CN111840299A

CN111840299A - Application of quinoline compound in preparation of cancer treatment drug

Info

Publication number: CN111840299A
Application number: CN202010342195.3A
Authority: CN
Inventors: 徐华
Original assignee: Wuhan Shengyun Biomedical Technology Co Ltd
Current assignee: Wuhan Shengyun Biomedical Technology Co Ltd
Priority date: 2019-04-30
Filing date: 2020-04-27
Publication date: 2020-10-30
Also published as: CN111848580B; CN111848580A

Abstract

The invention provides an application of a quinoline compound in preparing a medicament for treating cancers, wherein the quinoline compound takes an EGFR signal channel and/or a VEGFR signal channel as a target spot, inhibits the activity of tyrosine kinase EGFR and/or VEGFR1 to promote cancer cell apoptosis, and has a very good inhibition effect on cancers; the invention can act on lung cancer, liver cancer, stomach cancer, colon cancer, breast cancer and the like, and has wide application range; the quinoline compound provided by the invention is used as an EGFR or/and VEGFR signal channel target drug in the treatment of cancer, and can be prepared into various dosage forms, so that the quinoline compound is convenient to use.

Description

Application of quinoline compound in preparation of cancer treatment drug

Technical Field

The invention relates to the technical field of cancer drugs, in particular to application of a quinoline compound in preparing a drug for treating cancer.

Background

Egfr (epidermal Growth factor receptor) is a receptor for cell proliferation and signaling of the Epithelial Growth Factor (EGF) and belongs to one of the ErbB receptor family. EGFR is also known as HER1, ErbB1, and mutations or overexpression thereof induce tumors. EGFR is a tyrosine kinase type receptor with cell membrane penetration. EGFR dimerization can activate its intracellular kinase pathways, and this autophosphorylation can direct downstream phosphorylation, including MPAK, Akt, and JNK pathways, to induce cell proliferation. There is high or abnormal expression of EGFR in many solid tumors, such as pancreatic cancer, renal cancer, lung cancer, prostate cancer, breast cancer, etc. EGFR is involved in the inhibition of tumor cell proliferation, angiogenesis, tumor invasion, metastasis and apoptosis. It may be made with: high expression of EGFR leads to enhancement of downstream signaling; increased expression of mutant EGFR receptors or ligands results in sustained activation of EGFR; the effect of the autocrine loop is enhanced; disruption of receptor down-regulation mechanisms; activation of abnormal signaling pathways, etc. The search for drugs capable of inhibiting the kinase activity of EGFR may be an effective way to inhibit tumorigenesis development and metastasis.

Neuroinvasion, which is one of the characteristics of malignant tumors and is responsible for severe pain, metastatic spread and poor prognosis in tumor patients, originates in the secretion of GDNF and the activation of its RET tyrosine kinase receptor. In the process of tumor angiogenesis, numerous angiogenesis factors such as VEGF, FGF and PDGF are involved, wherein VEGF is the most important regulatory factor in the process of tumor angiogenesis. VEGF as angiogenesis promoting factor can increase permeability of blood vessel, stimulate proliferation and migration of endothelial cells, promote angiogenesis of tumor, and participate in growth, invasion and metastasis of tumor. VEGF has been found to have enhanced expression in a variety of human tumors, including breast cancer, colorectal cancer, non-small cell lung cancer, ovarian cancer, etc., and its enhanced expression is directly related to the number of new blood vessels in tumor tissues, so the search for drugs that inhibit VEGFR activity is also one of the potential cancer therapies. The inventor's earlier application, US8993566B2, discloses a process for the preparation of quinolines comprising formula 1 and pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof. In vitro activity screening shows that the compounds can inhibit the activity of C-Met kinase, thereby having anti-tumor activity on partial cancers; subtle differences in the structure of the compounds can lead to changes in the magnitude of the anti-cancer activity and/or the corresponding optimal target.

