WO2015127873A1 - 芳氨基嘧啶类化合物及其应用以及由其制备的药物组合物和药用组合物 - Google Patents

芳氨基嘧啶类化合物及其应用以及由其制备的药物组合物和药用组合物 Download PDF

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WO2015127873A1
WO2015127873A1 PCT/CN2015/073045 CN2015073045W WO2015127873A1 WO 2015127873 A1 WO2015127873 A1 WO 2015127873A1 CN 2015073045 W CN2015073045 W CN 2015073045W WO 2015127873 A1 WO2015127873 A1 WO 2015127873A1
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phenyl
amino
pyrimidin
acrylamide
ureido
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PCT/CN2015/073045
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French (fr)
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兰炯
崔玉敏
张亮
金云舟
周福生
程鹏飞
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上海海雁医药科技有限公司
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Priority to CN201580002151.3A priority Critical patent/CN105683168B/zh
Publication of WO2015127873A1 publication Critical patent/WO2015127873A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D239/00—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings
    • C07D239/02—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings
    • C07D239/24—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members
    • C07D239/28—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, directly attached to ring carbon atoms
    • C07D239/46—Two or more oxygen, sulphur or nitrogen atoms
    • C07D239/48—Two nitrogen atoms
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5375—1,4-Oxazines, e.g. morpholine
    • A61K31/5377—1,4-Oxazines, e.g. morpholine not condensed and containing further heterocyclic rings, e.g. timolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D239/00—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings
    • C07D239/02—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings
    • C07D239/24—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members
    • C07D239/28—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, directly attached to ring carbon atoms
    • C07D239/32—One oxygen, sulfur or nitrogen atom
    • C07D239/42—One nitrogen atom
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings

Definitions

  • the present invention relates to the field of medical technology, and in particular to an aromatic aminopyrimidine compound and its use as an EGFR tyrosine kinase inhibitor, and pharmaceutical compositions and pharmaceutical compositions prepared therefrom.
  • Lung cancer is the world's highest incidence of cancer. It ranks first among all cancers in China. It is also the cancer with the highest morbidity and mortality in China.
  • EGFR mutations of which L858R and Exon 19 deletion mutations account for more than 90%, and such patients are more sensitive to EGFR inhibitors.
  • the first-generation EGFR inhibitors such as erlotinib have been marketed, and gefitinib is effective in such patients, which can reduce tumors in more than 60% of patients and significantly prolong the progression-free survival of patients.
  • the second-generation irreversible pan-EGFR inhibitor (Afatinib (BIBW2992)) currently on the market is significantly better than the first-generation EGFR inhibitor in patients with EGFR-mutant lung cancer.
  • the second-generation inhibitor also has a strong wild-type EGFR inhibitory activity, and the inhibitory activity against wild-type EGFR is significantly higher than that of the resistant T790M mutation.
  • the patient's rash and other toxic side effects are serious and the curative effect on drug-resistant patients is poor.
  • a small number of first-generation EGFR inhibitor-resistant patients respond to these drugs.
  • An object of the present invention is to provide an arylaminopyrimidine compound, or a pharmaceutically acceptable salt thereof, a stereoisomer, a solvent compound or a prodrug thereof.
  • Another object of the present invention is to provide a pharmaceutical composition
  • a pharmaceutical composition comprising the above-described arylaminopyrimidine compound, or a pharmaceutically acceptable salt thereof, a stereoisomer, a solvent compound or a prodrug thereof.
  • a third object of the present invention is to provide a medicament for preparing an arylaminopyrimidine compound for regulating EGFR tyrosine kinase activity or for preparing a medicament for treating EGFR-related diseases.
  • the first aspect of the present invention provides a novel arylaminopyrimidine compound, or a pharmaceutically acceptable salt thereof, a stereoisomer, a solvate compound or a prodrug thereof, wherein the arylaminopyrimidine compound has a structure such as ) shown:
  • A is selected from: substituted or unsubstituted C 6-10 arene, substituted or unsubstituted C 5-8 heteroaryl, substituted or unsubstituted C 3-8 cycloalkyl, substituted or Unsubstituted C 3-8 heterocycloalkyl, substituted or unsubstituted 8-14 membered fused ring group; wherein said "substituted” means that one or more hydrogens in the group are selected from the group consisting of Substituted by a substituent: nitro, halogen, cyano, C 1-6 alkyl, halogenated C 1-6 alkyl, C 1-6 alkoxy, halogenated C 1-6 alkoxy;
  • Z 1 is selected from NR 9 or CR 10 R 11 ; wherein R 9 is selected from hydrogen or C 1-6 alkyl; R 10 and R 11 are each independently selected from hydrogen, halogen, C 1-6 alkyl; or R 10 and R 11 together with a carbon atom to which they are bonded form a C 3-8 cycloalkyl group or a C 3-8 heterocycloalkyl group;
  • Z 2 and Z 3 are each independently selected from CR 12 or N; wherein R 12 is selected from the group consisting of hydrogen, halogen, nitro, amino, C 1-6 alkyl substituted amino, C 1-6 acyl substituted amino, cyanide a group, a C 1-6 alkyl group, a halogenated C 1-6 alkyl group, a C 1-6 alkoxy group, a halogenated C 1-6 alkoxy group;
  • R 1 is selected from hydrogen or C 1-6 alkyl; or R 1 together with A and the nitrogen atom to which they are attached constitutes a group selected from substituted or unsubstituted C 5-8 heteroaryl, substituted or unsubstituted a substituted C 3-8 heterocycloalkyl group, or a substituted or unsubstituted 8-14 membered fused ring group, wherein said "substituted” means that one or more hydrogens in the group are selected from the group consisting of Substituted by a substituent: nitro, halogen, cyano, C 1-6 alkyl, halogenated C 1-6 alkyl, C 1-6 alkoxy, halogenated C 1-6 alkoxy;
  • R 2 is selected from the group consisting of hydrogen, halogen, C 1-6 alkyl, halogenated C 1-6 alkyl, or C 1-6 alkoxy;
  • R 3 , R 4 and R 5 are each independently selected from the group consisting of hydrogen, halogen, nitro, amino, C 1-6 alkyl substituted amino, C 1-6 acyl substituted amino, C 1-6 alkyl substituted Diamino, cyano, C 1-6 alkyl, halogenated C 1-6 alkyl, C 1-6 alkoxy, C 1-6 alkoxy substituted C 1-6 alkoxy, C 6 -10 arene-substituted C 1-6 alkoxy, halogenated C 1-6 alkoxy, -OC 6-10 arene, -CO-C 3-6 heterocycloalkyl, -CO-NR 13 R 14 , C 3-6 heteroaryl, C 3-6 heterocycloalkyl, C 3-6 heterocycloalkyl-CO-C 1-6 alkyl, C 3-6 heterocycloalkyl-NC 1- 6 alkyl, C 3-6 heterocycloalkyl-C 1-6 alkyl, C 3-6 heterocycloalkyl-C 3-6 hetero
  • R 7 and R 8 are each independently selected from the group consisting of hydrogen, halogen, C 1-6 alkyl, C 3-8 cycloalkyl, C 3-8 heterocycloalkyl, C 6-10 arene or C 5-8 Heteroaryl.
  • Z 1 is selected from NR 9 or CR 10 R 11 ;
  • R 9 is selected from hydrogen or methyl; and
  • R 10 and R 11 are each independently selected from hydrogen or methyl.
  • Z 2 and Z 3 are each independently selected from CH or N.
  • R 1 is selected from hydrogen or methyl, or R 1 together with A and the nitrogen atom to which they are attached constitutes piperidine.
  • R 2 is selected from the group consisting of hydrogen, fluorine, chlorine, methyl, ethyl, propyl, isopropyl, trifluoromethyl.
  • R 3 is hydrogen, halogen, C 1-6 alkoxy (preferably methoxy).
  • R 3 is hydrogen or methoxy.
  • R 4 is hydrogen, halogen, C 1-6 alkoxy (preferably methoxy).
  • R 4 is hydrogen, fluorine, chlorine or methoxy.
  • R 5 is hydrogen, halogen, amino, C 1-6 alkyl-substituted diamino, C 1-6 alkyl, C 1-6 alkoxy, C 3-6 heterocycloalkyl , -CO-C 3-6 heterocycloalkyl, -CO-NR 13 R 14 , C 3-6 heterocycloalkyl-C 3-6 heterocycloalkyl, said C 3-6 heterocycloalkyl Substituted by one or more substituents selected from the group consisting of C 1-6 alkyl, halogenated C 1-6 alkyl, C 1-6 alkyl substituted amino.
  • R 5 is hydrogen, fluorine, chlorine, diethylamino, N,N,N'-trimethylethylenediamine, methyl, ethyl, propyl, methoxy, Ethoxy, C 6 heterocycloalkyl, -CO-C 6 heterocycloalkyl, -CO-N(CH 2 CH 3 ) 2 , C 6 heterocycloalkyl-C 6 heterocycloalkyl, said C
  • the 6 heterocycloalkyl group may be substituted by one or more substituents selected from the group consisting of methyl, ethyl, propyl, fluoroethyl, N(CH 3 ) 2 , N(CH 2 CH 3 ) 2 .
  • R 5 is hydrogen, fluorine, chlorine, diethylamino, N,N,N'-trimethylethylenediamine, methyl, ethyl, propyl, methoxy, Ethoxy, morpholinyl, piperazinyl, piperidinyl, -CO-morpholinyl, -CO-piperazinyl, -CO-piperidinyl, -CO-N(CH 2 CH 3 ) 2 , piperazine Pyridyl-piperazinyl, piperidinyl-morpholinyl, the morpholinyl, piperazinyl, piperidinyl may be substituted by one or more substituents selected from the group consisting of methyl, ethyl, Propyl, fluoroethyl, N(CH 3 ) 2 , N(CH 2 CH 3 ) 2 .
  • R 5 is methoxy or is selected from the group consisting of:
  • the arylaminopyrimidine compound is a compound of formula (II):
  • A, Z 2 , Z 3 , R 1 , R 2 , R 3 , R 4 , R 5 , R 7 , R 8 and R 9 are as defined in the formula (I).
  • the arylaminopyrimidine compound is a compound of the formula (III):
  • A, Z 2 , Z 3 , R 1 , R 2 , R 3 , R 4 , R 5 , R 7 , R 8 , R 10 and R 11 are as defined in the formula (I).
  • R 7 and R 8 are each independently hydrogen.
  • the compound of formula (I), the compound of formula (II), or the compound of formula (III) provided by the present invention A is a phenyl group.
  • the arylaminopyrimidine compound is a compound of formula a:
  • Z 1 , Z 2 , Z 3 , R 1 , R 2 , R 3 , R 4 , R 5 , R 7 and R 8 are as defined in the formula (I).
  • A is a phenyl group, and R 7 and R 8 are each independently hydrogen.
  • R 1 together with A and the nitrogen atom to which they are attached constitute a piperidine.
  • the arylaminopyrimidine compound is a compound of formula b:
  • Z 1 , Z 2 , Z 3 , R 2 , R 3 , R 4 , R 5 , R 7 and R 8 are as defined in the formula (I).
  • the present invention provides a compound of formula (I), a compound of formula (II), or a compound of formula (III) wherein R 1 together with A and the nitrogen atom to which they are attached constitutes piperidine, and R 7 And R 8 are each independently hydrogen.
  • the invention provides a compound selected from the group consisting of: or a pharmaceutically acceptable salt, stereoisomer, solvent compound or prodrug thereof:
  • the alkyl group as a group or a part of other groups, such as a halogen-substituted alkyl group or a hydroxy-substituted alkyl group, may be linear or Branched.
  • C 1-6 alkyl denotes an alkyl group having from 1 to 6 carbons including, but not limited to, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl Base, n-pentyl, n-hexyl.
  • the "alkoxy group” means a group formed by linking an alkyl group to an oxygen atom, including but not limited to a methoxy group, an ethoxy group, an isopropoxy group. , cyclopropoxy, and the like.
  • the "diamino group” means a diamino group which is bonded through an alkyl group, such as ethylenediamine, propylenediamine, butanediamine, etc.; Alkyl substituted, for example N,N,N'-trimethylbutanediamine.
  • the "amino group” means NH 2 .
  • the "C 1-6 alkyl-substituted amino group” means that one or two hydrogens in NH 2 are substituted by a C 1-6 alkyl group. These include, but are not limited to, NH(CH 3 ), N(CH 3 ) 2 , N(CH 2 CH 3 ) 2 and the like.
  • halogen means fluorine, chlorine, bromine and iodine. Preferred are fluorine and chlorine.
  • aromatic hydrocarbon group means a monocyclic or fused aromatic hydrocarbon ring (for example, a phenyl group and a naphthyl group) containing six to ten carbon atoms.
  • heteroaryl means any fused or non-fused aromatic ring system in which at least one ring contains 1-4 selected from nitrogen, oxygen and sulfur.
  • a five to eight membered ring of a hetero atom, preferably at least one hetero atom is selected from the group consisting of nitrogen.
  • Heteroaryl includes, but is not limited to, thienyl, furyl, imidazolyl, pyrazolyl, thiazolyl, oxazolyl, isoxazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, benzimidazolyl , benzopyrazolyl, fluorenyl and the like.
  • cycloalkyl means a saturated or partially unsaturated monocyclic, fused ring or bridged ring containing a specified number of carbon atoms.
  • C 3-8 cycloalkyl refers to a cycloalkyl group of three to eight carbons including cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and the like.
  • heterocycloalkyl means a cycloalkyl group as defined in the present invention, wherein one or more carbon atoms on the ring are oxygen, nitrogen, -NR-, Substituted by a group such as sulfur, carbonyl, -S(O)- or -S(O) 2 -; heterocycloalkyl groups include, but are not limited to, morpholinyl, piperazinyl, piperidinyl, thiomorpholinyl and the like.
  • the "C 1-6 acyl group” includes a formyl group, an acetyl group, a propionyl group, a butyryl group, a valeryl group, or a hexanoyl group.
  • the "fused ring group” means a polycyclic organic compound in which two or more carbocyclic or heterocyclic rings are formed by a shared ring, such as naphthalene, anthracene, anthracene. ⁇ , Philippine, etc.
  • the “pharmaceutically acceptable salt” includes a pharmaceutically acceptable acid addition salt and a pharmaceutically acceptable base addition salt.
  • “Pharmaceutically acceptable acid addition salt” means a salt formed with an inorganic or organic acid which retains the bioavailability of the free base without any other side effects.
  • Inorganic acid salts include, but are not limited to, hydrochlorides, hydrobromides, sulfates, phosphates, and the like; organic acid salts include, but are not limited to, formate, acetate, propionate, glycolate, gluconate , lactate, oxalate, maleate, succinate, fumarate, tartrate, citrate, glutamate, aspartate, benzoate, methanesulfonate , p-toluenesulfonate and salicylate. These salts can be prepared by methods known in the art.
  • “Pharmaceutically acceptable base addition salts” including but not limited to salts of inorganic bases such as sodium, potassium, calcium and magnesium salts, and the like. These include, but are not limited to, salts of organic bases such as ammonium salts, triethylamine salts, lysine salts, arginine salts and the like. These salts can be prepared by methods known in the art.
  • the compounds of formula (I) of the present invention may exist in more than one crystalline form, including various crystalline forms and mixtures thereof.
  • solvate refers to a complex of a compound of the invention with a solvent. They either react in a solvent or precipitate out of the solvent or crystallize out. For example, a complex formed with water is referred to as a "hydrate.” Solvates of the compounds of formula (I) are within the scope of the invention.
  • the compounds of the invention may contain one or more chiral centers and exist in different optically active forms.
  • the compound contains a chiral center
  • the compound contains the enantiomer.
  • the invention includes mixtures of the two isomers and isomers, such as racemic mixtures. Enantiomers can be resolved by methods known in the art, such as crystallization and chiral chromatography. When the compound of formula (I) contains more than one chiral center, diastereomers may be present.
  • the present invention includes resolved optically pure specific isomers as well as mixtures of diastereomers. Diastereomers can be resolved by methods known in the art, such as crystallization and preparative chromatography.
  • the invention includes prodrugs of the above compounds.
  • Prodrugs include known amino protecting groups and carboxy protecting groups which are hydrolyzed under physiological conditions or released via an enzymatic reaction to give the parent compound.
  • Specific prodrug preparation methods can be referred to (Saulnier, MG; Frennesson, DB; Deshpande, MS; Hansel, SB and Vysa, DM Bioorg. Med. Chem Lett. 1994, 4, 1985-1990; and Greenwald, RB; Choe, YH; Conover, CD; Shum, K.; Wu, D.; Royzen, MJ Med. Chem. 2000, 43, 475.).
  • the second aspect of the present invention provides a process for producing the above aromatic aminopyrimidine compound, and the compound of the present invention can be easily produced by various synthetic operations, which are well known to those skilled in the art. Specific methods of preparation of these compounds can include, but are not limited to, the procedures described below.
  • the preferred synthetic route of the compound of the formula (I) of the present invention is as follows.
  • the specific compounds in the following examples can be prepared by referring to the following synthetic route, and the steps in the method can be expanded or combined as needed during the specific operation.
  • Step 1-1 Z 1 H at the 4-position of pyrimidine is reacted with an amine compound to be converted into the corresponding amide compound, and the reaction is carried out at a certain temperature using a suitable base and a suitable solvent.
  • the base used may be Not limited to triethylamine.
  • Step 1-2 The chlorine at the 2-position of the pyrimidine is substituted with an amine, and it can be carried out at a certain temperature using a suitable catalyst and a suitable solvent.
  • the catalyst can be, but is not limited to, TFA or p-toluenesulfonic acid.
  • the palladium catalyst used may be, but is not limited to, Pd 2 (dba) 3
  • the ligand used may be, but not limited to, XantPhos
  • the base used may be, but not limited to, cesium carbonate.
  • Step 1-3 conversion of the nitro compound to the corresponding amine compound can be carried out under acidic conditions with a metal (may be, but not limited to, iron powder, zinc powder) or stannous chloride; or under palladium carbon catalysis , hydrogenation reduction.
  • a metal may be, but not limited to, iron powder, zinc powder
  • stannous chloride or under palladium carbon catalysis , hydrogenation reduction.
  • Step 1-4 The amino compound can be condensed to the amide under basic conditions with the corresponding acid chloride, or condensed with the corresponding carboxylic acid in the presence of a condensing agent to form an amide.
  • Step 2-1 Z 1 H at the 4-position of pyrimidine is reacted with an amine compound to be converted into the corresponding amide compound, and the reaction is carried out at a certain temperature using a suitable base and a suitable solvent.
  • the base used may be Not limited to triethylamine.
  • Step 2-2 conversion of the nitro compound to the corresponding amine compound can be carried out under acidic conditions with a metal (may be, but not limited to, iron powder, zinc powder) or stannous chloride; or under palladium carbon catalysis , hydrogenation reduction.
  • a metal may be, but not limited to, iron powder, zinc powder
  • stannous chloride or under palladium carbon catalysis , hydrogenation reduction.
  • Step 2-3 the amine compound can be condensed with the corresponding acid chloride under basic conditions to form an amide, or in condensation The amide is condensed with the corresponding carboxylic acid in the presence of an agent.
  • Step 2-4 The chlorine at the 2-position of the pyrimidine is substituted with an amine, and it can be carried out at a certain temperature using a suitable catalyst and a suitable solvent.
  • the catalyst can be, but is not limited to, TFA or p-toluenesulfonic acid.
  • the palladium catalyst used may be, but is not limited to, Pd 2 (dba) 3
  • the ligand used may be, but not limited to, XantPhos (4,5-bis(diphenylphosphine)-9,9 - dimethylxanthene)
  • the base used may be, but not limited to, cesium carbonate.
  • Step 3-1 The chlorine at the 2-position of the pyrimidine is substituted with an amine, and it can be carried out at a certain temperature using a suitable catalyst and a suitable solvent.
  • the catalyst can be, but is not limited to, TFA or p-toluenesulfonic acid.
  • the palladium catalyst used may be, but is not limited to, Pd 2 (dba) 3
  • the ligand used may be, but not limited to, XantPhos
  • the base used may be, but not limited to, cesium carbonate.
  • Step 3-2 Z 1 H at the 4-position of pyrimidine is reacted with an amine compound to be converted into the corresponding amide compound, and the reaction is carried out at a certain temperature using a suitable base and a suitable solvent.
  • the base used may be Not limited to triethylamine.
  • Step 3-3 conversion of the nitro compound to the corresponding amine compound can be carried out under acidic conditions using a metal (which may be, but not limited to, iron powder, zinc powder) or stannous chloride; or under palladium carbon catalysis , hydrogenation reduction.
  • a metal which may be, but not limited to, iron powder, zinc powder
  • stannous chloride or under palladium carbon catalysis , hydrogenation reduction.
  • Step 3-4 The amino compound can be condensed to the amide under basic conditions with the corresponding acid chloride, or condensed with the corresponding carboxylic acid in the presence of a condensing agent to form an amide.
  • a third aspect of the invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising any one or more of the above compounds, or a pharmaceutically acceptable salt, solvate or prodrug thereof, and further comprising a pharmaceutically acceptable carrier.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a therapeutically effective amount of the above aromatic aminopyrimidine compound, such as the compound of the above formula (I), the compound of the formula (II), or the compound of the formula (III), or the above examples a compound, or a pharmaceutically acceptable salt, stereoisomer, solvate thereof or prodrug thereof; and a pharmaceutically acceptable carrier.
