N-methyl-2-(9-oxoazepin-10(9H)-yl)acetamide compounds, and preparation method and application thereof

Abstract

The application belongs to the technical field of medicine, and relates to N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamide compounds and a preparation method and application thereof. The preparation method of the compound is as follows: taking o-aminobenzoic acid as a starting material, performing substitution reaction, condensation reaction, alkylation reaction and hydrolysis reaction to generate a key intermediate acridone acetic acid, and finally performing condensation reaction on the acridone acetic acid and substituted pyrimidine in the presence of TCFH and NMI condensing agents to obtain the target compound. The preparation method of the N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamide compound provided in the application is simple and feasible, and good results are shown in in-vitro anti-tumor activity tests, and the compound has excellent anti-proliferation ability on human breast cancer cells BT-549 and MDA-MB-231 in particular.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamide compounds, their preparation methods, and applications. Background Technology

[0002] According to the IHC classification method, breast cancer can be divided into two groups: hormone receptor HR (ER, PR) positive and negative groups. Each group can be further divided into two subgroups based on HER2 expression levels, resulting in a total of four groups: luminal A (ER+, PR+, HER-), luminal B (ER+, PR+, HER+), HER2 (ER-, PR-, HER+), and TNBC (ER-, PR-, HER-). 10%–15% of breast cancer patients are negative for all three hormone receptors; this type is called triple-negative breast cancer (TNBC). TNBC is a highly heterogeneous disease compared to other breast cancers, commonly seen in young women. It typically presents as highly invasive ductal carcinoma, characterized by poor differentiation and high invasiveness. Its early recurrence rate is relatively high, often accompanied by distant metastasis; more than one-third of TNBC patients will experience recurrence or distant metastasis. Recurrent or metastatic TNBC generally has a poor prognosis, with a 5-year survival rate of less than 15%, significantly lower than the overall 5-year survival rate of 31% for breast cancer patients. Compared to other breast cancer subtypes, TNBC is negative for all three receptors, so conventional hormone receptor-targeting drugs and endocrine therapy are less effective for it. In current clinical practice, the standard treatment for TNBC is still mainly systemic chemotherapy. Although there are new treatments such as poly(ADP-ribose polymerase) (PARP) inhibitors, immune checkpoint inhibitors (PD-1, PDL-1), and antibody-drug conjugates (ADCs), many unknowns still exist and require further validation through clinical research. The treatment of TNBC remains a global challenge.

[0003] MYC genes are among the earliest discovered group of oncogenes, encoding a class of nuclear proteins including c-Myc, N-Myc, and L-Myc. The c-Myc proto-oncogene is one of the most common activated proto-oncogenes, belonging to the basic helical-loop-helical zipper (bHLHZip) transcription factor class. c-Myc is involved in a variety of cellular processes by regulating up to 15% of human genes. Its C-terminal domain contains a bHLHZip motif, which lacks a fixed structure before dimerizing with another bHLHZip protein—MAX. After dimerization, it forms an ordered α-helix structure, which undergoes various post-translational modifications and protein-protein interactions, regulating c-Myc function and binding to E-box elements in the promoter regions of target genes to regulate gene expression. Overexpression of c-Myc protein is commonly found in breast cancer, cervical cancer, and colon cancer. It has been detected in almost 70% of human cancers. As a downstream protein regulated by transcription factors and closely related to the cell cycle, c-Myc protein has become an attractive cancer treatment strategy as a potential regulatory target for inhibiting the malignant proliferation of tumor cells.

[0004] Some regions of the c-Myc protein lack a fixed secondary structure before dimerization, and the c-Myc protein molecule lacks an active pocket on its surface that can serve as a drug-binding region. Its short half-life has made it difficult to target as a drug, making it a recognized difficult-to-drug target. The gene encoding the c-Myc protein contains a nucleic acid hypersensitive element III located upstream of the P1 promoter. 1 (NHEⅢ) 1 This element controls approximately 85%–90% of the transcriptional activity of the c-Myc gene. The G4 structure of c-Myc consists of a single-stranded guanine-rich (G) sequence, typically with four consecutive G bases. Each group contains one G base, and these four G bases interact via HoogSteen hydrogen bonds to form a tetrap. Multiple tetrap planes are further stacked to form the G4 secondary structure of c-Myc. Several non-G bases are spaced between the consecutive G base groups, forming a loop region wrapped around the G tetrap. The G4 structure can appear in the promoter regions and telomere ends of some genes. Its formation or disintegration can regulate gene stability, replication, transcription, and telomere length. As a transient special DNA secondary structure, it may appear during the transcriptional amplification phase of the c-Myc gene and inhibits the interaction of RNA polymerase and NHEIII. 1 The binding of ligands restricts the expression of the c-Myc gene, which is an important way to silence c-Myc gene transcription. In addition, the G4 structure of c-Myc has a single strand of DNA that does not form a G4 strand coiled around the corresponding terminal tetrad at the 3' and 5' ends to form a capping structure, creating a potential binding site for small molecule ligands.

[0005] In cancer treatment and diagnosis, exploring the molecular-level effects of G4 structure on tumor-related mechanisms is a promising research avenue. G4 ligands inhibit tumor proliferation by inducing and interacting with G4 structures, maintaining telomeres, suppressing proto-oncogene transcription, and disrupting genomic stability. Currently, there are no marketed drugs targeting G4. The main challenge in drug development lies in the diversity of G4 types, structures, and morphologies. Compounds need to specifically target the G4 structure in the c-Myc gene promoter region while also possessing good drug-like properties, posing a significant challenge to the development of current G4 ligand drugs. Summary of the Invention

[0006] In view of the above-mentioned technical problems, the present invention provides N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamide compounds, their preparation methods and applications. The prepared compounds showed good results in in vitro antitumor activity tests, especially exhibiting excellent antiproliferative ability against human breast cancer cells (BT-549, MDA-MB-231).

[0007] N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamide compounds, as shown in general formulas I, II and III: in: Each R1 is independently methylpiperazinyl, morpholinyl, methylpiperidinyl, dimethylamino, N,N-diethylamino, N,N-dimethylethane-1,2-diamino, N,N-dimethylpropane-1,3-diamino, N,N,N-trimethylethane-1,2-diamino, N,N,N-trimethylpropane-1,3-diamino, pyrrolyl, 2-(piperazin-1-yl)ethanol-1-olyl, aniline, benzylamino; Each R2 is independently methylpiperazinyl, morpholinyl, methylpiperidinyl, dimethylamino, N,N-diethylamino, N,N-dimethylethane-1,2-diamino, N,N-dimethylpropane-1,3-diamino, N,N,N-trimethylethane-1,2-diamino, N,N,N-trimethylpropane-1,3-diamino, pyrrolyl, 2-(piperazin-1-yl)ethanol-1-olyl, aniline; Each R3 is independently methylpiperazinyl, morpholinyl, methylpiperidinyl, dimethylamino, N,N-diethylamino, N,N-dimethylethane-1,2-diamino, N,N-dimethylpropane-1,3-diamino, N,N,N-trimethylethane-1,2-diamino, N,N,N-trimethylpropane-1,3-diamino, cyclopentanamino, pyrrolyl, 2-(piperazin-1-yl)ethanol-1-ol, benzylamino.

[0008] Furthermore, the new compounds represented by general formulas I, II and III include N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamides and their derivatives, tautomers, and pharmaceutically acceptable salts, hydrates or solvates.

