Prmt7 target spot inhibitor and application thereof

Abstract

The application discloses a PRMT7 target point inhibitor and application thereof, wherein the compound is shown in any one of formulas (II)-(V), and the application discloses a plurality of compounds with novel structures, which can inhibit PRMT7 and further inhibit tumors, have the advantages of high activity and good treatment effect, and can be used for preparing antitumor drugs.

Description

[0001] This invention is a divisional application of the invention entitled "A PRMT7 target inhibitor and its application", application number: 2024112753072, application date: 2024.9.12. Technical Field

[0002] This invention relates to a PRMT7 target inhibitor and its application, belonging to the field of medicinal chemistry. Background Technology

[0003] Protein arginine methylation has attracted much attention due to its mediated cellular biological processes and its involvement in physiological and disease processes. As an enzymatic reaction of this post-translational modification, protein arginine methyltransferases (PRMTs) have gradually become important targets of interest in the field of drug discovery.

[0004] Nine sequence-related PRMT subtypes (PRMT1-9) have been identified in mammals, classified into three classes (Type I, Type II, and Type III) based on their methylation products. Type I PRMTs (PRMT1, PRMT2, PRMT3, PRMT4, PRMT6, and PRMT8) catalyze the formation of monomethylarginine (Rme1) and asymmetric dimethylarginine (Rme2a), Type II PRMTs (PRMT5 and PRMT9) catalyze the formation of Rme1 and symmetric dimethylarginine (Rme2s), while Type III PRMT (PRMT7) catalyzes only the formation of Rme1. The nine members of the PRMT protein family share a common Rossmann-like structure, comprising seven β-sheets linked by α-helices and a β-barrel domain. They, along with some non-SET domain lysine methyltransferases (e.g., DOT1L), are classified as class I S-adenosylmethionine (SAM)-dependent methyltransferases, which share the same basic domain. However, little is known about these methyltransferases. Except for PRMT7 and PRMT9, other PRMTs contain only one methyltransferase domain, while these two methyltransferases each contain two tandem domains generated by a repeating ancestral gene. The single C-terminal domain has no catalytic activity, but the pseudodimer formed by folding together with the N-terminal domain is essential for enzyme activity.

[0005] PRMT7 plays a vital biological role in normal physiological processes, and its dysregulation is closely related to the development and progression of various cancers. In breast cancer, high expression of PRMT7 promotes tumor cell proliferation and metastasis, making it a potential therapeutic target. In chronic myeloid leukemia (CML), PRMT7 regulates glycine metabolism to maintain the survival of cancer stem cells (CSCs). PRMT7 gene knockout can effectively kill CML cells without affecting normal cells, making it a promising therapeutic target for CML. To date, several PRMT inhibitors have been discovered, primarily type I PRMT inhibitors, including selective and non-selective inhibitors, as well as PRMT5 inhibitors. Szewczyk et al. reported the first selective PRMT7 inhibitor, SGC3027, in 2020, demonstrating significant inhibitory activity against PRMT7 (IC5). 50 While reaching 2.5 nM, it is an adenosine analogue with poor drug-like properties and significant toxic side effects. Recently, Yang et al. reported a novel class of 5-fluoro-4-aminopyrimidine derivatives with activity against PRMT7 only at the micromolar level. The optimal compound, JS1310, selectively inhibits PRMT7 within the PRMT family, with an IC50 concentration of 2.5 nM. 50 The concentration is 5 μM. Therefore, it is of great significance to design and synthesize novel PMRT7 inhibitors with high selectivity, high activity, low toxicity, and good drug-like properties through medicinal chemical methods and techniques. Summary of the Invention

[0006] Purpose of the invention: The technical problem to be solved by the present invention is to provide a PRMT7 target inhibitor with advantages such as high activity and good therapeutic effect and its application.

[0007] Technical solution: To solve the above-mentioned technical problems, the present invention provides a compound or a pharmaceutically acceptable salt thereof that inhibits the activity of PRMT7 protein, wherein the general formula of the compound is shown in any one of formulas (I) to (V):

[0008] ;

[0009] Among them, X1 and X2 are independently selected from sulfur atoms and CH;

[0010] R1 is selected from the following groups: ;

[0011] Among them, R2, R3, R4, R5, and R6 are independently selected from methoxy, ethyl, and hydrogen;

[0012] A and B are independently selected from CH or N.

[0013] Where X1 is a sulfur atom, X2 is CH; when X1 is CH, X2 is a sulfur atom.

[0014] R1 is selected from the following groups: .

[0015] The present invention also provides a compound or a pharmaceutically acceptable salt thereof for inhibiting PRMT7 protein activity, the structural formula of which is shown below:

[0016] .

[0017] Wherein, the pharmaceutically acceptable salt is the acid addition salt of the compound.

[0018] The acid used for salt formation is an inorganic acid or an organic acid. The inorganic acid includes hydrochloric acid, sulfuric acid, or phosphoric acid; the organic acid includes acetic acid, trichloroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid, tartaric acid, or methanesulfonic acid.

[0019] The present invention also provides a pharmaceutical composition comprising the compound or a pharmaceutically acceptable salt thereof, an enantiomer thereof, and a pharmaceutically acceptable carrier.

[0020] The present invention also provides the use of the said compound or a pharmaceutically acceptable salt thereof or the said pharmaceutical composition in the preparation of PRMT7 target inhibitor drugs.

[0021] The present invention also provides the use of the said compound or a pharmaceutically acceptable salt thereof or the said pharmaceutical composition in the preparation of a medicament for the prevention and / or treatment of cancer or tumor-related diseases.

[0022] The cancers or tumor-related diseases mentioned include prostate cancer, colorectal cancer, ovarian cancer, leukemia, lung cancer, liver cancer, breast cancer, esophageal cancer, glioma, or pancreatic cancer.

[0023] This invention designs and synthesizes several novel PRMT7 inhibitors using multiple medicinal chemistry methods and techniques, and tests their inhibitory activity against PRMT7 and their antiproliferative activity against various tumor cell lines.

[0024] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages: The present invention provides a variety of compounds with novel structures that can inhibit PRMT7 and thus fight tumors, and have the advantages of high activity and good therapeutic effect, and can be used to prepare anti-tumor drugs. Detailed Implementation

[0025] The technical solution of the present invention will be further described below.

[0026] Example 1: Preparation of 2-methyl-N 6 -(7-(3,4,5-trimethoxyphenyl)quinoxalin-2-yl)quinoline-4,6-diamine (I-1)

[0027] Synthesis route:

[0028]

[0029] Synthesis method:

[0030] Step 1: Take 6.53 g (40 mmol) of 2-cyano-4-nitroaniline, 30 mL of acetone, and 90 mL of toluene in a 250 mL round-bottom flask, cool to 0 °C in an ice bath, and then slowly add 6.0 mL of SnCl4 dropwise. After the addition is complete, slowly raise the reaction system to room temperature, then heat to 110 °C and reflux for 6 hours. Filter the mixture, wash three times with dichloromethane (20 mL × 3), wash three times with methanol (10 mL × 3), and dry to obtain a yellow solid Al. 1 H NMR (400 MHz, DMSO- d 6) δ 9.46 (d, J = 2.4 Hz, 1H), 9.21 (s, 2H), 8.61 (dd, J = 9.3, 2.4 Hz, 1H), 8.03 (d, J = 9.3 Hz,1H), 6.71 (s, 1H), 2.62 (s, 3H).

[0031] Step 2: Take A1 (6.0 g, 29.53 mmol), 200 mL of water, and 1.2 g of 10% Pd / C in a 500 mL round-bottom flask and react with hydrogen at room temperature overnight. Filter the mixture and concentrate the filtrate under reduced pressure to obtain a yellow solid A2 (4.86 g, 95%). 1 H NMR (400MHz, DMSO- d 6) δ 8.38 (brs, 2H), 7.78 (d, J= 8.9 Hz, 1H), 7.29 (dd, J = 9.0, 2.3Hz, 1H), 7.21 (d, J = 2.3 Hz, 1H), 6.51 (s, 1H), 5.76 (s, 2H), 2.54 (s, 3H).

