Heterocyclic Substituted Acardite Derivate and Application Thereof

Abstract

This present invention discloses a heterocyclic substituted acardite derivate and application thereof, namely compounds in the general formula (1) or the general formula (2) or pharmaceutically acceptable salts thereof, wherein A is monosubstituted or polysubstituted quinoline, isoquinoline, quinazoline, pyrrole or pyrimidine, and the substituent is halogen, C1-5alkyl, C1-5haloalkyl, C1-5alkoxy, C1-5haloalkoxy, C1-5alkylamino, C1-5haloalkylamino, amino or nitryl; R1 is C1-5alkyl; R2 is one or more selected from hydrogen, halogen, alkyl, alkoxy, haloalkyl or haloalkoxy; and R3 is one or more selected from hydrogen, halogen, alkyl, alkoxy, haloalkyl or haloalkoxy. The compound of the present invention and the pharmaceutically acceptable salt thereof can be used for treating tumor or leukemia.

Description

FIELD OF THE INVENTION

This present invention relates to an aromatic heterocyclic substituted acardite derivate and application thereof In addition, the present invention relates to application of aromatic heterocyclic substituted acardite derivate and pharmaceutically acceptable salts thereof in the treatment of tumor or leukemia.

BACKGROUND OF THE INVENTION

With better understanding of the tumor molecular mechanisms, the research on the targeted therapy of the tumor moleculars has achieved important advance. Protein kinase inhibitor is one of newly developed targeted therapy drugs, which affects the survival, proliferation and disease progression of tumor cells through blocking the intra-cellular molecular transduction pathway. Raf kinases play a crucial role in the signal transduction pathway of Raf/MEK/ERK. Although the function of the Raf kinase in normal tissues is not yet understood, but the existing basic and clinical research results have shown that the upregulation of Raf gene and overexpression of its protein are present in various solid tumors, including renal cell carcinoma, hepatocellular carcinoma, melanoma and non-small cell lung cancer. Currently, more and more single target point and multi-target point therapy drugs for Raf kinases are successfully developed and applied clinically, for example, sorafenib and erlotinib have achieved good clinical results, and the anti-tumor therapy has came into the “molecular targeted therapy” era. CN200810129360.6 disclosed that a kind of aromatic heterocyclic substituted acardite derivates with no substituent or only carbamyl in the A ring have prospect of inhibiting specific tumors, and the preliminary pharmacological experiments found that the effects of some compouns are better than sorafenib.

SUMMARY OF THE INVENTION

The objective of the present invention is to provide an aromatic heterocyclic substituted acardite derivate having more medicinal value through structural modification based on the existing technology. After the present invention adds specific substituents in the A ring, especially adding substituents in the quinazoline, pyrrole or pyrimidine rings, the inhibitory activity and selectivity of the compounds to specific tumors are greatly increased, and the absorptivity and utilization rate of the compounds are increased and the toxic side effects are reduced. The objective of the present invention is further to provide application of the compound or pharmaceutically acceptable salts thereof in the treatment of tumor or leukemia. The heterocyclic substituted acardite derivate of the present invention can be represented by the following formulas [1] and [2]:

• • wherein,

A is monosubstituted or polysubstituted quinoline, isoquinoline, quinazoline, pyrrole or pyrimidine, preferably monosubstituted or polysubstituted quinazoline, pyrrole or pyrimidine, further preferably monosubstituted or polysubstituted quinazoline; the substituent is halogen, alkyl, haloalkyl, alkoxy, haloalkoxy, alkylamino, haloalkylamino, amino or nitryl, preferably halogen, C_1-5 alkyl, C_1-5 haloalkyl, C_1-5 alkoxy, C_1-5 haloalkoxy, C_1-5 alkylamino, C_1-5haloalkylamino, amino or nitryl, more preferably halogen, C_1-5alkyl, C_1-5haloalkyl, C_1-5alkoxy, C_1-5haloalkoxy, amino or nitryl; still more preferably halogen, amino, C_1-5alkyl or C_1-3alkoxy, particularly preferably Cl, Br, F, amino, methoxy, methyl, ethyl, propyl, isopropyl, butyl or t-butyl in the present invention.

R₁ is alkyl, more preferably C_1-5alkyl, most preferably methyl, ethyl, propyl and isopropyl.

R₂ is one or more selected from hydrogen, halogen, alkyl, alkoxy, haloalkyl or haloalkoxy; preferably one or more selected from hydrogen, halogen, C_1-5alkyl, C_1-5alkoxy, C_1-5haloalkyl or C_1-5haloalkoxy, most preferably one or more selected from H, Cl, Br, F, methoxy, ethoxy, propoxy, methyl, ethyl, propyl, isopropyl, butyl, t-butyl or trifluoromethyl.

R₃ is one or more selected from hydrogen, halogen, alkyl, alkoxy, C_1-5 haloalkyl or C_1-5haloalkoxy, preferably one or more selected from hydrogen, halogen, C_1-5alkyl, C_1-5alkoxy, C_1-5haloalkyl or C_1-5haloalkoxy, most preferably one or more selected from H, Cl, Br, F, methoxy, ethoxy, propoxy, methyl, ethyl, propyl, isopropyl, butyl, t-butyl or trifluoromethyl.

The pharmaceutically acceptable salts of the compound in the present invention are selected from:

• a) basic salts of inorganic acids and organic acids, the described acid is selected from hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, mesylate, trifluoromethanesulfonic acid, benzene sulfonic acid, paratoluenesulfonic acid, 1-naphthalene sulfonic acid, 2-naphthalene sulfonic acid, acetic acid, trifluoroacetic acid, malic acid, tartaric acid, citric acid, lactic acid, oxalic acid, succinic acid, fumaric acid, maleic acid, benzoic acid, salicylic acid, phenylacetic acid or almonds acid;
• b) acid salts of organic and inorganic base, the described cation is selected from alkali metal cation, alkaline earth metal cation, ammonium cation, aliphatic-substituted ammonium cation or aromatic-substituted ammonium cation.

Preparation of the Compound of Formula 1

Method 1: the target compound is obtained from substituted heterocyclic 2-carboxylate as starting materials through acyl chlorination, aminoalkylation, two-step condensation and salt forming reaction and the route is as follows:

Method 2: the target compound is obtained from substituted heterocyclic 2-carboxylate as starting materials through acyl chlorination, aminoalkylation, condensation and salt forming reaction and the route is as follows:

Preparation of the Compound of Formula 2

The target compound is obtained from halogen substituted heterocyclic as starting materials through two-step condensation and salt forming reaction and the route is as follows:

The substituents A, R₁, R₂ and R₃ in the above menthioned reaction routes have the above described meanings.

The beneficial effects of the present invention are as follows:

The derivatives of the present invention have raf kinase inhibitory activity. The action mechanism of this compound is that this compound affects the survival, proliferation and disease progression of tumor cells through inhibiting raf kinase and blocking the ras protein signal transduction connection, thereby inhibiting the growth of achiblastomas, such as malignant tumors (for example, bladder cancer, lung cancer, pancreatic cancer), myelopathy (for example, myelogenous leukemia) or adenoma (for example, villous adenoma of colon).

The experiment results have shown that the compound with special substituents added in A ring in the present invention has stronger antitumor activity compared with the previously disclosed compound with no substituent or only carbamyl in A ring, which is obviously stronger than Sorafenib in the effects of tumor cell metastasis and tumor angiogenesis. The test on normal human umbilical vein endothelial cells found that this part of the compounds have lower toxicity to normal human cells, such as endothelial cell, which are safe and reliable, but which can inhibit the tumor angiogenesis to achieve anti-tumor activity. In vivo nude mice transplanted model experiment proved that the compound of the present invention has inhibitory effects to human liver and kidney cancer and the effects are stronger than Sorafenib, which has more obvious effects on lung cancer and the effects are significantly better than the positive control drug Sorafenib. The results show that the compound of the present invention or pharmaceutically acceptable salts thereof can be used in the drugs for the treatment of cancer or leukemia, particularly drugs used for treating lung cancer.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The melting point was measured by the electric melting point instrument and the thermometer was not corrected; the elemental analyzer was Foss-Heraeus type; and the mass spectrograph was electrospray ionization mass spectrometry.

A: preparing the aromatic heterocyclic substituted acardite derivate having the general formula 1 accroding to method 1

Embodiment 1

Preparation of 4-Chloro-6-Methoxyquinolinyl-2-Carbonyl Chloride

50g of 4-hydroxy-6-methoxy-2-quinolinecarboxylic acid and 100 m1 of thionyl chloride were added into a three-necked flask, heated and refluxed for 17 hours until the reaction finished. The filtrate was added with toluene and concentrated under vacuum to obtain yellow solid, namely 4-chloro-6-methoxyquinolinyl-2-carbonyl chloride, with dry weight of 50g.

Embodiment 2

Preparation of 4-Chloro-7-Fluoroquinazolinyl-2-Carbonyl Chloride

Prepared from 4-hydroxy-7-fluoro-2-quinazolinecarboxylic acid with reference to the method of embodiment 1.

Embodiment 3

Preparation of 4-Methoxy-5-Chloropyrimidine-2-Carbonyl Chloride

Prepared from 4-methoxy-5-hydroxy-2-pyrimidinecarboxylic acid with reference to the method of embodiment 1.

Embodiment 4

Preparation of 4-Chloro-7-Amino-Isoquinolyl-2-Carbonyl Chloride

Prepared from 4-hydroxy-7-amino-2-quinazolinecarboxylic acid with reference to the method of embodiment 1.

Embodiment 5

Preparation of 5-Methyl-4-Chloropyrrolyl-2-Carbonyl Chloride

Prepared from 5-methyl-4-hydroxy-2-pyrrolecarboxylic acid with reference to the method of embodiment 1.

Embodiment 6

Preparation of 4-Chloro-6-Methoxyl-N-methyl-2-Quinolinyl Formamide

10 g of 4-chloro-6-methoxyquinolinyl-2-carbonyl chloride (obtained from embodiment 1) was reacted with 200 ml of 2M methylamine ethanol solution under 0° C. for 36 hours until the reaction finished. The solvent was evaporated under vacuum and the residues were added with water followed by stirring evenly. Ethyl acetate was added for extracting and the ethyl acetate layer was dried with anhydrous sodium sulfate. The ethyl acetate layer was removed under vacuum to obtain 9 g of 4-chloro-6-methoxy-N-methyl-2-quinoline carboxamide.

Embodiment 7

Preparation of 4-Chloro-7-Fluoro-N-Methyl-2-Quinazoline Methanamide

Prepared from 4-chloro-7-fluoroquinazolinyl-2-carbonyl chloride with reference to the method of embodiment 6.

