WRN degradation agent as well as preparation method and application thereof

Abstract

The invention discloses a novel difunctional micromolecule (I) with WRN degradation activity and a preparation method and application thereof. The compound can significantly proliferate tumor cells at a micromolar concentration level, has dual functions of targeted inhibition and degradation, and has an application prospect of improving drug resistance and toxicity of antitumor drugs. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to a novel bifunctional small molecule and a preparation method and application thereof, and in particular to a bifunctional small molecule with WRN protein degradation activity and a preparation method and application thereof. Background Art

[0002] In tumor cells with mismatch repair deficiency (dMMR), damage to the microsatellite region cannot be corrected in time, resulting in deletion or amplification of repeated sequences, a phenomenon called microsatellite instability (MSI). In sporadic non-hereditary tumors, MSI is common in colorectal cancer, endometrial cancer, gastric cancer, and ovarian cancer. Currently, immune checkpoint inhibitors have been clinically proven to be effective against several tumors carrying MSI-H (high microsatellite instability). However, the response rate of MSI-H tumors to immune checkpoint inhibitors is limited, and the dosage of immunosuppressants is limited by toxicity.

[0003] Studies have found that after knocking out the WRN gene or depleting the WRN protein, dMMR / MSI tumor cells will experience synthetic lethality, leading to cell death. WRN (Werner helicase, WRN) belongs to the RecQ helicase, and its gene is located on chromosome 8p11-12 and consists of 34 exons. The WRN protein is composed of 1432 amino acids and has a total of 5 domains, namely the 3'→5' exonuclease domain located in the N-terminal region, the ATPase domain, the RQC (RecQ C-terminal) domain that binds to DNA, the HRDC (helicase and ribonuclease DC-terminal) domain that mediates protein interaction, and the nuclear localization signal domain at the C-terminal. Among them, the ATPase domain and the RQC domain are the core of WRN's unwinding function and are also the key regions where WRN participates in the synthetic lethality effect.

[0004] Currently, Novartis' small molecule inhibitor HRO-761 and Vividion Therapeutics' covalent small molecule inhibitor VVD-133214 are in Phase I clinical trials, both of which specifically target the helicase domain of WRN. However, long-term use of WRN small molecule inhibitors can lead to drug resistance, ultimately reducing the therapeutic effect of the drug. Summary of the invention

[0005] Purpose of the invention: The first purpose of the present invention is to provide a new bifunctional small molecule, the second purpose is to provide a method for preparing the compound, and the third purpose is to provide a pharmaceutical application of the compound and its pharmaceutical composition.

[0006] Technical solution: The compound represented by the general formula (I) of the present invention or a pharmaceutically acceptable salt thereof,

[0007]

[0008] in:

[0009] R 1 Selected from

[0010] R 2 for R 3 is selected from H, OCH3, OH, CN or halogen, R 4 is selected from H, F, Cl, Br, I or substituted C1-C4 alkyl, R 5 is selected from H, F, Cl, Br, I, SF3, C(O)H or substituted C1-C4 alkyl, substituted C1-C4 alkenyl, substituted C1-C4 alkynyl, substituted C3-C5 cycloalkyl or OCF3, R 6 is selected from H, F, Cl, Br, I, OH, CN, substituted C1-C4 alkyl, substituted C3-C5 cycloalkyl or substituted C1-C4 alkoxy, wherein the substituent is selected from at least one H, F, Cl, Br or I; X is selected from CR 7 or N, R 7 is selected from H, F, Cl, Br or I;

[0011] A is selected from

[0012] n is an integer selected from 1 to 11 (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11), m is an integer selected from 1 to 3 (1, 2, 3), R 8 is selected from H, F, Cl, Br, I or OH, Y, Z, K are independently selected from CH, N or CR 9 , R 9 Selected from H, F, Cl, Br, I, OCH3, OH, CN, CF3 or OCF3.

[0013] Preferably, R 1 Selected from

[0014] Preferably, R 2 Selected from

[0015] Preferably, A is selected from

[0016] n is selected from integers of 1 to 9 (1, 2, 3, 4, 5, 6, 7, 8, 9), and m is selected from integers of 1 to 3 (1, 2, 3).

[0017] Preferably, the compound of the present invention is selected from any one of the following compounds:

[0018]

[0019]

[0020]

[0021]

[0022] Protein degradation targeting chimeras (PROTACs) are hybrid bifunctional compounds that can recruit target proteins and target protein degrading enzymes, and specifically degrade target proteins using the ubiquitin-proteasome pathway. The present invention uses PROTAC technology to develop a degrader targeting WRN, which will provide a new option for the development of drugs targeting WRN, and is also expected to solve the potential drug resistance and toxic side effects of WRN inhibitors.

[0023] Preferably, the pharmaceutically acceptable salt is an acid addition salt formed by the compound and any one of the following acids: hydrogen chloride, hydrogen bromide, sulfuric acid, carbonic acid, oxalic acid, citric acid, malic acid, succinic acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid or ferulic acid.

[0024] "Pharmaceutically acceptable salts" refer to salts of compounds of the invention, prepared from compounds of the invention having specific substituents with relatively nontoxic acids or bases. When the compounds of the invention contain relatively acidic functional groups, base addition salts can be obtained by contacting the free form of such compounds with a sufficient amount of base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amino or magnesium salts or similar salts. When the compounds of the invention contain relatively basic functional groups, acid addition salts can be obtained by contacting the free form of such compounds with a sufficient amount of acid in a pure solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid (forming carbonates or bicarbonates), phosphoric acid (forming phosphates, monohydrogen phosphates, dihydrogen phosphates, sulfuric acid (forming sulfates or bisulfates), hydroiodic acid, phosphorous acid, and the like; and organic acid salts such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, Citric acid, tartaric acid and methanesulfonic acid and similar acids; organic acid salts also include amino acids (such as arginine, etc.), glucuronic acid and other organic acid salts. Certain specific compounds of the present invention contain basic and acidic functional groups, and can be converted into any base or acid addition salt. Preferably, the salt is contacted with a base or acid in a conventional manner, and the parent compound is separated, thereby regenerating the free form of the compound. The free form of the compound differs from its various salt forms in certain physical properties, such as different solubility in polar solvents.