[0032] Step 3: Take A2 (173 mg, 1.0 mmol), 7-bromo-2-chloroquinoxaline (292 mg, 1.2 mmol), tris(dibenzylideneacetone)palladium (Pd2dba3, 46 mg, 0.05 mmol), 4,5-bis(diphenylphosphine-9,9-dimethyloxanthracene) (Xantphos, 58 mg, 0.1 mmol), sodium tert-butoxide (290 mg, 3 mmol), and 5 mL of 1,4-dioxane into a 25 mL round-bottom flask, purge with argon three times, and react overnight at 50 °C under argon protection. Filter the reaction solution through diatomaceous earth, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain yellow solid A3 (61 mg, 16%). 1 H NMR (400 MHz, DMSO- d 6) δ 10.71 (s, 1H), 9.03 (d, J = 2.2 Hz, 1H), 8.73 (s, 1H), 8.46 – 8.22 (m, 3H), 8.08 (dd, J = 9.1, 2.2 Hz, 1H), 7.91 (d, J = 9.0 Hz, 1H), 7.83 (d, J = 8.6 Hz, 1H), 7.64 (dd, J = 8.7, 2.2 Hz, 1H), 6.64 (s, 1H), 2.58 (s, 3H).

[0033] Step 4: Take A3 (61 mg, 0.16 mmol), 3,4,5-trimethoxyphenylboronic acid (41 mg, 0.19 mmol), bis(triphenylphosphine) palladium dichloride (5.6 mg, 0.008 mmol, Titan reagent, catalog number 1020950), sodium carbonate (51 mg, 0.48 mmol), 3 mL of 1,4-dioxane and 0.2 mL of water into a 25 mL round-bottom flask, purge with argon three times, and react overnight at 80 °C under argon protection. Filter the reaction solution through diatomaceous earth, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain yellow solid I-1 (53 mg, 71%). 1H NMR (400 MHz, DMSO- d 6) δ 10.49 (s, 1H), 9.05 (d, J = 2.2 Hz, 1H),8.69 (s, 1H), 8.57 (brs, 2H), 8.27 (d, J = 2.0 Hz, 1H), 8.20 (dd, J = 9.1, 2.2Hz, 1H), 7.96 (d, J = 8.5 Hz, 1H), 7.93 – 7.87 (m, 2H), 7.12 (s, 2H), 6.64 (s,1H), 3.93 (s, 6H), 3.74 (s, 3H), 2.61 (s, 3H).

[0034] Example 2: Preparation of N-(1-methyl-1H-pyrazol-4-yl)-7-(3,4,5-trimethoxyphenyl)quinoxalin-2-amine (I-2)

[0035] Synthesis route:

[0036]

[0037] Synthesis method:

[0038] Step 1: 7-bromo-2-chloroquinoxaline (438 mg, 1.8 mmol), 1-methyl-1H-pyrazole-4-amine (146 mg, 1.5 mmol), tris(dibenzylacetone)palladium (69 mg, 0.075 mmol), 1,1'-binaphthyl-2,2'-bis(diphenylphosphine) (93 mg, 0.15 mmol), sodium carbonate (477 mg, 4.5 mmol), and 4 mL of 1,4-dioxane were placed in a 25 mL round-bottom flask. Argon gas was purged three times, and the reaction was carried out at 100 °C for 22 h under argon protection. The reaction solution was filtered through diatomaceous earth, concentrated under reduced pressure, and purified by rapid silica gel column chromatography to obtain a pale yellow solid A4 (51 mg, 11%). 1 H NMR (400 MHz, DMSO- d 6) δ 10.15 (s, 1H), 8.44 (s, 1H), 8.42 (s, 1H), 7.98 (d, J = 2.3 Hz, 1H), 7.74 (d, J = 8.7 Hz, 1H),7.62 (s, 1H), 7.51 (dd, J= 8.6, 2.2 Hz, 1H), 3.87 (s, 3H).

[0039] Step 2: Take A4 (51 mg, 0.17 mmol), 3,4,5-trimethoxyphenylboronic acid (54 mg, 0.26 mmol), bis(triphenylphosphine)palladium dichloride (5.6 mg, 0.008 mmol), sodium carbonate (51 mg, 0.48 mmol), 3 mL of 1,4-dioxane and 0.2 mL of water into a 25 mL round-bottom flask, purge with argon three times, and react at 100 °C for 15 hours under argon protection. Filter the reaction solution through diatomaceous earth, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain yellow solid I-2 (50 mg, 76%). 1 H NMR (400 MHz, DMSO-) d 6) δ 9.98 (s, 1H), 8.42 (s, 1H), 8.38 (s, 1H), 8.02 (s, 1H), 7.86 (d, J = 8.4 Hz, 1H), 7.75 (d, J = 8.5 Hz, 1H), 7.69 (s, 1H), 7.08 (s, 2H), 3.92 (s, 6H), 3.89 (s, 3H), 3.73 (s, 3H).

[0040] Example 3: Preparation of N 1 N 1 -dimethyl-N 4 -(7-(3,4,5-trimethoxyphenyl)quinoxalin-2-yl)phenyl-1,4-diamine (I-3)

[0041] Structural formula:

[0042] Following the synthesis method of Example 2, the starting material 1-methyl-1H-pyrazole-4-amine was replaced with an equimolar mass of 4-amino-N,N-dimethylaniline, with a yield of 83%. Intermediate 1 H NMR (400 MHz, DMSO- d 6) δ 9.81 (s, 1H), 8.47 (s, 1H), 7.83 – 7.67 (m, 4H), 7.50 (dd, J = 8.7, 2.2 Hz, 1H), 6.82 – 6.74 (m, 2H), 2.89 (s, 6H).

[0043] 1 H NMR (400 MHz, DMSO-d 6) δ 9.67 (s, 1H), 8.45 (s, 1H), 7.90 – 7.77(m, 4H), 7.74 (dd, J = 8.5, 2.0 Hz, 1H), 7.06 (s, 2H), 6.82 – 6.77 (m, 2H), 3.91 (s, 6H), 3.72 (s, 3H), 2.88 (s, 6H).

[0044] Example 4: Preparation of N-(4-amino-2-methylquinoline-6-yl)-2-(4-(5-fluoro-2-(3,4,5-trimethyloxyphenyl)pyrimidin-4-yl)piperazin-1-yl)acetamide (I-4)

[0045] Synthesis route:

[0046]

[0047] Synthesis method:

[0048] Step 1: Take the raw materials 2-(piperazin-1-yl)acetic acid (B1, 0.72 g, 5 mmol), 2,4-dichloro-5-fluoropyrimidine (B2, 1.67 g, 10 mmol), DIPEA (1.73 mL, 10 mmol) and 9 mL of ethanol into a 25 mL round-bottom flask, stir the mixture at room temperature for 12 hours, filter, and dry to obtain white solid B3 (0.71 g, 52%). 1 H NMR (400 MHz, Chloroform- d ) δ 7.97 (d, J = 6.0 Hz, 1H), 3.91 (t, J = 5.1 Hz, 4H), 3.28 (s, 2H), 2.82 –2.76 (m, 4H).

[0049] Step 2: Dissolve B3 (165 mg, 0.6 mmol) in 4 ml of DMF, add DIPEA (0.31 ml, 1.8 mmol), slowly add 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU, 239 mg, 0.63 mmol), stir at room temperature for half an hour, then slowly add A2 (104 mg, 0.6 mmol), stir at room temperature for 6 hours, concentrate the reaction solution under reduced pressure, and purify by rapid silica gel column chromatography to obtain white solid B4 (167 mg, 65%). 1 H NMR (400 MHz, DMSO- d6) δ 10.07 (s, 1H), 8.41 (d, J = 2.4 Hz, 1H), 8.22 (d, J = 6.5 Hz, 1H), 7.91 (dd, J = 9.1, 2.2 Hz, 1H), 7.84 – 7.57 (m, 3H), 3.83 (t, J = 4.9 Hz, 4H), 3.27 (s, 2H), 2.70 (t, J = 5.0 Hz, 4H), 2.50 (s, 3H).

