CN114573581A

CN114573581A - 5-substituted amino-1, 3-disubstituted phenyl pyrido [2,3-d ] pyrimidine compound and preparation and application thereof

Info

Publication number: CN114573581A
Application number: CN202210322190.3A
Authority: CN
Inventors: 胡春; 谢倩; 李乐瑢; 王宇航; 文杰; 杨小力; 黄二芳; 彭子俊; 姚语桐; 林柔嘉; 金逸丹; 邓杰元
Original assignee: Shenyang Pharmaceutical University
Current assignee: Shenyang Pharmaceutical University
Priority date: 2022-03-30
Filing date: 2022-03-30
Publication date: 2022-06-03
Anticipated expiration: 2042-03-30
Also published as: CN114573581B

Abstract

The 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] of the invention]Pyrimidine compounds and preparation and application thereof, belonging to the technical field of medicine. The 5-substituted amino-1, 3-disubstituted phenyl pyrido [2,3-d]The structural general formula of the pyrimidine compound is shown as the formula (I): r₁Independently selected from hydrogen, fluorine; r₂Independently selected from hydrogen, methyl, benzyl; r₃、R₄Independently selected from hydrogen, methyl, cyclopropyl, 2-dimethylaminoethyl, 3-morpholinopropyl, 3-trifluoromethylphenyl, 4-trifluoromethoxyphenyl, 3-chloro-4-fluorophenyl, 3-chloro-4-trifluoromethylphenyl, 2-furylmethylOr R is₃、R₄Together with the nitrogen atom to which they are attached form a morpholinyl group. The compound has simple synthesis method, is suitable for industrial production, and biological activity tests show that the compound has antitumor activity and can be applied to preparation of antitumor drugs.

Description

5-substituted amino-1, 3-disubstituted phenyl pyrido [2,3-d ] pyrimidine compound and preparation and application thereof

Technical Field

The invention belongs to the technical field of medicines, and particularly relates to a 5-substituted amino-1, 3-disubstituted phenyl pyrido [2,3-d ] pyrimidine compound, and a preparation method and an application thereof.

Background

Malignant tumor is a serious disease seriously threatening human health, has plagued human for many years, and is a big factor hindering the increase of human life span. With the discovery of oncogenes and the elucidation of cell signaling pathways, human understanding of the mechanism of canceration of cells has been greatly enriched, and by analyzing the function of oncogene products, many oncoproteins have been found to be located at different sites of normal cell signaling pathways, playing an important role in promoting cell division and proliferation (Klein G. science,1987(4833): 1539-.

The mitogen-activated protein kinase MAPK signaling pathway is a very important chain of information transfer in cells. The MAPK pathway has three stages of signal transmission processes: MAPK, MAPK kinases (MEK or MKK) and kinases of MAPK kinases (MEKK or mkkkk). These three kinases, which are sequentially activated and which together regulate many important physiological/pathological effects such as cell growth, differentiation, stress, inflammatory response, are the major areas of tumor development induction (Orton RJ, Sturm OE, Vyshemirsky V, Calder M, Gilbert DR, Kolch W. the Biochemical journal.2005,392(2): 249-61). The common MAPK signal pathways in a human body mainly comprise a cell signal external kinase (ERK1/2) signal pathway, a c-Jun-N terminal kinase (JNK)/Stress Activated Protein (SAPK) signal pathway, a p38MAPK signal pathway, mitogen activated extracellular signal-regulated kinase (MEK) and the like, and the MAPK pathway inhibitor small-molecule drugs which are currently on the market mainly comprise:

trametinib (Trametinib) is a MEK inhibitor drug with anticancer activity that inhibits the activity of MEK1 and MEK2 kinases. Trametinib has achieved good results in phase III clinical trials for metastatic melanoma carrying the BRAF V600E mutation. In this mutation, the amino acid valine at position 600 of the BRAF protein has been substituted by glutamic acid, thus rendering the mutated BRAF protein constitutively active (Flaherty KT, Infante JR, Daud A, Gonzalez R, Kefford RF, Sosman J, et al. the New England Journal of medicine.2012,367(18): 1694-703).

Bimetinib (Binimetinib) is a selective inhibitor of MEK, developed by Array Biopharma for the treatment of various cancers. Binimetinib is a central kinase in the tumor-promoting MAPK pathway in MEK. Inappropriate activation of this pathway has been shown to occur in many cancers. 6 months 2018, it was approved by the FDA for use in combination with encorafenib in the treatment of patients with unresectable or metastatic BRAF V600E or V600K mutation-positive melanoma (Koelbinger P, Dornbierer J, Dummer R. future Oncology.2017,13(20): 1755-.

Sorafenib is an orally taken Raf kinase inhibitor, can directly inhibit Raf kinase activity, blocks signal transmission of a Ras pathway, and can also act on a platelet-derived growth factor receptor and a vascular endothelial growth factor receptor to down-regulate the expression level of p-MEK, thereby blocking a downstream Ras-Raf-MEK-ERK pathway. Sorafenib causes very low incidence of fatal adverse reactions such as gastrointestinal upset, skin toxicity, etc. (Wilhelm SM, Carter C, Tang L, et al cancer research.2004,64(19): 7099-.

Dalafinib (Dabraafinib) is a drug used for treating non-surgical or metastatic melanoma and can specifically act on B-Raf^V600EThe structural domain blocks MAPK pathway, inhibits the growth and proliferation of tumor cells and promotes the apoptosis of the tumor cells. And researches show that the parallel passage causing the adverse reaction of the B-Raf inhibitor can be regulated and controlled by combining the B-Raf kinase inhibitor and the MEK inhibitor, and the combination of the B-Raf kinase inhibitor and the MEK inhibitor can improve the response speed and reduce the toxicity. Patients often develop resistance to dabrafenib more rapidly, and therefore this problem can be solved by combination with the MEK inhibitor trametinib (Khoja L, Hogg D.Extert Review of Anticancer therapy.2015,112(6): 536-545).

Disclosure of Invention

The invention aims to provide a 5-substituted amino-1, 3-disubstituted phenyl pyrido [2,3-d ] pyrimidine compound, and preparation and application thereof, wherein the compound is a 5-substituted amino-1, 3-disubstituted phenyl pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione compound, and correspondingly, the compound is applied to the preparation of medicaments for treating or/and preventing tumor diseases related to MAPK pathways such as B-Raf kinase, Ras/Raf/MEK/ERK signal pathway, p38MAPK signal pathway, JNK-SAPK signal pathway and the like, and comprises the application of the compound as a multi-target inhibitor in the aspect of tumor resistance.

