Isoxazole biphenyl compounds with CBP BRD and BRD4 BD2 inhibitory activity as well as preparation method and application of isoxazole biphenyl compounds

Abstract

The invention provides an isoxazole biphenyl compound with CBP BRD and BRD4 BD2 inhibitory activity as well as a preparation method and application of the isoxazole biphenyl compound, and also provides an isoxazole biphenyl compound with CBP BRD and BRD4 BD2 inhibitory activity or a pharmaceutically acceptable salt of the isoxazole biphenyl compound, and the isoxazole biphenyl compound has a structural formula shown as a general formula (I) and a general formula (II). The invention also provides application of the isoxazole biphenyl compound or the pharmaceutically acceptable salt thereof or the pharmaceutical composition containing the isoxazole biphenyl compound in preparation of drugs for preventing or treating cancers, inflammations and metabolic diseases related to CBP and BRD4. The invention has the following technical effects: a novel molecule is designed and synthesized, and the structure is optimized, so that a compound with specific inhibitory activity on two targets of CBP BRD and BRD4 BD2 at the same time is obtained. The invention further provides a synthesis method of the compound, and the target compound can be efficiently prepared.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and more specifically, to a class of isoxazole biphenyl compounds with CBP BRD and BRD4 BD2 inhibitory activities, their preparation methods, and applications. Background Technology

[0002] The bromodomain (BRD) is a conserved protein domain composed of four α-helices (αZ, αA, αB, αC). The ZA loop and BC loop between these helices together form a hydrophobic pocket, a key structural feature of the BRD. As a crucial "reader" in epigenetic regulation, the bromodomain primarily functions by specifically recognizing and binding to acetylated lysine residues (Kac) on histones or non-histone proteins. This binding event, in turn, regulates downstream biological processes, including gene transcription and chromatin remodeling. Based on amino acid sequence homology, structural differences, and functional associations, 61 bromodomains have been identified in the human genome, classified into eight major families. Bromodomains from different families exhibit significant differences in cellular localization, target binding specificity, and biological function.

[0003] BRD4 belongs to the II subfamily, namely the BET family (bromodomain and extra-terminal family). BRD4 recognizes and binds to acetylated histones through its bromodomain, thereby recruiting transcription elongation complexes (such as the positive transcription elongation factor P-TEFb) to regulate oncogene transcription. Furthermore, BET-BRD proteins can recruit mediators and transcription factors (such as the oncogene MYC and nuclear factor κB (NF-κB)) to form a super-elongation complex. P-TEFb promotes serine phosphorylation of the C-terminal domain of RNA polymerase II (RNA polymerase II), ultimately driving RNA polymerase II-mediated transcription of various genes (Filippakopoulos P., Nat. Rev. Drug Discov. 2014, 13, 337-356; Hung KL, Nature 2021, 600, 731-736). Numerous studies have demonstrated a broad association between BET protein dysregulation and the occurrence and development of cancer, inflammation, and metabolic diseases (Faivre E.J., Nature 2020, 578, 306-310; Latif AL, Nat. Commun. 2021, 12, 241; Wang Z.-Q., Signal Transduct. Tar. 2023, 8, 420.). Small molecule inhibitors targeting the bromine domain of BRD4 show great potential in medical applications. First-generation pan-BET inhibitors lack specificity for BRD protein and BD1 and BD2, leading to some adverse reactions. Therefore, improving selectivity by inhibiting only BD1 or BD2 may provide a potential solution to improve specificity, avoid adverse reactions, and enhance the clinical efficacy of BET inhibitors.

[0004] CREB-binding protein (CREBBP, also known as CBP) belongs to subfamily III. CBP is a multifunctional nuclear protein whose core function is as a transcriptional coactivator, possessing histone acetyltransferase activity. CBP binds to transcription factors through its specific functional domains, forming the crucial "transcription factor-CBP" complex to precisely initiate gene transcription. Notably, CBP can acetylate the tumor suppressor protein p53, thereby enhancing p53's DNA-binding ability, promoting the transcription of downstream apoptosis-related genes, and thus inhibiting tumor cell proliferation and inducing tumor cell apoptosis. Targeting the bromine domain of CBP has become a promising strategy in cancer treatment. Currently, small molecule inhibitors targeting the bromine domain of CBP (such as CCS1477 (Inobrodib) and FT-7051) have entered clinical trials to evaluate their efficacy in treating hematologic malignancies and castration-resistant prostate cancer.

[0005] Therefore, both targets mentioned above have potential in the field of anticancer treatment, but related small molecule inhibitors are relatively scarce. Developing novel small molecule compounds that can simultaneously inhibit CBP BRD and BRD4 is expected to provide candidates for drug development and activity validation. Therefore, this paper proposes a class of isoxazole biphenyl compounds with inhibitory activities against CBP BRD and BRD4 BD2, along with their preparation methods and applications. Summary of the Invention

[0006] The purpose of this invention is to provide corresponding solutions to the aforementioned problems existing in the prior art. To achieve the above-mentioned objective, this invention provides the following technical solution: providing a class of isoxazole biphenyl compounds or pharmaceutically acceptable salts thereof with CBP BRD and BRD4 BD2 inhibitory activities, said compounds having the structural formulas shown in general formulas (I) and (II): (I) (II) in, R is selected from It can be any one of heterocyclic group, C1-C6 alkyl, or C3-C6 cycloalkyl; wherein R1-R5 are selected from H, halogen, methoxy, nitro or trifluoromethoxy; and the heterocyclic group is selected from thiophene group.

[0007] The term “C1-C6 alkyl” refers to saturated alkyl groups with 1-6 carbon atoms, including straight-chain and branched groups.

[0008] The term "C3-C6 cycloalkyl" refers to a saturated cycloalkyl group with 3-6 carbon atoms.

[0009] Preferably, R is selected from any one of the following substituents: 4-chlorophenyl, 2-chloro-4-fluorophenyl, 2-thienyl, butyl, phenyl, 2-chlorophenyl, n-propyl, 4-fluorophenyl, 2,4-difluorophenyl, 2-fluorophenyl, 3-fluorophenyl, 4-bromophenyl, 4-methoxyphenyl, 4-nitrophenyl, ethyl, 5-chloro-2-thienyl, 2,4,5-trifluorophenyl, 4-(trifluoromethoxy)-phenyl, 3-chloro-2-fluorophenyl, 2,5-difluorophenyl, 4-(trifluoromethyl)phenyl, 3,5-difluorophenyl, isobutyl.

[0010] In a preferred embodiment of the invention, the compounds of general formula (I) and (II) are preferably any one of the following compounds: 4-Chloro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-3); 2-Chloro-4-fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-4); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)thiophene-2-sulfonamide (WJA-5); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)butane-1-sulfonamide (WJA-6); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-7); 2-Chloro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-8); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)butyramide (WJA-9); 4-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-12); 2,4-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-13); 2-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-14); 3-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-16); 4-Bromo-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-17); 4-Methoxy-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-21); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-nitrobenzenesulfonamide (WJA-22); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)ethanesulfonamide (WJA-23); 5-Chloro-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)thiophene-2-sulfonamide (WJA-26); 2,4,5-Trifluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-30); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-(trifluoromethoxy)benzenesulfonamide (WJA-31); 3-Chloro-2-fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-32); 2,5-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-33); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-(trifluoromethyl)benzenesulfonamide (WJA-35); 3,5-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-38); 2-Methyl-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)propane-1-sulfonamide (WJA-39).

[0011] The specific structures of each compound are shown in Table 1: Table 1. Compound Structures ; ; More preferably, The isoxazole diphenyl ether compound mentioned is any one of the following compounds: N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinophenyl)butyramide (WJA-9).

[0012] 2-Fluoro-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinophenyl)benzenesulfonamide (WJA-14).

[0013] 4-Bromo-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-17).

[0014] N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)ethanesulfonamide (WJA-23).

[0015] 2,5-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-33).