[0050] Step 3: Take B4 (167 mg, 0.39 mmol), 3,4,5-trimethoxyphenylboronic acid (124 mg, 0.58 mmol), bis(triphenylphosphine)palladium dichloride (14 mg, 0.02 mmol), potassium carbonate (166 mg, 1.2 mmol), 4 mL of 1,4-dioxane and 0.2 mL of water and place them in a 25 mL round-bottom flask. Replace the flask with argon gas three times and react at 100 °C for 15 hours under argon protection. Filter the reaction solution through diatomaceous earth, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain yellow solid I-4 (162 mg, 74%). 1 H NMR (400 MHz, Chloroform- d ) δ 9.33 (s, 1H), 8.52 (d, J = 2.3 Hz, 1H), 8.18 (d, J =6.3 Hz, 1H), 7.93 (d, J = 9.0 Hz, 1H), 7.59 (s, 2H), 7.49 (dd, J = 9.0, 2.2 Hz,1H), 6.50 (s, 1H), 5.12 (s, 2H), 3.99 (t, J = 5.0 Hz, 4H), 3.95 (s, 6H), 3.90 (s, 3H), 3.28 (s, 2H), 2.83 (t, J = 4.9 Hz (4H), 2.58 (s, 3H).

[0051] Example 5: Preparation of 2-methyl-N 6 -(3-(3,4,5-trimethoxyphenyl)quinoxalin-6-yl)quinoline-4,6-diamine (I-5)

[0052] Synthesis route:

[0053]

[0054] Synthesis method:

[0055] Step 1: Take 7-bromo-2-chloroquinoxaline (243 mg, 1 mmol), 3,4,5-trimethoxyphenylboronic acid (212 mg, 1 mmol), bis(triphenylphosphine)palladium dichloride (35 mg, 0.05 mmol), sodium carbonate (318 mg, 3 mmol), 5 mL of 1,4-dioxane and 0.5 mL of water into a 25 mL round-bottom flask, replace with argon gas three times, and react at 100 °C for 15 hours under argon protection. Filter to obtain yellow solid C1 (330 mg, 88%). 1 H NMR (400 MHz, DMSO- d 6) δ 9.69 (s, 1H), 8.37 (d, J = 2.2 Hz, 1H), 8.06 (d, J = 8.8 Hz, 1H), 7.96 (dd, J = 8.8, 2.2 Hz, 1H), 7.65(s, 2H), 3.94 (s, 6H), 3.77 (s, 3H).

[0056] Step 2: C1 (250 mg, 0.67 mmol), A2 (100 mg, 0.58 mmol), Pd2dba3 (23 mg, 0.025 mmol), 2-dicyclohexylphosphine-2',4',6'-triisopropylbiphenyl (Xphos, 24 mg, 0.05 mmol), cesium carbonate (490 mg, 1.5 mmol), and 4 mL of 1,4-dioxane were placed in a 25 mL round-bottom flask. Argon gas was purged three times, and the reaction was carried out at 100 °C for 20 h under argon protection. The reaction solution was filtered through diatomaceous earth, concentrated under reduced pressure, and purified by rapid silica gel column chromatography to obtain yellow solid I-5 (27 mg, 10%). 1 H NMR (400 MHz, DMSO- d 6) δ 9.27 (s, 1H), 8.96 (s, 1H), 7.99 (d, J =2.3 Hz, 1H), 7.94 (d, J = 9.1 Hz, 1H), 7.73 (d, J = 8.9 Hz, 1H), 7.60 – 7.51 (m,4H), 7.41 (d, J= 2.5 Hz, 1H), 6.67 (brs, 2H), 6.44 (s, 1H), 3.90 (s, 6H), 3.74(s, 3H), 2.42 (s, 3H).

[0057] Example 6: Preparation of 1-(4-amino-2-methylquinoline-6-yl)-3-(3',4',5'-trimethoxy-[1,1'-diphenyl]-3-yl)urea (I-6)

[0058] Synthesis route:

[0059]

[0060] Synthesis method:

[0061] Step 1: Take A2 (86 mg, 0.5 mmol), 3-bromophenyl isocyanate (D1, 0.18 ml, 1.5 mmol), 2.5 ml acetonitrile, and 1.5 ml DMF and place them in a 25 ml round-bottom flask. Stir at room temperature for 12 hours, filter, and wash the filter cake several times with ethyl acetate and methanol to obtain white solid D2 (176 mg, 95%). 1 H NMR (400 MHz, DMSO- d 6) δ 9.61 (s, 1H), 9.54 (s, 1H), 8.63 (brs, 2H), 8.38 (d, J = 2.1 Hz, 1H), 7.95 (t, J = 2.0 Hz, 1H), 7.89 (dd, J = 9.1, 2.2 Hz, 1H), 7.85 – 7.79 (m, 2H), 7.36 – 7.29 (m, 2H), 6.56 (s, 1H), 2.57 (s, 3H).

[0062] Step 2: Take D2 (100 mg, 0.27 mmol), 3,4,5-trimethoxyphenylboronic acid (92 mg, 0.43 mmol), tetrakis(triphenylphosphine)palladium (16 mg, 0.014 mmol), potassium carbonate (112 mg, 0.81 mmol), 4 mL of 1,4-dioxane and 0.2 mL of water into a 25 mL round-bottom flask, purge with argon three times, and react at 100 °C for 15 hours under argon protection. Filter the reaction solution through diatomaceous earth, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain white solid I-6 (69 mg, 56%). 1 H NMR (400MHz, DMSO- d6) δ 9.25 (s, 1H), 9.18 (s, 1H), 8.62 (s, 2H), 8.36 (d, J = 2.2 Hz, 1H), 7.95 (dd, J = 9.1, 2.2 Hz, 1H), 7.81 (d, J = 9.1 Hz, 1H), 7.71 (t, J = 2.0 Hz, 1H), 7.54 (dt, J = 8.3, 1.3 Hz, 1H), 7.39 (t, J = 7.9 Hz, 1H), 7.31 (dt, J = 7.7,1.4 Hz, 1H), 6.87 (s, 2H), 6.56 (s, 1H), 3.87 (s, 6H), 3.70 (s, 3H), 2.57 (s, 3H).

[0063] Example 7: Preparation of N-(4-amino-2-methylquinoline-6-yl)-2-(3,4,5-trimethoxyphenyl)benzo[d]thiazol-6-carboxamide (I-7)

[0064] Synthesis route:

[0065]

[0066] Synthesis method:

[0067] Step 1: 2-chloro-6-benzothiazolium carboxylic acid (214 mg, 1 mmol), 3,4,5-trimethoxyphenylboronic acid (318 mg, 1.5 mmol), tetrakis(triphenylphosphine)palladium (35 mg, 0.05 mmol), sodium carbonate (424 mg, 4 mmol), 5 mL of 1,4-dioxane and 0.3 mL of water were placed in a 25 mL round-bottom flask. Argon gas was purged three times, and the reaction was carried out at 100 °C for 8 hours under argon protection. The reaction solution was diluted with water (20 mL), and the pH was adjusted to neutral with 4 N HCl. The mixture was filtered, and the filter cake was washed with water, followed by washing with ethyl acetate and methanol to obtain a white solid product E1 (62 mg, 18%). 1 H NMR (400 MHz, DMSO- d 6) δ 13.20 (s, 1H), 8.77 (d, J = 1.7 Hz, 1H), 8.13 (d, J = 8.6 Hz, 1H), 8.07 (dd, J= 8.6, 1.7Hz, 1H), 7.38 (s, 2H), 3.93 (s, 6H), 3.77 (s, 3H).

[0068] Step 2: Dissolve E1 (165 mg, 0.6 mmol) in 4 ml DMF, add DIPEA (0.1 ml, 0.54 mmol), slowly add HATU (72 mg, 0.19 mmol), stir at room temperature for half an hour, then slowly add A2 (31 mg, 0.18 mmol), stir at room temperature overnight, concentrate the reaction solution under reduced pressure, and purify by rapid silica gel column chromatography to obtain yellow solid I-7 (31 mg, 34%). 1 H NMR (400 MHz, DMSO- d 6) δ 10.90 (s, 1H), 8.86 (d, J = 1.7 Hz, 1H), 8.80(d, J = 2.1 Hz, 1H), 8.56 (brs, 2H), 8.23 ​​(d, J = 8.6 Hz, 1H), 8.19 (dd, J = 8.5, 1.7 Hz, 1H), 8.07 (dd, J = 9.0, 2.1 Hz, 1H), 7.93 (d, J = 9.0 Hz, 1H), 7.41 (s,2H), 6.60 (s, 1H), 3.94 (s, 6H), 3.78 (s, 3H), 2.59 (s, 3H).