[0009] Furthermore, compounds of general formula I, or their pharmaceutically acceptable salts, hydrates, or solvates, are: N-[6-(4-methylpiperazin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(4-methylpiperidin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(N,N-dimethylamino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(N,N-diethyl-amino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-(6-{[2-(dimethylamino)ethyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-(6-{[3-(dimethylamino)propyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-[6-(5-methyl-2,5-diazahexyl-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(6-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-{6-[4-(2-hydroxyethyl)piperazin-1-yl]pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide; N-[6-(phenylamino)pyrimidin-4-yl]-2-(9-oxoylidene-10-yl)acetamide; N-[6-(benzylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

[0010] Furthermore, compounds of general formula II, or their pharmaceutically acceptable salts, hydrates, or solvates, are: N-[2-(4-methylpiperazin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(4-methylpiperidin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(N,N-dimethylamino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(N,N-diethyl-amino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-(2-{[2-(dimethylamino)ethyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-(2-{[3-(dimethylamino)propyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-[2-(5-methyl-2,5-diazahexyl-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(2-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-{2-[4-(2-hydroxyethyl)piperazin-1-yl]pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide; N-[2-(phenylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

[0011] Furthermore, compounds of general formula III, or their pharmaceutically acceptable salts, hydrates, or solvates, are: N-[6-(4-methylpiperazin-1-yl)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(methylthio)-6-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(4-methylpiperidin-1-yl)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(dimethylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(diethylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-(6-((2-(dimethylamino)ethyl)amino)-2-(methylthio)pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide N-(6-{[3-(dimethylamino)propyl]amino}-2-(methylthio)pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-[2-(methylthio)-6-(5-methyl-2,5-diazahex-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(methylthio)-6-(6-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(cyclopentylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxoylidene-10-yl)acetamide; N-[2-(methylthio)-6-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-{6-[4-(2-hydroxyethyl)piperazin-1-yl]-2-(methylthio)pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide; N-[6-(benzylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

[0012] The preparation method of the compounds represented by general formulas I, II and III of the present invention specifically includes the following steps: using anthranilic acid as the starting material, the compounds undergo substitution reaction, condensation reaction, alkylation reaction and hydrolysis reaction to generate the key intermediate acridinone acetic acid, and finally, in the presence of TCFH and NMI condensing agents, the compounds undergo a condensation reaction with substituted pyrimidines to obtain the target compounds represented by general formulas I, II and III.

[0013] Compared with the prior art, the beneficial effects of the present invention are as follows.

[0014] The preparation methods of the N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamide compounds represented by general formulas I, II and III provided by this invention are simple and feasible, and have shown good results in in vitro antitumor activity tests, especially with excellent antiproliferative ability against human breast cancer cells (BT-549, MDA-MB-231). Detailed Implementation

[0015] The present invention will be explained in detail through the following embodiments. These embodiments are merely illustrative and do not limit the scope of the present invention in any way.

[0016] Example 1.

[0017] Preparation of 2-(phenylamino)benzoic acid: o-aminobenzoic acid (1 g, 7.29 mmol) was placed in a 100 ml round-bottom flask with bromobenzene (1.72 g, 10.94 mmol), K2CO3 (3.02 g, 21.88 mmol), Cu (0.45 g, 7.29 mmol) and CuI (1.39 g, 7.29 mmol), and about 10 ml of DMF solvent was added. The mixture was heated to 120 °C and stirred for 12 h under nitrogen protection to obtain the crude product 2-(phenylamino)benzoic acid.

[0018] Example 2.

[0019] Preparation of 9,10-dihydroacridin-9-one: The crude product 2-(phenylamino)benzoic acid was added to concentrated sulfuric acid (98%) (1.5 g), heated to 100℃ and stirred for 2 h; after the reaction was completed, it was cooled to room temperature, water was added and allowed to stand to precipitate, filtered, and dried to obtain a white solid, namely 9,10-dihydroacridin-9-one.

[0020] Example 3.

[0021] Preparation of ethyl (9-oxoylide-8a,9,10,10a-tetrahydroacridin-10-yl) ethyl acetate: 1.5 g (7.68 mmol) of 9,10-dihydroacridin-9-one was placed in a 250 ml round-bottom flask, 15 ml of DMF was added, and the mixture was kept in an ice bath. Sodium hydride (60%) (1.54 g, 38.42 mmol) was added in portions, and the mixture was stirred for 1.5 h. Then, ethyl bromoacetate (2.57 g, 15.37 mmol) was added, and the mixture was heated to 30 °C and reacted for 12 h. After the reaction was complete, water was added, and the mixture was allowed to stand for 4 h before being filtered and dried to obtain a yellow-green solid, namely ethyl (9-oxoylide-8a,9,10,10a-tetrahydroacridin-10-yl) ethyl acetate.

[0022] Example 4.

[0023] Preparation of (9-oxoylidene-10-yl)acetic acid: Ethyl (9-oxoylidene-8a,9,10,10a-tetrahydroacryl-10-yl)acetate (1.5 g, 4.8 mmol) and NaOH (0.76 g, 19.20 mmol) were dissolved together in 15 ml of water:methanol = 1:1 and reacted at 50 °C for 4 h. After the reaction was completed, part of the methanol solvent was removed by vacuum distillation, and 50 ml of water was added. The pH was adjusted to 1-2 with 1 mol / L dilute hydrochloric acid, and a large amount of yellow solid precipitated. After standing for 2 h, the solid was filtered and dried to obtain the target compound acridinone acetic acid, namely (9-oxoylidene-10-yl)acetic acid.

[0024] Example 5.

[0025] The compound represented by general formula I was synthesized by the following preparation method.

[0026] (1) Preparation of N-(6-chloropyrimidin-4-yl)-2-(9-oxoylidene-10-yl)acetamide: 4-amino-6-chloropyrimidin (1 g, 7.72 mmol), (9-oxoylidene-10-yl)acetic acid (2.35 g, 9.26 mmol), N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH) (3.03 g, 10.81 mmol), and N-methylimidazolium (NMI) (1.96 g, 23.93 mmol) were placed in a 100 ml round-bottom flask, and about 20 ml of acetonitrile was added as solvent. The mixture was reacted at 65 °C for 12 h. After the reaction was complete, water was added, and the mixture was allowed to stand for 6 h before filtration and drying to obtain the target compound.

[0027] (2) Preparation of N-[6-(4-methylpiperazin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-a): N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol) and N-methylpiperazine (0.16 g, 1.64 mmol) were placed in a 100 ml round-bottom flask, 10 ml of DMF solvent was added, and triethylamine (0.28 g, 2.74 mmol) was added. The reaction mixture was heated to 80 °C and stirred for 6 h. After the reaction was completed, the mixture was cooled to room temperature, water was added, and the mixture was allowed to stand for 4 h. The mixture was then filtered, dried, and a deep yellow mixed solid was obtained. The solid was purified by acidic silica gel column chromatography to obtain the target compound.

[0028] 1H NMR (600 MHz, DMSO-d6) δ 2.17 (s, 3H), 2.32 (t, J = 5.0 Hz, 4H), 3.48 – 3.54 (m, 4H), 5.54 (s, 2H), 7.34 – 7.38 (m, 3H), 7.70 (d, J = 8.8 Hz,2H), 7.82 (ddd, J = 8.7, 6.9, 1.8 Hz, 2H), 8.35 – 8.41(m, 3H), 11.11 (s, 1H). 13 C NMR (150 MHz, DMSO-d6) δ 177.16, 168.36, 163.10, 158.11, 157.64, 142.87,134.70, 127.11, 122.02, 116.19, 89.83, 54.47, 49.34, 46.18, 45.97, 43.78. ESI-HRMS calcd. For C 24 H 24 N6O2[M+H] + 429.20, found: 429.19.

[0029] (3) Preparation of N-[6-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-b): Morpholine (0.16 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) were used as raw materials and the preparation method was the same as (2).

[0030] 1 H NMR (600 MHz, DMSO-d6) δ 3.42 – 3.50 (m, 4H), 3.56 – 3.64 (m, 4H), 5.53 (s, 2H), 7.35 (dd, J = 13.9, 6.4 Hz, 3H), 7.69 (d, J = 8.7 Hz, 2H), 7.82(ddd, J = 8.6, 6.8, 1.8 Hz, 2H), 8.32 – 8.43 (m, 3H), 11.11 (s, 1H). 13C NMR(150 MHz, DMSO-d6) δ 177.16, 168.38, 163.39, 158.09, 157.67, 142.88, 134.72,127.11, 122.05, 122.00, 116.20, 89.84, 66.17, 49.35, 44.32. ESI-HRMS calcd.forC 23 H 21 N5O3[M+H] + 416.16, found:416.16.

[0031] (4) Preparation of N-[6-(4-methylpiperidin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-c): Using 4-methylpiperidine (0.16 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0032] 1 H NMR (600 MHz, DMSO-d6) δ 0.85 (d, J = 6.0 Hz, 3H), 0.93 – 1.02 (m,2H), 1.54 – 1.64 (m, 3H), 2.81 (td, J = 12.8, 2.4 Hz, 2H), 4.12 – 4.28 (m,2H), 5.53 (s, 2H), 7.35 (dd, J= 14.9, 7.5 Hz, 3H), 7.70 (d, J = 8.9 Hz, 2H),7.82 (ddd, J = 8.8, 6.9, 1.8 Hz, 2H), 8.31 – 8.43 (m, 3H), 11.05 (s, 1H). 13 CNMR (150 MHz, DMSO-d6) δ 177.15, 168.31, 162.74, 158.18, 157.57, 142.87,134.71, 127.11, 122.03, 122.01,116.19, 89.58, 49.33, 44.33, 33.65, 30.88,22.05. ESI-HRMS calcd. for C 25 H 25 N5O2[M+H] +428.20, found:428.20.