Disclosure of Invention

In view of the above, the present invention provides an application of a quinoline compound in the preparation of a medicament for treating cancer on the basis of a previous study, wherein the quinoline compound has a structure shown in formula i, and inhibits the activity of EGFR kinase and/or VEGFR1 kinase by using an EGFR signal pathway as a target and/or a VEGFR signal pathway as a target, so as to inhibit cancers mediated by EGFR kinase and/or VEGFR1 kinase.

Further, the quinoline compounds inhibit phosphorylation of receptor tyrosine kinases EGFR or VEGFR 1.

Further, the cancer is any one of lung cancer, liver cancer, stomach cancer, colon cancer, breast cancer and pancreatic cancer.

Furthermore, the medicine comprises a quinoline compound as a main active ingredient, or is prepared by taking the quinoline compound as an ingredient.

Furthermore, the medicament is prepared into inclusion compound, liposome, microspheres, nanoparticles or emulsion by quinoline compound and pharmaceutically acceptable carrier.

Furthermore, the formulation of the quinoline compound comprises any one of injection, tablet, granule, pill, capsule, suspension and emulsion.

Compared with the prior art, the invention has the following advantages:

(1) The invention takes EGFR and/or VEGFR signal channel as drug target spot, promotes cancer cell apoptosis by inhibiting the activity of EGFR and/or VEGFR1, and has very good inhibition effect on cancer;

(2) the quinoline compound provided by the invention is used as an EGFR and VEGFR signal channel target spot drug for treating cancers, can be prepared into various dosage forms, is convenient to use, has an inhibiting effect on lung cancer, liver cancer, stomach cancer, colon cancer and breast cancer, and has a wide application range.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate an embodiment of the invention and, together with the description, serve to explain the invention and not to limit the invention. In the drawings:

FIG. 1 shows the EGFR kinase inhibitory activity of Cabozantinib, a compound of formula I;

FIG. 2 shows the inhibitory activity of Cabozantinib, a compound of formula I, on VEGFR1 kinase;

FIG. 3 is a graph showing the effect of intragastric administration of a compound of formula I on the mean body weight of rats.

It should be noted that the data in the following experimental examples are obtained by a lot of experiments, which are limited by the text, and only a part of the data is shown in the specification, and those skilled in the art can understand and implement the present invention under the data, and the rest of the data have the same trend and can draw the same conclusion as the experimental conclusion described below, and will not be described in detail later.

The technical effect of the invention is verified by the following experimental research:

EXPERIMENTAL EXAMPLE 1 inhibition of human EGFR, VEGFR1, MET kinase by Compounds of formula I

1.1 inhibition of human EGFR kinase by Compounds of formula I

1. Experimental Material

1.1 reagents

(1)EGFR Kinase Enzyme System、EGFR,Active(0.1μg/μL)、Poly(Glu4,Tyr1)(1mg/ml)、5×Reaction BufferA、DTT(0.1M)、MnCl₂Solution(2.5M)；

(2)ADP-Glo^TMKinase Assay Kit、Ultra-Pure ATP(10mM)、ADP(10mM)、ADP-Glo^TMReagent、Kinase Detection Buffer、Kinase Detection Substrate；

The above reagents were purchased from Promega corporation, USA.

1.2 drugs (1) Cabozantinib (5mM) (2) test drugs-1 (5mM) or more were provided by Hem-Han GmbH.

1.3 consumable cell culture plate (96 wells), gun head: corning, USA.

1.4 Main instruments (1) HERAMELL VIOS 160i cell culture box: thermo Scientific, usa; (2)1300Series A2 superclean bench: thermo Scientific, usa; (3)5424R Low temperature centrifuge: eppendorf, Germany; (4) the microporous plate multifunctional microplate reader: berthold, Germany; (5)4 ℃, 20 ℃ and 80 ℃ in a refrigerator: haier corporation;

1.5 preparation of the drug

Formula I: shaking with DMSO solution to dissolve, making into 5mM drug stock solution, storing at 4 deg.C, and using within two weeks; cabozantinib: the mixture was dissolved in DMSO with shaking to prepare a 5mM drug stock solution, which was stored at 4 ℃ for two weeks.