[0069] Example 8: Preparation of N-(4-amino-2-methylquinoline-6-yl)-2-(3',4',5'-trimethoxy-[1,1'-diphenyl]-3-yl)acetamide (I-8)

[0070] Synthesis route:

[0071]

[0072] Synthesis method:

[0073] Step 1: Take m-bromophenylacetic acid (F1, 215 mg, 1 mmol), 3,4,5-trimethoxyphenylboronic acid (339 mg, 1.6 mmol), tetrakis(triphenylphosphine)palladium (35 mg, 0.05 mmol), cesium carbonate (1300 mg, 4 mmol), 5 mL of 1,4-dioxane and 0.5 mL of water into a 25 mL round-bottom flask, purge with argon three times, and react at 100 °C for 17 hours under argon protection. Dilute the reaction solution with water, adjust the pH to neutral with 4 N HCl, extract with dichloromethane, collect the organic phase, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain white solid F2 (120 mg, 40%). 1 H NMR (400 MHz, DMSO- d 6) δ 12.37(s, 1H), 7.59 – 7.52 (m, 1H), 7.42 – 7.34 (m, 1H), 7.24 (dt, J = 7.5, 1.4 Hz,1H), 6.89 (s, 2H), 3.86 (s, 6H), 3.69 (s, 3H), 3.65 (s, 2H).

[0074] Step 2: Dissolve F2 (120 mg, 0.4 mmol) in 4 ml DMF, add DIPEA (0.26 ml, 1.5 mmol), slowly add HATU (167 mg, 0.44 mmol), stir at room temperature for half an hour, then slowly add A2 (69 mg, 0.4 mmol), stir at room temperature for 4 hours, concentrate the reaction solution under reduced pressure, and purify by rapid silica gel column chromatography to obtain yellow solid I-8 (71 mg, 39%). 1 H NMR (400 MHz, DMSO- d 6) δ 10.33 (s, 1H), 8.24 (s, 1H), 7.66 (t, J =1.8 Hz, 1H), 7.60 (s, 2H), 7.59 – 7.55 (m, 1H), 7.42 (t, J = 7.6 Hz, 1H), 7.36 (d, J = 7.6 Hz, 1H), 6.90 (s, 2H), 6.40 (s, 1H), 6.37 (s, 2H), 3.86 (s, 6H), 3.77 (s, 2H), 3.69 (s, 3H), 2.37 (s, 3H).

[0075] Example 9: Preparation of N-(4-amino-2-methylquinoline-6-yl)-2-((3',4',5'-trimethoxy-[1,1'-diphenyl]-3-yl)oxy)acetamide (I-9)

[0076] Structural formula:

[0077] Following the synthesis method of Example 8, the starting material m-bromophenylacetic acid was replaced with equimolar amounts of 2-(3-bromophenoxy)acetic acid, with a yield of 43%. 1 H NMR (400 MHz, DMSO- d 6) δ 10.67 (s, 1H), 8.64 (d, J = 2.1 Hz,1H), 8.55 (brs, 2H), 7.96 (dd, J = 9.1, 2.1 Hz, 1H), 7.88 (d, J = 9.1 Hz, 1H), 7.40 (t, J = 7.9 Hz, 1H), 7.34 – 7.27 (m, 2H), 7.06 – 7.00 (m, 1H), 6.89 (s,2H), 6.57 (s, 1H), 4.90 (s, 2H), 3.83 (s, 6H), 3.69 (s, 3H), 2.57 (s, 3H).

[0078] Example 10: Preparation of N-(4-amino-2-methylquinoline-6-yl)-2-(2-(4-(5-fluoro-2-(3,4,5-trimethoxyphenyl)pyrimidin-4-yl)piperazin-1-yl)acetamyl)acetamide (I-10)

[0079] Synthesis route:

[0080]

[0081] Synthesis method:

[0082] Step 1: Dissolve B1 (275 mg, 1 mmol) in 5 ml of dichloromethane, add DIPEA (0.42 ml, 3 mmol), slowly add HATU (399 mg, 1.05 mmol), stir at room temperature for half an hour, then slowly add glycine tert-butyl ester (0.15 ml, 1.1 mmol), stir at room temperature overnight, concentrate the reaction solution under reduced pressure, and purify by rapid silica gel column chromatography to obtain white oily solid G1 (320 mg, 82%). 1 H NMR (400 MHz, Chloroform- d) δ 7.94 (d, J = 6.1 Hz,1H), 7.51 (s, 1H), 3.99 (d, J = 5.5 Hz, 2H), 3.93 – 3.84 (m, 4H), 3.11 (s, 2H), 2.73 – 2.64 (m, 4H), 1.48 (s, 9H).

[0083] Step 2: Take G1 (252 mg, 0.65 mmol), 3,4,5-trimethoxyphenylboronic acid (220 mg, 1.04 mmol), bis(triphenylphosphine)palladium dichloride (23 mg, 0.03 mmol), potassium carbonate (269 mg, 1.95 mmol), 4 mL of 1,4-dioxane and 0.2 mL of water into a 25 mL round-bottom flask, replace with argon gas three times, and react at 100 °C for 12 hours under argon protection. Filter the reaction solution with diatomaceous earth, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain oily solid G2 (300 mg, 89%). 1 H NMR (400 MHz, DMSO-) d 6) δ 8.35 (d, J = 6.6 Hz, 1H), 8.16 (t, J = 6.1 Hz, 1H), 7.58 (s,2H), 3.89 – 3.82 (m, 10H), 3.77 (d, J = 6.1 Hz, 2H), 3.72 (s, 3H), 3.04 (s,2H), 2.64 (t, J = 4.9 Hz, 4H), 1.41 (s, 9H).

[0084] Step 3: Dissolve G2 (300 mg, 0.58 mmol) in 1 ml of dichloromethane, add 1 ml of trifluoroacetic acid, stir at room temperature for 2 hours, and then concentrate under reduced pressure to obtain yellow oily solid G3 (232 mg, 95%). 1 H NMR (400 MHz, DMSO- d 6) δ 8.97 (t, J = 5.8 Hz, 1H), 8.47 (d, J = 6.3 Hz, 1H), 7.59 (s, 2H), 4.89– 4.35 (m, 4H), 3.90 (d, J= 5.8 Hz, 2H), 3.86 (s, 6H), 3.73 (s, 3H), 3.67 –3.22 (m, 6H).

[0085] Step 4: Dissolve G3 (275 mg, 1 mmol) in 2 ml DMF, add DIPEA (0.35 ml, 2 mmol), slowly add HATU (200 mg, 0.53 mmol), stir at room temperature for half an hour, then slowly add A2 (86 mg, 0.5 mmol), stir at room temperature overnight, concentrate the reaction solution under reduced pressure, and purify by rapid silica gel column chromatography to obtain white solid I-10 (110 mg, 36%). 1 HNMR (400 MHz, DMSO- d 6) δ 10.15 (s, 1H), 8.35 (d, J = 6.7 Hz, 1H), 8.25 (s, 1H), 8.16 (t, J = 6.0 Hz, 1H), 7.68 – 7.60 (m, 2H), 7.58 (s, 2H), 6.73 (brs, 2H), 6.45 (s, 1H), 4.01 (d, J = 5.9 Hz, 2H), 3.92 – 3.79 (m, 10H), 3.72 (s, 3H), 3.09 (s, 2H), 2.68 (t, J = 4.9 Hz, 4H), 2.41 (s, 3H).

