Erianin PROTACs as well as preparation method and application thereof

Abstract

The invention discloses erianin PROTACs as well as a preparation method and application thereof, the structural formula of the erianin PROTACs is # imgabs0 #, E3 Ligand is pomalidomide, and Linker is an alkane chain or a PEG chain. The erianin PROTACs disclosed by the invention are superior to the prior art in the aspects of target protein degradation efficiency, anti-tumor activity, innovativeness, application prospect and the like, and show remarkable technical progress and practical value.

Description

Technical Field

[0001] The present invention belongs to the technical field of organic synthesis, and specifically relates to Erianin PROTACs and a preparation method and application thereof. Background Art

[0002] As cancer becomes one of the major threats to human health, the research and development of targeted anti-tumor drugs is gaining increasing attention. Traditional small molecule drugs face numerous challenges in clinical application due to issues such as drug resistance and side effects. For example, traditional drugs typically work by directly inhibiting the function of the target protein, but long-term use can easily lead to drug resistance in tumor cells and high toxicity to normal cells, limiting their efficacy and scope of application. In recent years, proteolysis-targeted chimera (PROTAC) technology has emerged as an emerging therapeutic strategy. By utilizing the intracellular ubiquitin-proteasome system to specifically degrade target proteins, it has provided new possibilities for overcoming the limitations of traditional drugs. The PROTAC molecule consists of three parts: an E3 ligase ligand (E3Ligand), a target protein ligand (POILigand), and a linker connecting the two. Its mechanism of action does not rely on direct inhibition of protein function, but rather achieves therapeutic effects by inducing target protein degradation. It has higher selectivity and lower toxicity, and has shown broad application prospects in the anti-tumor field.

[0003] At the same time, research on the use of effective ingredients from traditional Chinese medicine in tumor biotherapy has gradually become a hot topic. Erianin is a low-molecular-weight natural biphenyl compound extracted from Dendrobium officinale, which exhibits multiple pharmacological activities, including anti-tumor, anti-inflammatory, and antibacterial activities. It has low cytotoxicity and good biocompatibility, and is considered a potential anti-tumor drug candidate. Studies have shown that Erianin has inhibitory effects on various cancers (such as gastrointestinal malignancies, lung cancer, and prostate cancer), especially exhibiting significant anti-tumor activity in lung cancer cell lines. Erianin exerts its effects by inducing apoptosis and autophagy, arresting the cell cycle, inhibiting cell migration, and inhibiting tumor angiogenesis. Its anti-tumor activity involves multiple signaling pathways, including JNK, PI3K / AKT, and ERK. However, Erianin still has limitations in clinical application, such as insufficient biological activity, insufficient selectivity, and drug resistance, which restrict its further development and application.

[0004] In the existing technology, although PROTAC technology has made certain progress, the design of PROTAC molecules for natural compounds is still rare, especially the research on Erianin as a target ligand has not been fully carried out. In addition, the existing PROTAC molecules still need to be optimized in terms of target protein degradation efficiency, anti-tumor activity and application range for specific cancers (such as lung cancer). Therefore, it is urgent to develop a PROTAC compound based on Erianin to improve its biological activity, selectivity and anti-tumor effect, while solving the shortcomings of traditional small molecule drugs and Erianin itself, and providing a new technical solution for the development of anti-cancer drugs. Summary of the Invention

[0005] The present invention aims to overcome the shortcomings of the prior art and provide a single-cell temporal transcriptome sequencing method based on RNA dual metabolic labeling.

[0006] The technical solutions of the present invention are as follows:

[0007] A PROTACs of Erianin, having the structural formula Among them, E3Ligand is pomalidomide, Linker is an alkane chain or PEG chain,

[0008] The alkane chain is -CH2-CO-NH-(CH2)n1-(CH2)-NH-, n1 is 1, 2, 3, 4, 5 or 7; or the alkane chain is -CO-(CH2)n2-(CH2)-NH-, n2 is 1, 2, 3, 4, 5, 6 or 7,

[0009] The PEG chain is -CH2-CO-NH-CH2-(CH2-O-CH2) n3 -CH2-NH-, n3 is 1, 2 or 3.

[0010] In a preferred embodiment of the present invention, its structural formula is selected from one of the following:

[0011]

[0012]

[0013] The preparation method of the above-mentioned Erianin PROTACs has the following reaction scheme:

[0014]

[0015]

[0016] Use of the above-mentioned Erianin PROTACs or pharmacologically or physiologically acceptable salts thereof in the preparation of a cancer treatment composition.

[0017] In a preferred embodiment of the present invention, the cancer is lung cancer.

[0018] A cancer treatment composition, the active ingredient of which includes the above-mentioned Erianin PROTACs or a pharmacologically or physiologically acceptable salt thereof.

[0019] In a preferred embodiment of the present invention, the cancer is lung cancer.

[0020] Use of the above-mentioned Erianin PROTACs or pharmacologically or physiologically acceptable salts thereof in the preparation of an anti-inflammatory composition.

[0021] An anti-inflammatory composition, the active ingredient of which includes the above-mentioned Erianin PROTACs or a pharmacologically or physiologically acceptable salt thereof.

[0022] The beneficial effects of the present invention are:

[0023] 1. The Erianin PROTACs of the present invention use Erianin as a target ligand (POI Ligand) and pomalidomide as an E3 ligase ligand to specifically degrade target proteins (such as OTUB1 and KRAS) in tumor cells through the ubiquitin-proteasome pathway. Experiments have shown that its degradation efficiency is concentration-dependent, and compared with traditional small molecule inhibitors, it has higher selectivity and lower toxicity.

[0024] 2. The Erianin PROTACs of the present invention showed significant inhibitory effects on the proliferation of various lung cancer cell lines (such as NCI-H446, NCI-H460, and A549), with IC50 values ​​much lower than those of Erianin itself. For example, compound EC2 had an IC50 of 21.62 nM in NCI-H446 cells, compared to 1118.06 nM for Erianin, significantly improving its anti-tumor activity and demonstrating superior efficacy to existing technologies.

[0025] 3. This invention utilizes Erianin as the POI ligand, pomalidomide as the E3 ligand, and an alkane or PEG chain as the linker to design a series of novel PROTAC molecules. This design not only retains the natural advantages of Erianin but also combines the efficient degradation properties of PROTAC technology, providing new ideas for the application of natural compounds in the anti-tumor field.

[0026] 4. Experiments with the proteasome inhibitor MG132 confirmed that the Erianin PROTACs of the present invention degrade target proteins through the ubiquitin-proteasome pathway. For example, after the addition of MG132, the protein levels of OTUB1 and KRAS increased significantly, demonstrating the scientific and reliable mechanism of action.

[0027] 5. The Erianin PROTACs of the present invention not only perform well in anticancer drugs (such as lung cancer treatment), but can also be used to prepare anti-inflammatory compositions, broadening their potential uses in the medical field and having significant practicality and development value. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 This is one of the experimental result diagrams of Example 4 of the present invention.