2. Experimental methods

(1)ADP-Glo^TMDissolving the Reagent at room temperature, and mixing the Kinase Detection Buffer and the Kinase Detection Substrate to obtain the Kinase Detection Reagent; (2) EGFR Kinase enzyme system, ATP dissolved on ice; (3) 400 μ L of 5 × Reaction buffer A, 1 μ L of LDTT, 1.6 μ L of MnCl ₂And 597.4 μ L dH2O to prepare 1ml 2 XBuffer; (4) will be provided with25 μ LATP solution (10mM), 500 μ L2 XBuffer and 475 μ L dH₂O mixing to prepare 250 mu MATPASSAY Solution; (5) EGFR (0.1. mu.g/. mu.L) was diluted with 1 XBuffer to 1 ng/. mu.L, carboplatin (5mM) was diluted with 1 XBuffer to 5, 12.5, 25, 50, 100, 200. mu.M, formula I (5mM) was diluted with 1 XBuffer to 5, 12.5, 25, 50, 100. mu.M; (6) mu.L of diluted EGFR (1 ng/. mu.L), 5. mu.L of LPoly (Glu4, Tyr1) (1mg/ml) and 5. mu.L of 2 XBuffer were added to each well of a 96-well plate, 5. mu.L of Cabozantinib (5, 12.5, 25, 50, 100, 200. mu.M) at different concentrations was added to one group, and 5. mu.L of formula I (5, 12.5, 25, 50, 100. mu.M) at different concentrations was added to the other group. An additional 2 control wells were set, one of which was not EGFR but was 5. mu.L of 1 XBuffer; the other well is not added with medicine but added with 5 mu LDMSO; (7) mu.L of 250. mu.M ATPASSAY Solution was added to each well to make the total reaction volume 25. mu.L. Mixing, and incubating at room temperature for 30-60 min; (8) add 25. mu. LADP-Glo to each well^TMReagent, mixing, and incubating at room temperature for 40 min; (9) adding 50 mu LkingaseDeprotection Reagent into each hole, uniformly mixing, and incubating at room temperature for 30-60 min; (10) measuring and recording the fluorescence intensity of each well; (11) calculating an inhibition rate ═ 1- (fluorescence intensity with inhibitor added-blank fluorescence intensity without enzyme)/(fluorescence intensity without inhibitor-blank fluorescence intensity without enzyme)) × 100; (12) the logarithmic value of the inhibitor concentration and the inhibition ratio were inputted to Graphpadprism 6, and a relationship between the two was plotted, and IC50 was calculated.

3. The results are shown in FIG. 1, where IC50 ═ 4.537. mu.M, Cabozantinib is a positive control, and IC is shown as follows₅₀The kinase activity of EGFR can be obviously inhibited by the formula I which is 3.764 mu M.

1.2 inhibition of human VEGFR1 kinase by formula I

1. Experimental Material

1.1 reagents

(1)VEGFR Kinase Enzyme System、VEGFR,Active(0.05μg/μL)、IGF1Rtide(1mg/ml)5×Reaction BufferA、DTT(0.1M)、MnCl₂Solution(2.5M)

(2)ADP-Glo^TMKinase Assay Kit、Ultra-Pure ATP(10mM)、ADP(10mM)、ADP-Glo^TMReagents above Reagent kit, Detection Buffer kit, Detection Substrate were purchased from Promega corporation, USA.

1.2 drugs

(1) Cabozantinib (5 mM); (2) the test drug, a drug of formula I (5mM) or more, was provided by Wuhan Hengyuan.

