DNA-PK / PARP1 double-target inhibitor as well as preparation method and application thereof

Abstract

The invention discloses a DNA-PK / PARP1 double-target inhibitor as well as a preparation method and application of the DNA-PK / PARP1 double-target inhibitor. The DNA-PK / PARP1 double-target inhibitor disclosed by the invention is a compound of which the structure is # imgabs0 #. The compound disclosed by the invention is novel in structure, can selectively act on DNA-PK and PARP1, and shows good DNA-PK enzyme and PARP1 protein inhibitory activity; the preparation method of the compound is mature in process, safe and pollution-free, and has the advantage of industrial popularization; the compound can be widely applied to preparation of a DNA-PK inhibitor and / or a PARP1 inhibitor and preparation of drugs for treating and / or preventing cancers.

Description

Technical Field

[0001] The present invention belongs to the pharmaceutical field, and specifically relates to a DNA-PK / PARP1 dual-target inhibitor and a preparation method and application thereof. Background Art

[0002] Cancer seriously threatens human life and health and is the second leading cause of death. Effective treatment options for cancer have become the focus of scientists. In the past decade, a large number of molecular targeted drugs have been launched for cancer treatment. However, since cancer is a multi-gene related and highly complex disease, single-target drugs often have limited efficacy in treating cancer and are only effective for some patients. In response to the above problems, combination drug regimens are often used in clinical practice. Although the combination of drugs with different mechanisms of action has made up for the shortcomings of single-target drugs to a certain extent, it has also caused a series of problems, such as the superposition of toxic and side effects and unpredictable PK / PD properties.

[0003] In recent years, significant progress has been made in the field of tumor treatment, and one of the aspects that has attracted much attention is targeted therapy. DNA-PK (DNA-dependent protein kinase): is a protein kinase that plays an important role in the repair of double-strand breaks in DNA. When DNA is damaged, DNA-PK is rapidly recruited to the site of damage and initiates the repair mechanism. PARP1 (Poly (ADP-ribose) polymerase 1): is a poly ADP-ribose polymerase that plays a key role in DNA damage response. When DNA is damaged, PARP1 is activated, catalyzing the production of poly ADP-ribose, and then recruiting other repair proteins to participate in the DNA repair process.

[0004] The genome of tumor cells is usually unstable, and DNA damage occurs frequently. In order to survive, tumor cells activate various DNA repair mechanisms, including DNA-PK and PARP1-mediated repair. If DNA-PK and PARP1 are inhibited at the same time, the DNA repair ability of tumor cells will be further damaged, leading to the accumulation of DNA damage and ultimately inducing tumor cell death. DNA-PK / PAPR1 are both involved in the self-damage repair process of cancer cells. In particular, simultaneous inhibition of DNA-PK / PARP1 has potential synergistic anti-cancer activity in breast cancer cells with BRCA mutations.

[0005] DNA-PK / PARP1 dual-target inhibitors are a promising direction in the field of tumor treatment. By simultaneously inhibiting two key DNA repair proteins, tumor cells can be effectively killed, but the research and development of related drugs still needs to be improved. Summary of the invention

[0006] In order to overcome the problems existing in the above-mentioned prior art, one of the purposes of the present invention is to provide a compound; the second purpose of the present invention is to provide a method for preparing the above-mentioned compound; the third purpose of the present invention is to provide a pharmaceutical composition; the fourth purpose of the present invention is to provide the application of the above-mentioned compound. In order to achieve the above-mentioned purposes, the technical solutions adopted by the present invention are:

[0007] The first aspect of the present invention provides a compound, the structure of which is shown in Formula 1:

[0008]

[0009] In Formula 1, R1 is selected from any one of an alkyl group, an aryl group, and a heterocycle, and R2 is selected from any one of an alkyl group, an aryl group, and a heterocycle.

[0010] Preferably, in Formula 1, R1 is selected from any one of an alkyl group and an aryl group, and R2 is selected from any one of an alkyl group and a heterocycle.

[0011] Preferably, the compound includes any one of the following compounds (1) to (10):

[0012]

[0013] Preferably, the compound is a dual-target inhibitor of DNA-PK and PARP1.

[0014] The second aspect of the present invention provides a method for preparing the compound according to the first aspect of the present invention, comprising the following steps:

[0015] 7) Mixing the compound represented by formula I with 1-boc-piperazine, removing the protecting group, and then reacting with an amino acid to obtain the compound represented by formula II;

[0016]

[0017] Wherein R2 is selected from any one of an alkyl group, an aryl group, and a heterocycle;

[0018] 8) mixing the compound represented by formula III with methyl bromoacetate, and then reducing the mixture to obtain the compound represented by formula IV;

[0019]

[0020] Wherein R1 is selected from any one of an alkyl group, an aryl group, and a heterocycle;

[0021] 9) reacting the compound represented by formula V with 4-aminotetrahydropyran hydrochloride, followed by hydrolysis to obtain the compound represented by formula VI;

[0022]

[0023] 10) reacting the compound represented by formula VI with diphenylphosphoryl azide, and then reacting with iodomethane to obtain the compound represented by formula VII;

[0024]

[0025] 11) reacting the compound represented by formula VII with the compound represented by formula IV, followed by hydrolysis to obtain the compound represented by formula VIII:

[0026]

[0027] 12) reacting the compound represented by formula II with the compound represented by formula VIII to obtain the target compound.

[0028] Preferably, in step 1), the molar ratio of the compound represented by formula I to 1-boc-piperazine is 1:0.8-1.2, more preferably 1:1.

[0029] Preferably, in the step 2), the molar ratio of the compound represented by formula III to methyl bromoate is 1:1.1-1.5, more preferably 1:1.39.

[0030] Preferably, in step 3), the molar ratio of the compound represented by formula V to 4-aminotetrahydropyran hydrochloride is 1:0.8-1.2, more preferably 1:1.

[0031] Preferably, in step 4), the molar ratio of the compound represented by formula VI to diphenylphosphoryl azide is 1:0.8-1.2, more preferably 1:1.

[0032] Preferably, in step 5), the molar ratio of the compound represented by formula VII to the compound represented by formula IV is 1:1 to 1.4, more preferably 1:1.2.

[0033] Preferably, in step 6), the molar ratio of the compound represented by formula II to the compound represented by formula VIII is 1:1 to 1.4, more preferably 1:1.2.

[0034] The third invention of the present invention provides a pharmaceutical composition, which includes the compound described in the first aspect of the present invention or its stereoisomer, solvate, prodrug, metabolite, pharmaceutically acceptable salt or cocrystal.

[0035] The fourth aspect of the present invention provides the use of the compound according to the first aspect of the present invention or its stereoisomers, solvates, prodrugs, metabolites, pharmaceutically acceptable salts or cocrystals in the preparation of drugs for treating and / or preventing and / or delaying and / or assisting in the treatment of cancer.

[0036] Preferably, the cancer includes at least one of breast cancer, melanoma, and leukemia.

[0037] Preferably, the cancer drug comprises a DNA-PK inhibitor and / or a PARP1 inhibitor.

[0038] Compared with the prior art, the present invention has the following advantages and effects:

[0039] The compound disclosed in the present invention has a novel structure, can selectively act on DNA-PK and PARP1, and exhibits good DNA-PK enzyme and PARP1 protein inhibitory activity; the preparation method of the compound is mature, safe and pollution-free, and has the advantage of being industrially popularizable; the compound can be widely used in the preparation of DNA-PK inhibitors and / or PARP1 inhibitors, as well as the preparation of drugs for treating and / or preventing cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 is a chemical structure diagram of the compound of the present invention;

[0041] Figure 2 The following is a reaction scheme for preparing the compounds of the present invention. DETAILED DESCRIPTION

[0042] The specific implementation of the present invention is further described below in conjunction with examples, but the implementation and protection of the present invention are not limited thereto. It should be noted that if there are processes that are not particularly described in detail below, they can be implemented or understood by those skilled in the art with reference to the prior art. If the manufacturer of the reagents or instruments used is not indicated, they are deemed to be conventional products that can be purchased commercially.

[0043] The chemical structure of the compound of the present invention is as follows Figure 1 shown.