  • a compound of the invention can be administered in a suitable dosage form with one or more pharmaceutical carriers.
  • These dosage forms are suitable for oral, rectal, topical, intraoral, and other parenteral administration (eg, subcutaneous, intramuscular, intravenous, etc.).
  • dosage forms suitable for oral administration include capsules, tablets, granules, and syrups and the like.
  • the compound of the present invention contained in these preparations may be a solid powder or granule; a solution or suspension in an aqueous or non-aqueous liquid; a water-in-oil or oil-in-water emulsion or the like.
  • the above dosage form can be activated by The preparation is made with one or more carriers or excipients via conventional pharmacy methods.
  • the above carriers need to be compatible with the active compound or other excipients.
  • commonly used non-toxic carriers include, but are not limited to, mannitol, lactose, starch, magnesium stearate, cellulose, glucose, sucrose, and the like.
  • Carriers for liquid preparations include water, physiological saline, aqueous dextrose, ethylene glycol, polyethylene glycol, and the like.
  • the active compound can form a solution or suspension with the above carriers.
  • a fourth aspect of the present invention provides the use of the above compound or a pharmaceutically acceptable salt, solvate or prodrug thereof for the preparation of a medicament for regulating EGFR tyrosine kinase activity or for the preparation of a medicament for treating an EGFR-related disease.
  • the EGFR-related disease is cancer, diabetes, immune system disease, neurodegenerative disease or cardiovascular disease.
  • the cancer is non-small cell lung cancer, head and neck cancer, breast cancer, kidney cancer, pancreatic cancer, cervical cancer, esophageal cancer, pancreatic cancer, prostate cancer, bladder cancer, colorectal cancer, ovarian cancer, stomach cancer, brain.
  • Malignant tumors include glioblastoma and the like, or any combination thereof.
  • the above compounds of the present invention or pharmaceutically acceptable salts, solvates or prodrugs thereof may also be used for the preparation of a disease for the treatment of aberrant expression of EGFR, or for the preparation of acquired resistance during treatment with an EGFR modulator. disease.
  • the acquired resistance is resistance caused by a T790 mutation encoded by EGFR exon 20, such as T790M.
  • an EGFR modulator refers to a small molecule tyrosine kinase inhibitor that targets EGFR, such as gefitinib, erlotinib, ectinib, lapatinib or afatinib.
  • a pharmaceutical composition of the present invention comprising a therapeutically effective amount of any one or more of the above compounds, or a pharmaceutically acceptable salt, solvate or prodrug thereof, and one or more selected from the group consisting of the following : Gefitinib, Erlotinib, Ektorinib, Lapatinib, XL647, NVP-AEE-788, ARRY-334543, EKB-569, BIBW2992, HKI272, BMS-690514, CI-1033, Where Detani, PF00299804, WZ4002, cetuximab, trastuzumab, panituzumab, matuzumab, nimotuzumab, zarumumab, pertuzumab, MDX-214, CDX-110, IMC-11F8, Zemab, Her2 vaccine PX 1041, HSP90 inhibitor, CNF2024, tansone, aspironmycin, IPI-504, SNX-5422,
  • “Therapeutically effective amount” means an amount that is functional or active to a human and/or animal and that is acceptable to humans and/or animals.
  • the therapeutically effective amount of the compound of the present invention or a pharmaceutically acceptable salt, solvate or prodrug thereof contained in the pharmaceutical composition or the pharmaceutical composition of the present invention is preferably 0.1 mg to 5 g/kg (body weight). .
  • the pharmaceutical composition can be used to treat diseases in which EGFR is abnormally expressed, such as cancer, diabetes, immune system diseases, neurodegenerative diseases or cardiovascular diseases.
  • the acquired drug-resistant disease is caused by a T790 mutation encoded by EGFR exon 20 or by a T790 mutation encoded by EGFR exon 20.
  • the EGFR exon 20 encodes a T790 of T790M.
  • the compounds of the invention may be combined with other drugs in certain diseases to achieve the desired therapeutic effect.
  • An example of a joint application is to treat advanced NSCLC.
  • a therapeutically effective amount of the formula of the invention Compounds indicated by I are used in combination with mTOR inhibitors (eg, rapamycin); or in combination with Met inhibitors (including Met antibody MetMAb and Met small molecule inhibitor PF02341066); or in combination with IGF1R inhibitors (eg, OSI-) 906); or in combination with a heat shock protein inhibitor.
  • the invention discloses a method for preparing a compound and a compound, a drug composition and a treatment plan, and those skilled in the art can learn from the contents of the paper and appropriately improve the process parameters. It is to be understood that all such alternatives and modifications are obvious to those skilled in the art and are considered to be included in the present invention.
  • the products, methods and applications of the present invention have been described in terms of the preferred embodiments, and it is obvious that the methods and applications described herein may be modified or appropriately modified and combined without departing from the spirit, scope and scope of the invention. The techniques of the present invention are implemented and applied.
  • the inventors unexpectedly discovered an aromatic aminopyrimidine compound, which is a selective inhibitor of EGFR mutations and has the advantage of low toxicity.
  • the arylaminopyrimidine compound provided by the present invention is a selective inhibitor of EGFR mutation, and the enzyme and cell experiments in vitro inhibit the proliferation of L858R-T790M enzyme and double mutant cell line H1975 at nanomolar concentration.
  • the inhibition of wild-type EGFR and cell line A431 was relatively weak. Its mutation selectivity greatly reduces the side effects caused by inhibition of wild-type EGFR, and is suitable for the case of secondary drug resistance in the current EGFR-TKI treatment. It is the first and second generation EGFR tyrosine kinase inhibitors. An ideal alternative.
  • the structure of the compound is determined by nuclear magnetic resonance ( 1 H NMR) and/or liquid chromatography-mass spectrometry (LC-MS).
  • the 1 H NMR shift ( ⁇ ) is given in parts per million (ppm).
  • 1 H NMR was measured using a Bruker AVANCE-400 nuclear magnetic instrument with an internal standard of tetramethylsilane (TMS).
  • LC-MS Product purity determination was determined by LC-MS.
  • the LC-MS was measured using an Agilent 1200 HPLC System/6140 MS LC/MS mass spectrometer (manufacturer: Agilent), column Waters X-Bridge, 150 x 4.6 mm, 3.5 ⁇ m.
  • Preparative high performance liquid chromatography with Waters PHW007, column XBridge C18, 4.6*150 mm, 3.5 um.
  • ISCO Combiflash-Rf75 or Rf200 automatic column analyzer Use ISCO Combiflash-Rf75 or Rf200 automatic column analyzer, Agela 4g, 12g, 20g, 40g, 80g, 120g disposable silica gel column.
  • the specifications for the board are 0.4mm to 0.5mm.
  • Silica gel is generally used as a carrier for Yantai Huanghai silica gel 200-300 mesh silica gel.
  • the basic alumina column is generally used as a carrier by using FCP 200-300 mesh basic alumina as a carrier.
  • the known starting materials of the present invention can be synthesized by or according to methods known in the art, or can be used in ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, Accela ChemBio Inc and Dari Chemical Products and other companies to buy.
  • the reactions were all carried out under a nitrogen or argon atmosphere.
  • An argon atmosphere or a nitrogen atmosphere means that the reaction flask is connected to an argon or nitrogen balloon having a volume of about 1 L.
  • the hydrogen atmosphere means that the reaction flask is connected to a hydrogen balloon of about 1 L volume, a Parker-20H type hydrogen generator.
  • the hydrogenation reaction is usually evacuated, charged with hydrogen, and operated three times.
  • the solution means an aqueous solution.
  • the progress of the reaction in the examples was monitored by thin layer chromatography (TLC).
  • TLC thin layer chromatography
  • the system of the developing solvent used in the reaction was selected from the group consisting of dichloromethane and methanol systems, n-hexane and ethyl acetate systems, petroleum ether and ethyl acetate systems. And one or more of the acetone systems; the volume ratio of the solvent is adjusted depending on the polarity of the compound.
  • the system of the eluent for column chromatography or the system of the developer for thin layer chromatography may be selected from the group consisting of: A, dichloromethane and methanol systems, B, n-hexane and ethyl acetate systems and C, dichloro One or more of the methane and acetone systems, the volume ratio of the solvent is adjusted depending on the polarity of the compound, and may be adjusted by adding a small amount of an alkaline reagent such as triethylamine or an acidic reagent such as acetic acid.
  • DMF dimethylformamide
  • DMSO dimethyl sulfoxide
  • THF tetrahydrofuran
  • DIEA N, N-diisopropylethylamine
  • EA ethyl acetate
  • PE petroleum ether.
  • room temperature refers to about 25 °C.
  • Step 1 tert-Butyl 4-oxopiperidine-1-carboxylic acid tert-butyl ester (5 g, 29.2 mmol) and sodium cyanoborohydride (2.8 g, 43.8 mmol) were placed in a 500 mL single-mouth reaction flask, and methanol was added. After 200 ml was dissolved, 1-methylpiperazine (3.2 g, 32.1 mmol) and glacial acetic acid (1 ml) were added and the reaction was kept at room temperature overnight. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction mixture was extracted three times with ethyl acetate/water.
  • Step 2 4-(4-methylpiperazin-1-yl)piperidine-1-carboxylic acid tert-butyl ester (6.5 g, 22.9 mmol) was added to 100 ml of dioxane hydrochloride solution at room temperature. Stir vigorously for 4 h. After completion of the reaction, the reaction mixture was concentrated under reduced pressure to give 1-methyl-4-(piperidin-4-yl)piperazine (4.2 g, 100%). MS m/z (ESI): 184.1 [M+H] +
  • Step 3 1-Methyl-4-(piperidin-4-yl)piperazine (4.2 g, 22.9 mmol), 4-fluoro-2-methoxy-1-nitrobenzene (4.69 g, 25.2 mmol And potassium carbonate (9.48 g, 68.7 mmol) was placed in a 250 mL one-mouth reaction flask, DMF (100 mL) was added to partially dissolve the substrate, and the reaction system was maintained at 100 ° C for 4 h. TLC was used to detect the progress of the reaction.
  • Step 4 Palladium on carbon (0.6 g, 10% wt) was added to 1-(1-(3-methoxy-4-nitrophenyl)piperidin-4- at room temperature The solution of 4-methylpiperazine (6 g, 17.9 mmol) in 200 ml of methanol was stirred vigorously under a hydrogen atmosphere at room temperature for 20 h. After completion of the reaction, the palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give the product 2-methoxy-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl) phenylamine. In the next step. MS m/z (ESI): 305.1 [M+H]+
  • Step 1 2-Bromo-5-nitropyridine (2 g, 10 mmol) was added to 10 mL of morpholine solution at room temperature and stirred vigorously at room temperature for 4 h. After the reaction was completed, a yellow solid precipitated. After filtration, the yellow solid was washed with EtOAc (EtOAc) (EtOAc) MS m/z (ESI): 210.1 [M+H] +
  • Step 2 Palladium on carbon (100 mg, 10% by weight) was added to a solution of 4-(5-nitropyridin-2-yl)morpholine (1 g, 9.1 mmol) in 60 ml of methanol at room temperature under a hydrogen atmosphere. Stir vigorously for 20 h. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give the product, 3-amino-6-morpholinylpyridine, which was directly used for the next reaction. MS m/z (ESI): 180.1 [M+H] +
  • Step 1 4-terdopiperidine-1-carboxylic acid tert-butyl ester (5 g, 29.2 mmol) and sodium cyanoborohydride (2.8 g, 43.8 mmol) were placed in a 500 mL single-mouth reaction flask, and dissolved in 200 ml of methanol. Then morpholine (2.8 g, 32.1 mmol) and glacial acetic acid (1 ml) were added and the reaction was kept at room temperature overnight. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction mixture was extracted three times with ethyl acetate/water.
  • Step 2 4-morpholinopiperidine-1-carboxylic acid tert-butyl ester (6 g, 22.2 mmol) was added to 100 ml of dioxane hydrochloride solution and stirred vigorously at room temperature for 4 h. After completion of the reaction, the reaction mixture was concentrated under reduced pressure to give 4-(piperidin-4-yl)morpholine (3.8 g, 100%). MS m/z (ESI): 171.1 [M+H] +
  • Step 3 4-(piperidin-4-yl)morpholine (3.8 g, 22.2 mmol), 4-fluoro-2-methoxy-1-nitrobenzene (4.18 g, 24.42 mmol) and potassium carbonate ( 9.19 g, 66.6 mmol) was placed in a 250 mL single-mouth reaction flask, DMF (50 mL) was added to partially dissolve the substrate, and the reaction was maintained at 100 ° C for 4 h. The progress of the reaction was checked by TLC.
  • Step 4 Palladium on carbon (0.58 g, 10% wt) was added to 4-(1-(3-methoxy-4-nitrophenyl)piperidin-4-yl)morpholine (5.8 g) at room temperature. , 18.1 mmol) in 200 ml of methanol, vigorously stirred at room temperature under a hydrogen atmosphere for 20 h. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give the product, 2-methoxy-4-(4-morpholino-1-yl)aniline, which was directly used for the next reaction. MS m/z (ESI): 292.2 [M+H] +
  • Step 1 N,N-Dimethylpiperidin-4-amine (5 g, 39 mmol) and 4-fluoro-2-methoxy-1-nitrobenzene (10.95 g, 58.6 mmol) and potassium carbonate (16.15) g, 117 mmol) was placed in a 500 mL single-mouth reaction flask, DMF (150 mL) was added to partially dissolve the substrate, and the reaction was kept at 100 ° C for 4 h. TLC was used to detect the progress of the reaction.
  • Step 2 Palladium on carbon (0.75 g, 10% wt) was added to 1-(3-methoxy-4-nitrophenyl)-N,N-dimethylpiperidin-4-amine at room temperature ( 7.5 g, 26.8 mmol) in 200 ml of methanol was stirred vigorously under a hydrogen atmosphere at room temperature for 20 h. After completion of the reaction, the palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give 1-(4-amino-3-methoxyphenyl)-N,N-dimethylpiperidin-4-amine as directly.
  • Step 1 4-Fluoro-2-methoxy-1-nitrobenzene (4.4 g, 25.7 mmol) and cesium carbonate (18.1 g, 51.4 mmol) were placed in a 250 mL single-mouth reaction flask, and DMF (100 mL) was added. The substrate is partially dissolved. Then morpholine (3.3 mL, 38.6 mmol) was added and the reaction was maintained at 100 ° C for 2 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction mixture was extracted three times with ethyl acetate/water.
  • Step 2 Palladium on carbon (500 mg, 10% by weight) was added to a solution of 4-(3-methoxy-4-nitrophenyl)morpholine in 100 ml of methanol at room temperature, and the reaction was vigorously stirred at room temperature under a hydrogen atmosphere. overnight. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give the product, 2-methoxy-4-morpholine aniline (4.5 g, yield: 84%). MS m/z (ESI): 209.1 [M+H] +
  • Step 1 5-Bromo-2-nitropyridine (1 g, 5 mmol) was added to 10 mL of morpholine solution at room temperature and stirred vigorously at room temperature for 4 h. After the reaction was completed, a yellow solid precipitated. After filtration, the yellow solid was washed with 50 ml of petroleum ether to afford 4-(6-nitropyridin-3-yl)morpholine (530 mg, 53%). MS m/z (ESI): 210.1 [M+H] +
  • Step 2 Palladium on carbon (60 mg, 10% by weight) was added to a solution of 4-(6-nitropyridin-3-yl)morpholine (600 mg, 2.86 mmol) in 30 ml of methanol at room temperature under a hydrogen atmosphere. Stir vigorously for 20 h. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give the product 2-amino-5-(4-morpholinyl)pyridine, which was directly used for the next reaction.
  • Step 1 4-Fluoro-2-methoxy-1-nitrobenzene (1.9 g, 11.1 mmol) and potassium carbonate (4.6 g, 33.3 mmol) were placed in a 60 mL single-mouth reaction flask, and DMF (60 mL) was added. The substrate is partially dissolved. Piperidine (950 mg, 11.1 mmol) was then added and the reaction was maintained at 100 ° C for 6 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction mixture was extracted three times with ethyl acetate/water. The organic layer was separated, washed with water and brine, dried over anhydrous sodium sulfate . Purification by Combi-Flash column chromatography gave 1-(3-methoxy-4-nitrophenyl)piperidine (2.2 g, 85%). MS m/z (ESI): 237.1 [M+H] +
  • Step 2 Palladium on carbon (200 mg, 10% by weight) was added to a solution of 1-(3-methoxy-4-nitrophenyl)piperidine (2 g, 8.44 mmol) in 60 ml of methanol at room temperature. Stir vigorously for 20 h at room temperature under an atmosphere. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give the product, 2-methoxy-4-(piperidin-1-yl)aniline, which was directly used for the next reaction.
  • Step 1 4-Fluoro-2-methoxy-1-nitrobenzene (5 g, 29.2 mmol) and potassium carbonate (12.1 g, 87.7 mmol) were placed in a 500 mL single-mouth reaction flask, and DMF (100 mL) was added. The substrate partially dissolved. Then 1-tert-butoxycarbonylpiperazine (8.2 g, 43.8 mmol) was added and the reaction was maintained at 100 ° C for 4 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction mixture was extracted three times with ethyl acetate/water.
  • Step 2 4-(3-methoxy-4-nitrophenyl)piperazine-1-carboxylic acid tert-butyl ester (8 g, 23.7 mmol) was added to 100 ml of dioxane hydrochloride solution at room temperature. Stir vigorously for 1 h at room temperature. After completion of the reaction, the reaction mixture was evaporated tolulululululululululululululu MS m/z (ESI): 238.1 [M+H] +
  • Step 3 1-(3-Methoxy-4-nitrophenyl)piperazine (500 mg, 2.1 mmol) and potassium carbonate (900 mg, 6.3 mmol) were placed in a 25 mL sealed tube and acetonitrile was added. 6 mL) partially dissolved the substrate. Then 1-bromo-2-fluoroethane (290 mg, 2.3 mmol) was added and the reaction was maintained at 80 ° C for 7 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the insoluble material was filtered off and concentrated under reduced pressure to give 1-(2-fluoroethyl)-4-(3-methoxy-4-nitrophenyl)per Azine (350 mg, 59%). MS m/z (ESI): 284.1 [M+H] +
  • Step 4 Palladium on carbon (35 mg, 10% wt) was added to 1-(2-fluoroethyl)-4-(3-methoxy-4-nitrophenyl)piperazine (350 mg, 1.24) at room temperature. In a 50 ml methanol solution of mmol), vigorously stirred at room temperature under a hydrogen atmosphere for 20 h. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give 4-(4-(2-fluoroethyl)piperazin-1-yl)-2-methoxyaniline as the next. MS m/z (ESI): 254.1 [M+H] +
  • Step 1 Diethylamine (1 g, 13.7 mmol) and 4-fluoro-2-methoxy-1-nitrobenzene (3.84 g, 20.5 mmol) and potassium carbonate (5.67 g, 41.1 mmol) were placed in 500 mL In a single-mouth reaction flask, DMF (50 mL) was added to partially dissolve the substrate, and the reaction system was maintained at 100 ° C for 4 h. The progress of the reaction was detected by TLC. After the substrate was completely reacted, the insoluble matter was filtered off, and concentrated under reduced pressure to give a yellow solid which was purified by Combi-Flash column chromatography to obtain N,N-diethyl-3-methoxy-4-nitrate. Aniline (1.8 g, 58%). MS m/z (ESI): 225.1 [M+H] +
  • Step 2 Palladium on carbon (0.22 g, 10% by weight) was added to a solution of N,N-diethyl-3-methoxy-4-nitroaniline (1.8 g, 8.04 mmol) in 200 ml of methanol Stir vigorously for 20 h at room temperature under a hydrogen atmosphere. After completion of the reaction, the palladium-carbon was filtered off, the filtrate was concentrated under reduced pressure, the product N 1, N 1 - diethyl-3-methoxy-1,4-diamine, was used directly in the next reaction. MS m/z (ESI): 195.1 [M+H] +
  • Step 1 4-terdopiperidine-1-carboxylic acid tert-butyl ester (5 g, 29.2 mmol) and sodium cyanoborohydride (2.8 g, 43.8 mmol) were placed in a 500 mL single-mouth reaction flask, and dissolved in 200 ml of methanol. Then diethylamine (2.3 g, 32.1 mmol) and glacial acetic acid (1 ml) were added and the reaction was kept at room temperature overnight. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction mixture was extracted three times with ethyl acetate/water.
  • Step 2 tert-Butyl 4-(diethylamino)piperidine-l-carboxylate (5.2 g, 20.2 mmol) was added to 100 ml of dioxane hydrochloride solution and stirred vigorously at room temperature for 4 h. After completion of the reaction, the reaction mixture was concentrated under reduced pressure to give N,N-diethylpiperidine-4-amine (3.1 g, 100%). MS m/z (ESI): 157.1 [M+H] +
  • Step 3 N,N-Diethylpiperidin-4-amine (3.1 g, 20.2 mmol) 4-fluoro-2-methoxy-1-nitrobenzene (4.53 g, 24.2 mmol) and potassium carbonate ( 8.36 g, 60.6 mmol) was placed in a 250 mL single-mouth reaction flask, DMF (100 mL) was added to partially dissolve the substrate, and the reaction was maintained at 100 ° C for 4 h. TLC was used to detect the progress of the reaction.
