0201] The mass spectrometer results are as follows: HR-ESI-MS m / z 441.2340 [M+H] + ,(calcd for C 26 H 29 N6O,441.2325).

[0202] Example 17

[0203] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0204] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of pale yellow solid inhibitor 5c was 49%.

[0205] like Figure 4 As shown, the finally obtained light yellow solid inhibitor 5c is The NMR results of the inhibitor 5c are as follows: H spectrum: 1 HNMR (400 MHz, DMSO-d6): δ 10.90 (s, 1H), 10.16 (s, 1H), 9.17 (s, 1H), 7.87–7.73 (m, 3H), 7.57 (dq, J = 10.1, 3.3, 2.6 Hz, 3H), 7.42 (td, J = 8.8, 2.5 Hz, 1H), 7.30–7.17 (m, 2H), 6.95–6.88 (m, 2H), 3.08–3.02 (m, 4H), 2.47 (t, J = 5.0 Hz, 4H), 2.23 (s, 3H). 13C NMR (100MHz, DMSO-d6): δ164.07,163.63,161.21,138.29,138.22,133.78,130.98,13 0.90,124.24,124.21,119.01,118.42,116.92,114.94,114.72,55.21,49.57,46.23.

[0206] The mass spectrometer results are as follows: HR-ESI-MS m / z 445.2107 [M+H] + ,(calcd for C 25 H 26 FN6O,445.2074).

[0207] Example 18

[0208] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0209] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5d was 65%.

[0210] like Figure 4 As shown, the final white solid inhibitor 5d is The NMR results of the inhibitor 5d are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.30(s,1H),9.97(d,J=2.1Hz,1H),9.11(s,1H),8.21(d,J=2.0Hz,1H),7.70(s,1H),7.55(d,J=8.7Hz,2H),7.17( dd,J=8.4,2.8Hz,1H),7.12–7.01(m,1H),6.91(d,J=8.7Hz,2H),6.53(s,1H),3.06(t,J=5.0Hz,4H),2.51(d,J=4.9Hz,4H),2.25(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ163.21, 159.29, 152.40, 146.11, 133.76, 119.03, 116.94, 113.92, 88.79, 70.89, 60.22, 55.09, 49.43, 46.04, 23.77.

[0211] The mass spectrometer results are as follows: HR-ESI-MS m / z 495.2349 [M+H] + ,(calcd for C 26 H26 F3N6O,495.2042).

[0212] Example 19

[0213] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0214] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5e was 68%.

[0215] The final white solid inhibitor 5e is The NMR results of the inhibitor 5e are as follows: H spectrum: 1 HNMR (400 MHz, DMSO-d6): δ10.74 (s, 1H), 9.91 (s, 1H), 9.03 (s, 1H), 8.01–7.91 (m, 2H), 7.80 (s, 1H), 7.60–7.53 (m, 2H), 7.29–7.16 (m, 2H), 7.08–7.01 (m, 2H), 6.94–6.89 (m, 2H), 3.84 (s, 3H), 3.06 (dd, J=6.3, 3.7 Hz, 4H), 2.47 (d, J=5.5 Hz, 4H), 2.24 (s, 3H). 13 C NMR (100MHz, DMSO-d6): δ164.91,162.09,152.07,146.16,133.82,129.88,128.02,119.03,116.93,114.22,113.98,55.86,55.18,49.55,46.17.

[0216] The mass spectrometer results are as follows: HR-ESI-MS m / z 457.2324 [M+H] + ,(calcd for C 26 H 29 N6O2,457.2274).

[0217] Example 20

[0218] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0219] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5f was 69%.

[0220] like Figure 4 As shown, the final white solid inhibitor 5f is The NMR results of the inhibitor 5f are as follows: H spectrum: 1HNMR (400MHz, DMSO-d6): δ10.76(s,1H),9.96(s,1H),9.08(s,1H),8.03–7.81(m,1H),7.69(d,J=7.7Hz,1H),7.57(d,J=8.6Hz,2H),7.53–7.45(m ,1H),7.36–7.13(m,3H),7.07(t,J=7.5Hz,1H),6.92(d,J=8.8Hz,2H),3. 92(s,3H),3.05(t,J=4.9Hz,4H),2.46(t,J=4.9Hz,4H),2.22(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ164.10,156.99,146.17,133.80,132.28,130.29,120.99,118.95,116.90,112.48,56.40,55.23,49.60,49.36,46.27.

[0221] The mass spectrometer results are as follows: HR-ESI-MS m / z 457.2310 [M+H] + ,(calcd for C 26 H 29 N6O2,457.2274).