A 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidine compound represented by formula I, a prodrug and a pharmaceutically active metabolite thereof, and a pharmaceutically acceptable salt thereof:

R₁independently selected from hydrogen, halogen;

R₂independently selected from hydrogen, C1-C4 alkyl, benzyl;

R₃、R₄independently selected from hydrogen, C1-C4 alkyl, C3-C6 cycloalkyl, 2-dimethylaminoethyl, 3-morpholinopropyl, C1-C3 alkyl-substituted phenyl, halogen-substituted phenylSubstituted C1-C3 alkyl-substituted phenyl, or R₃、R₄Together with the nitrogen atom to which they are attached form a morpholinyl group.

Further, the compounds represented by formula I, prodrugs and pharmaceutically active metabolites thereof, and pharmaceutically acceptable salts thereof, according to the present invention:

R₁independently selected from hydrogen, fluorine;

R₂independently selected from hydrogen, methyl, benzyl;

R₃、R₄independently selected from hydrogen, methyl, cyclopropyl, 2-dimethylaminoethyl, 3-morpholinopropyl, 3-trifluoromethylphenyl, 4-trifluoromethoxyphenyl, 3-chloro-4-fluorophenyl, 3-chloro-4-trifluoromethylphenyl, 2-furylmethyl, or R₃、R₄Together with the nitrogen atom to which they are attached form a morpholinyl group.

Further, the present invention is preferably a compound as follows:

6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholinopropyl) amino ] pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T01), having the formula I-1:

6-benzyl-5- { [2- (dimethylamino) ethyl ] amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T02), having the formula I-2 below:

6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- (methylamino) pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T03), having the formula I-3 below:

5- { [ 3-chloro-4- (trifluoromethyl) phenyl ] amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T04), having the formula I-4 below:

1, 3-bis (4-fluorophenyl) -8-methyl-5- { [3- (trifluoromethyl) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T05), having the formula I-5 below:

1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethyl) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T06), having the formula I-6 below:

5- [ (3-chloro-4-fluorophenyl) amino ] -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T07), having the formula I-7 below:

1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethoxy) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T08), having the structural formula shown in the following formula I-8:

1, 3-bis (4-fluorophenyl) -5- [ (furan-2-ylmethyl) amino ] -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T09), having the formula I-9 below:

1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholinopropyl) amino ] pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T10), having the formula I-10 below:

6-benzyl-8-methyl-5- [ (3-morpholinopropyl) amino ] -1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T11), having the formula I-11:

6-benzyl-5- (cyclopropylamino) -8-methyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T12), which has the following formula I-12:

6-benzyl-8-methyl-5-morpholinyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T13), which has the following structural formula I-13:

6-benzyl-5- { [2- (dimethylamino) ethyl ] amino } -8-methyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T14), which has the following structural formula I-14:

6, 8-dimethyl-5- [ (3-morpholinopropyl) amino ] -1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T15), having the following structural formula I-15:

a pharmaceutical composition comprising as active ingredient a compound of any one of the compounds of the present invention, prodrugs and pharmaceutically active metabolites thereof, and pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable carrier or diluent.

"pharmaceutically acceptable salt" refers to conventional acid addition salts or base addition salts that retain the biological potency and properties of the compounds of formula I and are formed with non-toxic organic or inorganic acids or organic or inorganic bases. Acid addition salts include hydrochloride, hydrobromide, hydroiodide, nitrate, phosphate, sulfate, perchlorate, thiocyanate, bisulfate, persulfate, borate, formate, acetate, propionate, valerate, pivalate, hexanoate, heptanoate, octanoate, isooctanoate, undecanoate, laurate, palmitate, stearate, oleate, cyclopropionate, oxalate, malonate, succinate, maleate, fumarate, adipate, azelate, acrylate, strawberry, crotonate, glatironate, itaconate, sorbate, cinnamate, glycolate, lactate, malate, tartrate, citrate, tartrate, mandelate, diphenoxylate, troponate, ascorbate, gluconate, glucoheptonate, mandelate, dibenzolate, trogoplate, ascorbate, gluconate, glucoheptonate, and the like, Glucarate, mannonate, lactobionate, benzoate, phthalate, paraththalate, furoate, nicotinate, isonicotinate, salicylate, acetylsalicylate, caseinate, gallate, caffeate, ferulate, picrate, camphorate, camphorsulfonate, methanesulfonate, ethanesulfonate, propanesulfonate, benzenesulfonate, p-toluenesulfonate, sulfanilate, sulfamate, taurate, 2-hydroxyethanesulfonate, glycinate, alaninate, valine, leucine, isoleucine, phenylalanine, tryptophan, caseinate, aspartate, asparagine, glutamate, lysine, glutamine, methionine, serine, threonine, cysteine, proline, histidine, arginine, and salts thereof, Edetate, pyruvate, alpha-ketoglutarate, alginate, cyclopentanepropionate, 3-phenylpropionate, 3-cyclohexylpropionic acid, 2-naphthoate, 2-naphthalenesulfonate, pamoate, lauryl sulfate, glycerophosphate, lauryl sulfate, pectin oleate. Preferred acids for the formation of acid addition salts include hydrochloric acid, p-toluenesulfonic acid, methanesulfonic acid, maleic acid, malic acid, picric acid, citric acid, sulfanilic acid. Base addition salts include ammonium salts, alkali metal salts, such as sodium and potassium salts, alkaline earth metal salts, such as calcium and magnesium salts, salts with organic bases, such as dicyclohexylamine salts, N-methyl-D-glucamine salts, and basic nitrogen-containing groups may be quaternized with such agents as lower alkyl halides, such as methyl, ethyl, propyl and butyl chlorides, bromides and iodides; dialkyl sulfates such as dimethyl sulfate, diethyl sulfate, dibutyl sulfate and diamyl sulfate; long chain halides, such as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides; aralkyl halides such as benzyl and phenethyl bromides.

The invention also relates to a pharmaceutical composition for inhibiting MEK kinase and B-RAF kinase, which comprises the compound shown in the formula I or the derivative or the pharmaceutically acceptable acid addition salt thereof and a pharmaceutically acceptable carrier.

"pharmaceutically acceptable" such as pharmaceutically acceptable carriers, excipients, prodrugs, means pharmacologically acceptable and substantially non-toxic to a patient to whom a particular compound is administered.

"pharmaceutically active metabolite" refers to a pharmaceutically acceptable and effective metabolite of a compound of formula I.

The term "halogen" as used in the present invention includes fluorine, chlorine, bromine and iodine.

The compounds of the present invention may be administered to a patient by various methods, including orally in capsules or tablets, by administration as sterile solutions or suspensions, and in some cases, intravenously as solutions. The free base compounds of the present invention may be formulated and administered in the form of their pharmaceutically acceptable acid addition salts.

The compound is used as a small molecule inhibitor related to MAPK signal pathways such as B-Raf kinase, Ras/Raf/MEK/ERK signal pathways, p38MAPK signal pathways, JNK-SAPK signal pathways and the like in a brand new structure type, has the characteristics of novel structure type, can act on a plurality of targets and the like, can be used for preparing medicines for treating or preventing tumor diseases related to MAPK pathways such as lung cancer, liver cancer, melanoma, colon cancer, rectal cancer, breast cancer, ovarian cancer, cervical cancer and renal cancer, such as the B-Raf kinase, the Ras/Raf/MEK/ERK signal pathways, the p38MAPK signal pathways, the JNK-SAPK signal pathways and the like, and has good application value and development and application prospects.