  • Step 4 Palladium on carbon (0.5 g, 10% by weight) was added to N,N-diethyl-1-(3-methoxy-4-nitrophenyl)piperidin-4-amine (at room temperature) 5 g, 17.9 mmol) in 200 ml of methanol was stirred vigorously under a hydrogen atmosphere at room temperature for 20 h. After completion of the reaction, the palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give 1-(4-amino-3-methoxyphenyl)-N,N-diethylpiperidin-4-amine as directly.
  • Step 1 4-Fluoro-2-methoxy-1-nitrobenzene (5 g, 29.2 mmol) and potassium carbonate (12.1 g, 87.7 mmol) were placed in a 500 mL single-mouth reaction flask, and DMF (100 mL) was added. The substrate partially dissolved. Then 1-tert-butoxycarbonylpiperazine (8.2 g, 43.8 mmol) was added and the reaction was maintained at 100 ° C for 4 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction mixture was extracted three times with ethyl acetate/water.
  • Step 2 4-(3-methoxy-4-nitrophenyl)piperazine-1-carboxylic acid tert-butyl ester (8 g, 23.7 mmol) was added to 100 ml of dioxane hydrochloride solution at room temperature. Stir vigorously for 1 h at room temperature. After completion of the reaction, the reaction mixture was evaporated tolulululululululululululululu MS m/z (ESI): 238.1 [M+H] +
  • Step 3 1-(3-Methoxy-4-nitrophenyl)piperazine (500 mg, 2.1 mmol) and potassium carbonate (900 mg, 6.3 mmol) were placed in a 50 mL single-mouth reaction flask, and acetonitrile (6 mL) was added. ) partially dissolve the substrate. Then bromoethane (253 mg, 2.3 mmol) was added and the reaction was maintained at 80 ° C for 7 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the insoluble material was filtered off and concentrated under reduced pressure to give a yellow solid 1-ethyl-4-(3-methoxy-4-nitrophenyl)piperazine (400 mg, 72) %). MS m/z (ESI): 266.3 [M+H] +
  • Step 4 Palladium on carbon (30 mg, 10% by weight) was added to a solution of 1-ethyl-4-(3-methoxy-4-nitrophenyl)piperazine (270 mg, 1 mmol) in 50 ml of methanol The mixture was vigorously stirred at room temperature for 20 h under a hydrogen atmosphere. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give 4-(4-ethylpiperazin-1-yl)-2-methoxyaniline as the next. MS m/z (ESI): 236.3 [M+H] +
  • Step 1 N-methylpiperazine (30 ml) was added to a solution of 5-bromo-2-nitropyridine (5 g, 21.6 mmol) in EtOAc (EtOAc) After the substrate was completely reacted, it was diluted with water and extracted with a dichloromethane/water system. The organic layer was separated, washed with water and brine, dried over anhydrous sodium sulfate 4-(6-Nitropyridin-3-yl)piperazine (2.5 g, 52%). MS m/z (ESI): 223.1 [M+H] +
  • Step 2 Palladium on carbon (250 mg, 10% wt) was added to a solution of 1-methyl-4-(6-nitropyridin-3-yl)piperazine (2.5 g, 11.2 mmol) in 60 mL of MeOH. Stir vigorously for 20 h at room temperature under a hydrogen atmosphere. After completion of the reaction, palladium carbon was filtered off, and the filtrate was concentrated under reduced pressure to give 5-(4-methylpiperazin-1-yl)pyridin-2-amine as the next. MS m/z (ESI): 193.1 [M+H] +
  • Step 1 4-Nitro-3-methoxybenzoic acid (5 g, 25.4 mmol) was added to 50 ml of SOCI 2 and 4 drops of DMF was added dropwise and vigorously stirred at 90 ° C for 3 h. The progress of the reaction was checked by TLC. After the substrate was completely reacted, the yellow solid 3-methoxy-4-nitrobenzoyl chloride obtained after concentration of the product was directly used for the next reaction.
  • Step 2 DIPEA (4 mL, 25.1 mmol) and diethylamine (2.7 mL, 25.1 mmol) were added to 3-methoxy-4-nitrobenzoyl chloride (5.4 g, 25.1 mmol) at 0 °C. Stir vigorously for 3 h at room temperature in 50 ml of THF. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, it was diluted with water, adjusted to pH, and extracted three times with ethyl acetate/water system. The organic layer was separated, washed with water and saturated brine, dried over anhydrous sodium sulfate. Concentration under reduced pressure gave 6.6 g of crude material. Purification by Combi-Flash column chromatography gave compound N,N-diethyl-3-methoxy-4-nitrobenzamide (5.5 g, 86%). MS m/z (ESI): 253.1 [M+H] +
  • Step 3 Pd/C (200 mg) was added to a solution of N,N-diethyl-3-methoxy-4-nitrobenzamide (1 g, 3.97 mmol) in 50 ml of methanol. Stir vigorously for 16 h at room temperature under a hydrogen atmosphere. After completion of the reaction, the mixture was filtered, and then evaporated, mjjjjjj MS m/z (ESI): 223.2 [M+H] +
  • Step 1 4-Nitro-3-methoxybenzoic acid (5 g, 25.4 mmol) was added to 50 ml of SOCI 2 and 4 drops of DMF was added dropwise and vigorously stirred at 90 ° C for 3 h. The progress of the reaction was checked by TLC. After the substrate was completely reacted, the yellow solid 3-methoxy-4-nitrobenzoyl chloride obtained after concentration of the product was directly used for the next reaction.
  • Step 2 DIPEA (4 mL, 25.1 mmol) and morpholine (2.2 g, 25.1 mmol) were added to 50 ml of 3-methoxy-4-nitrobenzoyl chloride (5.5 g, 25.4 mmol) at 0 °C. The mixture was stirred vigorously for 3 h at room temperature in THF. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, it was diluted with water, adjusted to pH, and extracted three times with ethyl acetate/water system. The organic layer was separated, washed with water and saturated brine, dried over anhydrous sodium sulfate. Concentration under reduced pressure gave the compound (3-methoxy-4-nitrophenyl) (morpholino)methanone (6.6 g, 98%). MS m/z (ESI): 267.1 [M+H] +
  • Step 3 Pd/C (200 mg) was added to a solution of (3-methoxy-4-nitrophenyl)(morpholino)methanone (1 g, 3.76 mmol) in 50 ml of methanol. At room temperature, stirred vigorously for 16h H 2 atmosphere. After completion of the reaction, the mixture was filtered and evaporated tolulululululululululululululululululululu MS m/z (ESI): 237.2 [M+H] +
  • Step 1 4-Nitro-3-methoxybenzoic acid (3 g, 15.2 mmol) was added to 50 ml of SOCl 2 and 4 drops of DMF was added dropwise and stirred vigorously at 90 ° C for 3 h. The progress of the reaction was checked by TLC. After the substrate was completely reacted, the yellow solid 3-methoxy-4-nitrobenzoyl chloride obtained after concentration of the product was directly used for the next reaction.
  • Step 2 Add N-methylpiperazine (1.5 g, 15.2 mmol) to a solution of 3-methoxy-4-nitrobenzoyl chloride (3.3 g, 15.2 mmol) in 50 mL of dichloromethane at 0 °. Stir vigorously for 3 h at room temperature. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, it was diluted with water, adjusted to pH, and extracted three times with ethyl acetate/water system. The organic layer was separated, washed with water and saturated brine, dried over anhydrous sodium sulfate. Concentration under reduced pressure gave (3-methoxy-4-nitrophenyl)(4-methylpiperazin-1-yl)methanone (3.0 g, 71%). MS m/z (ESI): 280.0 [M+H] +
  • Step 3 Add Pd/C (150 mg) to (3-methoxy-4-nitrophenyl)(4-methylpiperazin-1-yl)methanone (1 g, 3.58 mmol) in 50 mL MeOH in. Stir vigorously for 16 h at room temperature under a hydrogen atmosphere. After the TLC detection reaction was completed, the mixture was filtered, and the filtrate was concentrated to give (4-amino-3-methoxyphenyl)(4-methylpiperazin-1-yl)methanone (780 mg, 87%). Next step.
  • Triethylamine (TEA) (11 g, 110 mmol) and 3-nitroaniline (5.6 g, 40.3 mmol) were added to a solution of 2-chloro-4-pyrimidine isocyanate (5.7 g, 36.6 mmol) in 200 ml of THF. Stir vigorously for 20 h at room temperature. After completion of the reaction, 500 ml of water was added to the reaction system, and a yellow solid precipitated. The yellow solid was washed with ethyl acetate (500 mL) and then EtOAc (EtOAc) Yield: 30%, purity: 86%, MS m/z (ESI): 294.1 [M+H] +
  • Step 4 1-(3-Aminophenyl)-3-(2-((2-methoxy-4-(4-methylpiperazin-1-yl)phenyl)amino)pyrimidin-4-yl) Urea synthesis
  • Step 5 N-(3-(3-(2-(2-methoxy-4-(4-methylpiperazin-1-yl)phenyl)amino)pyrimidin-4-yl)ureido) Synthesis of phenyl)acrylamide (cpd-1)
  • Step 2 Synthesis of 1-(2-((4-(4-methylpiperazin-1-yl)phenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea
  • Step 3 N-(3-(3-(2-(4-(4-methylpiperazin-1-yl)phenyl)amino)pyrimidin-4-yl)ureido)phenyl)acrylamide ( Synthesis of cpd-2)
  • Step 2 Synthesis of 1-(2-((5-methoxypyridin-2-yl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea
  • Step 3 Synthesis of N-(3-(3-(2-((5-methoxypyridin-2-yl)amino)pyrimidin-4-yl)ureido)phenyl)acrylamide (cpd-3)
  • Triethylamine (172 mg, 1.7 mmol) was added to 1-(2-((5-methoxypyridin-2-yl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea ( 200 mg, 0.56 mmol) of a mixed solution of 2 ml of THF and 2 ml of DMF was vigorously stirred at 0 °C.
  • Acryloyl chloride (62 mg, 0.68 mmol) was slowly added dropwise to the reaction mixture at 0 °C. Stir at 0 ° C for 4 h. TLC was used to detect the progress of the reaction.
  • Step 2 1-(2-((3-Chloro-4-((3-fluorobenzyl)oxy)phenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea synthesis
  • Step 1 1 - (2-((2-Methoxy-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl) Synthesis of -3-(3-nitrophenyl)urea
  • Step 2 1-(2-((2-Methoxy-4-phenyl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl Synthesis of -3-(3-aminophenyl)urea
  • Step 3 N-(3-(3-(2-(2-methoxy-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino) Synthesis of pyrimidin-4-yl)ureido)phenyl)acrylamide (cpd-5)
  • Step 2 Synthesis of 1-(2-((6-morpholinopyridin-3-yl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea
  • Step 3 Synthesis of N-(3-(3-(2-((6-morpholinopyridin-3-yl)amino)pyrimidin-4-yl)ureido)phenyl)acrylamide (cpd-6)
  • Triethylamine (90 mg, 0.9 mmol) was added to 1-(2-((6-morpholinopyridin-3-yl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea ( 120 mg, 0.30 mmol) in 5 ml of THF was stirred vigorously at 0 °C.
  • Acryloyl chloride (27 mg, 0.30 mmol) was slowly added dropwise to the reaction mixture at 0 °C. Stir at 0 ° C for 4 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, it was diluted with water and extracted three times with ethyl acetate/water.
  • Step 2 Synthesis of 1-(2-((2,4-dimethoxyphenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea
  • Step 3 Synthesis of N-(3-(3-(2-((2,4-dimethoxyphenyl)amino)pyrimidin-4-yl)ureido)phenyl)acrylamide (cpd-7)
  • Triethylamine (100 mg, 0.9 mmol) was added to 1-(2-((2,4-dimethoxyphenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea ( 100 mg, 0.3 mmol) in 5 ml of THF was stirred vigorously at 0 °C.
  • Acryloyl chloride (30 mg, 0.3 mmol) was slowly added dropwise to the reaction mixture at 0 °C. Stir at 0 ° C for 4 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, it was diluted with water and extracted three times with ethyl acetate/water.
  • Step 2 Synthesis of 1-(2-((3-chloro-4-fluorophenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea
  • Step 3 Synthesis of N-(3-(3-(2-(3-chloro-4-fluorophenyl)amino)pyrimidin-4-yl)ureido)phenyl)acrylamide (cpd-8)
  • Step 1 1-(2-((2-Methoxy-4-(4-morpholino-1-yl)phenyl)amino)pyrimidin-4-yl)-3-(3-nitrophenyl) Urea synthesis
  • Step 2 1-(2-((2-Methoxy-4-(4-morpholino-1-yl)phenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl) Synthesis of urea
  • Step 3 N-(3-(3-(2-(2-methoxy-4-(4-morpholinipridin-1-yl)phenyl)amino)pyrimidin-4-yl)ureido) Synthesis of phenyl)acrylamide (cpd-9)
  • Step 1 1-(2-((4-(4-(Dimethylamino)piperidin-1-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl)-3-(3) Synthesis of -nitrophenyl)urea
  • Step 2 1-(2-((4-(4-(Dimethylamino)piperidin-1-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl)-3-(3) Synthesis of -aminophenyl)urea
  • Step 3 N-(3-(3-(2-(4-(4-(dimethylamino)piperidin-1-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl) Synthesis of ureido)phenyl)acrylamide (cpd-10)
  • Step 2 Synthesis of 1-(2-((4-(piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea
  • Step 2 Synthesis of 1-(2-((4-(dimethylamino)phenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea
  • Step 3 Synthesis of N-(3-(3-(2-((4-(dimethylamino)phenyl)amino)pyrimidin-4-yl)ureido)phenyl)acrylamide (cpd-12)
  • Triethylamine (163 mg, 1.61 mmol) was added to 1-(2-((4-(dimethylamino)phenyl)amino)pyrimidin-4-yl)-3-(3-aminophenyl)urea ( A mixed solution of 195 mg, 0.54 mmol) of 2 ml of THF and 2 ml of DMF was vigorously stirred at 0 °C. Acryloyl chloride (58 mg, 0.65 mmol) was slowly added dropwise to the reaction mixture at 0 °C. Stir at 0 ° C for 4 h. TLC was used to detect the progress of the reaction.
  • 1-(2-Chloropyrimidin-4-yl)-3-(3-nitrophenyl)urea (6.0 g, 20 mmol) was placed in a 250 mL single-mouth reaction flask and mixed with THF/water (75 mL / 50 mL). The solution dissolves the substrate. Ammonium chloride (5.3 g, 100 mmol) and reduced iron powder (8.9 g, 160 mmol) were successively added to the stirred reaction flask at room temperature, and then the reaction was heated to 65 ° C and kept stirring for 5 h. The progress of the reaction was checked by TLC. After the substrate was completely reacted, excess iron powder was removed by filtration, and the filter cake was rinsed three times with ethyl acetate.
  • Step 3 Synthesis of N-(3-(3-(2-((5-morpholinopyridin-2-yl)amino)pyrimidin-4-yl)ureido)phenyl)acrylamide (cpd-13)
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide (200 mg, 0.62 mmol) and 2-methoxy-4-morpholinoaniline (131 mg, 0.62) Methyl) was placed in a 50 mL one-mouth reaction flask, and n-butanol (10 mL) was added to partially dissolve the substrate.
  • Trifluoroacetic acid (215 mg, 1.89 mmol) was then added and the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.62 mmol
  • 4-(4-(2-fluoroethyl)piperazine-1 2-Methoxy aniline 157 mg, 0.62 mmol
  • Trifluoroacetic acid 215 mg, 1.89 mmol
  • the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.62 mmol
  • N,N-diethyl-3-methoxybenzene-1 4-Diamine 157 mg, 0.62 mmol
  • Trifluoroacetic acid 215 mg, 1.89 mmol
  • the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.62 mmol
  • 1-(4-amino-3-methoxyphenyl)- N,N-Diethylpiperidin-4-amine 157 mg, 0.62 mmol
  • Trifluoroacetic acid 215 mg, 1.89 mmol
  • the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.62 mmol
  • 4-methylaniline 67 mg, 0.62 mmol
  • n-butanol 10 mL was added to partially dissolve the substrate.
  • Trifluoroacetic acid (215 mg, 1.89 mmol) was then added and the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.62 mmol
  • 5-(4-methylpiperazin-1-yl)pyridine- 2-Amine 69 mg, 0.62 mmol
  • Trifluoroacetic acid 215 mg, 1.89 mmol
  • the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.62 mmol
  • p-fluoroaniline 69 mg, 0.62 mmol
  • Trifluoroacetic acid 215 mg, 1.89 mmol
  • the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.62 mmol
  • p-ethoxyaniline 85 mg, 0.62 mmol
  • n-butanol 10 mL was added to partially dissolve the substrate.
  • Trifluoroacetic acid (215 mg, 1.89 mmol) was then added and the reaction was maintained at 110 ° C for 3 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • N-(3-(3-(2-chloropyrimidin-4-yl)ureido)phenyl)acrylamide 200 mg, 0.63 mmol
  • 4-morpholinylaniline 112 mg, 0.62 mmol
  • Trifluoroacetic acid 215 mg, 1.89 mmol
  • the reaction was maintained at 110 ° C for 6 h. After the reaction is completed, it is cooled to room temperature. After the substrate is completely reacted, the reaction mixture is extracted three times with ethyl acetate/water.
  • Step 2 Synthesis of N-(4-(4-(dimethylamino)piperidin-1-yl)-2-methoxyphenyl)-N-4-methylpyrimidine-2,4-diamine
  • Step 3 1-(2-(4-(4-(Dimethylamino)piperidin-1-yl)-2-methoxyphenylamino)pyrimidin-4-yl)-1-methyl-3 Synthesis of -(3-nitrophenyl)urea
  • Step 4 1-(2-(4-(4-(Dimethylamino)piperidin-1-yl)-2-methoxyphenylamino)pyrimidin-4-yl)-1-methyl-3 Synthesis of -(3-aminophenyl)urea
  • Step 5 N-(3-(3-(2-(4-(4-(dimethylamino)piperidin-1-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl) Synthesis of -3-methylureido)phenyl)acrylamide (cpd-32)
  • Step 2 Synthesis of tert-butyl 1-((2-chloropyrimidin-4-yl)carbamoyl)piperidin-3-ylcarbamate
  • Triethylamine (10 g, 97 mmol) and 3-tert-butoxycarbonylaminopiperidine (7.1 g, 35.4 mmol) were added to 100 ml of 2-chloro-4-isocyanate (5.0 g, 32.2 mmol) at room temperature. Stir vigorously at room temperature for 18 h in THF. After the completion of the reaction, 150 ml of ethyl acetate was added to the reaction mixture, and the organic layer was washed with EtOAc (EtOAc) 3.5g). Yield: 31%, purity: 90%, MS m/z (ESI): 356.2 [M+H] + .
  • Step 5 3-acrylamido-N-(2-(2-methoxy-4-(4-morpholino-1-yl)phenylamino)pyrimidin-4-yl)piperidine-1-methyl Synthesis of amide (cpd-33)
  • Example 36 3-acrylamido-N-(2-(4-((2-(dimethylamino)ethyl)(methyl)amino)-2-methoxyphenylamino)pyrimidine-4 Of -piperidine-1-carboxamide (cpd-36)
  • Diethyl malonate (6.5.0 g, 40.0 mmol) was placed in a 500 mL single-mouth reaction flask, and a mixed solution of anhydrous toluene (100 mL) was added to dissolve.
  • Sodium hydride (4.4 g, 80.0 mmol) was added to the stirred reaction flask at room temperature, then the reaction was stirred at room temperature for 0.5 h.
  • 2,4-dichloropyrimidine (5.0 g, 34.0 mmol) was added to the reaction system, and after the reaction system was replaced with nitrogen three times, Pd 2 (dba) 3 (600 mg, 0.68 mmol) and P (t-Bu) were added.
  • Diethyl 2-(2-chloropyrimidin-4-yl)malonate (2.9 g, 10.0 mmol) was placed in a 250 mL single-mouth reaction flask, and a mixed solution of DMSO/H 2 O (100 mL / 2 mL) was added to make a bottom. The substance dissolves.
  • Sodium chloride 600 mg, 10.0 mmol was sequentially added to the stirred reaction flask at room temperature, then the reaction was heated to 140 ° C and kept stirring for 5 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the ethyl acetate/water system was extracted three times. The organic layer was separated, washed with water and brine, dried over anhydrous sodium sulfate , 7.0 mmol), used directly in the next reaction.
  • Step 3 Synthesis of ethyl 2-(2-chloropyrimidin-4-yl)-2-methylpropanoate
  • Ethyl 2-(2-chloropyrimidin-4-yl)acetate (1.4 g, 7.0 mmol) was placed in a 100 mL single-mouth reaction flask, and acetone (50 mL) was added to dissolve the substrate.
  • Potassium carbonate (2.9 g, 21.0 mmol) and methyl iodide (1.5 g, 10.5 mmol) were sequentially added to the stirred reaction flask at room temperature, and then the reaction was heated to 60 ° C and kept stirring for 5 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the precipitate was removed by filtration. The filter cake was rinsed three times with ethyl acetate.