[0025] The "pharmaceutically acceptable salts" of the present invention can be synthesized from parent compounds containing acid radicals or bases by conventional chemical methods. In general, such salts are prepared by reacting these compounds in free acid or base form with a stoichiometric amount of an appropriate base or acid in water or an organic solvent or a mixture of the two. In general, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred.

[0026] The method for preparing the compound or a pharmaceutically acceptable salt thereof of the present invention is selected from any one of the following methods:

[0027] Method 1: When A is When the compound I-1 is acylated, iodinated, substituted, coupled, deprotected and substituted, the compound IA is obtained;

[0028]

[0029] Compound I-3 is prepared by acylation reaction of compounds I-1 and I-2, the base used is triethylamine, N,N-diisopropylethylamine, potassium carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide, sodium hydrogen hydride, potassium tert-butoxide, sodium methoxide or sodium ethoxide, preferably N,N-diisopropylethylamine; the reaction solvent used is one of dichloromethane, ethyl acetate, acetone, tetrahydrofuran, dichloromethane, acetonitrile, toluene, ethylene glycol dimethyl ether, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide or a mixture of any two thereof, preferably dichloromethane; the reaction temperature is 0 to 100°C, preferably 0 to 30°C.

[0030] Compound I-4 is prepared from compound I-3 by substitution reaction, the iodine salt used is potassium iodide or sodium iodide, preferably potassium iodide; the reaction solvent used is one of dichloromethane, ethyl acetate, acetone, tetrahydrofuran, dichloromethane, acetonitrile, toluene, ethylene glycol dimethyl ether, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide or a mixture of any two thereof, preferably acetone; the reaction temperature is 0 to 100°C, preferably 50 to 80°C.

[0031] Compound I-6 is prepared from compound I-4 and compound I-5, and the base used is triethylamine, N,N-diisopropylethylamine, potassium carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide, sodium hydrogen, potassium tert-butoxide, sodium methoxide or sodium ethoxide, preferably N,N-diisopropylethylamine; the reaction solvent used is ethyl acetate, acetone, tetrahydrofuran, dichloromethane, acetonitrile, toluene, ethylene glycol dimethyl ether, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide or a mixture of any two thereof, preferably dioxane; the reaction temperature is 0 to 100°C, preferably 20 to 30°C.

[0032] Compound I-8 is prepared by Suzuki coupling reaction of compound I-6 and organoboron reagent I-7, wherein the solvent used is selected from tetrahydrofuran, acetonitrile, N-methylpyrrolidone, N,N-dimethylacetamide, dimethyl sulfoxide, N,N-dimethylformamide, 1,4-dioxane or water, or a mixture of any two thereof, preferably a 3:1 mixed solvent of 1,4-dioxane and water; and the base used is selected from potassium bicarbonate, potassium carbonate, sodium bicarbonate, sodium carbonate, potassium phosphate, sodium acetate, triethylamine or N,N-diisopropylamine. Ethylamine, preferably potassium phosphate; the catalyst used is selected from tetrakis(triphenylphosphine)palladium (Pd(PPh3)4), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (Pd(dppf)Cl2), palladium acetate (Pd(OAc)2), tris(dibenzylideneacetone)dipalladium (Pd2(dba)3) or bis(triphenylphosphine)palladium dichloride (Pd(PPh3)2Cl2), preferably Pd(dppf)Cl2; the reaction temperature is 25°C to 150°C, preferably 60 to 100°C.

[0033] Compound I-9 is prepared from compound I-8 by acid deprotection, the acid used is hydrogen chloride, trifluoroacetic acid, trifluoromethanesulfonic acid, sulfuric acid, hydrobromic acid or phosphoric acid, preferably hydrogen chloride; the reaction solvent used is ethyl acetate, tetrahydrofuran, dioxane, methanol, ethanol or diethyl ether or a mixture of any two, preferably dioxane; the reaction temperature is 0 to 60°C, preferably 20 to 30°C.

[0034] Compound IA is prepared from compound I-9 and compound I-10, and the base used is triethylamine, N,N-diisopropylethylamine, potassium carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide, sodium hydrogen hydride, potassium tert-butoxide, sodium methoxide or sodium ethoxide, preferably N,N-diisopropylethylamine; the reaction solvent is ethyl acetate, acetone, tetrahydrofuran, dichloromethane, acetonitrile, toluene, ethylene glycol dimethyl ether, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide or a mixture of any two thereof, preferably N,N-dimethylformamide; the reaction temperature is 0 to 150°C, preferably 25 to 80°C.

[0035] Method 2: When A is When the compound I-9 is condensed with the compound I-11, the compound IB is obtained;

[0036]

[0037] Compound IB is prepared from compound I-9 and compound I-11, the base used is triethylamine, N,N-diisopropylethylamine, potassium carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide, sodium hydrogen, potassium tert-butoxide, sodium methoxide or sodium ethoxide, preferably N,N-diisopropylethylamine; the condensation agent used is N,N,N′,N′-tetramethyl-O-(7-azabenzotriazole-1-yl) urea hexafluorophosphate (HATU), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI), 1-Hydroxybenzotriazole (HOBt), propylphosphonic anhydride (T3P) and trichloro-s-triazine, preferably N,N,N′,N′-tetramethyl-O-(7-azabenzotriazole-1-yl)urea hexafluorophosphate (HATU); the reaction solvent is one of ethyl acetate, acetone, tetrahydrofuran, dichloromethane, acetonitrile, toluene, ethylene glycol dimethyl ether, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide or a mixture of any two thereof, preferably N,N-dimethylformamide; the reaction temperature is 0 to 60°C, preferably 0 to 25°C.