The preparation route of the compound of the invention is as follows:

the compound T01-T15, pharmaceutically acceptable salts and prodrugs thereof can act on Ras/Raf/MEK/ERK signal pathways and inhibit phosphorylation of ERK1/2, thereby achieving the purpose of inhibiting tumor cell proliferation.

The invention has the beneficial effects that:

the invention provides a structure of a novel antitumor compound, which has potential patent medicine value, simple preparation route, easy synthesis and low cost. Compared with the marketed drugs, the compound has more remarkable anti-tumor cell proliferation activity.

Detailed Description

The present invention is described in detail by the following examples. It should be understood, however, that the present invention is not limited to the following examples which are specifically set forth. Example 1: preparation of 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholinopropyl) amino ] pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T01)

Step I: 40mL of methylene chloride dissolved phosgene solid (9.5g,32mmol) was weighed out into a 500mL round-bottom flask, para-fluoroaniline (8.89g, 80mmol) was weighed out into 40mL methylene chloride, and this was dropped into the flask with a dropping funnel at room temperature within 30min, and triethylamine (20.24g,0.2mol) was weighed out into 50mL methylene chloride and then into the flask within 30min, and para-fluoroaniline (8.89g, 80mmol) was weighed out again into 40mL methylene chloride and again dropped into the flask as described above. The reaction is stirred for 30min after the dripping is finished, and the p-fluoroaniline reaction is monitored to be complete. And (3) carrying out reduced pressure distillation to remove about 3/4 volume of dichloromethane, stirring with 5% HCl solution for 5-10min to generate a large amount of solid, filtering out the solid, repeatedly washing with diethyl ether, and drying to obtain the 1, 3-di-p-fluorophenylurea as a white solid with the yield of 18.5g and the yield of 93.16%.

Step II: 1, 3-Di-p-fluorophenylurea (4.96g,20mmol), malonic acid (2.7g,26mmol), phosphorus oxychloride (3.4g,22mmol) and 15mL acetonitrile were weighed into a 35mL sealed tube. Sealing and heating to 120 ℃, reacting for 1h, monitoring the reaction completion of raw materials, pouring the reaction liquid in the sealed tube into an eggplant-shaped bottle, distilling under reduced pressure to remove all solvents, adding 50mL of 2mol/L NaOH and 20mL of ethyl acetate, and stirring and dissolving. The organic phase was extracted three more times with 2mol/L aqueous NaOH, the aqueous layers were combined, the aqueous layer was extracted once with a small amount of ethyl acetate, and the remaining aqueous layer was adjusted to pH with hydrochloric acid to give a large amount of solid. The solid was filtered off, the filter cake was washed with water and dried to give 1, 3-bis (4-fluorophenyl) pyrimidine-2, 4,6(1H,3H,5H) -trione as a yellow-white solid 3.87g, yield 61.18%.

Step III: 1, 3-bis (4-fluorophenyl) pyrimidine-2, 4,6(1H,3H,5H) -trione (5.12g,16.2mmol), phosphorus oxychloride (4.92g,32.4mmol) and 20mL acetonitrile were weighed into a 125mL sealed tube. Sealing and heating to 120 ℃ for reaction for 3H, monitoring the reaction completion of raw materials, pouring the reaction liquid in a sealed tube into an eggplant-shaped bottle, distilling under reduced pressure to remove all solvents, adding a small amount of ethanol to dissolve the mixture, adding water to precipitate a solid, filtering out the solid, washing a filter cake with water, and drying to obtain a yellow solid, namely 5.12g of 6-chloro-1, 3-bis (4-fluorophenyl) pyrimidine-2, 4(1H,3H) -diketone, with the yield of 94.43%.

Step IV: 6-chloro-1, 3-bis (4-fluorophenyl) pyrimidine-2, 4(1H,3H) -dione (3.34g,10mmol), 40% methylamine water solution (1.28g,15mmol), triethylamine (2.22g,20mmol) and 20mL ethanol were weighed into a 50mL round-bottomed flask, and the reaction was completed after stirring and refluxing for 8 hours. Most of the ethanol was distilled off under reduced pressure, and diethyl ether was added to stir the solid out. The solid was filtered off, washed with diethyl ether and dried to give 6-methylamino-1, 3-bis (4-fluorophenyl) pyrimidine-2, 4(1H,3H) -dione as a yellow solid in a yield of 3.16g, 95.96% yield.

Step V: 6-methylamino-1, 3-bis (4-fluorophenyl) pyrimidine-2, 4(1H,3H) -dione (1.65g,5mmol), diethyl benzylmalonate (1.50g,6mmol) and 15g diphenyl ether are weighed into a 100mL three-neck round-bottom flask, heated to 256 ℃ on a heating jacket and then refluxed for 40min, TLC monitors that the raw materials are completely reacted, the reaction liquid is poured into 30mL petroleum ether, a large amount of solid is separated out, filtered, dried and purified by column chromatography to obtain a pure product of 6-benzyl-5-hydroxy-1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione, a white solid, the yield is 2.00g, and the yield is 82.07%.

Step VI: 6-benzyl-5-hydroxy-1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (3.39g,6.95mmol), triethylamine (1.06g,10.44mmol), 0.1g DMAP and 15mL acetonitrile were weighed into a 100mL round-bottomed flask, 5mL acetonitrile was weighed to dissolve p-toluenesulfonyl chloride (2.54g,10.17mmol) in an Erlenmeyer flask, the solution was slowly dropped into the flask, stirred at room temperature for 10min and then heated to reflux for 5H, and the completion of the reaction was monitored by TLC. And (3) distilling under reduced pressure to remove the solvent, adding ethyl acetate, stirring to obtain a large amount of solid, carrying out suction filtration and drying to obtain the product 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydro-pyrido [2,3-d ] pyrimidine-5-yl p-methyl benzene sulfonate which is a yellow white solid with the yield of 4.22g and the yield of 94.63%.

Step VII: weighing 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.91g,1.41mmol), N-aminopropylmorpholine (0.28g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of N-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h, and the completion of the reaction was monitored by TLC. Cooling the reaction solution to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and dryingDrying to obtain 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholine propyl) amino]Pyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T01), white solid, yield 0.33g, yield 38.13%; m.p.: 112.4-116.2 deg.C; MS 614.4[ M + H ]]⁺；¹HNMR(400MHz,DMSO-d₆)δ9.03(t,J＝5.2Hz,1H),7.53(dd,J＝8.9,4.9Hz,2H),7.40–7.23(m,8H),7.15(dd,J＝7.4,4.1Hz,3H),4.01(s,2H),3.46(s,4H),3.26–3.30(m,2H),2.75(s,3H),2.18(s,6H),1.56(s,2H)。

Example 2: preparation of 6-benzyl-5- { [2- (dimethylamino) ethyl ] amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T02)

The compound 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ] pyrimidin-5-yl p-methylbenzenesulfonate was prepared by the method of example 1.