[0222] Example 21

[0223] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0224] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of 5g of white solid inhibitor was 43%.

[0225] like Figure 4 As shown, the final white solid inhibitor 5g is The NMR results of the inhibitor 5g are as follows: H spectrum: 1HNMR (400MHz, DMSO-d6): δ10.78(s,1H),9.93(s,1H),9.09(s,1H),7.84(s,1H),7.69(dd,J=8.2,1.7Hz,1H),7.61–7.54(m,2H),7.27(d,J=1.9Hz, 1H),7.20(s,2H),7.14(dd,J=8.2,1.9Hz,1H),6.95–6.89(m,2H),3.95(s ,3H),3.06(dd,J=6.3,3.6Hz,4H),2.49(d,J=8.0Hz,4H),2.24(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ163.20,157.81,152.16,146.15,136.44,133.79,1 31.66,124.63,120.96,119.01,116.93,112.93,56.98,55.15,49.51,46.12.

[0226] The mass spectrometer results are as follows: HR-ESI-MS m / z 491.3066 [M+H] + ,(calcd for C 26 H 28 ClN6O2,491.1884).

[0227] Example 22

[0228] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0229] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor in 5h was 47%.

[0230] like Figure 4 As shown, the final white solid inhibitor 5h is The NMR results of the inhibitor for 5h are as follows: 1 H NMR (400MHz, DMSO-d6): δ9.93(s,1H),9.10(s,1H),7.83(s,1H),7.60(d,J=8.2Hz,1H),7.57(d,J=9.0Hz,2H),7.39(d,J=1.8Hz,1H),7.2 7(dd,J=8.2,1.8Hz,1H),7.19(s,2H),6.92(d,J=9.1Hz,2H),3.94(s,3H),3.08(t,J=5.0Hz,4H),2.55(t,J=5.0Hz,4H),2.29(s,3H). Carbon spectrum:13 C NMR (100MHz, DMSO-d6): δ163.31,157.73,152.12,146.02,133.87,131.81,125.0 8,124.94,123.93,119.00,116.98,115.72,113.38,56.98,54.99,49.34,45.87.

[0231] The mass spectrometer results are as follows: HR-ESI-MS m / z 535.1790 [M+H] + ,(calcd for C 26 H 28 BrN6O2,535.1379).

[0232] Example 23

[0233] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0234] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5i was 64%.

[0235] like Figure 4 As shown, the final white solid inhibitor 5i is The NMR results of the inhibitor 5i are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.80(s,1H),9.98(s,1H),9.08(s,1H),7.75(s,1H),7.59–7.54(m,2H),7.29(s,2H),7.21 (s,2H),6.96–6.89(m,2H),3.88(s,6H),3.73(s,3H),3.07(t,J=5.0Hz,4H),2.51(d,J=2.3Hz,4H),2.25(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ166.68,164.14,152.31,145.37,139.73,133.06,131.43,131.24,130.31 ,128.39,118.28,116.20,104.88,64.75,59.84,55.81,54.37,48.72,45.33,29.73,18.38,13.27.

[0236] The mass spectrometer results are as follows: HR-ESI-MS m / z 517.3616 [M+H] + ,(calcd for C28 H 33 N6O4,517.2485).

[0237] Example 24

[0238] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0239] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5j was 42%.

[0240] like Figure 4 As shown, the final white solid inhibitor 5j is The NMR results of the inhibitor 5j are as follows: H spectrum: 1 H NMR (600MHz, DMSO-d6): δ10.17(s,1H),9.53(s,1H),8.03–7.94(m,3H),7.86(d,J=1.9Hz,1H),7.64–7.55(m,3H),7.52(t,J=7.6Hz,2H),7 .33(dd,J=8.5,2.0Hz,1H),7.24(d,J=8.4Hz,1H),7.22–7.15(m,2H),7.12–7.07(m,2H),6.99(d,J=7.9Hz,2H),6.73(t,J=7.2Hz,1H). Carbon spectrum: 13 C NMR (150MHz, DMSO-d6): δ165.68,151.65,145.11,137.84,135.79,132.92, 131.74,129.59,128.80,128.04,119.97,119.55,119.07,115.78,114.64.

[0241] The mass spectrometer results are as follows: HR-ESI-MS m / z 420.1810 [M+H] + ,(calcd for C 26 H 22 N5O,420.1764).

[0242] Example 25

[0243] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0244] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5k was 24%.