[0038] Method 3: When A is When the compound I-9 is condensed with the compound I-12, the compound IC is obtained;

[0039]

[0040] Compound IB is prepared from compound I-9 and compound I-12, the base used is triethylamine, N,N-diisopropylethylamine, potassium carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide, sodium hydrogen, potassium tert-butoxide, sodium methoxide or sodium ethoxide, preferably N,N-diisopropylethylamine; the condensation agent used is N,N,N′,N′-tetramethyl-O-(7-azabenzotriazole-1-yl) urea hexafluorophosphate (HATU), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI), 1-Hydroxybenzotriazole (HOBt), propylphosphonic anhydride (T3P) and trichloro-s-triazine, preferably N,N,N′,N′-tetramethyl-O-(7-azabenzotriazole-1-yl)urea hexafluorophosphate (HATU); the reaction solvent is one of ethyl acetate, acetone, tetrahydrofuran, dichloromethane, acetonitrile, toluene, ethylene glycol dimethyl ether, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide or a mixture of any two thereof, preferably N,N-dimethylformamide; the reaction temperature is 0 to 60°C, preferably 0 to 25°C.

[0041] Among them, R 1 , R 2 , n is defined as above;

[0042] The compounds IA, IB and IC prepared by the above methods are salified with a pharmaceutically acceptable acid to obtain the pharmaceutically acceptable salts.

[0043] The pharmaceutical composition of the present invention comprises the compound of the present invention or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

[0044] The pharmaceutically acceptable carrier can be an adjuvant widely used in the field of drug production. The adjuvant is mainly used to provide a safe, stable and functional pharmaceutical composition, and can also provide a method to dissolve the active ingredient at a desired rate after the subject receives the administration, or promote the effective absorption of the active ingredient after the subject receives the composition. The pharmaceutical adjuvant can be an inert filler, or provide a certain function, such as stabilizing the overall pH value of the composition or preventing the degradation of the active ingredient of the composition. The pharmaceutical adjuvant can include one or more of the following adjuvants: adhesives, suspending agents, emulsifiers, diluents, fillers, granulating agents, adhesives, disintegrants, lubricants, anti-adhesive agents, glidants, wetting agents, gelling agents, absorption delay agents, dissolution inhibitors, enhancers, adsorbents, buffers, chelating agents, preservatives, colorants, flavoring agents and sweeteners.

[0045] The pharmaceutical composition of the present invention can be prepared according to the disclosed content using any method known to those skilled in the art, such as conventional mixing, dissolving, granulating, emulsifying, grinding, encapsulating, embedding or lyophilizing processes.

[0046] The pharmaceutical composition of the present invention can be administered in any form, including injection (intravenous), mucosal, oral (solid and liquid preparations), inhalation, ocular, rectal, topical or parenteral (infusion, injection, implantation, subcutaneous, intravenous, intraarterial, intramuscular) administration. The pharmaceutical composition of the present invention can also be a controlled release or sustained release dosage form (e.g., liposomes or microspheres). Examples of solid oral preparations include, but are not limited to, powders, capsules, caplets, soft capsules, and tablets. Examples of liquid preparations for oral or mucosal administration include, but are not limited to, suspensions, emulsions, elixirs, and solutions. Examples of topical preparations include, but are not limited to, emulsions, gels, ointments, creams, patches, pastes, foams, lotions, drops, or serum preparations. Examples of preparations for parenteral administration include, but are not limited to, solutions for injection, dry powder preparations that can be dissolved or suspended in a pharmaceutically acceptable carrier, suspensions for injection, and emulsions for injection. Examples of other suitable formulations of the pharmaceutical composition include, but are not limited to, eye drops and other ophthalmic preparations; aerosols, such as nasal sprays or inhalers; liquid dosage forms suitable for parenteral administration; suppositories and lozenges.

[0047] The compound or pharmaceutically acceptable salt thereof or pharmaceutical composition thereof described in the present invention is used in the preparation of WRN protein degrading agent.

[0048] Preferably, the drug is a drug for treating tumors.

[0049] More preferably, the drug is a drug for treating colorectal cancer, gastric cancer, prostate cancer, endometrial cancer, adrenocortical carcinoma, uterine cancer, cervical cancer, esophageal cancer, breast cancer, kidney cancer or ovarian cancer.

[0050] The hydrate, solvate or crystal of the compound of general formula (I) of the present invention and the use thereof in the preparation of anti-tumor drugs also fall within the protection scope of the present invention.

[0051] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0052] The compounds designed by the present invention can significantly inhibit tumor cell proliferation, and the inhibitory activity reaches the micromolar concentration level, and have the dual functions of targeted inhibition and degradation to improve the drug resistance and toxicity of anti-tumor drugs. DETAILED DESCRIPTION

[0053] The technical solution of the present invention is further described below in conjunction with embodiments.

[0054] Example 1: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)2-(2-(3,6-dihydro-2H-pyran-4-yl)6-(4-(2-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amino)-2-oxoethyl)piperazin-1-yl)5-ethyl-7-oxo[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IA-1)

[0055] (1) Synthesis of 2-bromo-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide (I-3)

[0056] Dissolve 2-chloro-4-(trifluoromethyl)aniline (5.00g, 25.57mmol) and triethylamine (2.60g, 25.57mmol) in DCM (50mL), slowly add bromoacetyl bromide (5.40g, 26.84mmol) under ice bath, and react at room temperature for 6 hours. TLC (PE:EA=4:1) shows that the raw material reacts completely, and the reaction is stopped. Add about 200mL of water, extract three times with 50mL of DCM, combine the organic phases, dry with anhydrous sodium sulfate, and concentrate to obtain a crude product. The crude product is sanded and passed through a column (PE:EA=10:1) to obtain 6.96g of white solid, with a yield of 86.0%. 1 H NMR(300MHz, DMSO-d6)δ(ppm):10.18(s,1H),8.07(d,J=8.4Hz,1H),7.91(s,1H),7.73(d,J=8.1Hz,1H),4.24(s,2H).MS(ESI+)m / z:316.1[M+H] + .

[0057] (2) Synthesis of 2-iodo-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide (I-4)

[0058] Compound I-3 (10.00 g, 31.60 mmol) was dissolved in acetone (136 mL), KI (5.76 g, 34.70 mmol) was added, and the mixture was heated to reflux for 0.5 hours. TLC (PE:EA=8:1) showed that the reaction of the raw material was complete, and the reaction was stopped. The reaction solution was cooled and filtered, and the filtrate was dried to obtain 11.71 g of yellow solid, with a yield of 101.9%. MS (ESI+) m / z: 363.9 [M+H] + .