[0086] Example 11: Preparation of N-(4-amino-2-methylquinoline-6-yl)-7-(3,4,5-trimethoxyphenyl)quinoline-2-carboxamide (I-11)

[0087] Structural formula:

[0088] Following the synthesis method of Example 7, the starting material 2-chloro-6-benzothiazole carboxylic acid was replaced with an equimolar mass of 7-bromoquinoline-2-carboxylic acid, with a yield of 61%. 1 H NMR (400 MHz, DMSO- d 6) δ 11.13 (s, 1H), 8.92 – 8.61 (m,4H), 8.55 – 8.51 (m, 1H), 8.38 (dd, J = 9.1, 2.2 Hz, 1H), 8.28 (d, J = 8.4 Hz, 1H), 8.24 (d,J = 8.6 Hz, 1H), 8.20 (dd, J = 8.6, 1.8 Hz, 1H), 7.97 (d, J = 9.1 Hz,1H), 7.18 (s, 2H), 6.63 (s, 1H), 3.94 (s, 6H), 3.75 (s, 3H), 2.62 (s, 3H).

[0089] Example 12: Preparation of N-(4-amino-2-methylquinoline-6-yl)-3',4',5'-trimethoxy-[1,1'-diphenyl]-3-carboxamide (I-12)

[0090] Structural formula:

[0091] Referring to the synthesis method of Example 8, the raw material m-bromophenylacetic acid was replaced with an equimolar mass of m-bromobenzoic acid, with a yield of 47%. 1 H NMR (400 MHz, DMSO- d 6) δ 10.92 (s, 1H), 8.83 (d, J = 2.1 Hz, 1H), 8.70(brs, 2H), 8.33 (t, J = 1.8 Hz, 1H), 8.12 (dd, J = 9.1, 2.1 Hz, 1H), 8.03 – 7.98(m, 1H), 7.98 – 7.91 (m, 2H), 7.65 (t, J = 7.7 Hz, 1H), 7.06 (s, 2H), 6.61 (s,1H), 3.90 (s, 6H), 3.72 (s, 3H), 2.60 (s, 3H).

[0092] Example 13: 2-Methyl-N 6 -(2-(3,4,5-trimethoxyphenyl)thieno[3,2-d]pyrimidin-4-yl)quinoline-4,6-diamine (I-13)

[0093] Synthesis route:

[0094]

[0095] Synthesis method:

[0096] Step 1: Take raw material A2 (86 mg, 0.5 mmol), 2,4-dichlorothiopheno[3,2-d]pyrimidine (H1, 205 mg, 1 mmol), DIPEA (0.17 mL, 1 mmol) and 3 mL of ethanol and place them in a 25 mL round-bottom flask. Stir the mixture at 85 °C for 6 hours, filter, wash the filter cake with ethyl acetate and methanol, and dry to obtain white solid H2 (150 mg, 88%). 1 H NMR (400 MHz, DMSO- d 6) δ 10.59 (s, 1H), 8.40 (d, J = 2.2 Hz, 1H), 8.29 (d, J = 5.4 Hz, 1H), 8.23 ​​(brs, 2H), 7.98 (dd, J = 9.0, 2.1 Hz, 1H), 7.90 (d, J = 9.0 Hz, 1H), 7.44(d, J = 5.4 Hz, 1H), 6.59 (s, 1H), 2.58 (s, 3H).

[0097] Step 2: Take H2 (147 mg, 0.43 mmol), 3,4,5-trimethoxyphenylboronic acid (137 mg, 0.65 mmol), bis(triphenylphosphine)palladium dichloride (18 mg, 0.03 mmol), potassium carbonate (207 mg, 1.5 mmol), 4 mL of 1,4-dioxane and 0.3 mL of water and place them in a 25 mL round-bottom flask. Replace the flask with argon gas three times and react at 100 °C for 14 hours under argon protection. Filter the reaction solution with diatomaceous earth, concentrate under reduced pressure, and purify by rapid silica gel column chromatography to obtain a white solid I-13 (124 mg, 61%). 1 H NMR (400 MHz, DMSO-) d 6) δ 9.99 (s, 1H), 8.35 (d, J = 2.2 Hz, 1H), 8.22 (d, J = 5.4 Hz, 1H), 7.94 (dd, J = 8.9, 2.2 Hz, 1H), 7.78 – 7.70 (m, 3H), 7.55 (d, J = 5.4 Hz,1H), 6.77 (brs, 2H), 6.47 (s, 1H), 3.81 (s, 6H), 3.71 (s, 3H), 2.44 (s, 3H).

[0098] Example 14: Preparation of N 6 -(2-(benzo[d][1,3]dioxacyclopenten-5-yl)thieno[3,2-d]pyrimidin-4-yl)-2-methylquinoline-4,6-diamine (I-14)

[0099] Structural formula:

[0100] Referring to the synthesis method of Example 13, the starting material 3,4,5-trimethoxyphenylboronic acid was replaced with equimolar amounts of 3,4-methyleneoxyphenylboronic acid, with a yield of 70%. 1 H NMR (400 MHz, DMSO- d 6) δ 10.28 (s, 1H), 8.88 –8.47 (m, 3H), 8.28 (d, J = 5.4 Hz, 1H), 8.16 (dd, J = 9.0, 2.1 Hz, 1H), 8.06 –7.89 (m, 2H), 7.81 (d, J = 1.7 Hz, 1H), 7.54 (d, J = 5.4 Hz, 1H), 6.98 (d, J = 8.2Hz, 1H), 6.64 (s, 1H), 6.09 (s, 2H), 2.63 (s, 3H).

[0101] Example 15: Preparation of N 6 -(2-(1H-indol-5-yl)thieno[3,2-d]pyrimidin-4-yl)-2-methylquinoline-4,6-diamine (I-15)

[0102] Structural formula:

[0103] Referring to the synthesis method of Example 13, the starting material 3,4,5-trimethoxyphenylboronic acid was replaced with equimolar mass of 5-indoleboronic acid, with a yield of 23%. 1 H NMR (400 MHz, DMSO- d 6) δ 11.51 (s, 1H), 9.01 – 8.61 (m,4H), 8.43 (s, 1H), 8.21 (dd, J = 9.0, 2.1 Hz, 1H), 8.14 (d, J = 8.6 Hz, 1H), 8.04(d, J = 9.0 Hz, 1H), 7.70 (d,J = 5.3 Hz, 1H), 7.56 – 7.43 (m, 2H), 6.69 (s, 1H), 6.56 (s, 1H), 2.65 (s, 3H).

[0104] Example 16: Preparation of N 6 -(2-(3,4-dihydroisoquinoline-2(1H)-yl)thieno[3,2-d]pyrimidin-4-yl)-2-methylquinoline-4,6-diamine (I-16)

[0105] Structural formula:

[0106] Referring to the synthesis method of Example 13, the starting material 3,4,5-trimethoxyphenylboronic acid was replaced with equimolar mass of 1,2,3,4-tetrahydroisoquinoline, with a yield of 90%. 1 H NMR (400 MHz, DMSO- d 6) δ 9.94 (s, 1H), 8.90 – 8.70 (m, 2H), 8.52 (s, 1H), 8.13 – 8.02 (m, 2H), 7.94 (d, J = 9.1 Hz, 1H), 7.23 (d, J = 5.3 Hz, 1H), 7.21 – 7.11 (m, 4H), 6.64 (s, 1H), 4.85 (s, 2H), 3.96 (t, J =5.8 Hz, 2H), 2.86 (t, J = 5.9 Hz, 2H), 2.63 (s, 3H).

[0107] Example 17: Preparation of N 6 -(2-(4-methoxypiperidin-1-yl)thieno[3,2-d]pyrimidin-4-yl)-2-methylquinoline-4,6-diamine (I-17)

[0108] Structural formula:

[0109] Following the synthesis method of Example 13, the starting material 3,4,5-trimethoxyphenylboronic acid was replaced with an equimolar mass of 4-methoxypiperidine, with a yield of 54%. 1 H NMR (400 MHz, DMSO- d 6) δ 9.51 (s, 1H), 8.28 (d, J = 2.2Hz, 1H), 7.96 (d, J= 5.4 Hz, 1H), 7.70 (dd, J = 9.0, 2.2 Hz, 1H), 7.63 (d, J = 8.9Hz, 1H), 7.12 (d, J = 5.4 Hz, 1H), 6.47 – 6.36 (m, 3H), 4.22 (dt, J = 13.1, 4.7Hz, 2H), 3.43 – 3.37 (m, 1H), 3.29 – 3.19 (m, 5H), 2.40 (s, 3H), 1.90 – 1.80(m, 2H), 1.44 – 1.32 (m, 2H).