[0016] In this invention, pharmaceutically acceptable salts may include acid addition salts formed by compounds of general formulas (I) and (II) with the following acids: hydrobromic acid, hydrochloric acid, sulfuric acid, phosphoric acid, borate, methanesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, benzenesulfonic acid, citric acid, lactic acid, pyruvic acid, tartaric acid, acetic acid, maleic acid or succinic acid, mandelic acid, fumaric acid, salicylic acid or phenylacetic acid. Furthermore, acidic salts of inorganic bases, such as salts containing basic metal cations, alkaline earth metal cations, or ammonium cations, are also included.

[0017] In a second aspect of the invention, a pharmaceutical composition is provided, comprising a compound as described in the first aspect or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient. The compound or pharmaceutically acceptable salt is the key active ingredient of the pharmaceutical composition for treating a disease or improving symptoms. The pharmaceutically acceptable excipient is used to improve the physical properties, stability, solubility, bioavailability, and other characteristics of the drug, including but not limited to fillers, disintegrants, binders, lubricants, stabilizers, and flavoring agents. The pharmaceutical composition can be formulated into various dosage forms, such as tablets, capsules, injections, oral liquids, and granules, to meet the medication needs and routes of administration of different patients.

[0018] In a third aspect of the invention, the use of compounds of general formula (I) and (II) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions comprising thereof, in the preparation of medicaments for inhibiting BRD4 BD2 is provided. Specifically, the invention provides the use of a class of isoxazol diphenyl ether compounds of general formula (I) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions comprising thereof, in the preparation of medicaments for the prevention or treatment of cancers, inflammations, and metabolic diseases associated with BRD4 BD2, wherein the cancers are leukemia, prostate cancer, or colon cancer, and the metabolic diseases are diabetes or coronary artery disease.

[0019] In a fourth aspect of the invention, a method for preparing a class of isoxazole diphenyl ether compounds as described in the first aspect is provided.

[0020] The present invention uses the following synthetic route to prepare compounds of general formula (I):

[0021] The substituent R in compounds of general formulas (I) and (II) are as defined in the first aspect.

[0022] The preparation method of the compounds of general formula (I) and (II) specifically includes the following steps: 1) 4-Bromo-1-fluoro-2-nitrobenzene and morpholine were subjected to a substitution reaction to obtain compound 2, namely 4-(4-bromo-2-nitrophenyl)morpholine; the molar ratio of 4-bromo-1-fluoro-2-nitrobenzene and morpholine was 1:1.1-2, the solvent was dimethyl sulfoxide, the reaction temperature was 80~130℃, and the reaction time was 0.25~2h; 2) Compound 2, iron powder, acetic acid and ammonium chloride were subjected to a reduction reaction to obtain compound 3, namely 5-bromo-2-morpholinoaniline; the molar ratio of compound 2, iron powder, acetic acid and ammonium chloride was 1:4-10:1.8-3:0.4-1, the solvent was ethanol and water, the reaction temperature was 50-80℃, and the reaction time was 12h. 3) Compound 3, pinacol diborate, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride, and potassium acetate were subjected to a borate esterification reaction to obtain compound 4, namely 2-morpholino-5-(4,4,5,5-tetramethyl-1,3,2-dioxoborane-2-yl)aniline; the molar ratio of compound 3, pinacol diborate, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride, and potassium acetate was 1:1-3:0.02-0.2:1-3, the solvent was 1,4-dioxane and dimethyl sulfoxide, the reaction system was under vacuum, the reaction temperature was 80-110℃, and the reaction time was 8-12h; 4) Compound 4 was subjected to sulfonation or acylation reactions with different sulfonyl chlorides or acyl chlorides to obtain a series of intermediates 5-1 or 5-2; the molar ratio of compound 4 to different sulfonyl chlorides or acyl chlorides was 1:1.0-3; the base used in the sulfonation reaction was pyridine, and the molar ratio of compound 4 to pyridine was 1:4-10; the base used in the acylation reaction was triethylamine, and the molar ratio of compound 4 to triethylamine was 1:1.5-3.5; the solvent was dichloromethane, the reaction temperature was room temperature to 40℃, and the reaction time was 2-12 h.

[0023] 5) Compound 5-1 or 5-2, 5-bromo-3-methylbenzisoxazole, bis(triphenylphosphine) palladium dichloride (PdCl2(PPh3)2), triphenylphosphine (PPh3), and potassium bicarbonate (KHCO3) were subjected to a Suzuki coupling reaction to obtain the target compound; the molar ratio of compound 5-1 or 5-2, 5-bromo-3-methylbenzisoxazole, PdCl2(PPh3)2, PPh3, and KHCO3 was 1:1-1.3:0.025-0.1:0.05-0.2:1.5-3, the solvent was N,N-dimethylacetamide and water (volume ratio 8-14:1), the reaction system was under vacuum, the reaction temperature was 85-110℃, and the reaction time was 12h.

[0024] Preferably, In step 1), the molar ratio of 4-(4-bromo-2-nitrophenyl)morpholine, 4-bromo-1-fluoro-2-nitrobenzene, and morpholine is 1:1.5. In step 2), the molar ratio of compound 2, iron powder, acetic acid, and ammonium chloride is 1:9:2.2:0.5. In step 3), the molar ratio of compound 3, pinacol diboronate, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride, and potassium acetate is 1:1.22:0.034:2. In step 4), the molar ratio of compound 4 to pyridine is 1:6.2, and the molar ratio of compound 4 to triethylamine is 1:3. In step 5), the molar ratio of compound 5-1 or 5-2, 5-bromo-3-methylbenzisoxazole, PdCl2(PPh3)2, PPh3 and KHCO3 is 1:1.2:0.05:0.1:2.

[0025] In step 4), the sulfonyl chloride is selected from R1SO2Cl, and the acyl chloride is selected from R1COCl; wherein R1 is selected from... It can be any one of heterocyclic groups or C1-C6 alkyl groups; wherein R1-R5 are selected from H, halogen, methoxy, nitro or trifluoromethoxy; and the heterocyclic group is selected from thiophene group.

[0026] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. This invention designs and synthesizes novel molecules and optimizes their structures to obtain compounds that simultaneously exhibit specific inhibitory activities against two targets, CBP BRD and BRD4BD2.

[0027] 2. This invention also provides a method for synthesizing the compound-like compound, which can efficiently prepare the target compound. Attached Figure Description

[0028] Figure 1 This is a structural schematic diagram provided for the present invention. Detailed Implementation

[0029] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention.

[0030] Therefore, the following detailed description of the embodiments of the present invention is not intended to limit the scope of the claimed invention, but merely illustrates some embodiments of the invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without inventive effort are within the scope of protection of the present invention. It should be noted that, in the absence of conflict, the embodiments and features and technical solutions in the embodiments of the present invention can be combined with each other. It should be noted that similar reference numerals and letters in the following figures indicate similar items; therefore, once an item is defined in one figure, it does not need to be further defined and explained in subsequent figures.