[0029] Figure 2 This is the second diagram of the experimental results of Example 4 of the present invention.

[0030] Figure 3 This is one of the experimental result diagrams of Example 5 of the present invention.

[0031] Figure 4 This is the second diagram of the experimental results of Example 5 of the present invention. DETAILED DESCRIPTION

[0032] The technical solution of the present invention is further illustrated and described below through specific implementation methods in conjunction with the accompanying drawings.

[0033] The following Erianin PROTACs prepared in Examples 1 to 2 are listed as follows:

[0034]

[0035]

[0036]

[0037]

[0038] Example 1

[0039] (1) Preparation of intermediates F2-F8 (preparation of E3 ligand)

[0040] A. The structural formula of intermediate f2(n=1) / f3(n=2) / f4(n=3) / f5(n=4) / f6(n=5) / f8(n=7) is The specific synthesis method is as follows:

[0041] Fluorothalidamide (7.19 g, 26.01 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (9.059 mL, 52.01 mmol) and N-tert-butyloxycarbonyl-1,2-ethylenediamine (5 g, 31.207 mmol) were then added and refluxed at 90°C for 4 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain intermediate f2 (yellow-green powder, 41.2% yield).

[0042] Fluorothalidamide (6.61 g, 23.91 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (8.33 mL, 47.82 mmol) and N-tert-butyloxycarbonyl-1,3-propylenediamine (5 g, 28.69 mmol) were then added and refluxed at 90°C for 4 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO3 and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain intermediate f3 (yellow-green powder, 55.4% yield).

[0043] Fluorothalidamide (6.12 g, 22.13 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (7.709 mL, 44.26 mmol) and N-tert-butyloxycarbonyl-1,4-butanediamine (5 g, 26.56 mmol) were then added and refluxed at 90°C for 4 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain intermediate f4 (yellow-green powder, 48.3% yield).

[0044] Fluorothalidamide (5.69 g, 20.59 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (7.175 mL, 41.19 mmol) and N-tert-butyloxycarbonyl-1,5-pentanediamine (5 g, 24.71 mmol) were then added and the mixture was heated to 90°C and refluxed for 4 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain intermediate f5 (yellow-green powder, 47.5% yield).

[0045] Fluorothalidamide (5.32 g, 19.26 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (6.709 mL, 38.52 mmol) and N-tert-butyloxycarbonyl-1,6-hexanediamine (5 g, 23.11 mmol) were then added and refluxed at 90°C for 4 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO3 and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain intermediate f6 (yellow-green powder, 46.5% yield).

[0046] Fluorothalidamide (4.71 g, 17.05 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (5.939 mL, 34.10 mmol) and N-tert-butyloxycarbonyl-1,8-octanediamine (5 g, 20.46 mmol) were then added and refluxed at 90°C for 4 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO3 and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain intermediate f8 (yellow-green powder, 39.1% yield).

[0047] B. The structural formula of intermediate F2(n=1) / F3(n=2) / F4(n=3) / F5(n=4) / F6(n=5) / F8(n=7) is The specific synthesis method is as follows:

[0048] Intermediate f2 (1 g, 2.40 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise under an ice bath. After addition, the mixture was slowly brought to room temperature for reaction. The reaction was monitored by TLC and completed in approximately 12 h. After the reaction ceased, the solvent was removed by concentration under reduced pressure to obtain intermediate F2 (yellow-green powder) in a yield of 97%. The product was used directly in the next step without purification.

[0049] Intermediate f3 (1 g, 2.34 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise under an ice bath. After addition, the mixture was slowly brought to room temperature for reaction. The reaction was monitored by TLC and completed in about 12 h. After the reaction ceased, the solvent was removed by concentration under reduced pressure to obtain intermediate F3 (yellow-green powder) in a yield of 98%. The product was used directly in the next step without purification.

[0050] Intermediate f4 (1 g, 2.27 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise under an ice bath. After addition, the mixture was slowly brought to room temperature for reaction. The reaction was monitored by TLC and completed in about 12 h. After the reaction ceased, the solvent was removed by concentration under reduced pressure to obtain intermediate F4 (yellow-green powder) in a yield of 98%. The product was used directly in the next step without purification.

[0051] Intermediate f5 (1 g, 2.21 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise under an ice bath. After addition, the mixture was slowly brought to room temperature for reaction. The reaction was monitored by TLC and completed in approximately 12 h. After the reaction ceased, the solvent was removed by concentration under reduced pressure to obtain intermediate F5 (yellow-green powder) in a yield of 97%. The product was used directly in the next step without purification.

[0052] Intermediate f6 (1 g, 2.15 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise under an ice bath. After addition, the mixture was slowly brought to room temperature for reaction. The reaction was monitored by TLC and completed in approximately 12 h. After the reaction ceased, the solvent was removed by concentration under reduced pressure to obtain intermediate F6 (yellow-green powder) in a yield of 97%. The product was used directly in the next step without purification.

[0053] Intermediate f8 (1 g, 2.10 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise under an ice bath. After addition, the mixture was slowly brought to room temperature for reaction. The reaction was monitored by TLC and completed in approximately 12 h. After the reaction ceased, the solvent was removed by concentration under reduced pressure to obtain intermediate F8 (yellow-green powder) in a yield of 98%. The product was used directly in the next step without purification.

[0054] (2) Preparation of intermediates C2-C8 (preparation of E3 ligand)

[0055] A. The structural formula of intermediate s2(n=1) / s3(n=2) / s4(n=3) / s5(n=4) / s6(n=5) / s7(n=6) / s8(n=7) is The specific synthesis method is as follows:

[0056] Fluorothalidamide (1.59 g, 5.74 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (1.999 ml, 11.48 mmol) and tert-butyl 3-aminopropionate (1 g, 6.88 mmol) were then added sequentially, and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain S2 (yellow-green powder, 47.8% yield).

[0057] Fluorothalidamide (1.44 cg, 5.23 c mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (1.823 ml, 10.47 mmol) and tert-butyl 4-aminobutyrate (1 g, 6.28 mmol) were then added, and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain S3 (yellow-green powder, 45.3% yield).

[0058] Fluorothalidamide (1.33 g, 4.81 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (1.675 ml, 9.62 mmol) and tert-butyl 5-aminovalerate (1 g, 5.77 mmol) were then added and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain S4 (yellow-green powder, 37.2% yield).

[0059] Fluorothalidamide (1.23 g, 4.45 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (1.551 mL, 8.90 mmol) and tert-butyl 6-aminohexanoate (1 g, 5.34 mmol) were then added and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain S5 (yellow-green powder, 42.8% yield).

[0060] Fluorothalidamide (1.14 g, 4.14 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (1.44 mL, 8.28 mmol) and tert-butyl 7-aminoheptanoate (1 g, 4.96 mmol) were then added and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain S6 (yellow-green powder, 41.5% yield).