[0033] (5) Preparation of N-[6-(N,N-dimethyl-amino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-d): Using dimethylamine (30% toluene solution) (0.25 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0034] 1 H NMR (600 MHz, DMSO-d6) δ 3.47 (t, J = 4.8 Hz, 3H), 3.60 (t, J = 4.7Hz, 3H), 5.54 (s, 2H), 7.30 – 7.40 (m, 3H), 7.69 (d, J = 8.8 Hz, 2H), 7.82(t, J = 7.7Hz, 2H), 8.29 – 8.48 (m, 3H), 11.12 (s, 1H). 13 C NMR (150 MHz, DMSO-d6) δ 177.16, 168.37, 163.38, 158.08, 157.66, 142.88, 134.71, 127.11, 122.04,122.00,116.19, 89.85, 49.35, 44.31. ESI-HRMS calcd. for C 21 H 19 N5O2[M+H] + 374.15, found: 374.16.

[0035] (6) Preparation of N-[6-(N,N-diethyl-amino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-e): Using diethylamine (0.16 g, 2.19 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0036] 1H NMR (600 MHz, DMSO-d6) δ 1.04 (t, J = 7.0 Hz, 6H), 3.41 (s, 4H), 5.52 (s, 2H), 7.18 (s, 1H), 7.36 (t, J = 7.4 Hz, 2H), 7.70 (d, J = 8.8 Hz,2H), 7.82 (ddd, J = 8.7, 6.8, 1.8 Hz, 2H), 8.33 (s, 1H), 8.37 (dd, J = 8.0,1.7 Hz, 2H), 11.01 (s, 1H). 13 C NMR (150 MHz, DMSO-d6) δ 177.14, 168.27,161.79, 158.14, 157.07, 142.87, 134.76, 127.12, 122.05, 122.01, 116.22,89.13,49.36, 42.18, 12.97. ESI-HRMS calcd. for C 23 H 23 N5O2[M+H] + 402.19, found: 402.18.

[0037] (7) Preparation of N-(6-{[2-(dimethylamino)ethyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (A6-f): Using N,N-dimethyl-1,2-ethylenediamine (0.15 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0038] 1 H NMR (600 MHz, DMSO-d6) δ 2.13 (s, 6H), 2.35 (t, J = 6.6 Hz, 2H), 3.34 – 3.48 (m, 2H), 5.50 (s, 2H), 7.09 (s, 1H), 7.36 (t, J = 7.4 Hz, 3H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7, 6.8, 1.8 Hz, 2H), 8.27 (s, 1H), 8.37 (dd, J = 8.0, 1.8 Hz, 2H), 10.90 (s, 1H).13 C NMR (150 MHz, DMSO) δ 183.25,177.19, 167.95, 163.90, 158.36, 142.91, 134.68, 127.08, 122.03, 116.24,58.47, 49.53, 45.64, 38.67. ESI-HRMS calcd. for C 23 H 24 N6O2[M+H] + 417.20, found:417.20.

[0039] (8) Preparation of N-(6-{[3-(dimethylamino)propyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (A6-g): Using N,N'-dimethyl-1,3-propanediamine (0.17 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0040] 1 H NMR (600 MHz, DMSO-d6) δ 1.60 (p, J = 7.1 Hz, 2H), 2.09 (s, 6H), 2.21 (t, J = 7.0 Hz, 2H), 3.25 – 3.49 (m, 2H), 5.50 (s, 2H), 7.07 (s, 1H),7.36 (t, J =7.4 Hz, 3H), 7.69 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7, 6.7,1.8 Hz, 2H), 8.27 (s, 1H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H), 10.90 (s, 1H). 13 CNMR (150 MHz, DMSO) δ 177.19, 167.97,163.95, 158.37, 142.91, 134.68, 127.09,122.02, 116.24, 91.45, 57.07, 49.50, 45.58, 27.31, 14.36. ESI-HRMS calcd. forC 24 H 26 N6O2[M+H] +431.21, found:431.22.

[0041] (9) Preparation of N-[6-(5-methyl-2,5-diazahex-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-h): Using N,N,N'-trimethylethylenediamine (0.17 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0042] 1 H NMR (600 MHz, DMSO-d6) δ 2.12 (s, 6H), 2.35 (t, J = 6.8 Hz, 2H), 2.92 (s, 3H), 3.65 (d, J = 64.4 Hz, 2H), 5.52 (s, 2H), 7.18 (s, 1H), 7.36 (t,J = 7.4 Hz, 2H), 7.70 (d, J = 8.8 Hz, 2H), 7.82 (ddd, J = 8.7, 6.9, 1.8 Hz,2H), 8.33 (s, 1H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H), 11.03(s, 1H). 13 C NMR (150MHz, DMSO) δ 177.15, 168.25, 162.92, 157.86, 157.12, 142.88, 134.73, 127.11,122.04, 122.02, 116.21, 89.22, 56.57, 49.42, 47.01, 45.87, 35.97. ESI-HRMScalcd. for C 24 H 26 N6O2[M+H] + 431.21, found:431.22.

[0043] (10) Preparation of N-[6-(6-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-i): Using N,N,N'-trimethyl-1,3-propanediamine (0.19 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0044] 1 H NMR (600 MHz, DMSO-d6) δ 1.76 (p, J = 7.0 Hz, 2H), 2.46 (s, 6H), 2.65 (t, J = 8.0 Hz, 2H), 2.92 (s, 3H), 3.55 (s,2H), 5.55 (s, 2H), 7.21 (s,1H), 7.36 (t, J = 7.4 Hz, 2H), 7.70 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7,6.9, 1.8 Hz, 2H), 8.34 – 8.41(m, 3H), 11.07 (s, 1H). 13 C NMR (150 MHz, DMSO) δ177.16, 168.29, 162.94, 157.88, 157.18, 142.88, 134.71, 127.13, 122.05,122.02, 116.19, 89.37, 55.55, 49.35, 46.67, 43.87, 35.76,23.65. ESI-HRMScalcd. for C 25 H 28 N6O2[M+H] + 445.23, found:445.23.

[0045] (11) Preparation of N-[6-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-k): Using tetrahydropyrrole (0.12 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0046] 1H NMR (600 MHz, DMSO-d6) δ 1.87 (s, 4H), 3.16 (s, 2H), 3.48 (s, 2H), 5.52 (s, 2H), 7.02 (s, 1H), 7.36 (t, J = 7.4 Hz,2H), 7.69 (d, J = 8.7 Hz,2H), 7.82 (ddd, J = 8.7, 6.9, 1.8 Hz, 2H), 8.31 (d, J = 1.1 Hz, 1H), 8.37(dd, J = 8.0, 1.7 Hz, 2H), 11.00 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.15,168.17,161.12, 158.04, 156.53, 142.89, 134.72, 127.11, 122.04, 122.01,116.21, 90.11, 55.38, 49.34, 25.23. ESI-HRMS calcd. for C 23 H 21 N5O2[M+H] + 400.17, found: 400.18.

[0047] (12) Preparation of N-{6-[4-(2-hydroxyethyl)piperazin-1-yl]pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide (A6-l): Using N-hydroxyethylpiperazine (0.28 g, 2.19 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0048] 1 H NMR (600 MHz, DMSO-d6) δ 2.41 (d, J = 20.2 Hz, 6H), 3.04 (s, 1H), 3.49 (q, J = 5.9 Hz, 6H), 5.53 (s, 2H), 7.29 – 7.41 (m, 3H), 7.70 (d, J = 8.8Hz, 2H), 7.82 (ddd, J =8.7, 6.8, 1.7 Hz, 2H), 8.37 (d, J = 6.5 Hz, 3H), 11.10(s, 1H). 13C NMR (150 MHz, DMSO) δ 177.16, 168.37,163.08, 158.11, 157.62,142.88, 134.71, 127.11, 122.02, 116.21, 89.78, 60.49, 58.83, 53.08, 49.35,45.93, 43.94. ESI-HRMS calcd. for C 25 H 26 N6O3[M+H] + 459.21, found:459.21.