[0059] (3) Synthesis of tert-butyl 4-(2-bromo-4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amine)-2-oxoethyl)-5-ethyl-7-oxo-4,7-2H-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate (I-6)

[0060] Dissolve 4-(2-bromo-5-ethyl-7-oxo-4,7-2H-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester (1.50 g, 3.51 mmol) and compound I-4 in 1,4-dioxane (15 mL), add DIEA (1.36 g, 10.53 mmol), and stir at room temperature overnight. TLC (PE:EA=1:1) showed that the raw material reacted completely, and the reaction was stopped. Add about 60 mL of water, extract three times with 20 mL of EA, combine the organic phases, dry with anhydrous sodium sulfate, and concentrate to obtain a crude product. The crude product was sanded and passed through a column to obtain an intermediate light yellow solid of 1.32 g, with a yield of 56.7%. MS (ESI+) m / z: 662.10 [M+H] + .

[0061] (4) Synthesis of tert-butyl 4-(4-(2-(((2-chloro-4-(trifluoromethyl)phenyl)amine)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-ethyl-7-oxo-4,7-2H-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate (I-8)

[0062] Compound I-6 (1.32 g, 1.99 mmol), 3,6-dihydro-2H-pyran-4-boronic acid pinacol ester (0.63 g, 2.98 mmol), Pd(dppf)Cl2 (0.15 g, 0.20 mmol), K3PO4 (1.27 g, 6.00 mmol) were dissolved in a mixed solvent (10 mL) of 1,4-dioxane and water (1,4-dioxane: water = 4:1), N2 was protected, and the temperature was raised to 60 ° C for 3 hours. TLC (DCM: MeOH = 20:1) showed that the raw material reacted completely, and the reaction was stopped. About 50 mL of water was added, and 15 mL of EA was used for extraction three times. The organic phases were combined and dried with anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sanded and passed through a column to obtain 0.976 g of a light yellow solid with a yield of 77.1%. 1H NMR (400MHz, DMSO-d6) δ (ppm): 10.35 (s, 1), 8.06 (d, J = 4.8 Hz, 1H), 7.96 (d, J = 2.4H z,1H),7.72(dd,J=4.8Hz,2.4Hz,1H),6.83(s,1H),5.31(s,1H),4.25(q,J=4.8Hz, 2H),3.93(m,2H),3.80(t,J=5.6Hz,2H),3.38(m,2H),2.96(m,2H),2.65(d,J=10.8 Hz,2H),2.52(m,2H),1.43(s,9H),1.17(t,J=7.6Hz,3H).MS(ESI+)m / z:666.0[M+H] + .

[0063] (5) Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (I-9)

[0064] Compound I-8 (0.98 g, 1.47 mmol) was dissolved in DCM (10 mL), and 4M HCl in Dioxane solution (20 mL) was slowly added to the system under ice bath, and the reaction was allowed to proceed overnight at room temperature. TLC (DCM: MeOH = 15: 1) showed that the reaction of the raw material was complete, and the reaction was stopped. 0.67 g of white solid was obtained by spin drying, with a yield of 75.9%. MS (ESI+) m / z: 566.2 [M+H] + .

[0065] (6) Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)2-(2-(3,6-dihydro-2H-pyran-4-yl)6-(4-(2-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amino)-2-oxoethyl)piperazin-1-yl)5-ethyl-7-oxo[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IA-1)

[0066] Compound I-9 (0.15 g, 0.27 mmol) and 2-bromo-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)acetamide (0.13 g, 0.32 mmol) were dissolved in DMF (1.5 mL), and DIEA (0.17 g, 1.33 mmol) was added. The reaction was allowed to proceed overnight at room temperature. TLC (DCM:MeOH=10:1) showed that the reaction of the raw materials was complete, and the reaction was stopped. About 15 mL of water was added, and the mixture was extracted three times with 5 mL of EA. The organic phases were combined, dried with anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.14 g of a yellow solid with a yield of 62.0%. 1 H NMR(300MHz,DMSO-d6)δ(ppm):11.15(s,1),10.83(s,1),10.35(s,1),8.76(d,J=8.4Hz,1H) ,8.07(m,1H),7.97(s,1H),7.86(t,J=7.9Hz,1H),7.71(d,J=8.5Hz,1H),7.61(d,J=7.3Hz,1 H),6.82(s,1H),5.31(s,2H),5.17(m,1H),4.25(s,2H),3.81(m,4H),3.26(m,2H),2.96(m,6 H),2.65(m,5H),2.45(m,2H),2.11(s,1H),1.17(t,J=7.2Hz,3H).MS(ESI+)m / z:879.3[M+H] + .

[0067] Example 2: Synthesis of 4-(4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amine)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazin-1-yl)-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)butanamide (IA-2)

[0068] Compound I-9 (0.15 g, 0.27 mmol) and 4-bromo-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)butyramide (0.17 g, 0.40 mmol) were dissolved in DMF (2 mL), DIEA (0.14 g, 1.06 mmol) was added, and the mixture was heated to 60 °C for 4 h. TLC (DCM: MeOH = 10: 1) showed that the raw materials were basically reacted, and the reaction was stopped. About 15 mL of water was added, and the mixture was extracted three times with 5 mL of EA. The organic phases were combined, dried with anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.15 g of a yellow solid with a yield of 61.2%. 1 H NMR (400MHz, DMSO-d6) δ (ppm): 11.16 (s, 1H), 10.41 (s, 1H), 9.80 (s, 1H), 8.46 (t, J = 8.4Hz, 1H), 8.05 (t, J = 8.0Hz, 1H) ,7.96(d,J=2.1Hz,1H),7.85(t,J=8.4Hz,1H),7.72(dd,J=8.8,2.1Hz,1H),7.63(s,J=8.0Hz,1H),6.83(s,1H),5.32( s,2H),5.15(q,J=12.8,5.4Hz,1H),4.25(d,J=2.9Hz,2H),3.80(t,J=5.5Hz,2H),3.67(m,2H),3.37(m,6H),2.90(m,4 H),2.70(m,3H),2.56(m,3H),2.52(m,1H),2.07(m,1H),1.92(m,2H),1.16(t,J=7.5Hz,3H).MS(ESI+)m / z:907.3[M+H] + .