[0061] Fluorothalidamide (1.07 g, 3.87 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (1.35 mL, 7.74 mmol) and tert-butyl 8-aminooctanoate (1 g, 4.64 mmol) were then added and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain S7 (yellow-green powder, 39.5% yield).

[0062] Fluorothalidamide (1.00 g, 3.63 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (1.266 mL, 7.27 mol) and tert-butyl 9-aminononanoate (1 g, 4.36 mmol) were then added and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain S8 (yellow-green powder, 37.1% yield).

[0063] B. The structural formula of intermediate C2(n=1) / C3(n=2) / C4(n=3) / C5(n=4) / C6(n=5) / C7(n=6) / C8(n=7) is The specific synthesis method is as follows:

[0064] Intermediate s2 (1 g, 2.49 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate C2 (yellow-green powder) in a 97% yield. This product was used directly in the next reaction without purification.

[0065] Intermediate s3 (1 g, 2.42 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate C3 (yellow-green powder) in a 98% yield. This product was used directly in the next step without purification.

[0066] Intermediate s4 (1 g, 2.35 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate C4 (yellow-green powder) in a 98% yield. This product was used directly in the next reaction without purification.

[0067] Intermediate s5 (1 g, 2.29 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford Intermediate C5 (yellow-green powder) in a 97% yield. This product was used directly in the next reaction without purification.

[0068] Intermediate s6 (1 g, 2.25 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate C6 (yellow-green powder) in a 98% yield. This product was used directly in the next step without purification.

[0069] Intermediate s7 (1 g, 2.23 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate C7 (yellow-green powder) in a 98% yield. This product was used directly in the next reaction without purification.

[0070] Intermediate s8 (1 g, 2.17 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate C8 (yellow-green powder) in a 97% yield. This product was used directly in the next reaction without purification.

[0071] (3) Preparation of intermediates O1-O3 (preparation of E3 ligand)

[0072] A. The structural formula of intermediate p1(n=1) / p2(n=2) / p3(n=3) is The specific synthesis method is as follows:

[0073] Fluorothalidamide (5.63 g, 20.39 mmol) was dissolved in 20 mL of DMF and stirred to dissolve. DIPEA (7.320 mL, 10.47 mmol) and tert-butyl [2-(2-aminoethoxy)ethyl]carbamate (5 g, 24.47 mmol) were then added and the mixture was heated to 90°C and refluxed for 6 h. The mixture was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed sequentially with saturated NaHCO₃ and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain p1 (yellow-green powder, 42.7% yield).

[0074] Fluorothalidamide (4.64 g, 16.78 mmol) was dissolved in 20 mL of dry DMF and stirred until dissolved. DIPEA (5.845 mL, 33.55 mmol) and tert-butyl 2-(2-(2-aminoethoxy)ethoxy)ethylcarbamate (5 g, 20.13 mmol) were then added, and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed with saturated NaHCO3 and saturated brine, respectively, and then dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain p2 (yellow-green powder, 43.6% yield).

[0075] Fluorothalidamide (3.94 g, 14.25 mmol) was dissolved in 5 mL of dry DMF and stirred until dissolved. DIPEA (4.964 mL, 28.5 mmol) and 1,1-dimethylethyl 13-amino-5,8,11-trioxa-2-azatridecanoate (5 g, 17.10 mmol) were then added and the mixture was heated to 90°C and refluxed for 6 h. After completion, the reaction was quenched with ice water and extracted with ethyl acetate. The combined organic phases were washed with saturated NaHCO3 and saturated brine, respectively, and then dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA = 1:1) to obtain p3 (yellow-green powder, 39.2% yield).

[0076] B. The structural formula of the intermediate O1(n=1) / O2(n=2) / O3(n=3) is The specific synthesis method is as follows:

[0077] Intermediate p1 (1 g, 2.17 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate O1 (yellow-green powder, 96.0% yield), which was used directly in the next reaction without purification.

[0078] Intermediate p2 (1 g, 1.98 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 h. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate O2 (yellow-green powder, 91.0% yield), which was used directly in the next step without purification.

[0079] Intermediate p3 (1 g, 1.82 mmol) was dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise in an ice bath. After addition, the mixture was slowly brought to room temperature and monitored by TLC. The reaction was complete in approximately 12 hours. After cessation of the reaction, the solvent was removed by concentration under reduced pressure to afford intermediate O3 (yellow-green powder, 95.0% yield), which was used directly in the next step without purification.

[0080] (4) Preparation of intermediate Erianin-C1

[0081] The structural formula of the intermediate Erianin-C1 is The specific synthesis method is as follows:

[0082] Erianin (100 mg, 314.10 mmol) was stirred and dissolved in 2 mL of DMF. After stirring, K2CO3 (65.10 mg, 470.32 mmol) and tert-butyl bromoacetate (91.87 mg, 470.32 mmol) were added. The reaction mixture was heated to 60°C and stirred under reflux overnight. After the reaction was completed, it was quenched with ice water and the pH was adjusted to 6-7 with 1 M dilute hydrochloric acid solution. The organic layers were combined and washed with saturated sodium bicarbonate and saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the intermediate. The intermediate was then dissolved in 6 mL of DCM and TFA (3 mL) was added dropwise under ice bath. After the addition was complete, the mixture was slowly allowed to react at room temperature. The reaction was monitored by TLC. The reaction was complete in about 12 hours. After the reaction was terminated, the solvent was removed under reduced pressure to obtain Erianin-C1 (white powder, 56.9% yield), which was used directly in the next reaction without purification.

[0083] Example 2

[0084] (1) Preparation of final product EF2-8

[0085] The structural formula of the final product EF2 (n = 1) / EF3 (n = 2) / EF4 (n = 3) / EF5 (n = 4) / EF6 (n = 5) / EF8 (n = 7) is

[0086] The specific synthesis method is as follows:

[0087] Erianin-C1 (60 mg, 0.16 mmol) and DIPEA (76 μL, 0.43 mmol) were dissolved in 2 mL of dry DMF and stirred at room temperature for 5 min. HATU (75.03 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. F2 (75.63 mg, 0.24 mmol) was added and stirred at room temperature. TLC monitoring indicated that the reaction was complete in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined and washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) afforded the product EF2 (yellow powder, 44.2% yield). ESI-MS m / z 674.2588 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.23(s,1H),7.48–7.43(m,2H),7.08(d,J=7.0Hz,1H),6.98(d,J=8.6H z,1H),6.81–6.79(m,2H),6.73–6.70(m,1H),6.36(s,2H),4.90(dd,J=12.3,5.3Hz,1H),4.53(s,2H) ,3.82(s,6H),3.82(s,3H),3.81(s,3H),3.59(q,J=6.2Hz,2H),3.49(t,J=6.3Hz,2H),2.90–2.85(m, 1H), 2.80 (q, J=2.3Hz, 4H), 2.78–2.74 (m, 1H), 2.73–2.66 (m, 1H), 2.09 (ddt, J=10.5, 5.5, 2.5Hz, 1H). 13 C NMR(126MHz,Chloroform-d)δ171.19,169.86,169.46,168.44,167.65,153.23,148.00,147.04,146.81,137.33,136.38,135.00,132.59,12 3.09,116.76,116.27,112.26,112.11,110.49,105.52,69.98,61.01, 56.28,56.22,56.15,49.03,42.30,38.85,38.56,37.40,31.54,22.86.