[0049] (13) Preparation of N-[6-(phenylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-m): Using aniline as a raw material, aniline (0.13 g, 1.37 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol) and dilute hydrochloric acid (2 mol / L, 1.37 ml, 2.74 mmol) were placed in a 100 ml round-bottom flask, and about 15 ml of DMF solvent was added. The mixture was heated to 90 °C and stirred for 6 minutes. After the reaction was complete, the mixture was cooled to room temperature, water was added, and the mixture was allowed to stand for 4 hours to precipitate. The solid was then filtered, dried, and separated and purified using an acidic silica gel chromatography column to obtain the target compound N-[6-(phenylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

[0050] 1 H NMR (600 MHz, DMSO-d6) δ 5.54 (s, 2H), 6.94 – 7.01 (m, 1H), 7.24 –7.32 (m, 2H), 7.36 (t, J = 7.4 Hz, 2H), 7.47 (s, 1H),7.56 – 7.63 (m, 2H), 7.71 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7, 6.9, 1.8 Hz, 2H), 8.37 (dd, J =8.0, 1.7 Hz, 2H), 8.47 (d,J = 1.1 Hz, 1H), 9.60 (s, 1H), 11.16 (s, 1H). 13C NMR(150 MHz, DMSO-d6) δ 177.20, 168.26, 162.07, 158.34, 156.86, 142.92, 140.32,134.71, 129.18, 127.07, 122.86, 122.06,122.02, 120.51, 116.29, 93.38, 63.09.ESI-HRMS calcd. for C 25 H 19 N5O2[M+H] + 422.15, found:422.15.

[0051] (14) Preparation of N-[6-(benzylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A6-n): using benzylamine (0.18 g, 1.64 mmol), N-(6-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (2).

[0052] 1 H NMR (600 MHz, DMSO-d6) δ 2.98 (s, 2H), 4.52 (s, 1H), 5.51 (d, J =16.9 Hz, 2H), 6.98 – 7.35(m, 5H), 7.36 (t, J = 7.4 Hz, 2H), 7.69 (t, J = 8.8Hz, 2H), 7.81 (tt, J = 6.9, 2.3 Hz, 2H), 7.91 (t, J = 6.2 Hz, 1H), 8.31 (d, J= 24.6 Hz, 1H), 8.37 (dt, J =7.9, 2.1 Hz, 2H), 10.98 (d, J = 39.6 Hz, 1H). 13 CNMR (150 MHz, DMSO) δ 177.18, 168.25, 163.35, 157.82, 157.04, 142.90, 134.69, 128.75, 127.45, 127.12, 127.08, 122.04, 122.02, 116.24, 116.21, 89.25, 49.53,49.36. ESI-HRMS calcd. for C 26 H 21 N5O2[M+H]+ 436.17, found:436.18.

[0053] Example 6.

[0054] The compound represented by general formula II was synthesized by the following preparation method.

[0055] (15) Preparation of N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide: Using 4-amino-2-chloropyrimidin (1 g, 7.72 mmol), (9-oxomylidene-10-yl)acetic acid (2.35 g, 9.26 mmol), N,N,N',N'-tetramethylchloromethacinium hexafluorophosphate (TCFH) (3.03 g, 10.81 mmol) and N-methylimidazolium (NMI) (1.96 g, 23.93 mmol) as raw materials, the preparation method is the same as (1).

[0056] (16) Preparation of N-[2-(4-methylpiperazin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-a): Using N-methylpiperazine as the raw material, N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol) and N-methylpiperazine (0.16 g, 1.64 mmol) were placed in a 100 ml round-bottom flask, about 15 ml of DMF solvent was added, and triethylamine (0.28 g, 2.74 mmol) was added. The reaction mixture was heated to 80 °C and stirred for 6 h. After the reaction was complete, the mixture was cooled to room temperature, water was added, and the mixture was allowed to stand for 4 hours before being filtered and dried. The resulting dark yellow mixed solid was purified by acidic silica gel column chromatography to obtain the target compound N-[2-(4-methylpiperazin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

[0057] 1 H NMR (600 MHz, DMSO-d6) δ 2.23 (s, 3H), 2.37 (t, J = 5.1 Hz, 4H), 3.76 (d, J = 5.8 Hz, 4H), 5.51 (s, 2H), 7.14 (s, 1H), 7.36 (t, J = 7.5 Hz,2H), 7.67 (d, J = 8.7 Hz, 2H), 7.82 (t, J = 7.8 Hz, 2H), 8.24 (d, J = 5.5 Hz,1H), 8.33 – 8.40 (m, 2H),10.99 (s, 1H). 13C NMR (150 MHz, DMSO-d6) δ 177.18,168.28, 161.44, 159.94, 158.21, 142.86, 134.76, 127.10, 122.08, 122.00,116.22, 99.15, 54.87, 49.62, 46.33, 43.72. ESI-HRMScalcd. for C 24 H 24 N6O2[M+H] + 429.20, found: 429.19.

[0058] (17) Preparation of N-[2-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-b): Using morpholine (0.14 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0059] 1 H NMR (600 MHz, DMSO-d6) δ 3.69 (q, J = 4.9 Hz, 4H), 3.71 – 3.78 (m,4H), 5.53 (s, 2H), 7.16 – 7.20 (m, 1H), 7.36 (t, J = 7.4 Hz, 2H), 7.68 (d, J =8.7 Hz, 2H), 7.79 – 7.85 (m, 2H), 8.27 (d, J = 5.5 Hz, 1H), 8.34 – 8.39 (m,2H), 11.03 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.17, 168.33, 161.54, 159.94,158.26,142.87, 134.74, 127.10, 122.07, 122.01, 116.22, 99.52, 66.46, 49.66,44.38. ESI-HRMS calcd. for C 23 H 21 N5O3[M+H] + 416.16, found:416.17.

[0060] (18) Preparation of N-[2-(4-methylpiperidin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-c): Using 4-methylpiperidine (0.16 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0061] 1 H NMR (600 MHz, DMSO-d6) δ 0.94 (d, J = 6.1 Hz, 3H), 1.00 – 1.11 (m,2H), 1.62 – 1.73 (m, 3H), 2.88 (td, J = 12.6, 2.3 Hz, 2H), 4.68 (dt, J = 12.9,2.6 Hz, 2H), 5.52 (s, 2H), 7.11 (d, J = 5.5 Hz,1H), 7.36 (ddd, J = 7.8, 6.9,0.9 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7, 6.9, 1.7 Hz, 2H), 8.22 (d, J = 5.5 Hz, 1H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H), 10.95 (s, 1H). 13 C NMR(150 MHz, DMSO-d6) δ 177.17, 168.22, 161.33, 159.92, 158.22, 142.87, 134.73,127.10, 122.06, 122.01, 116.21, 98.62, 49.61, 43.98, 34.02, 31.19, 22.33.ESI-HRMS calcd. for C 25 H 25 N5O2[M+H] + 428.20, found:428.20.

[0062] (19) Preparation of N-[2-(N,N-dimethyl-amino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-d): Using dimethylamine (30% toluene solution) (0.25 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0063] 1 H NMR (600 MHz, DMSO-d6) δ 3.15 (s, 6H), 5.55 (s, 2H), 7.12 (s, 1H), 7.36 (t, J = 7.4 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7, 6.9,1.8 Hz, 2H), 8.23 ​​(d, J = 5.5 Hz, 1H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H), 10.95(s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.17, 168.30, 162.14, 159.70, 158.08,142.87, 134.73, 127.10, 122.05, 122.01, 116.23, 98.32, 49.64, 37.02. ESI-HRMScalcd. for C 21 H 19 N5O2[M+H] + 374.15, found:374.15.

[0064] (20) Preparation of N-[2-(diethylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-e): Using diethylamine (0.16 g, 2.19 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0065] 1H NMR (600 MHz, DMSO-d6) δ 1.16 (t, J = 6.9 Hz, 6H), 3.61 (q, J = 7.0Hz, 4H), 5.54 (d, J = 12.1 Hz, 2H), 7.09 (s, 1H),7.36 (ddt, J = 7.9, 6.9, 1.7Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.79 – 7.84 (m, 2H), 8.21 (d, J = 5.5 Hz, 1H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H), 10.89 (s, 1H). 13 C NMR (150 MHz, DMSO) δ177.17, 168.26, 160.85, 159.80, 158.25, 142.88, 134.73, 127.10, 122.16,122.05, 122.01, 116.23, 98.23, 49.64, 41.50, 13.60. ESI-HRMScalcd. forC 23 H 23 N5O2[M+H] + 402.19, found:402.19.