[0069] Example 3: Synthesis of 6-(4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amine)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazin-1-yl)-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)hexanamide (IA-3)

[0070] Compound I-9 (0.15 g, 0.27 mmol) and 6-bromo-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)hexanamide (0.14 g, 0.32 mmol) were dissolved in DMF (1 mL), DIEA (0.171 g, 1.33 mmol) was added, and the mixture was heated to 80 °C for 6 h. TLC (DCM: MeOH = 10: 1) showed that the raw materials were basically reacted, and the reaction was stopped. About 10 mL of water was added, and the mixture was extracted three times with 4 mL of EA. The organic phases were combined, dried with anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.15 g of a yellow solid with a yield of 62.1%. 1 H NMR (400MHz, DMSO-d6) δ (ppm): 11.15 (s, 1H), 10.41 (s, 1H), 9.72 (s, 1H), 8.47 (t, J = 8.8Hz, 1H), 8.05 (t, J = 7.4Hz, 1H), 7.9 6(d,J=2.1Hz,1H),7.84(t,J=7.9Hz,1H),7.72(dd,J=8.7,2.1Hz,1H),7.62(d,J=7.3Hz,1H),6.83(s,1H),5.32(s,2H),5.1 5(q,J=5.4Hz,1H),4.25(d,J=2.9Hz,2H),3.80(t,J=5.5Hz,2H),3.73(m,2H),3.36(m,6H),2.95(m,3H),2.91(m,2H),2.79 (m,2H),2.68(m,2H),2.54(m,2H),2.07(m,1H),1.66(m,4H),1.37(m,2H),1.16(t,J=7.4Hz,3H).MS(ESI+)m / z:935.3[M+H] + .

[0071] Example 4: Synthesis of 8-(4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amine)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazin-1-yl)-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)octanamide (IA-4)

[0072] Compound I-9 (0.15 g, 0.27 mmol) and 8-bromo-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)octanamide (0.15 g, 0.32 mmol) were dissolved in DMF (1.5 mL), DIEA (0.17 g, 1.33 mmol) was added, and the mixture was heated to 80 °C for 6 h. TLC (DCM: MeOH = 10: 1) showed that the raw materials were basically reacted, and the reaction was stopped. About 15 mL of water was added, and the mixture was extracted three times with 5 mL of EA. The organic phases were combined, dried with anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.15 g of a light yellow solid with a yield of 56.8%. 1 HNMR (400MHz, DMSO-d6) δ (ppm): 11.17 (s, 1H), 10.44 (s, 1H), 9.72 (s, 1H), 8.47 (t, J = 8.4Hz, 1H), 8.05 (t, J = 7.5Hz ,1H),7.97(d,J=2.1Hz,1H),7.83(t,J=8.0Hz,1H),7.72(dd,J=8.8,2.2Hz,1H),7.62(d,J=7.3Hz,1H),6.83(s,1H ),5.33(s,2H),5.15(q,J=5.4Hz,1H),4.25(d,J=2.9Hz,2H),3.80(m,4H),3.38(m,6H),2.90(m,6H),2.63(m,2H), 2.54(m,1H),2.46(m,2H),2.05(m,1H),1.65(m,4H),1.35(m,6H),1.16(t,J=7.5Hz,3H).MS(ESI+)m / z:963.3[M+H] + .

[0073] Example 5: Synthesis of 10-(4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amine)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazin-1-yl)-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)decanamide (IA-5)

[0074] Compound I-9 (0.15 g, 0.27 mmol), 10-bromo-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)decylamide (0.16 g, 0.32 mmol) were dissolved in DMF (1.5 mL), DIEA (0.17 g, 1.33 mmol) was added, and the mixture was heated to 80 °C for 6 h. TLC (DCM: MeOH = 10: 1) showed that the raw materials were basically reacted, and the reaction was stopped. About 15 mL of water was added, and the mixture was extracted three times with 5 mL of EA. The organic phases were combined, dried with anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.138 g of a light yellow solid with a yield of 52.5%. 1 H NMR (400MHz, DMSO-d6) δ (ppm): 11.15 (s, 1H), 10.36 (s, 1H), 9.69 (s, 1H), 8.48 (dd, J = 8.4, 5.4Hz, 1H), 8.06 (t, J = 7.2Hz, 1H), 7.9 6(d,J=2.1Hz,1H),7.82(t,J=8.4,1H),7.71(dd,J=8.7,2.2Hz,1H),7.60(d,J=7.3Hz,1H),6.81(m,1H),5.30(s,2H),5.14(dd,J =12.8,5.4Hz,1H),4.24(d,J=2.9Hz,2H),3.79(t,J=5.4Hz,2H),3.57(d,J=11.3Hz,2H),3.36(m,6H),2.90(m,5H),2.61(m,3H), 2.54(m,1H),2.46(m,2H),2.06(m,1H),1.63(m,2H),1.45(m,2H),1.30(m,10H),1.16(t,J=7.4Hz,3H).MS(ESI+)m / z:991.3[M+H] + .

[0075] Example 6: Synthesis of 2-(4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazin-1-yl)-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)butanamide (IA-6)

[0076] Compound I-9 (0.15 g, 0.27 mmol) and 2-bromo-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)acetamide (0.13 g, 0.32 mmol) were dissolved in DMF (1.5 mL), DIEA (0.171 g, 1.33 mmol) was added, and the reaction was carried out at room temperature for 6 h. TLC (DCM: MeOH = 10: 1) showed that the raw materials were basically reacted, and the reaction was stopped. About 15 mL of water was added, and 5 mL of EA was used for extraction three times. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.042 g of a light yellow solid with a yield of 17.2%. 1 H NMR (400MHz, CDCl3) δ (ppm): 11.14 (s, 1H), 10.53 (s, 1H), 10.38 (s, 1H), 8.32 (d, J = 1.8Hz, 1H), 8.04 (d, J = 8.4Hz ,1H),8.02(d,J=8.4Hz,1H),7.97(d,J=2.1Hz,1H),7.90(d,J=8.2Hz,1H),7.72(dd,J=8.9,2.1Hz,1H),6.83(m,1 H),5.32(s,2H),5.14(m,1H),4.25(m,2H),3.80(t,J=4.2Hz,2H),3.74(m,2H),3.29(m,1H),2.90(m,5H),2.56( m,4H),2.43(m,1H),2.06(m,1H),1.29(m,1H),1.21(m,2H),1.18(m,3H),0.84(m,1H).MS(ESI+)m / z:879.3[M+H] + .