[0088] Erianin-C1 (60 mg, 0.16 mmol) and DIPEA (76 μL, 0.43 mmol) were dissolved in 2 mL of dry DMF and stirred at room temperature for 5 min. HATU (75.03 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. F3 (78.95 mg, 0.24 mmol) was added and stirred at room temperature. TLC monitoring indicated that the reaction was complete in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined and washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) afforded the product EF3 (yellow powder, 47.3% yield). ESI-MS m / z 688.2744 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.25(s,1H),7.46(dd,J=8.5,7.1Hz,1H),7.19(t,J=6.2Hz,1H),7.08(d,J =7.0Hz,1H),6.85–6.82(m,3H),6.74(d,J=1.6Hz,1H),6.36(s,2H),4.90(dd,J=12.3,5.3Hz,1H),4.55(s ,2H),3.84(s,3H),3.82(d,J=1.9Hz,9H),3.46(q,J=6.6Hz,2H),3.31(t,J=6.8Hz,2H),2.84(d,J=4.5Hz ,1H),2.82–2.79(m,4H),2.79–2.74(m,1H),2.74–2.67(m,1H),2.12–2.06(m,1H),1.87(p,J=6.8Hz,2H). 13 C NMR(126MHz,DMSO-d6)δ171.21,169.48,169.40,168.48,167.70,153.22 ,147.95,147.05,146.77,137.31,136.39,136.27,135.05,132.71,122. 96,116.57,115.99,112.20,111.70,110.36,105.51,69.77,61.01,56.2 7,56.21,56.15,49.01,39.90,38.58,37.44,36.48,31.54,29.42,22.86.

[0089] Erianin-C1 (100 mg, 0.26 mmol) and DIPEA (125.58 μL, 0.72 mmol) were dissolved in dry DMF solution and stirred at room temperature for 5 min. HATU (125.05 mg, 0.33 mmol) was added. After approximately 0.5 h of reaction, F4 (137.23 mg, 0.24 mmol) was added and stirred at room temperature. TLC was used to monitor the reaction, and the reaction was complete in approximately 4 h. Crushed ice was added to the reaction solution to quench the reaction. The solution was extracted three times with ethyl acetate. The organic layers were combined and washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) was performed to obtain the product EF4 (yellow powder, 45.5% yield). ESI-MS m / z 702.2901 [M+Na] + . 1 H NMR (500MHz, DMSO-d6) δ11.10(s,1H),7.95(d,J=4.8Hz,1H),7.54(t,J=7.8Hz,1H),7.03(dd,J=28.8,7.8H z,2H),6.92–6.85(m,2H),6.80(dd,J=8.3,2.0Hz,1H),6.53(t,J=6.0Hz,1H),6.48(s,2H),5.04(dd,J=12. 8,5.4Hz,1H),4.43(s,2H),3.74(s,3H),3.72(s,6H),3.60(s,3H),3.29(q,J=6.4Hz,2H),3.18(q,J=6.3Hz ,2H),2.92–2.83(m,1H),2.74(s,4H),2.61–2.52(m,2H),2.01(dd,J=10.1,4.5Hz,1H),1.56–1.51(m,4H). 13 C NMR(126MHz,DMSO-d6)δ172.82,170.10,167.85,167.31,152.63,147.54, 147.11,146.37,137.26,136.25,135.58,134.14,132.19,121.87,117.17, 115.50,112.29,110.40,109.02,105.57,68.87,59.95,55.74,55.66,54. 92,48.54,41.50,37.90,37.58,36.51,35.79,30.98,26.49,26.11,22.16.

[0090] Erianin-C1 (60 mg, 0.159 mmol) and DIPEA (76 μL, 0.43 mmol) were dissolved in dry DMF and stirred at room temperature for 5 min. HATU (75.034 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. F5 (85.66 mg, 0.24 mmol) was added and stirred at room temperature. TLC followed the reaction, and the reaction was complete in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined and washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) was performed to obtain the product EF5 (yellow powder, 44.2% yield). ESI-MS m / z 716.7880 [M+Na] + . 1 H NMR (500MHz, DMSO-d6) δ11.10(s,1H),7.90(t,J=5.9Hz,1H),7.55(dd,J=8.6,7.1Hz,1H),7.05(d,J=8.6Hz,1H),7.01(d,J=7.1Hz,1H ),6.91–6.86(m,2H),6.81(dd,J=8.2,2.0Hz,1H),6.54–6.50(m,1H),6.49(s,2H),5.04(dd,J=12.8,5.4Hz,1H),4.43(s,2H),3.75(s, 3H),3.72(s,6H),3.60(s,3H),3.25(q,J=6.7Hz,2H),3.15(q,J=6.7Hz,2H),2.91–2.85(m,1H),2.75(d,J=2.5Hz,4H),2.61–2.56(m,1 H),2.56–2.51(m,1H),2.02(ddq,J=10.4,5.4,3.1,2.4Hz,1H),1.56(p,J=7.4Hz,2H),1.51–1.44(m,2H),1.31(tt,J=9.5,6.4Hz,2H). 13C NMR(126MHz,DMSO-d6)δ172.83,170.11,168.96,167.79,167.32,152.65, 147.55,147.13,146.40,137.26,136.28,135.60,134.15,132.20,121.86, 117.14,115.48,112.29,110.41,109.04,105.58,68.86,55.75,55.68,48. 56,41.80,37.61,36.54,30.99,28.81,28.36,23.63,22.16,20.77,14.09.

[0091] Erianin-C1 (60 mg, 0.159 mmol) and DIPEA (76 μL, 0.43 mmol) were dissolved in dry DMF and stirred at room temperature for 5 min. HATU (75.03 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. F6 (89.01 mg, 0.24 mmol) was added and stirred at room temperature. TLC followed the reaction, and the reaction was complete in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined and washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) was performed to obtain the product EF6 (yellow powder, 47.9% yield). ESI-MS m / z 730.8150 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.32(s,1H),7.47(dd,J=8.5,7.1Hz,1H),7.07(d,J=7.0Hz,2H),6.85(d,J=8.5Hz,1H), 6.84–6.79(m,2H),6.74(d,J=1.9Hz,1H),6.36(s,2H),6.21(d,J=7.4Hz,1H),4.93–4.88(m,1H),4.52(s,2H),3.85(s, 3H),3.82(d,J=2.4Hz,9H),3.32(q,J=6.8Hz,2H),3.24(q,J=6.2,5.7Hz,2H),2.90–2.84(m,2H),2.83–2.76(m,5H),2 .76–2.70(m,1H),2.11(ddd,J=12.1,6.1,3.4Hz,1H),1.65(q,J=7.2Hz,2H),1.56(p,J=7.3Hz,2H),1.47–1.32(m,4H). 13C NMR(126MHz,DMSO-d6)δ172.83,170.11,168.97,167.75,167.32,152.65,147 .52,147.13,146.41,137.26,136.28,135.60,134.14,132.20,121.83,117.15 ,115.44,112.27,110.39,109.03,105.58,68.84,59.96,55.75,55.66,48.55,41.77,38.18,37.59,36.53,30.99,29.03,28.98,28.63,26.04,22.16,22.10.