1.3 consumable cell culture plate (96 wells), gun head: corning Inc. of USA

1.5 formulation of the drug formula I: shaking with DMSO solution to dissolve, making into 5mM drug stock solution, storing at 4 deg.C, and using within two weeks; cabozantinib: the mixture was dissolved in DMSO with shaking to prepare a 5mM drug stock solution, which was stored at 4 ℃ for two weeks.

2. The experimental procedure was the same as 2.1 for the inhibition of human EGFR kinase.

3. The results of the experiment are shown in FIG. 2, in which IC50 is 2.833. mu.M, Cabozantinib as positive control, and IC is shown in the figure₅₀As 2.349 μ M, formula i significantly inhibited the kinase activity of VEGFR 1.

1.3 inhibition of human MET kinase by Compounds of formula I

1. Experimental Material

1.1 reagents

MET Kinase Enzyme System、MET,Active(0.1μg/μl)、Poly(Glu4,Tyr1)(1mg/ml)5×Reaction BufferA、DTT(0.1M)；(2)ADP-Glo^TMKinaseAssay Kit、Ultra-PureATP(10mM)、ADP(10mM)、ADP-Glo^TMReagent above Reagent was purchased from Promega corporation, USA.

1.2 drugs

(1) Cabozantinib (5mM), (2) test drug compound of formula i (5mM) the above drug was supplied by wuhan constant source.

1.3 consumable cell culture plate (96 wells), gun head: corning Inc. of USA

1.4 Main Instrument

(1) Heraccell vios 160i cell incubator: thermo Scientific, usa; (2)1300Series A2 superclean bench: thermo Scientific, usa; (3)5424R Low temperature centrifuge: eppendorf, Germany; (4) the microporous plate multifunctional microplate reader: berthold, Germany; (5)4 ℃, 20 ℃ and 80 ℃ in a refrigerator: haier corporation;

1.5 preparation of the drug

A compound of formula I: shaking with DMSO solution to dissolve, making into 5mM drug stock solution, storing at 4 deg.C, and using within two weeks; cabozantinib: the mixture was dissolved in DMSO with shaking to prepare a 5mM drug stock solution, which was stored at 4 ℃ for two weeks.

2. Experimental methods

(1)ADP-Glo^TMDissolving the Reagent at room temperature, and mixing the Kinase Detection Buffer and the Kinase Detection Substrate to obtain the Kinase Detection Reagent; (2) MET Kinase enzymeSystemm, ATP dissolved on ice; (3) 1ml of 2 XBuffer was prepared by mixing 400. mu.l of 5 × Reaction Buffer A, 1. mu.l of DTT and 599. mu.l of dH 2O; (4) mu.l ATP solution (10mM), 500. mu.l 2 XBuffer and 495. mu.l dH₂O mixing to prepare 50 mu MATPASSAY Solution; (5) MET (0.1. mu.g/. mu.l) diluted 1 XBuffer to 2 ng/. mu.l, carboplatin (5mM) diluted 1 XBuffer to 5, 10, 20, 30, 40, 80. mu.M, compound of formula I diluted 1 XBuffer to 5, 12.5, 25, 50, 100. mu.M; (6) mu.l of diluted MET (2 ng/. mu.l), 5. mu.l of Poly (Glu4, Tyr1) (1mg/ml) and 8. mu.l of 2 Xbuffer were added to each well of a 96-well plate, 5. mu.l of Cabozantinib (5, 10, 20, 30, 40, 80. mu.M) was added to one group and 5. mu.l of the compound of formula I (5, 12.5, 25, 50, 100. mu.M) was added to the other group; another 2 control wells were set, one well being filled with 5. mu.l 1 XBuffer without MET; the other well was dosed with 5. mu.l DMSO without drug; (7) adding 5 μ l of 250 μ M ATPASSAY Solution into each well to make the total reaction system to be 25 μ l, mixing, and incubating at room temperature for 30-60 min; (8) add 25. mu.l ADP-GloTM Reagent into each well, mix well, incubate 40min at room temperature; (9) adding 50 mul of Kinase Deprotecting reagent into each well, mixing uniformly, and incubating for 30-60min at room temperature; (10) measuring and recording the fluorescence intensity of each well; (11) the inhibition rate (1- (Fluorosa angustifolia plus inhibitor) was calculated Light intensity-enzyme-free blank fluorescence intensity)/(inhibitor-free fluorescence intensity-enzyme-free blank fluorescence intensity)) × 100; (12) the logarithmic value of the inhibitor concentration and the inhibition ratio were inputted to Graphpadprism 6, and a relationship between the two was plotted, and IC50 was calculated.