[0245] like Figure 4 As shown, the final white solid inhibitor 5k is The 5k NMR results of the inhibitor are as follows: H spectrum: 1 H NM (600MHz, DMSO-d6): δ10.81(d,J=39.5Hz,1H),9.96(s,1H),9.21(s,1H),8.00–7.96(m,2H),7.92(s,1H),7.63(d,J=8.3H z,2H),7.24(s,2H),7.20–7.15(m,2H),7.10–7.04(m,4H),6.99–6.95(m,2H),6.72(tt,J=7.2,1.2Hz,1H),3.84(s,3H). Carbon spectrum: 13 C NMR (150MHz, DMSO-d6): δ164.94,162.08,145.39,129.90,129.58,127.98,119.87,118.99,118.76,115.43,113.98,55.86.

[0246] The mass spectrometer results are as follows: HR-ESI-MS m / z 450.1908 [M+H] + ,(calcd for C 27 H 24 N5O2,450.1852).

[0247] Example 26

[0248] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0249] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of pale yellow solid inhibitor 51 was 29%.

[0250] like Figure 4 As shown, the finally obtained light yellow solid inhibitor 51 is The NMR results of the inhibitor 5l are as follows: H spectrum: 1HNMR (600MHz, DMSO-d6): δ10.89(s,1H),9.98(s,1H),9.28(s,1H),7.93(s, 1H),7.88(s,1H),7.68(d,J=8.2Hz,1H),7.65–7.61(m,2H),7.27(d,J=1.9H z,1H),7.22(s,2H),7.20–7.16(m,2H),7.13(dd,J=8.2,1.9Hz,1H),7.11–7 .08(m,2H),7.00–6.97(m,2H),6.73(tt,J=7.3,1.1Hz,1H),3.94(s,3H). Carbon spectrum: 13 CNMR (100MHz, DMSO-d6): δ162.87,157.33,151.46,144.89,136.73,135.98,134.25,132.02 ,131.16,129.14,124.24,120.49,119.35,118.80,118.41,115.09,113.08,112.45,56.50.

[0251] The mass spectrometer results are as follows: HR-ESI-MS m / z 484.1557 [M+H] + ,(calcd for C 27 H 23 ClN5O2,484.1462).

[0252] Example 27

[0253] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0254] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5m was 35%.

[0255] like Figure 4 As shown, the final white solid inhibitor 5m is The NMR results of the inhibitor 5m are as follows: H spectrum: 1H NMR (400MHz, DMSO-d6): δ9.99(s,1H),9.39(s,1H),7.95(s,1H),7.88(s,1H),7.62–7.58(m,3H),7.39(d,J=1.8Hz,1H),7.27(dd,J=8.2,1. 8Hz,1H),7.22–7.20(m,2H),7.19–7.16(m,2H),7.11–7.08(m,2H),7.00–6.97(m,2H),6.73(t,J=7.3Hz,1H),3.94(s,3H),3.84(s,1H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ167.08,163.45,159.35,157.70,151.75,145.22,132.75,131.75,129.58,1 25.13,124.94,123.91,123.49,119.67,118.95,116.18,115.69,115.66,65.39,56.95,56.71,15.64.

[0256] The mass spectrometer results are as follows: HR-ESI-MS m / z 528.1031 [M+H] + ,(calcd for C 27 H 23 BrN5O2, 528.0957).

[0257] Example 28

[0258] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0259] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 5n was 41%.

[0260] like Figure 4 As shown, the final white solid inhibitor 5n is The NMR results of the inhibitor 5n are as follows: H spectrum: 1H NMR (400MHz, DMSO-d6): δ10.95(s,1H),9.99(s,1H),9.23(s,1H),7.91(s,1H),7.78(s,1H),7.64–7.59(m,2H),7.29(s,2H),7.24(d,J=1 .3Hz,2H),7.18(dd,J=8.5,7.2Hz,2H),7.10–7.07(m,2H),7.00–6.97(m,2H),6.76–6.70(m,2H),3.88(s,6H),3.74(d,J=2.0Hz,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ164.94,153.08,151.94,145.33,140.58,131.05,129.56,119.79,119.42,118.90,115.63,105.73,60.59,56.60.

[0261] The mass spectrometer results are as follows: HR-ESI-MS m / z 510.2128 [M+H] + ,(calcd for C 29 H 28 N5O4,510.2063).

[0262] Example 29

[0263] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0264] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6a was 70%.