  • Ethyl 2-(2-chloropyrimidin-4-yl)-2-methylpropanoate (900 mg, 4.0 mmol) was placed in a 50 mL single-mouth reaction flask, and methanol (5 mL) was added to partially dissolve the substrate. Then 1N aqueous sodium hydroxide solution (5 mL) was added, and the reaction mixture was kept stirring at room temperature for 6 h. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction solution was extracted once with ethyl acetate/water system. The aqueous layer was separated, and the pH was adjusted to 5 with 2N aqueous hydrochloric acid and extracted three times with ethyl acetate/water.
  • the crude acid chloride was slowly added to a solution of benzene-1,3-diamine (365 mg, 3.38 mmol) and triethylamine (0.63 mL, 4.5 mmol) in dichloromethane (10 mL) at 0 ° C. Stirring was continued for 1 h at 0 °C. TLC was used to detect the progress of the reaction. After the substrate was completely reacted, the reaction solution was extracted three times with dichloromethane/water. The organic layer was separated, washed with brine, dried over anhydrous sodium sulfate and evaporated. , directly into the next step of the reaction.
  • N-(3-Aminophenyl)-2-(2-chloropyrimidin-4-yl)-2-methylpropanamide (700 mg) was placed in a 50 mL single-mouth reaction flask, and dichloromethane (10 mL) was added. The substrate is dissolved. Thereafter, triethylamine (1 mL) and acryloyl chloride (0.5 mL) were successively added at 0 ° C, and the reaction was kept stirring at 0 ° C for 1 h. TLC was used to detect the progress of the reaction.
  • Step 7 N-(3-(2-(2-(2-methoxy-4-(4-methylpiperazin-1-yl)phenylamino)pyrimidin-4-yl)-2-methyl) Synthesis of trifluoroacetate of propionylamino)phenyl)acrylamide
  • N-(3-(2-(2-chloropyrimidin-4-yl)-2-methylpropionylamino)phenyl)acrylamide (710 mg, 0.91 mmol) and 2-methoxy-4-(4) -Methylpiperazin-1-yl)aniline (402 mg, 1.82 mmol) was placed in a 50 mL single-mouth reaction flask, and n-butanol (10 mL) was added to partially dissolve the substrate. Trifluoroacetic acid (620 mg, 5.46 mmol) was then added and the reaction was maintained at 130 ° C for 16 h. After completion of the reaction, it was cooled to room temperature and concentrated under reduced pressure to give a crude material.
  • the preparation method was the same as in Example 23 except that 4-(4-ethylpiperazin-1-yl)-2-methoxyaniline was replaced by 4-(4-propylpiperazin-1-yl). 2-methoxyaniline (preparation method similar to intermediate 11).
  • the preparation method was the same as in Example 32 except that 1-(4-amino-3-methoxyphenyl)-N,N-dimethylpiperidin-4-amine in Step 2 was replaced with N 1 -( 2-(Dimethylamino)ethyl)-3-methoxy-N 1 methylbenzene-1,4-diamine (preparation method similar to intermediate 4).
  • the preparation method was the same as in Example 9, except that the starting material 1-(2-chloropyrimidin-4-yl)-3-(3-nitrophenyl)urea was replaced by 1-(2,5-dichloropyridine. 4-yl)-3-(3-nitrophenyl)urea.
  • the reagents used in the following z-lyte test methods were purchased from Invitrogen.
  • the inhibitory effect of the test substance on the kinase activity of the double mutant EGFR kinase T790M/L858R kinase (Invitrogen, PV4879) and wild type EGFR kinase (EGFR WT) (Invitrogen, PV3872) was determined by the z-lyte method.
  • the working concentrations of the components in the 10 ⁇ L T790M/L858R kinase reaction were: 25 ⁇ M ATP, 0.08 ng/ ⁇ L T790M/L858R kinase, 2 ⁇ M Tyr04 substrate (Invitrogen, PV3193).
  • the concentration of DMSO after addition of the compound prepared in the above examples of the present invention was 2%.
  • the working concentrations of the components in the 10 ⁇ L EGFR WT kinase reaction were: 10 ⁇ M ATP, 0.8 ng/ ⁇ L T790M WT kinase, 2 ⁇ M Tyr04 substrate (Invitrogen, PV3193).
  • the concentration of DMSO after adding the test substance was 2%.
  • the 10 mM drug stock solution was diluted at room temperature with a water gradient of 4% DMSO to a final concentration of 10-0.005 [mu]M.
  • 2.5 ⁇ L of the test substance solution and 5 ⁇ L of a mixture of T790M/L858R kinase (or EGFR WT kinase) diluted with the reaction buffer and Tyr04 substrate were added to each well, and 2.5 ⁇ L of ATP was added to initiate the reaction.
  • the C1 well was replaced with ATP with a reaction buffer, no drug was added to the C2 well, and the phosphorylated substrate was added to the C3 well as described. After shaking at room temperature for 60 min.
  • Phosphorylation rate (1-((ER ⁇ C3 520nm - C3 450nm ) / ((C1 450nm - C3 450nm ) + ER ⁇ (C3 520nm - C1 520nm )))) ⁇ 100%
  • Inhibition rate (IR) (1 - (phosphorylation rate of test compound) / (phosphorylation rate of C2)) ⁇ 100%
  • the half-inhibitory concentration IC 50 was calculated using XLFIT 5.0 software (IDBS, UK).
  • the inhibitory activity or selective inhibitory activity of the exemplified compounds on the enzymes is shown in Tables 1 to 2.
  • the exemplified compounds of the present invention showed strong inhibitory activity against EGFR mutant enzymes (T790M/L858R and L858R) and weaker inhibitory activity against EGFR wild-type enzyme (EGFR WT).
  • the compounds of the present invention have significant selective inhibitory activity against EGFR mutant enzymes compared to the positive control BIBW2992 (Afatinib).
  • the selective inhibitory activity of some of the exemplified compounds of the present invention exceeds that of the positive control CO1686 (see, in particular, Cancer Discovery, doi: 10.1158/2159-8290. CD-13-0314).
  • Example 42 MTT (3-(4,5-dimethylthiazole-2)-2,5-diphenyltetrazolium bromide) method for detecting cell viability
  • MTT test method steps are carried out using methods well known to those skilled in the art, and the reagents used in the methods are commercially available.
  • the medium was removed and 0.25% trypsin/EDTA (Gibco, 25200-056) was added.
  • the cells were then counted and the cells were diluted to the following concentrations: A431 and H1975 cells at 27,800 per ml and NIH3T3 at 33,300 per ml.
  • the cells were seeded in a 96-well plate (BD 3072), 90 ⁇ L per well, and cultured overnight.
  • A431 cell culture medium 10% FBS (Gibco, 10099-141) DMEM (Hyclone SH30243.01B);
  • NIH3T3 cell culture medium 10% FBS (Gibco, 10099-141) DMEM (Hyclone SH30243.01B);
  • H1975 cell culture medium 10% FBS (Gibco, 10099-141) RPMI-1640 (Hyclone SH30809.01B);
  • the cells were placed in an incubator. After 72 hours of culture, 10 ⁇ L of 5 mg/ml MTT (Sigma, M5655) solution was added to each well, and then the 96-well plate was incubated in a 37 ° C 5% CO 2 incubator for 4 hours.
  • the plate was again centrifuged at 2000 rpm for 5 min. After removing the supernatant, 150 ⁇ L of DMSO was added to each well, and the plate was shaken in a shaker until all crystal violet was dissolved (about 10-20 min). Finally, the 492 nm light absorption was measured using a fluorescence microplate reader, and the IC 50 was calculated using XLFIT 5.0 software (IDBS, UK). The inhibitory activity or selective inhibitory activity of the exemplified compounds on the cells is shown in Tables 3 to 6.
  • the compounds of the present invention showed strong inhibitory activity against EGFR mutant cells (H1975 cells), but showed weak inhibitory activity against EGFR wild-type cells (A431 cells), and a positive control.
  • the compounds of the present invention have significant selective inhibitory activity against EGFR mutant cells.
  • some of the exemplified compounds of the present invention have a selective inhibitory activity against EGFR mutant cells even exceeding the positive control CO1686.
  • the compounds of the present invention have higher IC50 values for NIH3T3 cells and thus exhibit less toxicity.
  • Example 43 EGFR T790M inhibitor cell viability assay by ELISA
  • the reagents, solution preparation methods, and cell treatment and lysate preparation steps and ELISA detection steps in the following methods were carried out in accordance with the instructions of R&D DYC3570, R&D DYC1095E and R&D DYC1095BE.
  • Cell lysis buffer 1% NP-40, 20 mM Tris (pH 8.0), 137 mM NaCl, 10% glycerol, 1 mM NaVO 3 , 2 mM EDTA.
  • Cell lysate cell lysis buffer + 10 ⁇ g/mL aprotinin (Sigma), 10 ⁇ g/mL Leupeptin (Sigma), now available.
  • 1x PBS buffer NaCl: 0.137 M, KCl: 0.0027 M, Na 2 PO 4 -12H 2 O: 0.01 M, KH 2 PO 4 : 0.0015 M, pH 7.4.
  • Wash Buffer PBS buffer containing 0.05% Tween-20.
  • Antibody dilutions were detected: 20 mM Tris, 137 mM NaCl, 0.05% Tween-20, 0.1% BSA, pH 7.2-7.4.
  • Blocking solution PBS buffer containing 1% BSA.
  • ELISA kits R&D DYC3570, R&D DYC1095E and R&D DYC1095BE.
  • H1975 cells were seeded at a density of 1 ⁇ 10 4 /well into 96-well plates at 90 ⁇ l of 10% FBS per well, 1640 medium, and cultured overnight at 37 ° C, 5% CO 2 .
  • the R&D capture antibody ((DYC1095BE or DYC1095E)) was diluted with PBS 1:180, and the diluted antibody 100 ⁇ L/well was added to an ELISA reaction plate (Corning costar 42592), and shaken overnight at 25 ° C;
  • the detection antibody is diluted with the detection antibody dilution solution in the prescribed ratio of the kit, and 100 ⁇ L is added to each well, and the mixture is incubated at 25 ° C for 1 hour in the dark;
  • Inhibition rate (%) 100% ⁇ (OD cell- OD drug treatment ) / (OD cell - OD blank )
  • A431 cells were seeded at a density of 1 ⁇ 10 4 /well into a 96-well plate, and 90 ⁇ l of each well was cultured in DMEM medium containing 10% FBS at 37 ° C, 5% CO 2 overnight.
  • the A431 cell culture medium was changed to 90 ⁇ l of serum-free DMEM medium, and cultivation was continued overnight.
  • R&D capture antibody (DYC3570E) was diluted with PBS 1:180, diluted antibody 100 ⁇ L/well was added to an ELISA reaction plate (Corning costar 42592), and shaken overnight at 25 ° C;
  • the detection antibody is diluted with the detection antibody dilution solution in the prescribed ratio of the kit, and 100 ⁇ L is added to each well, and the mixture is incubated at 25 ° C for 1 hour in the dark;
  • Inhibition rate (%) 100% ⁇ (OD EGF - OD drug ) / (OD EGF - OD cells )
  • the exemplified compounds of the present invention have significant selective inhibitory activity against cell-level targets compared to the positive control BIBW2992. And the selective inhibition activity of some of the exemplified compounds even exceeded the positive control CO1686, up to a factor of six.
  • the compounds of the present invention showed strong inhibitory activity against EGFR mutant enzymes and cells, and showed weak inhibitory activity against EGFR wild-type enzymes and cells, and thus such compounds have T790M mutations.
  • EGFR has better selective inhibitory activity and lower cytotoxicity.

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Abstract

提供一种对EGFR酪氨酸激酶具有抑制作用的芳氨基嘧啶类化合物及其药学上可接受的盐、立体异构体、溶剂化合物或前药，还提供含有该化合物的药物组合物、药用组合物及其应用。

Description

芳氨基嘧啶类化合物及其应用以及由其制备的药物组合物和药用组合物 技术领域
本发明涉及医药技术领域，特别涉及一种芳氨基嘧啶类化合物及其作为EGFR酪氨酸激酶抑制剂的应用，以及由其制备的药物组合物和药用组合物。
背景技术
肺癌是全球发病率最高的癌症，在中国肺癌发病率位居所有癌症中第一位，也是中国发病率和死亡率最高的癌症，在中国肺癌病人中，30％病人具有EGFR突变，其中L858R和外显子19缺失突变占90％以上，这类病人对EGFR抑制剂更为敏感。现有已上市第一代EGFR抑制剂例如厄洛替尼，吉非替尼对这类病人效果良好，能够使其中60％以上病人肿瘤缩小，明显延长病人无进展生存期。但绝大多数病人在6-12个月获得耐药，第一代EGFR抑制剂将不再起效，而这类病人目前处于无药可用状态。临床发现对第一代EGFR抑制剂产生耐药的病人中有50％检测到EGFR T790M突变。在T790M突变细胞系H1975中第一代EGFR抑制剂，吉非替尼和厄洛替尼，均大于3uM，基本没有活性。
目前开发上市的第二代不可逆pan-EGFR抑制剂(Afatinib(BIBW2992))对EGFR突变肺癌病人疗效显著好于第一代EGFR抑制剂。但第二代抑制剂同时也具有很强的野生型EGFR抑制活性，对野生型EGFR的抑制活性显著高于耐药T790M突变，病人皮疹等毒副作用严重且对耐药病人疗效较差，仅有小部分第一代EGFR抑制剂耐药病人对这类药物产生应答。
为了提高对耐药EGFR T790M突变抑制活性的同时降低对野生型EGFR的抑制活性，开发活性更高、选择性更好、毒性更低的第三代EGFR突变体选择性抑制剂具有重要的意义。
发明内容
本发明的目的是提供一种芳氨基嘧啶类化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药。
本发明的另一个目的是提供包含上述芳氨基嘧啶类化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药的药物组合物。
本发明的第三个目的是提供上述芳氨基嘧啶类化合物制备调控EGFR酪氨酸激酶活性的药物，或制备治疗EGFR相关疾病药物上的应用。
本发明第一方面提供了一种新的芳氨基嘧啶类化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药，所述芳氨基嘧啶类化合物结构如式(I)所示：
Figure PCTCN2015073045-appb-000001
式(I)中，A选自：取代或未取代的C6-10芳烃基、取代或未取代的C5-8杂芳基、取代或未取代的C3-8环烷基、取代或未取代的C3-8杂环烷基、取代或未取代的8-14元稠环基团；其中，所述的“取代”是指基团中的一个或多个氢被选自下组的取代基所取代：硝基、卤素、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、卤代的C1-6烷氧基；
Z1选自NR9或CR10R11；其中，R9选自氢或C1-6烷基；R10、R11各自独立地选自氢、卤素、C1-6烷基；或R10和R11与相连的碳原子共同形成C3-8环烷基或C3-8杂环烷基；
Z2、Z3各自独立地选自CR12或N；其中，R12选自氢、卤素、硝基、氨基、C1-6烷基取代的氨基、C1-6酰基取代的氨基、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、卤代的C1-6烷氧基；
R1选自氢或C1-6烷基；或者，R1与A及它们相连的氮原子一起组成选自下列的基团：取代或未取代的C5-8杂芳基、取代或未取代的C3-8杂环烷基、或者取代或未取代的8-14元稠环基团，其中，所述的“取代”是指基团中的一个或多个氢被选自下组的取代基所取代：硝基、卤素、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、卤代的C1-6烷氧基；
R2选自氢、卤素、C1-6烷基、卤代C1-6烷基、或C1-6烷氧基；
R3、R4、R5各自独立地选自：氢、卤素、硝基、氨基、C1-6烷基取代的氨基、C1-6酰基取代的氨基、C1-6烷基取代的二氨基、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、C1-6烷氧基取代的C1-6烷氧基、C6-10芳烃基取代的C1-6烷氧基、卤代的C1-6烷氧基、-O-C6-10芳烃基、-CO-C3-6杂环烷基、-CO-NR13R14、C3-6杂芳基、C3-6杂环烷基、C3-6杂环烷基-CO-C1-6烷基，C3-6杂环烷基-N-C1-6烷基，C3-6杂环烷基-C1-6烷基，C3-6杂环烷基-C3-6杂环烷基；任选地，所述C6-10芳烃基、C3-6杂芳基、或者C3-6杂环烷基可被一个或多个选自下组的取代基所取代：硝基、卤素、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、卤代的C1-6烷氧基、C1-6烷基取代的氨基；其中，R13、R14各自独立地选自氢、或C1-6烷基；
