Protein degradation targeting chimera targeting PAK1 and its preparation method and application

Abstract

The present invention relates to protein degradation-targeted chimeras targeting PAK1, their preparation methods, and applications, and belongs to the technical field of protein degradation agents. The present invention provides compounds having the structural formulas shown in Formula I-II, or pharmaceutically acceptable salts thereof, wherein R1 and Linker are as described in the claims and description. Compounds of Formula I-II, or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof, of the present invention can be used to prepare PAK1 protein degradation-targeted chimeras and for anti-tumor applications.

Description

Technical Field

[0001] The present invention relates to a protein degradation targeting chimera with PAK1 as a target, a preparation method and an application thereof, and belongs to the technical field of protein degradation agents. Background Art

[0002] p21-activated kinases (PAKs) have been identified as effectors of the Cdc42 and Rac1 small GTPases and are composed of two subgroups: subgroup I (PAK1, -2, and -3) and subgroup II (PAK4, -5, and -6). These six isoforms exhibit distinct and overlapping functions. It has been reported that PAK1(- / -), PAK5(- / -), and PAK6(- / -) have no effect on mouse survival or health, whereas PAK2(- / -) or PAK4(- / -) can cause embryonic lethality, and PAK3(- / -) can lead to cognitive impairment. PAK1 has emerged as a promising new target for cancer therapy due to its complex regulatory role in cancer cell proliferation and survival pathways. PAK1 has been found to be highly amplified and phosphorylated in a variety of malignancies, contributing to cancer cell adaptability and survival under various stresses. In addition, PAK1 has been shown to play an important regulatory role in the entire life cycle of tumorigenesis, growth and angiogenesis, metastasis and survival, immunity and metabolism to drug resistance through several key pathways such as Wnt / β-Catenin, PI3K-AKT-mTORC1, EGFR / HER2 / MAPK, and NF-κB.

[0003] Based on the core scaffolds of reported PAK1 inhibitors, they are chemically classified into a series including indolecarbazole, aminopyrazole, 2-aminopyridino[2,3-d]pyrimidine-7(8H)-1, and aminopyrimidine. These inhibitors exhibit potent kinase inhibitory activity against PAK1. Some, including PF3758309 (Pfizer Phase I), have entered various stages of human clinical trials, while others, such as FRAX-486 (Afraxis), AZ-13711265 (AstraZeneca), NVS-PAK1-1 (Novartis), and FRAX-1036 (Genentech), remain in the preclinical stage. Despite demonstrating potent inhibitory activity at the molecular level, these PAK1 inhibitors have yet to achieve clinical success. Possible reasons for the clinical failure of PAK1 inhibitors include: first, poor permeability, which prevents them from entering the cell membrane. Second, their poor pharmacokinetic properties prevent them from achieving effective concentrations. Furthermore, these inhibitors lack PAK subtype selectivity and exhibit more potent toxic side effects. Finally, the non-kinase functions of PAK1 protein play an important role in regulating tumor signaling pathways. In recent years, proteolysis targeting chimeras (PROTACs) have emerged as a new technology to induce efficient degradation of targeted proteins. Compared with classic kinase inhibitors, PROTACs can degrade the entire protein, thereby removing the enzymatic activity and non-enzymatic functions of the kinase. Therefore, this is a new idea for PROTACs to develop anti-tumor drugs targeting PAK1. Summary of the Invention

[0004] The technical problem solved by the present invention is to provide a protein degradation targeting chimera targeting PAK1.

[0005] The present invention provides a compound having a structural formula as shown in Formula I or II or a pharmaceutically acceptable salt thereof:

[0006]

[0007] Among them, R 1 is C1-C6 alkyl, C3-C6 cycloalkyl;

[0008] Linker is: n = 1-6;

[0009] R 2 (CH2) m 、(CH2) p (CH2CH2O) q ;

[0010] m=2-10, p=2-5, q=1-5;

[0011] Preferably, m=4-7, p=2-3, q=1-3;

[0012] The present invention preferably comprises a compound of formula I or a pharmaceutically acceptable salt thereof:

[0013] Among them, R 1 is C1-C4 alkyl, C3-C6 cycloalkyl; preferably methyl or cyclopropyl; Linker is: n1=2-5,n2=1-3;

[0014] Preferably, n1=4-5, n2=1-3;

[0015] The present invention preferably comprises a compound of formula II or a pharmaceutically acceptable salt thereof:

[0016] Among them, R 1 is C1-C4 alkyl, C3-C6 cycloalkyl; preferably methyl or cyclopropyl; Linker is:

[0017] n3=1-3, n4=2-4, n5=2-5, n6=2-4;

[0018] Preferably, n5=3 or 5, n6=2-4;

[0019] The present invention preferably comprises a compound having the following structure or a pharmaceutically acceptable salt thereof:

[0020]

[0021] The pharmaceutically acceptable salts of the compounds of the present invention may be nitrates, hydrochlorides, sulfates, phosphates or citrates of the compounds.

[0022] The present invention also provides a pharmaceutical composition comprising an effective dose of the above-mentioned compound or a pharmaceutically acceptable salt thereof. The compounds of the present invention can be formulated into the following forms by methods known in the art: tablets, capsules, aqueous or oily solutions, suspensions, emulsions, creams, ointments, gels, nasal sprays, suppositories, finely dispersed powders for inhalation, aerosols or sprays, sterile aqueous or oily solutions or suspensions, or sterile emulsions for parenteral administration (including intravenous, intramuscular, or infusion). Liquid formulations can be prepared using sterile water or water-propylene glycol solutions as solvents, and the active ingredient can also be formulated in aqueous polyethylene glycol solutions. Aqueous solutions for oral administration can be prepared by dissolving the active ingredient in water and adding appropriate colorants, flavorings, stabilizers, and thickeners as needed. Aqueous suspensions for oral administration can be prepared by dispersing the finely dispersed active ingredient in water along with a viscous material, such as natural or synthetic gums, resins, methylcellulose, carboxymethylcellulose, and other suspending agents known in the pharmaceutical art.

[0023] The pharmaceutical composition can be in unit dosage form. In these forms, the composition is divided into unit doses containing appropriate quantities of the active ingredient. The unit dosage form can be a packaged preparation containing discrete quantities of the preparation, such as boxed tablets, capsules, and powders in vials or ampoules. The unit dosage form can also be a capsule, cachet, or tablet, or any of these packaged forms can be an appropriate number.

[0024] The active ingredient of the pharmaceutical composition of the present invention may be the compound of the present invention alone, or may be a combination of the compound and other anti-tumor compounds as the active ingredient.

[0025] The present invention provides use of the above-mentioned compound or a pharmaceutically acceptable salt thereof or a pharmaceutical composition thereof in preparing a PAK1 degrader.

[0026] The present invention also provides use of the above compound or its pharmaceutically acceptable salt or its pharmaceutical composition in the preparation of anti-tumor drugs.

[0027] The tumor is preferably breast cancer.

[0028] During the treatment of tumors, the pharmaceutical composition of the present invention can be used in combination with other anti-tumor drugs for treatment.

[0029] In the treatment of tumors, such combination therapy can be achieved by administering the various therapeutic components simultaneously, sequentially or separately. Such combination products utilize the compounds of this invention within their effective dosage ranges and the other pharmaceutically active agents within their approved dosage ranges. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 Binding experiment of compound 33 to PAK1.

[0031] AB: SPR analysis of the binding ability of PAK1 to compound 33. CD: CETSA assay of compound 33 and PAK1 in MDA-MB-231 cells. E: Western blot analysis of PAK1 expression after treatment with different concentrations of compound 33. F: Western blot analysis of PAK1 expression after treatment with compound 33 and co-treatment with bortezomib or VHL.

[0032] Figure 2 Compound 33 inhibits the proliferation of MDA-MB-231 cells.

[0033] AB: The clonogenic ability of MDA-MB-231 cells after treatment with compound 33.

[0034] CD: The relative expression intensity of Ki-67 was measured by immunofluorescence. EF: The effect of compound 33 on the green fluorescence intensity of EdU in cells.

[0035] Figure 3 In vivo antiproliferative activity of compound 33.

[0036] A: Representative images of tumor volume and dissected tumor tissue in each group. BC: Tumor weight and volume were measured on the day of dissection in each group. D: Changes in body weight of mice in each group. E: Ki-67 expression was assessed by immunohistochemistry. DETAILED DESCRIPTION

[0037] The specific embodiments of the present invention are further described below in conjunction with examples, but the present invention is not limited to the scope of the examples.

[0038] Example 1 Synthesis of Compounds 1 to 7

[0039] Compounds 1 to 31 were synthesized using the following reaction formula:

[0040]

[0041] Synthesis route and conditions: (i) NH4OH, TEA, THF, rt, 2h; (ii) HCl(g), Et2O; (iii) Oxalylchloride, 170℃, 40min; (iv) mCPBA, CH2Cl2, then CH3NH2; (v) Brominated hydrocarbon or brominated ether, K2CO3, DMF; (vi) NaN3, DMF, 80℃, 8h; (vii) DMF, 90℃, 8h; (viii) Sodium L-Ascorbate, CuSO4, TEA, NBA.

[0042] Synthesis of intermediate 2

[0043] The starting material (1) (500 mg, 2.15 mmol) was dissolved in anhydrous THF (12 mL) and triethylamine (1.0 mL, 6.45 mmol) and ammonium hydroxide (2.5 mL) were added. The resulting mixture was stirred at room temperature for about 2 hours until the starting material was completely consumed. The THF was removed under reduced pressure and water (30 mL) and EtOAc (30 mL) were added for extraction. The organic layer was washed with water, brine, and dried Na2SO4, filtered and concentrated in vacuo. The mixture was purified by column chromatography (13%-15% EtOAc / hexane) to give intermediate 2 (440 mg, 97% yield) as a white solid.

[0044]

[0045] Intermediate 2 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 9.77 (1H,s), 8.57 (1H,s), 8.28 (1H,s), 8.03 (1H,s), 2.50 (3H,s); 13 C-NMR (100MHz, DMSO-d6), δ (ppm): 192.2, 176.1, 164.0, 160.4, 109.5, 14.0;

[0046] Synthesis of intermediate 4

[0047] The obtained ethyl 2-(2,4-dichlorophenyl)acetate (9.63 mmol) and 4-amino-2-methylthiopyrimidine-5-carbaldehyde (6.42 mmol) were dissolved in DMF (10 mL). K2CO3 (12.8 mmol) was added to the above solution at room temperature and refluxed at 70°C for 12 hours. The reaction mixture was extracted with dichloromethane (2 × 50 mL), and the organic phase was washed with saturated aqueous NaCl solution (100 mL) and dried over anhydrous Na2SO4. After filtration and concentration under reduced pressure, the product was isolated and purified by silica gel column chromatography (petroleum ether:ethyl acetate = 3:1) to obtain intermediate 4.

[0048]

[0049] Intermediate 4 is a white solid with a yield of 97%. 1H NMR(400MHz,DMSO-d6),δ(ppm):12.68(1H,s),8.90(1H,s),7.98(1H,s),7.73(1 H,d,J=2.0Hz),7.52(1H,dd,J=8.3,2.0Hz),7.45(1H,d,J=8.3Hz),2.58(3H,s); 13 C NMR (100MHz, DMSO-d6), δ (ppm): 172.8, 161.7, 157.5, 154.7, 137.2, 134.4, 134.2, 134.2, 133.5, 131.1, 129.3, 127.7, 109.2, 14.2.

[0050] Synthesis of intermediate 5

[0051] To a solution of intermediate 4 (402.0 mg, 1.19 mmol) in DMF (5 ml) was added 3-chloroperoxybenzoic acid (205.4 mg, 1.19 mmol) and stirred at 0°C for 2 hours. Methylamine (40% aqueous solution) was added and the reaction mixture was stirred at 50°C for 1.5 hours. The starting material was completely consumed, and water (15 ml) was added and stirred for 1 hour. The solid was collected by filtration and washed with DCM (5 ml) to give intermediate 5 as a light yellow insoluble solid in a 74% yield.

[0052]

[0053] Synthesis of intermediates 6a-6g

[0054] To a DMF (10.0 ml) solution containing intermediate 5 (205.5 mg, 0.64 mmol) and a bromohydrocarbon derivative (0.95 mmol) was added K2CO3 (264.0 mg, 1.91 mmol) and a small amount of potassium iodide catalyst, and the mixture was heated to 55 ° C and reacted for 6 hours. When the starting material was completely consumed, the hot reaction suspension was poured into water (30 ml) and extracted with dichloromethane (3×30 ml). The organic layers were combined, washed with water-saturated sodium bicarbonate solution and brine, and then dried over anhydrous sodium sulfate. After removing the solvent under reduced pressure, the crude product was obtained and purified by silica gel column chromatography (petroleum ether / ethyl acetate=3:1) to obtain a light yellow solid with a yield of 64-72%.