[0265] like Figure 5 As shown, the final white solid inhibitor 6a is The NMR results of the inhibitor 6a are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.27(s,1H),9.17(s,1H),8.00(dd,J=36.2,7.5Hz,3H),7.60(ddd,J=12.1,7.3,4.0Hz,4H),7.50(t,J= 7.6Hz, 2H), 7.16 (d, J = 7.8Hz, 1H), 6.99 (t, J = 7.9Hz, 1H), 6.94 (d, J = 8.7Hz, 2H), 3.74 (t, J = 4.7Hz, 4H), 3.03 (t, J = 4.7Hz, 4H). Carbon spectrum: 13C NMR (100MHz, DMSO-d6): δ167.85,165.63,151.42,146.31,135.01,133.76,133.29,13 2.18,131.33,129.73,129.02,128.99,128.22,118.84,116.76,113.99,66.67,49.97.

[0266] The mass spectrometer results are as follows: HR-ESI-MS m / z 414.1954 [M+H] + ,(calcd for C 24 H 24 N5O2,414.1852).

[0267] Example 30

[0268] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0269] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6b was 56%.

[0270] like Figure 5 As shown, the final white solid inhibitor 6b is The NMR results of the inhibitor 6b are as follows: H spectrum: 1 HNMR (600MHz, DMSO-d6): δ10.25(d,J=55.6Hz,1H),9.16(s,1H),8.04(d,J=8.3Hz,2H),7.60(d,J=8.3Hz,2H),7.17(s,1H),7 .13–7.09(m,2H),7.06(s,1H),7.01–6.96(m,1H),6.96–6.92(m,2H),3.86(s,3H),3.76–3.71(m,4H),3.05–3.00(m,4H). Carbon spectrum: 13 CNMR (150MHz, DMSO-d6): δ162.49,151.36,146.22,133.81,130.30,118.52,116.76,114.21,113.87,66.66,55.93,49.96.

[0271] The mass spectrometer results are as follows: HR-ESI-MS m / z 444.2053 [M+H] + ,(calcd for C 25 H 26 N5O3,444.1957).

[0272] Example 31

[0273] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0274] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6c was 62%.

[0275] like Figure 5 As shown, the final white solid inhibitor 6c is The NMR results of the inhibitor 6c are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.94(s,1H),10.75(s,1H),9.22(s,1H),8.12(dd,J=13.7,8.1Hz,1H),7.68(dd,J=14.9,8.3Hz,2H),7.60(d,J=2.0Hz,1 H),7.41(s,1H),7.27–7.19(m,1H),7.08–7.01(m,1H),6.96(dd,J=8.2,5 .3Hz,3H),4.19(s,3H),3.75(t,J=4.7Hz,4H),3.05(t,J=4.7Hz,4H). Carbon spectrum: 13 C NMR(100MHz,DMSO-d6)δ159.51,158.30,151.10,146.40,133.90,132.65,121.7 6,120.49,119.18,116.64,113.57,113.38,104.65,66.67,57.62,56.71,49.95.

[0276] The mass spectrometer results are as follows: HR-ESI-MS m / z 478.1665 [M+H] + ,(calcd for C 25 H 25 ClN5O3,478.1568).

[0277] Example 32

[0278] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0279] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6d was 59%.

[0280] like Figure 5 As shown, the final white solid inhibitor 6d is The NMR results of the inhibitor 6d are as follows: H spectrum: 1HNMR (600MHz, DMSO-d6): δ10.97(s,1H),10.75(s,1H),9.27(s,1H),8.07(d,J=31.2Hz,2H),7.69(d,J=8.3Hz,2H),7.53(s,1H),7.3 7(dd,J=8.4,1.8Hz,1H),7.03(d,J=7.8Hz,1H),6.96(dd,J=9.2,7.1Hz,3H),4.19(s,3H),3.78–3.73(m,4H),3.07–3.02(m,4H). Carbon spectrum: 13 C NMR (150MHz, DMSO-d6): δ158.18,151.04,146.41,119.17,116.62,116.33,66.67,65.39,57.60,49.91,15.64.

[0281] The mass spectrometer results are as follows: HR-ESI-MS m / z 522.1139 [M+H] + ,(calcd for C 25 H 25 BrN5O3,522.1063).

[0282] Example 33

[0283] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0284] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6e was 58%.

[0285] like Figure 5 As shown, the final white solid inhibitor 6e is The NMR results of the inhibitor 6e are as follows: H spectrum: 1 HNMR (400MHz, DMSO-d6): δ10.37(s,1H),10.19(s,1H),9.16(s,1H),7.66–7.59(m,2H),7.40(s,2H),7.18(s,1H),7.01 (d,J=7.5Hz,2H),6.93(d,J=8.5Hz,2H),3.90(d,J=1.5Hz,6H),3.76(s,3H),3.75–3.72(m,4H),3.05–3.00(m,4H). Carbon spectrum: 13C NMR (101MHz, DMSO-d6): δ164.88,153.16,151.40,146.22,140.86,133.81, 118.61,116.70,114.15,105.89,66.67,65.39,60.64,56.55,49.96,15.64.