R7、R8各自独立地选自：氢、卤素、C1-6烷基、C3-8环烷基、C3-8杂环烷基、C6-10芳烃基或C5-8杂芳基。
在另一优选例中，Z1选自NR9或CR10R11；R9选自氢或甲基；R10、R11各自独立地选自氢、或甲基。
在另一优选例中，Z2、Z3各自独立地选自CH或N。
在另一优选例中，R1选自氢或甲基，或R1与A及它们相连的氮原子一起组成哌啶。
在另一优选例中，R2选自氢、氟、氯、甲基、乙基、丙基、异丙基、三氟甲基。
在另一优选例中，R3为氢、卤素、C1-6烷氧基(优选甲氧基)。
在另一优选例中，R3为氢或甲氧基。
在另一优选例中，R4为氢、卤素、C1-6烷氧基(优选甲氧基)。
在另一优选例中，R4为氢、氟、氯或甲氧基。
在另一优选例中，R5为氢、卤素、氨基、C1-6烷基取代的二氨基、C1-6烷基、C1-6烷氧基、C3-6杂环烷基、-CO-C3-6杂环烷基、-CO-NR13R14、C3-6杂环烷基-C3-6杂环烷基，所述C3-6杂环烷基可被一个或多个选自下组的取代基所取代：C1-6烷基、卤代的C1-6烷基、C1-6烷基取代的氨基。
在另一优选例中，R5为氢、氟、氯、二乙基胺基、N,N,N’-三甲基乙二胺基、甲基、乙基、丙基、甲氧基、乙氧基、C6杂环烷基、-CO-C6杂环烷基、-CO-N(CH2CH3)2、C6杂环烷基-C6杂环烷基，所述C6杂环烷基可被一个或多个选自下组的取代基所取代：甲基、乙基、丙基、氟乙基、N(CH3)2、N(CH2CH3)2。
在另一优选例中，R5为氢、氟、氯、二乙基胺基、N,N,N’-三甲基乙二胺基、甲基、乙基、丙基、甲氧基、乙氧基、吗啉基、哌嗪基、哌啶基、-CO-吗啉基、-CO-哌嗪基、-CO-哌啶基、-CO-N(CH2CH3)2、哌啶基-哌嗪基、哌啶基-吗啉基，所述吗啉基、哌嗪基、哌啶基可被一个或多个选自下组的取代基所取代：甲基、乙基、丙基、氟乙基、N(CH3)2、N(CH2CH3)2。
在另一优选例中，R5为甲氧基、或为选择下组的结构：
Figure PCTCN2015073045-appb-000002
Figure PCTCN2015073045-appb-000003
在另一优选例中，所述芳氨基嘧啶类化合物为式(II)所示的化合物：
Figure PCTCN2015073045-appb-000004
式(II)中，A、Z2、Z3、R1、R2、R3、R4、R5、R7、R8和R9如式(I)中所定义。
在另一优选例中，所述芳氨基嘧啶类化合物为式(III)所示的化合物：
Figure PCTCN2015073045-appb-000005
式(III)中，A、Z2、Z3、R1、R2、R3、R4、R5、R7、R8、R10和R11如式(I)中所定义。
在另一优选例中，本发明提供的式(I)化合物、式(II)化合物、或式(III)化合物中，R7、R8各自独立地为氢。
在另一优选例中，本发明提供的式(I)化合物、式(II)化合物、或式(III)化合物中， A为苯基。
在另一优选例中，所述芳氨基嘧啶类化合物为式a所示的化合物：
Figure PCTCN2015073045-appb-000006
其中，Z1、Z2、Z3、R1、R2、R3、R4、R5、R7、R8如式(I)中所定义。
在另一优选例中，本发明提供的式(I)化合物、式(II)化合物、或式(III)化合物中，A为苯基，而且，R7、R8各自独立地为氢。
在另一优选例中，本发明提供的式(I)化合物、式(II)化合物、或式(III)化合物中，R1与A及它们相连的氮原子一起组成哌啶。
在另一优选例中，所述芳氨基嘧啶类化合物为式b所示的化合物：
Figure PCTCN2015073045-appb-000007
其中，Z1、Z2、Z3、R2、R3、R4、R5、R7、R8如式(I)中所定义。
在另一优选例中，本发明提供的式(I)化合物、式(II)化合物、或式(III)化合物其中，R1与A及它们相连的氮原子一起组成哌啶，而且，R7、R8各自独立地为氢。
在另一优选例中，本发明提供了选自下列的化合物，或其药学上可接受的盐、立体异构体、溶剂化合物或其前药：
N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-1)；
N-(3-(3-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-2)；
N-(3-(3-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-3)；
N-(3-(3-(2-((3-氯-4-((3-氟苄基)氧)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-4)；
N-(3-(3-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-5)；
N-(3-(3-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-6)；
N-(3-(3-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-7)；
N-(3-(3-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-8)；
N-(3-(3-(2-((2-甲氧基-4-(4-吗啉哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-9)；
N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基) 丙烯酰胺(cpd-10)；
N-(3-(3-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-11)；
N-(3-(3-(2-((4-(二甲氨基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-12)；
N-(3-(3-(2-((5-吗啉代吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-13)；
N-(3-(3-(2-((2-甲氧基-4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-14)；
N-(3-(3-(2-((2-甲氧基-4-(吗啉-4-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-15)；
N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-16)；
4-((4-(3-(3-丙烯酰基酰胺苯基)脲基)嘧啶-2-基)氨基)-N,N-二乙基-3-甲氧基苯甲酰胺(cpd-17)；
N-(3-(3-(2-((4-(2-甲氧基乙氧基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-18)；
N-(3-(3-(2-((2-甲氧基-4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-19)；
N-(3-(3-(2-((4-(4-(2-氟乙基)哌嗪-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-20)；
N-(3-(3-(2-((4-(二乙基氨基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-21)；
N-(3-(3-(2-((4-(4-(二乙基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-22)；
N-(3-(3-(2-((4-(4-乙基哌嗪-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-23)；
N-(3-(3-(2-(4-甲苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-24)；
N-(3-(3-(2-((5-(4-甲基哌嗪-1-基)吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-25)；
N-(3-(3-(2-((4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-26)；
N-(3-(3-(2-((4-乙氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-27)；
N-(3-(3-(2-((2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-28)；
N-(3-(3-(2-((4-氯苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-29)；
N-(3-(3-(2-((3,4,5-三甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-30)；
N-(3-(3-(2-((4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-31)；
N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)-3-甲基脲基)苯基)丙烯酰胺(cpd-32)；
3-丙烯酰氨基-N-(2-(2-甲氧基-4-(4-吗啉代-1-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-33)；
3-丙烯酰氨基-N-(2-(2-甲氧基-4-(4-甲基哌嗪-1-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-34)；
3-丙烯酰氨基-N-(2-(2-甲氧基-4-(1-甲基哌啶-4-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-35)；
3-丙烯酰氨基-N-(2-(4-((2-(二甲基氨基)乙基)(甲基)氨基)-2-甲氧基苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-36)；
N-(3-(2-(2-(2-甲氧基-4-(4-甲基哌嗪-1-基)苯基氨基)嘧啶-4-基)-2-甲基丙酰氨基)苯基)丙烯酰胺(cpd-37)；
1-(3-丙烯酰基酰胺基)-3-(2-(2-甲氧基-4-(4-丙基哌嗪-1-基)苯基氨基)嘧啶-4-基)脲(cpd-38)；
3-(3-丙烯酰基酰胺基)-1-(2-(4-((2-(二甲基氨基)乙基)(甲基)氨基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基脲(cpd-39)；或
1-(3-丙烯酰基酰胺基)-3-(5-氯-2-(2-甲氧基-4-(4-吗啉代哌啶-1-基)苯基氨基)嘧啶-4-基)脲(cpd-40)。
在本发明提供的上述芳氨基嘧啶类化合物中，所述的烷基，作为基团或是其他基团的一部分，例如卤素取代的烷基、羟基取代的烷基，可以是直链的或是支链的。例如，C1-6烷基表示具有1到6个碳的烷基，包括但不限于甲基、乙基、丙基、异丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、正己基。
在本发明提供的上述芳氨基嘧啶类化合物中，所述“烷氧基”是指烷基与氧原子连结后的生成基团，包括但不限于甲氧基、乙氧基、异丙氧基、环丙氧基等。
在本发明提供的上述芳氨基嘧啶类化合物中，所述“二氨基”是指通过烷基相连的二氨基，如乙二胺、丙二胺、丁二胺等；而且，末端的氨基可以被烷基取代，例如N,N,N’-三甲基丁二胺基。
在本发明提供的上述芳氨基嘧啶类化合物中，所述“氨基”指NH2。
在本发明提供的上述芳氨基嘧啶类化合物中，所述“C1-6烷基取代的氨基”指NH2中的一个或两个氢被C1-6烷基取代。包括但不限于NH(CH3)、N(CH3)2、N(CH2CH3)2等。
在本发明提供的上述芳氨基嘧啶类化合物中，“卤素”是指氟，氯，溴和碘。优选氟和氯。
在本发明提供的上述芳氨基嘧啶类化合物中，“芳烃基”是指包含六到十个碳原子的单环或稠合的芳烃环(例如苯基和萘基)。
在本发明提供的上述芳氨基嘧啶类化合物中，“杂芳基”是指任何稠合或非稠合的芳环系统，其中至少一个环是含有1-4个选自氮、氧和硫的杂原子的五到八元环，优选至少一个杂原子选自氮。杂芳基包括但不限于噻吩基、呋喃基、咪唑基、吡唑基、噻唑基、恶唑基、异恶唑基、吡啶基、嘧啶基、吡嗪基、哒嗪基、苯并咪唑基、苯并吡唑基、吲哚基等。
在本发明提供的上述芳氨基嘧啶类化合物中，“环烷基”是指包含指定数目的碳原子的饱和的或部分不饱和的单环、稠环或桥环。例如，C3-8环烷基是指三到八个碳的环烷基，包括环丙基、环丁基、环戊基、环己基等。
在本发明提供的上述芳氨基嘧啶类化合物中，“杂环烷基”是指本发明中所定义的环烷基，其中一个或多个环上的碳原子被氧、氮、-NR-、硫、羰基、-S(O)-或-S(O)2- 等基团取代；杂环烷基包括但不限于吗啉基、哌嗪基、哌啶基、硫代吗啉基等。
在本发明提供的上述芳氨基嘧啶类化合物中，“C1-6酰基”包括甲酰基、乙酰基、丙酰基、丁酰基、戊酰基、己酰基。
在本发明提供的上述芳氨基嘧啶类化合物中，“稠环基团”是指两个或两个以上碳环或杂环以共有环边而形成的多环有机化合物，如萘、蒽、蒽醌、菲等。
在本发明提供的上述芳氨基嘧啶类化合物中，“药学上可接受的盐”包括药学可接受的酸加成盐和药学可接受的碱加成盐。“药学上可接受的酸加成盐”是指能够保留游离碱的生物有效性而无其他副作用的，与无机酸或有机酸所形成的盐。无机酸盐包括但不限于盐酸盐、氢溴酸盐、硫酸盐、磷酸盐等；有机酸盐包括但不限于甲酸盐、乙酸盐、丙酸盐、乙醇酸盐、葡糖酸盐、乳酸盐、草酸盐、马来酸盐、琥珀酸盐、富马酸盐、酒石酸盐、柠檬酸盐、谷氨酸盐、天冬氨酸盐、苯甲酸盐、甲磺酸盐、对甲苯磺酸盐和水杨酸盐等。这些盐可通过本专业已知的方法制备。“药学可接受的碱加成盐”，包括但不限于无机碱的盐如钠盐，钾盐，钙盐和镁盐等。包括但不限于有机碱的盐，比如铵盐，三乙胺盐，赖氨酸盐，精氨酸盐等。这些盐可通过本专业已知的方法制备。
本发明的式(I)化合物可以存在多于一种晶型，包括各种晶型及其混合物。
本发明中提及的“溶剂化物”是指本发明的化合物与溶剂形成的配合物。它们或者在溶剂中反应或者从溶剂中沉淀析出或者结晶出来。例如，一个与水形成的配合物称为“水合物”。式(I)化合物的溶剂化物属于本发明范围之内。
本发明的化合物可以含有一个或多个手性中心，并以不同的光学活性形式存在。当化合物含有一个手性中心时，化合物包含对映异构体。本发明包括这两种异构体和异构体的混合物，如外消旋混合物。对映异构体可以通过本专业已知的方法拆分，例如结晶以及手性色谱等方法。当式(I)化合物含有多于一个手性中心时，可以存在非对映异构体。本发明包括拆分过的光学纯的特定异构体以及非对映异构体的混合物。非对映异构体可由本专业已知方法拆分，比如结晶以及制备色谱。
本发明包括上述化合物的前药。前药包括已知的氨基保护基和羧基保护基，在生理条件下被水解或经由酶反应释放得到母体化合物。具体的前药制备方法可参照(Saulnier，M.G.；Frennesson，D.B.；Deshpande，M.S.；Hansel，S.B and Vysa，D.M.Bioorg.Med.Chem Lett.1994，4，1985-1990；和Greenwald，R.B.；Choe，Y.H.；Conover，C.D.；Shum，K.；Wu，D.；Royzen，M.J.Med.Chem.2000，43，475.)。
本发明第二方面提供了上述芳氨基嘧啶类化合物的制备方法，本发明中的化合物可以通过多种合成操作容易地制备，这些操作是所属领域技术人员熟练掌握的。这些化合物的特定制备方法可以包括(但不限于)下文所述的流程。
本发明式(I)化合物优选的合成路线有如下三种，以下实施例中具体化合物可参照下述合成路线进行制备，在具体操作过程中，可以根据需要对方法中的步骤进行扩展或合并。
合成路线1
Figure PCTCN2015073045-appb-000008
步骤1-1：嘧啶4位的Z1H与胺类化合物反应，转化成相应的酰胺类化合物，反应需在一定温度下，使用合适的碱及适当的溶剂才能进行，所用的碱可以是但不限于三乙胺。
步骤1-2：嘧啶2位的氯以胺取代，需在一定温度下，使用合适的催化剂及适当的溶剂才能进行。使用酸催化，催化剂可以是但不限于TFA或对甲苯磺酸。使用Buchwald-Hartwig胺化法，所用的钯催化剂可以是但不限于Pd2(dba)3，所用的配体可以是但不限于XantPhos，所用的碱可以是但不限于碳酸铯。
步骤1-3：该硝基化合物转化为相应的胺基化合物可在酸性条件下，用金属(可以是但不限于铁粉，锌粉)或者氯化亚锡进行还原；或者在钯碳催化下，加氢还原。
步骤1-4：该胺基化合物可在碱性条件下与相应的酰氯缩合成酰胺，或者在缩合剂存在下与相应的羧酸缩合成酰胺。
合成路线2
Figure PCTCN2015073045-appb-000009
步骤2-1：嘧啶4位的Z1H与胺类化合物反应，转化成相应的酰胺类化合物，反应需在一定温度下，使用合适的碱及适当的溶剂才能进行，所用的碱可以是但不限于三乙胺。
步骤2-2：该硝基化合物转化为相应的胺基化合物可在酸性条件下，用金属(可以是但不限于铁粉，锌粉)或者氯化亚锡进行还原；或者在钯碳催化下，加氢还原。
步骤2-3：该胺基化合物可在碱性条件下与相应的酰氯缩合成酰胺，或者在缩合 剂存在下与相应的羧酸缩合成酰胺。
步骤2-4：嘧啶2位的氯以胺取代，需在一定温度下，使用合适的催化剂及适当的溶剂才能进行。使用酸催化，催化剂可以是但不限于TFA或对甲苯磺酸。使用Buchwald-Hartwig胺化法，所用的钯催化剂可以是但不限于Pd2(dba)3，所用的配体可以是但不限于XantPhos(4,5-双(二苯基膦)-9,9-二甲基氧杂蒽)，所用的碱可以是但不限于碳酸铯。
合成路线3
Figure PCTCN2015073045-appb-000010
步骤3-1：嘧啶2位的氯以胺取代，需在一定温度下，使用合适的催化剂及适当的溶剂才能进行。使用酸催化，催化剂可以是但不限于TFA或对甲苯磺酸。使用Buchwald-Hartwig胺化法，所用的钯催化剂可以是但不限于Pd2(dba)3，所用的配体可以是但不限于XantPhos，所用的碱可以是但不限于碳酸铯。
步骤3-2：嘧啶4位的Z1H与胺类化合物反应，转化成相应的酰胺类化合物，反应需在一定温度下，使用合适的碱及适当的溶剂才能进行，所用的碱可以是但不限于三乙胺。
步骤3-3：该硝基化合物转化为相应的胺基化合物可在酸性条件下，用金属(可以是但不限于铁粉，锌粉)或者氯化亚锡进行还原；或者在钯碳催化下，加氢还原。
步骤3-4：该胺基化合物可在碱性条件下与相应的酰氯缩合成酰胺，或者在缩合剂存在下与相应的羧酸缩合成酰胺。
本发明第三方面提供了一种药物组合物，它包括上述化合物中任一个或多个化合物或其药学可接受的盐、溶剂化物或前药，并且还包括药学可接受的载体。
本发明提供的一种药物组合物，所述组合物包括治疗有效量的上述芳氨基嘧啶类化合物，如上述式(I)化合物、式(II)化合物、或式(III)化合物，或上述示例化合物，或其药学上可接受的盐、立体异构体、溶剂化合物或其前药；以及药学可接受的载体。
通常，本发明化合物或其药学可接受的盐可以与一种或多种药用载体形成适合的剂型施用。这些剂型适用于口服、直肠给药、局部给药、口内给药以及其他非胃肠道施用(例如，皮下、肌肉、静脉等)。例如，适合口服给药的剂型包括胶囊、片剂、颗粒剂以及糖浆等。这些制剂中包含的本发明的化合物可以是固体粉末或颗粒；水性或非水性液体中的溶液或是混悬液；油包水或水包油的乳剂等。上述剂型可由活性化合 物与一种或多种载体或辅料经由通用的药剂学方法制成。上述的载体需要与活性化合物或其他辅料兼容。对于固体制剂，常用的无毒载体包括但不限于甘露醇、乳糖、淀粉、硬脂酸镁、纤维素、葡萄糖、蔗糖等。用于液体制剂的载体包括水、生理盐水、葡萄糖水溶液、乙二醇和聚乙二醇等。活性化合物可与上述载体形成溶液或是混悬液。
本发明第四方面提供了上述化合物或其药学上可接受的盐、溶剂化物或前药可用于制备调控EGFR酪氨酸激酶活性的药物，或制备治疗EGFR相关疾病的药物中的应用。
作为优选，所述EGFR相关疾病为癌症，糖尿病，免疫系统疾病，神经退行性疾病或心血管疾病。
作为优选，所述癌症为非小细胞肺癌、头颈癌、乳腺癌、肾癌、胰腺癌、子宫颈癌、食道癌、胰腺癌、前列腺癌、膀胱癌、结肠直肠癌、卵巢癌、胃癌、脑恶性肿瘤包括成胶质细胞瘤等，或它们的任何组合。
本发明的上述化合物或其可药学上可接受的盐、溶剂化物或其前药还可用于制备治疗EGFR异常表达的疾病，或用于制备在使用EGFR调节剂治疗期间具有获得性耐药性的疾病。
作为优选，所述获得性耐药性是由EGFR外显子20编码的T790突变引起的耐药性，如T790M。
在本发明中，EGFR调节剂是指靶向EGFR的小分子酪氨酸激酶抑制剂，如吉非替尼，厄洛替尼，埃克替尼，拉帕替尼或阿法替尼。
本发明的药用组合物，包括治疗有效量的上述化合物中任一个或多个化合物或其药学上可接受的盐、溶剂化物或前药，以及选自下组药物中的一种或多种：吉非替尼、厄洛替尼、埃克替尼、拉帕替尼、XL647、NVP-AEE-788、ARRY-334543、EKB-569、BIBW2992、HKI272、BMS-690514、CI-1033、凡德他尼、PF00299804、WZ4002、西妥昔单抗、曲妥珠单抗、帕尼突单抗、马妥珠单抗、尼妥珠单抗、扎鲁木单抗、帕妥珠单抗、MDX-214、CDX-110、IMC-11F8、Zemab、Her2疫苗PX 1041、HSP90抑制剂、CNF2024、坦螺旋霉素、阿螺旋霉素、IPI-504、SNX-5422、NVP-AUY922、或其组合。除本发明的化合物或其药学上可接受的盐、溶剂化物或前药以外，上述药用组合物中的其他药物均为本领域技术人员熟知的抗肿瘤药物。
“治疗有效量”是指可对人和/或动物产生功能或活性的且可被人和/或动物所接受的量。
本发明所述药物组合物或所述药用组合物中含有的本发明化合物或其药学上可接受的盐、溶剂化物或其前药的治疗有效量优选为0.1mg-5g/kg(体重)。
所述的药用组合物可用于治疗EGFR异常表达的疾病，如癌症，糖尿病，免疫系统疾病，神经退行性疾病或心血管疾病。
所述获得性耐药性的疾病是由EGFR外显子20编码的T790突变所引起的，或者是包含EGFR外显子20编码的T790突变所引起的。
在另一优选例中，所述的EGFR外显子20编码的T790为T790M。
本发明的化合物在某些疾病中可以与其他药物联合应用，以达到预期的治疗效果。一个联合应用的例子是用来治疗晚期NSCLC。例如，将治疗有效量的本发明式 I所示化合物与mTOR抑制剂联用(例如雷帕霉素)；或与Met抑制剂(包括Met抗体MetMAb和Met小分子抑制剂PF02341066)联用；或与IGF1R抑制剂联用(例如OSI-906)；或与热休克蛋白抑制剂联用等。
本发明公开了一类化合物以及化合物的制备方法、药物组成和治疗方案，本领域技术人员可以借鉴本文内容，适当改进工艺参数实现。特别需要指出的是，所有类似的替换和改动对本领域技术人员来说是显而易见的，它们都被视为包括在本发明。本发明的产品、方法及应用已经通过较佳实施例进行了描述，相关人员明显能在不脱离本发明内容、精神和范围内对本文所述的方法和应用进行改动或适当变更与组合，来实现和应用本发明技术。
应理解，在本发明范围内中，本发明的上述各技术特征和在下文(如实施例)中具体描述的各技术特征之间都可以互相组合，从而构成新的或优选的技术方案。
具体实施方式
发明人经过长期而深入的研究，意外地发现了一种芳氨基嘧啶类化合物，作为一类EGFR突变选择性的抑制剂，具有毒副作用低的优点。具体地，本发明提供的芳氨基嘧啶类化合物，作为一类EGFR突变选择性的抑制剂，体外酶、细胞实验表明其在纳摩尔浓度下可抑制L858R-T790M酶及双突变细胞株H1975的增殖；而对野生型EGFR及细胞株A431的抑制则相对较弱。其突变选择性大大减小了因抑制野生型EGFR而产生的毒副作用，适用于目前EGFR-TKI治疗中产生继发性耐药的病例，是第一、二代EGFR酪氨酸激酶抑制剂的理想替代物。
为了使本领域的技术人员更好地理解本发明的技术方案，下面结合具体实施例对本发明作进一步的详细说明。
本发明中，化合物的结构通过核磁共振(1H NMR)和/或液质联用质谱(LC-MS)来确定。1HNMR位移(δ)以百万分之一(ppm)的单位给出。1H NMR的测定是用Bruker AVANCE-400核磁仪，内标为四甲基硅烷(TMS)。
产物纯度测定通过LC-MS确定。LC-MS的测定用Agilent 1200HPLC System/6140MS液质联用质谱仪(生产商：安捷伦)，柱子Waters X-Bridge,150×4.6mm,3.5μm。
制备高效液相色谱(pre-HPLC)用Waters PHW007，柱子XBridge C18,4.6*150mm,3.5um。
采用ISCO Combiflash-Rf75或Rf200型自动过柱仪，Agela 4g、12g、20g、40g、80g、120g一次性硅胶柱。
薄层层析硅胶板使用烟台黄海HSGF254或青岛GF254硅胶板，薄层色谱法(TLC)，检测反应使用的硅胶板采用的规格是0.15mm～0.2mm，薄层色谱法分离纯化产品使用的硅胶板采用的规格是0.4mm～0.5mm。硅胶一般使用烟台黄海硅胶200～300目硅胶为载体。碱性氧化铝柱一般使用国药层析用FCP200～300目碱性氧化铝为载体。
本发明的已知的起始原料可以采用或按照本领域已知的方法来合成，或可以于ABCR GmbH&Co.KG，Acros Organics，Aldrich Chemical Company，韶远化学科技(Accela ChemBio Inc)和达瑞化学品等公司处购买。
实施例中无特殊说明，反应均在氮气或氩气氛下进行。氩气氛或氮气氛是指反应瓶连接一个约1L容积的氩气或氮气气球。
氢气氛是指反应瓶连接一个约1L容积的氢气气球，Parker-20H型氢气发生器。氢化反应通常抽真空，充入氢气，反复操作3次。
实施例中无特殊说明，溶液是指水溶液。
实施例中的反应进程的监测采用薄层色谱法(TLC)，反应所使用的展开剂的体系选自：二氯甲烷和甲醇体系、正己烷和乙酸乙酯体系、石油醚和乙酸乙酯体系和丙酮体系中的一种或多种；溶剂的体积比根据化合物的极性不同而进行调节。
纯化化合物采用的柱层析的洗脱剂的体系或薄层色谱法的展开剂的体系可选自：A、二氯甲烷和甲醇体系，B、正己烷和乙酸乙酯体系和C、二氯甲烷和丙酮体系中的一种或多种，溶剂的体积比根据化合物的极性不同而进行调节，也可以加入少量的三乙胺等碱性或醋酸等酸性试剂进行调节。
在本发明中，DMF：二甲基甲酰胺，DMSO：二甲基亚砜，THF：四氢呋喃，DIEA：N,N-二异丙基乙胺，EA：乙酸乙酯，PE：石油醚。
以下实施例中，室温指的是约25℃。
准备例
中间体1：2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯胺的制备
Figure PCTCN2015073045-appb-000011
步骤1：将叔丁基4-氧代哌啶-1-甲酸叔丁酯(5g,29.2mmol)和氰基硼氢化钠(2.8g，43.8mmol)置于500mL的单口反应瓶中，加入甲醇200ml溶解，之后加入1-甲基哌嗪(3.2g，32.1mmol)和冰醋酸(1ml)并保持反应体系在室温下过夜。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Combi-Flash柱层析纯化得到4-(4-甲基哌嗪-1-基)哌啶-1-甲酸叔丁酯(6.5g，79％)。MS m/z(ESI):284.1[M+H]+
步骤2：室温下，将4-(4-甲基哌嗪-1-基)哌啶-1-甲酸叔丁酯(6.5g,22.9mmol)加入到100ml盐酸二氧六环溶液中，室温下剧烈搅拌4h。反应结束后，将反应液减压浓缩，得到1-甲基-4-(哌啶-4-基)哌嗪(4.2g，100％)。MS m/z(ESI):184.1[M+H]+
步骤3：将1-甲基-4-(哌啶-4-基)哌嗪(4.2g,22.9mmol)，4-氟-2-甲氧基-1-硝基苯(4.69g，25.2mmol)和碳酸钾(9.48g,68.7mmol)置于250mL的单口反应瓶中，加入DMF(100mL)使底物部分溶解，保持反应体系在100℃下加热4h。TLC检测反应进度，待底物完全反应后，滤去不溶物，减压浓缩得到黄色固体1-(1-(3-甲氧基-4-硝基苯基)哌啶-4-基)-4-甲基哌嗪，经Combi-Flash柱层析纯化得到目标产品4a(6g，78％)。MS m/z(ESI):335.1[M+H]+
步骤4：室温下，将钯碳(0.6g,10％wt)加入到1-(1-(3-甲氧基-4-硝基苯基)哌啶-4- 基)-4-甲基哌嗪(6g,17.9mmol)的200ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯胺，直接用于下一步反应。MS m/z(ESI):305.1[M+H]+
中间体2：3-氨基-6-吗啉基吡啶的制备
Figure PCTCN2015073045-appb-000012
步骤1：室温下，将2-溴-5-硝基吡啶(2g,10mmol)加入到10ml吗啉溶液中，室温下剧烈搅拌4h。反应结束后，有黄色固体析出。过滤后，以50ml石油醚洗涤黄色固体，得到4-(5-硝基吡啶-2-基)吗啉(1.9g,92％)。MS m/z(ESI):210.1[M+H]+
步骤2：室温下，将钯碳(100mg,10％wt)加入到4-(5-硝基吡啶-2-基)吗啉(1g,9.1mmol)的60ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，得到产物3-氨基-6-吗啉基吡啶，直接用于下一步反应。MS m/z(ESI):180.1[M+H]+