[0031] Example 1: Preparation of 2-fluoro-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA14) Step 1: Preparation of 4-(4-bromo-2-nitrophenyl)morpholine (2) 16.0 g (72.7 mmol) of 4-bromo-1-fluoro-2-nitrobenzene was added to a 150 mL round-bottom flask containing 25 mL of dimethyl sulfoxide, followed by the addition of morpholine (9.5 g, 109.1 mmol). The resulting mixture was stirred at 130 °C for 20 minutes, and the reaction solution gradually turned orange-red. The reaction progress was monitored by thin-layer chromatography. After the reaction was complete, the reaction solution was poured into 90 mL of saturated sodium bicarbonate solution and extracted with ethyl acetate (EA) (100 mL × 3). The combined organic layers were washed successively with water (100 mL × 2) and brine (100 mL), dried over anhydrous Na₂SO₄, and concentrated under reduced pressure to give the target compound as an orange-red oily liquid. The yield was 19.2 g (92%). 1 H NMR spectrum (400 MHz, CDCl3), δ, ppm ( J ,Hz):8.00–7.83(1H,m,Ar H);7.59(1H,s,Ar H);7.11–6.96(1H,m,Ar H);3.83(4H,t, J =4.5 Hz,morpholine H);3.03(4H,t, J =4.5 Hz, morpholine H). Step 2 Preparation of 5-bromo-2-morpholinoaniline (3) Iron powder (26.3 g, 470.2 mmol), acetic acid (6.9 g, 114.9 mmol), and ammonium chloride (1.4 g, 26.1 mmol) were added to a round-bottom flask containing 135 mL of water and stirred at 50°C for 15 minutes. Then, 4-(4-bromo-2-nitrophenyl)morpholine (15.0 g, 52.2 mmol) dissolved in 45 mL of ethanol was added to the mixture, and stirring continued for 45 minutes. The reaction progress was monitored by thin-layer chromatography. After the reaction was complete, the mixture was cooled to room temperature, and the pH was adjusted to 7–9 with sodium bicarbonate solution. After filtration through diatomaceous earth, the filter cake was washed with ethyl acetate and water. The combined filtrates were extracted with ethyl acetate (150 mL × 3). The combined organic layers were washed successively with water (100 mL × 3) and saturated brine (100 mL), dried over anhydrous Na₂SO₄, and concentrated under reduced pressure to give the target compound as a grayish-white solid. Yield: 8.5 g (63%).

[0032] Step 3 Preparation of 2-morpholino-5-(4,4,5,5-tetramethyl-1,3,2-dioxoborane-2-yl)aniline (4) 1,4-Dioxane (30 mL) and dimethyl sulfoxide (3 mL) were added to a round-bottom flask, followed by the addition of 5-bromo-2-morpholinoaniline (7.5 g, 29.1 mmol), and the mixture was degassed under vacuum. Then, pinacol diborate (8.9 g, 35.0 mmol), potassium acetate (5.7 g, 58.3 mmol), and [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride (725.6 mg, 0.99 mmol) were added sequentially to the system. The reaction mixture was stirred at 100°C for 12 hours under nitrogen protection, and the reaction progress was monitored by thin-layer chromatography. After the reaction was complete, some of the solvent was removed under reduced pressure. The residual mixture was extracted with ethyl acetate (100 mL × 3). The combined organic layers were washed successively with water and brine, dried over anhydrous Na₂SO₄, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent system: petroleum ether / ethyl acetate, v / v ratio 4:1) to give the target product as a pale yellow solid. Yield: 6.75 g (76%). 1 H NMR spectrum (400 MHz, CDCl3), δ, ppm ( J ,Hz):7.24(1H,d, J =7.9 Hz,Ar H);7.19(1H,s,Ar H);6.98(1H,d, J=7.8 Hz,Ar H);3.89(2H,brs,NH2);3.86–3.83(4H,m,morpholine H);2.98–2.92(4H,m,morpholine H);1.32(12H,s,4×CH3). Step 4: Preparation of 4-chloro-N-(2-morpholino-5-(4,4,5,5-tetramethyl-1,3,2-dioxoborane-2-yl)phenyl)benzenesulfonamide (5-1-1) Compound 7 (200.0 mg, 0.60 mmol) and 4-chlorobenzenesulfonyl chloride (151.6 mg, 0.72 mmol) were added to a flask containing 6 mL of dichloromethane, followed by the dropwise addition of 0.3 mL of pyridine. The mixture was stirred at room temperature for 12 hours, monitored by thin-layer chromatography. After the reaction was complete, the solvent was removed under reduced pressure, and the crude product was purified by silica gel column chromatography (eluent system: petroleum ether / ethyl acetate, v / v ratio 4:1) to give the target compound as a white solid. Yield: 157 mg (55%). This intermediate was used directly in subsequent reaction steps.