Step VII: weighing 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.91g,1.41mmol), N-dimethylethylenediamine (0.17g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of N-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h, and the completion of the reaction was monitored by TLC. Cooling the reaction solution to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain 6-benzyl-5- { [2- (dimethylamino) ethyl]Amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T02), white solid, yield 0.31g, 39.43% yield; m.p.: 85.7-95.3 ℃; MS 558.22992[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ9.11(t,J＝4.6Hz,1H),7.55–7.50(m,2H),7.41–7.36(m,4H),7.35–7.31(m,2H),7.27–7.23(m,2H),7.16–7.11(m,3H),4.05(s,2H),3.37–3.27(m,2H),2.75(s,3H),2.25(t,J＝6.1Hz,2H),1.95(s,6H)。

Example 3: preparation of 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- (methylamino) pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T03)

Step VII: weighing 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl-p-methylbenzenesulfonate (0.91g,1.41mmol), 40% aqueous methylamine solution (0.17g,1.95mmol), triethylamine (0.23g,2.25mmol), and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottom flask and refluxed for 8h, and the reaction was monitored by TLC for completion. After the reaction liquid is cooled to room temperature, a large amount of crystals are separated out, the crystals are filtered out, repeatedly washed by methanol and dried to obtain the 6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- (methylamino) pyrido [2,3-d ]]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T03), white solid, yield 0.52g, yield 73.68%; m.p.: 257.9-262.0 ℃; MS 501.17163[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ9.03(q,J＝5.3Hz,1H),7.58–7.49(m,2H),7.41–7.24(m,8H),7.18–7.11(m,3H),4.07(s,2H),2.90(d,J＝5.3Hz,3H),2.75(s,3H)。

Example 4: preparation of 5- { [ 3-chloro-4- (trifluoromethyl) phenyl ] amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T04)

The compound 6-methylamino-1, 3-bis (4-fluorophenyl) pyrimidine-2, 4(1H,3H) -dione was prepared by the method of example 1.

Step V: weighing 6-methylamino-1, 3-bis (4-fluorophenyl) pyrimidine-2, 4(1H,3H) -diketone (1.65g,5mmol), diethyl malonate (0.96g,6mmol) and 15g diphenyl ether in a 100mL three-neck round-bottom flask, heating to 256 ℃ on a heating jacket, refluxing for 40min, monitoring the reaction of the raw materials by TLC, pouring the reaction liquid into 30mL petroleum ether, precipitating a large amount of solid, performing suction filtration, drying, and purifying by column chromatography to obtain a pure product 1, 3-bis (4-fluorophenyl) -5-hydroxy-8-methylpyrido [2,3-d ]]Pyrimidine-2, 4,7(1H,3H,8H) -trione (XQA20), white solid, yield 1.39g, 69.97% yield; m.p.: 246.3 to 253.6 ℃; MS 398.09290[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ12.00(s,1H),7.60–7.54(m,2H),7.43–7.34(m,6H),5.63(s,1H),2.73(s,3H)。

Step VI: 1, 3-bis (4-fluorophenyl) -5-hydroxy-8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (2.66g,6.69mmol), triethylamine (1.06g,10.44mmol), 0.1g DMAP and 15mL acetonitrile were weighed into a 100mL round-bottomed flask, 5mL acetonitrile was weighed to dissolve p-toluenesulfonyl chloride (2.54g,10.17mmol) in an Erlenmeyer flask, the solution was slowly dropped into the flask, stirred at room temperature for 10min and then heated under reflux for 5H, and the reaction was monitored by TLC for completion. The solvent is removed by reduced pressure distillation, ethyl acetate is added, a large amount of solid is stirred out, and the product 1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydro-pyrido [2,3-d ] pyrimidine-5-yl p-methyl benzene sulfonate is obtained after suction filtration and drying, and is yellow white solid, the yield is 3.49g, and the yield is 94.62%.

Step VII: weighing 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.83g,1.5mmol), 3-chloro-4-trifluoromethylaniline (0.38g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h and monitored by TLC for completion. Cooling the reaction solution to room temperature, precipitating a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain 5- { [ 3-chloro-4- (trifluoromethyl) phenyl]Amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T04), white solid, yield 0.56g, 64.94% yield; m.p.: 265.2-269.7 ℃; MS 575.08948[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ10.58(s,1H),7.78–7.65(m,3H),7.61–7.53(m,2H),7.46–7.31(m,6H),5.65(s,1H),2.74(s,3H)。

Example 5: preparation of 1, 3-bis (4-fluorophenyl) -8-methyl-5- { [3- (trifluoromethyl) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T05)

The compound 1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ] pyrimidin-5-yl p-methylbenzenesulfonate was obtained by the method of example 4.

Step VII: weighing 1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.83g,1.5mmol), 3-trifluoromethylaniline (0.31g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol in a 50mL round-bottomed flask, heating to 120 ℃ for reflux reaction for 8h, TLCThe reaction was monitored for completion. Cooling the reaction solution to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain the 1, 3-bis (4-fluorophenyl) -8-methyl-5- { [3- (trifluoromethyl) phenyl]Amino } pyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T05), white solid, yield 0.62g, 76.48% yield; m.p.: 197.2-201.3 deg.C; MS 541.12878[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ10.59(s,1H),7.70–7.53(m,6H),7.46–7.32(m,6H),5.60(s,1H),2.74(s,3H)。

Example 6: preparation of 1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethyl) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T06)

Step VII: weighing 1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.83g,1.5mmol), 4-trifluoromethylaniline (0.31g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h, and the completion of the reaction was monitored by TLC. Cooling the reaction solution to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain the 1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethyl) phenyl]Amino } pyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T06), white solid, yield 0.65g, yield 80.18%; m.p.: 199.0 to 201.7 ℃; MS 541.12787[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ10.74(s,1H),7.77–7.73(m,2H),7.61–7.51(m,4H),7.45–7.32(m,6H),5.81(s,1H),2.74(s,3H)。

Example 7: 5- [ (3-chloro-4-fluorophenyl) amino ] -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T07):

Step VII: the weighing is carried out by 1, and,3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d]Pyrimidin-5-yl-p-methylbenzenesulfonate (0.83g,1.5mmol), 3-chloro-4-fluoroaniline (0.29g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h and monitored by TLC for completion. Cooling the reaction solution to room temperature, precipitating a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain 5- [ (3-chloro-4-fluorophenyl) amino]-1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T07), white solid, yield 0.56g, yield 71.13%; m.p.: 250.3-254.2 ℃; MS 525.09283[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ10.36(s,1H),7.60–7.52(m,3H),7.49–7.31(m,8H),5.47(s,1H),2.73(s,3H)。