[0077] Example 7: Synthesis of 4-(4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazin-1-yl)-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)butanamide (IA-7)

[0078] Compound I-9 (0.15 g, 0.27 mmol) and 4-bromo-N-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)butyramide (0.13 g, 0.32 mmol) were dissolved in DMF (1.5 mL), DIEA (0.17 g, 1.33 mmol) was added, and the mixture was heated to 80 °C for 6 h. TLC (DCM: MeOH = 10: 1) showed that the raw materials were basically reacted, and the reaction was stopped. About 15 mL of water was added, and the mixture was extracted three times with 5 mL of EA. The organic phases were combined, dried with anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.052 g of a light yellow solid with a yield of 20.8%. 1 H NMR (400MHz, CDCl3) δ (ppm): 11.13 (s, 1H), 10.74 (s, 1H), 10.41 (s, 1H), 8.29 (s, 1H), 8 .05(d,J=8.6Hz,1H),7.92(m,3H),7.72(d,J=8.7Hz,1H),6.83(s,1H),5.31(m,2H),5.1 2(m,1H),4.25(m,2H),3.80(t,J=5.5Hz,2H),3.64(m,2H),2.90(m,5H),2.64(m,5H),2 .05(m,1H),1.89(m,2H),1.23(m,2H),1.16(t,J=7.4Hz,3H).MS(ESI+)m / z:907.3[M+H] + .

[0079] Example 8: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-((2-((2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amine)acetyl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IB-1)

[0080] The compound (2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)glycine (0.097g, 0.29mmol) was dissolved in DMF (2mL), HATU (0.11g, 0.29mmol) and DIEA (0.21g, 1.59mmol) were added, and after stirring at room temperature for half an hour, compound I-9 (0.15g, 0.27mmol) was added to the reaction system, and the reaction was carried out at room temperature for 6h. TLC (DCM:MeOH=15:1) showed that the raw materials were basically reacted completely, and the reaction was stopped. About 15mL of water was added, and 5mL of EA was used for extraction three times. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.15g of a yellow solid with a yield of 65.3%. 1 H NMR (400MHz, DMSO-d6) δ (ppm): 11.15 (s, 1H), 10.36 (s, 1H), 8.07 (t, J = 7.8Hz, 1H), 7.97 (d, J = 2.1Hz, 1H), 7.72 (m ,1H),7.61(t,J=7.8Hz,1H),7.10(m,2H),6.83(m,1H),5.33(s,2H),5.08(m,1H),4.40(m,2H),4.24(d,J=4Hz,2H) ,4.16(d,J=16.8Hz,1H),3.95(d,J=12.3Hz,1H),3.80(t,J=5.5Hz,2H),3.54(m,1H),3.44(s,1H),3.24(m,2H),3 .00(m,2H),2.88(m,2H),2.75(m,2H),2.60(m,2H),2.05(m,1H),1.20(t,J=7.4Hz,3H).MS(ESI+)m / z:879.3[M+H] + .

[0081] Example 9: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(4-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amine)butyryl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IB-2)

[0082] The compound 4-((2-(2,6-dioxopiperidin-3-yl)-dioxoisoindolin-4-yl)amine)butyric acid (0.070 g, 0.19 mmol) was dissolved in DMF (2 mL), HATU (0.072 g, 0.19 mmol) and DIEA (0.15 g, 1.17 mmol) were added, and after stirring at room temperature for half an hour, compound I-9 (0.11 g, 0.19 mmol) was added to the reaction system, and the reaction was carried out at room temperature for 6 hours. TLC (DCM: MeOH = 15: 1) showed that the raw materials were basically reacted completely, and the reaction was stopped. About 15 mL of water was added, and 5 mL of EA was used for extraction three times. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.11 g of a yellow solid with a yield of 65.0%. 1 H NMR (400MHz, CDCl3) δ (ppm): 9.00 (s, 1H), 8.46 (d, J = 8.7Hz, 1H), 8.22 (m, 1H), 7.63 (d, J = 2.0Hz, 1H), 7.53 (m, 2H), 7.0 9(d,J=7.1Hz,1H),7.00(d,J=8.6Hz,1H),6.95(m,1H),6.33(m,1H),5.10(m,2H),4.92(m,1H),4.64(d,J=12.8Hz,1H), 4.36(q,J=2.8Hz,2H),3.91(t,J=5.4Hz,2H),3.82(m,1H),3.64(m,2H),3.40(q,J=6.6Hz,2H),3.25(m,1H),3.12(m,2 H),2.82(m,4H),2.71(m,4H),2.49(m,2H),2.13(m,1H),2.04(m,2H),1.29(t,J=7.1Hz,3H).MS(ESI+)m / z:907.3[M+H] + .