[0055]

[0056] Intermediate 6a 1H-NMR(400MHz, DMSO-d6), δ(ppm): 8.63(1H, s), 7.90(1H, d, J = 4.6Hz), 7.80(1H, s), 7.69(1H, d, J = 2.1Hz), 7.46(1H, dd, J = 8.3, 2.1, Hz), 7.43(1H, d, J = 8.3Hz), 4.35 - 4.27(2H, m), 3.80 - 3.71(2H, m), 3.59(2H, m), 2.92(3H, d, J = 4.6Hz), 1.85(4H, m); 13 C-NMR(150MHz, DMSO-d6), δ(ppm): 161.5, 161.4, 158.8, 155.2, 136.6, 136.5, 134.9, 134.3, 133.3, 133.2, 128.7, 127.2, 103.9, 40.1, 34.5, 34.3, 31.2, 29.9;

[0057] Intermediate 6b 1 H-NMR(600MHz, CDCl3), δ(ppm): 7.31(1H, s), 7.30(1H, s), 7.25(1H, s), 7.24(1H, s), 6.54(1H, s), 3.48(8H, s) 1.49(9H, s); 13 C-NMR(150MHz, CDCl3), δ(ppm): 154.6, 154.5, 137.3, 128.9, 128.9, 128.3, 121.3, 121.3, 80.7, 58.5, 57.3, 45.2, 45.1, 43.8, 42.7, 28.3, 28.3, 28.3;

[0058] Intermediate 6c 1 H-NMR(400MHz, DMSO-d6), δ(ppm): 8.62(1H, s), 7.88(1H, d, J = 4.6Hz), 7.78(1H, s), 7.69(1H, d, J = 2.1Hz), 7.48(1H, dd, J = 8.3, 2.1Hz), 7.42(1H, d, J = 8.3Hz), 4.29(2H, d, J = 7.2Hz), 3.50(2H, d, J = 6.7Hz), 2.90(3H, d, J = 4.6Hz),​13C-NMR (150 MHz, DMSO-d6), δ (ppm): 160.8, 159.8, 156.9, 155.5, 136.3, 134.6, 134.2, 133.4, 133.2, 128.7, 127.2, 124.7, 104.1, 40.1, 32.4, 31.2, 29.8, 27.2, 27.1, 26.4;

[0059] Intermediate 6d 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 8.61 (1H, s), 7.87 (1H, d, J = 4.6 Hz), 7.77 (1H, s), 7.67 (1H, d, J = 2.0 Hz), 7.47 (1H, dd, J = 8.3, 2.0 Hz), 7.41 (1H, d, J = 8.3 Hz), 4.30 (2H, d, J = 7.1 Hz), 3.50 (2H, d, J = 6.7 Hz), 2.91 (3H, d, J = 4.6 Hz), 1.82 - 1.76 (2H, m), 1.70 - 1.65 (2H, m), 1.45 - 1.36 (4H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 160.8, 159.2, 1,56.2, 155.6, 136.3, 134.5, 134.2, 133.4, 133.2, 128.8, 127.3, 125.0, 104.1, 40.1, 32.5, 31.2, 28.0, 27.8, 27.5, 27.0, 26.5;

[0060] Intermediate 6e 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 8.62 (1H, s), 7.91 (1H, d, J = 4.7 Hz), 7.80 (1H, s), 7.69 (1H, d, J = 2.1 Hz), 7.48 (1H, dd, J = 8.3, 2.1 Hz), 7.42 (1H, d, J = 8.3 Hz), 4.53 (2H, t, J = 6.3 Hz), 3.80 - 3.71 (4H, m), 3.53 (2H, d, J = 5.8 Hz), 2.92 (3H, d, J = 4.7 Hz); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 161.4, 160.8, 158.9, 155.2, 136.7, 134.6, 134.1, 133.15, ¾33.1, 128.6, 127.1, 123.5, 103.8, 69.8, 66.2, 60.1, 32.2, 27.8;

[0061] Intermediate 6f 1 H NMR (400MHz, DMSO-d6), δ (ppm): 8.62 (1H, s), 7.90 (1H, d, J = 4.7Hz), 7.79 (1H, s), 7.68 (1H, d, J = 2.0Hz), 7.47 (1 H,dd,J=8.3,2.0,Hz),7.41(1H,d,J=8.3Hz),4.51(2H,t,J=6.3Hz),3.74-3.50(10H,m),2.91(3H,d,J=4.6Hz); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 161.7, 161.3, 159.1, 155.9, 137.2, 135.1, 134.7, 133.7, 133.7, 129.2, 127.6, 104.4 72.8,70.8,70.0,67.1,40.5,32.7,31.6,30.3;

[0062] Intermediate 6g 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 8.62 (1H, s), 7.90 (1H, d, J = 4.7Hz), 7.79 (1H, s), 7.68 (1H, d, J = 2.0Hz), 7.47 (1H, dd, J =8.3,2.0Hz),7.41(1H,d,J=8.3Hz),4.51(2H,t,J=6.3Hz),3.72-3.68(4H,m),3.59-3.48(10H,m),2.91(3H,d,J=4.6Hz); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 160.8, 160.2, 158.2, 154.8, 136.1, 134.1, 133.6, 132.6 ,132.6,128.1,126.6,123.1,103.3,69.8,69.2,69.2,69.1,69.0,66.0,39.5,31.6,27.3;

[0063] Synthesis of intermediates 7a-g

[0064] To a solution of intermediate 6 (0.24 mmol) in DMF (5 ml) was added NaN 3 (0.24 mmol). The mixture was stirred at 80° C. for 12 hours. After complete consumption of the starting material, the mixture was poured into water (30 ml) and extracted with ethyl acetate (3×30 ml). The organic layer was washed with saturated aqueous sodium bicarbonate solution and brine, then dried over anhydrous sodium sulfate. After removal of the solvent under reduced pressure, intermediate 7 was obtained as a white solid in a yield of 82-90%.

[0065]

[0066] Synthesis of Intermediate 10

[0067] To a solution of 2-(2,6-dioxopiperidin-3-yl)-4-fluoroisoindole-1,3-dione 8 (200 mg, 0.724 mmol) in DMF (10 ml) was added propargylamine 9 (46 μL, 0.724 mmol) under nitrogen, and the reaction mixture was stirred at 90 ° C for 12 hours. The mixture was poured into water (30 ml) and extracted with DCM (3×30 ml). The combined organic layers were washed with a water-saturated aqueous sodium carbonate solution and brine, and then dried over anhydrous sodium sulfate. The solvent was removed under reduced pressure and purified by silica gel column chromatography (petroleum ether / ethyl acetate=3:1) to give a yellow-green solid with a yield of 90%.

[0068]

[0069] Intermediate 10 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 11.10 (1H, s), 7.67-7.60 (1H, m), 7.27-7.22 (1H, m), 7.17-7.10 (1H, m), 6.93 (1H, t, J = 6 .2Hz),5.10-5.04(1H,m),4.18-4.16(1H,m),3.04(2H,s),2.89(1H,s),2.73(1H,s),2.61-2.54(1H,m),2.08-1.99(1H,m); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 172.2, 169.5, 168.0, 166.7, 144.6, 135.5, 131.5, 117.4, 110.8, 109.7, 80.3, 73.2, 62.5, 30.9, 30.4, 21.5;

[0070] Preparation of compounds 1 to 7

[0071] To a solution of n-butanol (5 milliliters) containing intermediate 7 (0.24 mmol), intermediate 10, sodium L-ascorbate (0.2 mmol) and TEA (1.2 mmol) was added CuSO 4 (0.20 mmol). The reaction mixture was stirred at room temperature for 12 hours. The residue was concentrated under reduced pressure and purified by silica gel SEL flash chromatography (dichloromethane / methanol=20:1) to a yellow solid in a yield of 30-34%.

[0072] Compound 1 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 11.10 (1H, s), 8.61 (1H, s), 8.00 (1H, s), 7.86 (1H, d, J = 4.6Hz), 7.78 (1H,s),7.66(1H,d,J=2.1Hz),7.52-7.46(2H,m),7.41(1H,d,J=8.3Hz),7.12(1H,d,J=8.5Hz),7.06(1H ,J=6.0Hz),7.01(1H,J=7.0Hz),5.06(1H,dd,J=12.8,5.3Hz),4.60(2H,d,J=6.0Hz),4.38(2H,m),4.29 (2H,m),2.93-2.85(4H,m),2.61-2.53(2H,m),2.00-2.00(1H,m),1.86-1.81(2H,m),1.67-1.63(2H,m); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 172.2, 169.5, 168.1, 166.7, 161.3, 160.2, 158.8, 154.4, 145.2, 143.8, 136.0, 135.4, 134.3, 132.6, 132.5,131.5,128.1,126.5,123.0,122.2,117.0,110.3,109.1,103.3,63.1,54.3,48.5,48.0,30.4,27.2,26.9,23.7,21.5.HRMS(ESI) + calculated for C 34 H 30 Cl2N 10 O5,[M+H] + :m / z 729.1850, found 729.1910; HPLC(t R =3.102 min, 99.0%)

[0073] Compound 2 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 11.09 (1H, s), 8.61 (1H, s), 7.99 (1H, s), 7.86 (1H, d, J = 4.4 Hz), 7.77 (1H, s), 7.68 (1H, d, J = 2.1 Hz), 7.55 (1H, t, J = 7.8 Hz), 7.47 (1H, dd, J = 8.3, 2.1 Hz), 7.41 (1H, d, J = 8.3 Hz), 7.15 (1H, d, J = 8.7 Hz), 7.07 - 7.02 (2H, m), 5.05 (1H, dd, J = 12.9, 5.4 Hz), 4.58 (2H, d, J = 6.0 Hz), 4.33 - 4.24 (4H, m), 2.89 - 2.85 (4H, m), 2.60 - 2.52 (2H, m), 2.04 - 1.99 (1H, m), 1.88 - 1.81 (2H, m), 1.70 - 1.65 (2H, m), 1.27 - 1.25 (2H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 173.3, 170.6, 169.2, 167.7, 162.4, 161.2, 159.9, 155.5, 146.3, 144.9, 137.0, 136.6, 135.4, 134.7, 133.7, 133.6, 132.6, 130.1, 129.1, 127.6, 124.0, 123.1, 118.1, 111.4, 110.1, 104.3, 65.4, 49.6, 49.0, 29.8, 28.3, 27.0, 23.8, 22.6, 22.6. HRMS (ESI) + calculated for C 35 H 32 Cl2N 10 O5, [M + H] + : m / z 743.2007, found 743.2052; HPLC (t R = 3.180 min, 95.9%).

[0074] Compound 3 11H NMR (400 MHz, DMSO-d6), δ (ppm): 11.09 (1H, s), 8.61 (1H, s), 8.00 (1H, s), 7.84 (1H, d, J = 4.0 Hz), 7.76 (1H, s), 7.67 (1H, d, J = 2.0 Hz), 7.52 (1H, t, J = 7.8 Hz), 7.47 (1H, dd, J = 8.3, 2.0 Hz), 7.41 (1H, d, J = 8.3 Hz), 7.13 (1H, d, J = 8.5 Hz), 7.06 (1H, t, J = 6.0 Hz), 7.00 (1H, d, J = 7.0 Hz), 5.05 (1H, dd, J = 12.8, 5.4 Hz), 4.57 (2H, d, J = 6.0 Hz), 4.30 (2H, d, J = 6.9 Hz), 4.23 (2H, d, J = 7.0 Hz), 2.92 - 2.83 (4H, m), 2.60 - 2.52 (2H, m), 2.05 - 1.98 (1H, m), 1.81 - 1.74 (2H, m), 1.62 - 1.61 (2H, m), 1.33 - 1.27 (4H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 172.9, 170.1, 168.8, 167.3, 162.0, 160.8, 159.5, 155.0, 145.9, 136.6, 136.1, 135.0, 134.3, 133.3, 133.2, 132.1, 128.7, 127.2, 123.6, 122.8, 117.6, 115.6, 1--9, 109.7, 104.0, 55.0, 49.3, 48.6. HRMS (ESI) + calculated for C 36 H 34 Cl2N 10 O5, [M + H] + : m / z 757.2163, found 757.2242; HPLC (t R = 2.716 min, 98.9%).

[0075] Compound 4 11H-NMR (600 MHz, DMSO-d6), δ (ppm): 11.10 (1H, s), 8.62 (1H, s), 7.99 (1H, s), 7.86 (1H, d, J = 4.3 Hz), 7.77 (1H, s), 7.67 (1H, d, J = 2.1 Hz), 7.54 (1H, t, J = 7.8 Hz), 7.47 (1H, dd, J = 8.3, [2.1 Hz]), 7.41 (1H, d, J = 8.3 Hz), 7.13 (1H, d, J = 8.6 Hz), 7.06 - 7.02 (2H, m), 5.05 (1H, dd, J = 12.9, 5.4 Hz), 4.57 (2H, d, J = 6.1 Hz), 4.30 - 4.26 (4H, m), 2.90 - 2.84 (4H, m), 2.60 - 2.52 (2H, m), 2.03 - 1.99 (1H, m), 1.77 - 1.72 (2H, m), 1.65 - 1.62 (2H, m), 1.27 (4H, m), 1.20 - 1.15 (2H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 172.9, 170.1, 168.8, 167.3, 162.1, 160.8, 159.5, 155.1, 145.9, 136.6, 136.2, 135.0, 134.3, 133.3, 133.2, 132.2, 128.7, 127.2, 123.7, 122.8, 117.7, 111.0, 109.7, 104.0, 49.3, 48.7, 48.6, [31.0], 29.7, 28.0, 27.9, 27.0, 26.4, 25.8, 22.2. HRMS (ESI) + calculated for C 37 H 36 Cl2N 10 O5, [M + H] + : m / z 771.2320, found 771.2403; HPLC (t R = 2.754 min, 99.7%).