[0044] The preparation reaction route of the compound of the present invention is as follows Figure 2 According to the above reaction route, the preparation of the compound includes the following steps:

[0045] Route A:

[0046]

[0047] According to the above reaction route, the preparation of the compound includes the following steps:

[0048] 1. Dissolve 4-methyl-3-nitrophenol (A) in N,N-dimethylformamide solution, add 2.5 molar times of potassium carbonate according to the molar amount of 4-methyl-3-nitrophenol, and react at 80°C for 30 minutes. Then add 1.39 molar times of compound B according to the molar amount of 4-methyl-3-nitrophenol, and react at 80°C for 30 minutes. After the reaction is complete as monitored by thin layer chromatography, the reaction solution is extracted with ethyl acetate, the organic phase is dried with saturated brine and anhydrous sodium sulfate, and vacuum desolventized to obtain intermediate C. The specific reaction formula is as follows:

[0049]

[0050] 2. Dissolve the intermediate C obtained in step 1 in methanol, add 2 spoons of Raney nickel under ice bath conditions, and add 2.5 molar times of hydrazine hydrate according to the molar amount of the intermediate. React at room temperature for 5 minutes. After the reaction is complete as monitored by a thin layer chromatography plate, remove the solvent, separate and purify by column chromatography to obtain intermediate D. The specific reaction formula is as follows:

[0051]

[0052] Route B:

[0053]

[0054] According to the above reaction route, the preparation of the compound includes the following steps:

[0055] 1. Dissolve 2,4-dichloro-5-pyrimidinecarboxylic acid ethyl ester (E) in acetonitrile, add 1 mole of 4-aminotetrahydropyran hydrochloride (F) according to the molar amount of 2,4-dichloro-5-pyrimidinecarboxylic acid ethyl ester, add 2.5 mole of potassium carbonate according to the molar amount of 2,4-dichloro-5-pyrimidinecarboxylic acid ethyl ester, react at room temperature for 10 hours, and monitor the reaction on a thin layer chromatography plate until it is complete. Remove the solvent, separate and purify by column chromatography to obtain intermediate G. The specific reaction formula is as follows:

[0056]

[0057] 2. Take the intermediate G obtained in step 1 and dissolve it in tetrahydrofuran, add 2.5 equivalents of sodium hydroxide solution, place it in a 60°C oil bath for reaction for 1 hour, and stop the reaction after the reaction is complete as monitored by a thin layer chromatography plate. The reaction solution is distilled under reduced pressure to remove the solvent to obtain a crude product. After adding water to the crude product, adjust the pH to soft acidity (PH≈5-6) with dilute hydrochloric acid under ice bath conditions. Precipitate and filter out the white solid, which is the intermediate H. The specific reaction formula is as follows:

[0058]

[0059] 3. Take the intermediate H obtained in step 2 and dissolve it in DMA, add 1 mol of diphenylphosphoryl azide and 1 mol of triethylamine, react at room temperature for one hour, transfer to a 120°C oil bath to react for 12 hours, and stop the reaction after the reaction is complete as monitored by a thin layer chromatography plate, add ice water and stir rapidly to precipitate, and filter out a light yellow solid, which is the intermediate I. The specific reaction formula is as follows:

[0060]

[0061] 4. Take the intermediate I obtained in step 3 and dissolve it in anhydrous tetrahydrofuran. Under ice bath conditions, add 2 molar times of sodium hydride. After removing the ice, place it at room temperature for 30 minutes, add iodomethane, react at room temperature for 8 hours, and stop the reaction after the reaction is complete as monitored by a thin layer chromatography plate. The reaction solution is extracted with ethyl acetate, and the organic phase is dried with saturated brine and anhydrous sodium sulfate, desolventized, and separated and purified by column chromatography to obtain intermediate J. The specific reaction formula is as follows:

[0062]

[0063] Route C:

[0064]

[0065] 1. Dissolve the amino acid (M) in tetrahydrofuran, add sodium hydroxide (2 mol), add water (THF: water = 1:1) and dicarbonic acid acetic anhydride in an ice bath, react at room temperature for 8 hours after removing the ice, and stop the reaction after monitoring the reaction completion with a thin layer chromatography plate. Extract the reaction solution with petroleum ether, adjust the pH of the aqueous phase to soft acidity (PH ≈ 5-6) with dilute hydrochloric acid under ice bath conditions, extract with ethyl acetate, take the organic phase, dry it with saturated brine and anhydrous sodium sulfate, and distill under reduced pressure to obtain intermediate N. The specific reaction formula is as follows:

[0066]

[0067] 2. Dissolve 2-fluoro-5-((4-oxy-3,4-dihydrophthalic acid-1-yl)methyl)benzoic acid (O) in DMF, add 1 mole of piperazine-1-carboxylic acid tert-butyl acetate (P) according to the molar amount of compound O, add 1.5 mole of HATU, 2.5 mole of N,N-diisopropylethylamine, react at room temperature for 3 hours, stop the reaction after the reaction is complete as monitored by thin layer chromatography, add ice water, beat, and filter under reduced pressure to obtain a flesh-pink intermediate Q. The specific reaction formula is as follows:

[0068]

[0069] 3. Take the intermediate Q obtained in step 2 and dissolve it in dichloromethane, add an appropriate amount of HCl·1,4-dioxane at room temperature, monitor the reaction on a thin layer chromatography plate, and filter the precipitated solid under reduced pressure to obtain a pink intermediate R. The specific reaction formula is as follows:

[0070]

[0071] 4. Take the intermediate N obtained in step 1 and the intermediate R obtained in step 3, add 1 mole of intermediate R, 1.5 mole of HATU, and 2.5 mole of N,N-diisopropylethylamine according to the molar amount of intermediate N, and react at room temperature for 3 hours. After the reaction is complete as monitored by thin layer chromatography, stop the reaction, extract the reaction solution with ethyl acetate, take the organic phase, dry it with saturated brine and anhydrous sodium sulfate, and distill it under reduced pressure to obtain a white intermediate S. The specific reaction formula is as follows:

[0072]

[0073] 5. Take the intermediate S obtained in step 4 and dissolve it in dichloromethane, add an appropriate amount of HCl·1,4-dioxane at room temperature, monitor the reaction on a thin layer chromatography plate, and after the reaction is complete, decompress and spin-dry the precipitated solid to obtain the intermediate T. The specific reaction formula is as follows:

[0074]

[0075] Route D:

[0076]

[0077] 1. Dissolve the intermediate J in 1,4-dioxane solution, add 1.2 mole times of intermediate D according to the mole of intermediate J, add 0.1 mole times of tris(dibenzylideneacetone)dipalladium, 0.3 mole times of 4,5-bis(diphenylphosphine)-9,9-dimethylxanthene and 2.5 mole times of cesium carbonate according to the mole of intermediate J, react at 105°C for 12 hours under nitrogen environment, and after monitoring the completion of the reaction on a thin layer chromatography plate, extract the reaction solution with dichloromethane, take the organic phase, dry it with saturated brine and anhydrous sodium sulfate, desolventize, separate and purify it by column chromatography to obtain intermediate K. The specific reaction formula is as follows:

[0078]

[0079] 2. Dissolve the intermediate K in a mixed solution of methanol and water, add 2.5 molar times of sodium hydroxide according to the molar amount of the intermediate K, react at room temperature for 30 minutes, and stop the reaction after the reaction is complete as monitored by a thin layer chromatography plate. Distill the reaction solution under reduced pressure to remove the solvent to obtain a crude product. After adding water to the crude product, adjust the pH to soft acidity (PH≈5-6) with dilute hydrochloric acid under ice bath conditions. Precipitate and filter out the white solid, which is the intermediate L. The specific reaction formula is as follows:

[0080]

[0081] 3. Take the intermediate L obtained in step 2 and add an appropriate amount of DMF as a solvent, then add 1.2 molar times of intermediate T according to the molar amount of intermediate L, and add 1.2 molar times of EDCI, 1.5 molar times of HOBT and 2.5 molar times of triethylamine according to the molar amount of intermediate L. Under room temperature conditions, react for 2 hours. After the reaction is complete as monitored by a thin layer chromatography plate, add ice water, beat and filter to obtain the final product. The specific reaction formula is as follows:

[0082]

[0083] Example 1

[0084] The specific preparation steps of the compound of this example are as follows:

[0085] 1) Synthesis of intermediate 5-(4-methyl-3-nitrophenoxy) methyl pentanoate (C): 1.0 g of 4-methyl-3-nitrophenol (A) (6.53 mmol) was dissolved in 10 ml of N,N-dimethylformamide solution, 2.26 g of potassium carbonate (16.33 mmol) was added, and the mixture was reacted at 80°C for 30 minutes, and 1.64 g of methyl 5-bromopentanoate (B) (9.08 mmol) was added and the mixture was reacted for 30 minutes. After the reaction was completed as monitored by thin layer chromatography, the reaction solution was extracted with ethyl acetate (30 mL × 3), allowed to stand for separation, and the organic phase was washed with water (5 mL × 1) and saturated brine (5 mL × 3) in turn, and then dried with anhydrous sodium sulfate, filtered, and ethyl acetate was removed under reduced pressure to obtain 1.55 g of solid, with a yield of 93.9%. The obtained solid was identified by nuclear magnetic resonance method, and the result showed that the solid was methyl 5-(4-methyl-3-nitrophenoxy) methyl pentanoate (C). Its structural formula is:

[0086]

[0087] 2) Synthesis of intermediate 5-(4-methyl-3-aminophenoxy) methyl pentanoate (D): Take 1.55g of 5-(4-methyl-3-nitrophenoxy) butyric acid methyl ester (C) in a reaction bottle, add 15mL of methanol to dissolve, and add 2 spoons of Raney nickel and 957.47mg of 80% Raney nickel (0.147mmol) in turn under ice bath conditions. React for 30 minutes at room temperature. After the reaction is complete as monitored by a thin layer chromatography plate, the reaction solution is decompressed to remove methanol to obtain a solid. The solid is subjected to column chromatography with dichloromethane: methanol to obtain 1.2g of a white solid with a yield of 87.6%. The white solid obtained by column chromatography is identified by nuclear magnetic resonance, and the result shows that the white solid is 5-(4-methyl-3-aminophenoxy) methyl pentanoate (D). Its structural formula is:

[0088]