[0033] Step 5: Preparation of 4-chloro-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA14) N,N-dimethylacetamide (14 mL) and water (1 mL) were added to a round-bottom flask, and the mixture was degassed under vacuum. Then, compound 5-1-1 (150.0 mg, 0.31 mmol), 5-bromo-3-methylbenzo[d]isoxazole (79.7 mg, 0.38 mmol), PdCl2(PPh3)2 (22.0 mg, 0.031 mmol), PPh3 (16.4 mg, 0.063 mmol), and KHCO3 (62.7 mg, 0.63 mmol) were added to the reaction flask. The reaction mixture was stirred at 100 °C for 12 hours under nitrogen protection. The reaction progress was monitored by thin-layer chromatography. After the reaction was complete, 30 mL of water was added to the mixture, followed by extraction with ethyl acetate. The organic layer was washed with water and saturated brine, dried over anhydrous Na2SO4, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluting system: petroleum ether / ethyl acetate, v / v ratio 4:1) to give the target compound as a white solid. Yield: 38 mg (25%). 1 H NMR(400 MHz,DMSO)δ9.40(s,1H,SO2NH),7.93(s,1H,Ar H),7.86–7.73(m,4H,Ar H),7.66(d, J =8.6 Hz,2H,Ar H),7.58–7.46(m,2H,Ar H),7.26(d,J =8.1 Hz,1H,Ar H),3.69–3.57((t, J =4.0 Hz,4H,morpholine H),2.61(s,3H,CH3),2.59(t, J =4.0 Hz, 4H, morpholine H). Example 2: 2-Chloro-4-fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-4) Prepared according to the method of Example 1, except that in step four, compound 4 and 2-chloro-4-fluorobenzenesulfonyl chloride are used as reactants. Yield: 20%, white solid, mp 205–206°C. 1 H NMR (400 MHz, DMSO- d 6) δ 9.41 (s, 1H, SO2NH), 8.13 (dd, J =8.9,5.9 Hz,1H,Ar H),7.88(s,1H,Ar H),7.81–7.71(m,3H,Ar H),7.54–7.46(m,2H,Ar H),7.42(td, J =8.7, 2.5 Hz, 1H, Ar H), 7.34 (d, J =8.9 Hz, 1H, Ar H), 3.69(t, J =4.5 Hz,4H,morpholine H),2.75(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3). 13 C NMR (101 MHz, DMSO)δ 165.40,162.86 (d, J F-C =256.5 Hz,1C,4'-ArC),161.57,155.58,144.21,136.23,135.23, 133.70,133.60 (d, J F-C =10.1 Hz, 1C, 2'-ArC), 132.77, 132.65 (d, J F-C =12.1 Hz,1C,6'-ArC),131.62,129.20,124.81,122.76(2×C),122.64,120.67, 119.77 119.60 119.51 (d,J F-C =26.3 Hz, 1C, 3'-ArC), 115.10,114.88 (d, J F-C =22.2Hz,1C,5'-ArC),110.10,66.23(2×C),52.26(2×C),9.63.HRMS(ESI):m / z[M+H] + calcdfor C 24 H 22 ClFN3O4S:502.1004,found:502.1007. Example 3: N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)thiophene-2-sulfonamide (WJA-5) Prepared according to the method of Example 1, except that in step four, compound 4 and thiophene-2-sulfonyl chloride are used as reactants. 1 H NMR (400 MHz, DMSO) δ9.35 (s, 1H, SO2NH), 7.97 (s, 1H, Ar H), 7.92 (dd, J =5.0,1.3Hz,1H,Ar H),7.85–7.75(m,2H,Ar H),7.63(d, J =2.1 Hz, 1H, Ar H), 7.60 (dd, J =3.7, 1.3 Hz, 1H, Ar H), 7.54 (dd, J =8.3,2.2 Hz,1H,Ar H),7.29(d, J =8.3 Hz, 1H, Ar H), 7.16(dd, J =4.9,3.8 Hz,1H,Ar H),3.74–3.61(m,4H,morpholine H),2.70–2.56(m,7H,morpholine H,CH3). 13 C NMR(101 MHz, DMSO) δ161.55,155.57,144.91,140.75,135.84,135.36,133.37,132.41,131.93,129.1 9,127.83,124.93,122.78,122.32,121.99,119.52,110.08,66.18(2×C),52.11(2×C),9.68. Example 4: N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)butane-1-sulfonamide (WJA-6) Prepared according to the method of Example 1, except that in step four, compound 4 and 1-butanol chloride are used as reactants. 1 H NMR (400 MHz, DMSO) δ8.58 (s, 1H, SO2NH), 8.04 (d, J =1.3 Hz, 1H, Ar H), 7.86 (dd, J =8.7,1.8 Hz,1H,Ar H),7.78(d, J =8.7 Hz, 1H, Ar H), 7.65 (d, J =2.1 Hz, 1H, Ar H), 7.52(dd, J =8.3,2.2 Hz,1H,Ar H),7.38(d, J =8.3 Hz,1H,Ar H),3.86–3.75(m,4H,morpholineH),3.30–3.25(m,2H,SO2 CH2 ),2.95–2.82(m,4H,morpholine H),2.61(s,3H,CH3),1.78–1.67(m,2H,SO2CH2 CH2 ), 1.45–1.35(m, 2H, CH2 CH3), 0.87(t, J =7.4 Hz, 3H, CH2 CH3 ). 13 C NMR(101 MHz, DMSO) δ161.55,155.56,143.86,136.19,135.57,132.86,129.41,124.07,122.73,122 .13,120.97,119.69,109.99,66.39(2×C),52.26(2×C),52.09,25.21,20.78,13.53,9.67. Example 5: N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-7) Prepared according to the method of Example 1, except that in step four, compound 4 and benzenesulfonyl chloride are used as reactants. 1 H NMR(400 MHz,DMSO)δ9.22(s,1H,SO2NH),7.92(s,1H,Ar H),7.82(t, J =7.0 Hz, 2H, ArH), 7.76(d, J=8.8 Hz,2H,Ar H),7.67–7.53(m,4H,Ar H),7.49(dd, J =8.3,2.1 Hz,1H,ArH),7.25(d, J =8.3 Hz,1H,Ar H),3.69–3.57(m,4H,morpholine H),2.61(s,3H,CH3),2.58–2.51(m,4H,morpholine H). 13 C NMR(101 MHz, DMSO) δ161.55,155.56,144.70,140.36,135.86,135.36,132.97,132.25,129.27,129.20(2×C ),126.77(2×C),124.65,122.77,122.31,121.90,119.47,110.06,66.13(2×C),52.09(2×C),9.68. Example 6: 2-Chloro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-8) Prepared according to the method of Example 1, except that in step four, compound 4 and 2-chlorobenzenesulfonyl chloride are used as reactants. 1 H NMR (400 MHz, DMSO) δ9.33 (s, 1H, SO2NH), 8.11 (dd, J =7.9,1.4 Hz,1H,Ar H),7.86(s,1H,Ar H),7.76(d, J =8.6 Hz,1H,Ar H),7.74–7.68(m,2H,Ar H),7.66(td, J =7.5,1.5 Hz,1H,Ar H),7.55(td, J =8.0, 2.0 Hz, 1H, Ar H), 7.49 (d, J =2.0 Hz, 1H, Ar H), 7.47 (dd, J =8.2,2.1 Hz,1H,Ar H),7.34(d, J =8.2 Hz,1H,Ar H),3.73–3.66(m,4H,morpholine H),2.80–2.70(m,4H,morpholine H),2.61(s,3H,CH3). 13C NMR(101 MHz, DMSO) δ161.56,155.57,143.71,136.87,136.27,135.27,134.84,132.05,131.85,131.40,130.7 0,129.17,127.78,124.48,122.75,122.69,119.83,119.56,110.09,66.27(2×C),52.24(2×C),9.66. Example 7: N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)butyramide (WJA-9) Step 1: Preparation of N-(2-morpholino-5-(4,4,5,5-tetramethyl-1,3,2-dioxoborane-2-yl)phenyl)butyramide Compound 4 (150 mg, 0.49 mmol) and butyryl chloride (63.1 mg, 0.59 mmol) were dissolved in 6 mL of dichloromethane, and triethylamine (150 mg, 1.48 mmol) was added. The mixture was reacted overnight at room temperature. The reaction was monitored by TLC. After the reaction was complete, 35 mL of water was added, and the mixture was extracted with ethyl acetate (50 mL × 3). The organic layer was washed three times with water and once with saturated brine. The solution was dried over anhydrous sodium sulfate and concentrated under reduced pressure to give the target compound as a white solid. The yield was 168.8 mg (91%). This compound was used directly in the next reaction.