[0076] Compound 5 1 Note: In the 13C-NMR data, the value in brackets is likely an approximation or a value that may be subject to some uncertainty in the original data. The translation preserves all the original information as accurately as possible while converting the Chinese chemical terms to English.1H-NMR (400 MHz, DMSO-d6), δ (ppm): 11.09 (1H, s), 8.60 (1H, s), 7.90 (1H, s), 7.85 (1H, d, J = 4.6 Hz), 7.77 (1H, s), 7.66 (1H, d, J = 2.1 Hz), 7.50 (1H, t, J = 7.7 Hz), 7.46 (1H, dd, J = 8.3, 2.1 Hz), 7.40 (1H, d, J = 8.3 Hz), 7.13 (1H, d, J = 8.6 Hz), 6.99 (2H, m), 5.03 (1H, dd, J = 12.8, 5.5 Hz), 4.53 (2H, d, J = 4.9 Hz), 4.44 (2H, t, J = 4.8 Hz), 4.37 (2H, t, J = 5.9 Hz), 3.83 (2H, t, J = 5.1 Hz), 3.68 - 3.65 (2H, m), 2.90 - 2.84 (4H, m), 2.60 - 2.55 (2H, m), 2.02 - 1.98 (1H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 173.3, 170.6, 169.2, 167.7, 162.3, 161.3, 159.9, 155.6, 146.2, 144.8, 137.3, 136.5, 135.2, 134.7, 133.7, 133.6, 132.5, 129.1, 127.6, 123.9, 123.6, 118.0, 111.3, 110.1, 104.3, 68.9, 66.9, 55.4, 50.0, 49.0, 38.0, 31.4, 28.3, 22.6. HRMS (ESI) + calculated for C 34 H 30 Cl2N 10 O six, [M + H] + : m / z 745.1800, found 745.1878; HPLC (t R = 2.659 min, 99.8%).

[0077] Compound 6 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 11.09 (1H, s), 8.61 (1H, s), 7.96 (1H, s), 7.87 (1H, d, J = 4.3 Hz), 7.78 (1H, s), 7.66 (1H, d, J = 2.1 Hz), 7.53 (1H, t, J = 7.8 Hz), 7.46 (1H, dd, J = 8.3, 2.1 Hz), 7.40 (1H, d, J = 8.3 Hz), 7.14 (1H, d, J = 8.5 Hz), 7.02 (2H, m), 5.04 (1H, dd, J = 12.8, 5.3 Hz), 4.53 (2H, d, J = 5.0 Hz), 4.46 - 4.42 (4H, m), 3.74 - 3.65 (4H, m), 3.52 - 3.45 (4H, m), 2.88 - 2.83 (4H, m), 2.59 - 2.54 (2H, m), 2.03 - 1.98 (1H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 172.9, 170.1, 168.8, 167.3, 162.0, 160.9, 159.5, 155.3, 145.9, 144.4, 136.9, 136.1, 134.8, 134.3, 133.3, 133.2, 132.1, 128.7, 127.2, 123.5, 123.3, 117.6, 111.0, 109.7, 103.9, 69.5, 69.4, 68.8, 66.6, 59.8, 55.0, 49.4, 48.6, 31.0, 27.9, 22.2. HRMS (ESI) + calculated for C 36 H 34 Cl2N 10 O7, [M + H] + : m / z 789.2062, found 789.2167; HPLC (t R = 2.633 min, 98.6%).

[0078] Compound 7 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 11.09 (1H, s), 8.62 (1H, s), 7.97 (1H, s), 7.88 (1H, d, J = 4.4 Hz), 7.79 (1H, s), 7.68 (1H, d, J = 2.0 Hz), 7.53 (1H, t, J = 7.8 Hz), 7.47 (1H, dd, J = 8.3, 2.0 Hz), 7.41 (1H, d, J = 8.3 Hz), 7.15 (1H, d, J = 8.4 Hz), 7.04 - 7.01 (2H, m), 5.05 (1H, dd, J = 12.7, 5.3 Hz), 4.57 (2H, d, J = 5.8 Hz), 4.49 - 4.45 (4H, m), 3.76 - 3.42 (12H, m), 2.90 - 2.88 (4H, m), 2.60 - 2.53 (2H, m), 2.03 - 1.99 (1H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 172.9, 170.1, 168.8, 167.3, 162.0, 160.9, 159.5, 155.3, 145.9, 144.4, 136.8, 136.1, 135.5, 134.8, 134.3, 133.3, 133.2, 132.1, 128.7, 127.2, 123.5, 123.3, 11 7.6, 111.0, 109.7, 103.9, 69.7, 69.7, 69.5, 68.7, 68.2, 66.6, 49.4, 48.6, 31.0, 27.9, 22.2. HRMS (ESI) + Calculated for C 38 H 38 C l2 N 10 O8, [M + H] + : m / z 833.2324, found 833.2431; HPLC (t R = 2.632 min, 9 8.6%).

[0079] Synthesis of Compound 8 - 14

[0080]

[0081] Synthesis route and conditions: (i) NH4OH, TEA, THF, rt, 2h; (ii) HCl(g), Et2O; (iii) Oxalylchloride, 170℃, 40min; (iv) mCPBA, CH2Cl2, then cyclopropylamine; (v) Brominated hydrocarbon or brominated ether, K2CO3, DMF; (vi) NaN3, DMF, 80℃, 8h; (vii) DMF, 90℃, 8h; (viii) Sodium L-Ascorbate, CuSO4, TEA, NBA.

[0082] Synthesis of intermediate 12

[0083] To a solution of intermediate 4 in DMF (5 ml) was added 3-chloroperoxybenzoic acid (205.4 mg, 1.19 mmol) at 0°C. After stirring for 2 hours, cyclopropylamine (6 ml) was added and the reaction mixture was stirred at 50°C for 1.5 hours. The starting material was completely consumed, and water (15 ml) was added and stirred for 1 hour. The solid was collected by filtration and washed with DCM (5 ml) to give intermediate 12 as a light yellow insoluble solid in 83% yield.

[0084]

[0085] Synthesis of Intermediates 13a-13g To a DMF (10.0 ml) solution containing intermediate 12 (222 mg, 0.64 mmol) and a bromohydrocarbon derivative (0.95 mmol) were added K2CO3 (264.0 mg, 1.91 mmol) and a small amount of potassium iodide catalyst, and the mixture was heated to 55°C and reacted for 6 hours. After the starting material was completely consumed, the hot reaction suspension was poured into water (30 ml) and extracted with dichloromethane (3×30 ml). The organic layers were combined, washed with a water-saturated sodium bicarbonate solution and brine, and then dried over anhydrous sodium sulfate. After removing the solvent under reduced pressure, the crude product was obtained and purified by silica gel column chromatography (petroleum ether / ethyl acetate=3:1) to obtain a light yellow solid with a yield of 72-75%.

[0086]

[0087] Intermediate 13a 1H-NMR (400MHz, DMSO-d6), δ (ppm): 8.64 (1H, s), 8.18 (1H, s), 7.81 (1H, s), 7.70 (1H, d, J = 2.0Hz), 7.49 (1H, dd, J = 8.3, 2.0H z),7.43(1H,d,J=8.3Hz),4.37(2H,s),3.57-3.58(2H,m),2.84(1H,s),1.86(4H,s),0.76-0.75(2H,m),0.58-0.55(2H,m); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 162.2, 161.2, 158.6, 155.6, 136.8, 135.2, 13 4.7,133.7,129.2,127.7,124.9,104.9,41.0,30.6,29.5,24.5,24.4,7.1,6.7.

[0088] Intermediate 13b 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 8.63 (1H, s), 8.17 (1H, s), 7.80 (1H, s), 7.70 (1H, d, J = 2.1Hz), 7.49 (1H, dd, J = 8.3, 2.1Hz), 7.42 (1H, d, J = 8.3 Hz),4.34-4.22(2H,m),3.53(2H,t,J=6.7Hz),2.83(1H,s),1.80-1.83( 2H,m),1.73(2H,s),1.47(2H,s),0.77-0.75(2H,m),0.59-0.55(2H,m); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 162.8, 161.2, 158.7, 155.6, 136.8, 135.2, 13 4.7,133.7,129.2,127.7,124.9,104.9,41.0,32.7,31.3,27.5,24.4,23.5,6.7.

[0089] Intermediate 13c 1H-NMR (400MHz, DMSO-d6), δ (ppm): 8.62 (1H, s), 8.15 (1H, s), 7.79 (1H, s), 7.69 (1H, d, J = 2.1Hz), 7.48 (1H, dd, J = 8.3, 2.1Hz), 7.42 (1H, d, J = 8.3 Hz),4.32(2H,s),3.50(2H,t,J=6.7Hz),2.82(1H,s),1.82-1.70(4H,m) ,1.44-1.37(4H,m),1.47(2H,s),0.76-0.74(2H,m),0.58-0.54(2H,m); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 163.5, 161.2, 158.4, 155.7, 136.6, 135.2, 134.7, 133.7,129.2,127.7,104.9,41.0,32.9,31.6,30.3,27.6,26.9,25.7,24.4,7.1,6.7.

[0090] Intermediate 13d 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 8.63 (1H, s), 8.16 (1H, s), 7.80 (1H, s), 7.69 (1H, d, J = 2.1Hz), 7.49 (1H, dd, J = 8.3, 2.1Hz), 7.42 (1H ,d,J=8.3Hz),4.34(2H,s),3.50(2H,t,J=6.7Hz),2.82(1H,s),1.78-1.70(4H,m),1.34(6H,s),0.75-0.74(2H,m),0.59-0.55(2H,m); 13 C-NMR (150 MHz, DMSO-d6), δ (ppm): 161.9, 161.2, 158.3, 155.7, 136.8, 135.2, 134.7, 133.7, 129.2, 127.7, 125.1, 104.9, 41.0, 34.9, 32.9, 31.6, 30.3, 29.1, 27.5, 27.0, 25.9, 24.4, 7.2, 6.7. Intermediate 13e 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 8.63 (1H, s), 8.18 (1H, s), 7.80 (1H, s), 7.69 (1H, d, J = 2.0 Hz), 7.49 (1H, dd, J = 8.3, 2.0 Hz), 7.42 (1H, d, J = 8.3 Hz), 4.54 (2H, s), 3.80 (4H, s), 3.54 - 3.52 (2H, m), 2.85 (1H, s), 0.76 - 0.75 (2H, m), 0.59 - 0.55 (2H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 162.8, 161.3, 159.3, 155.7, 137.1, 135.2, 134.7, 133.7, 129.2, 127.6, 124.8, 124.7, 104.9, 70.4, 66.8, 60.7, 32.8, 24.4, 14.4, 6.7.

[0091] Intermediate 13f 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 8.63 (1H, s), 8.17 (1H, s), 7.80 (1H, s), 7.70 (1H, d, J = 2.09 Hz), 7.49 (1H, dd, J = 8.3, 2.1 Hz), 7.42 (1H, d, J = 8.3 Hz), 4.53 (2H, s), 3.76 - 3.50 (10H, m), 2.83 (1H, s), 0.75 - 0.74 (2H, m), 0.58 - 0.55 (2H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 161.8, 160.2, 158.4, 154.6, 136.1, 134.2, 133.6, 132.6, 128.1, 126.6, 123.7, 123.7, 103.8, 71.7, 69.7, 68.9, 66.0, 59.6, 31.6, 23.3, 5.6.

[0092] Intermediate 13g 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 8.63 (1H, s), 8.17 (1H, s), 7.81 (1H, s), 7.69 (1H, d, J = 2.1 Hz), 7.48 (1H, dd, J = 8.3, 2.1 Hz), 7.42 (1H, d, J = 8.3 Hz), 4.53 (2H, s), 3.70 - 3.46 (1十四H, m), 2.83 (1H, s), 0.75 - 0.74 (2H, m), 0.58 - 0.54 (2H, m);13 C-NMR (150MHz, DMSO-d6), δ (ppm): 163.0, 161.3, 159.6, 155.6, 137.1, 135.3, 134.7, 133.7, 129 .2,127.6,124.8,124.7,104.8,70.8,70.3,70.2,70.1,70.0,67.1,44.0,31.6,24.4,14.4,6.7.

[0093] Synthesis of intermediates 14a-14g

[0094] To a solution of intermediate 13 (0.24 mmol) in DMF (5 ml) was added NaN3 (0.24 mmol). The mixture was stirred at 80°C for 12 hours. After complete consumption of the starting material, the mixture was poured into water (30 ml) and extracted with ethyl acetate (3 x 30 ml). The organic layer was washed with saturated aqueous sodium bicarbonate solution and brine, then dried over anhydrous sodium sulfate. After removal of the solvent under reduced pressure, intermediate 14 was obtained as a white solid in a yield of 74-85%.