[0092] Erianin-C1 (60 mg, 0.16 mmol) and DIPEA (76 μL, 0.43 mmol) were dissolved in dry DMF and stirred at room temperature for 5 min. HATU (75.03 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. F8 (95.71 mg, 0.24 mmol) was added and stirred at room temperature. TLC followed the reaction, and the reaction was complete in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined and washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) was performed to obtain the product EF8 (yellow powder, 39.6% yield). ESI-MS m / z 758.8690 [M+Na] + . 1H NMR(500MHz,Chloroform-d)δ8.17(s,1H),7.48(t,J=7.9Hz,1H),7.08(d,J=7.2Hz,1H),7.04(t,J=5.8Hz,1H),6.87(d,J= 8.5Hz,1H),6.83(t,J=6.2Hz,2H),6.75(s,1H),6.37(s,2H),6.22(s,1H),4.91(dd,J=12.2,5.3Hz,1H),4.53(s,2H),3.86 (s,3H),3.83(d,J=2.7Hz,9H),3.32(q,J=6.9Hz,2H),3.25(q,J=6.3Hz,2H),2.88(dd,J=12.5,7.2Hz,1H),2.82–2.80(m,4 H),2.77(dd,J=13.6,3.6Hz,1H),2.13(dt,J=10.6,4.3Hz,1H),1.53(t,J=7.3Hz,3H),1.39(d,J=8.6Hz,2H),1.32(s,9H). 13 C NMR (126MHz, DMSO-d6) δ172.82,170.10,167.72,167.32,152.65,147.15,146.43,136.28,135.60,134.12,132.19,121.80,117.15,115.37, 112.27,110.38,109.00,105.57,68.80,59.96,55.75,55.66,48.55,4 1.82,38.20,37.61,36.56,30.98,29.83,29.05,28.71,26.27,22.16.

[0093] (2) Preparation of final product EC2-8

[0094] The structural formula of the final product EC2 (n = 1) / E-C3 (n = 2) / E-C4 (n = 3) / E-C5 (n = 4) / E-C6 (n = 5) / E-C7 (n = 6) / E-C8 (n = 7) is

[0095] The specific synthesis method is as follows:

[0096] A 25 mL round-bottom flask was charged with Erianin (60 mg, 188.45 mmol), C2 (78.09 mg, 226.14 mmol), EDCL (48.05 mg, 250.65 mmol), 4-DMAP (6.22 mg, 50.91 mmol), and DCM (5 mL). The reaction was stirred at room temperature for 5 h, extracted with DCM, and the organic phase was washed with 5% NaHCO3 solution and saturated sodium chloride, and dried over anhydrous Na2SO4. The solid was removed by filtration, and the solvent was evaporated under reduced pressure to obtain the crude product. After concentration under reduced pressure, PTLC (PE:EA=1:2) was performed to obtain the final product EC2 (yellow powder, 48.2% yield). ESI-MS m / z 645.6650 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.08(s,1H),7.55–7.51(m,1H),7.14(d,J=7.1Hz,1H),7.01–6.96(m,2H) ,6.88(d,J=2.2Hz,1H),6.87(d,J=8.4Hz,1H),6.57(t,J=6.3Hz,1H),6.34(s,2H),4.90(dd,J=12.3,5. 3Hz,1H),3.82(s,3H),3.81(s,6H),3.77(s,3H),3.74(dd,J=13.8,7.3Hz,2H),2.93(t,J=6.6Hz,2H),2 .90–2.81(m,5H),2.78(dd,J=13.0,4.0Hz,1H),2.76–2.68(m,1H),2.10(dtd,J=11.9,4.5,2.1Hz,1H). 13 C NMR(126MHz,Chloroform-d)δ171.11,169.70,169.37,168.39,167.65,153.16,149.22,146.44,139.35,137.38,136.39,136.26,134.40,132.74 ,127.03,122.75,116.59,112.34,112.11,110.72,105.50,60.99,60.54 ,56.18,56.10,49.01,38.51,38.27,37.04,34.13,31.50,22.85,14.32.

[0097] A 25 mL round-bottom flask was charged with Erianin (60 mg, 188.45 mmol), C3 (81.21 mg, 226.14 mmol), EDCL (48.05 mg, 250.65 mmol), 4-DMAP (6.22 mg, 50.91 mmol) and DCM (5 mL). The reaction was stirred at room temperature for 5 h, extracted with DCM, and the organic phase was washed with 5% NaHCO3 solution and saturated sodium chloride, and dried over anhydrous Na2SO4. The solid was removed by filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. After concentration under reduced pressure, PTLC (PE:EA=1:2) was performed to obtain the final product EC3 (yellow powder, 44.1% yield). ESI-MS m / z 659.6520 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.07(s,1H),7.50(dd,J=8.6,7.1Hz,1H),7.11(d,J=7.1H z,1H),7.01–6.97(m,2H),6.90–6.85(m,2H),6.37(t,J=6.0Hz,1H),6.34(s,2H),4.92(d d,J=12.4,5.2Hz,1H),3.82(d,J=2.1Hz,9H),3.79(s,3H),3.49–3.43(m,2H),2.92–2.86 (m,1H),2.84(h,J=3.1Hz,4H),2.82–2.78(m,1H),2.77–2.69(m,3H),2.15–2.07(m,3H). 13 C NMR(126MHz,Chloroform-d)δ171.23,171.07,169.60,168.41,167.70,153.18,149.29,146.95,139.47,137.38,136.38,136.27,134.47,132 .64,126.94,122.84,116.82,112.37,111.86,110.29,105.49,61.01, 56.18,56.09,49.01,41.80,38.36,37.10,31.54,31.13,24.75,22.92.