[0066] (21) Preparation of N-(2-{[2-(dimethylamino)ethyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (A2-f): Using N,N-dimethyl-1,2-ethylenediamine (0.15 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0067] 1H NMR (600 MHz, DMSO-d6) δ 2.20 (s, 6H), 2.44 (t, J = 6.8 Hz, 2H), 3.41 (q, J = 6.6 Hz, 3H), 5.55 (s, 2H), 6.84 (s, 1H),7.03 – 7.17 (m, 1H),7.36 (t, J = 7.4 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7, 6.9,1.8 Hz, 2H), 8.15 (d, J =5.6 Hz, 1H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H), 10.87(s, 1H). 13 C NMR (150 MHz, DMSO) δ177.17, 168.31,162.51, 159.92, 158.30,142.87, 134.73, 127.09, 122.05, 122.01, 116.24, 99.02, 58.64, 49.71, 45.75,37.41. ESI-HRMS calcd. for C 23 H 24 N6O2[M+H] + 417.20, found:417.20.

[0068] (22) Preparation of N-(2-{[3-(dimethylamino)propyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (A2-g): Using N,N'-dimethyl-1,3-propanediamine (0.17 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0069] 1H NMR (600 MHz, DMSO-d6) δ 1.79 (q, J = 7.1 Hz, 2H), 2.35 (s, 6H), 2.56 (t, J = 7.4 Hz, 2H), 3.35 (q, J = 6.6 Hz, 2H), 5.56 (s, 2H), 6.97 – 7.21(m, 2H), 7.36 (t, J = 7.5 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J =8.8, 6.9,1.8 Hz, 2H), 8.16 (d, J = 5.6 Hz, 1H), 8.37 (dd, J = 8.0, 1.7 Hz,2H), 10.92 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.17, 168.31, 162.54, 159.88,158.32, 142.87, 134.71, 127.10, 122.05, 122.01,116.23, 98.91, 56.70, 49.68,46.05, 44.62, 26.42. ESI-HRMS calcd. for C 24 H 26 N6O2[M+H] + 431.21, found:431.22.

[0070] (23) Preparation of N-[2-(5-methyl-2,5-diazahex-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-h): Using N,N,N'-trimethylethylenediamine (0.17 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0071] 1H NMR (600 MHz, DMSO-d6) δ 2.21 (s, 6H), 2.47 (t, J = 6.7 Hz, 2H), 3.13 (s, 3H), 3.73 (t, J = 6.8 Hz, 2H), 5.56 (s, 2H), 7.02 – 7.16 (m, 1H),7.36 (t, J = 7.4 Hz, 2H), 7.68 (d, J = 8.8 Hz, 2H), 7.82 (ddd, J = 8.7, 6.9,1.8 Hz, 2H), 8.22 (d, J =5.5 Hz, 1H), 8.37 (dd, J = 7.9, 1.7 Hz, 2H), 10.90(s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.17, 168.31,161.68, 159.70, 158.11,142.88, 134.72, 127.10, 122.04, 122.01, 116.24, 98.36, 56.92, 49.70, 46.65,45.93, 35.50. ESI-HRMS calcd. for C 24 H 26 N6O2[M+H] + 431.21, found: 431.22.

[0072] (24) Preparation of N-[6-(2-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-i): Using N,N,N'-trimethyl-1,3-propanediamine (0.19 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0073] 1H NMR (600 MHz, DMSO-d6) δ 1.80 (t, J = 7.4 Hz, 2H), 2.34 (s, 6H), 2.98 (q, J = 7.0 Hz, 2H), 3.13 (s, 3H), 3.64 (t, J =7.1 Hz, 2H), 5.56 (s,2H), 7.11 (s, 1H), 7.36 (t, J = 7.4 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82(ddd, J = 8.7, 6.8, 1.7 Hz, 2H), 8.23 ​​(d, J = 5.5 Hz, 1H),8.37 (dd, J = 8.0,1.7 Hz, 2H), 10.93 (s, 1H). 13 C NMR (150 MHz, DMSO-d6) δ 177.17, 168.32,161.67, 159.70, 158.17, 142.87, 134.72, 127.11, 122.06, 122.01, 116.22,98.44,56.42, 49.68, 46.79, 46.20, 35.41, 24.62. ESI-HRMS calcd. for C 25 H 28 N6O2[M+H] + 445.23, found:445.23.

[0074] (25) Preparation of N-[2-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-k): Using tetrahydropyrrole (0.12 g, 1.64 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0075] 1H NMR (600 MHz, DMSO-d6) δ 1.99 (d, J = 6.0 Hz, 4H), 3.56 (d, J = 5.9Hz, 4H), 5.61 (s, 2H), 7.09 – 7.20 (m, 1H), 7.40 (t, J = 7.4 Hz, 2H), 7.72(d, J = 8.7Hz, 2H), 7.82 – 7.89 (m, 2H), 8.25 (d, J = 5.5 Hz, 1H), 8.38 –8.45 (m, 2H), 10.99 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.18, 168.35,160.32,159.67, 158.13, 142.87, 134.71, 127.10, 122.04, 122.01, 116.22, 98.36, 49.65,46.81, 25.49. ESI-HRMS calcd. for C 23 H 21 N5O2[M+H] + 400.17, found:400.18.

[0076] (26) Preparation of N-{2-[4-(2-hydroxyethyl)piperazin-1-yl]pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide (A2-l): Using N-hydroxyethylpiperazine (0.28 g, 2.19 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol), and triethylamine (0.28 g, 2.74 mmol) as raw materials, the preparation method is the same as (16).

[0077] 1 H NMR (600 MHz, DMSO-d6) δ 2.42 (d, J = 21.8 Hz, 6H), 2.98 – 3.07 (m,1H), 3.50 (q, J = 5.6 Hz, 6H), 5.55 (s, 2H), 7.35 (dd, J = 15.4, 7.9 Hz, 3H), 7.70 (d, J = 8.7 Hz, 2H), 7.82 (t, J = 7.9 Hz, 2H), 8.38 (d, J = 7.7 Hz, 3H), 11.11 (s, 1H). 13C NMR (150 MHz, DMSO) δ 177.17, 168.27, 161.45, 159.91, 158.21,142.87, 134.73, 127.10, 122.06, 122.02, 116.22,99.14, 60.78, 60.22, 53.47,49.64, 43.85. ESI-HRMS calcd. for C 25 H 26 N6O3[M+H] + 459.21, found:459.21.

[0078] (27) Preparation of N-[2-(phenylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (A2-m): Using aniline as a raw material, aniline (0.13 g, 1.37 mmol), N-(2-chloropyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.55 mmol) and dilute hydrochloric acid (2 mol / L, 1.37 ml, 2.74 mmol) were mixed in a 100 ml round-bottom flask, 15 mL of DMF solvent was added, the temperature was raised to 90 °C and stirred for 6 h. After the reaction was completed, the temperature was lowered to room temperature, water was added and allowed to stand for 4 h to precipitate, the mixture was filtered, dried to obtain a mixed solid, and purified by acidic silica gel chromatography column to obtain the target compound.

[0079] 1 H NMR (600 MHz, DMSO-d6) δ 5.57 (d, J = 19.8 Hz, 2H), 6.98 (t, J =7.3 Hz, 1H), 7.35 (dt, J = 17.9, 7.8 Hz, 5H), 7.71(d, J = 9.0 Hz, 2H), 7.78 –7.91 (m, 4H), 8.32 – 8.44 (m, 3H), 9.52 (s, 1H), 11.07 (s,1H). 13 C NMR (150MHz, DMSO) δ 177.18, 168.48, 160.19, 160.08, 158.08, 142.89, 140.95, 134.76,128.98, 127.11,122.08, 122.02, 121.92, 119.44, 116.28, 101.16. ESI-HRMScalcd. for C 25 H 19 N5O2[M+H] +422.15, found:422.15.

[0080] Example 7.

[0081] The compound represented by general formula III was synthesized by the following preparation method.

[0082] (28) Preparation of N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide: The preparation method was the same as (1), using 2-methylthio-4-amino-6-chloropyrimidine (1.0 g, 5.69 mmol), (9-oxoylidene-10-yl)acetic acid (1.73 g, 6.83 mmol), N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH) (3.03 g, 7.97 mmol) and N-methylimidazolium (NMI) (1.45 g, 17.65 mmol) as raw materials.