[0110] Example 18: Preparation of 2-methyl-N 6 -(2-(4-morpholinylphenyl)thieno[3,2-d]pyrimidin-4-yl)quinoline-4,6-diamine (I-18)

[0111] Structural formula:

[0112] Following the synthesis method of Example 13, the starting material 3,4,5-trimethoxyphenylboronic acid was replaced with equimolar amounts of 4-(4-morpholino)phenylboronic acid, with a yield of 57%. 1 H NMR (400 MHz, DMSO- d 6) δ 9.89 (brs, 1H), 8.35 (d, J = 2.3 Hz, 1H), 8.27 (d, J = 8.9 Hz, 2H), 8.13 (d, J = 5.4 Hz, 1H), 7.85 (dd, J =9.0, 2.3 Hz, 1H), 7.70 (d, J = 9.0 Hz, 1H), 7.45 (d, J = 5.4 Hz, 1H), 6.99 (d, J =9.1 Hz, 2H), 6.50 (s, 2H), 6.46 (s, 1H), 3.79 – 3.69 (m, 4H), 3.23 – 3.17 (m,4H), 2.43 (s, 3H).

[0113] Example 19: Preparation of N 6-(2-(6-methoxypyridin-3-yl)thieno[3,2-d]pyrimidin-4-yl)-2-methylquinoline-4,6-diamine (I-19)

[0114] Structural formula:

[0115] Referring to the synthesis method of Example 13, the starting material 3,4,5-trimethoxyphenylboronic acid was replaced with equimolar mass of 6-methoxypyridine-3-boronic acid, with a yield of 68%. 1 H NMR (400 MHz, DMSO- d 6) δ 10.35 (s, 1H), 9.07 (d, J = 2.3 Hz, 1H), 8.74 – 8.61 (m, 3H), 8.55 (dd, J = 8.8, 2.4 Hz, 1H), 8.31 (d, J =5.5 Hz, 1H), 8.18 (dd, J = 9.1, 2.1 Hz, 1H), 8.01 (d, J = 8.8 Hz, 1H), 7.57 (d, J =5.4 Hz, 1H), 6.90 (d, J = 8.7 Hz, 1H), 6.65 (s, 1H), 3.91 (s, 3H), 2.63 (s, 3H).

[0116] Example 20: Preparation of 2-methyl-N 6 -(2-(3,4,5-trimethoxyphenyl)benzo[d]thiazolyl)quinoline-4,6-diamine (I-20)

[0117] Structural formula:

[0118] Referring to the synthesis method of Example 5, the starting material 7-bromo-2-chloroquinoxaline was replaced with an equimolar mass of 6-bromo-2-chlorobenzothiazole, with a yield of 28%. 1 H NMR (400 MHz, DMSO- d 6) δ 8.81 (s, 1H), 7.94 (d, J = 8.8Hz, 1H), 7.90 – 7.83 (m, 2H), 7.71 (d, J= 9.0 Hz, 1H), 7.59 – 7.38 (m, 3H), 7.34 – 7.25 (m, 3H), 6.47 (s, 1H), 3.91 (s, 6H), 3.75 (s, 3H), 2.47 (s, 3H).

[0119] Example 21: Preparation of N 6 -(2-(benzo[d][1,3]dioxacyclopenten-5-yl)benzo[d]thiazo-6-yl)-2-methylquinoline-4,6-diamine (I-21)

[0120] Structural formula:

[0121] Referring to the synthesis method of Example 5, the raw material 3,4,5-trimethoxyphenylboronic acid was replaced with equimolar amounts of 3,4-methyleneoxyphenylboronic acid, with a yield of 32%. 1 H NMR (400 MHz, DMSO- d 6) δ 9.08 (s, 1H), 8.57 (brs,2H), 7.98 – 7.89 (m, 3H), 7.87 (d, J = 8.9 Hz, 1H), 7.68 (dd, J = 9.1, 2.3 Hz,1H), 7.61 – 7.51 (m, 2H), 7.33 (dd, J = 8.8, 2.3 Hz, 1H), 7.09 (d, J = 8.0 Hz,1H), 6.55 (s, 1H), 6.16 (s, 2H), 2.57 (s, 3H).

[0122] Example 22: Preparation of N 6 -(2-(3,4-dihydroisoquinoline-2(1H)-yl)benzo[d]thiazoline-6-yl)-2-methylquinoline-4,6-diamine (I-22)

[0123] Synthesis route:

[0124]

[0125] Synthesis method:

[0126] Step 1: Dissolve 6-bromo-2-chlorobenzothiazole (J1, 497 mg, 2 mmol) and 1,2,3,4-tetrahydroisoquinoline (J2, 0.5 ml, 4 mmol) in 6 ml of ethanol, add DIPEA (0.69 ml, 4 mmol), stir at 85 °C overnight, cool to room temperature, a large amount of solid precipitates, filter, wash with a small amount of ethyl acetate, to obtain white solid J3 (560 mg, 81%). 1 H NMR (400MHz, Chloroform- d ) δ 7.73 (d, J = 1.9 Hz, 1H), 7.44 (d, J = 8.5 Hz, 1H), 7.39(dd, J = 8.6, 2.0 Hz, 1H), 7.25 – 7.18 (m, 4H), 4.81 (s, 2H), 3.86 (t, J = 5.9Hz, 2H), 3.03 (t, J = 5.9 Hz, 2H).

[0127] Step 2: J3 (224 mg, 0.65 mmol), A2 (86 mg, 0.5 mmol), Pd2dba3 (23 mg, 0.025 mmol), Xphos (24 mg, 0.05 mmol), cesium carbonate (490 mg, 1.5 mmol), and 4 mL of 1,4-dioxane were placed in a 25 mL round-bottom flask, purged with argon three times, and reacted at 100 °C for 15 h under argon protection. The reaction solution was filtered through diatomaceous earth, concentrated under reduced pressure, and purified by rapid silica gel column chromatography to obtain yellow solid I-5 (92 mg, 42%). 1 H NMR (400 MHz, DMSO- d 6) δ8.62 (s, 1H), 8.40 (brs, 2H), 7.74 – 7.68 (m, 2H), 7.66 (d, J = 2.4 Hz, 1H), 7.56 (dd, J = 9.1, 2.3 Hz, 1H), 7.47 (d, J = 8.5 Hz, 1H), 7.33 – 7.29 (m, 1H), 7.27 – 7.20 (m, 3H), 7.15 (dd, J = 8.6, 2.3 Hz, 1H), 6.49 (s, 1H), 4.77 (s,2H), 3.82 (t,J = 6.0 Hz, 2H), 2.99 (t, J = 5.9 Hz, 2H), 2.54 (s, 3H).

[0128] Example 23: Preparation of N 6 -(2-(4-methoxypiperidin-1-yl)benzo[d]thiazo-6-yl)-2-methylquinoline-4,6-diamine (I-23)

[0129] Structural formula:

[0130] Following the synthesis method of Example 22, the starting material 1,2,3,4-tetrahydroisoquinoline was replaced with an equimolar mass of 4-methoxypiperidine, with a yield of 48%. 1 H NMR (400 MHz, DMSO- d 6) δ 8.62 (s, 1H), 8.41 (brs, 2H), 7.71 (d, J = 9.1 Hz, 1H), 7.66 (t, J = 2.8 Hz, 2H), 7.56 (dd, J = 9.2, 2.3 Hz, 1H), 7.42 (d, J = 8.5 Hz, 1H), 7.13 (dd, J = 8.6, 2.3 Hz, 1H), 6.50 (s, 1H), 3.77 (dt, J = 13.2, 5.1 Hz, 2H), 3.48 (tt, J = 7.9, 3.4 Hz, 1H), 3.41 – 3.36 (m, 1H), 3.30 (s, 3H), 2.54 (s, 3H), 1.99 – 1.90 (m, 2H), 1.61 – 1.48 (m, 2H).