[0034] Step 2: Preparation of N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)butyramide (WJA-9) Prepared according to the method in step 5 of Example 1. 1 H NMR (400 MHz, DMSO) δ8.97(s,1H,SO2NH),8.26(s,1H,Ar H),8.01(s,1H,Ar H),7.84(dd, J =8.7,1.6 Hz,1H,Ar H),7.76(d, J =8.7 Hz, 1H, Ar H), 7.45 (dd, J =8.2,1.9 Hz,1H,Ar H),7.27(d, J =8.3 Hz,1H,Ar H),3.91–3.74(m,4H,morpholine H),2.97–2.77(m,4H,morpholine H),2.61(s,3H,CH3),2.42(t, J =7.3 Hz, 2H, COCH2), 1.64(q, J=7.3 Hz, 2H, COCH2 CH2 ),0.94(t, J =7.3 Hz, 3H, CH2 CH3 ). 13 C NMR(101 MHz, DMSO) δ171.23,161.48,155.56,142.56,136.00,135.14,132.84,129.33,123.03,122 .73,120.93,120.43,119.39,109.92,66.32(2×C),51.69(2×C),38.33,18.62,13.66,9.68. Example 8: 4-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-12) Prepared according to the method of Example 1, except that in step four, compound 4 and 4-fluorobenzenesulfonyl chloride are used as reactants. Yield: 27%, white solid, mp 222–223°C. 1 H NMR (400 MHz, DMSO- d 6)δ9.31(s,1H,SO2NH),7.93(s,1H,Ar H),7.88(dd, J =8.8,5.2 Hz,2H,Ar H),7.83–7.74(m,2H,Ar H),7.56(d, J =2.1 Hz, 1H, Ar H), 7.51(dd, J =8.3,2.1 Hz,1H,Ar H),7.43(t, J =8.8 Hz, 2H, ArH), 7.26(d, J =8.3 Hz, 1H, Ar H), 3.64(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3),2.58(t, J =4.5 Hz, 4H, morpholine H). 13 C NMR (101 MHz, DMSO)δ 165.62,163.12 (d, J F-C =252.5 Hz,1C,4'-ArC),161.56,155.57,144.99, 136.88,136.85 (d, J F-C=3.0 Hz,1C,1'-ArC),135.84,135.32,131.97, 129.93,129.83 (d, J F-C =10.1 Hz,2C,2',6'-ArC),129.21,124.91,122.78, 122.40,122.28 (d, J F-C =12.1 Hz,2C,3',5'-ArC),119.50,116.53,116.30,110.07,66.11(2×C),52.14(2×C),9.64.HRMS(ESI):m / z[M+H] + calcd for C 24 H 23 FN3O4S:468.1393,found:468.1396. Example 9: 2,4-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-13) Prepared according to the method of Example 1, except that in step four, compound 4 and 2,4-difluorobenzenesulfonyl chloride are used as reactants. 1 H NMR(400 MHz,DMSO)δ9.55(s,1H,SO2NH),7.95(s,1H,Ar H),7.87(td, J =8.7,6.4Hz,1H,Ar H),7.83–7.73(m,2H,Ar H),7.64–7.57(m,1H,Ar H),7.55(d, J =7.2 Hz, 2H, ArH), 7.29(d, J =8.8 Hz, 1H, Ar H), 7.25(td, J =8.5,2.1 Hz,1H,Ar H),3.66–3.60(m,4H,morpholine H),2.78–2.64(m,4H,morpholine H),2.61(s,3H,CH3). Example 10: 2-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA14) Prepared according to the method of Example 1, except that in step four, compound 4 and 2-fluorobenzenesulfonyl chloride were used as reactants. Yield: 45 mg (30%), melting point: 184–185°C. 1 H NMR (400 MHz, DMSO- d6)δ9.44(s,1H,SO2NH),7.92(s,1H,Ar H),7.83(td, J =7.6,1.6 Hz,1H,Ar H),7.79–7.75(m,2H,Ar H),7.73–7.67(m,1H,Ar H),7.55(d, J =2.1 Hz,1H,Ar H),7.51(dd, J =8.3,2.2 Hz,1H,Ar H),7.49–7.42(m,1H,Ar H),7.36(t, J =7.6 Hz,1H,Ar H),7.29(d, J =8.3 Hz,1H,Ar H),3.64(t, J =4.5 Hz,4H,morpholine H),2.67(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3). 13 C NMR(101MHz,DMSO)δ161.56, 159.54,157.01 (d, J F-C =255.5 Hz,1C,2'-ArC),155.57,145.12, 135.93 (1×C),135.85 (d, J F-C =8.1 Hz,1C,4'-ArC),135.24,131.53, 130.32(1×C),130.20 (d, J F-C =12.1 Hz,1C,6'-ArC),129.18, 127.91,127.77 (d, J F-C =14.1 Hz,1C,1'-ArC),125.13, 124.93,124.90 (d, J F-C =3.0 Hz,1C,5'-ArC),f 122.77,122.37,119.54, 117.51,117.30 (d, J F-C =21.2 Hz,1C,3'-ArC),110.07,66.18(2×C),52.22(2×C),9.67.HRMS(ESI):m / z[M+H] + calcd for C 24 H 23FN3O4S:468.1393,found:468.1393. Example 11: 3-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-16) Prepared according to the method of Example 1, except that in step four, compound 4 and 3-fluorobenzenesulfonyl chloride are used as reactants. Yield: 42%, white solid, mp 204–205°C. 1 H NMR(400 MHz, CDCl3)δ8.04(s,1H,SO2NH),7.84(d, J =2.1 Hz,1H,Ar H),7.73(s,1H,Ar H),7.72–7.68(m,1H,Ar H),7.65(dt, J =8.1,1.2 Hz,1H,Ar H),7.63–7.59(m,1H,Ar H),7.57(dt, J =8.0,2.2 Hz,1H,Ar H),7.44(td, J =8.1,5.2 Hz,1H,Ar H),7.32(dd, J =8.2,2.1 Hz,1H,Ar H),7.26–7.19(m,2H,Ar H),3.80(t, J =4.6 Hz,4H,morpholine H),2.64(s,3H,CH3),2.61(t, J =4.6 Hz, 4H, morpholine H). 13 C NMR (101 MHz, CDCl3)δ 163.64 162.54 161.13 (d, J F-C =253.5 Hz, 1C, 3'-ArC), 155.29, 141.59,141.53 (d, J F-C =6.1 Hz,1C,1'-ArC),141.10,138.88,135.98,133.19, 131.07, 130.99 (d, J F-C =8.1 Hz,1C,5'-ArC),129.48,123.94,123.06, 122.80,122.76 (d, J F-C=4.0Hz, 1C, 6'-ArC), 122.66, 120.55,120.34 (d, J F-C =21.2 Hz,1C,4'-ArC),119.33,117.83, 114.59,114.35 (d, J F-C =24.2 Hz,1C,2'-ArC),110.23,67.30(2×C),53.01(2×C),10.17.HRMS(ESI):m / z[M+H] + calcd for C 24 H 23 FN3O4S:468.1393,found:468.1397. Example 12: 4-Bromo-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-17) Prepared according to the method of Example 1, except that in step four, compound 4 and 4-bromobenzenesulfonyl chloride are used as reactants. 1 H NMR (400 MHz, Chloroform- d )δ8.02(s,1H,SO2NH),7.83(d, J =2.0 Hz, 1H, Ar H), 7.71(dd, J =7.9,2.3 Hz,4H,Ar H),7.64–7.56(m,3H,Ar H),7.32(dd, J =8.2,2.1 Hz,1H,Ar H),7.22(d, J =8.2 Hz,1H,Ar H),3.89–3.76(m,4H,morpholine H),2.64(s,3H,CH3),2.64–2.54(m,4H,morpholine H). 13 C NMR(101 MHz, CDCl3)δ162.54,155.28,141.04,138.85,138.60,135.99,133.24,132.46(2×C),129.47,128.5 0(2×C),128.29,123.87,123.08,122.67,119.33,117.67,110.25,67.33(2×C),53.00(2×C),10.22. Example 13: 4-Methoxy-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-21) Prepared according to the method of Example 1, except that in step four, compound 4 and 4-methoxybenzenesulfonyl chloride are used as reactants. 1 H NMR (400 MHz, DMSO- d 6) δ 9.00 (s, 1H, SO2NH), 7.92 (d, J =1.3 Hz, 1H, Ar H), 7.77(d, J =8.9 Hz, 4H, Ar H), 7.58 (d, J =2.2 Hz, 1H, Ar H), 7.47 (dd, J =8.3,2.2 Hz,1H,Ar H),7.26(d, J =8.3 Hz,1H,Ar H),7.16–7.00(m,2H,Ar H),3.79(s,3H,OCH3),3.68(t, J =4.5Hz,4H,morpholine H),2.61(s,3H,CH3),2.60–2.53(m,4H,morpholine H). 13 C NMR (101MHz, DMSO) δ162.62,161.56,155.58,144.13,135.98,135.46,132.59,131.83,129.23,129.05 (2× C),124.27,122.78,122.35,120.95,119.50,114.39(2×C),110.07,66.19(2×C),55.73,52.14(2×C),9.64. Example 14: N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-nitrobenzenesulfonamide (WJA-22) Prepared according to the method of Example 1, except that in step four, compound 4 and 4-nitrobenzenesulfonyl chloride are used as reactants. 1 H NMR (400 MHz, DMSO- d 6)δ9.72(s,1H,SO2NH),8.54–8.31(m,2H,Ar H),8.13–8.01(m,2H,Ar H),7.95(d, J =1.8 Hz, 1H, Ar H), 7.82(dd, J =8.7,1.8 Hz,1H,Ar H),7.76(d, J =8.7 Hz,1H,Ar H),7.60–7.47(m,2H,Ar H),7.26(d,J =8.7 Hz, 1H), 3.58(t, J =4.5 Hz,4H,morpholine H),2.70–2.55(m,7H,morpholine H,CH3). 13 C NMR(101 MHz, DMSO) δ161.57,155.60,149.77,146.20,145.81,135.73,135.17,131.28,129.23,128.43(2×C ),125.53,124.58(2×C),123.69,122.80,122.18,119.57,110.06,66.05(2×C),52.17(2×C),9.62. Example 15: N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)ethanesulfonamide (WJA-23) Prepared according to the method of Example 1, except that in step four, compound 4 and ethylsulfonyl chloride are used as reactants. 1 H NMR (400 MHz, DMSO- d 6) δ 8.54 (s, 1H, SO2NH), 8.04 (d, J =1.8 Hz, 1H, Ar H), 7.86 (dd, J =8.8,1.8 Hz,1H,Ar H),7.78(d, J =8.7 Hz, 1H, Ar H), 7.65 (d, J =2.2 Hz, 1H, Ar H), 7.52 (dd, J =8.3,2.2 Hz,1H,Ar H),7.38(d, J =8.3 Hz, 1H, Ar H), 3.80(t, J =4.6 Hz,4H,morpholine H),3.30–3.25(m,2H, CH2 CH3), 2.89(t, J =4.6Hz,4H,morpholine H),2.61(s,3H,CH3),1.29(s,3H,CH2 CH3 ). 13C NMR(101 MHz, DMSO) δ161.57,155.57,143.87,136.26,135.58,132.91,129.44,124.09,122 .74,122.18,120.90,119.72,109.98,66.38(2×C),52.25(2×C),46.83,9.67,8.14. Example 16: 5-Chloro-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)thiophene-2-sulfonamide (WJA-26) Prepared according to the method of Example 1, except that in step four, compound 4 and 5-chlorothiophene-2-sulfonyl chloride are used as reactants. 1 H NMR (400 MHz, DMSO- d 6)δ9.63(s,1H,SO2NH),7.99(s,1H,Ar H),7.90–7.69(m,2H,Ar H),7.66–7.50(m,2H,Ar H),7.46(d,J=4.1 Hz,1H,Ar H),7.30(d, J =8.5 Hz, 1H, ArH), 7.23(d, J =4.1 Hz, 1H, Ar H), 3.67(t, J =4.5 Hz,4H,morpholine H),2.68(t, J =4.6 Hz,4H,morpholine H),2.62(s,3H,CH3). 13 C NMR(101 MHz, DMSO) δ161.55,155.53,145.81,139.42,135.71,135.20,135.06,132.12,131.19,129.1 7,127.97,125.58,123.49,122.78,122.21,119.49,110.04,66.13(2×C),52.20(2×C),9.61. Example 17: 2,4,5-Trifluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-30) Prepared according to the method of Example 1, except that in step four, compound 4 and 2,4,5-trifluorobenzenesulfonyl chloride are used as reactants. Yield: 20%, white solid, mp 195–196°C. 1 H NMR (400 MHz, DMSO- d6)δ9.73(s,1H,SO2NH),8.07–7.85(m,3H,Ar H),7.83(dd, J =8.8,1.8 Hz,1H,Ar H),7.76(d, J =8.6 Hz,1H,Ar H),7.57(d, J =6.4 Hz,2H,Ar H),7.30(d, J =8.9 Hz,1H,Ar H),3.64(t, J =4.5 Hz,4H,morpholine H),2.72(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3). 