Embodiment 8

Preparation of 4-Methoxyl-5-Chloro-N-Methyl-2-Pyrimidinecarboxamide

Prepared from 4-methoxyl-5-chloropyrimidinyl-2-carbonyl chloride with reference to the method of Embodiment 6.

Embodiment 9

Preparation of 4-Chloro-7-Amino-N-Methyl-2-Isoquinolinecarboxamide

Prepared from 4-chloro-7-aminoisoquinolyl-2-carbonyl chloride with reference to the method of embodiment 6.

Embodiment 10

Preparation of 5-Methyl-4-Chloro-N-Methyl-2-Pyrrole Carboxamide

Prepared from 5-methyl-4-chloropyrryl-2-carbonyl chloride with reference to the method of embodiment 6.

Embodiment 11

Preparation of

4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl) oxy))aniline 10 g of 4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide (obtained from embodiment 6) was dissolved in DMF, added with 20 g of potassium tert-butylate and 10 g of 4-aminophenol and kept at 70° C. under the protection of nitrogen for 8 hours. After the reaction finished, the reaction solution was poured into 250 ml of ethyl acetate and 250 ml of saturated saline solution and stirred evenly for separation. The water solution was extracted with ethyl acetate again. The ethyl acetate layer was washed with saturated saline solution and dried with anhydrous sodium sulfate. The solvent was evaporated and removed under vacuum to obtain 6 g of 4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))aniline.

Embodiment 12

Preparation of 4-(2-(N-Methylaminoformoxyl)-4-(7Fluoroquinazolinyl) Oxy)Aniline

Prepared from 4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide with reference to the method of embodiment 11.

Embodiment 13

Preparation of

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl) oxy)aniline Prepared from 4-methoxyl-5-chloro-N-methyl-2-pyrimidinecarboxamide with reference to the method of embodiment 11.

Embodiment 14

Preparation of 4-(2-(N-Methylaminoformoxyl)-4-(7-Amino-Isoquinolyl) Oxy)Aniline

Prepared from 4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide with reference to the method of embodiment 11.

Embodiment 15

Preparation of 4-(2-(N-Methylaminoformoxyl)-4-(5-Methyl-Pyrryl)Oxy) Aniline

Prepared from 5-methyl-4-chloro-N-methyl-2-pyrrole carboxamide with reference to the method of embodiment 11.

Embodiment 16

Synthesis of 4-Chloro-3-(Trifluoromethyl)Phenyl Isocyanate

20 g of 4-chloro-3-(trifluoromethyl)aniline was mixed with 100 ml benzene, added with 20g of diphosgene and refluxed for 12 hours. The reaction solution was added with toluene, and the solvent was evaporated and removed under vacuum to obtain the product 4-chloro-3-(trifluoromethyl)phenyl isocyanate.

Embodiment 17

Synthesis of 4-Bromo-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-bromo-3-(trifluoromethyl)aniline with reference to the method of embodiment 16.

Embodiment 18

Synthesis of 4-Fluoro-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-fluoro-3-(trifluoromethyl)aniline with reference to the method of embodiment 16.

Embodiment 19

Synthesis of Compound 1

7 g of 4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))aniline (obtained from embodiment 11), 5 g of 4-chloro-3-(trifluoromethyl)phenyl isocyanate (obtained from embodiment 16) and 50 ml of methylene dichloride were stirred at room temperature for 24 hours, and the crystals were separated out followed by air pump filtration and collection to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea.

Embodiment 20

Synthesis of Compound 2

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl) oxy))anilineaniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 21

Synthesis of Compound 3

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquin olinyl)oxy))phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 22

Synthesis of Compound 4

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazol inyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)aniline (obtained from embodiment 12) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 23

Synthesis of Compound 5

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazoli nyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 24

Synthesis of Compound 6

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazol inyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 25

Synthesis of Compound 7

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyri midinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)aniline (obtained from embodiment 13) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 26

Synthesis of Compound 8

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrim idinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 27

Synthesis of Compound 9

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyri midinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 28

Synthesis of Compound 10

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquin olyl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)aniline (obtained from embodiment 14) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 29

Synthesis of Compound 11

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquin olyl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-isoquinolyl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 30

Synthesis of Compound 12

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoqui nolyl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-isoquinolyl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 31

Synthesis of Compound 13

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl) oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)aniline (obtained from embodiment 15) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 32

Synthesis of Compound 14

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl) oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 33

Synthesis of Compound 15

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl) oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 19.

Embodiment 34

Synthesis of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea mesylate

10 g of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea free base was dissolved in 300 ml of ether and added with methanesulfonic acid/ethanol solution in drops at room temperature until pH=2, and white crystal was precipitated followed by air pump filtration and collection to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea mesylate.

Embodiment 35

Synthesis of Pharmaceutically Acceptable Salts of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquin olinyl)oxy))phenyl)urea

With reference to the method of embodiment 34, fluoromethanesulfonic acid/ethanol solution, benzene sulfonic acid/ethanol solution, p-toluenesulfonic acid/ethanol solution,

1-naphthalenesulfonic acid/ethanol solution, 2-naphthalenesulfonic acid/ethanol solution, acetic acid/ethanol solution, trifluoroacetic acid/ethanol solution, malic acid/ethanol solution, tartaric acid/ethanol solution, citric acid/ethanol solution, lactic acid/ethanol solution, oxalic acid/ethanol solution, succinic acid/ethanol solution, fumaric acid/ethanol solution, maleic acid/ethanol solution, benzoic acid/ethanol solution, salicylic acid/ethanol solution, phenylacetic acid/ethanol solution or mandelic acid/ethanol solution were added in drops to synthesize trifluoromethylsulfonate, benzene sulfonate, tosilate, 1-naphthalenesulfenesulfonate, 2-naphthalenesulfenesulfonate, acetate, trifluoroactate, malate, tartrate, citrate, lactate, oxalate, succinate, fumarate, maleate, benzoate, salicylate, phenylacetate or mandelate of N-(4-chloro-3 -(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea.

The pharmaceutically acceptable salts of compounds 2-15 can be also synthesized according to the above mentioned method.

B: preparing the aromatic heterocyclic substituted acardite derivate having the general formula 1 accroding to method 2

Embodiment 36

Synthesis of

N-(4-chloro-3 -(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea

20 g of 4-chloro-3-(trifluoromethyl)phenyl isocyanate (obtained from embodiment 16), 15 g of 4-aminophenol and 500 ml of dichloromethane were stirred at room temperature for 2 h, and the crystal was precipitated, followed by air pump filtration and collection, washing with dichloromethane and vacuum drying to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea.

Embodiment 37

Synthesis of

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea

Prepared from 4-bromo-3-(trifluoromethyl)phenyl isocyanate (obtained from embodiment 17) with reference to embodiment 36.

Embodiment 38

Synthesis of

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea

Prepared from 4-fluoro-3-(trifluoromethyl)phenyl isocyanate (obtained from embodiment 18) with reference to embodiment 36.

Embodiment 39

Synthesis of

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea (compound 1)

10 g of N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea (obtained from embodiment 36), 8.2 g of 4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide (obtained from embodiment 6) and 50 ml dichloromethane were stirred at room temperature for 24 h, and the crystal was precipitated, followed by air pump filtration and collection to obtain 12 g of N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea.

Embodiment 40

Synthesis of Compound 2

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquino linyl)oxy))phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide according to the method of Embodiment 39.

Embodiment 41

Synthesis of Compound 3

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquin olinyl)oxy))phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide according to the method of Embodiment 39.

Embodiment 42

Synthesis of Compound 4

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazol inyl)oxy) phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide (obtained from embodiment 7) according to the method of Embodiment 39.

Embodiment 43

Synthesis of Compound 5

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazoli nyl)oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide according to the method of Embodiment 39.

Embodiment 44

Synthesis of Compound 6

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazol inyl)oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide according to the method of Embodiment 39.

Embodiment 45

Synthesis of Compound 7

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyri midinyl)oxy) phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-methoxyl-5-chloro-N-methyl-2-pyrimidinecarboxamide (obtained from embodiment 8) according to the method of Embodiment 39.

Embodiment 46

Synthesis of Compound 8

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrim idinyl)oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and 4-methoxyl-5-chloro-N-methyl-2-pyrimidinecarboxamide according to the method of Embodiment 39.

Embodiment 47

Synthesis of Compound 9

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyri midinyl)oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-N-methyl-2-pyrimidinecarboxamide according to the method of Embodiment 39.

Embodiment 48

Synthesis of Compound 10

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquin olyl)oxy)phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide (obtained from embodiment 9) according to the method of Embodiment 39.

Embodiment 49

Synthesis of Compound 11

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquin olyl)oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide according to the method of Embodiment 39.

Embodiment 50

Synthesis of Compound 12

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoqui nolyl)oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide according to the method of Embodiment 39.

Embodiment 51

Synthesis of Compound 13

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(2-methyl-pyrryl) oxy)phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

2-methyl-4-chloro-N-methyl-2-pyrrole carboxamide (prepared from embodiment 10) according to the method of Embodiment 39.

Embodiment 52

Synthesis of Compound 14

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(2-methyl-pyrryl) oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

2-methyl-4-chloro-N-methyl-2-pyrrole carboxamide according to the method of Embodiment 39.

Embodiment 53

Synthesis of Compound 15

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(2-methyl-pyrryl) oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

2-methyl-4-chloro-N-methyl-2-pyrrole carboxamide according to the method of Embodiment 39.

C: preparing the aromatic heterocyclic substituted acardite derivate having the general formula 2 accroding to method 3

Embodiment 54

Preparation of 4-(4-(6-Methoxyquinolinyl)Oxy))Aniline

8 g of 4-chloro-6-methoxyquinoline was dissolved in DMF, added with 20 g of potassium tert-butylate and 10 g of 4-aminophenol and reacted under the protection of nitrogen at 70° C. for 8 hours. After the end of the reaction, the reaction liquid was poured into 250 ml of ethyl acetate and 250 ml of saturated salt water and mixed evenly followed by liquid separation. The water solution was extracted with ethyl acetate.

The ethyl acetate layer was added with saturated salt water for washing and dried by anhydrous sodium sulfate. The solvent was evaporated under vacuum to obtain 6 g of 4-(4-(6-methoxyquinolinyl)oxy))aniline.

Embodiment 55

Preparation of 4-(4-(7Fluoroquinazolinyl)Oxy)Aniline

Prepared from 4-chloroquinazoline with reference to the method of embodiment 54.

Embodiment 56

Preparation of 4-(5-(4-Methoxypyrimidinyl)Oxy)Aniline

Prepared from 5-chloro-4-methoxypyrimidine with reference to the method of embodiment 54.