[0083] (29) Preparation of N-[6-(4-methylpiperazin-1-yl)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-a): Using N-methylpiperazine as a raw material, N-methylpiperazine (0.14 g, 1.46 mmol) and N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol) were mixed and dissolved in 10 ml of DMF. Triethylamine (0.25 g, 2.43 mmol) was added, and the mixture was heated to 65 °C and stirred for 6 h. After the reaction was completed, the mixture was cooled to room temperature, and water was added to precipitate a solid. The mixture was allowed to stand for 4 hours. h, filter and dry to obtain a mixed solid, which is then separated and purified by acid silica gel chromatography column to obtain the target compound N-[6-(4-methylpiperazin-1-yl)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

[0084] 1 H NMR (600 MHz, DMSO-d6) δ 2.18 (s, 3H), 2.32 (t, J = 5.0 Hz, 4H), 2.48 (s, 3H), 3.50 (s, 4H), 5.51 (s, 2H), 7.06 (s,1H), 7.36 (t, J = 7.4 Hz,2H), 7.68 (d, J = 8.7 Hz, 2H), 7.81 (ddd, J = 8.8, 6.8, 1.8 Hz, 2H), 8.36(dd, J = 7.8, 1.7 Hz, 2H), 11.07 (s, 1H).13 C NMR (150 MHz, DMSO-d6) δ 177.15,170.04, 168.31, 162.85, 157.51, 142.86, 134.73, 127.11, 122.05, 121.99,116.20, 85.71, 54.47, 49.33, 45.98, 43.88, 13.72. ESI-HRMS calcd. forC 25 H 26 N6O2S[M+H] + 475.18, found:475.18.

[0085] (30) Preparation of N-[2-(methylthio)-6-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-b): Using morpholine (0.13 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0086] 1 H NMR (600 MHz, DMSO-d6) δ 2.48 (s, 3H), 3.48 (s, 4H), 3.59 (t, J =4.8 Hz, 4H), 5.51 (s, 2H), 7.06 (s, 1H), 7.36 (t, J= 7.4 Hz, 2H), 7.68 (d, J= 8.7 Hz, 2H), 7.81 (ddd, J = 8.6, 6.8, 1.8 Hz, 2H), 8.36 (dd, J = 8.0, 1.8Hz, 2H), 11.09 (s, 1H). 13 C NMR (150 MHz, DMSO-d6) δ 177.15, 170.04, 168.31,163.11, 157.54, 142.86, 134.72, 127.11, 122.05, 122.00, 116.19, 85.70,66.17,49.35, 44.39, 13.72. ESI-HRMS calcd. for C 24 H 23 N5O3S[M+H] + 462.15, found:462.15.

[0087] (31) Preparation of N-[6-(4-methylpiperidin-1-yl)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-c): Using 4-methylpiperidine (0.14 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0088] 1 H NMR (600 MHz, DMSO-d6) δ 0.86 (d, J = 6.0 Hz, 3H), 0.91 – 1.04 (m,2H), 1.52 – 1.61 (m, 2H), 1.63 (d, J = 3.8 Hz, 1H), 2.47 (s, 3H), 2.82 (td, J= 12.8, 2.4 Hz, 2H), 4.18 (s, 2H), 5.51 (s, 2H), 7.05(s, 1H), 7.36 (t, J =7.4 Hz, 2H), 7.69 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.8, 6.9, 1.8 Hz, 2H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H), 11.03 (s, 1H). 13 C NMR (150 MHz, DMSO) δ177.14, 169.99,168.26, 162.47, 157.46, 142.86, 134.71, 127.11, 122.03,122.00, 116.19, 85.52, 49.33, 44.45, 33.68, 30.84, 22.03, 13.71. ESI-HRMScalcd. for C 26 H 27 N5O2S[M+H] + 474.19, found:474.20.

[0089] (32) Preparation of N-[6-(dimethylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-d): Dimethylamine (30% toluene solution) (0.22 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) were used as raw materials, and the preparation method was the same as (29).

[0090] 1 H NMR (600 MHz, DMSO-d6) δ 2.49 (s, 3H), 2.98 (s, 6H), 5.51 (s, 2H), 6.93 (s, 1H), 7.36 (t, J = 7.4 Hz, 2H), 7.69 (d, J= 8.8 Hz, 2H), 7.82 (ddd, J= 8.6, 7.0, 1.8 Hz, 2H), 8.37 (dd, J= 8.0, 1.7 Hz, 2H), 11.02 (s, 1H). 13 C NMR(150 MHz, DMSO) δ 177.14, 169.59, 168.19, 163.15, 156.92, 142.87, 134.73,127.11, 122.04, 122.00, 116.20, 85.23, 49.35, 37.20, 13.66. ESI-HRMS calcd.forC 22 H 21 N5O2S[M+H] + 420.14, found: 420.15.

[0091] (33) Preparation of N-[6-(diethylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-e): Diethylamine (0.14 g, 1.95 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) were used as raw materials, and the preparation method was the same as (29).

[0092] 1H NMR (600 MHz, DMSO-d6) δ 1.05 (t, J = 7.0 Hz, 6H), 2.47 (s, 3H), 3.40 (s, 4H), 5.50 (s, 2H), 6.92 (s, 1H), 7.32 – 7.41 (m, 2H), 7.69 (d, J =8.7 Hz, 2H), 7.82 (ddd, J =8.7, 6.9, 1.7 Hz, 2H), 8.37 (dd, J = 8.0, 1.8 Hz, 2H), 11.01 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.14, 169.79,168.20, 161.61,156.99, 142.86, 134.74, 127.12, 122.04, 122.01, 116.20, 85.11, 49.35, 42.42,13.62, 13.03. ESI-HRMS calcd. for C 24 H 25 N5O2S[M+H]+448.17, found:448.18.

[0093] (34) Preparation of N-(6-{[2-(dimethylamino)ethyl]amino}-2-(methylthio)pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (AS-f): Using N,N-dimethyl-1,2-ethylenediamine (0.14 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0094] 1 H NMR (600 MHz, DMSO-d6) δ 2.14 (s, 6H), 2.34 (t, J = 6.7 Hz, 2H), 2.47 (s, 3H), 3.38 (dd, J = 13.1, 5.9 Hz, 2H), 5.47(s, 2H), 6.81 (s, 1H),7.36 (t, J = 7.4 Hz, 2H), 7.43 (s, 1H), 7.67 (d, J = 8.8Hz, 2H), 7.78 – 7.85(m, 2H), 8.33 – 8.40 (m, 2H), 10.86 (s, 1H).13 C NMR (150 MHz, DMSO) δ 183.28,177.18, 169.96, 167.87, 163.63,142.90, 134.69, 127.08, 122.04, 122.01,116.24, 89.00, 58.52, 52.47, 49.53, 45.68, 13.58. ESI-HRMS calcd. forC 24 H 26 N6O2S[M+H] + 463.18, found:463.19.

[0095] (35) Preparation of N-(6-{[3-(dimethylamino)propyl]amino}-2-(methylthio)pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide (AS-g): Using N,N'-dimethyl-1,3-propanediamine (0.15 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0096] 1 H NMR (600 MHz, DMSO-d6) δ 1.61 (q, J = 7.0 Hz, 2H), 2.11 (s, 6H), 2.23 (d, J = 6.6 Hz, 2H), 2.47 (s, 3H), 3.31 (s,2H), 5.48 (s, 2H), 6.80 (s,1H), 7.29 – 7.50 (m, 3H), 7.67 (d, J = 8.8 Hz, 2H), 7.82 (ddd, J = 8.8, 6.8,1.8 Hz, 2H), 8.36(dd, J = 8.0, 1.8 Hz, 2H), 10.86 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.18, 170.00, 167.88, 163.61, 155.60, 142.90, 134.69, 127.08,122.04, 122.01, 116.23, 88.14, 57.04, 49.51, 45.51, 38.80, 27.28,13.58. ESI-HRMS calcd. for C 25H 28 N6O2S[M+H] + 477.20, found: 477.21.