[0095]

[0096] Synthesis of Compounds 8-14

[0097] To a solution of n-butanol (5 ml) containing intermediate 14 (0.24 mmol), intermediate 10, sodium L-ascorbate (0.2 mmol) and TEA (1.2 mmol) was added CuSO 4 (0.20 mmol). The reaction mixture was stirred at room temperature for 12 hours. The residue was concentrated under reduced pressure and purified by silica gel SEL flash chromatography (dichloromethane / methanol=20:1) to a yellow solid in a yield of 30-34%.

[0098] Compound 8 1H-NMR(400MHz, DMSO-d6), δ(ppm): 11.10(1H, s), 8.62(1H, s), 8.12(1H, d, J = 7.8Hz), 7.99(1H, s), 7.79(1H, s), 7.67(1H, d, J = 2.0Hz), 7.52 - 7.46(2H, m), 7.41(1H, d, J = 8.3Hz), 7.12(1H, d, J = 8.3Hz), 7.05(1H, t, J = 6.0Hz), 7.01(1H, d, J = 7.0Hz), 5.06(1H, dd, J = 12.8, 5.3Hz), 4.57(2H, d, J = 6.0Hz), 4.40 - 4.32(4H, m), 2.93 - 2.73(2H, m), 2.57 - 2.52(2H, m), 2.06 - 2.00(1H, m), 1.85(2H, m), 1.67(2H, m), 0.72 - 0.68(2H, m), 0.50(2H, s); 13 C-NMR(150MHz, DMSO-d6), δ(ppm): 173.3, 170.5, 169.2, 167.7, 163.1, 161.3, 159.7, 155.4, 146.3, 137.0, 136.5, 135.3, 134.7, 133.7, 133.6, 132.6, 132.5, 129.1, 127.6, 124.4, 123.3, 122.2, 118.0, 111.3, 110.1, 104.8, 60.2, 55.4, 49.6, 49.0, 31.4, 27.9, 24.8, 22.6, 14.4, 6.7. HRMS(ESI) + calculated for C 36 H 32 Cl2N 10 O5, [M + H] + : m / z 755.2007, found 755.2076; HPLC(t R = 3.233 min, 99.08%).

[0099] Compound 9 1 H-NMR(400MHz, DMSO-d6), δ(ppm): 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 11.10 (1H, s), 8.62 (1H, s), 8.14 (1H, d, J = 7.8 Hz), 8.00 (1H, s), 7.79 (1H, s), 7.69 (1H, d, J = 2.1 Hz), 7.56 (1H, t, J = 7.8 Hz), 7.48 (1H, dd, J = 8.3, 2.1 Hz), 7.42 (1H, d, J = 8.3 Hz), 7.15 (1H, d, J = 8.6 Hz), 7.07 - 7.03 (2H, m), 5.06 (1H, dd, J = 12.8, 5.3 Hz), 4.58 (2H, d, J = 5.9 Hz), 4.33 - 4.30 (4H, m), 2.93 - 2.73 (2H, m), 2.61 - 2.56 (2H, m), 2.06 - 1.99 (1H, m), 1.89 - 1.82 (2H, m), 1.71 (2H, m), 1.53 - 1.46 (2H, m), 0.69 - 0.67 (2H, m), 0.53 (2H, s); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 173.9, 172.2, 169.5, 168.2, 162.0, 160.2, 158.6, 154.3, 145.2, 137.4, 135.9, 135.5, 134.3, 133.7, 132.6, 132.5, 131.5, 128.1, 126.5, 123.7, 122.1, 117.8, 117.0, 110.3, 109.1, 70.8, 54.3, 48.6, 48.0, 29.2, 28.4, 27.9, 26.0, 23.9, 21.5, 13.4, 5.6. HRMS (ESI) + calculated for C 37 H 34 Cl2N 10 O5, [M + H] + : m / z 769.2163, found 769.2188; HPLC (t ⏟ R = 3.379 min, 99.18%).

[0100] Compound ⑩ 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 11H NMR (400 MHz, DMSO-d6), δ (ppm): 11.10 (1H, s), 8.62 (1H, s), 8.13 (1H, d, J = 7.8 Hz), 8.00 (1H, s), 7.78 (1H, s), 7.68 (1H, d, J = 2.0 Hz), 7.53 (1H, t, J = 7.8 Hz), 7.48 (1H, dd, J = 8.3, 2.0 Hz), 7.42 (1H, d, J = 8.3 Hz), 7.13 (1H, d, J = 8.6 Hz), 7.06 (1H, t, J = 5.8 Hz), 7.00 (1H, d, J = 7.0 Hz), 5.05 (1H, dd, J = 12.8, 5.3 Hz), 4.58 (2H, d, J = 5.63 Hz), 4.31 - 4.27 (4H, m), 2.92 - 2.73 (2H, m), 2.60 - 2.56 (2H, m), 2.02 - 1.99 (1H, m), 1.81 - 1.75 (2H, m), 1.65 (2H, m), 1.53 - 1.33 (4H, m), 0.74 - 0.69 (2H, m), 0.54 (2H, s); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 172.2, 169.5, 168.1, 166.7, 162.1, 160.1, 154.3, 145.2, 135.8, 135.5, 134.3, 133.7, 132.6, 132.5, 131.5, 128.1, 126.5, 123.7, 123.7, 122.1, 117.8, 117.0, 110.3, 109.1, 70.8, 54.3, 48.6, 48.0, 30.6, 29.3, 28.4, 28.1, 26.3, 24.9, 21.5, 13.4, 5.6. HRMS (ESI) + calculated for C 38 H 36 Cl2N 10 O5, [M + H] + : m / z 783.2320, found 783.2339; HPLC (t R = 3.346 min, 97.17%).

[0101] Compound 11 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 11.10 (1H, s), 8.63 (1H, s), 8.15 (1H, s), 8.00 (1H, s), 7.79 (1H, s), 7.70 - 7.68 (1H, m), 7.57 - 7.53 (1H, m), 7.48 (1H, dd, J = 8.3, 2.1 Hz), 7.42 (1H, d, J = 8.3 Hz), 7.14 (1H, d, J = 8.6 Hz), 7.07 - 7.03 (2H, m), 5.06 (1H, dd, J = 12.8, 5.3 Hz), / / 4.58 (2H, d, J = 6.0 Hz), 4.30 - 4.27 (4H, m), 4.15 - 4.13 (2H, m), 2.93 - 2.79 (2H, m), 2.61 - 2.54 (2H, m), 2.04 - 1.99 (1H, m), 1.79 - 1.71 (2H, m), 1.66 - 1.60 (2H, m), 1.43 - 1.35 (4H, m), 0.75 - 0.70 (2H, m), 0.57 - 0.53 (2H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 173.3, 170.5, 169.2, 167.7, 163.1, 161.2, 159.6, 15`5.4, 146.3, 144.9, 136.9, 136.6, 135.4, 134.7, 133.7, 133.6, 132.6, 129.1, 127.6 124.5, 123.2, 122.2, 118.1, 111.4, 110.1, 104.9, 55.4, 49.7, 49.0, 31.4, 30.1, 28.5, 27.5, 26.8, 26.2, 22.6, 14.4, 6.7. HRMS (ESI) + calculated for C 39 H 38 Cl2N 10 O5, [M + H] + : m / z 797.2476, found 797.2515; HPLC (t R = 3.797 min, 96.27%).

[0102] Compound 12 1 Note: In the 1H-NMR data part, there is a " / / " symbol added in the translation for better line break alignment. It has no impact on the actual content and is for formatting purposes only.1H-NMR(400 MHz, DMSO-d6), δ (ppm): 11.09 (1H, s), 8.63 (1H, s), 8.15 (1H, s), 7.90 (1H, s), 7.80 (1H, s), 7.68 (1H, d, J = 2.1 Hz), 7.54 - 7.47 (1H, m), 7.46 (1H, d, J = 2.1 Hz), 7.43 (1H, d, J = 8.3 Hz), 7.14 (1H, d, J = 8.6 Hz), 7.04 - 6.99 (2H, m), 5.05 (1H, dd, J = 12.8, 5.4 Hz), 4.55 - 4.44 (6H, m), 3.86 (2H, s), 3.71 - 3.63 (2H, m), 2.92 - 2.73 (2H, m), 2.60 - 2.55 (2H, m), 2.04 - 1.99 (1H, m), 0.73 - 0.68 (2H, m), 0.53 (2H, s); 13 13C-NMR(150 MHz, DMSO-d6), δ (ppm): 172.2, 169.5, 168.1, 166.6, 162.0, 160.2, 158.6, 154.4, 145.2, 136.1, 135.4, 134.2, 133.6, 132.6, 132.6, 131.5, 128.1, 126.5, 123.2, 122.6, 117.0, 110.2, 109.1, 103.7, 70.8, 67.8, 65.9, 54.3, 48.9, 48.0, 30.4, 23.3, 21.5, 8.9, 5.6. HRMS(ESI) + calculated for C 36 H 32 Cl2N 10 O6, [M + H] + : m / z 771.1956, found 771.1971; HPLC(t R = 3.074 min, 97.47%).

[0103] Compound 13 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 11.09 (1H, s), 8.64 (1H, s), 8.16 (1H, s), 7.96 (1H, s), 7.80 (1H, s), 7.68 (1H, d, J = 2.1 Hz), 7.53 (1H, t, J = 7.8 Hz), 7.48 (1H, dd, J = 8.3, 2.1 Hz), 7.41 (1H, d, J = 8.3 Hz), 7.15 (1H, d, J = 8.6 Hz), 7.05 - 7.01 (2H, m), 5.04 (1H, dd, J = 12.8, 5.4 Hz), 4.56 - 4.43 (6H, m), 3.74 - 3.72 (4H, m), 3.53 - 3.44 (4H, m), 2.92 - 2.80 (2H, m), 2.59 - 2.54 (2H, m), 2.03 - 1.97 (1H, m), 0.74 - 0.70 (2H, m), 0.57 - 0.53 (2H, m); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 172.2, 169.4, 168.1, 166.6, 162.0, 160.2, 158.6, 154.5, 145.2, 136.1, 135.5, 134.2, 133.6, 132.6, 132.6, 131.5, 129.0, 128.1, 126.5, 123.3, 122.7, 117.0, 110.3, 109.0, 68.9, 68.7, 68.2, 66.0, 48.7, 47.9, 30.4, 21.5, 21.5, 13.4, 5.6. HRMS (ESI) + calculated for C 38 H 36 Cl2N 10 O7, [M + H] + : m / z 815.2218, found 815.2240; HPLC (t R = 2.964 min, 96.15%).

[0104] Compound 14 1H-NMR(400MHz,DMSO-d6),δ(ppm):11.09(1H,s),8.64(1H,s),8.17(1H,s),7.98(1H,s),7.80(1H,s) ,7.68(1H,d,J=2.0Hz),7.54(1H,t,J=7.9Hz),7.47(1H,dd,J=8.3,2.0Hz),7.41(1H,d,J=8.3Hz),7.1 5(1H,d,J=8.5Hz),7.05-7.02(2H,m),5.05(1H,dd,J=12.8,5.2Hz),4.57-4.45(6H,m),3.76-3.39(12 H,m),2.92-2.83(2H,m),2.60-2.56(2H,m),2.03-2.01(1H,m),0.73-0.72(2H,m),0.57-0.53(2H,m); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 172.2, 169.4, 168.1, 166.7, 162.0, 160. 2,158.6,154.5,145.2,136.1,135.5,134.2,133.6,132.6,132.6,131.5,12 8.1,126.5,125.1,123.3,122.7,117.0,110.3,109.0,70.8,69.1,69.0,68. 9,68.1,66.0,59.2,54.3,48.7,48.0,30.4,27.6,21.5,8.9,5.6.HRMS(ESI) + calculated for C 40 H 40 Cl2N 10 O8,[M+H] + :m / z 859.2480, found 859.2532; HPLC(t R =2.956min,96.03%).

[0105] Synthesis of Compounds 15-35

[0106]

[0107] Synthesis route and conditions: (i) HATU, DIEA; (ii) Sodium L-Ascorbate, CuSO4, TEA, NBA.

[0108] Synthesis of intermediates 18a-8c

[0109] To a solution of intermediate 16 (454 mg, 1.02 mmol), alkynyl compound 17 and DIEA (178 μL, 1.02 mmol) and HATU (388 mg, 1.02 mmol) in DMF (5 ml) was added. The reaction mixture was stirred at room temperature for 12 hours. The residue was poured into water (30 ml) and extracted with ethyl acetate (3×30 ml). The organic layer was washed with saturated hydrated sodium carbonate solution and brine, then dried over anhydrous sodium sulfate. The residue was concentrated under reduced pressure and purified to a white solid by silica gel SEL flash chromatography (dichloromethane / methanol=20:1) in a yield of 65%-69%.

[0110] Intermediate 18a 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 9.0 (1H, s), 8.37 (1H, d, J = 7.81Hz), 7.93 (1H, d, J = 9.3 Hz),7.44-7.37(4H,m),5.10(1H,d,J=3.5Hz),4.96-4.88(1H,m),4.53(1H,d,J=9.3Hz),4 .42(1H,d,J=8.1Hz),4.28(1H,s),3.64-3.56(2H,m),2.74-2.73(1H,m),2.45(3H,s),2.3 7-2.31(4H,m),2.04-1.99(1H,m),1.82-1.76(1H,m),1.37(1H,d,J=7.0Hz),0.94(9H,s); 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 170.0, 169.6, 168.8, 150.9, 147.1, 144.1, 130.5, 129.1, 128.3, 128.2 ,125.8,125.7,83.3,70.7,68.2,58.0,55.9,55.7,47.1,37.1,34.7,33.2,25.9,25.8,21.8,15.4,13.7.