Embodiment 57

Preparation of 4-(4-(7-Amino-Isoquinolyl)Oxy)Aniline

Prepared from 4-chloro-7-aminoisoquinoline with reference to the method of embodiment 54.

Embodiment 58

Preparation of 4-(4-(2-Methyl-Pyrryl)Oxy)Aniline

Prepared from 4-chloro-2-methylpyrrol with reference to the method of embodiment 54.

Embodiment: 59

Preparation of 4-(4-(6-Methoxyl-7-Fluoro-Quinolinyl)Oxy)Aniline

Prepared from 4-chloro-6-methoxyl-7-fluoro-quinoline with reference to the method of embodiment 54.

Embodiment: 60

Preparation of 4-(4-(6-Methyl-7-Fluoro-Quinolinyl)Oxy))Aniline

Prepared from 4-chloro-6-methyl-7-fluoro-quinazoline with reference to the method of embodiment 54.

Embodiment 61

Synthesis of 4-Chloro-3-(Trifluoromethyl)Phenyl Isocyanate

100 ml of diphosgene is mixed with 20 g of 4-chloro-3-(trifluoromethyl)aniline and refluxed for 12 hours. The reaction liquid was added into toluene, and the solvent was evaporated under vacuum to obtain the product 4-chloro-3-(trifluoromethyl)phenyl isocyanate.

Embodiment 62

Synthesis of 4-Bromo-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-bromo-3-(trifluoromethyl)aniline with reference to the method of embodiment 61.

Embodiment 63

Synthesis of 4-Fluoro-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-fluoro-3-(trifluoromethyl)aniline with reference to the method of embodiment 61.

Embodiment 64

Synthesis of 4-Chloro-3-Ethylphenyl Isocyanate

Prepared from 4-chloro-3-ethylaniline with reference to the method of embodiment 61.

Embodiment 65

Synthesis of 4-Ethyl-3-Trifluoromethyl

Prepared from 4-ethyl3-trifluoromethylaniline with reference to the method of embodiment 61.

Embodiment 66

Synthesis of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-quinolinyl)oxyphenyl)urea (compound 16) 7 g of 4-(4-(6-methoxyquinolinyl)oxy))aniline4-(4-quinolinyl)oxyaniline (prepared from embodiment 54), 5 g of 4-chloro-3-(trifluoromethyl)phenyl isocyanate (prepared from embodiment 61) and 50 ml of methylene dichloride were mixed and reacted at room temperature for 24 hours, and the crystal was precipitated, followed by air pump filtration and collection to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea.

Embodiment 67

Synthesis of Compound 17

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea was prepared from 4-(4-(6-methoxyquinolinyl)oxy))aniline and 4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method of Embodiment 66.

Embodiment 68

Synthesis of Compound 18

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea was prepared from 4-(4-(6-methoxyquinolinyl)oxy))aniline and 4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 69

Synthesis of Compound 19

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7fluoroquinazolinyl)oxy)phenyl)urea was prepared from 4-(4-(7fluoroquinazolinyl)oxy)aniline (prepared from embodiment 55) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 70

Synthesis of Compound 20

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7fluoroquinazolinyl)oxy)phenyl)urea was prepared from 4-(4-(7fluoroquinazolinyl)oxy)aniline and 4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 71

Synthesis of Compound 21

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7fluoroquinazolinyl)oxy)phenyl)urea was prepared from 4-(4-(7fluoroquinazolinyl)oxy)aniline and 4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 72

Synthesis of Compound 22

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)urea was prepared from 4-(5-(4-methoxypyrimidinyl)oxy)aniline (prepared from embodiment 56) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 73

Synthesis of Compound 23

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)urea was prepared from 4-(5-(4-methoxypyrimidinyl)oxy)aniline and 4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 74

Synthesis of Compound 24

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)urea was prepared from 4-(5-(4-methoxypyrimidinyl)oxy)aniline and 4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 75

Synthesis of Compound 25

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from 4-(4-(7-amino-isoquinolyl)oxy)aniline (prepared from embodiment 57) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 76

Synthesis of Compound 26

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from 4-(4-(7-amino-isoquinolyl)oxy)aniline and 4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 77

Synthesis of Compound 27

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from 4-(4-(7-amino-isoquinolyl)oxy)aniline and 4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 78

Synthesis of Compound 28

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy)phenyl)urea was prepared from 4-(4-(2-methyl-pyrryl)oxy)aniline (prepared from embodiment 58) and 4-chloro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 79

Synthesis of Compound 29

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy)phenyl)urea was prepared from 4-(4-(5-methyl-pyrryl)oxy)aniline and 4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 80

Synthesis of Compound 30

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy)phenyl)urea was prepared from 4-(4-(5-methyl-pyrryl)oxy)aniline and 4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd of Embodiment 66.

Embodiment 81

Synthesis of Compound 31

N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea was prepared from 4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)aniline (prepared from embodiment 59) and 4-chloro-3-ethylphenyl isocyanate (prepared from embodiment 64) according to the methd of Embodiment 66.

Embodiment 82

Synthesis of Compound 32

N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea was prepared from 4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)aniline (prepared from embodiment 59) and 4-ethyl-3-trifluoromethyl isocyanate (prepared from embodiment 65) according to the methd of Embodiment 66.

Embodiment 83

Synthesis of Compound 33

N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))phenyl)urea was prepared from 4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))aniline (prepared from embodiment 60) and 4-chloro-3-ethylphenyl isocyanate (prepared from embodiment 64) according to the methd of Embodiment 66.

Embodiment 84

Synthesis of Compound 34

N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))phenyl)urea was prepared from 4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))aniline (prepared from embodiment 60) and 4-ethyl-3-trifluoromethyl isocyanate (prepared from embodiment 65) according to the methd of Embodiment 66.

Embodiment 85

Preparation of 4-Chloro-7-Nitrylquinoline-2-Carbonyl Chloride

Prepared from 4-hydroxyl-7-nitryl-2-quinoline carboxylic acid with reference to the method of embodiment 1.

Embodiment 86

Preparation of 4-Chloro-7-Trifluoromethylquinazoline-2-Carbonyl Chloride

Prepared from 4-hydroxyl-7-trifluoromethyl-2-quinazoline carboxylic acid with reference to the method of embodiment 1.

Embodiment 87

Preparation of 4-Chloro-7-Nitryl-N-Ethyl-2-Quinoline Carboxamide

Prepared from 4-chloro-7-nitrylquinoline-2-carbonyl chloride (prepared from embodiment 58) and 2M ethylamine ethanol solution with reference to the method of embodiment 6.

Embodiment 88

Preparation of 4-Chloro-7-Trifluoromethyl-N-Propyl-2-Quinazoline Methanamide

Prepared from 4-chloro-7-trifluoromethylquinazoline-2-carbonyl chloride (prepared from embodiment 86) and 2M propylamine ethanol solution with reference to the method of embodiment 6.

Embodiment 89

Preparation of

2-methyl-4-(2-(N-ethylcarbamyl)-4-(7-nitrylquinolinyl)oxy))aniline

Prepared from 4-chloro-7-nitryl-N-ethyl-2-quinoline carboxamide (prepared from embodiment 87) and 3-methyl-4-aminophenol with reference to the method of embodiment 11.

Embodiment 90

Preparation of 2-Methoxyl-4-(2-(N-Ethylcarbamyl)-4-(7-Nitrylquinolinyl)Oxy))Aniline

Prepared from 4-chloro-7-nitryl-N-ethyl-2-quinoline carboxamide (prepared from embodiment 87) and 3-methoxyl-4-aminophenol with reference to the method of embodiment 11.

Embodiment 91

Preparation of 2-Fluoro-4-(2-(N-Propylcarbamyl)-4-(7-Trifluoromethylquinolinyl)Oxy)) Aniline

Prepared from 4-chloro-7-trifluoromethyl-N-propyl-2-quinazoline methanamide (prepared from embodiment 88) and 3-fluoro-4-aminophenol with reference to the method of embodiment 11.

Embodiment 92

Preparation of 2-Trifluoromethyl-4-(2-(N-Propylcarbamyl)-4-(7-Trifluoromethylquinolinyl)Oxy))Aniline

Prepared from 4-chloro-7-trifluoromethyl-N-propyl-2-quinazoline methanamide (prepared from embodiment 88) and 4-amino-3-trifluoromethyl phenol with reference to the method of embodiment 11.

Embodiment 93

Synthesis of 4-Chloro-3-Methoxyphenyl Isocyanate

Prepared from 4-chloro-3-methoxyaniline with reference to the method of embodiment 61.

Embodiment 94

Preparation of Compound 35

N-(4-chloro-3-methoxyphenyl)-N′-(2-methyl-4-(2-(N-ethylcarbamyl)-4-(7-nitryl-quinolinyl)ox y)phenyl)urea was sythesized from 2-methyl-4-(2-(N-ethylcarbamyl)-4-(7-nitrylquinolinyl)oxy))aniline (prepared from embodiment 89) and 4-chloro-3-methoxyphenyl isocyanate (prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 95

Preparation of Compound 36

N-(4-chloro-3-methoxyphenyl)-N′-(2-methoxyl-4-(2-(N-ethylcarbamyl)-4-(7-nitryl-quinolinyl) oxy)phenyl)urea was sythesized from 2-methoxyl-4-(2-(N-ethylcarbamyl)-4-(7-nitrylquinolinyl)oxy))aniline (prepared from embodiment 90) and 4-chloro-3-methoxyphenyl isocyanate (prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 96

Preparation of Compound 37

N-(4-chloro-3-methoxyphenyl)-N′-(2-fluoro-4-(2-(N-propylcarbamyl)-4-(7-trifluoromethylqui nolinyl)oxy))phenyl)urea was sythesized from 2-fluoro-4-(2-(N-propylcarbamyl)-4-(7-trifluoromethylquinolinyl)oxy))aniline (prepared from embodiment 91) and 4-chloro-3-methoxyphenyl isocyanate (prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 97

Preparation of Compound 38

N-(4-chloro-3-methoxyphenyl)-N′-(2-trifluoromethyl-4-(2-(N-propylcarbamyl)-4-(7-trifluorom ethylquinolinyl)oxy))phenyl)urea was sythesized from

2-trifluoromethyl-4-(2-(N-propylcarbamyl)-4-(7-trifluoromethylquinolinyl)oxy))aniline (prepared from embodiment 92) and 4-chloro-3-methoxyphenyl isocyanate (prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 98

Synthesis of N-(4-Chloro-3-(Trifluoromethyl)Phenyl)-N′-(4-(4-(6-Methoxyquinolinyl)Oxy))Phenyl)Urea Mesylate

10 g of N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea free base was dissolved in 300 ml of ether and added with methanesulfonic acid/ethanol solution in drops at room temperature until pH=2, and white crystal was precipitated followed by air pump filtration and collection to obtain N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea mesylate.