[0089] 3) Synthesis of ethyl 2-chloro-4-((tetrahydro-2h-pyran-4-yl)amino)pyrimidine-5-carboxylate (G): Take 5g (22.62mol) of ethyl 2,4-dichloro-5-pyrimidinecarboxylate (E) in a reaction bottle, add 3.11g (22.62mol) of 4-aminotetrahydropyran hydrochloride (F), add 7.82g (56.55mol) of potassium carbonate, and add 30ml of acetonitrile to dissolve. React at room temperature for 10 hours. After the reaction is complete as monitored by a thin layer chromatography plate, wash the insoluble solid with 25ml of dichloromethane and ethanol respectively, remove the liquid, and obtain a solid. The solid is subjected to column chromatography with dichloromethane: petroleum ether to obtain 6g of a white solid with a yield of 87.6%. The white solid obtained by column chromatography is identified by nuclear magnetic resonance method, and the result shows that the white solid is 5-(4-methyl-3-aminophenoxy) pentanoic acid methyl ester (G). Its structural formula is:

[0090]

[0091] 4) Synthesis of 2-chloro-4-((tetrahydro-2h-pyran-4-yl)amino)pyrimidine-5-carboxylic acid (H): 6 g (21 mol) of ethyl 2-chloro-4-((tetrahydro-2h-pyran-4-yl)amino)pyrimidine-5-carboxylate (G) was dissolved in tetrahydrofuran, 1.68 g (41.07 mol) of sodium hydroxide was dissolved in 5 ml of water, and then added to the reaction solution, the reaction bottle was placed in a 60°C oil bath for 1 hour, and the reaction was stopped after the reaction was complete as monitored by a thin layer chromatography plate. The reaction solution was distilled under reduced pressure to remove the solvent to obtain a crude product. After adding water to the crude product, the pH was adjusted to soft acidity (PH≈5-6) with dilute hydrochloric acid under ice bath conditions. The precipitate was separated and 4.4 g of a white solid was filtered out with a yield of 81%. The solid was identified by nuclear magnetic resonance, and the result showed that the solid was 2-chloro-4-((tetrahydro-2h-pyran-4-yl)amino)pyrimidine-5-carboxylic acid (H). Its structural formula is:

[0092]

[0093] 5) Synthesis of 2-chloro-9-(tetrahydro-2H-pyran-4-yl)-7,9-dihydro-8H-purin-8-one (I): 2 g (7.76 mol) of 2-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)pyrimidine-5-carboxylic acid (H) was dissolved in 2 ml of DMA, and 1.67 ml (7.76 mol) of diphenylphosphoryl azide and 1.08 ml (7.76 mol) of triethylamine were added. The mixture was reacted at room temperature for one hour, and then in an oil bath at 120°C for 12 hours. After the reaction was complete as monitored by a thin layer chromatography plate, the reaction was stopped, ice water was added for rapid stirring, and a precipitate was separated. 0.92 g of a light yellow solid was filtered out with suction, and the yield was 46.54%. The solid was identified by nuclear magnetic resonance, and the result showed that the solid was 2-chloro-9-(tetrahydro-2H-pyran-4-yl)-7,9-dihydro-8H-purin-8-one (I). Its structural formula is:

[0094]

[0095] 6) Synthesis of 2-chloro-7-methyl-9-(tetrahydro-2H-pyran-4-yl)-7,9-dihydro-8H-purin-8-one (J): 920 mg (3.61 mol) of 2-chloro-9-(tetrahydro-2H-pyran-4-yl)-7,9-dihydro-8H-purin-8-one (I) was dissolved in anhydrous tetrahydrofuran. 362 mg (9.03 mol) of sodium hydride was added under ice bath condition. After removing ice, the mixture was allowed to react at room temperature for 30 min. 563 μl (9.03 mol) of iodomethane was added and the reaction was continued at room temperature for 8 hours. After monitoring the completion of the reaction on a thin layer chromatography plate, the reaction was stopped. The reaction solution was extracted with ethyl acetate, and the organic phase was dried with saturated brine and anhydrous sodium sulfate, desolventized, and purified by column chromatography to obtain 673 mg of a white solid with a yield of 69.33%. The obtained solid was identified by nuclear magnetic resonance, and the results showed that the solid was 2-chloro-7-methyl-9-(tetrahydro-2H-pyran-4-yl)-7,9-dihydro-8H-purine-8-one (J). Its structural formula is:

[0096]

[0097] 7) Synthesis of 4-((tert-butyloxycarbonyl)amino)butyric acid (N): 500 mg (4.85 mol) of 4-aminobutyric acid (M) was dissolved in 5 ml of tetrahydrofuran, 484 mg (12.12 mol) of sodium hydroxide was added, 5 ml of water was added under ice bath, 1.17 ml (5.09 mol) of dicarbonic acetic anhydride was slowly added dropwise, and the reaction was allowed to react for 8 hours. The thin layer chromatography plate was developed with ninhydrin. After monitoring the completion of the reaction, the reaction solution was extracted with ethyl acetate (30 mL×1), allowed to stand for separation, the aqueous phase was adjusted to acidic, and then extracted with ethyl acetate (30 mL×3), the organic phase was washed with water (5 mL×1) and saturated brine (5 mL×3), then dried over anhydrous sodium sulfate, filtered, and ethyl acetate was removed under reduced pressure to obtain 830 mg of a white solid with a yield of 84.23%. The white solid was identified by nuclear magnetic resonance method, and the results showed that it was 4-((tert-butyloxycarbonyl)amino)butyric acid (N). Its structural formula is:

[0098]

[0099] 8) Synthesis of tert-butyl 4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazine-1-carboxylate (Q): 2 g (6.71 mol) of 2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoic acid (O) was dissolved in DMF, 1.25 g (6.71 mol) of tert-butyl piperazine-1-carboxylate acetate (P) was added, 3.82 g (10.06 mol) of HATU, 2 .34ml (13.41mol) N,N-diisopropylethylamine, react at room temperature for 3 hours. After the reaction is complete as monitored by thin layer chromatography, the reaction is stopped, ice water is added, slurry is beaten, and vacuum filtration is performed to obtain 2.7g of flesh-pink solid with a yield of 86.31%. The obtained solid is identified by nuclear magnetic resonance method, and the result shows that the solid is tert-butyl 4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazine-1-carboxylate (Q). Its structural formula is:

[0100]

[0101] 9) Synthesis of 4-(4-fluoro-3-(piperazine-1-carbonyl)benzyl)phthalazin-1(2H)-one (R): 2.7 g of tert-butyl 4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazine-1-carboxylate (Q) was dissolved in 6 ml of dichloromethane, and 3 ml of HCl·1,4-dioxane was added at room temperature. After the reaction was completed as monitored by thin layer chromatography, the precipitated solid was filtered under reduced pressure to obtain 1.6 g of a pink solid with a yield of 75.45%. The obtained solid was identified by nuclear magnetic resonance method, and the result showed that the solid was 4-(4-fluoro-3-(piperazine-1-carbonyl)benzyl)phthalazin-1(2H)-one (R). Its structural formula is:

[0102]

[0103] 10) Synthesis of tert-butyl (4-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-4-oxobutyl)carbamate (S): 830 mg (4.08 mol) of 4-((tert-butyloxycarbonyl)amino)butyric acid (N) was added with 1.5 g (4.08 mol) of 4-(4-fluoro-3-(piperazine-1-carbonyl)benzyl)phthalazine -1(2H)-ketone (R), add 2.33g (6.13mol) of HATU and 1.78 (10.21mol) of N,N-diisopropylethylamine, react at room temperature for 3 hours, and stop the reaction after the reaction is complete as monitored by thin layer chromatography. The reaction solution is extracted with ethyl acetate, and the organic phase is dried with saturated brine and anhydrous sodium sulfate, and distilled under reduced pressure to obtain 1.85g of a white intermediate with a yield of 82.12%. The obtained solid was identified by nuclear magnetic resonance method, and the result showed that the solid was tert-butyl (4-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-4-oxobutyl)carbamate (S). Its structural formula is:

[0104]

[0105] 11) Synthesis of 4-(3-(4-(4-aminobutyryl)piperazine-1-carbonyl)-4-fluorobenzyl)phthalazin-1(2H)-one (T): 1.85 g (3.35 mol) of tert-butyl (4-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-4-oxobutyl)carbamate (S) was dissolved in 4 ml of dichloromethane, and 4 ml of HCl·1,4-dioxane was added at room temperature. After the reaction was completed as monitored by thin layer chromatography, the precipitated solid was filtered under reduced pressure at low temperature to obtain 1.1 g of a pink solid with a yield of 72.64%. The obtained solid was identified by nuclear magnetic resonance method, and the result showed that the solid was 4-(3-(4-(4-aminobutyryl)piperazine-1-carbonyl)-4-fluorobenzyl)phthalazin-1(2H)-one (T), and its structural formula was:

[0106]