中间体3：2-甲氧基-4-(4-吗啉代-1-基)苯胺的制备
Figure PCTCN2015073045-appb-000013
步骤1：将4-氧代哌啶-1-甲酸叔丁酯(5g,29.2mmol)和氰基硼氢化钠(2.8g，43.8mmol)置于500mL的单口反应瓶中，加入甲醇200ml溶解，之后加入吗啉(2.8g，32.1mmol)和冰醋酸(1ml)并保持反应体系在室温下过夜。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Combi-Flash柱层析纯化得到4-吗啉代哌啶-1-甲酸叔丁酯(6g，76％)。MS m/z(ESI):271.1[M+H]+
步骤2：室温下，将4-吗啉代哌啶-1-甲酸叔丁酯(6g,22.2mmol)加入到100ml盐酸二氧六环溶液中，室温下剧烈搅拌4h。反应结束后，将反应液减压浓缩，得到4-(哌啶-4-基)吗啉(3.8g，100％)。MS m/z(ESI):171.1[M+H]+
步骤3：将4-(哌啶-4-基)吗啉(3.8g,22.2mmol)，4-氟-2-甲氧基-1-硝基苯(4.18g，24.42mmol)和碳酸钾(9.19g,66.6mmol)置于250mL的单口反应瓶中，加入DMF(50mL)使底物部分溶解，保持反应体系在100℃下加热4h。TLC检测反应进度，待底物完全反应后，滤去不溶物，减压浓缩得到黄色固体，经Combi-Flash柱层析纯化得到4-(1-(3-甲氧基-4-硝基苯基)哌啶-4-基)吗啉(5.8g，81％)。MS m/z(ESI):322.1[M+H]+
步骤4：室温下，将钯碳(0.58g,10％wt)加入到4-(1-(3-甲氧基-4-硝基苯基)哌啶-4-基)吗啉(5.8g,18.1mmol)的200ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应 结束后，滤去钯碳，将滤液减压浓缩，产物2-甲氧基-4-(4-吗啉代-1-基)苯胺，直接用于下一步反应。MS m/z(ESI):292.2[M+H]+
中间体4：1-(4-氨基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2015073045-appb-000014
步骤1：将N,N-二甲基哌啶-4-胺(5g,39mmol)和4-氟-2-甲氧基-1-硝基苯(10.95g，58.6mmol)和碳酸钾(16.15g,117mmol)置于500mL的单口反应瓶中，加入DMF(150mL)使底物部分溶解，保持反应体系在100℃下加热4h。TLC检测反应进度，待底物完全反应后，滤去不溶物，减压浓缩得到黄色固体，经Combi-Flash柱层析纯化得到1-(3-甲氧基-4-硝基苯基)-N,N-二甲基哌啶-4-胺(7.5g，69％)。MS m/z(ESI):280.1[M+H]+
步骤2：室温下，将钯碳(0.75g,10％wt)加入到1-(3-甲氧基-4-硝基苯基)-N,N-二甲基哌啶-4-胺(7.5g,26.8mmol)的200ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物1-(4-氨基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺，直接用于下一步反应。MS m/z(ESI):250.2[M+H]+
中间体5：2-甲氧基-4-吗啉代苯胺的制备
步骤1：将4-氟-2-甲氧基-1-硝基苯(4.4g,25.7mmol)和碳酸铯(18.1g,51.4mmol)置于250mL的单口反应瓶中，加入DMF(100mL)使底物部分溶解。之后加入吗啉(3.3mL,38.6mmol)并保持反应体系在100℃下加热2h。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物4-(3-甲氧基-4-硝基苯基)吗啉，直接用于下步反应。MS m/z(ESI):238.1[M+H]+
步骤2：室温下，将钯碳(500mg,10％wt)加入到4-(3-甲氧基-4-硝基苯基)吗啉的100ml甲醇溶液中，在氢气气氛下室温剧烈搅拌反应过夜。反应结束后，滤去钯碳，滤液减压浓缩，产物2-甲氧基-4-吗啉苯胺(4.5g，两步产率84％)，直接用于下一步反应。MS m/z(ESI):209.1[M+H]+
中间体6：2-氨基-5-(4-吗啉基)吡啶的制备
Figure PCTCN2015073045-appb-000016
步骤1：室温下，将5-溴-2-硝基吡啶(1g,5mmol)加入到10ml吗啉溶液中，室温下剧烈搅拌4h。反应结束后，有黄色固体析出。过滤后，以50ml石油醚洗涤黄色固 体，得到4-(6-硝基吡啶-3-基)吗啉(530mg，53％)。MS m/z(ESI):210.1[M+H]+
步骤2：室温下，将钯碳(60mg,10％wt)加入到4-(6-硝基吡啶-3-基)吗啉(600mg,2.86mmol)的30ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，得到产物2-氨基-5-(4-吗啉基)吡啶，直接用于下一步反应。MS m/z(ESI):180.1[M+H]+
中间体7：2-甲氧基-4-(哌啶-1-基)苯胺的制备
Figure PCTCN2015073045-appb-000017
步骤1：将4-氟-2-甲氧基-1-硝基苯(1.9g,11.1mmol)和碳酸钾(4.6g,33.3mmol)置于60mL的单口反应瓶中，加入DMF(60mL)使底物部分溶解。之后加入哌啶(950mg,11.1mmol)并保持反应体系在100℃下加热6h。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Combi-Flash柱层析纯化得到1-(3-甲氧基-4-硝基苯基)哌啶(2.2g，85％)。MS m/z(ESI):237.1[M+H]+
步骤2：室温下，将钯碳(200mg,10％wt)加入到1-(3-甲氧基-4-硝基苯基)哌啶(2g,8.44mmol)的60ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物2-甲氧基-4-(哌啶-1-基)苯胺，直接用于下一步反应。MS m/z(ESI):207.1[M+H]+
中间体8：4-(4-(2-氟乙基)哌嗪-1-基)-2-甲氧基苯胺的制备
Figure PCTCN2015073045-appb-000018
步骤1：将4-氟-2-甲氧基-1-硝基苯(5g,29.2mmol)和碳酸钾(12.1g,87.7mmol)置于500mL的单口反应瓶中，加入DMF(100mL)使底物部分溶解。之后加入1-叔丁氧羰基哌嗪(8.2g,43.8mmol)并保持反应体系在100℃下加热4h。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Combi-Flash柱层析纯化得到4-(3-甲氧基-4-硝基苯基)哌嗪-1-羧酸叔丁酯(8.1g，82％)。MS m/z(ESI):338.2[M+H]+
步骤2：室温下，将4-(3-甲氧基-4-硝基苯基)哌嗪-1-羧酸叔丁酯(8g,23.7mmol)加入到100ml盐酸二氧六环溶液中，室温下剧烈搅拌1h。反应结束后，将反应液减压浓缩，得到1-(3-甲氧基-4-硝基苯基)哌嗪(5.6g，95％)。MS m/z(ESI):238.1[M+H]+
步骤3：将1-(3-甲氧基-4-硝基苯基)哌嗪(500mg,2.1mmol)和碳酸钾(900mg,6.3mmol)置于25mL的封管反应瓶中，加入乙腈(6mL)使底物部分溶解。之后加入1-溴-2-氟乙烷(290mg,2.3mmol)并保持反应体系在80℃下加热7h。TLC检测反应进度，待底 物完全反应后，滤去不溶物，减压浓缩得到黄色固体1-(2-氟乙基)-4-(3-甲氧基-4-硝基苯基)哌嗪(350mg，59％)。MS m/z(ESI):284.1[M+H]+
步骤4：室温下，将钯碳(35mg,10％wt)加入到1-(2-氟乙基)-4-(3-甲氧基-4-硝基苯基)哌嗪(350mg,1.24mmol)的50ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物4-(4-(2-氟乙基)哌嗪-1-基)-2-甲氧基苯胺，直接用于下一步反应。MS m/z(ESI):254.1[M+H]+
中间体9：N1，N1-二乙基-3-甲氧基苯-1,4-二胺的制备
Figure PCTCN2015073045-appb-000019
步骤1：将二乙胺(1g,13.7mmol)和4-氟-2-甲氧基-1-硝基苯(3.84g，20.5mmol)和碳酸钾(5.67g,41.1mmol)置于500mL的单口反应瓶中，加入DMF(50mL)使底物部分溶解，保持反应体系在100℃下加热4h。TLC检测反应进度，待底物完全反应后，滤去不溶物，减压浓缩得到黄色固体，经Combi-Flash柱层析纯化得到N,N-二乙基-3-甲氧基-4-硝基苯胺(1.8g，58％)。MS m/z(ESI):225.1[M+H]+
步骤2：室温下，将钯碳(0.22g,10％wt)加入到N,N-二乙基-3-甲氧基-4-硝基苯胺(1.8g,8.04mmol)的200ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物N1，N1-二乙基-3-甲氧基苯-1,4-二胺，直接用于下一步反应。MS m/z(ESI):195.1[M+H]+
中间体10：1-(4-氨基-3-甲氧基苯基)-N,N-二乙基哌啶-4-胺的制备
Figure PCTCN2015073045-appb-000020
步骤1：将4-氧代哌啶-1-甲酸叔丁酯(5g,29.2mmol)和氰基硼氢化钠(2.8g，43.8mmol)置于500mL的单口反应瓶中，加入甲醇200ml溶解，之后加入二乙胺(2.3g，32.1mmol)和冰醋酸(1ml)并保持反应体系在室温下过夜。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Combi-Flash柱层析纯化得到4-(二乙基氨基)哌啶-1-甲酸叔丁酯(5.2g，70％)。MS m/z(ESI):257.2[M+H]+
步骤2：室温下，将4-(二乙基氨基)哌啶-1-甲酸叔丁酯(5.2g,20.2mmol)加入到100ml盐酸二氧六环溶液中，室温下剧烈搅拌4h。反应结束后，将反应液减压浓缩，得到N,N-二乙基哌啶-4-胺(3.1g，100％)。MS m/z(ESI):157.1[M+H]+
步骤3：将N,N-二乙基哌啶-4-胺(3.1g,20.2mmol)4-氟-2-甲氧基-1-硝基苯(4.53g，24.2mmol)和碳酸钾(8.36g,60.6mmol)置于250mL的单口反应瓶中，加入DMF(100mL) 使底物部分溶解，保持反应体系在100℃下加热4h。TLC检测反应进度，待底物完全反应后，滤去不溶物，减压浓缩得到黄色固体，经Combi-Flash柱层析纯化得到N,N-二乙基-1-(3-甲氧基-4-硝基苯基)哌啶-4-胺(5g，80％)。MS m/z(ESI):308.1[M+H]+
步骤4：室温下，将钯碳(0.5g,10％wt)加入到N,N-二乙基-1-(3-甲氧基-4-硝基苯基)哌啶-4-胺(5g,17.9mmol)的200ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物1-(4-氨基-3-甲氧基苯基)-N,N-二乙基哌啶-4-胺，直接用于下一步反应。MS m/z(ESI):278.1[M+H]+
中间体11：4-(4-乙基哌嗪-1-基)-2-甲氧基苯胺的制备
Figure PCTCN2015073045-appb-000021
步骤1：将4-氟-2-甲氧基-1-硝基苯(5g,29.2mmol)和碳酸钾(12.1g,87.7mmol)置于500mL的单口反应瓶中，加入DMF(100mL)使底物部分溶解。之后加入1-叔丁氧羰基哌嗪(8.2g,43.8mmol)并保持反应体系在100℃下加热4h。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Combi-Flash柱层析纯化得到4-(3-甲氧基-4-硝基苯基)哌嗪-1-羧酸叔丁酯(8.1g，82％)。MS m/z(ESI):338.2[M+H]+
步骤2：室温下，将4-(3-甲氧基-4-硝基苯基)哌嗪-1-羧酸叔丁酯(8g,23.7mmol)加入到100ml盐酸二氧六环溶液中，室温下剧烈搅拌1h。反应结束后，将反应液减压浓缩，得到1-(3-甲氧基-4-硝基苯基)哌嗪(5.6g，95％)。MS m/z(ESI):238.1[M+H]+
步骤3：将1-(3-甲氧基-4-硝基苯基)哌嗪(500mg,2.1mmol)和碳酸钾(900mg,6.3mmol)置于50mL的单口反应瓶中，加入乙腈(6mL)使底物部分溶解。之后加入溴乙烷(253mg,2.3mmol)并保持反应体系在80℃下加热7h。TLC检测反应进度，待底物完全反应后，滤去不溶物，减压浓缩得到黄色固体1-乙基-4-(3-甲氧基-4-硝基苯基)哌嗪(400mg，72％)。MS m/z(ESI):266.3[M+H]+
步骤4：室温下，将钯碳(30mg,10％wt)加入到1-乙基-4-(3-甲氧基-4-硝基苯基)哌嗪(270mg,1mmol)的50ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物4-(4-乙基哌嗪-1-基)-2-甲氧基苯胺，直接用于下一步反应。MS m/z(ESI):236.3[M+H]+
中间体12：5-(4-甲基哌嗪-1-基)吡啶-2-胺的制备
Figure PCTCN2015073045-appb-000022
步骤1：室温下，将N-甲基哌嗪(30ml)加入到5-溴-2-硝基吡啶(5g,21.6mmol)的80ml二氯甲烷溶液中，45℃下剧烈搅拌2h。待底物完全反应后，加水稀释，用二氯甲烷/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后， 减压浓缩得黄色固体1-甲基-4-(6-硝基吡啶-3-基)哌嗪(2.5g，52％)。MS m/z(ESI):223.1[M+H]+
步骤2：室温下，将钯碳(250mg,10％wt)加入到1-甲基-4-(6-硝基吡啶-3-基)哌嗪(2.5g,11.2mmol)的60ml甲醇溶液中，在氢气气氛下室温剧烈搅拌20h。反应结束后，滤去钯碳，将滤液减压浓缩，产物5-(4-甲基哌嗪-1-基)吡啶-2-胺，直接用于下一步反应。MS m/z(ESI):193.1[M+H]+
中间体13：4-氨基-N,N-二乙基-3-甲氧基苯甲酰胺的制备
Figure PCTCN2015073045-appb-000023
步骤1：将4-硝基-3-甲氧基苯甲酸(5g,25.4mmol)加入到50ml的SOCl2中，滴加4滴DMF，90℃下剧烈搅拌3h。TLC检测反应进度，待底物完全反应后，产物浓缩后得到的黄色固体3-甲氧基-4-硝基苯甲酰氯直接用于下一步反应。
步骤2：在0℃下，将DIPEA(4mL,25.1mmol)与二乙胺(2.7mL,25.1mmol)加入到3-甲氧基-4-硝基苯甲酰氯(5.4g,25.1mmol)的50ml THF溶液中，室温下剧烈搅拌3h。TLC检测反应进度,待底物完全反应后,加水稀释,调节pH至碱性,用乙酸乙酯/水体系萃取三次,分离出有机层,经水和饱和食盐水洗涤,无水硫酸钠干燥后,减压浓缩得到粗产物6.6g。经Combi-Flash柱层析纯化得到化合物N,N-二乙基-3-甲氧基-4-硝基苯甲酰胺(5.5g,86％)。MS m/z(ESI):253.1[M+H]+
步骤3：将Pd/C(200mg)加入到N,N-二乙基-3-甲氧基-4-硝基苯甲酰胺(1g,3.97mmol)的50ml甲醇溶液中。室温下,在氢气氛围中剧烈搅拌16h。反应结束后,过滤,滤液浓缩,得化合物4-氨基-N,N-二乙基-3-甲氧基苯甲酰胺(840mg,95％),产物直接用于下一步。MS m/z(ESI):223.2[M+H]+
中间体14：(4-氨基-3-甲氧基苯基)(吗啉代)甲酮的制备
Figure PCTCN2015073045-appb-000024
步骤1：将4-硝基-3-甲氧基苯甲酸(5g,25.4mmol)加入到50ml的SOCl2中，滴加4滴DMF，90℃下剧烈搅拌3h。TLC检测反应进度,待底物完全反应后,产物浓缩后得到的黄色固体3-甲氧基-4-硝基苯甲酰氯直接用于下一步反应。
步骤2：在0℃下，将DIPEA(4mL,25.1mmol)与吗啉(2.2g,25.1mmol)加入到3-甲氧基-4-硝基苯甲酰氯(5.5g,25.4mmol)的50ml THF溶液中，室温下剧烈搅拌3h。TLC检测反应进度,待底物完全反应后,加水稀释,调节pH至碱性,用乙酸乙酯/水体系萃取三次,分离出有机层,经水和饱和食盐水洗涤,无水硫酸钠干燥后,减压浓缩得到化合物(3-甲氧基-4-硝基苯基)(吗啉代)甲酮(6.6g,98％)。MS m/z(ESI):267.1[M+H]+
步骤3：将Pd/C(200mg)加入到(3-甲氧基-4-硝基苯基)(吗啉代)甲酮(1g,3.76mmol)的50ml甲醇溶液中。室温下,在H2氛围中剧烈搅拌16h。反应结束后,过滤,滤 液浓缩,得化合物(4-氨基-3-甲氧基苯基)(吗啉代)甲酮(900mg,99％),产物直接用于下一步。MS m/z(ESI):237.2[M+H]+
中间体15：(4-氨基-3-甲氧基苯基)(4-甲基哌嗪-1-基)甲酮的制备
Figure PCTCN2015073045-appb-000025
步骤1：将4-硝基-3-甲氧基苯甲酸(3g,15.2mmol)加入到50ml的SOCl2中，滴加4滴DMF，90℃下剧烈搅拌3h。TLC检测反应进度,待底物完全反应后,产物浓缩后得到的黄色固体3-甲氧基-4-硝基苯甲酰氯直接用于下一步反应。
步骤2：在0℃下,将N-甲基哌嗪(1.5g,15.2mmol)加入到3-甲氧基-4-硝基苯甲酰氯(3.3g,15.2mmol)的50ml二氯甲烷溶液中，室温下剧烈搅拌3h。TLC检测反应进度,待底物完全反应后,加水稀释,调节pH至碱性,用乙酸乙酯/水体系萃取三次,分离出有机层,经水和饱和食盐水洗涤,无水硫酸钠干燥后,减压浓缩得到化合物(3-甲氧基-4-硝基苯基)(4-甲基哌嗪-1-基)甲酮(3.0g,71％)。MS m/z(ESI):280.0[M+H]+
步骤3：将Pd/C(150mg)加入到(3-甲氧基-4-硝基苯基)(4-甲基哌嗪-1-基)甲酮(1g,3.58mmol)的50ml甲醇溶液中。室温下,在氢气氛围中剧烈搅拌16h。TLC检测反应结束后,过滤,滤液浓缩,得(4-氨基-3-甲氧基苯基)(4-甲基哌嗪-1-基)甲酮(780mg,87％),产物直接用于下一步。
实施例1：N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-1)的制备
Figure PCTCN2015073045-appb-000026
步骤1：2-氯-4-嘧啶异氰酸酯的合成
在0℃下，将4-氨基-2-氯嘧啶(4.8g,37.0mmol)加入到三光气(5.5g,18.5mmol)的200ml THF溶液中，然后缓慢滴加N,N-二异丙基乙胺(DIEA)(6.23g,48.2mmol)。室温下剧烈搅拌4h。TLC检测反应进度，待底物完全反应后，产物2-氯-4-嘧啶异氰酸酯直接用于下一步反应。
步骤2：1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲的合成
室温下，将三乙胺(TEA)(11g,110mmol)与3-硝基苯胺(5.6g,40.3mmol)加入到2- 氯-4-嘧啶异氰酸酯(5.7g,36.6mmol)的200ml THF溶液中，室温下剧烈搅拌20h。反应结束后，向反应体系加入500ml水，有黄色固体析出。以500ml乙酸乙酯洗涤黄色固体，然后以30ml乙酸乙酯打浆，过滤得到化合物1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(2.8g)。产率：30％，纯度：86％，MS m/z(ESI):294.1[M+H]+
步骤3：1-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将2-甲氧基-4-(4-甲基哌嗪-1-基)苯胺(739mg,3.34mmol)与三氟乙酸(TFA)(1.14g,10mmol)加入到化合物1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(980mg,3.34mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得化合物1-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(700mg)，产物直接用于下一步。产率：44％，纯度：74％，MS m/z(ESI):479.2[M+H]+
步骤4：1-(3-氨基苯基)-3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲的合成
将二氯化锡(554mg,2.93mmol)加入到1-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(700mg,1.46mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体化合物1-(3-氨基苯基)-3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲(300mg)，产品直接用于下一步。产率：45％，纯度：70％，MS m/z(ESI):449.2[M+H]+
步骤5：N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-1)的合成
将三乙胺(70mg,0.67mmol)加入到1-(3-氨基苯基)-3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲(100mg,0.23mmol)的2ml THF与2mlDMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(25mg,0.27mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得化合物N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-1)(2.4mg)。产率：2％，纯度：100％，MS m/z(ESI):503.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.52(s,1H),10.14(s,1H),9.61(s,1H),8.30(d,1H),8.10(d,1H),7.94(s,1H),7.48(d,J＝8.7Hz,1H),7.35(d,J＝8.7Hz,1H),7.15(t,J＝5.5Hz,1H),6.64(s,2H)，6.47–6.43(m,2H)，6.26(d,J＝16.9Hz,1H)，5.75(d,J＝12.0Hz,1H)，5.33(m,1H)，3.75(s,3H)，3.11(m,4H)，2.47(m,4H)，2.33(s,3H)。
实施例2：N-(3-(3-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-2)的制备
Figure PCTCN2015073045-appb-000027
步骤1：1-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将4-(4-甲基哌嗪-1-基)苯胺(392mg,2.00mmol)与三氟乙酸(700mg,6.0mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(600mg,2.00mmol)的14ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以50ml乙醇洗涤棕色固体，得1-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(500mg)，产物直接用于下一步。产率：56％，纯度：72％，MS m/z(ESI):449.2[M+H]+
步骤2：1-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(755mg,3.33mmol)加入到1-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(500mg,1.11mmol)的10ml甲醇与5ml DMSO的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(180mg)，产品直接用于下一步。产率：39％，纯度：72％，MS m/z(ESI):419.2[M+H]+
步骤3：N-(3-(3-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-2)的合成
将三乙胺(131mg,1.29mmol)加入到(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(180mg,0.43mmol)的5ml THF溶液中，0℃下剧烈搅拌。将丙烯酰氯(60mg,0.65mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得化合物N-(3-(3-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-2)(12.4mg)。产率：6％，纯度：95％，MS m/z(ESI):473.0[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.31(s,1H),10.17(s,1H),9.55(s,1H),9.20(s,1H),8.18(d,J＝5.6Hz,1H),7.94(s,1H),7.46(dd,J＝13.8,9.1Hz,3H),7.19(t,J＝8.1Hz,1H),6.89(d,J＝9.0Hz,2H),6.75(d,J＝5.6Hz,2H),6.47(dd,J＝17.0,10.1Hz,1H),6.26(dd,J＝16.9,2.0Hz,1H),5.75(dd,J＝10.1,2.0Hz,1H),3.09(m,4H),2.53(m,4H),2.33(s,3H)。
实施例3：N-(3-(3-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-3)的制备
Figure PCTCN2015073045-appb-000028
步骤1：1-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将2-氨基-5-甲氧基吡啶(414mg,3.34mmol)与三氟乙酸(1.14g,10mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(980mg,3.34mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得化合物1-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)-3-(3-硝基苯基)(650mg)，产物直接用于下一步。产率：51％，纯度：78％，MS m/z(ESI):382.1[M+H]+
步骤2：1-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(1157mg,5.13mmol)加入到1-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(650mg,1.71mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(500mg)，产品直接用于下一步。产率：83％，纯度：78％，MS m/z(ESI):352.1[M+H]+
步骤3：N-(3-(3-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-3)的合成
将三乙胺(172mg,1.7mmol)加入到1-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg,0.56mmol)的2ml THF与2mlDMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(62mg,0.68mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-3)(26.3mg)。产率：2％，纯度：94.5％，MS m/z(ESI):406.0[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.17(s,1H),9.85(s,1H),9.49(s,1H),9.37(s,1H),8.49(d,J＝2Hz,1H),8.21(d,J＝5.6Hz,1H),7.90(t,J＝2.4Hz,2H),7.44(d,J＝8Hz,1H),7.22(t,J＝8.4Hz,1H),6.95(d,J＝5.2Hz,2H),6.78(d,J＝8.4Hz,1H),6.45(t,J＝12.4Hz,1H),6.25(dd,J＝2Hz,1H),5.75(dd,J＝1.6Hz,1H),3.81(s,3H)。
实施例4：N-(3-(3-(2-((3-氯-4-((3-氟苄基)氧)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-4)的制备
Figure PCTCN2015073045-appb-000029