13 C NMR(101MHz,DMSO)δ161.59, 155.68 ,155.59, 153.69,153.18 (d, J F-C =252.5 Hz,1C,2'-ArC), 151.15 (d, J F-C =256.5 Hz,1C,4'-ArC), 146.75 ,146.10, 144.29 (d, J F-C =248.5 Hz,1C,5'-ArC),135.86,135.13,130.82,129.25,125.84, 125.34,125.17 (d, J F-C =17.2 Hz,1C,1'-ArC),124.10,122.80,122.20,119.63, 118.52,118.31 (d, J F-C =21.2 Hz,1C,6'-ArC),110.05, 108.75,108.69,108.47,108.41,108.19 (td, J F-C =22.2,6.1 Hz,1C,3'-ArC),66.16(2×C),52.30(2×C),9.60.HRMS(ESI):m / z[M+H] + calcd for C 24 H 21 F3N3O4S:504.1205,found:504.1208. Example 18: N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-(trifluoromethoxy)benzenesulfonamide (WJA-31) Prepared according to the method of Example 1, except that in step four, compound 4 and 4-trifluoromethoxybenzenesulfonyl chloride are used as reactants. Yield: 17%, white solid, mp 214–215°C. 1 H NMR (400 MHz, DMSO- d 6) δ 9.43 (s, 1H, SO2NH), 7.98 (d, J =1.8 Hz,1H,Ar H),7.95–7.86(m,2H,Ar H),7.81(dd, J =8.7,1.8 Hz,1H,Ar H),7.76(d, J =8.7 Hz,1H,Ar H),7.66–7.56(m,3H,Ar H),7.54(dd, J =8.3,2.3 Hz,1H,Ar H),7.26(d, J =8.3 Hz, 1H, Ar H), 3.58(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3),2.53(t, J =4.5 Hz, 4H, morpholine H). 13 C NMR (101 MHz, DMSO) δ161.59,155.60,151.05,145.57,139.53,135.90,135.28,131.71,129.43(2×C),129.24,125.38, 123.68 (To be specified), 123.58, 122.80, 122.39, 121.61 (2×C), 121.12 ,119.57, 118.55,115.99 (q, J F-C =258.6 Hz,1C,OCF3),110.06,66.07(2×C),52.18(2×C),9.63.HRMS(ESI):m / z[M+H] + calcdfor C 25 H 23 F3N3O5S:534.1311,found:534.1313. Example 19: 3-Chloro-2-fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-32) Prepared according to the method of Example 1, except that in step four, compound 4 and 3-chloro-2-fluorobenzenesulfonyl chloride were used as reactants. Yield: 24%, white solid, mp 197–198°C. 1 H NMR (400 MHz, DMSO- d 6) δ 9.78 (s, 1H, SO2NH), 7.99 (d, J =1.8 Hz, 1H, Ar H), 7.90(ddd, J =8.3,6.8,1.7 Hz,1H,Ar H),7.83(dd, J =8.7,1.8 Hz,1H,Ar H),7.77(d, J =8.7 Hz, 1H, Ar H), 7.72(ddd, J =8.0,6.4,1.7 Hz,1H,Ar H),7.63–7.52(m,2H,Ar H),7.36(td, J =8.0,1.0 Hz,1H,Ar H),7.27(d, J =8.9 Hz, 1H, Ar H), 3.57(t, J =4.5 Hz,4H,morpholine H),2.66(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3). 13 C NMR (101 MHz, DMSO) δ161.55,155.54, 154.96,152.42 (d, J F-C =256.5Hz,1C,2'-ArC),146.47,135.72,135.46,135.06,130.82, 130.17,130.03 (d, J F-C =14.1 Hz,1C,1'-ArC),129.16,128.80,125.95, 125.64,125.60 (d, J F-C =4.0 Hz,1C,4'-ArC),124.92,122.77,122.08, 121.56,121.39 (d, J F-C=17.1 Hz,1C,3'-ArC),119.52,110.01,66.09(2×C),52.26(2×C),9.62.HRMS(ESI):m / z[M+H] + calcd for C 24 H 22 ClFN3O4S:502.1004,found:502.1006. Example 20: 2,5-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-33) Prepared according to the method of Example 1, except that in step four, compound 4 and 2,5-difluorobenzenesulfonyl chloride are used as reactants. Yield: 29%, white solid, mp 192–193°C. 1 H NMR (400 MHz, DMSO- d 6) δ 9.69 (s, 1H, SO2NH), 7.96 (d, J =1.9 Hz,1H,Ar H),7.84–7.73(m,2H,Ar H),7.70–7.47(m,5H,Ar H),7.29(d, J =8.9 Hz, 1H, Ar H), 3.62(t, J =4.5 Hz,4H,morpholine H),2.70(t, J =4.6 Hz,4H,morpholine H),2.61(s,3H,CH3). 13 C NMR (101 MHz, DMSO) δ 161.54, 158.36,155.92 (d, J F-C =246.4 Hz, 1C, 5'-ArC), 155.79,155.77 155.54 153.30,153.27 (d, J F-C =251.5,3.1Hz,1C,2'-ArC),145.84,135.78,135.14,131.07,129.17,125.58,123.66,122.76, 122.37, 122.29 ,122.16, 122.12 (supplement), 122.04 (dd, J F-C =25.3,8.1 Hz,1C,1'-ArC), 119.58,119.50 (repair) ,119.52,119.34,119.26 (dd, J F-C =24.3,8.1 Hz,1C,6'-ArC), 116.58,116.31 (d, J F-C =27.1 Hz,1C,4'-ArC),115.05,114.80(d, J F-C =25.3 Hz,1C,3'-ArC),110.02,66.11(2×C),52.20(2×C),9.59.HRMS(ESI):m / z[M+H] + calcd for C 24 H 22 F2N3O4S:486.1299,found:486.1302. Example 21: N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)-4-(trifluoromethyl)benzenesulfonamide (WJA-35) Prepared according to the method of Example 1, except that in step four, compound 4 and 4-trifluoromethylbenzenesulfonyl chloride are used as reactants. Yield: 25%, white solid, mp 227–228°C. 1 H NMR (400 MHz, DMSO- d 6)δ9.60(s,1H,SO2NH),8.03–7.93(m,5H,Ar H),7.86–7.73(m,2H,H),7.62–7.52(m,2H,Ar H),7.25(d, J =8.9 Hz, 1H, Ar H), 3.54(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3),2.55(t, J =4.4 Hz, 4H, morpholine H). 13 C NMR (101 MHz, DMSO) δ161.55,155.54,145.84,144.51,135.74,135.16, 132.62,132.30 (d, J F-C =32.3 Hz,1C,4'-ArC),131.37,129.18,127.75(2×C), 127.48 , 126.46,126.42 (d, J F-C =4.0 Hz, 2C, 3', 5'-ArC), 125.54, 124.77 ,124.02,122.77,122.23, 122.05 119.50 119.35 (q, J F-C =273.7 Hz,1C,CF3),110.01,66.01(2×C),52.13(2×C),9.60.HRMS(ESI):m / z[M+H] + calcd for C 25 H 23 F3N3O4S:518.1361,found:518.1365. Example 22: 3,5-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-38) Prepared according to the method of Example 1, except that in step four, compound 4 and 3,5-difluorobenzenesulfonyl chloride are used as reactants. Yield: 17%, white solid, mp 219–220°C. 1 H NMR (400 MHz, DMSO- d 6) δ 9.65 (s, 1H, SO2NH), 7.96 (d, J =1.8 Hz, 1H, Ar H), 7.83 (dd, J =8.7,1.8 Hz,1H,Ar H),7.76(d, J =8.7Hz,1H,Ar H),7.68–7.60(m,1H,Ar H),7.60–7.48(m,4H,Ar H),7.28(d, J =8.2 Hz, 1H, ArH), 3.64(t, J =4.5 Hz,4H,morpholine H),2.65(t, J =4.5 Hz,4H,morpholine H),2.61(s,3H,CH3). 13 C NMR (101 MHz, DMSO)δ 163.46,163.34 161.57 160.96,160.84 (dd, J F-C =252.5,12.1 Hz,2C,3',5'-ArC),155.56,145.63, 144.14,144.06,143.97 (t, J F-C=8.5 Hz,1C,1'-ArC),135.77,135.15,131.25,129.20,125.49,123.52,122.77,122.20,119.52, 110.75, 110.66,110.55,110.47 (dd, J F-C =20.2,9.1 Hz,2C,2',6'-ArC),110.02, 108.92,108.67, 108.41 (t, J F-C =25.3 Hz,1C,4'-ArC),66.05(2×C),52.22(2×C),9.58.HRMS(ESI):m / z[M+H] + calcd for C 24 H 22 F2N3O4S:486.1299,found:486.1300. Example 23: 2-Methyl-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)propane-1-sulfonamide (WJA-39) Prepared according to the method of Example 1, except that in step four, compound 4 and 2-methylbenzenesulfonyl chloride are used as reactants. 1 H NMR (400 MHz, DMSO- d 6) δ 8.57 (s, 1H, SO2NH), 8.04 (d, J =1.7 Hz, 1H, Ar H), 7.86 (dd, J =8.8,1.8 Hz,1H,Ar H),7.78(d, J =8.7 Hz, 1H, Ar H), 7.65 (d, J =2.2 Hz, 1H, Ar H), 7.52 (dd, J =8.3,2.2 Hz,1H,Ar H),7.38(d, J =8.3 Hz, 1H, Ar H), 3.80(t, J =4.6 Hz,4H,morpholine H),3.21(d, J =6.5 Hz, 2H, SO2CH2), 2.89(t, J =4.6 Hz,4H,morpholine H),2.61(s,3H),2.25–2.15(m,1H,SO2CH2 CH ),1.05(s,3H,CH( CH3)2),1.03(s,3H,CH( CH3 )2). 13 C NMR(101 MHz, DMSO) δ161.56,155.55,143.65,136.22,135.60,132.87,129.38,123.97,122.74,12 2.15,120.54,119.69,110.00,66.39(2×C),59.94,52.19(2×C),24.40,22.08(2×C),9.64. Example 24: CBP TR-FRET target activity assay Compound dilution buffers were prepared in ECHO plates (Beckman) using DMSO. Biochemical reactions were performed in 20 μL samples in 384-well plates (OptiPlate-384, PerkinElmer). Each well contained 20 nL of the compound, 5 μL of protein (N-terminal GST-labeled CREBBP, amino acid numbers 1081-1197) (final concentration 3 nM), 5 μL of biotin (sequence: GTAVLREIRRYQK(Ac)STEL-GG-K(Biotin)-OH), synthesized by GL Corporation; final concentration 20 nM), and 10 μL of assay mixture (from EPIgeneous Binding Domain kit B, Revvity; kit included GST-Eu). 3+ -Cryptate antibody and Streptavidin-XL665 reagent (final concentration 1.25 nM). The reaction mixture was shaken for 30 seconds and incubated at room temperature for 3 hours. Readings were taken on an EnVision reader (PerkinElmer) at excitation wavelength of 340 nm and emission wavelengths of 615 nm and 665 nm. Blank control group (DMSO, detection buffer, peptide) and high-concentration control group (DMSO, protein, peptide) were used as references. Results were calculated from the fluorescence signals at 665 nm and 615 nm, expressed as the HTRF ratio: HTRF ratio = F(665 nm) / F(615 nm). In the following formula, the HTRF ratio is abbreviated as F. Inhibition rate = [1-(F inhibitor -F blank control ) / F high control -F blank control [×100. Inhibition curve and half-maximum inhibitory concentration (IC50)] 50 The values ​​were calculated using GraphPad Prism 7 software.