[0107] 12) Synthesis of intermediate tert-butyl 5-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanoic acid methyl ester (K): 0.4 g of 2-chloro-7-methyl-9-(tetrahydro-2H-pyran-4-yl)-7,9-dihydro-8H-purin-8-one (J) (1.49 mmol) was added to a reaction flask, and 0.365 g of 5-(4-methyl-3-aminophenoxy)pentanoic acid methyl ester (1.64 mmol), 0.258 g of 4,5-bis(diphenylphosphine)-9,9-dimethyloxy Anthracene (0.447 mmol), 0.97 g of cesium carbonate (2.98 mmol), 0.12 g of tris(dibenzylideneacetone)dipalladium and 15 mL of 1,4-dioxane solution were reacted at 105°C under a nitrogen environment for 12 hours. After the reaction was completed as monitored by a thin layer chromatography plate, the reaction solution was extracted with ethyl acetate (30 mL×3), allowed to stand for separation, and the organic phase was washed with water (5 mL×2) and saturated brine (5 mL×5) in turn, then dried over anhydrous sodium sulfate, filtered, and the ethyl acetate was removed under reduced pressure to obtain a solid. The solid was subjected to column chromatography using dichloromethane:methanol to obtain 0.3 g of a white solid with a yield of 44.2%. The colorless liquid obtained by column chromatography was identified by nuclear magnetic resonance method, and the results showed that the white solid was methyl 4,4-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanoate (K). Its structural formula is:

[0108]

[0109] 13) Synthesis of intermediate 5-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanoic acid (L): 0.3 g of methyl 4-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanoate (K) was placed in a reaction flask (0.658 mmol) and 5 ml of methanol was added to dissolve. 0.065 mg of NaOH (1.65 mmol) and 2 ml of water were added to dissolve. The NaOH aqueous solution was slowly added dropwise to the reaction flask. The reaction was carried out at 60 degrees for 30 minutes. After the reaction was completed as monitored by a thin layer chromatography plate, the reaction solution was distilled under reduced pressure to remove the solvent to obtain a crude product. After adding water to the crude product, stirring rapidly, filtering to obtain 260 mg of white solid, the yield is 89.7%. The white solid obtained by filtering was identified by nuclear magnetic resonance method, and the result showed that the white solid was 4-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanoic acid (L). Its structural formula:

[0110]

[0111] 14) Synthesis of the product N-(4-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-4-oxobutyl)-5-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanamide: 0.04 g of the intermediate 5-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanamide was taken. 4-(3-(4-(4-aminobutyryl)piperazine-1-carbonyl)-4-fluorobenzyl)phthalazin-1(2H)-one (T) (87.81 μmol), 0.012 g of EDCI (105.38 μmol), 0.018 g of HOBT (131.72 μmol), 0.022 g of triethylamine (219.53 μmol) and 1 ml of DMF were added in sequence. The mixture was reacted at room temperature for 2 hours. After the reaction was completed as monitored by a thin layer chromatography plate, ice water was added to the reaction bottle, which was rapidly stirred and filtered to obtain 0.015 g of a white solid with a yield of 19.2%. The white solid obtained by filtration was identified by nuclear magnetic resonance method, and the result showed that the white solid was N-(4-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-4-oxobutyl)-5-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanamide. Its structural formula:

[0112]

[0113] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-1 are as follows: 11H NMR (400 MHz, CDCl3) δ 8.44 - 8.32 (m, 1H), 7.84 (d, J = 1.7 Hz, 1H), 7.75 (d, J = 2.4 Hz, 1H), 7.69 (dt, J = 5.4, 2.0 Hz, 2H), 7.64 (dt, J = 8.4, 3.7 Hz, 1H), 7.34 - 7.21 (m, 2H), 7.02 - 6.86 (m, 2H), 6.81 (s, 1H), 6.43 (dd, J = 8.3, 2.8 Hz, 1H), 6.37 - 6.18 (m, 1H), 4.47 (tt, J = 12.2, 4.1 Hz, 1H), 4.21 (s, 2H), 4.03 (dd, J = 11.6, 4.5 Hz, 2H), 3.92 (d, J = 3.8 Hz, 2H), 3.46 (qd, J = 8.1, 4.3 Hz, 4H), 3.32 (s, 3H), 3.21 (dd, J = 11.1, 5.6 Hz, 3H), 2.78 - 2.61 (m, 2H), 2.18 (d, J = 6.7 Hz, 3H), 1.71 - 1.66 (m, 1H), 1.21 (s, 1H), 0.85 - 0.75 (m, 1H). 13 13C NMR (101 MHz, CDCl3) δ 172.12, 170.41, 164.18, 159.62, 156.80, 154.72, 154.33, 151.70, 149.25, 144.39, 137.80, 133.41, 132.63, 131.66, 130.78, 130.58, 129.70, 128.49, 128.15, 127.28, 126.09, 123.96, 117.68, 115.92, 107.22, 105.17, 76.38, 76.06, 75.74, 66.40, 48.98, 45.70, 44.05, 41.04, 38.18, 36.64, 35.36, 29.72, 28.91, 28.67, 27.86, 26.29, 23.54, 21.56, 16.29, 0.00. MS (ESI) m / z (M + H) + : calculated for C 47 H 53 FN 10 O7: 889.4156, found: 889.4166。

[0114] Example 2

[0115] In the preparation of the compound of this embodiment, step 1) compound (B) uses methyl 4-bromobutyrate, step 7) compound (M) uses 2-aminobutyric acid, and the remaining steps are the same as in Example 1 to obtain the compound of this embodiment N-(1-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-1-oxobutan-2-yl+)-4-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)butanamide, 0.027 g of white solid product, yield 34.56%, recorded as compound LSQ-2. The structure is:

[0116]

[0117] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-2 are as follows: 1 H NMR (400MHz, CDCl3) δ7.94-7.74(m,2H),7.74-7.57(m,2H),7.25(s,1H),7.03-6.72(m,3H),6.56-6. 31(m,1H),4.57-4.39(m,1H),4.21(s,1H),4.06(qd,J=13.4,8.5Hz,2H),3.94(dt,J=10.3,5.5Hz,2H ),3.62(d,J=16.4Hz,2H),3.57-3.38(m,3H),3.38-3.24(m,3H),2.77-2.67(m,2H),2.45(dt,J=21.6 ,6.0Hz,2H),2.20(s,3H),2.09-1.96(m,2H),1.67(t,J=11.8Hz,2H),1.51(s,1H),0.92-0.70(m,2H). 13C NMR (101MHz, CDCl3) δ173.86,172.77,170.79,165.14,160.62,157.83,1 57.75,155.72,155.36,155.25,152.72,150.27,150.07,145.38,138.86, 134.48,133.61,132.96,132.63,131.57,130.71,130.62,129.51,129.2 4,128.31,127.12,124.99,123.43,118.69,118.06,116.93,116.77,116. 29,108.30,106.66,106.20,106.12,77.43,77.11,76.79,67.40,67.34, 66.69,53.16,52.18,51.61,50.03,49.57,47.28,46.74,45.33,42.23,41 .91,37.69,32.96,32.86,30.63,30.04,29.92,27.34,27.29,25.93,25. 70,25.57,24.77,17.31,17.27,9.84,9.76,9.70,1.03.MS(ESI)m / z(M+H) + :calculated for C 46 H 51 FN 10 O7:875.3999,found:875.4022.

[0118] Example 3

[0119] In the preparation of the compound of this embodiment, step 1) compound (B) uses methyl 4-bromobutyrate, step 7) compound (M) uses 3-aminopropionic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this embodiment N-(3-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-3-oxopropyl)-4-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)butanamide, 0.030 g of white solid product, yield 12.5%, recorded as compound LSQ-3. The structure is:

[0120]

[0121] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-3 are as follows: 1H NMR(400MHz,CDCl3)δ8.45-8.25(m,1H),7.87-7.76(m,2H),7.71-7.59(m,2H),7.25(dt,J=6.3,3.6Hz,2H),6.95(td,J=7.9,4.6Hz,2H),6.93-6.79(m,2H),6.48-6.32(m,1H),4.54-4.42(m,1H),4.21(s,2H),4.04(ddd,J=11.7,5.7,3.9Hz,2H),3.95-3.82(m,2H),3.63(td,J=21.9,12.9Hz,3H),3.45(q,J=11.4Hz,6H),3.31(d,J=7.6Hz,4H),3.23-3.12(m,1H),2.70(qd,J=12.3,4.6Hz,2H),2.45(dq,J=24.4,7.0Hz,2H),2.36-2.26(m,2H),2.19(t,J=3.6Hz,3H),2.01(p,J=5.0Hz,2H),1.67(d,J=11.8Hz,2H),1.18(t,J=7.1Hz,1H). 13 C NMR(101MHz,CDCl3)δ172.22,172.04,169.24,169.06,164.17,164.04,159.74,156.79,154.69,154.35,154.22,151.70,149.15,144.43,137.80,137.73,133.48,132.62,131.75,131.54,130.81,130.57,129.68,129.63,128.50,128.20,127.23,126.06,123.96,117.36,115.85,115.81,115.28,115.06,106.12,106.06,105.45,105.25,76.44,76.32,76.12,75.88,75.80,66.39,65.82,65.71,49.02,43.81,41.00,40.80,40.59,36.59,34.17,34.01,32.22,32.07,31.84,28.99,28.90,26.32,26.27,24.42,16.29,16.24,13.18,-0.00.MS(ESI)m / z(M+H)+:calculated for C 45 H 49 FN 10O7:861.3843,found:861.3849.