步骤1：1-(2-((3-氯-4-((3-氟苄基)氧基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将3-氯-4-((3-氟苄基)氧)苯胺(414mg,3.34mmol)与三氟乙酸(1.14g,10mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(980mg,3.34mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((3-氯-4-((3-氟苄基)氧基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(650mg)，产物直接用于下一步。产率：51％，纯度：78％，MS m/z(ESI):382.1[M+H]+
步骤2：1-(2-((3-氯-4-((3-氟苄基)氧基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(1157mg,5.13mmol)加入到1-(2-((3-氯-4-((3-氟苄基)氧基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(650mg,1.71mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((3-氯-4-((3-氟苄基)氧基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(500mg)，产品直接用于下一步。产率：83％，纯度：78％，MS m/z(ESI):352.1[M+H]+
步骤3：N-(3-(3-(2-((3-氯-4-((3-氟苄基)氧)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-4)的合成
将三乙胺(172mg,1.7mmol)加入到1-(2-((3-氯-4-((3-氟苄基)氧基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg,0.56mmol)的2ml THF与2mlDMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(62mg,0.68mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((3-氯-4-((3-氟苄基)氧)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-4)(26.3mg)。产率：2％，纯度：100％，MS m/z(ESI):533.1[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.17(s,1H),9.85(s,1H),9.49(s,1H),9.37(s,1H),8.49(d,J＝2Hz,1H),8.21(d,J＝5.6Hz,1H),7.90(t,J＝2.4Hz,2H),7.44(d,J＝8Hz,1H),7.22(t,J＝8.4Hz,1H),6.95(d,J＝5.2Hz,2H),6.78(d,J＝8.4Hz,1H),6.45(t,J＝12.4Hz,1H),6.25(t,J＝2Hz,1H),5.75(t,J＝1.6Hz,1H),3.81(s,3H)。
实施例5：N-(3-(3-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-5)的制备
Figure PCTCN2015073045-appb-000030
步骤1：1-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将2-甲氧基-4(4-(4-甲基哌嗪)哌啶)苯胺(338mg,1.12mmol)与三氟乙酸(380mg,3.33mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(328mg,1.12mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(281mg)，产物直接用于下一步。产率：45％，纯度：79％，MS m/z(ESI):562.1[M+H]+
步骤2：1-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(339mg,1.5mmol)加入到1-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(281mg,0.5mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg)，产品直接用于下一步。产率：76％，纯度：82％，MS m/z(ESI):532.1[M+H]+
步骤3：N-(3-(3-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-5)的合成
将三乙胺(114mg,1.13mmol)加入到1-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg,0.38mmol)的2ml THF与2mlDMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(42mg,0.46mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-5)(3.3mg)。产率：1.5％，纯度：94％，MS m/z(ESI):586.1[M+H]+；1H NMR(400MHz,DMSO-d6):δ11.02(s,1H),10.13(s,1H),8.28(s,1H),8.09(d,J＝5.6Hz,1H),7.89(s,1H),7.48(d,J＝8.4Hz,1H),7.31(t,J＝9.2Hz,1H),7.13(m,J＝7.2Hz,1H),6.(m,6H),6.27(t,J＝2Hz,1H),5.74(m,1H),3.74(s,3H),3.66(m,1H),2.66(m,4H),2.33(m,4H),2.14(s,3H),1.82(m,4H),1.50(m,4H)。
实施例6：N-(3-(3-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-6)的制备
Figure PCTCN2015073045-appb-000031
步骤1：1-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将6-吗啉代吡啶-3-胺(427mg,2.38mmol)与三氟乙酸(815mg,7.15mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(700mg,2.38mmol)的8ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以柱层析纯化得棕色固体1-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(120mg)。产率：10％，纯度：85％，MS m/z(ESI):436.1[M+H]+
步骤2：1-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(190mg,0.82mmol)加入到1-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(120mg,0.27mmol)的3ml甲醇与3mlDMF的混合溶液中，90℃下剧烈搅拌7h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(120mg)，产品直接用于下一步。产率：90％，纯度：70％，MS m/z(ESI):406.2[M+H]+
步骤3：N-(3-(3-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-6)的合成
将三乙胺(90mg,0.9mmol)加入到1-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(120mg,0.30mmol)的5ml THF溶液中，0℃下剧烈搅拌。将丙烯酰氯(27mg,0.30mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-6)(1.66mg)。产率：2％，纯度：97％，MS m/z(ESI):461.3[M+H]+。
实施例7：N-(3-(3-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-7)的制备
Figure PCTCN2015073045-appb-000032
步骤1：1-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将2,4-二甲氧基苯胺(261mg,1.7mmol)与三氟乙酸(582mg,5.1mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(500mg,1.7mmol)的8ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(120mg)，产物直接用于下一步。产率：18％，纯度：76％，MS m/z(ESI):411.1[M+H]+
步骤2：1-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(231mg,1.0mmol)加入到1-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(140mg,0.34mmol)的3ml甲醇与3mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(100mg)，产品直接用于下一步。产率：90％，纯度：69％，MS m/z(ESI):381.2[M+H]+
步骤3：N-(3-(3-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-7)的合成
将三乙胺(100mg,0.9mmol)加入到1-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(100mg,0.3mmol)的5ml THF溶液中，0℃下剧烈搅拌。将丙烯酰氯(30mg,0.3mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-7)(19.08mg)。产率：15％，纯度：100％，MS m/z(ESI):435.3[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.45(s,1H),10.12(s,1H),9.59(s,1H),8.35(s,1H),8.11(d,J＝5.6Hz,1H),7.88(s,1H),7.45(dd,J＝14.0,8.5Hz,2H),7.12(t,J＝8.1Hz,1H),6.69–6.35(m,5H),6.25(dd,J＝17.0,1.9Hz,1H),5.74(dd,J＝10.1,1.9Hz,1H),3.75(s,3H),3.73(s,3H)。
实施例8：N-(3-(3-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-8)的制备
Figure PCTCN2015073045-appb-000033
步骤1：1-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将3-氯-4-氟苯胺(162mg,1.12mmol)与三氟乙酸(380mg,3.33mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(328mg,1.12mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(230mg)，产物直接用于下一步。产率：51％，纯度：74％，MS m/z(ESI):403.1[M+H]+
步骤2：1-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(388mg,1.72mmol)加入到1-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(230mg,0.57mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg)，产品直接用于下一步。产率：94％，纯度：80％，MS m/z(ESI):373.1[M+H]+
步骤3：N-(3-(3-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-8)的合成
将三乙胺(163mg,1.61mmol)加入到1-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg,0.54mmol)的2ml THF与2ml DMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(58mg,0.65mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-8)(3.7mg)。产率：1.6％，纯度：96.8％，MS m/z(ESI):427.0[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.17(s,1H),9.91(s,1H),9.62(s,1H),9.47(s,1H),8.28(d,J＝6Hz,1H),8.01(m,1H),7.93(s,1H),7.64(m,1H),7.42(d,J＝8.4Hz,1H),7.33(t,J＝8.8Hz,1H),7.25(t,J＝8Hz,1H),7.08(m,2H),6.47(m,1H),6.26(m,1H),5.75(m,1H)。
实施例9：N-(3-(3-(2-((2-甲氧基-4-(4-吗啉哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-9)的制备
Figure PCTCN2015073045-appb-000034
步骤1：1-(2-((2-甲氧基-4-(4-吗啉代-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将2-甲氧基-4(4-吗啉哌啶)苯胺(326mg,1.12mmol)与三氟乙酸(380mg,3.33mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(328mg,1.12mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((2-甲氧基-4-(4-吗啉代-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(270mg)，产物直接用于下一步。产率：49％，纯度：65％，MS m/z(ESI):549.1[M+H]+
步骤2：1-(2-((2-甲氧基-4-(4-吗啉代-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(339mg,1.5mmol)加入到1-(2-((2-甲氧基-4-(4-吗啉代-1-基)苯基)氨 基)嘧啶-4-基)-3-(3-硝基苯基)脲(270mg,0.49mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((2-甲氧基-4-(4-吗啉代-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg)，产品直接用于下一步。产率：79％，纯度：75％，MS m/z(ESI):519.1[M+H]+
步骤3：N-(3-(3-(2-((2-甲氧基-4-(4-吗啉哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-9)的合成
将三乙胺(114mg,1.13mmol)加入到1-(2-((2-甲氧基-4-(4-吗啉代-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg,0.38mmol)的2ml THF与2mlDMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(42mg,0.46mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((2-甲氧基-4-(4-吗啉哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-9)(70.98mg)。产率：32.7％，纯度：99％，MS m/z(ESI):573.3[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.28(s,1H),10.19(s,1H),10.10(s,1H),9.87(s,1H),8.12(d,J＝8Hz,1H),7.94(s,1H),7.50(d,J＝8Hz,1H),7.36(d,J＝8Hz,1H),7.18(t,J＝8Hz,1H),6.75(s,1H),6.69(d,J＝4Hz,1H),6.28(m,3H),6.25(m,1H),5.75(m,1H),4.03(d,J＝12Hz,2H),3.89(d,J＝12Hz,2H),3.89(s,3H),3.69(t,J＝12Hz,2H),3.51(d,J＝12Hz,2H),3.38(s,1H),3.14(m,2H),2.65(m,2H),2.15(d,J＝12Hz,2H),1.70(m,2H)。
实施例10：N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-10)的制备
Figure PCTCN2015073045-appb-000035
步骤1：1-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将1-(4-氨基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺(279mg,1.12mmol)与三氟乙酸(380mg,3.33mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(328mg,1.12mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(230mg)，产物直接用于下一步。产率：41％，纯度：65％，MS m/z(ESI):507.1[M+H]+
步骤2：1-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(305mg,1.35mmol)加入到1-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(230mg,0.45mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(180mg)，产品直接用于下一步。产率：84％，纯度：78％，MS m/z(ESI):477.2[M+H]+
步骤3：N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-10)的合成
将三乙胺(114mg,1.13mmol)加入到1-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(180mg,0.38mmol)的2ml THF与2mlDMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(42mg,0.46mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得化合物N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-10)(25.36mg)。产率：12.6％，纯度：99％，MS m/z(ESI):531.3[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.25(m,3H),10.00(s,1H),9.49(m,1H),8.14(d,J＝4Hz,1H),7.93(s,1H),7.48(m,2H),7.20(m,1H),7.82(s,1H),6.70(d,J＝4Hz,1H),6.57(m,2H),6.28(t,J＝12Hz,1H),5.74(t,J＝12Hz,1H),4.04(d,J＝20Hz,2H),3.99(s,3H),3.33(m,1H),2.79(s,6H),2.70(t,J＝12Hz,2H),2.08(d,J＝12Hz,2H),1.72(m,2H)。
实施例11：N-(3-(3-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-11)的制备
Figure PCTCN2015073045-appb-000036
步骤1：1-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将4-(哌啶-1-基)-苯胺(197mg,1.12mmol)与三氟乙酸(380mg,3.33mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(328mg,1.12mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(230mg)，产物直接用于下一步。产率：47％，纯度：80％，MS m/z(ESI):434.2[M+H]+
步骤2：1-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲合成
将二氯化锡(380mg,1.59mmol)加入到1-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(230mg,0.53mmol)的60ml甲醇与40ml DMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合 物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg)，产品直接用于下一步。产率：94％，纯度：85％，MS m/z(ESI):404.2[M+H]+
步骤3：N-(3-(3-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-11)的合成
将三乙胺(155mg,1.50mmol)加入到1-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(200mg,0.50mmol)的2ml THF与2ml DMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(54mg,0.6mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-11)(1.80mg)。产率：0.8％，纯度：93.7％，MS m/z(ESI):458.3[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.28(s,1H),10.14(s,1H),9.52(s,1H),9.17(s,1H),8.16(d,J＝5.2Hz,1H),7.91(s,1H),7.45(m,3H),7.18(t,J＝7.6Hz,1H),6.85(d,J＝8.8Hz,2H),6.72(d,J＝5.2Hz,1H),6.45(m,1H),6.28(m,1H),5.76(m,1H),3.02(m,4H),1.61(m,4H),1.51(m,2H)。
实施例12：N-(3-(3-(2-((4-(二甲氨基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-12)的制备
Figure PCTCN2015073045-appb-000037
步骤1：1-(2-((4-(二甲基氨基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲的合成
将4-氨基-N,N-二甲基苯胺(152mg,1.12mmol)与三氟乙酸(380mg,3.33mmol)加入到1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(328mg,1.12mmol)的20ml正丁醇溶液中，130℃下剧烈搅拌7h。反应结束后，体系冷却至室温，过滤得棕色固体。以30ml乙醇洗涤棕色固体，得1-(2-((4-(二甲基氨基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(221mg)，产物直接用于下一步。产率：50％，纯度：78％，MS m/z(ESI):394.3[M+H]+
步骤2：1-(2-((4-(二甲基氨基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将二氯化锡(380mg,1.68mmol)加入到1-(2-((4-(二甲基氨基)苯基)氨基)嘧啶-4-基)-3-(3-硝基苯基)脲(221mg,0.56mmol)的60ml甲醇与40mlDMF的混合溶液中，90℃下剧烈搅拌4h。反应结束后，体系冷却至室温，加入等当量碳酸氢钠固体。将混合物剧烈搅拌30min，滤去不溶物。将滤液减压浓缩得棕色固体1-(2-((4-(二甲基氨基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(195mg)，产品直接用于下一步。产率：96％，纯度：79％，MS m/z(ESI):364.2[M+H]+
步骤3：N-(3-(3-(2-((4-(二甲氨基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-12)的合成
将三乙胺(163mg,1.61mmol)加入到1-(2-((4-(二甲基氨基)苯基)氨基)嘧啶-4-基)-3-(3-氨基苯基)脲(195mg,0.54mmol)的2ml THF与2ml DMF的混合溶液中，0℃下剧烈搅拌。将丙烯酰氯(58mg,0.65mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌4h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((4-(二甲氨基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-12)(14.20mg)。产率：6.3％，纯度：99.7％，MS m/z(ESI):418.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.42(s,1H),10.14(s,1H),9.56(s,1H),9.09(s,1H),8.14(d,J＝5.2Hz,1H),7.92(s,1H),7.47(d,J＝8Hz,1H),7.35(d,J＝8.8Hz,2H),7.13(t,J＝8Hz,1H),6.67(m,4H),6.45(m,1H),6.25(m,1H),5.75(m,1H),2.83(s,6H)。
实施例13：N-(3-(3-(2-((5-吗啉代吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-13)的制备
Figure PCTCN2015073045-appb-000038
步骤1：1-(2-氯嘧啶-4-基)-3-(3-氨基苯基)脲的合成
将1-(2-氯嘧啶-4-基)-3-(3-硝基苯基)脲(6.0g,20mmol)置于250mL的单口反应瓶中，加入THF/水(75mL/50mL)混合溶液使底物溶解。在室温下，向搅拌的反应瓶中依次加入氯化铵(5.3g,100mmol)和还原铁粉(8.9g,160mmol)，随后将反应体系加热至65℃并保持搅拌5h。TLC检测反应进度，待底物完全反应后，过滤除去多余的铁粉，滤饼用乙酸乙酯淋洗三次。滤液用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到1-(2-氯嘧啶-4-基)-3-(3-氨基苯基)脲(5.4g)，直接用于下一步反应。
步骤2：N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺的合成
将1-(2-氯嘧啶-4-基)-3-(3-氨基苯基)脲(5.4g,20mmol)置于100mL的单口反应瓶中，加入二氯甲烷(50mL)使底物溶解。在0℃下，向搅拌的反应瓶中依次加入三乙胺(4.3mL,30mmol)和丙烯酰氯(2.0mL,24mmol)，随后将反应体系维持在0℃并持续搅拌1h。TLC检测反应进度，待底物完全反应后，反应液用THF/水体系萃取三次，分离出有机层，饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经乙醇回流打浆得N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(5.2g)，直接用于下一 步反应。产率：80％(两步)，纯度：90％，MS m/z(ESI):318.1[M+H]+.