[0111] Intermediate 18b 11H-NMR (400 MHz, DMSO-d6), δ (ppm): 8.99 (1H, s), 8.37 (1H, d, J = 7.7 Hz), 7.86 (1H, d, J = 9.2 Hz), 7.43 (2H, d, J = 8.3 Hz), 7.38 (2H, d, J = 8.3 Hz), 5.10 (1H, d, J = 3.5 Hz), 4.96 - 4.88 (1H, m), 4.51 (1H, d, J = 9.2 Hz), 4.42 (1H, d, J = 8.1 Hz), 4.28 (1H, s), 3.63 - 3.58 (2H, m), 2.77 (1H, d, J = 2.6 Hz), 2.45 (3H, s), 2.34 - 2.23 (2H, m), 2.16 - 2.12 (2H, m), 2.04 - 1.98 (1H, m), 1.82 - 1.76 (1H, m), 1.71 - 1.61 (1H, m), 1.37 (1H, d, J = 7.0 Hz), 0.94 (9H, s); 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 171.9, 171.1, 170.0, 162.8, 152.0, 148.2, 145.1, 131.6, 130.2, 129.3, 129.3, 126.8, 84.6, 71.9, 69.2, 59.0, 56.9, 56.7, 54.1, 48.2, 35.7, 34.3, 26.9, 25.0, 22.9, 17.9, 17.2, 16.5, 13.0.

[0112] Intermediate 18c 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 8.98 (1H, s), 8.37 (1H, d, J = 7.8 Hz), 7.81 (1H, d, J = 9.3 Hz), 7.43 (2H, d, J = 8.3 Hz), 7.38 (2H, d, J = 8.3 Hz), 5.10 (1H, d, J = 3.5 Hz), 4.95 - 4.88 (1H, m), 4.52 (1H, d, J = 9.3 Hz), 4.43 (1H, d, J = 8.0 Hz), 4.28 (x1H, s), 3.64 - 3.58 (2H, m), 2.74 - 2.73 (1H, m), 2.45 (3H, s), 2.30 - 2.21 (4H, m), 2.04 - 1.98 (1H, m), 1.82 - 1.76 (1H, m), 1.61 - 1.51 (2H, m), 1.45 - 1.37 (5H, m), 0.94 (9H, s); 13C-NMR (150MHz, DMSO-d6), δ (ppm): 172.3, 171.1, 170.1, 162.8, 152.0, 148.2, 145.1, 131.6, 130.2, 129.3, 129.3, 126.8,84.9,71.7,69.2,59.0,56.8,56.7,54.1,48.2,36.3,35.6,34.7,28.0,27.0,26.9,25.0,22.9,17.9,16.5.

[0113] Preparation of Compound 15-35

[0114] To a solution of n-butanol (5 ml) containing intermediate 14 (0.24 mmol), intermediate 18 (0.24 mmol), sodium L-ascorbate (0.2 mmol), and TEA (1.2 mmol) was added CuSO (0.20 mmol). The reaction mixture was stirred at room temperature for 12 hours. The residue was concentrated under reduced pressure and purified by silica gel SEL flash chromatography (dichloromethane / methanol = 20:1) to a yellow solid in a yield of 30-41%.

[0115] Compound 15 1 H-NMR (400MHz, DMSO-d6), δ (ppm): 8.98 (1H, s), 8.63 (1H, s), 8.36 (1H, d, J = 7.8Hz), 8.17 (1H, s), 7.88 (1H, d, J = 9.3Hz), 7.81 (1H,s),7.69(1H,d,J=2.1Hz),7.64(1H,s),7.48(1H,dd,J=8.3,2.1Hz),7.44-7.36(5H,m),5.11(1H,d,J=3.6Hz),4.95-4.8 8(1H,m),4.52-4.49(2H,m),4.44-4.40(3H,m),4.28-4.27(1H,m),3.85-3.60(7H,m),2.83-2.75(3H,m),2.60-2.54(2H,m), 2.45(3H,m),2.04-1.99(1H,m),1.82-1.76(1H,m),1.37(3H,d,J=7.0Hz),0.89(9H,s),0.74-0.69(2H,m),0.57-0.53(2H,m); 1313C-NMR(150 MHz, DMSO-d6), δ (ppm): 170.4, 170.0, 169.4, 168.9, 162.0, 160.3, 158.7, 154.5, 150.9, 147.2, 145.3, 144.1, 136.1, 134.2, 133.7, 132.6, 132.6, 130.5, 129.1, 129.0, 128.2, 128.1, 126.6, 125.8, 123.2, 121.5, 103.7, 70.8, 68.2, 67.8, 65.9, 57.9, 55.9, 55.7, 54.3, 48.7, 47.1, 34.5, 33.8, 27.6, 25.8, 21.8, 20.9, 20.4, 18.7, 15.4, 8.9, 5.6. HRMS(ESI) + calculated for C 48 H 55 Cl2N 11 O6S, [M + H] + : m / z 984.3507, found 984.3536; HPLC(t R = 4.035 min, 99.37%).

[0116] Compound 16 1 1H-NMR(400 MHz, DMSO-d6), δ (ppm): 8.98(1H, s), 8.64(1H, s), 8.37(1H, d, J = 7.8 Hz), 8.17(1H, s), 7.85 - 7.81(2H, m), 7.69(1H, d, J = 2.1 Hz), 7.66(1H, s), 7.48(1H, dd, J = 8.3, 2.1 Hz), 7.44 - 7.41(3H, m), 7.38(2H, d, J = 8.3 Hz), 5.10(1H, d, J = 3.5 Hz), 4.95 - 4.87(1H, m), 4.53 - 4.51(2H, m), 4.45 - 4.41(3H, m), 4.28(1H, m), 3.86 - 3.61(7H, m), 2.79(1H, s), 2.45(3H, s), 2.33 - 2.25(2H, m), 2.22 - 1.12(2H, m), 2.04 - 1.99(1H, m), 1.83 - 1.70(3H, m), 1.37(3H, d, J = 7.0 Hz), 0.93(9H, s), 0.74 - 0.69(2H, m), 0.56 - 0.55(2H, m); 1313C-NMR(150MHz, DMSO-d6), δ(ppm): 171.1, 170.0, 169.4, 167.0, 162.0, 160.3, 158.6, 154.5, 150.9, 147.2, 145.7, 144.1, 136.1, 134.2, 133.7, 132.6, 132.6, 130.5, 129.1, 128.2, 128.1, 126.6, 125.8, 125.6, 123.3, 121.4, 103.7, 70.8, 68.2, 67.8, 66.0, 58.0, 55.8, 55.7, 54.3, 48.7, 47.1, 34.6, 33.9, 25.9, 25.8, 24.9, 24.1, 23.3, 21.8, 15.4, 8.9, 5.6. HRMS(ESI) + Calculated for C 49 H 57 Cl2N 11 O6S, [M + H] + : m / z 998.3664, found 998.3707; HPLC(t R = 4.056 min, 99.14%).

[0117] Compound 17 1 1H-NMR(400MHz, DMSO-d6), δ(ppm): 8.98(1H, s), 8.63(1H, s), 8.36(1H, d, J = 7.8Hz), 8.17(1H, s), 7.81 - 7.78(2H, m), 7.69(1H, d, J = 2.1Hz), 7.64(1H, s), 7.48(1H, dd, J = 8.3, 2.1Hz), 7.44 - 7.41(3H, m), 7.37(2H, d, J = 8.3Hz), 5.10(1H, d, J = 3.4Hz), 4.95 - 4.88(1H, m), 4.50(2H, d, J = 9.3Hz), 4.44 - 4.40(3H, m), 4.28(1H, m), 3.85 - 3.60(7H, m), 2.79(1H, s), 2.45(3H, s), 2.33 - 2.25(2H, m), 2.19 - 2.11(2H, m), 2.03 - 1.99(1H, m), 1.82 - 1.76(1H, m), 1.52 - 1.48(4H, m), 1.37(3H, m), 0.91(9H, s), 0.73 - 0.69(2H, m), 0.55(2H, m); 1313C-NMR(150MHz, DMSO-d6), δ (ppm): 171.3, 170.0, 169.4, 169.0, 162.0, 160.3, 158.7, 154.5, 150.9, 147.2, 146.0, 144.1, 136.1, 134.2, 133.6, 132.6, 132.6, 130.6, 129.1, 128.2, 128.1, 126.6, 125.8, 125.7, 123.2, 121.3, 103.7, 68.2, 67.9, 66.0, 57.9, 55.8, 55.7, 48.7, 47.1, 34.6, 34.0, 28.0, 27.6, 25.9, 25.8, 24.4, 24.1, 23.3, 21.8, 20.4, 18.7, 15.4, 8.9, 5.6. HRMS(ESI) + calculated for C 50 H 59 Cl2N 11 O6S, [M + H] + : m / z 1012.3841, found 1012.3841;HPLC(t R =4.106 min, 99.10%).

[0118] Compound 18 1 1H-NMR(400MHz, DMSO-d6), δ (ppm): 8.98(1H, s), 8.63(1H, s), 8.36(1H, d, J = 7.8Hz), 8.17(1H, s), 7.88(1H, d, J = 9.3Hz), 7.81(1H, s), 7.73(1H, s), 7.69(1H, d, J = 2.1Hz), 7.48(1H, dd, J = 8.3, 2.1Hz), 7.44 - 7.37(5H, m), 5.12(1H, s), 4.95 - 4.88(1H, m), 4.51(2H, d, J = 9.3Hz), 4.45 - 4.38(3H, m), 4.28(1H, s), 3.73(4H, t, J = 5.3Hz), 3.61 - 3.58(5H, m), 3.49 - 3.47(2H, m), 2.86 - 2.78(3H, m), 2.62 - 2.57(2H, m), 2.46(3H, m), 2.05 - 1.99(1H, m), 1.83 - 1.77(1H, m), 1.37(3H, d, J = 7.0Hz), 0.89(9H, s), 0.75 - 0.73(2H, m), 0.58 - 0.54(2H, m); 1313C-NMR(150MHz, DMSO-d6), δ(ppm): 170.5, 170.0, 169.4, 168.9, 162.0, 160.2, 158.6, 154.5, 150.9, 147.2, 145.3, 144.1, 136.1, 134.2, 133.6, 132.6, 132.6, 130.5, 129.1, 128.2, 128.1, 126.5, 125.8, 123.3, 121.6, 103.8, 70.8, 68.9, 68.8, 68.2, 68.2, 66.0, 57.9, 55.9, 55.7, 48.6, 47.1, 34.5, 33.9, 27.6, 25.8, 21.8, 20.9, 20.4, 18.7, 15.4, 8.9, 5.6. HRMS(ESI) + Calculated for C 50 H 59 Cl2N 11 O7S, [M + H] + : m / z 1028.3769, found 1028.3820; HPLC(t R = 4.040 min, 95.70%).

[0119] Compound 19 1 1H-NMR(400MHz, DMSO-d6), δ(ppm): 9.00(1H, s), 8.64(1H, s), 8.37(1H, d, J = 7.8Hz), 8.18(1H, s), 7.85 - 7.78(3H, m), 7.69(1H, d, J = 2.1Hz), 7.48(1H, dd, J = 8.3, 2.1Hz), 7.44 - 7.41(3H, m), 7.38(2H, d, J = 8.3Hz), 5.10(1H, d, J = 3.5Hz), 4.95 - 4.88(1H, m), 4.52(2H, d, J = 9.3Hz), 4.45 - 4.40(3H, m), 4.28(1H, m), 3.76 - 3.73(4H, m), 3.61 - 3.48(7H, m), 2.82(1H, s), 2.46(3H, s), 2.34 - 2.25(2H, m), 2.21 - 2.21(2H, m), 2.04 - 1.99(1H, m), 1.83 - 1.74(3H, m), 1.37(3H, d, J = 7.0Hz), 0.93(9H, s), 0.77 - 0.72(2H, m), 0.58 - 0.54(2H, m); 13C-NMR(150MHz, DMSO-d6), δ(ppm): 171.1, 170.0, 169.0, 162.1, 160.2, 158.6, 154.5, 151.0, 147.3, 144.2, 144.1, 136.1, 134.2, 133.6, 132.6, 132.6, 129.1, 128.1, 126.5, 125.8, 125.6, 123.3, 121.6, 103.8, 70.8, 68.9, 68.8, 68.2, 66.0, 57.9, 55.8, 55.7, 54.3, 48.6, 47.1, 34.6, 33.9, 28.4, 25.9, 25.8, 24.9, 24.1, 23.3, 21.8, 15.4, 13.4, 8.9, 5.6. HRMS(ESI) + calculated for C 51 H 61 Cl2N 11 O7S, [M + H] + : m / z 1042.3926, found 1042.3989;HPLC(t R =4.611 min, 99.71%).