[0097] (36) Preparation of N-[2-(methylthio)-6-(5-methyl-2,5-diazahex-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-h): Using N,N,N'-trimethylethylenediamine (0.15 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0098] 1 H NMR (600 MHz, DMSO-d6) δ 2.13 (s, 6H), 2.36 (t, J = 6.8 Hz, 2H), 2.48 (s, 3H), 2.90 (s, 3H), 3.65 (s, 2H), 5.50 (s,2H), 6.91 (s, 1H), 7.36(ddd, J = 7.8, 6.9, 0.8 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J =8.7, 6.9, 1.8 Hz, 2H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H),11.02 (s, 1H). 13 C NMR(150 MHz, DMSO) δ 177.15, 169.59, 168.19, 162.74, 157.02, 142.86, 134.73,127.11, 122.05, 122.01,116.20, 85.18, 56.49, 49.40, 47.09, 45.85, 35.98,13.63. ESI-HRMS calcd. for C 25 H 28 N6O2S[M+H] + 477.20, found:477.21.

[0099] (37) Preparation of N-[2-(methylthio)-6-(6-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-i): Using N,N,N'-trimethyl-1,3-propanediamine (0.17 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0100] 1 H NMR (600 MHz, DMSO-d6) δ 1.55 – 1.65 (m, 2H), 2.05 (s, 6H), 2.14(t, J = 6.9 Hz, 2H), 2.48 (s, 3H), 2.92 (s, 3H), 3.57 (d, J = 52.7 Hz, 2H),5.50 (s, 2H), 6.92 (s,1H), 7.36 (t, J = 7.4 Hz, 2H), 7.68 (d, J = 8.7 Hz,2H), 7.82 (ddd, J = 8.7, 6.9, 1.8 Hz, 2H), 8.37 (dd, J = 8.0, 1.7 Hz, 2H),11.00 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.15, 169.60,168.15, 162.65, 156.96,142.87, 134.73, 127.11, 122.05, 122.01, 116.21, 85.22, 56.75, 49.38, 47.64,45.56, 35.92, 25.18, 13.62. ESI-HRMS calcd. for C 26 H 30 N6O2S[M+H] + 491.22, found:491.22.

[0101] (38) Preparation of N-[6-(cyclopentylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-j): Using cyclopentylamine (0.12 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0102] 1 H NMR (600 MHz, DMSO-d6) δ 1.30 – 1.56 (m, 4H), 1.62 (s, 2H), 1.86(s, 2H), 2.47 (s, 3H), 2.98 (s, 1H), 5.49 (d, J = 13.3 Hz, 2H), 6.81 (s, 1H),7.30 – 7.48 (m, 3H), 7.68 (t, J = 7.7 Hz, 2H), 7.82 (ddd, J =8.7, 6.9, 1.8Hz, 2H), 8.36 (dd, J = 8.0, 1.8 Hz, 2H), 10.85 (s, 1H). 13 C NMR (150 MHz, DMSO)δ 177.18, 169.89,163.16, 162.34, 156.92, 142.90, 134.69, 127.11, 127.08,122.04, 122.01, 116.24, 85.23, 49.54, 32.74, 23.89, 13.66, 13.60. ESI-HRMScalcd. for C 25 H 25 N5O2S[M+H] + 460.17, found:460.18.

[0103] (39) Preparation of N-[2-(methylthio)-6-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-k): Using tetrahydropyrrole (0.10 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0104] 1H NMR (600 MHz, DMSO-d6) δ 1.86 (s, 4H), 2.48 (s, 3H), 3.10 – 3.23(m, 2H), 3.50 (s, 2H), 5.50 (s, 2H), 6.76 (s, 1H), 7.36 (t, J = 7.4 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J = 8.7, 6.9, 1.8 Hz, 2H), 8.37 (dd, J =8.0, 1.7 Hz, 2H), 10.99 (s, 1H). 13 C NMR (150 MHz, DMSO) δ 177.15, 169.68,168.11, 160.92, 156.37, 142.87, 134.72, 127.11, 122.04, 122.00, 116.20,86.15, 49.34, 24.95, 13.62. ESI-HRMS calcd. forC 24 H 23 N5O2S[M+H] + 446.16, found:446.18.

[0105] (40) Preparation of N-{6-[4-(2-hydroxyethyl)piperazin-1-yl]-2-(methylthio)pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide (AS-l): Using N-hydroxyethylpiperazine (0.25 g, 1.95 g), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0106] 1H NMR (600 MHz, DMSO-d6) δ 2.40 (dd, J = 14.9, 7.0 Hz, 6H), 2.48 (s,3H), 3.49 (q, J = 6.0, 5.5 Hz, 6H), 4.43 (s, 1H),5.51 (s, 2H), 7.05 (s, 1H),7.36 (ddd, J = 7.9, 6.9, 0.8 Hz, 2H), 7.68 (d, J = 8.7 Hz, 2H), 7.82 (ddd, J= 8.7, 6.9, 1.8 Hz, 2H), 8.37 (dd, J = 8.0, 1.7 Hz,2H), 11.07 (s, 1H). 13 C NMR(150 MHz, DMSO) δ 177.15, 170.02, 168.30, 162.81, 157.49, 142.86, 134.72,127.10, 122.04, 122.00, 116.20, 85.66, 60.48, 58.84, 53.09, 49.34, 46.09,13.72. ESI-HRMS calcd. for C 26 H 28 N6O3S[M+H] + 505.19, found:505.20.

[0107] (41) Preparation of N-[6-(benzylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (AS-n): using benzylamine (0.16 g, 1.46 mmol), N-[6-chloro-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide (0.2 g, 0.48 mmol), and triethylamine (0.25 g, 2.43 mmol) as raw materials, the preparation method is the same as (29).

[0108] 1H NMR (600 MHz, DMSO-d6) δ 2.43 (s, 3H), 2.98 (s, 2H), 4.52 (s, 1H), 5.48 (d, J = 16.5 Hz, 2H), 6.91 (d, J = 14.4 Hz,1H), 7.25 (dt, J = 23.5, 7.7Hz, 4H), 7.36 (d, J = 7.4 Hz, 2H), 7.67 (t, J = 9.2 Hz, 2H), 7.81 (ddd, J =8.8, 7.0, 1.9 Hz, 2H), 8.01 (s, 1H), 8.36 (dt, J =7.9, 2.0 Hz, 2H), 10.96 (d,J = 42.2 Hz, 1H). 13 C NMR (150 MHz, DMSO) δ 177.17, 169.27, 168.19, 163.16,156.92,142.88, 134.74, 134.70, 128.75, 127.60, 127.21, 127.08, 122.05,122.01, 116.23, 85.24, 49.53, 49.07, 13.60. ESI-HRMS calcd. for C27H23N5O2S[M+H] + 482.16, found:482.16.

[0109] Cytotoxicity evaluation of the compound in Example 8.

[0110] Screening method: MTT (microtetrozolium) reduction method; Cell lines: BT-549, MDA-MB-231.

[0111] Test method: (1) Cell culture: Remove the cryovials from the -80℃ freezer and thaw them in a 37℃ water bath. After the cryopreservation solution has completely thawed, centrifuge at 1000 r / min for 5 min at room temperature. Discard the cryopreservation solution, add complete culture medium and transfer to cell culture flasks. Culture MDA-MB-231 and BT-549 cells in a 37℃, 5% CO2 incubator using DMEM medium containing 10% FBS and RPMI-1640, respectively. Perform experiments when the cells reach the logarithmic growth phase.

[0112] (2) Cell seeding: Wash cells with PBS, digest cells with trypsin, resuspend cells, and count cells using a hemocytometer to obtain the cell concentration of the cell suspension. Then, dilute the cell suspension to a specific concentration using complete culture medium. Sow cells at a concentration of 1×10⁻⁶. 4 Up to 5×10 4 Cells were seeded at a density of 100 μL of culture medium per well in 96-well plates. The cells were incubated at 37°C and 5% CO2 for 12 h to allow them to adhere.

[0113] (3) Compound treatment: The target compounds were prepared at different concentrations (0.1 μM, 1 μM, 10 μM, 100 μM, 1000 μM). Each treatment group included 3 parallel wells, and 100 μl of the compound treatment solution was added to each well. The blank control group was added only with culture medium, and the negative control group was added with the same volume of DMSO. The mixture was incubated in an incubator for 48 h.

[0114] (4) MTT assay: After incubation for 48 h, add 10 μL of MTT (thiazolyl blue) solution (5 mg / mL) and continue incubation for 4 h. After incubation, discard the culture medium and add 150 μL of DMSO to dissolve the blue-purple crystalline formazan. Measure the absorbance (OD value) of each well at 490 nm using a microplate reader.