Embodiment 99

Synthesis of Pharmaceutically Acceptable Salts of N-(4-Chloro-3-(Trifluoromethyl)Phenyl)-N′-(4-(4-(6-Methoxyquinolinyl)Oxy))Phenyl)Urea

With reference to the method of embodiment 85, fluoromethanesulfonic acid/ethanol solution, benzene sulfonic acid/ethanol solution, p-toluenesulfonic acid/ethanol solution, 1-naphthalenesulfonic acid/ethanol solution, 2-naphthalenesulfonic acid/ethanol solution, acetic acid/ethanol solution, trifluoroacetic acid/ethanol solution, malic acid/ethanol solution, tartaric acid/ethanol solution, citric acid/ethanol solution, lactic acid/ethanol solution, oxalic acid/ethanol solution, succinic acid/ethanol solution, fumaric acid/ethanol solution, maleic acid/ethanol solution, benzoic acid/ethanol solution, salicylic acid/ethanol solution, phenylacetic acid/ethanol solution or mandelic acid/ethanol solution were added in drops to synthesize trifluoromethylsulfonate, benzene sulfonate, tosilate, 1-naphthalenesulfenesulfonate, 2-naphthalenesulfenesulfonate, acetate, trifluoroactate, malate, tartrate, citrate, lactate, oxalate, succinate, fumarate, maleate, benzoate, salicylate, phenylacetate or mandelate of N-(4-chloro-3 -(trifluoromethyl)phenyl)-N′-(4-(4-quinolinyl)oxyphenyl)urea.

The pharmaceutically acceptable salts of compounds 17-38 can be also synthesized according to the above mentioned method.

The compounds in table 1 to 14 were prepared according to methods of the above mentioned embodiments, and the characteristics are shown in the following tables.

TABLE 1
substituted quinoline derivatives
Compound		Elementary		Mass spectrum
No.	R3	analysis	Name	M + 1
1	4-chloro-3-	C: 57.3	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	545.5
	trifluoromethyl	H: 3.8	(4-(2-(N-methylaminoformoxyl)-4-(6-
		N: 10.3	methoxyquinolinyl)oxy))phenyl)urea
2	4-fluoro-3-	C: 59.2	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	529
	trifluoromethyl	H: 3.9	(4-(2-(N-methylaminoformoxyl)-4-(6-
		N: 10.5	methoxyquinolinyl)oxy))phenyl)urea
3	4-bromo-3-	C: 53.0	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	590
	trifluoromethyl	H: 3.31	(4-(2-(N-methylaminoformoxyl)-4-(6-
		N: 9.38	methoxyquinolinyl)oxy))phenyl)urea

TABLE 2
substituted quinazoline derivatives
Compound		Elementary		Mass spectrum
No.	R3	analysis	Name	M + 1
4	4-chloro-3-	C: 54.1	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	534.5
	trifluoromethyl	H: 3.11	(4-(2-(N-methylaminoformoxyl)-4-
		N: 13.0	(7 fluoroquinazolinyl)oxy)phenyl)urea
5	4-fluoro-3-	C: 55.5	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	518
	trifluoromethyl	H: 3.30	(4-(2-(N-methylaminoformoxyl)-4-
		N: 13.4	(7 fluoroquinazolinyl)oxy)phenyl)urea
6	4-bromo-3-	C: 49.7	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	579
	trifluoromethyl	H: 2.91	(4-(2-(N-methylaminoformoxyl)-4-
		N: 12.2	(7 fluoroquinazolinyl)oxy)phenyl)urea

TABLE 3
substituted pyrimidine derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
7	4-chloro-3-	C: 50.9	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	496.5
	trifluoromethyl	H: 3.48	(4-(2-(N-methylaminoformoxyl)-5-(4-
		N: 14.0	methoxypyrimidinyl)oxy)phenyl)urea
8	4-fluoro-3-	C: 52.5	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	480
	trifluoromethyl	H: 3.67	(4-(2-(N-methylaminoformoxyl)-5-(4-
		N: 14.5	methoxypyrimidinyl)oxy)phenyl)urea
9	4-bromo-3-	C: 46.8	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	541
	trifluoromethyl	H: 3.00	(4-(2-(N-methylaminoformoxyl)-5-(4-
		N: 13.1	methoxypyrimidinyl)oxy)phenyl)urea

TABLE 4
substituted isoquinoline derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
10	4-chloro-3-	C: 56.5	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	530.5
	trifluoromethyl	H: 3.70	(4-(2-(N-methylaminoformoxyl)-
		N: 13.2	4-(7-amino-isoquinolyl)oxy)phenyl)urea
11	4-fluoro-3-	C: 58.5	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	514
	trifluoromethyl	H: 3.81	(4-(2-(N-methylaminoformoxyl)-
		N: 13.8	4-(7-amino-isoquinolyl)oxy)phenyl)urea
12	4-bromo-3-	C: 52.4	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	574
	trifluoromethyl	H: 3.44	(4-(2-(N-methylaminoformoxyl)-
		N: 12.4	4-(7-amino-isoquinolyl)oxy)phenyl)urea

TABLE 5
substituted pyrrole derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
13	4-chloro-3-	C: 54.1	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	467.5
	trifluoromethyl	H: 4.01	(4-(2-(N-methylaminoformoxyl)-4-(5-
		N: 12.2	methyl-pyrryl)oxy)phenyl)urea
14	4-fluoro-3-	C: 55.8	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	451
	trifluoromethyl	H: 4.02	(4-(2-(N-methylaminoformoxyl)-4-(5-
		N: 12.6	methyl-pyrryl)oxy)phenyl)urea
15	4-bromo-3-	C: 49.5	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	511
	trifluoromethyl	H: 3.70	(4-(2-(N-methylaminoformoxyl)-4-(5-
		N: 11.0	methyl-pyrryl)oxy)phenyl)urea

TABLE 6
substituted quinoline derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
16	4-chloro-3-	C: 59.2	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	488.5
	trifluoromethyl	H: 3.70	(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea
		N: 8.77
17	4-fluoro-3-	C: 61.3	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	472
	trifluoromethyl	H: 3.81	(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea
		N: 8.80
18	4-bromo-3-	C: 54.0	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	532
	trifluoromethyl	H: 3.40	(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea
		N: 7.79

TABLE 7
substituted quinazoline derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
19	4-chloro-3-	C: 55.5	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	477.5
	trifluoromethyl	H: 2.91	(4-(4-(7 fluoroquinazolinyl)oxy)phenyl)urea
		N: 11.6
20	4-fluoro-3-	C: 57.7	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	461
	trifluoromethyl	H: 2.75	(4-(4-(7 fluoroquinazolinyl)oxy)phenyl)urea
		N: 12.2
21	4-bromo-3-	C: 50.5	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	522
	trifluoromethyl	H: 2.71	(4-(4-(7 fluoroquinazolinyl)oxy)phenyl)urea
		N: 10.8

TABLE 8
substituted pyrimidine derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
22	4-chloro-3-	C: 52.1	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	439.5
	trifluoromethyl	H: 3.30	(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)
		N: 12.8
23	4-fluoro-3-	C: 54.2	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	423
	trifluoromethyl	H: 3.38	(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)
		N: 13.3
24	4-bromo-3-	C: 47.2	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	484
	trifluoromethyl	H: 2.99	(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)
		N: 11.4

TABLE 9
substituted isoquinoline derivatives
Compound		Elementary		Mass
No.	R3	analysis %	Name	spectrum m/e
25	4-chloro-3-	C: 58.4	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	473.5
	trifluoromethyl	H: 3.38	(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea
		N: 11.81
26	4-fluoro-3-	C: 60.7	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	457
	trifluoromethyl	H: 3.70	(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea
		N: 13.5
27	4-bromo-3-	C: 53.5	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	518
	trifluoromethyl	H: 3.40	(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea
		N: 10.7

TABLE 10
substituted pyrrole derivatives
Compound		Elementary		Mass
No.	R3	analysis %	Name	spectrum m/e
28	4-chloro-3-	C: 55.7	N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-	410.5
	trifluoromethyl	H: 3.75	(4-(4-(2-methyl-pyrryl)oxy)phenyl)urea
		N: 10.4
29	4-fluoro-3-	C: 58.2	N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-	395
	trifluoromethyl	H: 4.01	(4-(4-(2-methyl-pyrryl)oxy)phenyl)urea
		N %: 10.5
30	4-bromo-3-	C: 50.3	N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-	455
	trifluoromethyl	H: 3.49	(4-(4-(2-methyl-pyrryl)oxy)phenyl)urea
		N: 9.41

TABLE 11
polysubstituted quinoline derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
31	4-chloro-3-	C: 64.6	N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-	466.5
	ethyl	H: 4.71	methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea
		N: 9.20
32	4-ethyl-3-	C: 62.7	N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-	501.5
	trifluoromethyl	H: 4.36	methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea
		N: 8.22

TABLE 12
polysubstituted quinazoline derivatives
Compound		Elementary		Mass
No.	R3	analysis	Name	spectrum m/e
33	4-chloro-3-	C: 64.1	N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-	451.5
	ethyl	H: 4.70	methyl-7-fluoro-quinolinyl)oxy)phenyl)urea
		N: 12.3
34	4-ethyl-3-	C: 61.8	N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-	485
	trifluoromethyl	H: 4.01	methyl-7-fluoro-quinolinyl)oxy)phenyl)urea
		N: 11.38

TABLE 13
substituted quinoline derivatives
Compound		Elementary		Mass
No.	R2	analysis	Name	spectrum m/e
35	2-methyl	C: 59.1	N-(4-chloro-3-methoxyphenyl)-N′-	550.5
		H: 4.32	(2-methyl-4-(2-(N-ethylcarbamyl)-4-
		N: 12.5	(7-nitryl-quinolinyl)oxy)phenyl)urea
36	2-methoxy	C: 57.2	N-(4-chloro-3-methoxyphenyl)-N′-	566.5
		H: 4.36	(2-methoxyl-4-(2-(N-ethylcarbamyl)-4-
		N: 12.2	(7-nitryl-quinolinyl)oxy)phenyl)urea

TABLE 14
substituted quinazoline derivatives
Compound		Elementary		Mass
No.	R2	analysis	Name	spectrum m/e
37	2-fluoro	C: 56.1	N-(4-chloro-3-methoxyphenyl)-N′-	579.5
		H: 4.17	(2-fluoro-4-(2-(N-propylcarbamyl)-4-
		N: 9.72	(7-trifluoromethylquinolinyl)oxy))phenyl)urea
38	2-trifluoro-	C: 53.4	N-(4-chloro-3-methoxyphenyl)-N′-	629.5
	methyl	H: 3.91	(2-trifluoromethyl-4-(2-(N-propylcarbamyl)-4-
		N: 8.58	(7-trifluoromethylquinolinyl)oxy))phenyl)urea