3. Results of the experiment

TABLE 1 inhibition data of Cabozantinib, Compounds of formula I on MET kinase

As shown in Table 1, the results of the kinase assay show that the compounds of formula I have no inhibitory activity against MET kinase.

EXAMPLE 2 in vitro pharmacodynamic assay of Compounds of formula I (in vitro anti-tumor Effect on cancer cells)

Culturing cancer cells to logarithmic phase, adjusting the concentration of cell suspension, adding 100 mu L of cell suspension into each hole of a 96-hole plate, and enabling the cell density to be 1000-10000 holes; at 5% CO₂Culturing in a cell culture box at 37 ℃ until cells adhere to the wall, and adding compounds of formula I with different concentration gradients; continuously culturing for different time, and observing cell morphology under an inverted microscope; adding 20 mu of LMTT (5mg/ml) into each well, and culturing for 4 h; absorbing culture solution in the holes, adding 150 mu LDMSO into each hole, and shaking to fully dissolve crystals; measuring the light absorption value of each hole at 490nm by using an enzyme-labeling instrument; each experiment was repeated at least twice.

TABLE 2 IC50 values (. mu.g/ml) of compounds of formula I on different cancer cells for 48h

	First experiment	Second experiment
			Human epidermal carcinoma A431 cells	5.32	7.73
Human lung cancer A549 cell	45.19	44.46
			Human lung carcinoma H460 cell	9.84	11.76
Human liver cancer HepG2 cell	15.54	12.95
			Human cervical carcinoma Hela cell	42.78	41.46
SW620 cell of human colon cancer	0.483	0.506
			Human ovarian cancer ES-2 cells	14.04	13.20
Human breast cancer MCF-7 cells	0.49	0.819
			Human gastric cancer BGC823 cell	16.68	19.17
Human gastric cancer MKN45 cell	8.68	13.20
			SW1990 cell of human pancreatic cancer	40.17	35.00
Human rectal cancer Caco2 cell	9.23	12.60

As shown in Table 2, the compound of the formula I has inhibition effect on various cancer cells, wherein the effect on human epidermal cancer A431 cells, human breast cancer MCF-7 cells, human gastric cancer MKN45 cells and human colon cancer SW620 cells is optimal; in addition, the growth inhibition of the compounds of formula I on cancer cells is significantly dependent on concentration, with the rate of growth inhibition increasing with increasing concentration.

Detailed Description

It should be noted that the embodiments and features of the embodiments may be combined with each other without conflict.

EXAMPLE 1 preparation of the Compound of formula I

Step A-1- [4- (3-chloropropyloxy) -3-methoxy ] acetophenone (II)

3-methoxy-4-hydroxyacetophenone (600g, 3.61mol), anhydrous potassium carbonate (698g, 5.055mol) was added to DMF (5v/w, 2500mL), stirred well for 30min at 25 deg.C, then 1, 3-bromochloropropane (795.9g, 1.4mol) was slowly added dropwise, after the addition was complete, the reaction was stirred at 25 deg.C for 10 h. After the reaction is finished, performing suction filtration, washing a filter cake with a small amount of DMF, collecting filtrate, slowly pouring the filtrate into ice water, stirring vigorously to precipitate white solid, performing suction filtration, and drying the filter cake to obtain 827.2g of white solid with the yield of 93.8%.