[0035] Example 25: BRD4 BD2 TR-FRET target activity assay Compound dilution buffers were prepared in ECHO plates (Beckman) using DMSO. Biochemical reactions were performed in 20 μL samples in 384-well plates (OptiPlate-384, PerkinElmer). Each well contained 15 nL of compound, 5 μL of protein (N-terminal GST-labeled BRD4 BD2, amino acid numbers 349-460) (final concentration 5 nM), 5 μL of biotin (sequence: SGRG-K(Ac)-GG-K(Ac)-GLG-K(Ac)-GGA-K(Ac)-RHRKVGG-K(Biotin), synthesized by GL Corporation, China; final concentration 10 nM), and 10 μL of assay mixture (from EPIgeneous Binding Domain kit B, Revvity; kit included GST-Eu). 3+ -Cryptate antibody and Streptavidin-XL665 reagent (final concentration 1.25 nM). The reaction mixture was shaken for 30 seconds and incubated at room temperature for 3 hours. Readings were taken on an EnVision reader (PerkinElmer) at excitation wavelength of 340 nm and emission wavelengths of 615 nm and 665 nm. Blank control group (DMSO, detection buffer, peptide) and high-concentration control group (DMSO, protein, peptide) were used as references. Results were calculated from the fluorescence signals at 665 nm and 615 nm, expressed as HTRF ratio: HTRF ratio = F(665 nm) / F(615 nm). In the following formula, HTRF ratio is abbreviated as F. Inhibition rate = [1-(F inhibitor -F blank control ) / F high control -F blank control [×100. Inhibition curve and half-maximum inhibitory concentration (IC50)] 50 The values ​​were calculated using GraphPad Prism 7 software.