[0122] Example 4

[0123] In the preparation of the compound of this example, step 1) compound (B) uses methyl 7-bromoheptanoate, step 7) compound (M) uses 7-aminoheptanoic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this example N-(7-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-7-oxoheptyl)-7-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)heptylamide, 0.019 g of white solid product, yield 10.1%, recorded as compound LSQ-4. The structure is:

[0124]

[0125] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-4 are as follows: 1 H NMR (400MHz, CDCl3) δ8.48-8.22(m,1H),7.83(s,1H),7.75(d,J=2.6Hz,1H),7.71-7.62(m,2H),7.26(dd,J=9.2,4.3Hz,2H),6.9 8(dd,J=8.9,4.0Hz,2H),6.78(s,1H),4.47(s,1H),4.22(s,2H),4.03(dd,J=11.6,4.5Hz,2H),3.89(t,J=6.3Hz,2H),3.62(d,J= 41.1Hz,3H),3.54-3.37(m,4H),3.32(s,4H),3.14(q,J=6.6Hz,2H),2.69(dd,J=12.5,4.6Hz,2H),2.28(t,J=7.5Hz,1H),2.20(s ,4H),2.09(t,J=7.6Hz,2H),1.66(ddd,J=19.3,14.8,10.9Hz,5H),1.60-1.45(m,4H),1.40(q,J=7.4Hz,4H),1.33-1.20(m,6H). 13C NMR (101MHz, CDCl3) δ160.69,158.01,155.42,152.74,150.30,138.79,134.49,133. 66,132.65,131.62,130.70,129.53,128.31,127.14,125.03,118.66,116.95,108.38 ,106.34,77.42,77.30,77.10,76.78,67.82,67.45,50.02,39.27,36.69,29.91,29. 43,29.25,29.01,28.90,27.31,26.63,25.86,25.73,24.96,17.33.MS(ESI)m / z(M+H) + :calculated for C 52 H 63 FN 10 O7:959.4938,found:959.4967.

[0126] Example 5

[0127] In the preparation of the compound of this example, step 1) compound (B) uses 3-(bromoethyl) methyl benzoate, step 7) compound (M) uses glycine, and the remaining steps are the same as in Example 1 to obtain the compound of this example N-(2-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-2-oxoethyl)-3-((4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)methyl)benzamide, 0.021 g of white solid product, yield 10.7%, recorded as compound LSQ-5. The structure is:

[0128]

[0129] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-5 are as follows: 11H NMR (400 MHz, CDCl3) δ 10.91 (d, J = 12.0 Hz, 1H), 8.50 - 8.40 (m, 1H), 8.00 - 7.90 (m, 2H), 7.89 (s, 1H), 7.81 - 7.73 (m, 3H), 7.73 - 7.67 (m, 1H), 7.64 (d, J = 7.7 Hz, 1H), 7.48 - 7.38 (m, 2H), 7.34 (d, J = 4.5 Hz, 2H), 7.09 - 6.99 (m, 2H), 6.82 (s, 1H), 6.59 (dd, J = 8.3, 2.6 Hz, 1H), 5.11 (s, 2H), 4.53 (tt, J = 12.2, 4.0 Hz, 1H), 4.33 - 4.19 (m, 4H), 4.07 (dd, J = 11.5, 4.2 Hz, 2H), 3.65 - 3.54 (m, 3H), 3.54 - 3.43 (m, 3H), 3.39 (s, 3H), 2.77 (qd, J = 12.5, 4.5 Hz, 2H), 2.28 (s, 3H), 1.75 - 1.62 (m, 2H). 13 13C NMR (101 MHz, CDCl3) δ 167.20, 167.03, 160.47, 157.50, 155.27, 152.74, 150.28, 145.45, 138.90, 138.04, 133.71, 132.64, 131.68, 130.78, 129.51, 128.87, 128.33, 127.20, 126.66, 126.31, 124.99, 119.02, 117.02, 116.35, 116.13, 108.51, 106.44, 77.37, 77.26, 77.05, 76.73, 69.61, 67.38, 50.02, 41.74, 37.65, 29.93, 27.32, 17.36. MS (ESI) m / z (M + H) + : calculated for C 48 H 47 FN 10 O7: 895.3686, found: 895.3693。

[0130] Example 6

[0131] In the preparation of the compound of this example, step 1) compound (B) uses 3-(bromoethyl) methyl benzoate, step 7) compound (M) uses 3-aminopropionic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this example N-(3-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-3-oxopropyl)-3-((4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)methyl)benzamide, 0.027 g of white solid product, yield 11.6%, recorded as compound LSQ-6. The structure is:

[0132]

[0133] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-6 are as follows: 1 H NMR (400MHz, CDCl3) δ11.11(d,J=9.2Hz,1H),8.48-8.35(m,1H),7.98-7.82(m,3H),7.72(ddd,J=16.9,6.2,3.0Hz,4H), 7.59(d,J=7.6Hz,1H),7.40(dt,J=13.2,7.0Hz,2H),7.34-7.27(m,2H),7.08-6.97(m,2H),6.87(s,1H),6.58(dd,J=8.3 ,2.5Hz,1H),5.09(s,2H),4.52(ddt,J=12.1,8.0,3.9Hz,1H),4.27(s,2H),4.07(dd,J=11.5,4.0Hz,2H),3.80-3.65(m, 4H),3.64(s,1H),3.60-3.41(m,6H),3.39(s,3H),3.28(s,2H),2.81-2.56(m,4H),2.28(s,3H),1.72(d,J=10.0Hz,2H). 13C NMR (101MHz, CDCl3) δ169.46,169.26,166.36,166.27,164.16,164.02,159.48,156.47,154.23,151.69,149.23,144.40, 137.87,136.84,133.59,133.42,132.63,131.63,130.72,130.60,129.73,128.47,128.19,127.74,127.27,126.12,125.6 0,125.22,123.95,118.03,115.98,115.28,115.06,107.42,105.45,76.34,76.23,76.02,75.71,68.66,66.34,48.97,45 .90,45.62,44.44,43.88,40.97,40.82,40.66,40.17,36.63,34.60,31.87,28.90,26.29,16.32,-0.00.MS(ESI)m / z(M+H) + :calculated for C 49 H 49 FN 10 O7:909.3843,found:909.3816.

[0134] Example 7

[0135] In the preparation of the compound of this embodiment, step 1) compound (B) uses methyl 4-(bromoethyl)benzoic acid, step 9) compound (R) and step 13 compound (L) are reacted in step 14), and the remaining steps are the same as in Example 1 to obtain the compound of this embodiment 4-(4-fluoro-3-(4-(4-((4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)methyl)benzoyl)piperazine-1-carbonyl)benzyl)phthalazin-1(2H)-one, 0.018 g of white solid product, yield 23.04%, recorded as compound LSQ-7. The structure is:

[0136]

[0137] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-7 are as follows: 1H NMR (400MHz, CDCl3) δ8.28 (s, 2H), 8.06 (s, 1H), 7.94 (d, J = 7.9Hz, 2H), 7.85 (s, 1H), 7. 59-7.35(m,5H),7.29(d,J=49.1Hz,1H),7.06(d,J=8.4Hz,1H),6.63(dd,J=8.4,2.6Hz ,1H),5.12(d,J=15.5Hz,2H),4.36(d,J=30.9Hz,2H),3.92(dd,J=11.6,4.3Hz,2H),3. 65(s,3H),3.30(s,3H),2.61-2.52(m,2H),2.17(s,3H),1.66(dd,J=12.7,4.0Hz,2H). 13 C NMR (101MHz, DMSO) δ159.85,156.87,155.97,152.48,150.08,139.89,135.28,133.99,131.02,129.88,127.94,127.69,125.93,122 .60,116.95,109.59,69.03,66.98,49.60,40.57,40.37,40.16,39.95,39.74,39.53,39.32,29.95,27.55,17.77.MS(ESI)m / z(M+H) + :calculated for C 46 H 44 FN9O6:838.3472,found:838.3455.

[0138] Example 8

[0139] In the preparation of the compound of this example, step 7) compound (M) uses 5-aminopentanoic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this example N-(5-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-5-oxopentyl)-5-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)pentanamide, 0.021 g of white solid product, yield 26.88%, recorded as compound LSQ-8. The structure is:

[0140]

[0141] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-8 are as follows: 11H NMR (400 MHz, CDCl3) δ 8.44 (d, J = 7.1 Hz, 1H), 7.90 (s, 1H), 7.75 (q, J = 11.3 Hz, 4H), 7.32 (p, J = 7.8 Hz, 3H), 7.04 (d, J = 9.0 Hz, 2H), 6.51 (d, J = 8.8 Hz, 2H), 4.53 (s, 1H), 4.28 (s, 2H), 4.13 - 4.05 (m, 2H), 4.03 - 3.91 (m, 2H), 3.84 - 3.61 (m, 3H), 3.60 - 3.46 (m, 4H), 3.41 (d, J = 14.7 Hz, 4H), 3.34 - 3.14 (m, 4H), 2.83 - 2.66 (m, 2H), 2.37 (td, J = 17.4, 8.1 Hz, 2H), 2.26 (s, 5H), 1.77 (q, J = 15.8 Hz, 7H), 1.67 - 1.63 (m, 1H), 1.59 - 1.46 (m, 2H). 13 13C NMR (101 MHz, CDCl3) δ 173.40, 171.67, 165.27, 160.77, 158.17, 157.77, 155.71, 155.19, 152.70, 150.46, 145.51, 142.06, 138.56, 134.46, 133.66, 132.04, 131.84, 131.60, 130.74, 129.49, 128.20, 127.04, 126.46, 125.10, 124.99, 119.19, 118.19, 116.86, 116.31, 110.47, 108.57, 106.64, 77.46, 77.15, 76.83, 67.39, 51.60, 50.07, 47.10, 46.75, 45.63, 45.06, 42.16, 41.94, 41.72, 41.21, 38.98, 38.79, 37.63, 36.28, 33.46, 32.43, 29.91, 28.84, 27.34, 22.58, 22.05, 21.72, 17.31. MS (ESI) m / z (M + H) + : calculated for C 48 H 55 FN 10 O7: 903.4312, found: 903.4315。