Step B-1- [4- (3-chloropropyloxy) -5-methoxy-2-nitro ] acetophenone (III)

Intermediate II (200g, 0.82Mol) was added to CH₂Cl₂(5v/w, 1000mL) and fully stirring to completely dissolve the intermediate II, then cooling the reaction solution to-20 ℃, slowly dropwise adding fuming nitric acid (130g, 2.06mol), keeping the temperature of the reaction solution not to exceed-10 ℃, and reacting for 2 hours at-20 ℃ after dropwise adding. After the reaction, the reaction solution was poured into ice water, the organic layer was collected, and the organic layer was washed with saturated brine until the aqueous layer was neutral, the organic layer was collected, dried with anhydrous sodium sulfate, and then the solvent was evaporated to dryness to obtain a reddish brown oil, which was sufficiently cooled to obtain 210g of a yellow solid with a yield of 89%.

Step C- (E) -1- [4- (3-chloropropoxy) -5-methoxy-2-nitrophenyl ] -3- (dimethylamino) propyl-2-en-1-one (IV)

Intermediate III (200g, 0.695mol) was added to toluene (5v/w, 1000mL), heated to 110 ℃ to completely dissolve intermediate III, DMF-DMA (414.2g, 3.476mol) was added, and the reaction was refluxed for 16 hours. After the reaction is finished, the reaction solution is cooled to room temperature and then is put into a cold trap to be stirred, yellow solid is separated out, suction filtration is carried out, and 180g of yellow solid is obtained after a filter cake is dried, and the yield is 75.8%.

Step D-7- (3-chloropropyloxy) -6-methoxy-4 (1H) -quinolinone (V)

The intermediate IV (150g, 0.44mol) is added into glacial acetic acid (8v/w, 1200mL), the temperature is raised to 40 ℃, after the intermediate IV is completely dissolved, iron powder (123.1g, 2.20mol) is added in portions slowly, the temperature is raised to 80 ℃, and the reaction is mechanically stirred for 2 hours. After the reaction is finished, the reaction solution is filtered while the reaction solution is hot, filtrate is collected, a large amount of solid is separated out after the filtrate is cooled, and the solid is filtered to obtain a solid in the shape of earthy yellow. Dissolving the filter cake in glacial acetic acid, stirring at 80 ℃ for about 30min, performing suction filtration while the solution is hot, collecting filtrate, cooling the filtrate to separate out solid, performing suction filtration to obtain khaki solid, combining the solid, washing the solid with water for multiple times until the filtrate is neutral, and drying the filter cake to obtain 79g of light yellow solid with the yield of 65%.

Step E-6-methoxy-7- [3- (1-morpholinyl) propoxy ] -4(1H) -quinolinone (VI)

Intermediate V (62g, 0.232mol), morpholine (100.92g, 1.16mol) was added to acetonitrile (620mL) and heated at reflux for 8 h. After the reaction, a part of the solvent was evaporated, the reaction flask was placed in a cold trap, filtered, and the filter cake was washed with ethyl acetate to obtain 66.74g of a yellow solid with a yield of 95.3%.

Step F-4-chloro-6-methoxy-7- [3- (1-morpholinyl) propoxy ] quinoline (VII)

Intermediate VI (63g, 0.198mol), POCl₃(5v/w, 315mL) was added to acetonitrile (5v/w, 315mL) and the reaction was refluxed at 85 ℃ for 6 h. After the reaction, the reaction solution was cooled and POCl was evaporated to dryness₃And acetonitrile to give a grey viscous solid, which is added to a large volume of ice-water mixture and adjusted to pH 10 with 10% KOH solution. By CH₂Cl₂Extraction (200mL x 3), collection of organic layer, anhydrous sodium sulfate drying, evaporation of solvent, cooling to obtain an off-white solid 58g, yield 87.3%.