[0286] The mass spectrometer results are as follows: HR-ESI-MS m / z 504.2218 [M+H] + ,(calcd for C 27 H 30 N5O5,504.2169).

[0287] Example 34

[0288] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0289] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6f was 51%.

[0290] like Figure 5 As shown, the final white solid inhibitor 6f is The NMR results of the inhibitor 6f are as follows: H spectrum: 1 H NMR (400MHz, DMSO-d6): δ10.31(s,1H),9.14(s,1H),8.10–7.99(m,2H),7.66–7.60(m,1H),7.57(dd,J=8.1,5.9Hz,4H),7.1 6(d,J=7.8Hz,2H),6.98(t,J=7.9Hz,1H),6.96–6.91(m,2H),3.08(t,J=5.0Hz,4H),2.55(t,J=5.0Hz,4H),2.28(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ165.12,150.96,145.61,134.52,133.13,131.67,128.48,127.73,118.27,116.57,113.44,54.47,48.82,45.36.

[0291] The mass spectrometer results are as follows: HR-ESI-MS m / z 427.2256 [M+H] + ,(calcd for C 25 H 27 N6O,427.2168).

[0292] Example 36

[0293] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0294] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of 6 g of white solid inhibitor was 55%.

[0295] like Figure 5 As shown, the final white solid inhibitor 6g is The NMR results of the inhibitor 6g are as follows: H spectrum: 1 HNMR (400 MHz, DMSO-d6): δ10.23 (s, 1H), 9.13 (s, 1H), 8.03 (d, J = 8.3 Hz, 2H), 7.62–7.53 (m, 2H), 7.19–7.07 (m, 3H), 7.06–6.81 (m, 4H), 3.86 (s, 3H), 3.06 (dd, J = 6.3, 3.7 Hz, 4H), 2.47 (d, J = 5.0 Hz, 4H), 2.23 (s, 3H). 13 C NMR (100MHz, DMSO-d6): δ162.00,150.95,145.74,133.07,131.25,129.71,118.20,116.52,113.73,113.31,55.45,54.67,49.04,45.67.

[0296] The mass spectrometer results are as follows: HR-ESI-MS m / z 457.2298 [M+H] + ,(calcd for C 26 H 29 N6O2,457.2274).

[0297] Example 37

[0298] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0299] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor in 6h was 58%.

[0300] like Figure 5 As shown, the final white solid inhibitor 6h is The NMR results of the inhibitor at 6h are as follows: 1HNMR (600MHz, DMSO-d6): δ10.96(s,1H),10.76(s,1H),9.24(s,1H),8.13(dd,J=19.8,8.2Hz,2H),7.68(d,J=8.6Hz,2H),7.42(d,J=1.9Hz,1H),7.24( dd,J=8.4,2.0Hz,1H),7.02(d,J=7.8Hz,1H),6.95(dd,J=8.4,6.2Hz,3H), 4.20(s,3H),3.07(t,J=5.0Hz,4H),2.49(t,J=6.2Hz,4H),2.24(s,3H). Carbon spectrum: 13 C NMR (150MHz, DMSO-d6): δ161.51,158.32,151.10,146.38,138.18,133.89,133.63,133.43,133.01,1 27.25,121.77,120.74,120.13,119.14,116.87,113.61,110.50,105.66,57.65,55.17,49.49,46.19.

[0301] The mass spectrometer results are as follows: HR-ESI-MS m / z 491.1949 [M+H] + ,(calcd for C 26 H 28 ClN6O2,491.1884).

[0302] Example 38

[0303] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0304] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6i was 49%.

[0305] like Figure 5 As shown, the final white solid inhibitor 6i is The NMR results of the inhibitor 6i are as follows: H spectrum: 1HNMR (600MHz, DMSO-d6): δ10.96(s,1H),10.76(s,1H),9.24(s,1H),8.13(dd,J=19.8,8.2Hz,2H),7.68(d,J=8.6Hz,2H),7.42(d,J=1.9Hz,1H),7.24( dd,J=8.4,2.0Hz,1H),7.02(d,J=7.8Hz,1H),6.95(dd,J=8.4,6.2Hz,3H), 4.20(s,3H),3.07(t,J=5.0Hz,4H),2.49(t,J=6.2Hz,4H),2.24(s,3H). Carbon spectrum: 13 C NMR (150MHz, DMSO-d6): δ161.51,158.32,151.10,146.38,138.18,133.89,133.63,133.43,133.01,1 27.25,121.77,120.74,120.13,119.14,116.87,113.61,110.50,105.66,57.65,55.17,49.49,46.19.