[0083] Example 10: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(6-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amine)hexanoyl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IB-3)

[0084] The compound 6-((2-(2,6-dioxopiperidin-3-yl)-dioxoisoindolin-4-yl)amine)hexanoic acid (0.12 g, 0.32 mmol) was dissolved in DMF (2 mL), and HATU (0.12 g, 0.32 mmol) and DIEA (0.21 g, 1.59 mmol) were added. After stirring at room temperature for half an hour, the compound IA-1-9 (0.15 g, 0.27 mmol) was added to the reaction system and reacted at room temperature for 6 hours. TLC (DCM: MeOH = 15: 1) showed that the raw materials were basically reacted completely, and the reaction was stopped. About 15 mL of water was added, and the mixture was extracted three times with 5 mL of EA. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.22 g of a yellow solid with a yield of 87.6%. 1 H NMR (400MHz, CDCl3) δ (ppm): 9.02 (s, 1H), 8.47 (d, J = 8.7Hz, 1H), 8.17 (s, 1H), 7.63 (d, J = 2.0Hz, 1H), 7.51 (m, 2H), 7.07 (d, J = 7 .0Hz,1H),6.96(m,1H),6.89(d,J=8.5Hz,1H),6.23(t,J=5.6Hz,1H),5.11(d,J=6.1Hz,2H),4.90(m,1H),4.64(d,J=12.3Hz,1H ),4.36(d,J=2.9Hz,2H),3.91(t,J=5.4Hz,2H),3.84(d,J=13.0Hz,1H),3.71(m,1H),3.59(m,1H),3.27(m,3H),3.15(m,2H),2 .83(m,4H),2.71(m,4H),2.39(m,2H),2.13(m,1H),1.74(m,4H),1.49(m,2H),1.30(t,J=7.5Hz,3H).MS(ESI+)m / z:935.3[M+H] + .

[0085] Example 11: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(8-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amine)octanoyl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IB-4)

[0086] The compound 8-((2-(2,6-dioxopiperidin-3-yl)-dioxoisoindolin-4-yl)amine)octanoic acid (0.13 g, 0.32 mmol) was dissolved in DMF (2 mL), HATU (0.12 g, 0.32 mmol) and DIEA (0.21 g, 1.59 mmol) were added, and after stirring at room temperature for half an hour, compound I-9 (0.150 g, 0.265 mmol) was added to the reaction system, and the reaction was carried out at room temperature for 6 hours. TLC (DCM: MeOH = 15: 1) showed that the raw materials were basically reacted completely, and the reaction was stopped. About 15 mL of water was added, and 5 mL of EA was used for extraction three times. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.13 g of a yellow solid with a yield of 49.8%. 1 H NMR (400MHz, CDCl3) δ (ppm): 9.03 (s, 1H), 8.46 (d, J = 8.7Hz, 1H), 8.21 (s, 1H), 7.63 (d, J = 2.0Hz, 1H), 7.51 (m, 2H), 7 .06(d,J=7.0Hz,1H),6.95(m,1H),6.88(d,J=8.5Hz,1H),6.22(m,1H),5.11(m,2H),4.89(m,1H),4.64(d,J=12.5Hz ,1H),4.36(q,J=2.8Hz,2H),3.82(m,3H),3.70(t,J=11.3Hz,1H),3.59(t,J=10.9Hz,1H),3.26(m,3H),3.14(m,2H) ,2.79(m,8H),2.39(m,2H),2.12(m,1H),1.66(m,4H),1.39(m,6H),1.30(t,J=7.3Hz,3H).MS(ESI+)m / z:963.3[M+H] + .

[0087] Example 12: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(11-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amine)undecanoyl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IB-5)

[0088] Compound 11-((2-(2,6-dioxopiperidin-3-yl)-dioxoisoindolin-4-yl)amine)undecanoic acid (0.14 g, 0.32 mmol) was dissolved in DMF (2 mL), HATU (0.12 g, 0.32 mmol) and DIEA (0.21 g, 1.59 mmol) were added, and after stirring at room temperature for half an hour, compound I-9 (0.15 g, 0.27 mmol) was added to the reaction system, and the reaction was carried out at room temperature for 6 hours. TLC (DCM: MeOH = 15: 1) showed that the raw materials were basically reacted completely, and the reaction was stopped. About 15 mL of water was added, and 5 mL of EA was used for extraction three times. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.18 g of a yellow solid with a yield of 66.8%. 1 H NMR (400MHz, CDCl3) δ (ppm): 9.03 (s, 1H), 8.46 (d, J = 8.7Hz, 1H), 8.25 (d, J = 7.3Hz, 1H), 7.63 (d, J = 2.0Hz, 1H), 7.50 (m, 2H), 7. 06(d,J=7.1Hz,1H),6.96(m,1H),6.87(d,J=8.6Hz,1H),6.22(m,1H),5.11(d,J=5.7Hz,2H),4.90(m,1H),4.64(d,J=12.7Hz,1H ),4.35(q,J=2.8Hz,2H),3.90(t,J=5.4Hz,2H),3.85(d,J=12.7Hz,1H),3.70(t,J=11.3Hz,1H),3.59(t,J=11.5Hz,1H),3.25( m,3H),3.14(m,2H),2.73(m,8H),2.36(m,2H),2.12(m,1H),1.66(m,4H),1.40(m,2H),1.32(m,3H).MS(ESI+)m / z:1005.3[M+H] + .

[0089] Example 13: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)piperidin-4-yl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IC-1)

[0090] The compound 1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)piperidine-4-carboxylic acid (0.12 g, 0.32 mmol) was dissolved in DMF (2 mL), HATU (0.12 g, 0.32 mmol) and DIEA (0.21 g, 1.59 mmol) were added, and after stirring at room temperature for half an hour, compound I-9 (0.15 g, 0.27 mmol) was added to the reaction system, and the reaction was carried out at room temperature for 6 hours. TLC (DCM: MeOH = 15: 1) showed that the raw materials were basically reacted completely, and the reaction was stopped. About 15 mL of water was added, and 5 mL of EA was used for extraction three times. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.23 g of a yellow solid with a yield of 92.2%. 1 H NMR (400MHz, CDCl3) δ (ppm): 9.03 (s, 1H), 8.47 (d, J = 8.7Hz, 1H), 8.20 (m, 1H), 7.63 (d, J = 2.0Hz, 1 H),7.53(m,2H),7.35(d,J=7.3Hz,1H),7.18(d,J=8.3Hz,1H),6.95(m,1H),5.13(m,2H),5.95(m, 1H),4.66(m,1H),4.36(m,2H),3.91(t,J=5.3Hz,3H),3.78(m,3H),3.60(m,1H),3.32(m,1H),3.0 9(m,4H),2.79(m,9H),2.10(m,3H),1.87(m,2H),1.32(d,J=7.0Hz,3H).MS(ESI+)m / z:933.3[M+H] + .