[0098] Erianin (60 mg, 188.45 mmol), C4 (84.38 mg, 226.14 mmol), EDCL (48.05 mg, 250.65 mmol), 4-DMAP (6.22 mg, 50.91 mmol) and DCM (5 mL) were added to a 25 mL round-bottom flask. The reaction was stirred at room temperature for 5 h, extracted with DCM, and the organic phase was washed with 5% NaHCO3 solution and saturated sodium chloride, and dried over anhydrous Na2SO4. The solid was removed by filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. After concentration under reduced pressure, PTLC (PE:EA=1:2) was performed to obtain the final product EC4 (yellow powder, 42.9% yield). ESI-MS m / z 701.7290 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.08(s,1H),7.49(dd,J=8.5,7.1Hz,1H),7.10(d,J=7.1Hz,1H),6.97(dd,J=8.3,2.2 Hz,1H),6.91(d,J=8.5Hz,1H),6.88–6.84(m,2H),6.34(s,2H),6.28(t,J=5.6Hz,1H),4.93–4.88(m,1H),3.82(s,3 H),3.81(s,6H),3.78(s,3H),3.35(q,J=6.4Hz,2H),2.91–2.85(m,1H),2.83(h,J=3.1Hz,4H),2.81–2.77(m,1H),2 .76–2.71(m,1H),2.65(t,J=7.0Hz,2H),2.15–2.10(m,1H),1.89(dtd,J=14.1,7.1,3.2Hz,2H),1.86–1.79(m,2H). 13 C NMR(126MHz,Chloroform-d)δ171.45,171.09,169.60,168.42,167.72,153. 16,149.35,147.00,139.55,137.41,136.31,136.25,134.41,132.63,126.8 4,122.88,116.74,112.35,111.69,110.14,105.49,61.00,60.55,56.17,56.09,48.99,42.44,38.36,37.09,33.65,31.53,28.77,22.91,22.47,14.33.

[0099] A 25 mL round-bottom flask was charged with Erianin (60 mg, 188.45 mmol), C5 (87.55 mg, 226.14 mmol), EDCL (48.05 mg, 250.65 mmol), 4-DMAP (6.22 mg, 50.91 mmol) and DCM (5 mL). The reaction was stirred at room temperature for 5 h, extracted with DCM, and the organic phase was washed with 5% NaHCO3 solution and saturated sodium chloride, and dried over anhydrous Na2SO4. The solid was removed by filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. After concentration under reduced pressure, PTLC (PE:EA=1:2) was performed to obtain the final product EC5 (yellow powder, 43.6% yield). ESI-MS m / z 687.7460 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.03(s,1H),7.49(dd,J=8.6,7.1Hz,1H),7.09(d,J=7.0Hz,1H),6. 96(dd,J=8.4,2.2Hz,1H),6.90–6.84(m,3H),6.34(s,2H),6.25(t,J=5.7Hz,1H),4.93–4.88(m,1 H),3.82(d,J=3.1Hz,9H),3.79(s,3H),3.34–3.28(m,2H),2.92–2.68(m,7H),2.61(t,J=7.4Hz,2 H),2.15–2.09(m,1H),1.83(p,J=7.5Hz,2H),1.75(p,J=7.3Hz,2H),1.57(dd,J=10.1,4.8Hz,2H). 13 CNMR(126MHz,Chloroform-d)δ172.04,171.07,169.64,168.43,167.77,153. 17,149.45,147.16,139.73,137.46,136.27,134.40,132.62,126.71,122.95, 116.78,112.41,111.62,111.52,109.96,105.51,61.00,56.18,56.11,48.99,42.78,38.36,37.11,34.14,31.54,29.34,29.28,29.10,27.03,25.09,22.94.

[0100] Erianin (60 mg, 188.45 mmol), C6 (90.72 mg, 226.14 mmol), EDCL (48.05 mg, 250.65 mmol), 4-DMAP (6.22 mg, 50.91 mmol) and DCM (5 mL) were added to a 25 mL round-bottom flask. The reaction was stirred at room temperature for 5 h, extracted with DCM, and the organic phase was washed with 5% NaHCO3 solution and saturated sodium chloride, and dried over anhydrous Na2SO4. The solid was removed by filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. After concentration under reduced pressure, PTLC (PE:EA=1:2) was performed to obtain the final product EC6 (yellow powder, 41.8% yield). ESI-MS m / z 701.2948 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.05(s,1H),7.48(dd,J=8.6,7.1Hz,1H),7.09(d,J=7.0Hz,1H),6.96(dd,J=8.3 ,2.2Hz,1H),6.90–6.85(m,3H),6.34(s,2H),6.24(t,J=5.6Hz,1H),4.93–4.88(m,1H),3.82(d,J=3.1Hz,8H),3 .79(s,3H),3.28(q,J=6.9Hz,2H),2.91–2.86(m,1H),2.83(h,J=3.2Hz,4H),2.81–2.76(m,1H),2.76–2.69(m, 1H), 2.59 (t, J = 7.4Hz, 2H), 2.15–2.10 (m, 1H), 1.82–1.75 (m, 2H), 1.71 (h, J = 7.2Hz, 3H), 1.49 (p, J = 3.5Hz, 4H). 13 C NMR(126MHz,Chloroform-d)δ172.03,171.11,169.63,168.46,167.76,153.16,149.44,147.14,139.72,137.45,136.25,134.39,132.61,126.70,1 22.94,116.77,112.40,111.50,109.95,105.50,60.99,56.17,56.11,48. 98,42.77,38.36,37.10,34.13,31.53,29.28,29.09,27.02,25.08,22.93.

[0101] Erianin (60 mg, 188.45 mmol), C7 (93.89 mg, 226.14 mmol), EDCL (48.05 mg, 250.65 mmol), 4-DMAP (6.22 mg, 50.91 mmol) and DCM (5 mL) were added to a 25 mL round-bottom flask. The reaction was stirred at room temperature for 5 h, extracted with DCM, and the organic phase was washed with 5% NaHCO3 solution and saturated sodium chloride, and dried over anhydrous Na2SO4. The solid was removed by filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. After concentration under reduced pressure, PTLC (PE:EA=1:2) was performed to obtain the final product EC7 (yellow powder, 43.2% yield). ESI-MS m / z 715.3105 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.20(s,1H),7.48(dd,J=8.5,7.1Hz,1H),7.08(d,J=7.1Hz,1H),6.96(dd,J=8.4,2.2Hz,1H) ,6.89–6.86(m,2H),6.85(s,1H),6.34(s,2H),6.23(t,J=5.5Hz,1H),4.93–4.88(m,1H),3.81(d,J=3.2Hz,9H),3.78(s,3H ),3.26(td,J=7.0,5.5Hz,2H),2.89–2.84(m,1H),2.83(h,J=3.2Hz,4H),2.81–2.76(m,1H),2.73(ddd,J=16.5,4.6,2.5Hz ,1H),2.57(t,J=7.5Hz,2H),2.14–2.08(m,1H),1.76(p,J=7.2Hz,2H),1.68(t,J=6.8Hz,3H),1.45(dd,J=12.9,7.2Hz,6H). 13 C NMR(126MHz,Chloroform-d)δ171.98,171.10,169.63,168.44,167.76,153.1 6,149.42,147.12,139.69,137.45,136.27,136.24,134.38,132.60,126.72,1 22.93,116.77,112.39,111.53,109.95,105.48,61.00,56.17,56.10,48.97,42.74,38.36,37.10,34.08,31.53,29.31,29.11,29.04,26.92,25.02,22.92.