[0036] Table 2. Inhibitory activity of the compounds against CBP BRD and BRD4 BD2 proteins determined by TR-FRET method. ; Note: The control drugs are SGC-CPB30 and ABBV-744.

[0037] The results of the bioactivity test (Table 2) show that the 23 compounds prepared in this invention all exhibited certain binding effects (inhibitory effects) on CBP BRD and BRD4 BD2 proteins.

[0038] The above embodiments are only used to illustrate the present invention and are not intended to limit the technical solutions described herein. Although the present invention has been described in detail with reference to the above embodiments, the present invention is not limited to the specific embodiments described above. Therefore, any modifications or equivalent substitutions to the present invention, as well as all technical solutions and improvements that do not depart from the spirit and scope of the invention, are covered within the scope of the claims of the present invention.

Claims

1. A class of isoxazolidine compounds or pharmaceutically acceptable salts thereof having CBP BRD and BRD4 BD2 inhibitory activity, characterized in that, The compound has the structural formulas shown in general formulas (I) and (II): (AND) (II) in, R is selected from It can be any one of heterocyclic groups or C1-C6 alkyl groups; wherein R1-R5 are selected from H, halogen, methoxy, nitro or trifluoromethoxy; and the heterocyclic group is selected from thiophene group.

2. The isoxazolidine compound or a pharmaceutically acceptable salt thereof having CBP BRD and BRD4 BD2 inhibitory activity according to claim 1, characterized in that, R is selected from any of the following substituents: 4-chlorophenyl, 2-chloro-4-fluorophenyl, 2-thienyl, butyl, phenyl, 2-chlorophenyl, n-propyl, 4-fluorophenyl, 2,4-difluorophenyl, 2-fluorophenyl, 3-fluorophenyl, 4-bromophenyl, 4-methoxyphenyl, 4-nitrophenyl, ethyl, 5-chloro-2-thienyl, 2,4,5-trifluorophenyl, 4-(trifluoromethoxy)-phenyl, 3-chloro-2-fluorophenyl, 2,5-difluorophenyl, 4-(trifluoromethyl)phenyl, 3,5-difluorophenyl, isobutyl.

3. The isoxazolidine compound or a pharmaceutically acceptable salt thereof having CBP BRD and BRD4 BD2 inhibitory activity according to claim 1, characterized in that, Compounds of general formula (I) are preferably any one of the following compounds: 4-Chloro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-3); 2-Chloro-4-fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-4); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)thiophene-2-sulfonamide (WJA-5); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)butane-1-sulfonamide (WJA-6); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-7); 2-Chloro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-8); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)butyramide (WJA-9); 4-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-12); 2,4-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-13); 2-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-14); 3-Fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-16); 4-Bromo-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-17); 4-Methoxy-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-21); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-nitrobenzenesulfonamide (WJA-22); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)ethanesulfonamide (WJA-23); 5-Chloro-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)thiophene-2-sulfonamide (WJA-26); 2,4,5-Trifluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-30); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-(trifluoromethoxy)benzenesulfonamide (WJA-31); 3-Chloro-2-fluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-32); 2,5-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-33); N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)-4-(trifluoromethyl)benzenesulfonamide (WJA-35); 3,5-Difluoro-N-(5-(3-methylbenzo[d]isoxazo-5-yl)-2-morpholinylphenyl)benzenesulfonamide (WJA-38); 2-Methyl-N-(5-(3-methylbenzo[d]isoxazol-5-yl)-2-morpholinylphenyl)propane-1-sulfonamide (WJA-39).

4. A pharmaceutical composition comprising a compound as described in any one of claims 1-3 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.

5. The use of any one of the isoxazolidine compounds according to any one of claims 1-4, or pharmaceutically acceptable salts thereof, or pharmaceutical compositions comprising such compounds, in the preparation of medicaments for the prevention or treatment of cancer, inflammation, and metabolic diseases associated with CBP, BRD, BRD4, and BD2.

6. The application according to claim 5, characterized in that, The cancers mentioned are leukemia, prostate cancer, or colon cancer, and the metabolic diseases are diabetes or coronary artery disease.

7. The method for preparing isoxazole biphenyl compounds according to any one of claims 1-3, characterized in that: Compounds of general formulas (I) and (II) were prepared using the following synthetic route: ； The substituent R in compounds of general formula (I) and (II) is as defined in any one of claims 1-3.

8. The method for preparing isoxazole biphenyl compounds according to claim 7, characterized in that, The method for preparing the compound of general formula (I) specifically includes the following steps: Step 1: 4-Bromo-1-fluoro-2-nitrobenzene and morpholine were subjected to a substitution reaction to obtain compound 2, namely 4-(4-bromo-2-nitrophenyl)morpholine; the molar ratio of 4-bromo-1-fluoro-2-nitrobenzene to morpholine was 1:1.1-2, the solvent was dimethyl sulfoxide, the reaction temperature was 80-130℃, and the reaction time was 0.25-2h; Step 2: Take compound 2, iron powder, acetic acid and ammonium chloride and carry out a reduction reaction to obtain compound 3, namely 5-bromo-2-morpholinoaniline; the molar ratio of compound 2, iron powder, acetic acid and ammonium chloride is 1:4-10:1.8-3:0.4-1, the solvent is ethanol and water, the reaction temperature is 50-80℃, and the reaction time is 12h. Step 3: Compound 3, pinacol diborate, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride, and potassium acetate were subjected to a borate esterification reaction to obtain compound 4, namely 2-morpholino-5-(4,4,5,5-tetramethyl-1,3,2-dioxoborane-2-yl)aniline; the molar ratio of compound 3, pinacol diborate, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride, and potassium acetate was 1:1-3:0.02-0.2:1-3, the solvent was 1,4-dioxane and dimethyl sulfoxide, the reaction system was under vacuum, the reaction temperature was 80-110℃, and the reaction time was 8-12h. Step 4: Compound 4 is subjected to sulfonation or acylation reactions with different sulfonyl chlorides or acyl chlorides to obtain a series of intermediates 5-1 or 5-2; the molar ratio of compound 4 to different sulfonyl chlorides or acyl chlorides is 1:1.0-3; the base used in the sulfonation reaction is pyridine, and the molar ratio of compound 4 to pyridine is 1:4-10; the base used in the acylation reaction is triethylamine, and the molar ratio of compound 4 to triethylamine is 1:1.5-3.5; the solvent is dichloromethane, the reaction temperature is room temperature to 40℃, and the reaction time is 2-12 h; Step 5: Take compound 5-1 or 5-2, 5-bromo-3-methylbenzisoxazole, bis(triphenylphosphine) palladium dichloride (PdCl2(PPh3)2), triphenylphosphine (PPh3), and potassium bicarbonate (KHCO3) and carry out a Suzuki coupling reaction to obtain the target compound; the molar ratio of compound 5-1 or 5-2, 5-bromo-3-methylbenzisoxazole, PdCl2(PPh3)2, PPh3, and KHCO3 is 1:1-1.3:0.025-0.1:0.05-0.2:1.5-3, the solvent is N,N-dimethylacetamide and water (volume ratio of 8-14:1), the reaction system is under vacuum, the reaction temperature is 85-110℃, and the reaction time is 12h.

9. The method for preparing a class of isoxazole diphenyl ether compounds according to claim 8, characterized in that: In step 1, the molar ratio of 4-(4-bromo-2-nitrophenyl)morpholine, 4-bromo-1-fluoro-2-nitrobenzene, and morpholine is 1:1.

5. In step 2, the molar ratio of compound 2, iron powder, acetic acid, and ammonium chloride is 1:9:2.2:0.

5. In step 3, the molar ratio of compound 3, pinacol diborate, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride, and potassium acetate is 1:1.22:0.034:

2. In step 4, the molar ratio of compound 4 to pyridine is 1:6.2, and the molar ratio of compound 4 to triethylamine is 1:

3.

10. The method for preparing a class of isoxazole diphenyl ether compounds according to claim 9, characterized in that: In step 5, the molar ratio of compound 5-1 or 5-2, 5-bromo-3-methylbenzisoxazole, PdCl2(PPh3)2, PPh3 and KHCO3 is 1:1.2:0.05:0.1:2; In step 4, the sulfonyl chloride is selected from R1SO2Cl, and the acyl chloride is selected from R1COCl; wherein R1 is selected from... It can be any one of heterocyclic groups or C1-C6 alkyl groups; wherein R1-R5 are selected from H, halogen, methoxy, nitro or trifluoromethoxy; and the heterocyclic group is selected from thiophene group.

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