Determination of Antitumor Activity

1. Inhibitory activity of the compound of the present invention on raf kinase

[Test Method]

Raf-1 inhibitor screening by chemoluminescence method

[Instruments]

Westernblot electrophoresis apparatus Rotaryshaker

[Test Materials]

Raf-1(truncated), Magnesium/ATP Cocktail, MEK1 unactive

[Tested Samples]

Compounds 1-38

[Positive Control]

Sorafenib

$\mathrm{Inhibiton}\mathrm{rate}\%=\frac{\begin{array}{c}\mathrm{Gray}\mathrm{value}\mathrm{of}\mathrm{the}\mathrm{negative}\mathrm{control}\mathrm{group}-\\ \mathrm{gray}\mathrm{value}\mathrm{of}\mathrm{the}\mathrm{drug}\text{-}\mathrm{treated}\mathrm{group}\end{array}}{\mathrm{Gray}\mathrm{value}\mathrm{of}\mathrm{the}\mathrm{negative}\mathrm{control}\mathrm{group}}\times 100\%$

[Results]

TABLE 13
Inhibition of compounds 1-16 and positive control medicine on raf kinase
Compound	Final concentration	Inhibition		Compound	Final concentration	Inhibition
No.	1.0*10−5 mol/ml	rate %	Activity	No.	1.0*10−5 mol/ml	rate %	Activity
1	1	75.0	+	9	1	99.2	+
2	1	61.1	+	10	1	3.5
3	1	56.3	+	11	1	11.2
4	1	82.2	+	12	1	12.1
5	1	98.9	+	13	1	55.0	+
6	1	80.1	+	14	1	41.3
7	1	99.1	+	15	1	35.5
8	1	44..5		16	1	62.1	+
Positive	1	85.7	+
control
medicine

TABLE 14
Inhibition of compounds 17-38 and positive control medicine on raf kinase
Compound	Final concentration	Inhibition		Compound	Final concentration	Inhibition
No.	1.0*10−5 mol/ml	rate %	Activity	No.	1.0*10−5 mol/ml	rate %	Activity
17	1	58.2	+	28	1	33.5
18	1	34.4		29	1	85.3	+
19	1	93.5	+	30	1	16.8
20	1	87.7	+	31	1	89.4	+
21	1	98.9	+	32	1	90.5	+
22	1	88.1	+	33	1	92.3	+
23	1	89.9	+	34	1	96.2	+
24	1	91.3	+	35	1	45.3
25	1	11.5		36	1	81.2	+
26	1	15.3		37	1	81.5	+
27	1	8.8		38	1	88.1	+
Positive	1	85.7	+
control
medicine

The test results of inhibitory activity of the compound on raf kinase showed that the inhibitory activity of the compound in the present invention is better than or equivalent to positive control medicine sorafenib. The test results indicate that these compounds can affect the survival, proliferation and disease progression of tumor cells through inhibiting the raf kinase and blocking the ras protein signal transduction cascade of tumor cells. The compound of the present invention has potential of being applied to treat tumor and leukemia.

2. Experimental therapeutic action of the compound in the present invention on S180 sarcoma mice

[Test Materials]

Test animals: ICR mice, 18-25 g

Tumor types: mice S180 sarcoma, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences.

• • Positive control medicine: Sorafenib
  • Tested samples: compounds 1-38

[Test Method]

18-25 g female ICR mice and well grown 7-11 day old mice sarcoma S180 tumor seeds were selected, and the seeds were inoculated into the subcutaneous at the right axillary. After inoculated 24 hours, these mice were randomly divided into cages and orally administrated 60 mg/kg for 9 days. On 10 day, the animals were killed and weighed, and the tumor weights were weighed to calculate average tumor weight in each group, followed by calculating the tumor inhibition rate according to the following formula and T test.

$\mathrm{Tumor}\mathrm{growth}\mathrm{inhibition}\mathrm{rate}=\frac{\begin{array}{c}\mathrm{Average}\mathrm{tumor}\mathrm{weight}\mathrm{in}\mathrm{the}\mathrm{control}\mathrm{group}-\\ \mathrm{average}\mathrm{tumor}\mathrm{weight}\mathrm{in}\mathrm{the}\mathrm{treatment}\mathrm{group}\end{array}}{\mathrm{average}\mathrm{tumor}\mathrm{weight}\mathrm{in}\mathrm{the}\mathrm{treatment}\mathrm{group}}\times 100\%$

[Determination Results]

TABLE 15
Tumor growth inhibition rate of compounds 1-3 and sorafenib on mice S180 sarcoma
	Administration	Animal number	Weight (g)	Tumor weight	Inhibition
Groups	Dosage	methods	Start	Final	Start	Final	x ± SD(g)	rate (%)	P value
Normal	0.4	ml/mouse	ig	20	20	18.9 ± 1.5	22.0 ± 3.4	1.61 ± 0.36
saline
Sora	60	mg/kg	ig	10	10	18.8 ± 1.2	21.7 ± 2.4	0.71 ± 0.30	55.9	<0.05
Compound 1	60	mg/kg	ig	10	10	18.7 ± 1.9	22.3 ± 1.3	0.99 ± 0.20	38.5	<0.05
Compound 2	60	mg/kg	ig	10	10	18.9 ± 1.7	20.9 ± 2.3	0.87 ± 0.24	46.0	<0.05
Compound 3	60	mg/kg	ig	10	10	18.0 ± 1.1	20.2 ± 2.5	0.75 ± 0.36	53.4	<0.05

TABLE 16
Tumor growth inhibition rate of compounds 1-12 and sorafenib on mice S180 sarcoma (%)
	1	2	3	4	5	6	7	8	9	10	11	12	Sorafenib
Mice	38.5	24.1	53.4	50.1	49.3	51.2	21.5	58.2	55.9	55.2	54.2	45.7	55.9%
S180
sarcoma

TABLE 17
Tumor growth inhibition rate of compounds 13-26 and sorafenib on mice S180 sarcoma (%)
	13	14	15	16	17	18	19	20	21	22	23	24	25	26
Mice	21.2	18.2	56.7	33.6	44.2	35.7	50.8	54.6	59.7	52.1	51.5	54.6	55.8	7.6
S180
sarcoma

TABLE 18
Tumor growth inhibition rate of compounds 27-38
and sorafenib on mice S180 sarcoma (%)
	27	28	29	30	31	32	33	34	35	36	37	38
Mice	55.1	11.5	14.2	55.2	50.3	49.8	55.3	52.6	55.8	55.1	24.1	59.2
S180
sarcoma

3. Experimental therapeutic action of the compound in the present invention on Human colon cancer HT-29 transplantable tumor in nude mice.

[Test Materials]

• • Test animals: Female BALB/cA nude mice, 35-40 day old, with weight of 18-22 g.
  • Tumor seeds: Human colon cancer HT-29 transplantable tumor in nude mice, established by inoculating human colon cancer HT-29 cell strains subcutaneously in nude mice
  • Positive control medicine: Sorafenib
  • Tested samples: Compounds 1-38

[Test Method]

Take eugenic tumor tissues and cut into about 1.5 mm³, and then incoculate subcutaneously at the right armpit of nude mice under the sterile conditions. The diameter of the transplantable tumor in nude mice was determined with a vernier caliper, and the animals were divided into groups after the tumors were grown to 100-300 mm³. Using the method of measuring the tumor diameter, dynamically observe the antitumor effects of tested materials. The diameter of the tumor was determined three times every week and the mouse weight was weighed at the same time. The mice were intragastrically administrated with Sorafenib and tested drugs, 60 mg/kg, for continuous 9 times. The solvent was intragastrically administrated as the control for continuous 9 times. Equal amount of control was administrated in the negative control group. Tumor volume (TV) is calculated as: TV=1/2×a×b², wherein a and b respectively represent length and width.

Relative tumor volume (RTV) is calculated as: RTV=TV_t/TV₀, wherein TV₀ is the tumor volume when administrated according to different cages and TV_t is the tumor volume measured each time.

Relative tumor reproduction rate T/C (%) is calculated as follows:

$T\text{/}C\left(\%\right)=\frac{T_{\mathrm{RTV}}}{C_{\mathrm{RTV}}}\times 100$

T_RTV: RTV in the treatment group; C_RTV: RTV in the negative control group.

The test results used relative tumor reproduction rate T/C (%) as evaluating indicator of anti-tumor activity.

Evaluation of in vivo anti-tumor activity

T/C % Evaluation
≧60   (−) No activity
60-50 (+/−) Marginal activity
50-40 (+) Moderate-strength activity
40-10 (++) High-strength activity
≦10   (+++)Extremenly high-strength activity

[Determination results]

TABLE 19
Experimental treatment of compounds 1-3 and Sorafenib on human
colon cancer HT-29 transplantable tumor in nude mice
		Animal
	Dosage	number	Weight (g)	TV		T/C
Groups	mg/kg	Start	Final	d0	d13	d0	d13	RTV	(%)
Control		6	6	18.8 ± 1.1	19.6 ± 0.9	133 ± 60	626 ± 226	5.07 ± 1.39
Solvent control		6	6	19.7 ± 0.6	20.8 ± 0.8	133 ± 32	547 ± 172	4.15 ± 0.93	81.85
Sorafenib	60	6	6	19.9 ± 1.1	20.4 ± 1.4	133 ± 33	308 ± 86	2.36 ± 0.57	46.48**
Compound 1	60	6	6	19.5 ± 1.0	20.5 ± 1.1	128 ± 34	359 ± 108	2.81 ± 1.05	55.42**
Compound 2	60	6	6	19.9 ± 0.8	21.0 ± 0.8	133 ± 23	265 ± 100	2.00 ± 0.41	39.45**
Compound 3	60	6	6	19.1 ± 1.0	19.7 ± 1.4	133 ± 18	322 ± 129	2.40 ± 0.67	47.34**

TABLE 20
Relative tumor reproduction rate of compounds 1-10 and Sorafenib on
Human colon cancer HT-29 transplantable tumor in nude mice T/C (%)
	1	2	3	4	5	6	7	8	9	10	Sorafenib
Human colon cancer	55.42	39.45	47.34	33.15	38.24	39.58	40.1	37.6	35.2	78.9	46.48
HT-29 transplantable
tumor in nude mice