[0306] The mass spectrometer results are as follows: HR-ESI-MS m / z 491.1949 [M+H] + ,(calcd for C 26 H 28 ClN6O2,491.1884).

[0307] Example 39

[0308] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0309] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6j was 43%.

[0310] like Figure 5 As shown, the final white solid inhibitor 6j is The NMR results of the inhibitor 6j are as follows: H spectrum: 1H NMR (400MHz, DMSO-d6): δ12.79(s,1H),10.36(s,1H),10.18(s,1H),7.62–7.59(m,3H),7.41(s,2H),7.06–7.01(m,1H),6.92(d ,J=8.7Hz,2H),6.26(t,J=2.0Hz,1H),3.90(s,6H),3.77(s,3H),3.05(t,J=4.9Hz,4H),2.45(t,J=5.0Hz,4H),2.21(s,3H). Carbon spectrum: 13 C NMR (100MHz, DMSO-d6): δ164.46,152.70,151.04,145.77,140.51,133.05,129.32,125.19,1 21.21,120.25,118.04,116.48,113.60,105.61,104.17,60.17,56.11,54.72,49.10,45.75.

[0311] The mass spectrometer results are as follows: HR-ESI-MS m / z 517.3605 [M+H] + ,(calcd for C 28 H 33 N6O4,517.2485).

[0312] Example 40

[0313] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0314] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6k was 44%.

[0315] like Figure 5 As shown, the final white solid inhibitor 6k is The NMR results of the inhibitor 6k are as follows: H spectrum: 1 H NMR (400 MHz, DMSO-d6): δ 10.42 (s, 1H), 10.28 (s, 1H), 9.21 (s, 1H), 8.05 (d, J = 7.4 Hz, 2H), 7.95 (dt, J = 6.1, 1.4 Hz, 1H), 7.90 (s, 1H), 7.68–7.60 (m, 3H), 7.58 (dd, J = 8.2, 6.4 Hz, 2H), 7.24–7.14 (m, 3H), 7.14–7.05 (m, 3H), 6.97 (d, J = 7.7 Hz, 3H), 6.72 (t, J = 7.2 Hz, 1H). 13C NMR (100MHz, DMSO-d6): δ162.78,151.34,145.38,134.66,133.29,132.19, 129.72,129.56,129.02,128.98,128.31,119.94,118.84,115.55,113.97.

[0316] The mass spectrometer results are as follows: HR-ESI-MS m / z 420.1864 [M+H] + ,(calcd for C 26 H 22 N5O,420.1746).

[0317] Example 41

[0318] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0319] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 61 was 39%.

[0320] like Figure 5 As shown, the final white solid inhibitor 61 is The NMR results of the inhibitor 6l are as follows: H spectrum: 1 H NMR (400 MHz, DMSO-d6): δ 10.25 (s, 1H), 9.22 (s, 1H), 8.05 (d, J = 8.4 Hz, 2H), 7.90 (s, 1H), 7.64 (d, J = 8.3 Hz, 2H), 7.21–7.14 (m, 3H), 7.12–7.08 (m, 4H), 7.06–6.92 (m, 4H), 6.73 (t, J = 7.1 Hz, 1H), 3.86 (s, 3H). 13 C NMR (100MHz, DMSO-d6): δ162.52,151.31,145.40,134.70,131.80,129.56,119.97,118.83,115.53,114.27,114.21,55.94.

[0321] The mass spectrometer results are as follows: HR-ESI-MS m / z 450.1942 [M+H] + ,(calcd for C 27 H 24 N5O2,450.1852).

[0322] Example 42

[0323] Compared with Example 1, this embodiment has the following differences, and the rest are the same:

[0324] The types of carboxylic acid containing R1 group and aniline containing R2 group were adjusted; the yield of white solid inhibitor 6m was 41%.

[0325] like Figure 5 As shown, the final white solid inhibitor 6m is The NMR results of the inhibitor 6m are as follows: H spectrum: 1 H NMR (400 MHz, DMSO-d6): δ 10.44–10.11 (m, 2H), 9.21 (s, 1H), 7.90 (s, 1H), 7.71–7.57 (m, 3H), 7.40 (s, 2H), 7.18 (t, J = 7.7 Hz, 3H), 7.13–7.04 (m, 3H), 6.98 (d, J = 8.0 Hz, 3H), 6.73 (t, J = 7.2 Hz, 1H), 3.90 (s, 6H), 3.77 (s, 3H). 13 C NMR (100MHz, DMSO-d6): δ153.18,145.37,134.68,129.56,119.95,118.86,115.56,104.64,60.66,56.61.