Step G-4- (2-fluoro-4-nitrophenoxy) -6-methoxy-7- [3- (1-morpholinyl) propoxy ] quinoline (VIII)

2-fluoro-4-nitrophenol (36.73g, 0.234mol) was added to dry chlorobenzene (5v/w, 250mL), heated to 145 deg.C, intermediate VII (12.5g, 0.039mol) was added to the reaction and reacted at this temperature for 20 hours. After the reaction, the solvent was evaporated to dryness to obtain a grey solid, which was dissolved in CH₂Cl₂In, with saturated K₂CO₃The solution was washed several times, the organic layer was collected and the solvent was evaporated to dryness to give a solid which was recrystallized from ethanol to give 49.16g of a yellow solid with a yield of 71.4%.

Step H-3-fluoro-4- [ 6-methoxy-7- [3- (1-morpholinyl) propoxy ] quinolin-4-yloxy ] aniline (A-1)

Iron powder (61.42g, 1.1mol) and 6mL of hydrochloric acid were added to 90% EtOH (25v/w, 1210mL), the temperature was raised to 80 ℃ and stirred for 15min, then intermediate VIII (48.42g, 0.11mol) was added to the reaction mixture in portions, and after the addition, the reaction was refluxed for 2 h. After the reaction, the reaction mixture was filtered while hot, the filtrate was collected, and the solvent was evaporated to dryness to obtain 43g of a yellow solid with a yield of 95%.

Step I-Ethyl 2- (cyanoacetamido) carboxylate (a)

2-cyanoacetic acid (85g, 1mol) and ethyl carbamate (89g, 1mol) were added to 500mL of toluene, and after slowly adding phosphorus oxychloride (45mL, 0.5mol), 5mL of DMF was added and reacted at 70 ℃ for 2 h. Cooling, adding 500mL of water into the reaction solution to quench the phosphorus oxychloride, performing suction filtration, washing a filter cake with diethyl ether, and drying to obtain 104g of a white solid with the yield of 67%.

Step J- [ 2-cyano-2- (2-phenylhydrazone group) acetylamino ] carboxylic acid ethyl ester (b)

Aniline (10.3g, 0.11mol), hydrochloric acid (45mL, 0.56mol), was added to 400mL of water and stirred. Cooling to 0 deg.C, controlling the temperature to 0-5 deg.C, and dripping NaNO₂(7.73g, 0.11mol) and reacting at 0-5 ℃ for 30min for later use after dripping. Adding the intermediate a and sodium acetate (80g, 0.98mol) into ethanol (400mL), stirring, cooling to 0 ℃, controlling the temperature to be between 0 and 5 ℃, dropwise adding the prepared solution into the solution of the intermediate a, and stirring and reacting for 2 hours at 0 to 5 ℃ after dropwise adding. Suction filtration, filter cake washing with water, drying, orange solid 21.5g, yield 91.1%.

Step K-2-phenyl-3, 5-dioxo-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carbonitrile (c)

Intermediate b (21g, 0.08mol), anhydrous sodium acetate (29g, 0.35mol) were added to glacial acetic acid (20v/w, 420mL) and the temperature was raised to reflux for 2 h. After the reaction is finished, cooling to 0 ℃, pouring the reaction solution into 1200mL of water, stirring for half an hour, carrying out suction filtration, washing a filter cake with water and petroleum ether, and drying to obtain 12g of orange-red solid with the yield of 67.8%.

Step L-2-phenyl-4-methyl-3, 5-dioxo-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carbonitrile (d)

Intermediate c (9g, 0.042mol) was added to DMF (100mL), NaH (1.3g, 0.054mol) was added in portions under ice bath, stirred for half an hour and then methyl iodide (2.74mL, 0.042mol) was added dropwise. After dropping, the reaction was carried out for 2 hours. After the reaction is completed, the reaction solution is poured into 600mL of water, a large amount of red solid is separated out, the suction filtration is carried out, a filter cake is washed by water, and the orange-red solid 7.6g is obtained after drying, wherein the yield is 80.0%.