[0142] Example 9

[0143] In the preparation of the compound of this example, step 1) compound (B) uses methyl 6-bromohexanoate, and the remaining steps are the same as in Example 1 to obtain the compound of this example N-(4-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-4-oxobutyl)-6-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)hexanamide, 0.024 g of white solid product, yield 30.72%, recorded as compound LSQ-9. The structure is:

[0144]

[0145] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-9 are as follows: 1 H NMR (400MHz, CDCl3) δ8.42-8.33(m,1H),7.84(s,1H),7.74(d,J=2.6Hz,1H),7.69( tq,J=8.8,4.4Hz,2H),7.65(t,J=3.7Hz,1H),7.25(dd,J=8.2,4.8Hz,2H),7.00-6. 91(m,2H),6.79(s,1H),6.44(dt,J=8.7,2.4Hz,1H),6.14(d,J=20.9Hz,1H),4.47( t,J=4.1Hz,1H),4.21(s,2H),4.04(dd,J=11.6,4.4Hz,2H),3.91(tt,J=6.3,2.8Hz, 2H),3.67(s,2H),3.58(s,2H),3.48(dt,J=12.3,3.0Hz,3H),3.42(d,J=4.2Hz,1H) ,3.33(s,3H),3.24-3.18(m,3H),2.69(dd,J=12.5,4.6Hz,1H),2.35(t,J=6.6Hz,1H ),2.20(s,2H),2.12(t,J=7.5Hz,2H),1.78(d,J=6.2Hz,2H),1.68(d,J=3.7Hz,1H) ,1.65(d,J=3.5Hz,1H),1.44(d,J=7.8Hz,1H),1.28(d,J=15.6Hz,1H),1.18(s,2H). 13CNMR (101MHz, CDCl3) δ171.47,160.49,157.93,155.42,152.76,150.31,145.46,138.80,134. 42,133.70,132.67,131.67,130.73,129.53,128.34,127.18,125.01,118.76,116.96,108.35, 106.34,77.38,77.27,77.07,76.86,76.75,70.53,67.63,67.46,50.04,45.11,39.20,37.68,3 6.70,31.94,30.91,29.94,29.71,29.08,27.32,25.84,25.51,24.56,17.33.MS(ESI)m / z(M+H) + :calculated for C 48 H 55 FN 10 O7:903.4312,found:903.4315.

[0146] Example 10

[0147] In the preparation of the compound of this embodiment, step 1) compound (B) uses methyl 4-bromobutyrate, step 7) compound (M) uses 3-azetidine carboxylic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this embodiment 4-(4-fluoro-3-(4-(1-(4-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)butanoyl)azinocyclobutane-3-carbonyl)piperazine-1-carbonyl)benzyl)phthalazin-1(2H)-one, 0.029 g of white solid product, yield 37.12%, recorded as compound LSQ-10. The structure is:

[0148]

[0149] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-10 are as follows: 1H NMR(400MHz,CDCl3)δ8.41-8.31(m,1H),7.83(q,J=4.7Hz,1H),7.75(dd,J=8.9,2.6Hz,1H),7.72-7.61(m,3H),7.26(dt,J=10.3,5.1Hz,2H),7.00-6.90(m,2H),6.86(d,J=5.3Hz,1H),6.44(dd,J=8.3,2.9Hz,1H),4.46(tt,J=13.2,4.7Hz,2H),4.36(d,J=7.0Hz,1H),4.21(s,2H),4.17-4.08(m,1H),4.03(dd,J=9.4,5.3Hz,2H),3.97-3.90(m,2H),3.88(s,1H),3.75-3.50(m,4H),3.44(dd,J=11.3,7.4Hz,2H),3.33-3.30(m,3H),3.26-3.04(m,3H),2.74-2.64(m,2H),2.49(d,J=7.4Hz,1H),2.36(d,J=7.2Hz,1H),2.19(q,J=4.2Hz,3H),2.06-1.93(m,3H),1.79-1.58(m,3H). 13 C NMR(101MHz,CDCl3)δ171.90,168.55,164.20,159.72,156.74,156.69,154.33,154.27,151.69,149.31,144.44,137.80,137.71,133.52,132.64,131.46,130.94,130.60,129.74,128.48,127.21,126.07,123.94,117.85,117.77,115.95,107.33,105.45,105.05,76.41,76.29,76.09,75.87,75.77,66.37,65.82,65.69,50.76,49.20,48.99,43.73,41.04,40.96,36.61,29.65,29.05,28.89,26.64,26.35,26.29,23.56,16.28,16.24,0.00.MS(ESI)m / z(M+H) + :calculated for C 46 H 49 FN 10 O7:873.3843,found:873.3815。

[0150] Comparative Example 1

[0151] In the preparation of the compound of this example, step 1) compound (B) uses 7-bromo-2,2-dimethylheptanoate, step 7) compound (M) uses 6-aminohexanoic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this example N-(6-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-6-oxohexyl)-2,2-dimethyl-7-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)heptylamide, 0.013 g of white solid product, yield 16.64%, recorded as compound LSQ-11. The structure is:

[0152]

[0153] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-11 are as follows: 1 H NMR (400MHz, CDCl3) δ8.39 (dd, J=7.0, 2.1Hz, 1H), 7.83 (s, 1H), 7.77-7.56 (m, 3H), 7.30-7.22 (m, 1H), 6.98 (dd, J=8.2, 4.9Hz, 2H),6.79(s,1H),6.47-6.33(m,1H),5.76(d,J=5.5Hz,1H),3.32(s,3H),2.69(dd,J=12.5,4.4Hz,2H),1.08(d,J=2.3Hz,5H). 13C NMR (101MHz, CDCl3) δ177.75,171.60,165.22,160.52,157.97,155.74,155.42,152.74,150.31,145.43,138.78,134.43,134.40, 133.66,132.62,131.77,131.70,131.62,130.70,129.52,129.16,128.33,127.16,125.01,118.75,116.99,116.96,116.12,108. 44,106.47,106.42,77.38,77.27,77.07,76.75,67.88,67.44,50.02,46.82,45.09,42.19,42.02,41.67,41.26,39.12,37.69,35 .53,32.97,29.91,29.70,29.35,29.26,27.30,26.71,26.66,26.49,26.19,25.50,24.73,24.44,17.33,1.04..MS(ESI)m / z(M+H) + :calculated for C 50 H 59 FN 10 O7:931.4625,found:931.4655.

[0154] Comparative Example 2

[0155] In the preparation of the compound of this example, step 1) compound (B) uses 7-bromo-2,2-dimethylheptanoate, step 7) compound (M) uses 3-azetidine carboxylic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this example 4-(3-(4-(1-(2,2-dimethyl-7-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)heptanoyl)azetidine-3-carbonyl)piperazine-1-carbonyl)-4-fluorobenzyl)phthalazin-1(2H)-one, 0.016 g of white solid product, yield 20.48%, recorded as compound LSQ-12. The structure is:

[0156]

[0157] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-12 are as follows: 1H NMR(400MHz,CDCl3)δ8.38(dt,J=7.1,3.0Hz,1H),7.88-7.79(m,1H),7.78-7.60(m,4H),7.26(dq,J=9.0,3.2Hz,2H),6.98(td,J=8.7,3.9Hz,2H),6.80(d,J=8.5Hz,1H),6.46(dt,J=8.2,2.4Hz,1H),4.47(tt,J=12.3,4.1Hz,1H),4.21(d,J=7.6Hz,2H),4.03(dd,J=11.8,4.3Hz,3H),3.89(td,J=6.6,2.4Hz,3H),3.57(s,3H),3.50-3.39(m,4H),3.32(d,J=2.7Hz,4H),3.27-3.07(m,3H),2.69(qd,J=12.4,4.3Hz,2H),2.19(d,J=2.8Hz,3H),1.68(ddd,J=24.1,10.5,5.3Hz,5H),1.48-1.34(m,4H),1.27-1.16(m,4H),1.13-1.02(m,6H). 13 C NMR(101MHz,CDCl3)δ176.29,168.79,168.59,164.20,159.70,156.98,154.69,154.36,151.70,149.28,144.40,137.74,133.51,132.64,131.56,130.91,130.83,130.60,129.67,128.48,128.19,127.24,126.10,123.94,122.40,117.72,115.94,115.31,115.05,107.49,105.36,76.39,76.28,76.08,75.76,66.80,66.40,53.26,48.98,45.94,45.64,44.17,43.73,41.14,41.07,41.01,40.77,40.54,39.26,36.63,30.30,28.87,28.66,28.19,26.28,25.62,24.51,24.25,23.55,16.29,16.26,-0.00.MS(ESI)m / z(M+H) + :calculated forC 51 H 59 FN 10 O7:943.4625,found:943.4612。