TABLE 21
Relative tumor reproduction rate of compounds 11-20 and Sorafenib on
human colon cancer HT-29 transplantable tumor in nude mice T/C (%)
	11	12	13	14	15	16	17	18	19	20
Human colon	81.2	80.5	81.2	40.1	85.2	55.2	54.7	40.2	39.4	40.1
cancer HT-29
transplantable
tumor in nude
mice

TABLE 22
Relative tumor reproduction rate of compounds 21-30 and Sorafenib on
human colon cancer HT-29 transplantable tumor in nude mice T/C (%)
	21	22	23	24	25	26	27	28	29	30
Human colon	38.7	41.2	39.7	37.8	79.5	41.1	80.3	39.1	78.3	79.5
cancer HT-29
transplatable
tumor in nude
mice

TABLE 23
Relative tumor reproduction rate of compounds
31-38 and Sorafenib on human colon cancer HT-29
transplantable tumor in nude mice T/C (%)
	31	32	33	34	35	36	37	38
Human colon	50.3	46.2	39.9	45.9	51.2	41.5	39.5	41.2
cancer HT-29
transplantable
tumor in
nude mice

The results of the above in vivo and vitro tumor inhibition tests showed that the inhibiting effects of such derivatives on S180 sarcoma in mice and human colon cancer HT-29 transplantable tumor in nude mice were better than or equivalent to positive control medicine sorafenib.The test results showed that the compound of the present invention or the pharmaceutically acceptable salt thereof can be used for treating tumor or leukemia. The pharmacodynamic experiments of the compounds in the present invention, positive control medicine sorafenib and compounds A′, B′ and C′ with no substituent or only amino formyl in A ring on human lung cancer cell strain A549, human high-metastic lung cancer cell strain 95D, lung cancer cell A549, human umbilical vein endothelial cell HUVEC cell growth and lumen formation, human lung cancer A549 cell transplantation tumor model in nude mice, human liver cancer cell bel-7402 transplantation tumor model in nude mice, and renal carcinoma cell line GCR-1 transplantation tumor model in nude mice were carried out to verify the effect of the compounds of the present invention.

Sorafenib was abbreviated as Sorafenib hereafter, and the compounds A′, B′ and C′ were respectively prepared according the method of CN200810129360.6, which were compounds with no substituent in A ring, wherein A′ is

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-quinolinyl)oxy)phenyl)urea, B′is

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-pyrimidinyl)oxy)phenyl)urea, and C′

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-pyrryl)oxy)phenyl)urea.

4. Using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay (MTT) to detect the inhibition effects on the growth of human lung cancer cell strain A549

[Test materials] MTT working solution, F12 medium containing 10% FBS, continuous injectors

[Tested compound] Compounds 1-38

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with no substituent or only amino formyl in A ring

The inhibition rate is calculated as follows:

$\mathrm{Cell}\mathrm{reproduction}\mathrm{rate}\%=1-\frac{\left(\begin{array}{c}\mathrm{Relative}\mathrm{OD}\mathrm{value}\mathrm{of}\mathrm{conrol}\mathrm{well}-\\ \mathrm{Relative}\mathrm{OD}\mathrm{value}\mathrm{of}\mathrm{drug}\mathrm{well}\end{array}\right)}{\mathrm{Relative}\mathrm{OD}\mathrm{value}\mathrm{of}\mathrm{conrol}\mathrm{well}}\times 100\%$

Relative OD value of conrol well=OD value of control well−OD value of blank well

Relative OD value of drug well=OD value of drug well−OD value of blank well

[Screening Results]

TABLE 24
Inhibition effects of compounds 1-18 on
the growth of human lung cancer cell A549
	Compound	Final concentration	Inhibition
	No.	1.0*10(−5) mol/L	rate	Activity
	1	1	50.20%	+
	2	1	61.11%	++
	3	1	69.67%	++
	4	1	91.12%	+++
	5	1	93.50%	+++
	6	1	92.50%	+++
	7	1	45.30%	+
	8	1	60.67%	++
	9	1	61.54%	++
	10	1	74.50%	++
	11	1	69.20%	++
	12	1	42.31%	+
	13	1	64.23%	++
	14	1	67.25%	++
	15	1	72.17%	++
	16	1	89.71%	+++
	17	1	88.52%	+++
	18	1	90.71%	+++

TABLE 25
Inhibition effects of compounds 19-38 on
the growth of human lung cancer cell A549
Compound	Final concentration	Inhibition
No.	1.0*10(−5) mol/ml	rate	Activity
19	1	91.23%	+++
20	1	88.32%	+++
21	1	89.21%	+++
22	1	82.49%	+++
23	1	55.41%	+
24	1	86.32%	+++
25	1	63.26%	+
26	1	55.50%	+
27	1	64.71%	+
28	1	88.52%	+++
29	1	86.42%	+++
30	1	85.47%	+++
31	1	46.78%	+
32	1	45.76%	+
33	1	57.53%	+
34	1	59.52%	+
35	1	55.74%	+
36	1	72.45%	++
37	1	71.65%	++
38	1	74.56%	++
Posive Sorafenib	1	54.60%	+
Compound A′	1	58.51%	+
Compound B′	1	61.62%	+
Compound C′	1	62.25%	+

5 Inhibition effects of compounds on human high-metastic lung cancer cell 95D migration

[Test materials] Boyden Chamber Transwell chamber (with pore size of 8 μm), human high-metastic lung cancer cell 95D cell strain, 1640 medium containing 10% FBS, 1640 medium containing no serum

[Tested compound] Compounds 1-38

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with no substituent or only amino formyl in A ring

The inhibition rate is calculated as follows:

$\mathrm{Cell}\mathrm{migration}\mathrm{inhibition}\mathrm{rate}\%=\frac{\left(\begin{array}{c}\mathrm{migrated}\mathrm{cell}\mathrm{number}\mathrm{in}\mathrm{the}\mathrm{chamber}\mathrm{containing}\mathrm{no}\mathrm{drug}-\\ \mathrm{migrated}\mathrm{cell}\mathrm{number}\mathrm{in}\mathrm{the}\mathrm{chamber}\mathrm{containing}\mathrm{drug}\end{array}\right)}{\mathrm{migrated}\mathrm{cell}\mathrm{number}\mathrm{in}\mathrm{the}\mathrm{chamber}\mathrm{containing}\mathrm{no}\mathrm{drug}}\times 100\%$

[Screening Results]

TABLE 26
Inhibition effects of compounds 1-18 on human high-
metastic lung cancer 95D cell strain migration
	Compound	Final concentration	Inhibition
	No.	1.0*10(−5) mol/L	rate	Activity
	1	1	80.21%	++
	2	1	85.17%	+++
	3	1	96.64%	+++
	4	1	96.38%	+++
	5	1	97.51%	+++
	6	1	93.71%	+++
	7	1	89.34%	+++
	8	1	89.56%	+++
	9	1	91.42%	+++
	10	1	61.43%	+
	11	1	78.66%	++
	12	1	66.79%	++
	13	1	65.45%	++
	14	1	57.57%	+
	15	1	63.68%	+
	16	1	89.31%	+++
	17	1	90.52%	+++
	18	1	93.73%	+++

TABLE 27
Inhibition effects of compounds 19-38 on human high-
metastic lung cancer 95D cell strain migration
Compound	Final concentration	Inhibition
No.	1.0*10(−5) mol/L	rate	Activity
19	1	90.21%	+++
20	1	89.12%	+++
21	1	88.76%	+++
22	1	88.77%	+++
23	1	85.53%	+++
24	1	85.48%	+++
25	1	59.76%	+
26	1	60.52%	+
27	1	59.77%	+
28	1	70.53%	+
29	1	61.44%	+
30	1	69.62%	++
31	1	76.18%	++
32	1	66.92%	++
33	1	77.52%	++
34	1	63.65%	+
35	1	68.47%	++
36	1	84.59%	+++
37	1	79.25%	+
38	1	80.53%	+
Positive Sorafenib	1	62.32%	+
Compound A′	1	63.51%	+
Compound B′	1	61.60%	+
Compound C′	1	63.20%	++

6. Effects of tested compounds on the adhesive ability of lung cancer cell A549

[Test materials] gelatin, CCK8, poly-lysine (PLL), A549 cell stains, 1640 medium containing 10% FBS

[Tested compound] Compounds 1-38 to be tested

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with no substituent or only amino formyl in A ring

[Screening Results]

The inhibition rate is calculated as follows:

$\mathrm{Inhibition}\mathrm{rate}\mathrm{of}\mathrm{cell}\mathrm{adhesion}\%=\frac{\begin{array}{c}\mathrm{Cell}\mathrm{group}\mathrm{without}\mathrm{treatment}\\ (\mathrm{glutin}\mathrm{adhesion}\mathrm{OD}/\mathrm{PLL}\mathrm{adhesion}\mathrm{OD}\mathrm{value})-\\ \mathrm{dosing}\mathrm{cell}\mathrm{group}\left(\mathrm{glutin}\mathrm{adhesion}\mathrm{OD}/\mathrm{PLL}\mathrm{adhesion}\mathrm{OD}\mathrm{value}\right)\end{array}}{\begin{array}{c}\mathrm{Cell}\mathrm{group}\mathrm{without}\mathrm{treatment}\\ \left(\mathrm{glutin}\mathrm{adhesion}\mathrm{OD}/\mathrm{PLL}\mathrm{adhesion}\mathrm{OD}\mathrm{value}\right)\end{array}}\times 100\%$

Dosing dosing cells without cell group (gelatin adhesive OD/PLL adhesion OD)−Dosing cell group (gelatin adhesive OD/PLL adhesion OD value)

Dosing dosing cells without cell group (gelatin adhesive OD/PLL adhesion OD value)

TABLE 28
Inhibition effects of compounds 1-18 on the adhesion
ability of human lung cancer cell A549
	Compound	Final concentration	Inhibition
	No.	1.0*10(−5) mol/L	rate	Activity
	1	1	60.22%	+
	2	1	75.15%	++
	3	1	79.66%	++
	4	1	89.71%	+++
	5	1	87.58%	+++
	6	1	93.59%	+++
	7	1	68.34%	++
	8	1	61.56%	+
	9	1	85.32%	+++
	10	1	64.57%	+
	11	1	59.63%	+
	12	1	62.30%	+
	13	1	63.39%	+
	14	1	67.51%	++
	15	1	68.63%	++
	16	1	90.77%	+++
	17	1	97.50%	+++
	18	1	93.72%	+++