Step M-2-phenyl-4-methyl-3, 5-dioxo-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carboxylic acid (e)

Intermediate d (7.6g, 0.033mol) was added to a mixed solvent of glacial acetic acid (100mL), HCl (72mL, 0.86mol), and the temperature was raised to 120 ℃ for reflux reaction for 4 h. The solvent was evaporated to dryness, 400mL of water was added to the fraction, filtered and dried to give 3.7g of a solid with a yield of 45.7%.

Step N-2-phenyl-4-methyl-3, 5-dioxo-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carbonyl chloride (C-1)

Intermediate e (3g, 0.012mol) was added to thionyl chloride (10V/W, 30mL), and the mixture was stirred and warmed to reflux for 6 hours. The solvent was evaporated to dryness to give 2.8g of yellow transparent crystals, yield 87.5%.

Step O-N- { 3-fluoro-4- { 6-methoxy-7- [3- (4-morpholinyl) propoxy ] quinolin-4-yloxy } phenyl } -2-phenyl-4-methyl-3, 5-dioxo-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carboxamide (formula I)

Intermediate A-1(3g, 0.007mol) was dissolved in DCM (20v/m,60mL), DIPEA (1.4g,0.01mol) was added and the temperature was reduced to 0 ℃ under ice bath. C-1(2.8g,0.01mol) was dissolved in DCM (8v/m,22mL) and added dropwise to the reaction mixture, the temperature was controlled not to exceed 0 ℃ and the reaction was carried out at 0 ℃ for 30min after completion of the dropwise addition. After the reaction, the reaction solution was washed with 5% NaOH solution, water, saturated brine, evaporated to dryness, and anhydrous ethanol (5v/m) was added to the residue, and the mixture was heated to reflux, stirred for 1 hour, filtered with suction while hot, and washed with anhydrous ethanol to obtain a pale yellow solid (3.8 g), with a yield of 83.6%.

EXAMPLE 2 formulation of quinolines of formula I

The medicine comprises a compound shown in the formula I as a main active ingredient, or a medicine prepared by taking the compound shown in the formula I as an ingredient. The medicament is prepared into inclusion compound, liposome, microspheres, nanoparticles or emulsion by quinoline compound and pharmaceutically acceptable carrier. The dosage form of the compound comprises any one of injection, tablets, granules, pills, capsules, suspending agents and emulsion.

And (3) tablet preparation: 10g of the compound of the formula I in example 2 is mixed with 20g of auxiliary materials according to a general pharmaceutical tabletting method and then pressed into 100 tablets, each tablet weighs 300 mg.

And (3) capsule preparation: 10g of the compound containing the formula I in example 2 is mixed with 20g of auxiliary materials according to the requirements of pharmaceutical capsules, and then the mixture is filled into empty capsules, wherein each capsule weighs 300 mg.

Although embodiments of the present invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made in the embodiments without departing from the spirit and scope of the invention.

Claims

1. The application of the quinoline compound in preparing the medicine for treating cancer is characterized in that the structure of the quinoline compound is shown as formula I, namely

The quinoline compound takes an EGFR signal path and/or a VEGFR signal path as a target spot and inhibits the activity of EGFR kinase and/or VEGFR1 kinase.

2. The use of claim 1, wherein the cancer is any one of lung cancer, liver cancer, stomach cancer, colon cancer, breast cancer, pancreatic cancer.

3. The use of claim 1, wherein the medicament comprises a quinoline compound as a main active ingredient, or a medicament prepared from a quinoline compound as an ingredient.

4. The use of claim 3, wherein the medicament is a clathrate, liposome, microsphere, nanoparticle, or emulsion of the quinoline compound and a pharmaceutically acceptable carrier.

5. The use of claim 3, wherein the quinoline compound is in the form of injection, tablet, granule, pill, capsule, suspension, or emulsion.