[0158] Comparative Example 3

[0159] In the preparation of the compound of this example, step 1) compound (B) uses methyl 4-(bromoethyl)phenylacetic acid, step 7) compound (M) uses 7-aminoheptanoic acid, and the remaining steps are the same as in Example 1 to obtain the compound of this example N-(6-(4-(2-fluoro-5-((4-oxo-3,4-dihydrophthalazin-1-yl)methyl)benzoyl)piperazin-1-yl)-6-oxohexyl)-4-(4-methyl-3-((7-methyl-8-oxo-9-(tetrahydro-2H-pyran-4-yl)-8,9-dihydro-7H-purin-2-yl)amino)phenoxy)benzamide, 0.019 g of white solid product, yield 24.32%, recorded as compound LSQ-13. The structure is:

[0160]

[0161] The hydrogen nuclear magnetic resonance spectrum, carbon nuclear magnetic resonance spectrum and mass spectrum of compound LSQ-13 are as follows: 1 H NMR (400MHz, CDCl3) δ8.50-8.40(m,1H),7.96(d,J=2.7Hz,1H),7.88(s,1H),7.80-7.71(m,2H),7.71-7.64(m,1H),7.47-7.42(m, 2H),7.31(q,J=7.3Hz,3H),7.06(dd,J=13.4,8.5Hz,3H),6.89-6.79(m,1H),6.59(dd,J=8.4,2.5Hz,1H),4.53(tt,J=12.4,4.2Hz, 1H),4.27(s,2H),4.06(dd,J=11.6,4.5Hz,3H),3.71(d,J=4.2Hz,4H),3.64(d,J=3.1Hz,2H),3.59-3.44(m,8H),3.39(s,3H),3.1 7(t,J=6.6Hz,3H),2.75(s,3H),2.28(s,3H),2.16-1.95(m,4H),1.72(dd,J=13.0,4.1Hz,2H),1.58(d,J=8.0Hz,3H),1.42(s,1H). 13C NMR (101MHz, CDCl3) δ160.52,157.63,155.30,152.73,150.31,145.44,138.89,136.47,1 34.66,134.40,133.66,132.58,131.63,130.76,129.61,129.52,128.33,127.16,125.01, 119.10,117.04,108.51,106.52,77.39,77.07,76.76,69.72,67.39,50.05,43.58,42.20 ,39.52,37.70,33.06,29.94,29.24,28.84,27.33,26.52,24.93,17.36.MS(ESI)m / z(M+H) + :calculated for C 46 H 49 FN 10 O7:979.4625,found:979.4606.

[0162] Performance Testing

[0163] 1. Test of the inhibitory activity of compounds on DNA-PK

[0164] The enzyme inhibitory activity of the compounds was determined by fluorescence analysis, including DNA-PK0(V4106) enzyme, ADP-Glo TM Kinase Assay (V9101) and ATP (V915B) were purchased from Pormega, 384well small volume white plate (784075) was purchased from Greiner, and Envision was purchased from PerkinElmer. The specific test method is as follows:

[0165] The experimental process is as follows:

[0166] 1. Thaw DNA-PK enzyme, DNA-Dependent Protein Kinase Substrate, 5X Reaction Buffer A, DNA-PK Activation Buffer, DTT (100Mm) and ATP (10mM) on ice, and keep the above reagents on ice throughout the experiment;

[0167] 2. Prepare 1× buffer by mixing 5× Reaction Buffer A and 10× DNA-PK Activation Buffer with deionized water, and add DTT to it. The concentration of DTT in 1× buffer is 50 μM.

[0168] 3. Prepare the test compound storage solution with 1× buffer into a 5-fold final concentration gradient, take 1 μl / well of the test compound and add it to the white microplate. At this time, the DMSO concentration of the compound should not be greater than 5%, and the final DMSO concentration of the experiment should not be greater than 1%. Centrifuge the microplate at 1000 rpm for 1 minute.

[0169] Positive control well (Pos.Ctrl): 1 μL / well 1× buffer containing 5% DMSO;

[0170] Blank control wells (Blank): 1 μL / well 1× buffer containing 5% DMSO;

[0171] 4. After the DNA-PK enzyme is completely thawed, use 1X buffer to dilute the DNA-PK enzyme to 2.5 unit / μL, take 2μL / well and add it to the white microplate. At this time, the amount of DNA-PK enzyme in each well is 5 units; add 2μL / well 1× buffer to the blank control well; this step should be performed on ice. After adding, centrifuge the microplate at 1000 rpm for 1 minute;

[0172] 5. Prepare DNA-PK peptide substrate / ATP mixture:

[0173] DNA-PK peptide substrate / ATP mixture: dilute DNA-PK peptide substrate 20 times with 1× buffer and dilute ATP (10mM) 80 times. The concentration of DNA-PK peptide substrate is 0.5ug / ul and the concentration of ATP is 125μM. Please perform this step on ice;

[0174] 6. Take 2 μL / well of DNA-PK peptide substrate / ATP mixed solution into the white microplate. At this time, the concentration of DNA-PK peptide substrate is 0.2 ug / ul, and the concentration of ATP is 50 μM. After adding, centrifuge the microplate at 1000 rpm for 1 minute.

[0175] 7 After centrifugation, stick the membrane to the microplate, press the membrane tightly, and incubate at 25℃ for 1 hour;

[0176] 8. After the incubation is completed, use Envison to perform chemiluminescence detection and read the luminescence value (RLU);

[0177] 9. Calculation of inhibition rate:

[0178] %Inhibtion=(RLU(Sample)-RLU(Pos.Ctrl)) / (RLU(Blank)-RLU(Pos.Ctrl))×100%

[0179] The results are shown in Table 1.

[0180] Table 1 Test results of DNA-PK inhibitory activity of compounds

[0181] Compound <![CDATA[DNA-PK IC 50 (nM)]]> LSQ-1 10.84 LSQ-2 23.21 LSQ-3 15.04 LSQ-4 1.99 LSQ-5 1.98 LSQ-6 4.51 LSQ-7 8.28 LSQ-8 8.38 LSQ-9 29.58 LSQ-10 27.04 LSQ-11 ＞50 LSQ-12 ＞50 LSQ-13 ＞50 Wortmannin 15.35

[0182] Among them, the positive control Wortmannin was compared with samples LSQ-1-10 respectively. The experimental results in Table 1 show that compounds LSQ-1 to LSQ-10 have strong inhibitory effects on DNA-PK, and their activities are comparable to those of Wortmannin. However, compounds LSQ-11 to LSQ-13 have poor inhibitory effects.

[0183] 2. Test of compound inhibitory activity on PARP1

[0184] The enzyme inhibition activity of the compound was determined by fluorescence analysis, where PARP1 Chemiluminescent AssayKit (80569) was purchased from BPS and Envision was purchased from PerkinElmer. The specific test method is as follows:

[0185] The experimental process is as follows:

[0186] 1. One day before the experiment, thaw 5×histone mixture on ice;

[0187] 2. Dilute 5×histone mixture 5 times with 1×PBS to prepare 1×histone mixture;

[0188] 3. Take 25 μL / well of 1×histone mixture and add it to the test plate. Centrifuge the test plate at 1000 rpm for 1 minute. Incubate at 4°C overnight.

[0189] 4. After the incubation, dry the liquid in the test plate and wash the test plate 3 times with 100 μL / well PBST buffer;

[0190] 5. Add 100 μL / well of Blocking buffer 3 to the test plate and incubate at 25°C for 60-90 minutes;

[0191] 6. Dilute 10× PARP Buffer 10 times with deionized water and place on ice for later use;

[0192] 7. After the incubation, dry the liquid in the test plate and repeat the plate washing 3 times;

[0193] 8. Take 2.5 μL / well of compound working solution with 10 times final concentration gradient and add it to the well plate according to the experimental arrangement diagram;

[0194] Positive control well (Pos.Ctrl): 2.5 μL / well 1X PARP buffer containing 10% DMSO;

[0195] Blank control wells: 2.5 μL / well 1X PARP buffer containing 10% DMSO;

[0196] 9. After the enzyme is completely dissolved, dilute the enzyme stock solution to 2 ng / μL with 1×PARP buffer, take 10μL / well enzyme solution and add it to the test plate. At this time, the amount of PARP1 enzyme in each well is 20 ng / well; add 10μL / well 1XPARP buffer to the blank control well; please do this step on ice, and centrifuge at 1000 rpm for 1 minute after adding;

[0197] 10. Add 12.5 μL of master mixture to each well of the PARP test plate. The 12.5 μL master mixture includes 1.25 μL 10× PARP buffer, 1.25 μL 10× PARP Assay mixture, 2.5 μL Activated DNA (5×), and 7.5 μL water. Seal the test plate and incubate at 25°C for 60 minutes.