[0115] (5) Data analysis: Cell viability for each experimental group was calculated using the following algorithm with Graphpad Prism9 software: Record the drug concentration and cell viability. Plot a concentration-response curve with the logarithm of the drug concentration as the independent variable X and cell viability as the dependent variable Y, and calculate the IC50. 50 value.

[0116] All the compounds in the examples exhibited good cytotoxicity against BT549 and MDA-MB-231 cells, with most compounds showing IC50 cytotoxicity against both BT549 and MDA-MB-231 cells. 50 <100 μM, IC50 of all example compounds and control compound CX-3453 against BT549 and MDA-MB-231 cytotoxicity. 50 The data is shown in Table 1.

[0117] Table 1. Cytotoxicity of the compounds.

[0118] Example 9 Compound Pair c-Myc Stability of the G4 structure (FRET experiment).

[0119] (1) Preparation of probe solution: Dilute the Pu27 probe from the original solution to the required concentration (400 nM) using Tris-HCl buffer (10 mM, pH = 7.2) containing KCl (2 μM).

[0120] (2) Probe and compound mixing: Mix 10 μL of dual probe Pu27 solution (400 nM) with 10 μL of compound solution (2 μM). Heat the mixture at 90 °C for 5 minutes, then cool it to room temperature using a thermal cycler. The annealing process ensures proper pairing of the probe and the target sequence. Add the final 20 μL of solution to the capillary of a LightCycler, ready for FRET measurements.

[0121] (3) FRET Measurement: Fluorescence detection was performed using a real-time fluorescence PCR instrument. The donor fluorophore (FAM) was excited using a 470 nm excitation source, and the emission signal of the acceptor fluorophore (TAMRA) was detected at a wavelength of 530 nm. Fluorescence signals were recorded every 1 °C at a heating rate of 1 °C / min within a temperature range of 37–99 °C. A constant temperature was maintained for 30 seconds before each measurement to ensure stable fluorescence values. Measurements were repeated three times for each treatment group.

[0122] (4) Data Analysis: The experimental data were finally analyzed using Origin 6.0 software. The fluorescence intensity as a function of temperature (ΔT) was analyzed. m Using Origin fitting, the corresponding ΔT is when the fluorescence intensity is 0.5. m value.

[0123] The effects of synthesized compounds on [the target / the environment] were studied using FRET melting experiments. c-Myc This study examines the binding affinity of the G4 structure in the promoter region and changes in its thermal stability. FRET is a rapid, effective, and reproducible semi-quantitative method for evaluating ligand-G4 interactions. The principle behind this method is to determine the stabilizing effect of the ligand on the G4 structure by observing the temperature rise during G4 unwinding after the addition of the ligand molecule. The G4 sequence consists of a fluorescent donor (FAM) and a fluorescent acceptor (TAMRA) group at both ends of the DNA strand. When the sequence successfully forms the G4 structure, the donor and acceptor groups are close together. The donor group is excited by a light source and fluoresces, which is absorbed by the acceptor group. At this point, the detector can detect the fluorescence emitted by the acceptor group but cannot detect the fluorescence excited by the donor group. As the temperature increases, the G4 structure gradually unwinds, and the donor and acceptor groups move away. The fluorescence of the acceptor group is then undetectable. This process is recorded by the instrument, yielding the FRET melting curve and ΔTm value of G4. m It is the temperature at which 50% of the G4 structure is unwound, and this is used to assess the stabilizing effect of small molecules on the G4 structure.

[0124] All examples of acridinone derivatives can improve c-Myc Thermal stability of the promoter region G4 structure (FPu27T), ΔT m The values ​​range from 0.03℃ to 4.7℃. ΔT values ​​for all example compounds and the control compound CX-3543 are shown. m The data is shown in Table 2.

[0125] Table 2. Melting range determined in the FRET melting experiment.

Claims

1. N-methyl-2-(9-oxoacridin-10(9H)-yl)acetamide compounds, characterized in that, The compounds are shown in general formulas I, II and III: in: Each R1 is independently methylpiperazinyl, morpholinyl, methylpiperidinyl, dimethylamino, N,N-diethylamino, N,N-dimethylethane-1,2-diamino, N,N-dimethylpropane-1,3-diamino, N,N,N-trimethylethane-1,2-diamino, N,N,N-trimethylpropane-1,3-diamino, pyrrolyl, 2-(piperazin-1-yl)ethanol-1-olyl, aniline, benzylamino; Each R2 is independently methylpiperazinyl, morpholinyl, methylpiperidinyl, dimethylamino, N,N-diethylamino, N,N-dimethylethane-1,2-diamino, N,N-dimethylpropane-1,3-diamino, N,N,N-trimethylethane-1,2-diamino, N,N,N-trimethylpropane-1,3-diamino, pyrrolyl, 2-(piperazin-1-yl)ethanol-1-olyl, aniline; Each R3 is independently methylpiperazinyl, morpholinyl, methylpiperidinyl, dimethylamino, N,N-diethylamino, N,N-dimethylethane-1,2-diamino, N,N-dimethylpropane-1,3-diamino, N,N,N-trimethylethane-1,2-diamino, N,N,N-trimethylpropane-1,3-diamino, cyclopentanamino, pyrrolyl, 2-(piperazin-1-yl)ethanol-1-ol, benzylamino.

2. The compound according to claim 1, characterized in that, The compounds of general formula I, or their pharmaceutically acceptable salts, hydrates, or solvates, are: N-[6-(4-methylpiperazin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(4-methylpiperidin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(N,N-dimethylamino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(N,N-diethyl-amino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-(6-{[2-(dimethylamino)ethyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-(6-{[3-(dimethylamino)propyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-[6-(5-methyl-2,5-diazahexyl-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(6-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-{6-[4-(2-hydroxyethyl)piperazin-1-yl]pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide; N-[6-(phenylamino)pyrimidin-4-yl]-2-(9-oxoylidene-10-yl)acetamide; N-[6-(benzylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

3. The compound according to claim 1, characterized in that, The compounds of general formula II, or their pharmaceutically acceptable salts, hydrates, or solvates, are: N-[2-(4-methylpiperazin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(4-methylpiperidin-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(N,N-dimethylamino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(N,N-diethyl-amino-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-(2-{[2-(dimethylamino)ethyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-(2-{[3-(dimethylamino)propyl]amino}pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-[2-(5-methyl-2,5-diazahexyl-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(2-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-{2-[4-(2-hydroxyethyl)piperazin-1-yl]pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide; N-[2-(phenylamino)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

4. The compound according to claim 1, characterized in that, The compounds of general formula III, or their pharmaceutically acceptable salts, hydrates, or solvates, are: N-[6-(4-methylpiperazin-1-yl)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(methylthio)-6-(morpholin-4-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(4-methylpiperidin-1-yl)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(dimethylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(diethylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-(6-((2-(dimethylamino)ethyl)amino)-2-(methylthio)pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide N-(6-{[3-(dimethylamino)propyl]amino}-2-(methylthio)pyrimidin-4-yl)-2-(9-oxomylidene-10-yl)acetamide; N-[2-(methylthio)-6-(5-methyl-2,5-diazahex-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[2-(methylthio)-6-(6-methyl-2,6-diazahept-2-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-[6-(cyclopentylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxoylidene-10-yl)acetamide; N-[2-(methylthio)-6-(pyrrolidone-1-yl)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide; N-{6-[4-(2-hydroxyethyl)piperazin-1-yl]-2-(methylthio)pyrimidin-4-yl}-2-(9-oxomylidene-10-yl)acetamide; N-[6-(benzylamino)-2-(methylthio)pyrimidin-4-yl]-2-(9-oxomylidene-10-yl)acetamide.

5. A pharmaceutical composition comprising the compound of claim 1 or a derivative thereof, a tautomer, and a pharmaceutically acceptable salt, hydrate, or solvate.

6. A method for preparing the compound as claimed in claim 1, characterized in that, Specifically, the process includes the following steps: using anthranilic acid as the starting material, the intermediate acridinone acetic acid is generated through substitution reaction, condensation reaction, alkylation reaction, and hydrolysis reaction. Finally, in the presence of TCFH and NMI condensing agents, it undergoes a condensation reaction with a substituted pyrimidine to obtain the target compound shown.

7. The use of the compound according to any one of claims 1-4 or the composition according to claim 5 in the preparation of an antitumor drug.

Images