TABLE 29
Inhibition effects of compounds 19-38 on the adhesion
ability of human lung cancer cell A549
Compound	Final concentration	Inhibition
No.	1.0*10(−5) mol/L	rate	Activity
19	1	90.28%	+++
20	1	95.31%	+++
21	1	90.22%	+++
22	1	72.44%	++
23	1	77.56%	++
24	1	73.30%	++
25	1	63.51%	+
26	1	68.57%	++
27	1	94.77%	+++
28	1	90.53%	+++
29	1	91.40%	+++
30	1	92.44%	+++
31	1	66.77%	++
32	1	64.73%	+
33	1	77.59%	++
34	1	76.54%	++
35	1	75.72%	++
36	1	74.50%	++
37	1	71.74%	++
38	1	75.53%	++
Positive medicine	1	72.66%	++
Sorafenib
Compound A′	1	71.55%	++
Compound B′	1	69.26%	++
Compound C′	1	68.62%	++

7. Effects of tested compounds on the growth of human umbilical vein endothelial cell HUVEC cell by CCK8 method

[Test materials] CCK8, human umbilical vein endothelial cell HUVEC cell, 1640 medium containing 10% FBS

[Tested compound] Compounds 1-38 to be tested

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with no substituent or only amino formyl in A ring

[Screening Results]

$\mathrm{Cell}\mathrm{reproduction}\mathrm{rate}\%=1-\frac{\left(\begin{array}{c}\mathrm{Relative}\mathrm{OD}\mathrm{value}\mathrm{of}\mathrm{conrol}\mathrm{well}-\\ \mathrm{Relative}\mathrm{OD}\mathrm{value}\mathrm{of}\mathrm{drug}\mathrm{well}\end{array}\right)}{\mathrm{Relative}\mathrm{OD}\mathrm{value}\mathrm{of}\mathrm{conrol}\mathrm{well}}\times 100\%$

Relative OD value of conrol well=OD value of control well−OD value of blank well

Relative OD value of drug well=OD value of drug well−OD value of blank well

TABLE 30
Inhibition effects of compounds 1-18 on the growth
of human umbilical vein endothelial cell HUVEC cell
	Compound	Final concentration	Inhibition
	No.	1.0*10(−5) mol/L	rate	Activity
	1	1	9.20%
	2	1	13.11%
	3	1	27.67%
	4	1	5.12%
	5	1	8.50%
	6	1	7.50%
	7	1	8.30%
	8	1	13.67%
	9	1	11.54%
	10	1	14.50%
	11	1	29.20%
	12	1	12.31%
	13	1	24.23%
	14	1	17.25%
	15	1	38.17%
	16	1	10.71%
	17	1	13.52%
	18	1	5.71%

TABLE 31
Inhibition effects of compounds 19-38 on the growth
of human umbilical vein endothelial cell HUVEC cell
Compound	Final concentration	Inhibition
No.	1.0*10(−5) mol/L	rate	Activity
19	1	5.23%
20	1	7.32%
21	1	11.21%
22	1	12.49%
23	1	17.41%
24	1	13.32%
25	1	23.26%
26	1	32.50%
27	1	24.73%
28	1	20.55%
29	1	21.40%
30	1	25.46%
31	1	26.70%
32	1	24.77%
33	1	17.50%
34	1	19.52%
35	1	5.74%
36	1	12.45%
37	1	19.65%
38	1	14.56%
Positive medicine	1	22.61%
Sorafenib
Compound A′	1	23.11%
Compound B′	1	31.64%
Compound C′	1	22.27%

8. Inhibition effects of compounds on the lumen formation ability of human umbilical vein endothelial cell HUVEC

[Experimental principles] The human umbilical vein endothelial cells have ability of spontaneously forming blood lumen on Matrigel, which can be used to simulate the process of angiogenesis in vivo. We used Matrigel method to investigate the effects of the compound on the lumen formation ability of human umbilical vein endothelial cell HUVEC.

[Test materials] HUVEC (taking generation 3 to 5 cells for experiments after obtained from primary separation and cultured at 37 under the conditions of 5% CO₂), Matrigel, cell culture medium M199.

[Tested compound] Compounds 1-38

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with no substituent or only amino formyl in A ring

[Screening Results]

The inhibition rate is calculated as follows:

$\mathrm{Lumen}\mathrm{formation}\mathrm{inhibition}\mathrm{rate}\%=\frac{\left(\begin{array}{c}\mathrm{length}\mathrm{sum}\mathrm{of}\mathrm{lumen}\mathrm{without}\mathrm{dosing}-\\ \mathrm{length}\mathrm{sum}\mathrm{of}\mathrm{lumen}\mathrm{after}\mathrm{dosing}\end{array}\right)}{\mathrm{length}\mathrm{sum}\mathrm{of}\mathrm{lumen}\mathrm{without}\mathrm{dosing}}\times 100\%$

TABLE 32
Inhibition effects of compounds 1-18 on the lumen formation
ability of human umbilical vein endothelial cell HUVEC
	Compound	Final concentration	Inhibition
	No.	1.0*10(−5) mol/L	rate	Activity
	1	1	72.22%	++
	2	1	75.14%	++
	3	1	76.61%	++
	4	1	86.77%	+++
	5	1	87.50%	+++
	6	1	90.55%	+++
	7	1	60.34%	+
	8	1	78.59%	++
	9	1	91.45%	+++
	10	1	50.34%	+
	11	1	79.63%	++
	12	1	52.37%	+
	13	1	53.36%	+
	14	1	67.53%	++
	15	1	68.64%	++
	16	1	80.79%	++
	17	1	67.50%	++
	18	1	63.71%	+

TABLE 33
Inhibition effects of compounds 19-38 on the lumen formation
ability of human umbilical vein endothelial cell HUVEC
Compound	Final concentration	Inhibition
No.	1.0*10(−5) mol/L	rate	Activity
19	1	89.22%	+++
20	1	88.54%	+++
21	1	89.47%	+++
22	1	86.72%	+++
23	1	97.56%	+++
24	1	93.78%	+++
25	1	79.79%	++
26	1	70.53%	++
27	1	89.75%	+++
28	1	89.52%	+++
29	1	86.43%	+++
30	1	90.66%	+++
31	1	66.77%	++
32	1	54.74%	+
33	1	67.51%	++
34	1	66.52%	++
35	1	65.70%	++
36	1	64.50%	+
37	1	71.72%	++
38	1	65.55%	++
Positive medicine	1	55.60%	++
Sorafenib
Compound A′	1	60.51%	++
Compound B′	1	61.62%	++
Compound C′	1	60.50%	++

9. Tumor inhibition rate of tested compounds on human lung cancer A549 cell transplantation model in nude mice

[Test animals] female BALB/cA nude mice, 35-40 days old, with weight of 18-22g. There were 12 mice in the negative conrol group and 6 mice in the treatment group.

[Test method] Take eugenic tumor tissues and cut into about 1.5 mm³, and then incoculate subcutaneously at the right armpit of nude mice under the sterile conditions. The diameter of the transplantable tumor in nude mice was determined with a vernier caliper, and the animals were divided into groups after the tumors were grown to 100-300 mm³.Using the method of measuring the tumor diameter, dynamically observe the antitumor effects of tested materials.

The diameter of the tumor was determined three times every week and the mouse weight was weighed at the same time. The dosage of the compound was 60 mg/kg, 6 times every week for 3 weeks. Sorafenib was oral administrated with dosage of 60 mg/kg, 6 times every week for 3 weeks. Equal amount of normal saline was administrated in the negative control group. Observe for one week after administration.

[Detection Indicators and Calculation Methods]

(1) Tumor volume (TV) is calculated as:

• • TV=1/2×a×b²
  • wherein a and b respectively represents length and width.

(2) Relative tumor volume (RTV) is calculated as:

• • RTV=TV_t/TV_0°

wherein TV₀ is the tumor volume when administrated according to different cages and TV_t is the tumor volume measured each time.

(3) Relative tumor reproduction rate T/C (%) is calculated as follows:

$T\text{/}C\left(\%\right)=\frac{T_{\mathrm{RTV}}}{C_{\mathrm{RTV}}}\times 100$

T_RTV: RTV in the treatment group; C_RTV: RTV in the negative control group. The test results used relative tumor reproduction rate T/C (%) as evaluating indicator of anti-tumor activity.

[Screening results] There was no mortality for the animals in the group of compounds and Sorafenib in the experiments with less toxicity.

TABLE 34
Relative tumor reproduction rate of compounds and Sorafenib on human
lung cancer A549 transplantation tumor in nude mice T/C (%)
Relative tumor reproduction rate of compounds 1-10 and Sorafenib on human lung
cancer A549 transplantation tumor in nude mice T/C (%)
	1	2	3	4	5	6	7	8	9	10
human lung	38.7	35.3	37.2	12.3	11.6	15.9	39.3	30.5	38.7	30.1
cancer A549
transplantation
tumor
Relative tumor reproduction rate of compounds 11-20 and Sorafenib on human lung
cancer A549 transplantation tumor in nude mice T/C (%)
	11	12	13	14	15	16	17	18	19	20
human lung	43.7	36.3	38.4	32.2	38.7	9.9	7.3	12.5	7.7	12.1
cancer A549
transplantation
tumor
Relative tumor reproduction rate of compounds 21-30 and Sorafenib on human lung
cancer A549 transplantation tumor in nude mice T/C (%)
	21	22	23	24	25	26	27	28	29	30
human lung	13.7	15.3	17.1	15.6	39.4	33.1	36.3	13.6	14.7	13.1
cancer A549
transplantation
tumor
Relative tumor reproduction rate of compounds 31-38 and Sorafenib on human lung
cancer A549 transplantation tumor in nude mice T/C (%)
	31	32	33	34	35	36	37	38	Sorafenib
human lung	38.7	35.3	37.5	22.3	23.6	37.9	38.3	36.5	41.8
cancer A549
transplantation
tumor
		Compound A′	Compound B′	Compound C′
	human lung	43.2	40.4	36.6
	cancer A549
	transplantation
	tumor

10. Tumor inhibition rate of tested compounds on human liver cancer cell bel-7402 transplantation tumor model in nude mice

[Test animals] female BALB/cA nude mice, 35-40 days old, with weight of 18-22g. There were 12 mice in the negative conrol group and 6 mice in the treatment group.

[Test method] Take eugenic tumor tissues and cut into about 1.5 mm³, and then incoculate subcutaneously at the right armpit of nude mice under the sterile conditions. The diameter of the transplantable tumor in nude mice was determined with a vernier caliper, and the animals were divided into groups after the tumors were grown to 100-300 mm³.Using the method of measuring the tumor diameter, dynamically observe the antitumor effects of tested materials.

The diameter of the tumor was determined three times every week and the mouse weight was weighed at the same time. The dosage of the compound was 60 mg/kg, 6 times every week for 3 weeks. Sorafenib was oral administrated with dosage of 60 mg/kg, 6 times every week for 3 weeks. Equal amount of normal saline was administrated in the negative control group. Observe for one week after administration.