[0198] 11. After the incubation, dry the liquid in the test plate and repeat the plate washing 3 times;

[0199] 12. Dilute the Streptavidin-HRP in the kit 50 times with the Blocking buffer solution, add 25 μL / well to the test plate, and incubate at 25°C for 30 minutes;

[0200] 13. After the incubation, dry the liquid in the test plate and repeat the washing 3 times;

[0201] 14. Mix HRP chemiluminescent substrate A and HRP chemiluminescent substrate B in the kit at a ratio of 1:1, add 50 μL / well of the mixture to the test plate, and immediately perform Luminescence detection using Envision to read the luminescence value (RLU);

[0202] 15. Calculation of inhibition rate:

[0203] %Inhibtion=(RLU(Sample)-RLU(Pos.Ctrl)) / (RLU(Blank)-RLU(Pos.Ctrl))×100%

[0204] The results are shown in Table 2.

[0205] Table 2 Test results of the inhibitory activity of compounds on PARP1

[0206]

[0207]

[0208] Comparison of positive control Niraparib and samples LSQ-1-10. The experimental results in Table 2 show that compounds LSQ-1 to LSQ-10 have strong inhibitory effects on PARP1, and their activity is comparable to that of Niraparib. However, compounds LSQ-11 to LSQ-13 have poor inhibitory effects.

[0209] 3. Activity test of compounds on breast cancer cells

[0210] (1) Test of compound toxicity to 4T1 cells

[0211] MTT was used to determine the cytotoxicity of the compound, where 4T1 cells were purchased from Fuheng Biological Company, MTT was purchased from Bid Pharmaceutical Company, 1640 was purchased from Fuheng Biological Company, bovine serum solution was purchased from Fuheng Biological Company, and penicillin-streptomycin antibacterial solution was purchased from Fuheng Biological Company. The specific test method is as follows:

[0212] The experimental process is as follows:

[0213] 1. Prepare 4T1 cell suspension: count cells; 2. Inoculate into 96-well plates, about 3000 cells per well, about 100ul cell suspension per well; 3. Culture in a 37°C incubator overnight; 4. Aspirate the culture medium, add drugs LSQ-1 to LSQ-13 diluted to 25μmol, and positive drugs AZD-7648 and Olaparib to the 98-well plate in sequence, and set up three replicate wells; 5. Culture in a 37°C incubator for 48h; 6. Add 20ul MTT in a dark environment, immerse the tip of the pipette in the culture medium and add it, and gently tap the culture plate after adding the reagent to help mix; 7. Culture in a 37°C incubator for 4 hours; 8. Measure the absorbance at 490nm and 570nm; 9. Calculate the inhibition rate.

[0214] Table 3 Results of cytotoxicity test of compounds on 4T1 cells

[0215] Compound 4T1 cell 25μmol inhibition rate (%) LSQ-1 14 LSQ-2 44 LSQ-3 81 LSQ-4 19 LSQ-5 50 LSQ-6 54 LSQ-7 40 LSQ-8 36 LSQ-9 35 LSQ-10 24 LSQ-11 ＜10 LSQ-12 ＜10 LSQ-13 ＜10 AZD-7648 32 Olaparib 53

[0216] Positive control substances AZD-7648 and Olaparib were compared with samples LSQ-1-10. The experimental results in Table 3 show that compounds LSQ-1 to LSQ-10 have strong toxicity to 4T1 cells, and most of their activities are comparable to those of AZD-7648. However, compounds LSQ-11 to LSQ-13 have poor inhibitory effects.

[0217] (2) Cytotoxicity test of compounds on B16-F10 cells

[0218] MTT was used to determine the cytotoxicity of the compound, where B16-F10 cells were purchased from Fuheng Biotechnology Co., Ltd., MTT was purchased from Bidex Pharmaceuticals Co., Ltd., and B16-F10 special culture medium was purchased from Prosai Co., Ltd. The specific test method is as follows:

[0219] The experimental process is as follows:

[0220] 1. Prepare B16-F10 cell suspension: count cells; 2. Inoculate into 96-well plates, about 3000 cells per well, about 100ul cell suspension per well; 3. Culture in a 37°C incubator overnight; 4. Aspirate the culture medium, add drugs LSQ-1 to LSQ-13 diluted to 25μmol, and positive drugs AZD-7648 and Olaparib to the 98-well plate in sequence, and set up three replicate wells; 5. Culture in a 37°C incubator for 48h; 6. Add 20ul MTT in a dark environment, immerse the tip of the pipette in the culture medium and add it, and gently tap the culture plate after adding the reagent to help mix; 7. Culture in a 37°C incubator for 4 hours; 8. Measure the absorbance at 490nm and 570nm; 9. Calculate the inhibition rate.

[0221] Table 4 Test results of cytotoxicity of compounds to B16-F10 cells

[0222] Compound B16-F10 cell 25μmol inhibition rate (%) LSQ-1 79 LSQ-2 79 LSQ-3 94 LSQ-4 80 LSQ-5 83 LSQ-6 84 LSQ-7 84 LSQ-8 75 LSQ-9 93 LSQ-10 82 LSQ-11 ＜10 LSQ-12 ＜10 LSQ-13 ＜10 AZD-7648 84 Olaparib 59

[0223] Positive control substances AZD-7648 and Olaparib were compared with samples LSQ-1-10. The experimental results in Table 4 show that compounds LSQ-1 to LSQ-10 have strong toxicity to B16-F10 cells, and their activity is comparable to that of AZD-7648. However, compounds LSQ-11 to LSQ-13 have poor inhibitory effects.

[0224] The weights of the relevant components mentioned in the embodiments of the present invention may not only refer to the specific content of each component, but also indicate the proportional relationship between the weights of the components. Therefore, as long as the content of the relevant components is proportionally enlarged or reduced according to the embodiments of the present invention, it is within the scope disclosed in the embodiments of the present invention.

Claims

1. A compound, characterized in that: The structure of the compound is shown in Formula 1; In Formula 1, R1 is selected from any one of an alkyl group, an aryl group, and a heterocycle, and R2 is selected from any one of an alkyl group, an aryl group, and a heterocycle.

2. The compound according to claim 1, characterized in that: In Formula 1, R1 is selected from any one of an alkyl group and an aryl group, and R2 is selected from any one of an alkyl group and a heterocycle.

3. The compound according to claim 1, characterized in that: The compound includes any one of the following compounds (1) to (10):

4. The method for preparing the compound according to claim 1, characterized in that: The following steps are involved: 1) Mixing the compound represented by formula I with 1-boc-piperazine, removing the protecting group, and then reacting with an amino acid to obtain the compound represented by formula II; Wherein R2 is selected from any one of an alkyl group, an aryl group, and a heterocycle; 2) mixing the compound represented by formula III with methyl bromoacetate, and then reducing the mixture to obtain the compound represented by formula IV; Wherein R1 is selected from any one of an alkyl group, an aryl group, and a heterocycle; 3) reacting the compound represented by formula V with 4-aminotetrahydropyran hydrochloride, followed by hydrolysis to obtain the compound represented by formula VI; 4) reacting the compound represented by formula VI with diphenylphosphoryl azide, and then reacting with iodomethane to obtain the compound represented by formula VII; 5) reacting the compound represented by formula VII with the compound represented by formula IV, followed by hydrolysis to obtain the compound represented by formula VIII: 6) reacting the compound represented by formula II with the compound represented by formula VIII to obtain the target compound.

5. The preparation method according to claim 4, characterized in that: In step 1), the molar ratio of the compound represented by formula I to 1-boc-piperazine is 1:0.8-1.2; In step 2), the molar ratio of the compound represented by formula III to methyl bromoate is 1:1.1-1.5; In step 3), the molar ratio of the compound represented by formula V to 4-aminotetrahydropyran hydrochloride is 1:0.8-1.2; In step 4), the molar ratio of the compound represented by formula VI to diphenylphosphoryl azide is 1:0.8-1.2; In step 5), the molar ratio of the compound represented by formula VII to the compound represented by formula IV is 1:1 to 1.4; In step 6), the molar ratio of the compound represented by formula II to the compound represented by formula VIII is 1:1 to 1.

4.

6. The use according to claim 8, characterized in that: In step 1), the molar ratio of the compound represented by formula I to 1-boc-piperazine is 1:1; In step 2), the molar ratio of the compound represented by formula III to methyl bromoate is 1:1.39; In step 3), the molar ratio of the compound represented by formula V to 4-aminotetrahydropyran hydrochloride is 1:1; In step 4), the molar ratio of the compound represented by formula VI to diphenylphosphoryl azide is 1:1; In step 5), the molar ratio of the compound represented by formula VII to the compound represented by formula IV is 1:1.2; In step 6), the molar ratio of the compound represented by formula II to the compound represented by formula VIII is 1:1.

2.

7. A pharmaceutical composition, characterized in that: The pharmaceutical composition comprises the compound described in claim 1 or its stereoisomer, solvate, prodrug, metabolite, pharmaceutically acceptable salt or cocrystal.

8. Use of the compound according to claim 1 or its stereoisomers, solvates, prodrugs, metabolites, pharmaceutically acceptable salts or cocrystals in the preparation of drugs for treating and / or preventing and / or delaying and / or assisting in the treatment of cancer.

9. The use according to claim 8, characterized in that: The cancer includes at least one of breast cancer, melanoma and leukemia.

10. The use according to claim 8, characterized in that: The cancer drugs include DNA-PK inhibitors and / or PARP1 inhibitors.

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