[0326] The mass spectrometer results are as follows: HR-ESI-MS m / z 510.2175 [M+H] + ,(calcd for C 29 H 28 N5O4,510.2063).

[0327] Related experiments and effect data:

[0328] According to different R1 groups and R2 groups, each example was subjected to in vitro SYK enzyme inhibition activity and anti-proliferation activity of cells experiment, and the specific steps are as follows:

[0329] First, a total reaction volume of 50 μL was prepared, wherein the reaction system included: 40 mM Tris buffer (pH 7.4), 10 mM MgCl2, 0.1 mg / mL bovine serum albumin (BSA), 1 mM dithiothreitol (DTT), 10 μM ATP, and 0.2 mg / mL Poly(Glu, Tyr) substrate. The inhibitor to be tested in each example was diluted with a 10% dimethyl sulfoxide (DMSO) solution, and then 5 μL of the dilution was added to 50 μL of the reaction system, resulting in a final DMSO concentration of 1%.

[0330] The in vitro SYK enzyme is then added to the reaction system, and the kinase activity of the reaction system is measured using the Kinase-Glo Plus Luminescent Kinase Assay Kit. This kit assesses kinase activity by quantitatively measuring the amount of ATP remaining in the solution after the kinase reaction. The luminescent signal intensity of the solution is generally positively correlated with the ATP content and negatively correlated with kinase activity.

[0331] Anti-proliferative activity of cells: MCF-7, MDA-MB-231, and BT-549 cells were cultured in an incubator at 37°C and 5% CO2. 3 Cells were uniformly seeded at a density of 100 cells / well in a 96-well plate and incubated for 24 hours. Various concentrations of the compound were added to the cells and incubated for another 24 hours. MTT solution was then added to each well and incubated in an incubator for another 4 hours. Finally, the absorbance of each well was measured at 490 nm using a microplate reader, and the cell inhibition rate was calculated. Data were analyzed using GraphPad Prism software.

[0332] The test results of the detection kit are shown in Tables 1, 2 and 3.

[0333] Table 1 Results of in vitro SYK enzyme inhibition and cell anti-proliferative activity of Examples 1 to 14

[0334]

[0335]

[0336] In Table 1, a indicates that each compound was tested three times, and the values ​​are mean ± error. b indicates that the target cells were treated for 24 h.

[0337] Table 2 Results of in vitro SYK enzyme inhibition and cell anti-proliferative activity of Examples 15 to 28

[0338]

[0339] In Table 2, a indicates that each compound was tested three times, and the values ​​are mean ± error. b indicates that the target cells were treated for 24 h.

[0340] Table 3 Results of in vitro SYK enzyme inhibition and cell anti-proliferative activity of Examples 29 to 42

[0341]

[0342]

[0343] In Table 3, a indicates that each compound was tested three times, and the values ​​are mean ± error. b indicates that the target cells were treated for 24 h.

[0344] 2. For inhibitors 5g, 5h, and 5i, perform the following experiments:

[0345] All enzymatic reactions were incubated at 30°C for 40 minutes. The total reaction volume was 50 μL, containing 40 mM Tris buffer (pH 7.4), 10 mM MgCl2, 0.1 mg / mL bovine serum albumin (BSA), 1 mM dithiothreitol (DTT), 10 μM ATP, and 0.2 mg / mL Poly (Glu, Tyr) substrate. First, the test compound was diluted with a 10% DMSO solution, and then 5 μL of the dilution was added to the 50 μL reaction system, so that the final DMSO concentration of all reactions was 1%. The experiment was performed using the Kinase-Glo Plus Luminescent Kinase Assay Kit. This assay assesses kinase activity by quantitatively detecting the amount of ATP remaining in the solution after the kinase reaction. The luminescent signal intensity is positively correlated with the ATP content in the solution and negatively correlated with the kinase activity. The results are shown in Table 4.

[0346] Table 4 SYK kinase activity assay of inhibitors 5g, 5h, and 5i

[0347] Compd. <![CDATA[IC 50 (nM)]]> Inhibitor 5g 34 Inhibitor 5h 27 Inhibitor 5i 16

[0348] As can be seen from Tables 1, 2, 3 and 4, the inhibitors provided in the examples of the present application have a certain effect on the inhibitory effect of SYK enzyme and the proliferation of tumor cells. Among these inhibitors, inhibitor 5i has better effect and lower cost.