Example 8: 1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethoxy) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T08):

Step VII: weighing 1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.83g,1.5mmol), 4-trifluoromethoxyaniline (0.35g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h, and the completion of the reaction was monitored by TLC. Cooling the reaction solution to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain the 1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethoxy) phenyl]Amino } pyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T08), white solid, yield 0.62g, yield 74.28%; m.p.: 212.7-220.2 ℃; MS 557.12305[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ10.49(s,1H),7.60–7.52(m,2H),7.47–7.30(m,10H),5.56(s,1H),2.73(s,3H)。

Example 9: preparation of 1, 3-bis (4-fluorophenyl) -5- [ (furan-2-ylmethyl) amino ] -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T09)

Step VII: weighing 1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.83g,1.5mmol), 2-furanmethanamine (0.19g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and refluxed for 8h, and the completion of the reaction was monitored by TLC. Cooling the reaction liquid to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing the crystals with methanol, and drying to obtain the 1, 3-bis (4-fluorophenyl) -5- [ (furan-2-ylmethyl) amino]-8-methylpyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T09), white solid, yield 0.43g, yield 60.17%; m.p.: 195.2-198.4 ℃; MS 477.13617[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ8.96(t,J＝5.6Hz,1H),7.62–7.48(m,3H),7.41–7.26(m,6H),6.43–6.36(m,2H),5.40(s,1H),4.41(d,J＝5.5Hz,2H),2.68(s,3H)。

Example 10: preparation of 1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholinopropyl) amino ] pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T10)

Step VII: weighing 1, 3-bis (4-fluorophenyl) -8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.83g,1.5mmol), N-aminopropylmorpholine (0.28g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of N-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h and monitored by TLC for completion. Cooling the reaction liquid to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing the crystals with methanol, and drying to obtain the 1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholinepropyl) amino]Pyrido [2,3-d]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T29), white solid, yield 0.41g, yield 52.21%; m.p.: 91.8-93.6 deg.C; MS 524.2[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ8.69(t,J＝5.3Hz,1H),7.51(dd,J＝8.9,4.8Hz,2H),7.38(dd,J＝8.7,4.5Hz,4H),7.32(d,J＝9.2Hz,2H),5.25(s,1H),3.56(t,J＝4.6Hz,4H),3.18(q,J＝6.4Hz,2H),2.68(s,3H),2.35(s,6H),1.73(p,J＝6.8Hz,2H)。

Example 11: preparation of 6-benzyl-8-methyl-5- [ (3-morpholinopropyl) amino ] -1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T11)

Step I: weigh 40mL of methylene chloride dissolved phosgene solid (9.5g,32mmol) into a 500mL round bottom flask, weigh aniline (7.45g,80mmol) into 40mL methylene chloride, drop it into the flask with a dropping funnel over 30min at room temperature, weigh triethylamine (20.24g,0.2mol) into 50mL methylene chloride and drop it into the flask over 30min, weigh aniline (7.4g,80mmol) again into 40mL methylene chloride, and drop it into the flask again as described above. The reaction is stirred for 30min after the dripping is finished, and the p-fluoroaniline reaction is monitored to be complete. And (3) carrying out reduced pressure distillation to remove about 3/4 volume of dichloromethane, stirring with 5% HCl solution for 5-10min to generate a large amount of solid, filtering out the solid, repeatedly washing with diethyl ether, and drying to obtain the 1, 3-diphenyl urea as a white solid with the yield of 16.17g and the yield of 93.25%.

Step II: 1, 3-diphenylurea (4.25g,20mmol), malonic acid (2.7g,26mmol), phosphorus oxychloride (3.4g,22mmol) and 15mL acetonitrile were weighed into a 35mL sealed tube. Sealing and heating to 120 ℃ for reacting for 1h, monitoring the reaction of the raw materials, pouring the reaction liquid in the sealed tube into an eggplant-shaped bottle, distilling under reduced pressure to remove all solvents, adding 50mL of 2mol/L NaOH and 20mL of ethyl acetate, and stirring to dissolve. The organic phase was extracted three more times with 2mol/L aqueous NaOH, the aqueous layers were combined, the aqueous layer was extracted once with a small amount of ethyl acetate, and the remaining aqueous layer was adjusted to pH with hydrochloric acid to give a large amount of solid. The solid was filtered off, the filter cake was washed with water and dried to give 1, 3-diphenylpyrimidine-2, 4,6(1H,3H,5H) -trione as a yellowish white solid 3.75g, in 66.90% yield.

Step III: 1, 3-diphenylpyrimidine-2, 4,6(1H,3H,5H) -trione (4.54g,16.2mmol), phosphorus oxychloride (4.92g,32.4mmol) and 20mL of acetonitrile were weighed into a 125mL sealed tube. Sealing and heating to 120 ℃ for reaction for 3H, monitoring the reaction completion of raw materials, pouring the reaction liquid in a sealed tube into an eggplant-shaped bottle, distilling under reduced pressure to remove all solvents, adding a small amount of ethanol to dissolve the mixture, adding water to precipitate a solid, filtering out the solid, washing a filter cake with water, and drying to obtain a yellow solid, namely 4.58g of 6-chloro-1, 3-diphenylpyrimidine-2, 4(1H,3H) -diketone, with the yield of 94.64%.

Step IV: 6-chloro-1, 3-diphenylpyrimidine-2, 4(1H,3H) -dione (2.99g,10mmol), 40% methylamine aqueous solution (1.28g,15mmol), triethylamine (2.22g,20mmol) and 20mL of ethanol were weighed out in a 50mL round-bottomed flask, and the reaction was completed after stirring and refluxing for 8 hours. Most of the ethanol was distilled off under reduced pressure, and diethyl ether was added to stir the solid out. The solid was filtered off, washed with diethyl ether and dried to give 6-methylamino-1, 3-diphenylpyrimidine-2, 4(1H,3H) -dione as a yellow solid in a yield of 2.76g, 94.09%.

Step V: weighing 6-methylamino-1, 3-diphenylpyrimidine-2, 4(1H,3H) -dione (1.47g,5mmol), diethyl benzylmalonate (1.50g,6mmol) and 15g diphenyl ether in a 100mL three-neck round-bottom flask, heating to 256 ℃ on a heating jacket, carrying out reflux reaction for 40min, after the TLC monitors that the raw materials are completely reacted, pouring the reaction liquid into 30mL petroleum ether, separating out a large amount of solid, carrying out suction filtration, drying, and purifying by column chromatography to obtain a pure product of 6-benzyl-5-hydroxy-1, 3-diphenyl-8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione, wherein the yield is 1.39g and the yield is 61.57%.