[0131] Example 24: Preparation of (6-((4-amino-2-methylquinoline-6-yl)amino)benzo[d]thiazol-2-yl)-L-phenylalanine tert-butyl ester (I-24)

[0132] Structural formula:

[0133] Referring to the synthesis method of Example 22, the starting material 1,2,3,4-tetrahydroisoquinoline was replaced with an equimolar mass of L-phenylalanine tert-butyl ester, with a yield of 23%. 1 H NMR (400 MHz, DMSO-d 6) δ 8.58 (s, 1H), 8.39 (brs,2H), 8.32 (d, J = 7.7 Hz, 1H), 7.71 (d, J = 9.1 Hz, 1H), 7.62 (d, J = 2.3 Hz, 1H), 7.59 (d, J = 2.3 Hz, 1H), 7.55 (dd, J = 9.1, 2.3 Hz, 1H), 7.34 (d, J = 8.5 Hz, 1H),7.33 – 7.28 (m, 4H), 7.27 – 7.21 (m, 1H), 7.08 (dd, J = 8.6, 2.3 Hz, 1H), 6.49(s, 1H), 4.56 (td, J = 8.1, 6.5 Hz, 1H), 3.13 – 3.00 (m, 2H), 2.54 (s, 3H), 1.33 (s, 9H).

[0134] Example 25: Preparation of 2-methyl-N 6 -(2-(3,4,5-trimethoxyphenyl)thieno[2,3-d]pyrimidin-4-yl)quinoline-4,6-diamine (I-25)

[0135] Structural formula:

[0136] Referring to the synthesis method of Example 13, the raw material 2,4-dichlorothiophene[3,2-d]pyrimidine was replaced with equimolar mass of 2,4-dichlorothiophene[2,3-d]pyrimidine, with a yield of 73%. 1 H NMR (400 MHz, DMSO- d 6) δ 10.20 (s,1H), 8.54 (s, 1H), 8.15 (d, J = 9.0 Hz, 1H), 7.93 (d, J = 5.9 Hz, 1H), 7.86 (d, J =9.0 Hz, 1H), 7.82 – 7.53 (m, 5H), 6.54 (s, 1H), 3.81 (s, 6H), 3.71 (s, 3H), 2.53 (s, 3H).

[0137] Example 26: Preparation of 2-methyl-N6 -(2-(3,4,5-trimethoxyphenyl)-5,7-dihydrofurano[3,4-d]pyrimidin-4-yl)quinoline-4,6-diamine (I-26)

[0138] Structural formula:

[0139] Referring to the synthesis method of Example 13, the starting material 2,4-dichlorothieno[3,2-d]pyrimidine was replaced with equimolar amounts of 2,4-dichloro-5,7-dihydrofurano[3,4-d]pyrimidine, with a yield of 75%. 1 H NMR (400 MHz, DMSO- d 6) δ 9.52 (s, 1H), 8.21 (d, J = 2.3 Hz, 1H), 7.86 (dd, J = 9.0, 2.3 Hz, 1H), 7.69 (d, J = 9.0 Hz, 1H), 7.66 (s, 2H), 6.50 (s, 2H), 6.43 (s, 1H), 5.01 (s, 2H), 4.98 –4.92 (m, 2H), 3.80 (s, 6H), 3.71 (s, 3H), 2.41 (s, 3H).

[0140] Example 27: Preparation of tert-butyl 4-((4-amino-2-methylquinoline-6-yl)amino)-2-(3,4,5-trimethoxyphenyl)-5,7-dihydro-6H-pyrrolo[3,4-d]pyrimidine-6-carboxylic acid (I-27)

[0141] Structural formula:

[0142] Referring to the synthesis method of Example 13, the starting material 2,4-dichlorothieno[3,2-d]pyrimidine was replaced with an equimolar mass of 6-Boc-2,4-dichloro-5,7-dihydropyrrolo[3,4-d]pyrimidine, with a yield of 60%. 1 H NMR (400 MHz, DMSO- d 6)δ 9.49 (s, 1H), 8.24 (t, J = 2.2 Hz, 1H), 7.88 (dd, J = 9.0, 2.3 Hz, 1H), 7.68(d, J= 9.1 Hz, 1H), 7.65 (s, 2H), 6.48 (s, 2H), 6.44 (s, 1H), 4.64 – 4.49 (m,4H), 3.80 (s, 6H), 3.70 (s, 3H), 2.40 (s, 3H), 1.49 (d, J = 5.4 Hz, 9H).

[0143] Example 28: Preparation of 2-methyl-N 6 -(2-(3,4,5-trimethoxyphenyl)-6,7-dihydro-5H-pyrrolo[3,4-d]pyrimidin-4-yl)quinoline-4,6-diamine hydrochloride (I-28)

[0144] Structural formula:

[0145] Synthesis method: 84 mg of I-27 was placed in a 25 ml round-bottom flask, 2 ml of 2 mol / L ethyl hydrochloride solution was added, the mixture was stirred at room temperature for 4 hours, filtered, washed several times with ethyl acetate, and dried to obtain a pale yellow solid I-28 (71 mg, 96%). 1 HNMR (400 MHz, DMSO- d 6) δ 10.40 – 10.28 (m, 2H), 10.24 (s, 1H), 8.82 (s, 1H), 8.69 – 8.54 (m, 2H), 8.16 (dd, J = 9.1, 2.1 Hz, 1H), 8.00 (d, J = 9.1 Hz, 1H),7.61 (s, 2H), 6.63 (d, J = 1.4 Hz, 1H), 4.65 – 4.58 (m, 2H), 4.51 (t, J = 5.8 Hz, 2H), 3.78 (s, 6H), 3.71 (s, 3H), 2.61 (s, 3H).

[0146] Example 29: Preparation of 2-methyl-N 6 -(2-(3,4,5-trimethoxyphenyl)quinazolin-4-yl)quinolin-4,6-diamine (I-29)

[0147] Structural formula:

[0148] Following the synthesis method of Example 13, the starting material 2,4-dichlorothiophene[3,2-d]pyrimidine was replaced with an equimolar mass of 2,4-dichloroquinazoline, with a yield of 82%. 1 H NMR (400 MHz, DMSO- d 6) δ 10.15 (s, 1H), 8.58 (d, J = 8.3 Hz, 1H), 8.39 (d, J = 2.3 Hz, 1H), 8.00 (dd, J = 9.0, 2.2 Hz, 1H), 7.90 –7.83 (m, 2H), 7.80 – 7.71 (m, 3H), 7.65 – 7.58 (m, 1H), 6.59 (s, 2H), 6.46(s, 1H), 3.80 (s, 6H), 3.71 (s, 3H), 2.43 (s, 3H).

[0149] Example 30: Preparation of 2-methyl-N 6 -(2-(3,4,5-trimethoxyphenyl)pyrrolo[2,1-f][1,2,4]triazin-4-yl)quinoline-4,6-diamine (I-30)

[0150] Structural formula:

[0151] Referring to the synthesis method of Example 13, the starting material 2,4-dichlorothieno[3,2-d]pyrimidine was replaced with equimolar mass of 2,4-dichloropyrrolo[2,1-f][1,2,4]triazine, with a yield of 71%. 1 H NMR (400 MHz, DMSO- d 6) δ 10.62(s, 1H), 8.90 – 8.60 (m, 3H), 8.31 (dd, J = 9.1, 2.1 Hz, 1H), 8.02 (d, J = 9.0Hz, 1H), 7.91 (dd, J = 2.6, 1.5 Hz, 1H), 7.55 (s, 2H), 7.31 (dd, J = 4.4, 1.6 Hz, 1H), 6.82 (dd, J = 4.4, 2.6 Hz, 1H), 6.65 (s, 1H), 3.81 (s, 6H), 3.71 (s, 3H), 2.63 (s, 3H).