[Detection Indicators and Calculation Methods]

(1) Tumor volume (TV) is calculated as:

• • TV=1/2×a×b²
  • wherein a and b respectively represents length and width.

(2) Relative tumor volume (RTV) is calculated as:

• • RTV=TV_t/TV_0°

wherein TV₀ is the tumor volume when administrated according to different cages (d₀) and TV_t is the tumor volume measured each time.

(3) Relative tumor reproduction rate T/C (%) is calculated as follows:

$T\text{/}C\left(\%\right)=\frac{T_{\mathrm{RTV}}}{C_{\mathrm{RTV}}}\times 100$

T_RTV: RTV in the treatment group; C_RTV: RTV in the negative control group. The test results used relative tumor reproduction rate T/C (%) as evaluating indicator of anti-tumor activity.

[Screening results] There was no mortality for the animals in the group of compounds and Sorafenib in the experiments with less toxicity.

TABLE 35
Relative tumor reproduction rate of compounds and Sorafenib on human liver
cancer cell bel-7402 transplantation tumor model in nude mice T/C (%)
Relative tumor reproduction rate of compounds 1-10 and Sorafenib on human liver
cancer cell bel-7402 transplantation tumor model in nude mice T/C (%)
	1	2	3	4	5	6	7	8	9	10
human liver	33.7	35.4	30.1	15.3	15.7	19.5	34.2	30.6	36.8	28.1
cancer cell
bel-7402
transplantation
tumor
	11	12	13	14	15	16	17	18	19	20
human liver	23.7	31.6	29.8	36.1	33.6	14.9	12.5	12.7	16.3	17.2
cancer cell
bel-7402
transplantation
tumor
	21	22	23	24	25	26	27	28	29	30
human liver	16.4	15.3	17.7	16.8	29.1	36.1	29.2	17.5	16.2	16.1
cancer cell
bel-7402
transplantation
tumor
	31	32	33	34	35	36	37	38	Sorafenib
human liver	33.4	31.4	32.7	31.5	34.5	37.9	36.3	31.5	35.9
cancer cell
bel-7402
transplantation
tumor
		Compound A′	Compound B′	Compound C′
	human liver	37.2	30.4	32.6
	cancer cell
	bel-7402
	transplantation
	tumor

11. Tumor inhibition rate of medicines on renal carcinoma cell line GCR-1 transplanted tumor model in nude mice

[Test animals] female BALB/cA nude mice, 35-40 days old, with weight of 18-22g. There were 12 mice in the negative conrol group and 6 mice in the treatment group.

[Test method] Take eugenic tumor tissues and cut into about 1.5 mm³, and then incoculate subcutaneously at the right armpit of nude mice under the sterile conditions. The diameter of the transplantable tumor in nude mice was determined with a vernier caliper, and the animals were divided into groups after the tumors were grown to 100-300 mm³.Using the method of measuring the tumor diameter, dynamically observe the antitumor effects of tested materials. The diameter of the tumor was determined three times every week and the mouse weight was weighed at the same time. The dosage of the medicine was 60 mg/kg, 6 times every week for 3 weeks. Sorafenib was oral administrated with dosage of 60 mg/kg, 6 times every week for 3 weeks. Equal amount of normal saline was administrated in the negative control group. Observe for one week after administration.

[Detection Indicators and Calculation Methods]

(1) Tumor volume (TV) is calculated as:

• • TV=1/2×a×b²
  • wherein a and b respectively represents length and width.

(2) Relative tumor volume (RTV) is calculated as:

• • RTV=TV_t/TV_0°
  • wherein TV₀ is the tumor volume when administrated according to different cages (d₀) and TV_t is the tumor volume measured each time.

(3) Relative tumor reproduction rate T/C (%) is calculated as follows:

$T\text{/}C\left(\%\right)=\frac{T_{\mathrm{RTV}}}{C_{\mathrm{RTV}}}\times 100$

• • T_RTV: RTV in the treatment group; C_RTV: RTV in the negative control group. The test results used relative tumor reproduction rate T/C (%) as evaluating indicator of anti-tumor activity.

[Screening results] There was no mortality for the animals in the group of compounds and Sorafenib in the experiments with less toxicity.

TABLE 36
Relative tumor reproduction rate of compounds and Sorafenib on human renal
carcinoma GCR-1 cell transplanted tumor model in nude mice T/C (%)
Relative tumor reproduction rate of compounds 11-20 and Sorafenib on human renal
carcinoma GCR-1 cell transplanted tumor model in nude mice T/C (%)
	1	2	3	4	5	6	7	8	9	10
human renal	30.5	23.2	31.3	9.6	11.2	12.5	21.2	29.1	28.3	27.1
carcinoma
GCR-1 cell
transplanted
tumor
	11	12	13	14	15	16	17	18	19	20
human renal	32.1	20.5	28.3	31.1	22.5	9.9	11.1	12.3	13.2	13.1
carcinoma
GCR-1 cell
transplanted
tumor
Relative tumor reproduction rate of compounds 21-30 and Sorafenib on human renal
carcinoma GCR-1 cell transplanted tumor model in nude mice T/C (%)
	21	22	23	24	25	26	27	28	29	30
human renal	7.8	8.1	9.1	10.8	31.2	24.2	26.2	11.4	12.8	10.2
carcinoma
GCR-1 cell
transplanted
tumor
	31	32	33	34	35	36	37	38	Sorafenib
human renal	31.4	32.5	33.4	36.1	32.3	25.9	32.3	20.5	33.9
carcinoma
GCR-1 cell
transplanted
tumor
	Compound A′	Compound B′	Compound C′
	human renal	34.2	30.4	35.6	34.2	30.4
	carcinoma
	GCR-1 cell
	transplanted
	tumor

According to the experimental results, the compound added with specific substituents in A ring have stronger anti-tumor activity than the compouns with no substituent or only amino formyl in A ring, especially the 4#-6# 16#-18# 19#-24# 28#-30# compounds have stonger anti-tumor activity which are stonger than the positive conrol Sorafenib, which have particularly evident effects on the tumor cell metastasis and tumor angiogenesis that are significantly stronger than Sorafenib. The test on normal human umbilical vein endothelial cells CCK8 found that these compounds have less toxicity to normal human cells like endothelial cells, which are relatively safe and reliable, but these compounds can achieve the antitumor activity through inhibiting the tumor angiogenesis. The in vivo transplantation experiments in nude mice showed that 4#-6# 16#-18# 19#-24# 28#-30# compounds have inhibition effects on human liver cancer and renal caner and their effects are better than Sorafenib, but these compounds have very significant effects on lung cancer and the effects obviously exceed the positive control medicine Sorafenib, which is an unexpected result.

The above results indicate that the compounds added with specific substituents in A-ring have more advantages than previously found compounds with no substituent or only amino formyl in A ring, and these new compounds have broader prospects in the treatment of cancer.

Claims

1. A compound or pharmaceutically acceptable salts thereof, wherein the compound is represented by the following formula [1] or [2]

2. The compound or pharmaceutically acceptable salts thereof according to claim 1, wherein A is monosubstituted or polysubstituted quinazoline, pyrrole or pyrimidine, and the substituent is halogen, C1-5alkyl, C1-5haloalkyl, C1-5alkoxy, C1-5haloalkoxy, C1-5alkylamino, C1-5haloalkylamino, amino or nitryl.

3. The compound or pharmaceutically acceptable salts thereof according to claim 2, wherein A is monosubstituted or polysubstituted quinazoline, and the substituent is halogen, C1-5alkyl, C1-5haloalkyl, C1-5alkoxy, C1-5haloalkoxy, amino or nitryl.

4. The compound or pharmaceutically acceptable salts thereof according to claim 1, wherein R1 is methyl or ethyl, propyl, isopropyl.

5. The compound or pharmaceutically acceptable salts thereof according to claim 1, wherein R2 is one or more selected from hydrogen, halogen, C1-5alkyl, C1-5 alkoxy or C1-5haloalkyl.

6. The compound or pharmaceutically acceptable salts thereof according to claim 5, wherein R2 is one or more selected from H, Cl, Br, F, methoxy, ethoxy, propoxy, methyl, ethyl, propyl, isopropyl, butyl, t butyl or trifluoromethyl.

7. The compound or pharmaceutically acceptable salts thereof according to claim 1, wherein R3 is one or more selected from hydrogen, halogen, C1-5alkyl, C1-5alkoxy, or C1-5haloalkyl.

8. The compound or pharmaceutically acceptable salts thereof according to claim 7, wherein R3 is one or more selected from H, Cl, Br, F, methoxy, ethoxy, propoxy, methyl, ethyl, propyl, isopropyl, butyl, t-butyl or trifluoromethyl.

9. The compound or pharmaceutically acceptable salts thereof according to claim 1, wherein the salts are selected from: a) basic salts of inorganic acids and organic acids, the described acid is selected from hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, mesylate, trifluoromethanesulfonic acid, benzene sulfonic acid, paratoluenesulfonic acid, 1-naphthalene sulfonic acid, 2-naphthalene sulfonic acid, acetic acid, trifluoroacetic acid, malic acid, tartaric acid, citric acid, lactic acid, oxalic acid, succinic acid, fumaric acid, maleic acid, benzoic acid, salicylic acid, phenylacetic acid or almonds acid;b) acid salts of organic and inorganic base, the described cation is selected from alkali metal cation, alkaline earth metal cation, ammonium cation, aliphatic-substituted ammonium cation or aromatic-substituted ammonium cation.

10. The compound or pharmaceutically acceptable salts thereof according to any one of claims 1 to 9, wherein the compound is selected from: the following N′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea:N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea;the following N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy) phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy) phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy) phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy) phenyl)urea;the following N′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy)) phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy)) phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy)) phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy)) phenyl)urea;the following N′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea;the following N′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)urea;the following N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)urea;the following N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)urea;the following N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)urea;the following N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea;the following N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)urea;the following substituted quinoline derivativesN-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea;N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-methyl-4-(2-(N-ethylcarbamoyl)-4-(7-nitryl-quinolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-methoxyl-4-(2-(N-ethylcarbamoyl)-4-(7-nitryl-quinolinyl)oxy)phenyl)urea;the following substituted quinazoline derivativesN-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinazolinyl)oxy)phenyl)urea;N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinazolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-fluoro-4-(2-(N-propylcarbamoyl)-4-(7-trifluoromethylquinazolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-trifluoromethyl-4-(2-(N-propylcarbamoyl)-4-(7-trifluoromethylquinazolinyl)oxy)phenyl)urea.

11. A method of treating tumor or leukemia in subject, the method comprising administering the subject an effective dosage amount of the compound or pharmaceutically acceptable salts thereof according to any one of claims 1 to 9.