[0349] In summary, the embodiments of the present application provide a benzimidazole spleen tyrosine kinase inhibitor, which can optimize the structure of the benzimidazole inhibitor by introducing an amide R1 group and a variety of cyclic R2 groups to achieve efficient and selective inhibition of spleen tyrosine kinase (Syk).

[0350] In addition, the present invention provides a benzimidazole spleen tyrosine kinase inhibitor, which is a new type of SYK small molecule inhibitor with high selectivity, good pharmacokinetic properties and low toxicity, and is of great significance for improving the treatment effect of cancer, overcoming drug resistance and combined immunotherapy.

[0351] In addition, a benzimidazole spleen tyrosine kinase inhibitor provided in the examples of the present application can specifically inhibit human breast cancer cells MDA-MB-231 based on the inhibition principle of the SYK signaling pathway, which is of great significance for the treatment of breast cancer.

[0352] The foregoing is merely a detailed description of the present invention, intended to enable those skilled in the art to understand or implement the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention is not limited to the embodiments shown herein, but rather is intended to conform to the broadest scope consistent with the principles and novel features claimed herein.

Claims

1. A benzimidazole spleen tyrosine kinase inhibitor having a molecular structure as shown in Formula 1, In Formula 1, the R0 group is an R1 group containing an amide group, and the R1 group is selected from at least one of the following: The R3 group is selected from at least one of the following: H, F, Cl and Br; The R2 group is selected from at least one of the following:

2. The inhibitor according to claim 1, characterized in that The inhibitor has a molecular structure as shown in Formula 2, 3. The inhibitor according to claim 1, characterized in that The inhibitor has a molecular structure as shown in Formula 3, 4. The inhibitor according to any one of claims 1 to 3, characterized in that The inhibitor has a molecular structure as shown in Formula 4, 5. A method for preparing the inhibitor according to any one of claims 1 to 3, comprising: Aniline containing an R2 group and 1,1-thiocarbonyldiimidazole are subjected to a first reaction to obtain a first intermediate; Conducting a second reaction between 4-nitro-o-phenylenediamine, 1,3-dicyclohexylcarbodiimide and the first intermediate to obtain a second intermediate; A third intermediate is obtained by reacting oxalyl chloride, triethylamine, 4-dimethylaminopyridinepyrazole and a carboxylic acid containing an R1 group; subjecting tetrahydroxydiboron, 4-4′-bipyridine, the second intermediate and the third intermediate to a fourth reaction to obtain an inhibitor; Wherein, the first intermediate has a molecular structure as shown in Formula 5, The second intermediate has a molecular structure as shown in Formula 6, The third intermediate has a molecular structure as shown in Formula 7, 6. The preparation method according to claim 5, characterized in that The temperature of the first reaction is 20° C. to 30° C., and the time of the first reaction is ≥ 4 h; and / or The temperature of the second reaction is 80° C. to 85° C., and the time of the second reaction is ≥16 h; and / or The temperature of the third reaction is 20°C to 30°C, and the time of the third reaction is ≥ 2h; and / or The temperature of the fourth reaction is 100° C. to 105° C., and the time of the fourth reaction is 24 h to 26 h.

7. The preparation method according to claim 5, characterized in that The amount n1 of the aniline and the amount n2 of the 1,1-thiocarbonyldiimidazole satisfy: n1:n2=2:1; and / or The amount n3 of the 4-nitro-o-phenylenediamine, the amount n4 of the 1,3-dicyclohexylcarbodiimide, and the amount n5 of the first intermediate satisfy: n3:n4:n5=1.6:1.92:1.92; and / or The amount n6 of the oxalyl chloride, the amount n7 of the triethylamine, the amount n8 of the 4-dimethylaminopyridinepyrazole and the amount n9 of the carboxylic acid satisfy: n6:n7:n8:n9=7.5:10.0:1.25:5.

0.

8. The preparation method according to claim 5, characterized in that The amount n10 of the tetrahydroxydiboron, the amount n11 of the 4-4′-bipyridine, the amount n12 of the second intermediate and the amount n13 of the third intermediate satisfy: n10:n11:n12:n13=0.8:0.01:0.2:0.

4.

9. A pharmaceutical composition for inhibiting spleen tyrosine kinase, comprising the inhibitor according to any one of claims 1 to 3.

10. The pharmaceutical composition according to claim 9, characterized in that The mass of the inhibitor is greater than or equal to 80% of the pharmaceutical composition.

Images