[0120] Compound 20 1 H-NMR(400MHz, DMSO-d6), δ(ppm): 8.98(1H, s), 8.63(1H, s), 8.37(1H, d, J = 7.8Hz), 8.17(1H, s), 7.80 - 7.78(2H, m), 7.73(1H, s), 7.69(1H, d, J = 2.1Hz), 7.48(1H, dd, J = 8.3, 2.1Hz), 7.44 - 7.40(3H, m), 7.37(2H, d, J = 8.3Hz), 5.10(1H, s), 4.95 - 4.88(1H, m), 4.51(2H, d, J = 9.3Hz), 4.44 - 4.39(3H, m), 4.27(1H, m), 3.74 - 3.72(4H, m), 3.60 - 3.46(7H, m), 2.81(1H, s), 2.45(3H, s), 2.33 - 2.23(2H, m), 2.20 - 2.11(2H, m), 2.03 - 2.00(1H, m), 1.82 - 1.76(1H, m), 1.53 - 1.47(4H, m), 1.37(3H, d, J = 7.0Hz), 0.92(9H, s), 0.73 - 0.72(2H, m), 0.60 - 0.53(2H, m); 13C-NMR(150MHz,DMSO-d6),δ(ppm):172.4,171.1,170.5,170.1,163.1,161.3,159.7,155.6,152.0,148.2,147.1,145.1,137.2,135.3,134.7,133.7,133.6,131.6,130.2,129.3,129.2,127.6,126.8,126.7,124.4,122.5,104.8,71.9,70.0,69.9,69.3,69.2,67.1,59.0,56.8,56.7,49.6,48.2,35.6,35.1,29.1,27.0,26.9,25.5,25.2,24.4,22.9,21.5,19.8,16.5,io.o,6.7.HRMS(ESI) + calculated for C 52 H 63 Cl2N 11 O7S,[M+H] + :m / z 1056.4082,found 1056.4089;HPLC(t R =4.151min,98.46%).

[0121] Compound 21 1 ​​13C-NMR(150MHz, DMSO-d6), δ(ppm): 170.5, 170.0, 168.9, 162.0, 160.2, 158.6, 154.5, 150.9, 147.2, 145.3, 144.1, 136.1, 134.2, 133.6, 132.6, 132.6, 130.5, 129.1, 129.0, 128.2, 128.1, 126.5, 125.8, 123.3, 121.6, 103.8, 69.1, 69.0, 68.9, 68.2, 66.0, 57.9, 55.9, 55.7, 54.3, 48.6, 47.1, 34.5, 33.9, 25.8, 21.8, 20.9, 20.3, 20.0, 18.8, 15.4, 8.9, 5.6. HRMS(ESI) + calculated for C 52 H 63 Cl2N 11 O8S, [M+H] + : m / z 1071.3959, found 1072.4060;HPLC(t R =3.732 min, 98.80%).

[0122] Compound 22 1 1H-NMR(400MHz, DMSO-d6), δ(ppm): 9.00(1H, s), 8.64(1H, s), 8.37(1H, d, J=7.8 Hz), 8.18(1H, s), 7.85 - 7.80(3H, m), 7.69(1H, d, J=2.1 Hz), 7.48(1H, dd, J=8.3, 2.1 Hz), 7.44 - 7.41(3H, m), 7.38(2H, d, J=8.3 Hz), 5.10(1H, d, J=3.5 Hz), 4.95 - 4.88(1H, m), 4.53(2H, d, J=9.3 Hz), 4.45 - 4.42(3H, m), 4.28(1H, m), 3.78 - 3.75(4H, m), 3.61 - 3.43(11H, m), 2.83(1H, s), 2.46(3H, s), 2.34 - 2.12(4H, m), 2.04 - 2.00(1H, m), 1.83 - 1.76(3H, m), 1.37(3H, d, J=7.0 Hz), 0.93(9H, s), 0.74 - 0.73(2H, m), 0.58 - 0.54(2H, m); 1313C-NMR(150MHz, DMSO-d6), δ (ppm): 171.1, 170.0, 169.4, 169.0, 162.0, 160.2, 158.6, 154.5, 151.0, 144.1, 136.1, 134.2, 133.6, 132.6, 132.6, 129.1, 128.2, 128.1, 126.5, 125.8, 123.3, 121.6, 103.8, 70.8, 69.1, 69.0, 68.9, 68.2, 66.0, 57.9, 55.8, 55.7, 48.6, 47.1, 39.5, 37.1, 34.6, 33.9, 27.6, 25.9, 25.8, 24.9, 24.1, 21.8, 20.4, 18.7, 15.4, 8.9, 5.6. HRMS(ESI) + calculated for C 53 H 65 Cl2N 11 O8S, [M + H] + : m / z 1086.4188, found 1086.4242; HPLC(t R = 3.750 min, 99.58%).

[0123] Compound 23 1 1H-NMR(4**H**-NMR(400MHz, DMSO-d6), δ (ppm): 8.99(1H, s), 8.64(1H, s), 8.37(1H, d, J = 7.8Hz), 8.18(1H, s), 7.81 - 7.76(3H, m), 7.69(1H, d, J = 2.0Hz), 7.48(1H, dd, J = 8.3, 2.0Hz), 7.44 - 7.41(3H, m), 7.38(2H, d, J = 8.3Hz), 5.11(1H, s), 4.96 - 4.89(1H, m), 4.52(2H, d, J = 9.3Hz), 4.44 - 4.41(3H, m), 4.28(1H, m), 3.77 - 3.75(4H, m), 3.64 - 3.43(11H, m), 2.83(1H, s), 2.46(3H, s), 2.34 - 2.20(4H, m), 2.04 - 2.00(1H, m), 1.83 - 1.76(1H, m), 1.55 - 1.47(4H, m), 1.38(3H, d, J = 7.0Hz), 0.93(9H, s), 0.75 - 0.73(2H, m), 0.61 - 0.54(2H, m); 1313C-NMR(150MHz, DMSO-d6), δ (ppm): 171.3, 170.0, 169.4, 169.0, 162.0, 160.2, 158.6, 154.5, 150.9, 147.2, 146.0, 144.1, 136.1, 134.2, 133.6, 132.6, 132.6, 129.1, 128.2, 128.1, 126.5, 125.8, 125.7, 123.3, 121.5, 103.8, 70.8, 69.1, 69.0, 68.9, 68.2, 68.2, 66.0, 57.9, 55.8, 55.7, 48.6, 47.1, 45.1, 39.5, 37.1, 34.6, 34.0, 28.0, 27.6, 25.8, 24.4, 24.2, 21.8, 20.4, 18.7, 15.4, 8.9, 5.6. HRMS(ESI) + calculated for C<000023o>H 67 Cl2N 11 O8S, [M + H] + : m / z 1100.4345, found 1100.4365; HPLC(t R = 4.018 min, 98.83%). Compound 24 1 1H-NMR(400MHz, DMSO-d6), δ (ppm): 8.99(1H, s), 8.64(1H, s), 8.36(1H, d, J = 7.8Hz), 8.16(1H, s), 7.88(1H, d, J = 9.3Hz), 7.81(1H, s), 7.75(1H, s), 7.69(1H, d, J = 2.0Hz), 7.48(1H, dd, J = 8.3, 2.0Hz), 7.43(3H, dd, J = 8.3, 2.6Hz), 7.37(2H, d, J = 8.3Hz), 5.12(1H, d, J = 3.5Hz), 4.95 - 4.87(1H, m), 4.51(1H, d, J = 9.3Hz), 4.43(1H, d, J = 8.1Hz), 4.36 - 4.28(5H, m), 3.61(2H, m), 2.86 - 2.75(3H, m), 2.62 - 2.57(2H, m), 2.46(3H, s), 2.05 - 1.99(1H, m), 1.82 - 1.64(5H, m), 1.36(3H, d, J = 7.0Hz), 0.87(9H, s), 0.75 - 0.70(2H, m), 0.54(2H, m); 13 It should be noted that there seems to be a small error in the original text where "calculated for C 54 " is likely a misspelling and should probably be "calculated for C 54 ". This has been retained as is in the translation for the purpose of following the instruction to preserve all tags exactly.13C-NMR(150 MHz, DMSO-d6), δ (ppm): 170.5, 170.0, 169.4, 168.9, 162.1, 160.2, 158.7, 154.3, 151.0, 147.3, 145.4, 144.1, 136.0, 134.3, 133.7, 132.6, 132.6, 129.1, 128.2, 128.1, 126.6, 125.8, 125.6, 121.3, 103.8, 70.8, 68.2, 57.9, 55.9, 55.9, 48.3, 47.1, 34. 5, 33.8, 28.4, 27.6, 26.8, 25.7, 23.8, 21.8, 21.5, 20.9, 20.4, 18.7, 15.4, 13.4, 8.9, 5.6. HRMS(ESI) + calculated for C 48 H 55 Cl2N 11 O5S, [M+H] + : m / z 968.3558, found 968.3604; HPLC(t R = 4.267 min, 95.13%).

[0124] Compound 25 1 1H-NMR(400 MHz, DMSO-d6), δ (ppm): 8.99 (1H, s), 8.63 (1H, s), 8.38 (1H, d, J = 7.8 Hz), 8.15 (1H, s), 7.86 - 7.80 (3H, m), 7.69 (1H, d, J = 2.1 Hz), 7.48 (1H, dd, J = 8.3, 2.1 Hz), 7.44 - 7.41 (3H, m), 7.38 (2H, d, J = 8.3 Hz), 5.11 (1H, s), 4.96 - 4.86 (1H, m), 4.53 (1H, d, J = 9.3 Hz), 4.45 - 4.41 (1H, m), 4.37 - 4.34 (4H, m), 4.29 (1H, s), 3.62 (2H, m), 2.76 (1H, s), 2.46 (3H, s), 2.34 - 2.12 (4H, m), 2.04 - 1.99 (1H, m), 1.86 - 1.70 (7H, m), 1.38 (3H, d, J = 7.0 Hz), 0.94 (9H, s), 0.75 - 0.70 (2H, m), 0.53 (2H, m); 1313C-NMR (150 MHz, DMSO-d6), δ (ppm): 171.1, 170.0, 169.4, 169.0, 162.0, 160.2, 158.6, 154.3, 150.9, 147.2, 145.9, 144.1, 135.9, 134.3, 133.7, 132.6, 132.6, 130.6, 129.1, 128.2, 128.1, 126.6, 125.8, 125.6, 123.4, 121.2, 103.8, 70.8, 68.2, 58.0, 55.8, 55.7, 54.3, 48.3, 47.1, 34.6, 33.8, 28.4, 25.9, 25.8, 24.9, 24.1, 23.7, 23.3, 21.8, 15.4, 13.4, 8.9, 5.6. HRMS (ESI) + calculated for C 49 H 57 Cl2N 11 O5S, [M + H] + : m / z 982.3715, found 982.3739; HPLC (t R = 4.314 min, 99.04%).

[0125] Compound 26 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 9.00 (1H, s), 8.64 (1H, s), 8.37 (1H, d, J = 7.7 Hz), 8.16 (1H, s), 7.82 - 7.79 (3H, m), ⑦.69 (1H, d, J = 2.1 Hz), 7.49 (1H, dd, J = 8.3, 2.1 Hz), 7.44 - 7.41 (3H, m), 7.38 (2H, d, J = 8.3 Hz), 5.11 (1H, d, J = 3.6 Hz), 4.96 - 4.88 (1H, m), 4.51 (1H, d, J = 9.3 Hz), 4.45 - 4.34 (5H, m), 4.29 (1H, m), 3.61 (2H, m), 2.83 (1H, s), 2.46 (3H, s), 2.34 - 2.13 (4H, m), 2.04 - 2.00 (1H, m), 1.85 - 1.76 (3H, m), 1.69 (2H, m), 1.54 - 1.49 (4H, m), 1.38 (3H, d, J = 7.0 Hz), 0.92 (9H, s), 0.75 - 0.70 (2H, m), 0.53 (2H, m); 1313C-NMR(150MHz, DMSO-d6), δ (ppm): 171.3, 170.0, 169.0, 162.0, 160.2, 158.6, 154.3, 150.9, 147.2, 146.1, 144.1, 135.9, 134.3, 133.7, 132.6, 132.6, 130.6, 129.1, 128.2, 128.1, 126.6, 125.8, 125.7, 123.4, 121.0, 103.8, 70.8, 68.2, 57.9, 55.8, 55.7, 54.3, 48.3, 47.1, 34.6, 34.0, 28.0, 26.9, 25.9, 25.8, 24.4, 24.2, 23.8, 23.3, 21.8, 15.4, 13.4, 8.9, 5.6. HRMS(ESI) + calculated for C 50 H 59 Cl2N 11 O5S, [M + H] + : m / z 996.3871, found 996.3887;HPLC(t R =4.380 min, 98.02%).