[0102] Erianin (60 mg, 188.45 mmol), C8 (97.06 mg, 226.14 mmol), EDCL (48.05 mg, 250.65 mmol), 4-DMAP (6.22 mg, 50.91 mmol) and DCM (5 mL) were added to a 25 mL round-bottom flask. The reaction was stirred at room temperature for 5 h, extracted with DCM, and the organic phase was washed with 5% NaHCO3 solution and saturated sodium chloride, and dried over anhydrous Na2SO4. The solid was removed by filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. After concentration under reduced pressure, PTLC (PE:EA=1:2) was performed to obtain the final product EC8 (yellow powder, 44.6% yield). ESI-MS m / z 729.3261 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.13(s,1H),7.48(dd,J=8.5,7.1Hz,1H),7.08(d,J=7.1Hz,1H),6.96(dd,J=8.3,2.2Hz,1H ),6.89–6.85(m,3H),6.34(s,2H),6.23(t,J=5.6Hz,1H),4.91(dd,J=12.3,5.3Hz,1H),3.81(d,J=3.0Hz,9H),3.79(s,3H) ,3.26(td,J=7.0,5.5Hz,2H),2.88(ddd,J=16.3,4.6,2.8Hz,1H),2.83(h,J=3.2Hz,4H),2.79(dd,J=12.6,3.9Hz,1H),2.7 6–2.70(m,1H),2.57(t,J=7.5Hz,2H),2.15–2.09(m,1H),1.76(p,J=7.4Hz,2H),1.67(p,J=7.1Hz,2H),1.46–1.35(m,8H). 13 C NMR(126MHz,Chloroform-d)δ171.88,171.13,169.63,168.47,167.75,153.1 5,149.40,147.09,139.65,137.44,136.27,136.24,134.39,132.60,126.73,1 22.92,116.76,112.38,111.55,111.49,109.98,105.47,60.99,56.16,56.10,48.97,42.68,38.35,37.10,34.00,31.52,29.21,28.83,26.76,24.96,22.92.

[0103] (3) Preparation of final product EO1-3

[0104] The structural formula of the final product EO1(n=1) / EO2(n=2) / EO3(n=3) is

[0105] The specific synthesis method is as follows:

[0106] Erianin-C1 (60 mg, 0.16 mmol) and DIPEA (76 μL, 0.43 mmol) were sequentially dissolved in dry DMF and stirred at room temperature for 5 min. HATU (75.03 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. O1 (86.14 mg, 0.24 mmol) was added and stirred at room temperature. TLC monitoring of the reaction indicated completion in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined and washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) afforded the product EO1 (yellow powder, 37.5% yield). ESI-MS m / z 718.2850 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.17(s,1H),7.47(dd,J=8.5,7.1Hz,1H),7.34(t,J=5.8Hz,1H),7.08(d,J=7.2Hz,1H),6 .90(d,J=8.5Hz,1H),6.80(d,J=1.4Hz,2H),6.74(d,J=1.4Hz,1H),6.37(s,2H),4.85(dd,J=12.3,5.3Hz,1H),4.53(s,2 H),3.83(s,6H),3.82(s,3H),3.82(s,3H),3.67(t,J=5.4Hz,2H),3.60(t,J=5.5Hz,2H),3.56(dd,J=6.3,4.6Hz,2H),3. 43(t,J=5.4Hz,2H),2.87–2.82(m,1H),2.80(q,J=3.0Hz,4H),2.78–2.71(m,1H),2.71–2.63(m,1H),2.08–2.02(m,1H). 13CNMR(126MHz,DMSO-d6)δ171.11,169.45,169.14,168.43,167.67,153.2 2,148.13,147.22,146.91,137.36,136.39,136.23,134.89,132.61,122. 86,116.85,116.11,112.18,111.93,110.53,105.52,69.96,69.86,69.4 5,61.01,56.22,56.11,48.99,42.43,38.96,38.57,37.42,31.50,22.85.

[0107] Erianin-C1 (60 mg, 0.16 mmol) and DIPEA (76 μL, 0.43 mmol) were sequentially dissolved in dry DMF and stirred at room temperature for 5 min. HATU (75.03 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. O1 (96.65 mg, 0.24 mmol) was added and stirred at room temperature. TLC followed the reaction, indicating completion in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined, washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) afforded the product EO2 (yellow powder, 39.1% yield). ESI-MS m / z 762.3112 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.61(s,1H),7.46(dd,J=8.5,7.1Hz,1H),7.40(t,J=5.7Hz,1H),7.08(d,J=7.0Hz,1H) ,6.86(d,J=8.5Hz,1H),6.83–6.77(m,2H),6.74(d,J=1.8Hz,1H),6.37(s,2H),4.90(dd,J=12.0,5.5Hz,1H),4.52(s ,2H),3.83(d,J=3.2Hz,9H),3.82(s,3H),3.70(t,J=5.3Hz,2H),3.64(s,4H),3.62(t,J=4.8Hz,2H),3.54(q,J=5.5H z,2H),3.44–3.39(m,2H),2.84(dd,J=13.1,3.0Hz,1H),2.80(q,J=2.6Hz,4H),2.78–2.69(m,2H),2.14–2.08(m,1H). 13C NMR(126MHz,DMSO-d6)δ171.33,169.43,169.11,168.64,167.72,153.21,14 8.11,147.20,146.88,137.41,136.36,136.14,134.91,132.67,122.84,116 .79,116.11,112.18,111.80,110.50,105.50,70.93,70.38,70.00,69.85,69.44,61.00,56.21,56.10,49.00,42.42,39.08,38.58,37.44,31.49,23.00.

[0108] Erianin-C1 (60 mg, 0.16 mmol) and DIPEA (76 μL, 0.43 mmol) were sequentially dissolved in dry DMF and stirred at room temperature for 5 min. HATU (75.03 mg, 0.20 mmol) was added and the reaction was monitored by TLC for approximately 0.5 h. O1 (107.17 mg, 0.24 mmol) was added and stirred at room temperature. TLC followed the reaction, indicating completion in approximately 4 h. The reaction solution was quenched by the addition of crushed ice and extracted three times with ethyl acetate. The organic layers were combined, washed sequentially with saturated ammonium chloride solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. PTLC (PE:EA = 1:2) was performed to obtain the product EO3 (yellow powder, 37.7% yield). ESI-MS m / z 806.3374 [M+Na] + . 1 H NMR(500MHz,Chloroform-d)δ8.47(s,1H),7.47(dd,J=8.5,7.1Hz,1H),7.39(t,J=5.7Hz,1H),7.09(d,J=7.1H z,1H),6.89(d,J=8.5Hz,1H),6.83–6.78(m,2H),6.75(d,J=1.8Hz,1H),6.37(s,2H),4.90(dd,J=12.3,5.3Hz, 1H),4.53(s,2H),3.85(s,3H),3.83(s,6H),3.82(s,2H),3.69(t,J=5.4Hz,2H),3.65–3.57(m,8H),3.53(q,J= 5.1Hz,2H),3.44(t,J=5.4Hz,2H),2.88–2.82(m,1H),2.82–2.78(m,4H),2.77–2.67(m,2H),2.13–2.06(m,1H). 13C NMR(126MHz,DMSO-d6)δ171.28,169.40,169.12,168.55,167.74,153.22,148 .09,147.19,146.93,137.42,136.36,136.16,134.89,132.66,122.81,116.87 ,116.00,112.15,111.80,110.44,105.51,70.92,70.73,70.48,69.86,69.78,69.58,61.01,56.22,56.11,49.01,42.51,39.00,38.61,37.47,31.55,22.93.