Step VI: 6-benzyl-5-hydroxy-1, 3-diphenyl-8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (2.42g,5.36mmol), triethylamine (1.06g,10.44mmol), 0.1g DMAP and 15mL acetonitrile were weighed into a 100mL round-bottomed flask, 5mL acetonitrile was weighed to dissolve p-toluenesulfonyl chloride (2.54g,10.17mmol) in an Erlenmeyer flask, the solution was slowly dropped into the flask, stirred at room temperature for 10min and then heated under reflux for 5H, and the reaction was monitored by TLC for completion. The solvent is removed by reduced pressure distillation, ethyl acetate is added, a large amount of solid is stirred out, and the product 6-benzyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydro-pyrido [2,3-d ] pyrimidine-5-yl p-methyl benzene sulfonate is obtained after suction filtration and drying, and is yellow white solid, the yield is 3.07g, and the yield is 94.62%.

Step VII: weighing 6-benzyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl-p-methylBenzenesulfonate (0.91g,1.5mmol), N-aminopropylmorpholine (0.28g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of N-butanol were heated to 120 ℃ in a 50mL round bottom flask to reflux the reaction for 8h and TLC monitored for reaction completion. Cooling the reaction liquid to room temperature, separating out a great amount of crystals, filtering out the crystals, repeatedly washing the crystals with methanol, and drying to obtain the 6-benzyl-8-methyl-5- [ (3-morpholinopropyl) amino]-1, 3-diphenylpyrido [2,3-d ]]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T11), white solid, yield 0.4g, 46.16% yield; m.p.: 180.6 to 185.1 ℃; MS 578.27502[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ9.06(t,J＝5.2Hz,1H),7.57–7.37(m,8H),7.36–7.23(m,4H),7.20–7.10(m,3H),4.01(s,2H),3.44(t,J＝4.5Hz,4H),3.35–3.26(m,3H),2.71(s,3H),2.18(t,J＝4.5Hz,4H),2.13(t,J＝6.8Hz,2H),1.54(p,J＝6.9Hz,2H)。

Example 12: preparation of 6-benzyl-5- (cyclopropylamino) -8-methyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T12)

The compound 6-benzyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ] pyrimidin-5-yl p-methylbenzenesulfonate was prepared by the method of example 11.

Step VII: weighing 6-benzyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.91g,1.5mmol), cyclopropylamine (0.12g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h and monitored by TLC for completion. Cooling the reaction liquid to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing the crystals with methanol, and drying to obtain the 6-benzyl-5- (cyclopropylamino) -8-methyl-1, 3-diphenylpyrido [2,3-d ]]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T12), white solid, yield 0.29g, 39.41% yield; m.p.: 152.7-159.4 ℃; MS 491.20706[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ9.32(d,J＝3.5Hz,1H),7.57–7.37(m,8H),7.35–7.23(m,4H),7.20–7.10(m,3H),4.33(s,2H),2.71(s,3H),2.66–2.58(m,1H),0.63(d,J＝6.7Hz,2H),0.56–0.46(m,2H)。

Example 13: preparation of 6-benzyl-8-methyl-5-morpholinyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T13)

Step VII: weighing 6-benzyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.91g,1.5mmol), morpholine (0.17g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of n-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h and monitored by TLC for completion. Cooling the reaction liquid to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing the crystals with methanol, and drying to obtain the 6-benzyl-8-methyl-5-morpholinyl-1, 3-diphenylpyrido [2,3-d ]]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T13), white solid, yield 0.29g, 37.14% yield; m.p.: 85.3-89.7 ℃; MS 521.21753[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ7.48(d,J＝8.1Hz,2H),7.42–7.22(m,8H),7.18(dd,J＝12.5,6.9Hz,4H),7.09–6.98(m,1H),4.06(d,J＝15.5Hz,1H),3.95(d,J＝15.5Hz,1H),3.30(s,3H),3.08–2.87(m,4H)。

Example 14: preparation of 6-benzyl-5- { [2- (dimethylamino) ethyl ] amino } -8-methyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T14)

Step VII: weighing 6-benzyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl p-methylbenzenesulfonate (0.91g,1.5mmol), N-dimethylethylenediamine (0.17g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL of N-butanol were heated to 120 ℃ in a 50mL round-bottomed flask and the reaction was refluxed for 8h, and the completion of the reaction was monitored by TLC. Cooling the reaction solution to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain 6-benzyl-5- { [2- (dimethylamino) ethyl]Amino } -8-methyl-1, 3-diphenylpyrido [2, 3-d)]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T14),white solid, yield 0.34g, 43.45% yield; m.p.: 181.4-186.4 ℃; MS 522.24884[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ9.16(t,J＝4.7Hz,1H),7.56–7.41(m,8H),7.36–7.26(m,4H),7.19–7.12(m,3H),4.07(s,2H),3.38–3.33(m,2H),2.73(s,3H),2.28(t,J＝6.2Hz,2H),1.97(s,6H)。

Example 15: preparation of 6, 8-dimethyl-5- [ (3-morpholinopropyl) amino ] -1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (T15)

The compound 6-methylamino-1, 3-bis (4-fluorophenyl) pyrimidine-2, 4(1H,3H) -dione was prepared by the method of example 11.

Step V: weighing 6-methylamino-1, 3-diphenylpyrimidine-2, 4(1H,3H) -dione (1.47g,5mmol), diethyl methylmalonate (1.04g,6mmol) and 15g diphenyl ether in a 100mL three-neck round-bottom flask, heating to 256 ℃ on a heating jacket, carrying out reflux reaction for 40min, after the TLC monitors that the raw materials are completely reacted, pouring the reaction liquid into 30mL petroleum ether, separating out a large amount of solid, carrying out suction filtration, drying, and purifying by column chromatography to obtain a pure product of 6-methyl-5-hydroxy-1, 3-diphenyl-8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione, wherein the yield is 1.05g and the yield is 55.94%.

Step VI: 6-methyl-5-hydroxy-1, 3-diphenyl-8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione (2.51g,6.66mmol), triethylamine (1.06g,10.44mmol), 0.1g DMAP and 15mL acetonitrile were weighed into a 100mL round-bottomed flask, 5mL acetonitrile was weighed to dissolve p-toluenesulfonyl chloride (2.54g,10.17mmol) in an Erlenmeyer flask, the solution was slowly dropped into the flask, stirred at room temperature for 10min and then heated under reflux for 5H, and the reaction was monitored by TLC for completion. The solvent is removed by reduced pressure distillation, ethyl acetate is added, a large amount of solid is stirred out, and the product 6-benzyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydro-pyrido [2,3-d ] pyrimidine-5-yl p-methyl benzene sulfonate is obtained after suction filtration and drying, and is yellow white solid, the yield is 3.34g, and the yield is 94.62%.