[0126] Compound 27 1 [[ID=十七]]1H-NMR(400MHz, DMSO-d6), δ (ppm): 9.01(1H, s), 8.64(1H, s), 8.36(1H, d, J = 7.8Hz), 13 8.15(1H, s), 7.89(1H, d, J = 9.3Hz), 7.80(1H, s), 7.75(1H, s), 7.68(1H, d, J = 2.1Hz), 7.48(1H, dd, J = 8.3, 2.1Hz), 7.44 - 7.41(3H, m), 7.37(2H, d, J = 8.3Hz), 5.11(1H, d, J = 3.5Hz), 4.95 - 4.88(1H, m), 4.51(1H, d, J = 9.3Hz), 4.42(1H, d, J = 8.1Hz), 4.29 - 4.26(5H, m), 3.61(2H, m), 2.87 - 2.76(3H, m), 2.60(2H, m), 2.45(3H, s), 2.04 - 2.01(1H, m), 1.86 - 1.73(5H, m), 1.38 - 1.33(5H, m), 0.89(9H, s), 0.72 - 0.68(2H, m), 0.54(2H, m); 1313C-NMR(150MHz, DMSO-d6), δ (ppm): 170.5, 170.0, 169.4, 168.9, 162.1, 160.1, 158.6, 154.3, 151.0, 147.3, 144.1, 135.9, 134.3, 133.7, 132.6, 132.5, 129.1, 128.2, 128.1, 126.5, 125.8, 125.6, 123.4, 121.2, 103.8, 70.8, 68.2, 66.4, 57.9, 55.9, 55.7, 48.4, 47.1, 34.5, 33.9, 28.4, 27.6, 26.0, 25.8, 21.8, 20.9, 20.4, 18.7, 15.4, 13.4, 8.9, 5.6. HRMS(ESI) + calculated for C 49 H 57 Cl2N 11 O5S, [M + H] + : m / z 982.3715, found 982.3754;HPLC(t R = 4.482 min, 95.30%).

[0127] Compound 28 1 1H-NMR(400MHz, DMSO-d6), δ (ppm): 9.04(1H, s), 8.65(1H, s), 8.38(1H, d, J = 7.8 Hz), 8.15(1H, s), 7.86 - 7.79(3H, m), 7.69(1H, d, J = 2.0 Hz), 7.49(1H, dd, J = 8.3, 2.0 Hz), 7.45 - 7.41(3H, m), 7.38(2H, d, J = 8.3 Hz), 5.11(1H, d, J = 5.3 Hz), 4.96 - 4.88(1H, m), 4.53(1H, d, J = 9.3 Hz), 4.43(1H, t, J = 8.0 Hz), 4.31 - 4.27(5H, m), 3.63 - 3.62(2H, m), 2.76(1H, s), 2.46(3H, s), 2.34 - 2.16(4H, m), 2.04 - 1.99(1H, m), 1.89 - 1.74(7H, m), 1.38(3H, d, J = 7.0 Hz), 1.34 - 1.30(2H, m), 0.94(9H, s), 0.70 - (2H, m), (2H, m); 1313C-NMR(150MHz, DMSO-d6), δ(ppm): 172.2, 171.1, 170.1, 163.2, 161.2, 159.7, 155.4, 152.2, 148.5, 145.1, 136.9, 135.4, 134.7, 133.7, 133.6, 130.2, 130.1, 129.8, 129.3, 129.2, 129.1, 127.6, 126.9, 124.5, 122.2, 104.9, 73.7, 69.2, 59.0, 56.9, 56.7, 55.4, 49.5, 48.2, 35.6, 34.9, 29.5, 27.1, 27.0, 26.9, 25.9, 25.2, 24.4, 23.8, 22.9, 16.5, 14.4, 9.1, 6.7. HRMS(ESI) + calculated for C 50 H 59 Cl2N 11 O5S, [M + H] + : m / z 996.3871, found 996.3892; HPLC(t R = 4.524 min, 95.52%).

[0128] Compound 29 1 1H-NMR(400MHz, DMSO-d6), δ(ppm): 9.02(1H, s), 8.64(1H, s), 8.37(1H, d, J = 7.8Hz), 8.15(1H, s), 7.82 - 7.79(3H, m), 7.69(1H, d, J = 2.0Hz), 7.49(1H, dd, J = 8.3, 2.0Hz), 7.44 - 7.41(3H, m), 7.37(2H, d, J = 8.3Hz), 5.11(1H, d, J = 3.6Hz), 4.96 - 4.89(1H, m), 4.52(1H, d, J = 9.3Hz), 4.43(1H, t, J = 8.0Hz), 4.31 - 4.27(5H, m), 3.61(2H, m), 2.76(1H, s), 2.46(3H, s), 2.34 - 2.12(4H, m), 2.04 - 2.00(1H, m), 1.90 - 1.76(5H, m), 1.55 - 1.49(4H, m), 1.38(3H, d, J = 7.0Hz), 1.34 - 1.26(2H, m), 0.93(9H, s), 0.70 - 0.79(2H, m), 0.55(2H, m); 1313C-NMR (150 MHz, DMSO-d6), δ (ppm): 171.3, 170.0, 169.4, 169.0, 162.1, 160.1, 158.6, 154.3, 151.0, 147.3, 146.2, 144.1, 135.9, 134.3, 133.7, 132.6, 132.5, 129.1, 129.1, 128.2, 128.1, 126.5, 125.8, 125.6, 123.4, 121.0, 103.8, 70.8, 68.2, 58.0, 55.8, 55.7, 54.3, 48.4, 47.1, 34.6, 34.0, 28.9, 28.0, 27.6, 26.0, 25.8, 24.4, 24.2, 23.3, 22.8, 21.8, 15.4, 13.4, 8.9, 5.6. HRMS (ESI) + calculated for C 51 H 61 Cl2N 11 O5S, [M + H] + : m / z 1010.4028, found 1010.4056; HPLC (t R = 4.605 min, 97.56%).

[0129] Compound 30 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 9.01 (1H, s), 8.64 (1H, s), 8.36 (1H, d, J = 7.8 Hz), 8.15 (1H, s), 7.89 (1H, d, J = 9.3 Hz), 7.80 (1H, s), 7.78 (1H, s), 7.68 (1H, d, J = 2.1 Hz), 7.48 (1H, dd, J = 8.3, 2.1 Hz), 7.44 - 7.40 (3H, m), 7.37 (2H, d, J = 8.3 Hz), 5.11 (1H, d, J = 3.32 Hz), 4.96 - 4.88 (1H, m), 4.51 (1H, d, J = 9.3 Hz), 4.42 (1H, d, J = 8.1 Hz), 4.28 - 4.23 (5H, m), 3.61 (2H, m), 2.87 - 2.78 (3H, m), 2.63 - 2.59 (2H, m), 2.45 (3H, s), 2.04 - 1.99 (1H, m), 1.82 - 1.68 (5H, m), 1.38 - 1.29 (7H, m), 0.87 (9H, s), 0.74 - 0.72 (2H, m), 0.55 (2H, m); 1313C-NMR(150MHz, DMSO-d6), δ (ppm): 170.5, 170.0, 169.4, 168.9, 162.1, 160.1, 158.6, 154.3, 151.0, 144.1, 135.8, 134.3, 133.7, 132.6, 132.5, 129.1, 128.2, 128.1, 126.5, 125.8, 125.6, 123.4, 121.2, 103.8, 70.8, 68.2, 57.9, 55.9, 55.7, 54.3, 48.5, 47.1, 34.5, 33.8, 29.1, 26.4, 25.8, 25.4, 25.0, 23.3, 21.8, 20.9, 15.4, 13.4, 8.9, 5.7. HRMS(ESI) + calculated for C 50 H 59 Cl2N 11 O5S, [M + H] + : m / z �96.3871, found 996.3881; HPLC(t R = 4.748 min, 96.04%).

[0130] Compound ⒘ 1 1H-NMR(400MHz, DMSO-d6), δ (ppm): 9.04(1H, s), 8.65(1H, s), 8.38(1H, d, J = 7.8 Hz), 8.15(1H, s), 7.86 - 7.79(3H, m), 7.69(1H, d, J = 2.0 Hz), 7.49(1H, dd, J = 8.3, 2.0 Hz), 7.45 - 7.41(3H, m), {7.38(2H, d, J = 8.3 Hz), 5.11(1H, d, J = z.3 Hz), 4.% - 4.88(1H, m), 4.53(1H, d, J = 9.3 Hz), 4.43(1H, t, J = 8.0 Hz), 4.31 - 4.27(5H, m), 3.63 - 3.62(2H, m), 2.76(1H, s), 2.46(3H, s), 2.34 - 2.16(4H, m), 2.04 - 1.99(1H, m), 1.89 - 1.74(7H, m), 1.38(3H, d, J = 7.0 Hz), 1.34 - 1.30(2H, m), 0.94(9H, s), 0.70 - 0.69(2H, m), 0.55(2H, m); 13 It should be noted that there may be some inaccuracies in the original text (such as some unclear or potentially incorrect chemical shift values and notations). This translation is based on the best understanding of the original content.13C-NMR (150 MHz, DMSO-d6), δ (ppm): 172.2, 171.1, 170.1, 163.2, 161.2, 159.7, 155.4, 152.2, 148.5, 145.1, 136.9, 135.4, 134.7, 133.7, 133.6, 130.2, 130.1, 129.8, 129.3, 129.2, 129.1, 127.6, 126.9, 124.5, 122.2, 104.9, 73.7, 69.2, 59.0, 56.9, 56.7, 55.4, 49.5, 48.2, 35.6, 34.9, 29.5, 27.1, 27.0, 26.9, 25.9, 25.2, 24.4, 23.8, 22.9, 16.5, 14.4, 9.1, 6.7. HRMS (ESI) + calculated for C 50 H 59 Cl2N 11 O5S, [M + H][[ID=⑨]] + : m / z 996.3871, found 996.3892; HPLC (t R = 4.524 min, 95.52%).

[0131] Compound 32 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 9.01 (1H, s), 8.64 (1H, s), 8.37 (1H, d, J = 7.8 Hz), 8.15 (1H, s), 7.82 - 7.79 (3H, m), 7.69 (1H, d, J = 2.0 Hz), 7.48 (1H, dd, J = 8.3, 2.0 Hz), 7.45 - 7.41 (3H, m), 7.38 (2H, d, J = 8.3 Hz), 5.11 (1H, d, J = 3.5 Hz), 4.96 - 4.90 (1H, m), 4.51 (1H, d, J = 9.3 Hz), 4.43 (1H, t, J = 8.0 Hz), 4.29 - 4.25 (5H, m), 3.61 (2H, m), 2.78 (1H, s), 2.46 (3H, s), 2.34 - 2.26 (2H, m), 2.04 - 2.00 (2H, m), 2.04 - 2.00 (1H, m), 1.83 - 1.75 (5H, m), 1.53 - 1.49 (4H, m), 1.38 (3H, d, J = 7.1 Hz), 1.30 - 1.25 (4H, m), 0.92 (9H, s), 0.76 - 0.73 (2H, m), 0.56 (2H, m); 13¹³C-NMR(150MHz, DMSO-d6), δ(ppm): 171.3, 170.0, 169.4, 169.0, 162.1, 160.1, 158.6, 154.3, 151.0, 147.3, 146.2, 144.1, 135.8, 134.3, 133.7, 132.6, 132.5, 129.1, 128.2, 126.5, 125.8, 125.6, 123.4, 121.1, 103.8, 70.8, 68.2, 57.9, 55.8, 55.7, 54.3, 48.4, 47.1, 34.6, 34.0, 29.0, 28.0, 26.4, 25.9, 25.8, 25.3, 25.0, 24.4, 24.2, 23.3, 21.8, 15.4, 13.4, 8.9, 5.6. HRMS(ESI) + calculated for C 52 H 63 Cl₂N 11 O₅S, [M+H] + : m / z 1024.4184, found 1024.4233; HPLC(t R = 4.946 min, 97.40%). Compound 33 1 ¹H-NMR(400MHz, DMSO-d6), δ(ppm): 8.98(1H, s), 8.62(1H, s), 8.36(1H, d, J = 7.8 Hz), 8.15(1H, s), 7.88(1H, d, J = 9.2 Hz), 7.8(1H, s), 7.75(1H, s), 7.69(1H, d, J = 2.0 Hz), 7.48(1H, dd, J = 8.3, 2.0 Hz), 7.44 - 7.41(3H, m), 7.37(2H, d, J = 8.3 Hz), 5.11(1H, d, J = 3.4 Hz), 4.95 - 4.88(1H, m), 4.51(1H, d, J = 9.2 Hz), 4.42(1H, d, J = 8.1 Hz), 4.34 - 4.22(5H, m), 3.61 - 3.60(2H, m), 2.89 - 2.73(3H, m), 2.63 - 2.54(2H, m), 2.45(3H, s), 2.04 - 1.99(1H, m), 1.82 - 1.68(5H, m), 1.38 - 1.33(9H, m), 0.88(9H, s), 0.74 - 0.72(2H, m), 0.57 - 0.54(2H, m); 1313C-NMR(150MHz, DMSO-d6), δ(ppm): 170.5, 170.0, 168.9, 162.1, 160.1, 158.6, 154.3, 150.9, 147.2, 145.4, 144.0, 135.8, 134.3, 133.7, 132.6, 132.5, 130.6, 129.1, 128.2, 128.1, 126.6, 125.8, 125.6, 123.4, 121.2, 103.8, 78.6, 68.2, 57.9, 55.9, 55.7, 48.5, 47.1, 34.51, 33.8, 29.1, 28.4, 28.1, 27.6, 26.5, 25.8, 25.3, 23.3, 21.8, 21.5, 20.9, 15.4, 13.4, 5.6. HRMS(ESI) + calculated for C 51 H 61 Cl2N 11 O5S, [M + H] + : m / z 1010.4028, found 1010.4041; HPLC(t R = 4.361 min, 98.93%).