[0109] Example 3: In vitro anti-tumor activity test of the Erianin PROTACs compound of the present invention (IC 50 )

[0110] The Erianin PROTACs prepared in Examples 1 and 2 of the present invention were tested for their ability to inhibit tumor cell proliferation using the conventional CCK-8 method. When the tumor cells (NCI-H441 cells (human lung adenocarcinoma cells)) grew to 80%-90% of the culture dish, the cells were digested from the culture dish, centrifuged and resuspended in fresh DMEM / 1640 complete medium, counted under a microscope, and then the cell suspension was diluted to 50 cells / μL. A circle of sterile PBS was spread on the outermost periphery of the 96-well plate, 100 μL per well, and 100 μL of the diluted cell suspension was spread on each well of the remaining wells and cultured in a 37°C, 5% CO2 incubator. After 24 hours, the medium in the 96-well plate was removed, and the drug was diluted with fresh complete medium in a certain concentration gradient, and then added to the 96-well plate in sequence. After 48 hours, the medium containing the drug was removed, and CCK-8 liquid diluted with the medium was added (100 μL medium + 10 μL CCK-8), and the cells were placed in a 37 ° C, 5% CO2 incubator for 30-40 minutes. The absorbance was detected at OD 450 on a microplate reader, and the inhibition rate was calculated based on the absorbance at OD450. The data were used to calculate the half-maximal inhibitory concentration (IC) using SPSS software. 50 .

[0111] The calculation formula is as follows:

[0112] Inhibition rate = [(Ac-As) / (Ac-Ab)] × 100%

[0113] As: absorbance of experimental wells (containing cells, culture medium, CCK-8 solution, and drug solution);

[0114] Ac: absorbance of control well (containing cells, culture medium, and CCK-8 solution, but no drug);

[0115] Ab: absorbance of blank wells (containing culture medium and CCK-8 solution, but not cells or drugs).

[0116] Experimental results: To further evaluate the in vitro anti-tumor activity of the target compound, NCI-H446 cells (human lung adenocarcinoma cells), NCI-H460 cells (human large cell lung cancer cells), and A549 cells (lung cancer cells) were selected as test tumor lines, and Erianin was used as the control group. The test results are shown in Table 1. Overall, the anti-tumor activity of the above-mentioned Erianin PROTACs compounds is better than that of Erianin itself. Combined with IC 50 and compound structures, EC2 was selected as the dosing group for further western blotting studies.

[0117] Table 1 In vitro anti-lung cancer activities of Erianin, EF2-8, EC2-8 and EO1-3 (IC 50 )

[0118]

[0119] Example 4: Degradation efficiency test of target protein by different concentrations of EC2 in the Erianin PROTACs compound of the present invention

[0120] Compound EC2 exhibited good anti-tumor activity in various lung cancer cells. Therefore, different concentrations of EC2 were used to act on NCI-H460 cells to study the target protein degradation efficiency at different concentrations and to explore whether the degradation of OTUB1 is dependent on the action concentration of EC2. The test method used was conventional Western blotting.

[0121] Experimental results: The degradation of OTUB1 on EC2 is concentration-dependent ( Figure 1 ).

[0122] At the same time, the degradation efficiency of target proteins by EC2 at different concentrations was investigated in NCI-H441 cells, specifically to explore whether the degradation of KRAS is dependent on the action concentration of EC2. The test method used was conventional WB method.

[0123] Experimental results: KRAS degradation has a concentration-dependent effect on EC2 ( Figure 2 ).

[0124] Example 5: Exploration of the degradation mechanism of target protein by EC2 in the Erianin PROTACs compound of the present invention

[0125] Since the degradation of the two target proteins by EC2 is concentration-dependent, the proteasome inhibitor MG132 was selected to verify the ubiquitination degradation, and the test method used was the conventional WB method.

[0126] Experimental results: In the groups that were treated with different concentrations of MG132 and then EC2 (20nM), the protein level of OTUB1 was significantly increased compared to the group treated with EC2 (20nM) alone, and the administration of MG132 alone did not cause significant changes in the protein. EC2 degrades OTUB1 protein through the ubiquitin proteasome pathway ( Figure 3 ).

[0127] Experimental results: In the groups that were treated with different concentrations of MG132 and then EC2 (200μM), the protein level of KRAS was significantly increased compared to the group treated with EC2 (200μM) alone, and the administration of MG132 alone did not cause significant changes in the protein. EC2 degrades KRAS protein through the ubiquitin proteasome pathway ( Figure 4 ).

[0128] The above description is merely a preferred embodiment of the present invention and therefore cannot be used to limit the scope of the present invention. In other words, equivalent changes and modifications made within the scope of the present invention and the contents of the specification should still fall within the scope of the present invention.

Claims

1. A PROTACs for Erianin, characterized by: Its structural formula is Among them, E3 Ligand is pomalidomide, Linker is an alkane chain or PEG chain, The alkane chain is -CH2-CO-NH-(CH2) n1 -(CH2)-NH-, n1 is 1, 2, 3, 4, 5 or 7; or the alkane chain is -CO-(CH2) n2 -(CH2)-NH-, n2 is 1, 2, 3, 4, 5, 6 or 7, The PEG chain is -CH2-CO-NH-CH2-(CH2-O-CH2) n3 -CH2-NH-, n3 is 1, 2 or 3.

2. The Erianin PROTACs according to claim 1, wherein: Its structural formula is selected from one of the following:

3. The method for preparing Erianin PROTACs according to claim 1 or 2, characterized in that: The reaction route is:

4. Use of the Erianin PROTACs or pharmacologically or physiologically acceptable salts thereof according to claim 1 or 2 in the preparation of a cancer therapeutic composition.

5. The use according to claim 4, characterized in that: The cancer is lung cancer.

6. A cancer treatment composition, characterized in that: The active ingredient comprises the Erianin PROTACs according to claim 1 or 2 or a pharmacologically or physiologically acceptable salt thereof.

7. The cancer therapeutic composition according to claim 6, wherein: The cancer is lung cancer.

8. Use of the Erianin PROTACs or pharmacologically or physiologically acceptable salts thereof according to claim 1 or 2 in the preparation of an anti-inflammatory composition.

9. An anti-inflammatory composition, characterized in that: The active ingredient comprises the Erianin PROTACs according to claim 1 or 2 or a pharmacologically or physiologically acceptable salt thereof.

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