Step VII: weighing 6-methyl-1, 3-diphenyl-8-methyl-2, 4, 7-trioxo-1, 2,3,4,7, 8-hexahydropyrido [2,3-d ]]Pyrimidin-5-yl-p-methylbenzenesulfonate (0.79g,1.5mmol), N-aminopropylmorpholine (0.28g,1.95mmol), triethylamine (0.23g,2.25mmol) and 15mL n-butanol was heated to 120 ℃ in a 50mL round bottom flask and refluxed for 8h, and the completion of the reaction was monitored by TLC. Cooling the reaction solution to room temperature, separating out a large amount of crystals, filtering out the crystals, repeatedly washing with methanol, and drying to obtain 6, 8-dimethyl-5- [ (3-morpholinepropyl) amino]-1, 3-diphenylpyrido [2,3-d ]]Pyrimidine-2, 4,7(1H,3H,8H) -trione (T15), white solid, yield 0.29g, yield 38.54%; m.p.: 82.4-84.3 ℃; MS 502.2[ M + H ]]⁺；¹H NMR(400MHz,DMSO-d₆)δ8.62–8.64(m 1H),7.51–7.41(m,8H),7.34–7.28(m,2H),3.52(t,J＝4.6Hz,4H),3.43(q,J＝6.5Hz,2H),2.68(s,3H),2.36–2.24(m,6H),2.06(s,3H),1.65(p,J＝7.0Hz,2H)。

Pharmacological examples

Example 16: inhibitory Activity of test Compounds on MCF7, A375, SK-Mel-2 cell proliferation

(1) Experimental materials

Cell line: MCF7, A375 and SK-Mel-2 cells were plated in 96-well plates at densities of 4000, 5000 and 4000/well, 200 ul/well, and used after 24 h.

Code T01-T15 target compound: dissolved in DMSO, diluted with culture medium to obtain six different concentrations of 100. mu.M, 50. mu.M, 25. mu.M, 12.5. mu.M, 6.25. mu.M and 3.125. mu.M, and stored at-20 deg.C for use, with the final concentration of DMSO in the culture medium being less than 0.1%.

Positive control drug: 5-Fluorouracil (5-Fu, Fluorouracil).

MTT: dissolved in PBS to 2mg/mL and stored at-20 ℃.

(2) Experimental methods

MCF7, A375, SK-Mel-2 cells were selected to evaluate the anti-tumor proliferation activity of test samples using the MTT method. MCF7, A375, SK-Mel-2 cell lines were cultured in DMEM medium containing 10% bovine serum (FBS). When the cells proliferated to 80-90% they were pooled and subsequently subcultured for no more than 20 passages, and then they were acclimatized for 24h before the next disposal. These cells were plated in 96-well plates and then incubated in a medium containing 5% CO₂Culturing in an incubator at constant temperature of 37 ℃ until the cells are completely attached to the wall. After 24h, various concentrations of representative compounds of the invention were added. After an additional 24h of incubation, MTT (2mg/mL) was added and incubation continued for 4 h. The culture medium was removed, the crystals were dissolved in DMSO, and the absorbance was measured at a wavelength of 570nm using a microplate reader (Thermo Multiskan GO, Thermo Fisher, USA). According to the formula: the cell growth inhibition rate is (1-drug group OD value/control group OD value) × 100%, the cell growth inhibition rate under the corresponding concentration is calculated, and the IC corresponding to the tested compound is calculated according to the logarithmic curve of the inhibition rate of the tested compound to the cell and the different concentrations of the tested compound₅₀The value is obtained. Representative compounds of the invention were tested according to the methods described above.

Most of the 15 compounds prepared above showed moderate tumor cell proliferation inhibition activity on MCF-7, A375 and SK-MEL-2 cell lines, wherein T06 showed strong proliferation inhibition activity on MCF7, A375 and SK-MEL-2, IC₅₀As low as 24.02 + -0.45 μ M, 25.81 + -0.69 μ M and 29.28 + -0.53 μ M.

Formulation examples

The following formulation examples are merely illustrative of the scope of the invention and are not to be construed as limiting in any way. The active compounds described in the following examples are the compounds T01 to T15 prepared in the above examples.

Example 17: tablet formulation

25-1000mg of active compound, 45mg of starch, 35mg of microcrystalline cellulose, 4mL of polyvinylpyrrolidone (which is a 10% aqueous solution), 4.5mg of sodium carboxymethyl cellulose, 0.5mg of magnesium stearate, and 1mg of talc.

Example 18: suspending agent formula

0.1-1000mg of active compound, 50mg of sodium carboxymethylcellulose, 1.25mg of syrup, 0.1mg of sodium benzoate, 25mg of flavoring agent and 5mg of coloring agent, and pure water is added to the volume of 5 mL.

Example 19: aerosol formulations

0.25mg of active compound, 25-75mL of ethanol and 70mg of propellant 22 (chlorodifluoromethane).

Example 20: suppository formula

250mg of active compound, 2000mL of saturated fatty acid glycerides.

Example 21: injectable formulation

50mg of active compound, 1000mL of isotonic saline solution.

Example 22: ointment formulation

0.025g of micronized active compound, 10g of liquid paraffin, and 100g of soft white wax.

Example 23: ointment formulation

0.025g of active compound, 5g of propylene glycol, 5g of sorbitan sesquioleate, 10g of liquid paraffin and 100g of soft white wax.

Example 24: oil-in-water cream formulation

0.025g of active compound, 5g of cetyl alcohol, 5g of glycerol monostearate, 10g of liquid paraffin, 2g of cetyl polyoxyethylene ether, 0.1g of citric acid, 0.2g of sodium citrate, 35g of propylene glycol and water to 100 g.

Example 25: oil-in-water cream formulation

0.025g of micronized active compound, 15g of soft white wax, 5g of liquid paraffin, 5g of cetyl alcohol, 2g of Sorbimcarol stearate (Tween 65 of the particular pharmaceutical excipient grade), 0.5g of sorbitan monostearate, 0.2g of sorbic acid, 0.1g of citric acid, 0.2g of sodium citrate, and water to 100 g.

Example 26: water-in-oil cream formulation

0.025g of active compound, 35g of soft white wax, 5g of liquid paraffin, 5g of sorbitan sesquioleate, 0.2g of sorbic acid, 0.1g of citric acid and 0.2g of sodium citrate, and water is added until the weight is 100 g.

Example 27: lotion formulation

0.25g of active compound, 0.5mL of isopropanol, 3mg of carboxyvinyl polymer, 2mg of NaOH and water to 1 g.

Example 28: suspension formulation for injection

10mg of the active compound, 7mg of sodium carboxymethylcellulose, 7mg of NaCl, 0.5mg of polyoxyethylene (20) sorbitan monooleate, 8mg of benzyl alcohol, and sterile water to 1 mL.

Example 29: aerosol formulation for oral and nasal inhalation

0.1% w/w active compound, 0.7% w/w sorbitan trioleate, 24.8% w/w trichlorofluoromethane, 24.8% w/w dichlorotetrafluoroethane and 49.6% w/w dichlorodifluoromethane.