[0091] Example 14: Synthesis of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)piperidin-4-yl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (IC-2)

[0092] The compound 1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)piperidine-4-carboxylic acid (0.12 g, 0.32 mmol) was dissolved in DMF (2 mL), HATU (0.12 g, 0.32 mmol) and DIEA (0.21 g, 1.59 mmol) were added, and after stirring at room temperature for half an hour, compound I-9 (0.15 g, 0.27 mmol) was added to the reaction system, and the reaction was carried out at room temperature for 6 hours. TLC (DCM: MeOH = 15: 1) showed that the raw materials were basically reacted completely, and the reaction was stopped. About 15 mL of water was added, and 5 mL of EA was used for extraction three times. The organic phases were combined and dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was sand-coated to obtain 0.21 g of a yellow solid with a yield of 85.3%. 1 H NMR (400MHz, CDCl3) δ (ppm): 9.03 (s, 1H), 8.42 (m, 2H), 7.63 (m, 2H), 7.51 (dd, J = 8.8, 2.1Hz, 1H), 7.25(d,J=2.3Hz,1H),7.03(dd,J=8.6,2.3Hz,1H),6.93(m,1H),5.13(m,2H),4.93(m,1H),4.63(d ,J=12.8Hz,1H),4.34(m,2H),3.95(m,5H),3.73(m,1H),3.58(m,1H),3.33(t,J=11.9Hz,1H),3.0 8(m,4H),2.79(m,9H),2.12(m,1H),1.95(m,4H),1.30(t,J=7.6Hz,3H).MS(ESI+)m / z:933.3[M+H] + .

[0093] Example 15: Evaluation of the inhibitory activity of compounds on tumor cell proliferation

[0094] 1. Experimental methods

[0095] (1) Human colon adenocarcinoma cell line (SW48) cultured to the logarithmic growth phase was plated into a 96-well plate at a predetermined density in a medium containing fetal bovine serum;

[0096] (2) cells were treated with compound or vehicle (DMSO) 24 h later, and day 0 plates were harvested for analysis;

[0097] (3) After drug administration, the 96-well plate was placed in a 37°C, 5% CO2 constant temperature incubator. After 5 days, 20 μL of 1.0% MTT thiazolyl blue solution was added to each well;

[0098] (4) Continue to place in a constant temperature incubator. After 4 hours, remove the supernatant culture medium by aspiration, add 150 μL DMSO to each well, and place on a decolorizing shaker to mix until the crystals dissolve;

[0099] (5) Measure the absorbance at 570 nm using a multifunctional microplate reader and calculate the IC according to the modified Koch method 50 value:

[0100] lgIC 50 =Xm-I[P-(3-Pm-Pn) / 4].

[0101] 2. Experimental results

[0102] The specific results are shown in Table 1. 50 <1μM (denoted as: A); IC 50 =1~10μM (denoted as: B); IC 50 >10μM (denoted as: C).

[0103] Table 1 Inhibitory activity of compounds on SW48 cell line

[0104] Number <![CDATA[IC 50 ]]> Number <![CDATA[IC 50 ]]> I-A-1 C I-B-1 C I-A-2 B I-B-2 C I-A-3 C I-B-3 C I-A-4 B I-B-4 C I-A-5 C I-B-5 C I-A-6 B I-C-1 C I-A-7 B I-C-2 C

[0105] The results in Table 1 show that the compounds of the present invention exhibit good inhibitory activity against human colon adenocarcinoma SW48 cell line, and the inhibitory IC 50 The concentration value reaches micromolar level.

Claims

1. A compound represented by general formula (I) or a pharmaceutically acceptable salt thereof, in: R 1 Selected from R 2 for R 3 is selected from H, OCH3, OH, CN or halogen, R 4 is selected from H, F, Cl, Br, I or substituted C1-C4 alkyl, R 5 is selected from H, F, Cl, Br, I, SF3, C(O)H or substituted C1-C4 alkyl, substituted C1-C4 alkenyl, substituted C1-C4 alkynyl, substituted C3-C5 cycloalkyl or OCF3, R 6 is selected from H, F, Cl, Br, I, OH, CN, substituted C1-C4 alkyl, substituted C3-C5 cycloalkyl or substituted C1-C4 alkoxy, wherein the substituent is selected from at least one H, F, Cl, Br or I; X is selected from CR 7 or N, R 7 is selected from H, F, Cl, Br or I; A is selected from n is an integer selected from 1 to 11, m is an integer selected from 1 to 3, R 8 is selected from H, F, Cl, Br, I or OH, Y, Z, K are independently selected from CH, N or CR 9 , R 9 Selected from H, F, Cl, Br, I, OCH3, OH, CN, CF3 or OCF3.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: R 1 Selected from 3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: R 2 Selected from 4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: A is selected from n is selected from integers of 1-9, and m is selected from integers of 1-3.

5. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: The compound is selected from any one of the following compounds:

6. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: The pharmaceutically acceptable salt is an acid addition salt formed by the compound and any one of the following acids: hydrogen chloride, hydrogen bromide, sulfuric acid, carbonic acid, oxalic acid, citric acid, malic acid, succinic acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid or ferulic acid.

7. A method for preparing the compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Choose from any of the following methods: Method 1: When A is When the compound I-1 is acylated, iodinated, substituted, coupled, deprotected and substituted, the compound IA is obtained; Method 2: When A is When the compound I-9 is condensed with the compound I-11, the compound IB is obtained; Method 3: When A is When the compound I-9 is condensed with the compound I-12, the compound IC is obtained; Among them, R 1 , R 2 , n is defined as in claim 1; The compounds IA, IB and IC prepared by the above methods are salified with a pharmaceutically acceptable acid to obtain the pharmaceutically acceptable salts.

8. A pharmaceutical composition, characterized in that Comprising the compound according to claim 1 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

9. Use of the compound according to claim 1 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to claim 8 in the preparation of a WRN protein degrader.

10. The use according to claim 9, characterized in that: The medicine is a medicine for treating tumors.

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