[0152] Example 31: Preparation of tert-butyl 4-((4-amino-2-methylquinoline-6-yl)amino)-2-(3,4,5-trimethoxyphenyl)-7,8-dihydropyrido[4,3-d]pyrimidine-6(5H)-carboxylic acid (I-31)

[0153] Structural formula:

[0154] Referring to the synthesis method of Example 13, the starting material 2,4-dichlorothiopheno[3,2-d]pyrimidine was replaced with an equimolar mass of N-Boc-2,4-dichloro-5,7,8-trihydropyrido[4,3-D]pyrimidine (Leyan, catalog number 1061570), with a yield of 34%. 1 HNMR (400 MHz, DMSO- d 6) δ 8.99 (s, 1H), 8.31 (s, 1H), 7.93 (d, J = 9.0 Hz, 1H), 7.75 (d, J = 9.0 Hz, 1H), 7.57 (s, 2H), 7.15 (brs, 2H), 6.48 (s, 1H), 4.53 (s,2H), 3.75 (s, 6H), 3.73 – 3.65 (m, 5H), 2.84 (t, J = 5.8 Hz, 2H), 2.47 (s, 3H), 1.48 (s, 9H).

[0155] Example 32: Preparation of 2-methyl-N 6 -(2-(3,4,5-trimethoxyphenyl)-5,6,7,8-tetrahydropyrido[4,3-d]pyrimidin-4-yl)quinoline-4,6-diamine hydrochloride (I-32)

[0156] Structural formula:

[0157] Referring to the synthesis method of Example 28, the raw material I-28 was replaced with an equimolar mass of I-31, and the yield was 97%. 1 HNMR (400 MHz, DMSO- d 6) δ 10.17 (s, 2H), 9.78 (s, 1H), 8.83 (s, 1H), 8.74 –8.58 (m, 2H), 8.12 (dd, J = 9.0, 2.1 Hz, 1H), 8.00 (d, J= 9.1 Hz, 1H), 7.59 (s,2H), 6.62 (d, J = 1.4 Hz, 1H), 4.29 (t, J = 4.8 Hz, 2H), 3.77 (s, 6H), 3.70 (s, 3H), 3.55 – 3.47 (m, 2H), 3.23 – 3.12 (m, 2H), 2.62 (s, 3H).

[0158] Example 33 Biological Evaluation Experiment

[0159] (1) Small molecule assay method for PRMT7 enzyme activity

[0160] 1. Prepare 1×assay buffer (10 mM Tirs-HCl (pH=8.5), 0.01% Tween-20, 0.01% BSA and 1 mM dithiothreitol).

[0161] 2. First, prepare a stock solution (20 mM) of the compound by dissolving it in DMSO. Prepare serial dilutions of the compound using 1×assay buffer (compound concentrations of 10 μM, 5 μM, 2.5 μM, 1.25 μM, 0.63 μM, 0.31 μM, 0.16 μM, 0.08 μM, and 0.04 μM), keeping the final concentration of DMSO below 1%.

[0162] 3. Prepare assay buffer containing PRMT7 protein (15 nM, Active Motif, Cat. No. 31795) and... 3 A solution of H-SAM (0.5 μM, PerkinElmer, Cat. No. NET155V001MC).

[0163] 4. Prepare a 0.5 μM solution of biotin-labeled substrate peptide H4 (1-21) (sequence: Ac-SGRGKGGKGLGKGGAKRHRKVGG-K(Biotin), GL Biochem (Shanghai) Ltd., 415263) using 1× assay buffer.

[0164] 5. First, add 200 nL of a serially diluted solution of the compound to each well of a 384-well plate. Then, add 10 μL of a solution containing PRMT7 protein and... 3Add 10 μL of 1× assay buffer to the H-SAM solution and incubate at room temperature for 15 minutes.

[0165] 6. Add 10 μL of substrate peptide H4 solution to each well to initiate the enzyme-catalyzed reaction and incubate at room temperature for 240 minutes.

[0166] 7. Prepare the stop solution: 1 × assay buffer containing SAM (10 mM, Sigma, Cat. No. A7007-100MG).

[0167] 8. Add 10 μL of stop solution to each well to stop the enzyme-catalyzed reaction.

[0168] 9. Transfer 25 μL of the above reaction solution to a rapid plate and incubate at room temperature for 1 hour.

[0169] 10. Use a Microbeta instrument to read the signal value of each well.

[0170] 11. Use GraphPad 8.0 software to fit the inhibition rate-concentration curve based on the signal values ​​and calculate the IC. 50 value.

[0171] The results are shown in Table 1 below. As can be seen from the experimental results, the compound of the present invention has significant inhibitory activity against PRMT7 protein.

[0172] Table 1. Inhibitory activity of the compounds of the present invention against PRMT7 protein.

[0173]

[0174] (2) Small molecule inhibition of tumor cell proliferation experiment

[0175] Pancreatic cancer cells PATU-8988 (FH0544), breast cancer cells MDA-MB-231 (FH0213), colorectal cancer cells RKO (FH0030), lung cancer cells A549 (FH0045), ovarian cancer cells SKOV-3 (FH0135), prostate cancer cells DU-145 (FH0197), 22Rv1 (FH0198), VCaP (FH0194), and PC-3 (FH0195), esophageal cancer cells ECA109 (FH0225), liver cancer cells HEPG2 (FH0076), leukemia cells MOLM13 (FH1018), and glioma cells U251 (FH0159) were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum at 37 °C and 5% CO2. All cells were derived from Shanghai Fuheng Biotechnology Co., Ltd. Cell suspensions were diluted to 50,000 cells / mL with complete cell culture medium, and 100 μL of medium containing 5,000 cells were seeded into 96-well plates. The plates were incubated at 37 °C for one day at 5% CO2. Subsequently, 100 μL of culture medium containing a gradient concentration (10 μM, 5 μM, 2.5 μM, 1.25 μM, 0.63 μM, 0.31 μM, 0.16 μM, 0.08 μM, and 0.04 μM) of the test compound was added to each well, with three replicates for each dose group. DMSO was included as a control. After 48 hours of treatment, 10 μL of CCK-8 solution (10 mg / mL, Share-bio, SB-CCK8) was added to each well, and the plates were incubated at 37 °C for 4 hours. The OD value at 450 nm was read from each well using a microplate reader (Bio-Tek Synergy H1). Data were processed using Excel software, and nonlinear fitting and IC50 calculation were performed using GraphPad 8.0 software. 50 Value. Inhibition rate of cell proliferation = (OD value of control group) 450 -Experimental group OD 450 ) / Control group OD 450 *100%. The results are shown in Tables 2 and 3 below:

[0176] Table 2. Inhibitory activity of the compounds of the present invention against the PATU-8988 cell line.

[0177]

[0178] As can be seen from Table 2, most of the compounds in the embodiments of the present invention have strong inhibitory activity against tumor cell proliferation.

[0179] Table 3. Inhibitory activity of compound I-13 of the present invention against various cancer cell lines.

[0180]

[0181] As can be seen from Table 3, Example 13 (I-13) of the present invention has significant killing power against a variety of tumor cell lines, a wide range of treatments, and great potential for anti-tumor therapy.

Claims

1. A compound or a pharmaceutically acceptable salt thereof that inhibits the activity of PRMT7 protein, characterized in that, The general formula of the compound is shown in any one of formulas (II) to (V): ； R1 is selected from the following groups: ； Among them, R2, R3, R4, R5, and R6 are independently selected from methoxy, ethyl, and hydrogen; A and B are independently selected from CH or N.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, R1 is selected from the following groups: 。 3. A compound or a pharmaceutically acceptable salt thereof that inhibits the activity of PRMT7 protein, characterized in that, Its structural formula is shown below: 。 4. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, characterized in that, The pharmaceutically acceptable salt is the acid addition salt of the compound.

5. The compound according to claim 4 or a pharmaceutically acceptable salt thereof, characterized in that, The acid used for salt formation is an inorganic acid or an organic acid, wherein the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; and the organic acid is selected from acetic acid, trichloroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid, tartaric acid or methanesulfonic acid.

6. A pharmaceutical composition, characterized in that, It comprises the compound of any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable carrier thereof.

7. The use of the compound of any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition of claim 6 in the preparation of a PRMT7 target inhibitor drug.

8. The use of the compound of any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition of claim 6 in the preparation of a medicament for the prevention and / or treatment of cancer or tumor-related diseases.

9. The application according to claim 8, characterized in that, The cancer or tumor-related disease is selected from prostate cancer, colorectal cancer, ovarian cancer, leukemia, lung cancer, liver cancer, breast cancer, esophageal cancer, glioma, or pancreatic cancer.

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