Example 30: atomized solution formulation

7mg of active compound, 5mg of propylene glycol, water to 10 g.

Example 31: powder formulations for inhalation

Gelatine capsules were filled with a mixture of the following ingredients, micronised active compound 0.1mg, lactose 20mg and the powder was inhaled with the aid of an inhalation device.

Example 32: powder formulations for inhalation

The spheronized powder was loaded into a multi-dose powder inhaler containing 0.1mg of micronized active compound per dose.

Example 33: powder formulations for inhalation

The spheronized powder was loaded into a multi-dose powder inhaler containing 0.1mg of micronized active compound and 1mg of micronized lactose per dose.

Example 34: capsule formulation

1.0mg of active compound, 321mg of small sugar spheres, 306.6 mg of Aquacoat ECD, 0.5mg of acetyl tributyl citrate, 800.1 mg of Tween-800, 100-5517.5 mg of Eudragit L, 1.8mg of triethyl citrate, 8.8mg of talcum powder and 0.lmg of defoamer MMSg.

Example 35: capsule seedling formula

2.0mg of active compound, 305mg of small sugar spheres, Aquocoat ECD 305.0 mg, acetyl tributyl citrate 0.4mg, Tween-800.14 mg, Eudragit NE 30D 12.6mg, Eudragit S10012.6 mg, talc 0.l6 mg.

Example 36: enema formula

2mg of active compound, 25mg of sodium carboxymethylcellulose, 0.5mg of disodium ethylenediaminetetraacetate, 0.8mg of methylparaben, 0.2mg of propylparaben, 7mg of sodium chloride, 1.8mg of citric acid, 800.01 mg of tween-tween and 1mL of pure water.

Example 37: formulations containing liposomes

A. Preparation of the instillation formulation

Dipalmitoyl lecithin (45mg), dimyristoyl lecithin (7mg), dipalmitoyl phosphatidyl glycerol (1mg) and active compound (5mg) were placed in a glass tube, all components were dissolved in chloroform and N was used₂Adding an aqueous solution (0.9% NaCl) to the lipids, forming liposomes at a temperature above the phase inversion temperature of the lipids, the resulting suspension containing liposomes ranging in size from very small vesicles to 2 μm.

B. Preparation of formulations for inhalation

Liposomes were prepared as in example A, with an aqueous solution containing 10% lactose at a 7:3 lactose to lipid ratio. The liposome suspension was frozen with dry ice and freeze-dried, and the dried product was micronized to give particles with a mass-mean aerodynamic diameter (MMAD) of about 2 μm.

While the invention has been described with reference to a preferred embodiment, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof without departing from the scope of the invention. Any simple modification, equivalent change and modification of the above embodiments according to the technical essence of the present invention, without departing from the technical solution of the present invention, still belong to the protection scope of the technical solution of the present invention.

Claims

1. A 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidines of formula I, their prodrugs and pharmaceutically active metabolites, and pharmaceutically acceptable salts thereof, wherein the formula I has the structure:

R₁independently selected from hydrogen, halogen;

R₂independently selected from hydrogen, C1-C4 alkyl, benzyl;

R₃、R₄independently selected from hydrogen, C1-C4 alkyl, C3-C6 cycloalkyl, 2-dimethylaminoethyl, 3-morpholinopropyl, C1-C3 alkyl-substituted phenyl, halogen-substituted C1-C3 alkyl-substituted phenyl, or R₃、R₄Together with the nitrogen atom to which they are attached form a morpholinyl group.

2. The 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidines, prodrugs and pharmaceutically active metabolites thereof, and pharmaceutically acceptable salts thereof, of claim 1 wherein:

R₁independently selected from hydrogen, fluorine;

R₂independently selected from hydrogen, methyl, benzyl;

3. The 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidines of claim 2, prodrugs and pharmaceutically active metabolites thereof, and pharmaceutically acceptable salts thereof, wherein the compounds are any one of the following compounds:

6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholinopropyl) amino ] pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

6-benzyl-5- { [2- (dimethylamino) ethyl ] amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

6-benzyl-1, 3-bis (4-fluorophenyl) -8-methyl-5- (methylamino) pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

5- { [ 3-chloro-4- (trifluoromethyl) phenyl ] amino } -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

1, 3-bis (4-fluorophenyl) -8-methyl-5- { [3- (trifluoromethyl) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethyl) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

5- [ (3-chloro-4-fluorophenyl) amino ] -1, 3-bis (4-fluorophenyl) -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

1, 3-bis (4-fluorophenyl) -8-methyl-5- { [4- (trifluoromethoxy) phenyl ] amino } pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

1, 3-bis (4-fluorophenyl) -5- [ (furan-2-ylmethyl) amino ] -8-methylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

1, 3-bis (4-fluorophenyl) -8-methyl-5- [ (3-morpholinopropyl) amino ] pyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

6-benzyl-8-methyl-5- [ (3-morpholinopropyl) amino ] -1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

6-benzyl-5- (cyclopropylamino) -8-methyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

6-benzyl-8-methyl-5-morpholinyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

6-benzyl-5- { [2- (dimethylamino) ethyl ] amino } -8-methyl-1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione;

6, 8-dimethyl-5- [ (3-morpholinopropyl) amino ] -1, 3-diphenylpyrido [2,3-d ] pyrimidine-2, 4,7(1H,3H,8H) -trione.

4. A pharmaceutical composition comprising as an active ingredient a compound of any one of the 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidines of claims 1 to 3, prodrugs and pharmaceutically active metabolites thereof, and pharmaceutically acceptable salts thereof and a pharmaceutically acceptable carrier or diluent.

5. The pharmaceutical composition according to claim 4, wherein the pharmaceutical composition is a pharmaceutical composition for inhibiting mitogen-activated protein kinase.

6. A process for preparing 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidines, prodrugs and pharmaceutically active metabolites thereof, and pharmaceutically acceptable salts thereof as claimed in claim 3, wherein said 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidines are prepared by the following route:

7. use of a 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidine as claimed in any of claims 1 to 3, a prodrug and a pharmaceutically active metabolite thereof, and a pharmaceutically acceptable salt thereof or a pharmaceutical composition as claimed in any of claims 4 to 5 for the preparation of a mitogen-activated protein kinase MAPK, a mitogen-activated protein kinase MEK, an extracellular regulated protein kinase ERK, a Raf protein kinase inhibitor.

8. Use of a 5-substituted amino-1, 3-disubstituted phenylpyrido [2,3-d ] pyrimidine according to any one of claims 1 to 3, a prodrug and a pharmaceutically active metabolite thereof, and a pharmaceutically acceptable salt thereof or a pharmaceutical composition according to any one of claims 4 to 5 for the manufacture of a medicament for the treatment of cancer.

9. The use of claim 8, wherein the cancer is lung cancer, liver cancer, melanoma, colon cancer, rectal cancer, breast cancer, ovarian cancer, cervical cancer, or renal cancer.