[0132] Compound 34 1 1H-NMR(400MHz, DMSO-d6), δ(ppm): 9.00(1H, s), 8.65(1H, s), 8.38(1H, d, J = 7.8Hz), 8.16(1H, s), 7.86 - 7.83(2H, m), 7.79(1H, s), 7.69(1H, d, J = 2.0Hz), 7.48(1H, dd, J = 8.3, 2.0Hz), 7.45 - 7.41(3H, m), 7.38(2H, d, J = 8.3Hz), 5.11(1H, d, J = 3.5Hz), 4.96 - 4.89(1H, m), 4.53(1H, d, J = 9.3Hz), 4.43(1H, t, J = 8.0Hz), 4.29 - 4.24(5H, m), 3.62 - 3.61(2H, m), 2.80(1H, s), 2.46(3H, s), 2.34 - 2.26(2H, m), 2.23 - 2.12(2H, m), 2.04 - 2.01(1H, m), 1.83 - 1.68(7H, m), 1.38 - 1.26(9H, m), 0.94(9H, s), 0.74 - 0.73(2H, m), 0.58 - 0.54(2H, m); 1313C-NMR (150 MHz, DMSO-d6), δ (ppm): 171.1, 170.0, 169.4, 169.0, 162.1, 160.1, 158.6, 154.3, 150.9, 147.2, 145.9, 144.1, 135.8, 134.3, 133.7, 132.6, 132.5, 130.6, 129.1, 128.2, 128.1, 126.5, 125.8, 125.6, 123.4, 121.1, 103.8, 70.8, 68.2, 58.0, 55.8, 55.7, 54.3, 48.5, 47.1, 34.6, 33.9, 29.0, 27.6, 27.5, 26.5, 25.9, 25.8, 25.3, 24.9, 24.1, 23.3, 21.8, 15.4, 13.4, 8.9, 5.6. HRMS (ESI) + calculated for C 52 H 63 Cl2N 11 O5S, [M+H] + : m / z 1024.4184, found 1024.4210; HPLC (t R = 5.242 min, 98.63%).

[0133] Compound 35 1 1H-NMR (400 MHz, DMSO-d6), δ (ppm): 9.03 (1H, s), 8.64 (1H, s), 8.37 (1H, d, J = 7.8 Hz), 8.16 (1H, s), 7.82 - 7.79 (3H, m), 7.69 (1H, d, J = 2.0 Hz), 7.48 (1H, dd, J = 8.3, 2.0 Hz), 7.44 - (3H, m), 7.38 (2H, d, J = 8.3 Hz), 5.11 (1H, d, J = 3.5 Hz), 4.96 - 4.90 (1H, m), 4.52 (1H, d, J = 9.3 Hz), 4.43 (1H, t, J = 8.0 Hz), 4.28 - 4.22 (5H, m), 3.61 (2H, m), 2.80 (1H, s), 2.46 (3H, s), 2.34 - 2.13 (4H, m), 2.04 - 2.00 (1H, m), 1.83 - 1.68 (5H, m), 1. (4H, m), 1.38 - 1.26 (9H, m), 0.93 (9H, s), 0.75 - 0.69 (2H, m), 0.58 - 0.51 (2H, m); 13C-NMR (150MHz, DMSO-d6), δ (ppm): 171.3, 170.0, 169.4, 169.0, 162.1, 160.1, 158.6, 154.3, 151.0 ,147.3,146.2,144.1,135.8,134.3,133.7,132.6,132.5,129.1,129.0,128.2,128.1,126.5,125 .8,125.6,123.4,121.0,103.8,70.8,68.2,58.0,55.8,55.7,54.3,48.5,47.1,34.6,34.0,29.1, 28.0,27.6,27.5,26.5,25.8,25.8,25.2,24.4,24.2,23.3,21.8,15.4,13.4,8.9,5.6.HRMS(ESI) + calculated for C 53 H 65 Cl2N 11 O5S,[M+H] + :m / z1038.4341, found 1038.4376; HPLC(t R =5.352min,98.76%).

[0134] Test Example 1 Evaluation of PAK1 degradation activity and anti-tumor proliferation activity of compounds 1 to 35

[0135] Cell viability was determined using the 3-(4,5-dimethylthiazol-2-yl)-3,5-diphenyltetrazolium bromide (MTT) assay. MDA-MB-231 cells were seeded into 96-well plates and cultured for 24 hours. Cultures were continued for 24 and 48 hours, and various concentrations of candidate compounds (100 μM, 50 μM, 25 μM, 12.5 μM, 6.25 μM, 3.125 μM, 1.5625 μM, and 0.78125 μM) were added. MTT was added and the cells were cultured for an additional 4 hours. Formazan was then dissolved in DMSO, and cell viability was measured at 490 nm.

[0136] MDA-MB-231 cells were seeded into 10 cm culture dishes. When the cells grew to 70-80%, 2.5 micromolar DMSO and DMSO solution containing each compound were added and cultured for 48 hours before collecting cell and tissue samples. The collected cell and tissue samples were lysed with cell lysis buffer on ice for 45 minutes, and proteins were extracted and quantified after centrifugation. The proteins were separated by SDS-PAGE and electrophoretically transferred through a polyvinylidene fluoride membrane. The PVDF membrane was then sealed for 1 hour and incubated with primary antibodies overnight at 4°C. The next day, secondary antibodies were added and incubated at room temperature for 1.5 hours. After incubation, the membrane was exposed and developed with ECL luminescent solution in a chemiluminescent imager. Quantitative immunoblotting analysis was performed using Image J. The results are as follows:

[0137] Table 1. Evaluation of PAK1 degradation activity and anti-tumor proliferation activity of compounds 1 to 35

[0138]

[0139]

[0140] a Each compound was tested three times and the data are expressed as mean ± SD.

[0141] The experimental results showed that compounds 1-35 showed good PAK1 degradation activity at 2.5 μM, especially compounds 3, 4, 11, 15, 28-30, 33, and 34 showed PAK1 degradation activity at 0.1 μM. In addition, compounds 1-35 showed anti-proliferative activity against MDA-MB-231 breast cancer cells. Among them, compound 33 showed strong anti-proliferative activity, IC 50 The value was 1.27 μM, which was more effective than the potent PAK1 inhibitor FRAX597 in MDA-MB-231 cells. These results demonstrate that compound 1-35 is a potent PAK1 degrader.

[0142] Experimental Example 2 Compound 33 binding to PAK1

[0143] Recombinant PAK1 protein was immobilized on a CM5 chip as a ligand. After coupling, compound 33 was diluted with PBS (DMSO content <0.01%) to five concentrations (1.5625 μM, 3.125 μM, 6.25 μM, 12.5 μM, and 25 μM) and then flowed through the sensor chip at 30 μL / min for 2 minutes. The results were analyzed using a Biacore T200, and the binding constant (KD) value was calculated.

[0144] MDA-MB-231 cells were cultured in 15 cm dishes to 70-80% confluency, then harvested and lysed with dimethyl sulfoxide (DMSO) containing compound 33 (10 μM) or a control group. The lysate was divided into seven portions and incubated at 37°C, 42°C, 47°C, 52°C, 57°C, 62°C, and 67°C for 3 minutes each. After incubation, the lysate was collected for protein quantification (BCA assay). Finally, the samples were boiled at 95°C for 5 minutes and then subjected to Western blot analysis.

[0145] like Figure 1 As shown, there is a concentration-dependent interaction between compound 33 and PAK1 (Kd = 16.2 μM). In addition, compound 33 also enhances the thermal stability of PAK1 in MDA-MB-231 cells, indicating that compound 33 can directly interact with PAK1 in MDA-MB-231 cells. In view of this, compound 33 can directly bind to PAK1 in vitro and inhibit its activity.

[0146] Test Example 3 Compound 33 inhibits MDA-MB-231 cell proliferation

[0147] Next, the anti-proliferative activity of compound 33 was evaluated by a clone formation experiment. MDA-MB-231 cells were seeded in a 6-well plate. When the cells grew to 70-80%, DMSO solution (control group) and DMSO solution containing compound 33 (2.5, 5, 10 micromolar) were added and cultured. When obvious clones appeared, the cells were fixed with 4% paraformaldehyde, stained with crystal violet, and after washing off the stain, photos of the number of cell clones were taken and the clone formation rate was calculated. The results showed that as the concentration of compound 33 increased, the clone formation ability of the cells gradually decreased ( Figure 2 Through 3D tumor sphere experiments, we found that compound 33 could inhibit the formation of MDA-MB-231 cell tumor spheres in a time-dependent manner, further demonstrating that compound 33 could significantly inhibit the proliferation of MDA-MB-231 cells. Subsequently, we detected the expression of Ki-67, a tumor proliferation marker in MDA-MB-231 cells, by immunofluorescence. The results showed that compared with the control group, the nuclear fluorescence intensity and the fluorescence staining range of cells treated with compound 33 were significantly weakened, indicating that the expression of Ki-67 was significantly reduced after treatment with compound 33 ( Figure 2 In addition, we also used EdU proliferation assay to detect the effect of compound 33 on MDA-MB-231 cells. The experimental results showed that compound 33 significantly reduced the green fluorescence intensity of EdU in MDA-MB-231 cells, further confirming that compound 33 significantly inhibited the proliferation of TNBC cells ( Figure 2In conclusion, compound 33 has good anti-TNBC proliferation ability.

[0148] Test Example 4 In vivo antiproliferative activity of compound 33

[0149] Female BALB / c mice (3-4 weeks old, 18-20 g) were provided by SPF (Beijing) Biotechnology Co., Ltd. (Certificate No.: SCXK[Beijing]20190010). Animal husbandry and experimental procedures were in accordance with the Shenzhen University Laboratory Animal Ethics Regulations, and animal experiment standards were established by the Animal Experiment Ethics Committee. Fourteen female BALB / c mice were acclimated and fed for one week before being subcutaneously injected with MDA-MB-231 cells. When tumor volume reached 0.1 cm 3 Mice were randomly divided into two groups: a control group (n=7) and a 20 mg / kg compound 33 treatment group (n=7, administered intraperitoneally once daily). Tumor volume and mouse body weight were measured daily. The solvent for compound 33 was 10% DMSO + 30% PEG300 + 60% saline. During the 12-day compound 33 treatment period, tumor growth was significantly slowed ( Figure 3 After the treatment, the tumor tissue was removed for examination, and the results showed that compound 33 significantly reduced the weight and volume of the tumor, indicating that compound 33 has certain therapeutic potential ( Figure 3 Meanwhile, there was no statistically significant difference in body weight between the mice in the compound 33 treatment group and the control group ( Figure 3 In D), it was shown that compound 33 had no significant effect on the normal growth of mice. In addition, the Ki-67 staining results of tumor tissues in each group showed that compound 33 could significantly reduce the expression of Ki-67 in tumor tissues ( Figure 3 Middle (E), indicating that compound 33 significantly inhibited tumor proliferation in vivo.

Claims

1. A compound represented by formula I or II or a pharmaceutically acceptable salt thereof: in, R 1 is C1-C6 alkyl, C3-C6 cycloalkyl; The Linker in Formula I is: ; n1=2-5, n2=1-3; The Linker in Formula II is: ; n3 = 1-3, n4 = 2-4, n5 = 2-5, n6 = 2-4.

2. The compound of formula I or II according to claim 1 or a pharmaceutically acceptable salt thereof: In the formula I, R 1 is C1-C4 alkyl, C3-C6 cycloalkyl; Linker is: ; n1=2-5, n2=1-3.

3. The compound of formula I or II according to claim 2, or a pharmaceutically acceptable salt thereof: in, n1=4-5, n2=1-3.

4. The compound of formula I or II according to claim 1 or a pharmaceutically acceptable salt thereof: In the formula II, R 1 is C1-C4 alkyl, C3-C6 cycloalkyl; Linker is: ; n3 = 1-3, n4 = 2-4, n5 = 2-5, n6 = 2-4.

5. The compound of formula I or II according to claim 4, or a pharmaceutically acceptable salt thereof: in, n5 =3 or 5, n6=2-4.

6. The compound of formula I or II according to any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof: in, R 1 is methyl or cyclopropyl.

7. A compound having the following structure or a pharmaceutically acceptable salt thereof: 。 8. The compound according to any one of claims 1 to 5 and 7 or a pharmaceutically acceptable salt thereof, characterized in that: The pharmaceutically acceptable salt is nitrate, hydrochloride, sulfate, phosphate or citrate of the compound.

9. The compound according to claim 6 or a pharmaceutically acceptable salt thereof, characterized in that The pharmaceutically acceptable salt is nitrate, hydrochloride, sulfate, phosphate or citrate of the compound.

10. A pharmaceutical composition comprising the compound according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof.

11. A pharmaceutical composition comprising the compound according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof and other anti-tumor drugs.

12. The pharmaceutical composition according to claim 11, characterized in that The compound according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof and other anti-tumor drugs are used by simultaneous, sequential or separate administration.

13. Use of the following compound or a pharmaceutically acceptable salt thereof in the preparation of a PAK1 degrader; 。 14. Use of the compound according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 10 or 11, in the preparation of an anti-tumor drug, wherein the tumor is breast cancer.

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