步骤3：N-(3-(3-(2-((5-吗啉代吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-13)的合成
室温下，将Pd2(dba)3(30mg,0.03mmol)，BINAP(2,2'-双二苯膦基-1,1'-联萘)(40mg,0.06mmol)，t-BuONa(91mg,0.96mmol)和5-吗啉代吡啶-2-胺(102mg,0..57mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)的5ml二氧六环溶液中，氮气保护下，封管120℃剧烈搅拌8h。反应结束后，过滤，减压浓缩滤液得黄色固体，以制备液相纯化得到N-(3-(3-(2-((5-吗啉代吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-13)(5.17mg)。产率：3％，纯度：97.67％，MS m/z(ESI):461.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.57(s,1H),10.17(s,1H),9.80(s,1H),9.70(s,1H),8.23(d,J＝5.6Hz,1H),7.86(d,J＝12.3Hz,3H),7.50(d,J＝8.0Hz,1H),7.44–7.33(m,1H),7.25(t,J＝8.1Hz,1H),7.10(d,J＝7.8Hz,1H),6.73(d,J＝5.5Hz,1H),6.46(dd,J＝17.0,10.2Hz,1H),6.26(d,J＝16.9Hz,1H),5.75(d,J＝10.3Hz,1H),3.76–3.70(m,4H),3.06–3.00(m,4H)。
实施例14：N-(3-(3-(2-((2-甲氧基-4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-14)的制备
Figure PCTCN2015073045-appb-000039
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和2-甲氧基-4-吗啉代苯胺(131mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到N-(3-(3-(2-((2-甲氧基-4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-14)5(15.85mg)。产率：5％，纯度：100％，MS m/z(ESI):490.7[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.47(s,1H),10.13(s,1H),9.59(s,1H),8.29(s,1H),8.10(d,J＝5.6Hz,1H),7.92(s,1H),7.47(d,J＝8.3Hz,1H),7.39(d,J＝8.6Hz,1H),7.14(t,J＝8.1Hz,1H),6.60(dd,J＝12.7,4.0Hz,2H),6.49–6.42(m,3H),6.25(dd,J＝17.0,2.0Hz,1H),5.74(dd,J＝10.1,2.0Hz,1H),3.81–3.70(m,7H),3.13–3.02(m,4H)。
实施例15：N-(3-(3-(2-((2-甲氧基-4-(吗啉-4-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-15)的制备
Figure PCTCN2015073045-appb-000040
将(4-氨基-3-甲氧基苯基)(吗啉代)甲酮(200mg,0.63mmol)与TFA(216mg，1.89mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(149mg,0.63mmol)的4ml正丁醇溶液中，120℃下剧烈搅拌6h。反应结束后，体系冷却至室温，加入适量的碳酸氢钠饱和溶液，用乙酸乙酯/水体系萃取三次，有机层减压浓缩得到粗产物。经制备液相分离纯化得N-(3-(3-(2-((2-甲氧基-4-(吗啉-4-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-15)(66.7mg)。产率：21％，纯度：99％，MS m/z(ESI):518.2[M+H]+；1H NMR(400MHz,DMSO)δ10.18(s,1H),9.90(s,1H),9.76(s,1H),8.26(d,J＝5.9Hz,1H),8.09(d,J＝8.1Hz,1H),7.93(s,1H),7.41(d,J＝7.6Hz,1H),7.25(t,J＝8.1Hz,1H),7.05(ddd,J＝9.7,7.2,1.6Hz,5H),6.46(dd,J＝16.9,10.1Hz,1H),6.26(dd,J＝17.0,1.9Hz,1H),5.76(dd,J＝10.1,1.9Hz,1H),3.56(d,J＝36.0Hz,11H)。
实施例16：N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-16)的制备
Figure PCTCN2015073045-appb-000041
将(4-氨基-3-甲氧基苯基)(4-甲基哌嗪-1-基)甲酮(200mg,0.63mmol)与TFA(216mg，1.89mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(157mg,0.63mmol)的4ml正丁醇溶液中，120℃下剧烈搅拌6h。反应结束后，体系冷却至室温，加入适量的碳酸氢钠饱和溶液，用乙酸乙酯/水体系萃取三次，有机层减压浓缩得到粗产物。经制备液相分离纯化得N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-16)(17.31mg)。产率：5％，纯度：100％，MS m/z(ESI):531.2[M+H]+；1H NMR(400MHz,CDCl3)δ10.17(s,1H),9.91(s,1H),9.54(s,1H),8.26(d,J＝5.8Hz,2H),8.15(t,J＝8.6Hz,1H),7.92(s,1H),7.42(d,J＝8.2Hz,1H),7.25(t,J＝8.1Hz,1H),7.15–6.89(m,5H),6.47(dd,J＝16.9,10.1Hz,1H),6.27(dd,J＝17.0,1.9Hz,1H),5.93–5.44(m,1H),3.89(s,4H),3.44(d,J＝57.3Hz,5H),2.32(s,5H),2.21(s,4H)。
实施例17：4-((4-(3-(3-丙烯酰基酰胺苯基)脲基)嘧啶-2-基)氨基)-N,N-二乙基-3-甲氧基苯甲酰胺(cpd-17)的制备
Figure PCTCN2015073045-appb-000042
将4-氨基-N,N-二乙基-3-甲氧基苯甲酰胺(200mg,0.63mmol)与TFA(216mg，1.89mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(140mg,0.63mmol)的4ml正丁醇溶液中，110℃下剧烈搅拌6h。反应结束后，体系冷却至室温，加入适量的碳酸氢钠饱和溶液，用乙酸乙酯/水体系萃取三次，有机层减压浓缩得到粗产物。经制备板和Combi-Flash反相柱层析纯化得到4-((4-(3-(3-丙烯酰基酰胺苯基)脲基)嘧啶-2-基)氨基)-N,N-二乙基-3-甲氧基苯甲酰胺(cpd-17)(8.14mg)。产率：2％，纯度：95％，MS m/z(ESI):504.2[M+H]+；1H NMR(400MHz,DMSO)δ10.16(s,1H),9.92(s,1H),9.53(s,1H),8.60–8.21(m,2H),8.10(t,J＝8.7Hz,1H),7.90(s,1H),7.42(d,J＝8.0Hz,1H),7.25(t,J＝8.1Hz,1H),7.01(s,4H),6.46(dd,J＝17.0,10.2Hz,1H),6.26(d,J＝16.9Hz,1H),5.75(d,J＝10.4Hz,1H),3.88(s,3H),3.29(s,4H),1.12(s,6H)。
实施例18：N-(3-(3-(2-((4-(2-甲氧基乙氧基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-18)的制备
Figure PCTCN2015073045-appb-000043
将4-(2-甲氧基乙氧基)苯胺(105mg,0.63mmol)与三氟乙酸(216mg,1.9mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)的5ml正丁醇溶液中，110℃下剧烈搅拌4h。反应结束后，体系冷却至室温，减压浓缩得棕色固体。经制备液相纯化得到N-(3-(3-(2-((4-(2-甲氧基乙氧基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-18)(4.70mg)。产率：2％，纯度：96.2％，MS m/z(ESI):449.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.16(d,J＝17.1Hz,2H),9.49(s,1H),9.24(s,1H),8.18(d,J＝5.6Hz,1H),7.92(s,1H),7.48(dd,J＝20.8,8.4Hz,3H),7.19(t,J＝8.1Hz,1H),6.84(dd,J＝28.6,7.3Hz,4H),6.46(dd,J＝17.0,10.1Hz,1H),6.25(dd,J＝17.0,2.0Hz,1H),5.75(dd,J＝10.1,2.0Hz,1H),4.06–4.00(m,2H),3.73–3.58(m,2H),3.32(s,3H)。
实施例19：N-(3-(3-(2-((2-甲氧基-4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-19)的制备
Figure PCTCN2015073045-appb-000044
将2-甲氧基-4-(哌啶-1-基)苯胺(130mg,0.63mmol)与三氟乙酸(216mg,1.9mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)的4ml正丁醇溶液中，110℃下剧烈搅拌4h。反应结束后，体系冷却至室温，减压浓缩得棕色固体。经制备液相纯化得到N-(3-(3-(2-((2-甲氧基-4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-19)(41.19mg)。产率：14％，纯度：98.65％，MS m/z(ESI):488.3[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.47(s,1H),10.14(d,J＝18.9Hz,1H),9.58(s,1H),8.38–8.19(m,1H),8.10(d,J＝5.5Hz,1H),7.90(s,1H),7.48(d,J＝7.9Hz,1H),7.35(t,J＝7.9Hz,1H),7.15(dd,J＝17.7,9.7Hz,1H),6.66–6.55(m,2H),6.54–6.36(m,3H),6.25(dd,J＝17.0,1.9Hz,1H),5.74(dd,J＝10.1,1.9Hz,1H),3.74(s,3H),3.15–3.05(m,4H),1.66–1.50(m,6H)。
实施例20：N-(3-(3-(2-((4-(4-(2-氟乙基)哌嗪-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-20)的制备
Figure PCTCN2015073045-appb-000045
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和4-(4-(2-氟乙基)哌嗪-1-基)-2-甲氧基苯胺(157mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到N-(3-(3-(2-((4-(4-(2-氟乙基)哌嗪-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-20)(22.42mg)。产率：6.8％，纯度：100％，MS m/z(ESI):535.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.48(s,1H),10.13(s,1H),9.58(s,1H),8.28(s,1H),8.10(d,J＝5.6Hz,1H),7.93(s,1H),7.47(d,J＝8.4Hz,2H),7.35(d,J＝9.2Hz,1H),7.13(t,J＝8Hz,1H),6.61(m,2H),6.47(m,3H),6.25(m,1H),5.75(m,1H),4.66(m,1H),4.52(m,1H),3.75(s,1H),3.13(m,4H),2.73(m,1H),2.65(m,1H),2.51(m,4H)。
实施例21：N-(3-(3-(2-((4-(二乙基氨基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-21)的制备
Figure PCTCN2015073045-appb-000046
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和N,N-二乙基-3-甲氧基苯-1,4-二胺(157mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到N-(3-(3-(2-((4-(二乙基氨基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-21)(86.24mg)。产率：29.2％，纯度：100％，MS m/z(ESI):476.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.66(s,1H),10.12(s,1H),9.61(s,1H),8.24(s,1H),8.07(d,J＝5.6Hz,1H),7.94(s,1H),7.49(d,J＝8Hz,1H),7.18(d,J＝8.4Hz,1H),7.07(t,J＝8Hz,1H),6.28(m,6H),5.74(m,1H),3.72(t,3H),3.32(m,4H),1.08(m,6H)。
实施例22：N-(3-(3-(2-((4-(4-(二乙基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-22)的制备
Figure PCTCN2015073045-appb-000047
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和1-(4-氨基-3-甲氧基苯基)-N,N-二乙基哌啶-4-胺(157mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到N-(3-(3-(2-((4-(4-(二乙基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-22)(26.31mg)。产率：7.6％，纯度：100％，MS m/z(ESI):559.3[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.45(s,1H), 10.12(s,1H),9.61(s,1H),8.25(s,1H),8.09(d,J＝8Hz,1H),7.88(s,1H),7.48(d,8Hz,1H),7.34(d,J＝8.4Hz,1H),7.14(m,1H),6.56(m,5H),6.43(m,1H),5.73(m,1H),3.74(s,3H),3.65((d,J＝12Hz,2H),2.62(m,4H),2.55(s,3H),1.75(d,J＝11.2Hz,2H),1.51(m,2H),0.97(t,J＝7.2Hz,6H)。
实施例23：N-(3-(3-(2-((4-(4-乙基哌嗪-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-23)的制备
Figure PCTCN2015073045-appb-000048
将4-(4-乙基哌嗪-1-基)-2-甲氧基苯胺(150mg,0.63mmol)与三氟乙酸(216mg,1.9mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)的3ml正丁醇溶液中，110℃下剧烈搅拌3h。反应结束后，体系冷却至室温，减压浓缩得棕色固体。经制备液相纯化得到N-(3-(3-(2-((4-(4-乙基哌嗪-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-23)(60.25mg)。产率：19％，纯度：98.21％，MS m/z(ESI):517.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.48(s,1H),10.12(s,1H),9.58(s,1H),8.28(s,1H),8.10(d,J＝5.6Hz,1H),7.93(s,1H),7.48(d,J＝8.2Hz,1H),7.36(t,J＝7.9Hz,1H),7.13(t,J＝8.1Hz,1H),6.59(dd,J＝13.9,3.8Hz,2H),6.51–6.31(m,3H),6.25(dd,J＝17.0,2.0Hz,1H),5.74(dd,J＝10.1,2.0Hz,1H),3.75(s,3H),3.12(s,4H),2.50(s,4H),2.39(d,J＝6.6Hz,2H),1.05(t,J＝7.2Hz,3H)。
实施例24：N-(3-(3-(2-(4-甲苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-24)的制备
Figure PCTCN2015073045-appb-000049
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和4-甲基苯胺(67mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到目标产品N-(3-(3-(2-(4-甲苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-24)(97.43 mg)。产率：40％，纯度：97.42％，MS m/z(ESI):389.1[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.16(s,1H),10.12(s,1H),9.55(s,1H),9.40(s,1H),8.21(d,J＝6Hz,1H),7.91(s,1H),7.53(d,J＝8Hz,2H),7.45(d,J＝8Hz,1H),7.21(t,J＝8Hz,1H),7.09(d,J＝8.4Hz,2H),6.88(m,2H),6.46(m,1H),6.25(m,1H),5.75(m,1H),2.25(s,3H)。
实施例25：N-(3-(3-(2-((5-(4-甲基哌嗪-1-基)吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-25)的制备
Figure PCTCN2015073045-appb-000050
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和5-(4-甲基哌嗪-1-基)吡啶-2-胺(69mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。反应结束后，过滤，减压浓缩滤液得黄色固体，以制备液相纯化得到N-(3-(3-(2-((5-(4-甲基哌嗪-1-基)吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-25)(16.04mg)。产率：4％，纯度：98.37％，MS m/z(ESI):474.1[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.63(s,1H),10.18(s,1H),9.76(s,1H),8.39–8.16(m,2H),7.86(dd,J＝10.8,7.4Hz,3H),7.51(d,J＝8.0Hz,1H),7.37(dd,J＝9.1,2.9Hz,1H),7.25(t,J＝8.1Hz,1H),7.09(d,J＝8.4Hz,1H),6.73(d,J＝5.6Hz,1H),6.57–6.39(m,1H),6.26(dd,J＝17.0,1.9Hz,1H),5.75(dd,J＝10.1,1.9Hz,1H),3.06(d,J＝4.4Hz,4H),2.48–2.40(m,4H),2.22(s,3H)。
实施例26：N-(3-(3-(2-((4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-26)的制备
Figure PCTCN2015073045-appb-000051
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和对氟苯胺(69mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到 粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到N-(3-(3-(2-((4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-26)(8.22mg)。产率：3.4％，纯度：97.42％，MS m/z(ESI):393.1[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.16(s,1H),9.99(s,1H),9.46(d,J＝5.2Hz,1H),7.92(s,1H),7.69(m,2H),7.43(d,J＝8.4Hz,1H),7.23(t,J＝8Hz,1H),7.11(t,J＝8.8Hz,2H),7.00(m,2H),6.47(m,1H),6.26(d,J＝16.8Hz,1H),5.75(m,J＝7.8Hz,1H)。
实施例27：N-(3-(3-(2-((4-乙氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-27)的制备
Figure PCTCN2015073045-appb-000052
将N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.62mmol)和对乙氧基苯胺(85mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热3h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到N-(3-(3-(2-((4-乙氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-27)(38.46mg)。产率：12.7％，纯度：100％，MS m/z(ESI):419.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.20(s,1H),10.14(s,1H),9.50(s,1H),9.23(s,1H),8.18(d,J＝5.6Hz),7.91(s,1H),7.50(m,3H),7.19(t,J＝8Hz,1H),6.82(m,4H),6.25(m,1H),5.74(m,1H),3.95(m,2H),3.30(s,3H),1.31(t,J＝2.8Hz,3H)。
实施例28：N-(3-(3-(2-((2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-28)的制备
Figure PCTCN2015073045-appb-000053
将2-甲氧基苯胺(116mg,0.63mmol)与TFA(216mg，1.89mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)的4ml正丁醇溶液中，110℃下剧烈搅拌6h。反应结束后，体系冷却至室温，加入适量的碳酸氢钠饱和溶液，用乙酸乙酯/水体系萃取三次，有机层减压浓缩得到粗产物。经制备液相分离纯化得 N-(3-(3-(2-((2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-28)(11.68mg)。产率：5％，纯度：100％，MS m/z(ESI):405.1[M+H]+；1H NMR(400MHz,DMSO)δ10.12(d,J＝18.3Hz,1H),9.55(s,1H),8.40–8.10(m,1H),7.90(d,J＝7.5Hz,1H),7.37(t,J＝35.9Hz,1H),7.19(t,J＝8.1Hz,1H),7.13–7.02(m,1H),7.01–6.86(m,1H),6.78(d,J＝6.9Hz,1H),6.46(dd,J＝16.9,10.1Hz,1H),6.26(d,J＝16.9Hz,1H),5.82–5.65(m,1H),3.82(s,2H),2.07(s,1H)。
实施例29：N-(3-(3-(2-((4-氯苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-29)的制备
Figure PCTCN2015073045-appb-000054
将4-氯苯胺(80mg,0.63mmol)与三氟乙酸(216mg,1.9mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)的4ml正丁醇溶液中，110℃下剧烈搅拌3h。反应结束后，体系冷却至室温，减压浓缩得棕色固体。经制备液相纯化得到N-(3-(3-(2-((4-氯苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-29)(9.37mg)。产率：4％，纯度：100.0％，MS m/z(ESI):409.0[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.18(s,1H),9.93(s,1H),9.61(s,1H),9.49(s,1H),8.27(d,J＝5.6Hz,1H),7.93(s,1H),7.75(dd,J＝9.0,2.8Hz,2H),7.32(tt,J＝16.2,8.1Hz,4H),7.04(dd,J＝14.7,6.6Hz,2H),6.46(dd,J＝17.0,10.1Hz,1H),6.26(dd,J＝17.0,1.8Hz,1H),5.75(dd,J＝10.1,1.8Hz,1H)。
实施例30：N-(3-(3-(2-((3,4,5-三甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-30)的制备
Figure PCTCN2015073045-appb-000055
将3,4,5-三甲氧基苯胺(116mg,0.63mmol)与TFA(216mg，1.89mmol)加入到N-(3-(3-(2-氯嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)的4ml正丁醇溶液中，110℃下剧烈搅拌6h。反应结束后，体系冷却至室温，加入适量的碳酸氢钠饱和溶液，用乙酸乙酯/水体系萃取三次，有机层减压浓缩得到粗产物。经制备液相分离纯化得N-(3-(3-(2-((3,4,5-三甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-30)(50.84mg)。产率：17％，纯度：100％，MS m/z(ESI):465.1[M+H]+；1H NMR(400MHz,DMSO)δ10.20(d,J＝31.9Hz,1H),9.50(s,1H),9.34(s,1H),8.24(d,J＝5.6Hz,1H),7.93(s, 1H),7.45(d,J＝8.2Hz,1H),7.21(t,J＝8.1Hz,1H),7.11–6.92(m,2H),6.86(d,J＝5.6Hz,1H),6.46(dd,J＝17.0,10.1Hz,1H),6.26(dd,J＝17.0,1.9Hz,1H),5.75(dd,J＝10.1,2.0Hz,1H),3.74(s,6H),3.61(s,3H)。
实施例31：N-(3-(3-(2-((4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-31)的制备
Figure PCTCN2015073045-appb-000056
将N-(3-(3-(2-氯代嘧啶-4-基)脲基)苯基)丙烯酰胺(200mg,0.63mmol)和4-吗啉基苯胺(112mg,0.62mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(215mg,1.89mmol)，并保持反应体系在110℃下加热6h。待反应结束后，冷却至室温，待底物完全反应后，反应液用乙酸乙酯/水体系萃取三次，分离出有机层，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(10mM NH4HCO3):CH3CN＝65:35～5:95]纯化得到N-(3-(3-(2-((4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺(cpd-31)(4.22mg)。产率：2％，纯度：100％，MS m/z(ESI):460.7[M+H]+；1H NMR(400MHz,DMSO-d6):δ10.28(s,1H),10.16(s,1H),9.22(s,1H),8.17(d,J＝5.6Hz,1H),7.93(s,1H),7.56–7.39(m,3H),7.19(t,J＝8.1Hz,1H),6.89–6.74(m,4H),6.46(dd,J＝16.9,10.1Hz,1H),6.26(dd,J＝17.0,1.9Hz,1H),5.75(dd,J＝10.1,1.9Hz,1H),3.80–3.67(m,4H),3.08–2.97(m,4H)。
实施例32：N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)-3-甲基脲基)苯基)丙烯酰胺(cpd-32)的制备
Figure PCTCN2015073045-appb-000057
步骤1：2-氯-4-甲氨基嘧啶的合成
将30％甲胺醇溶液(7.63g,73.8mmol)加入到2,4-二氯嘧啶(11g,73.8mmol)的 200ml乙醇和80ml甲醇的混合溶液中，80℃下剧烈搅拌3h。反应结束后，体系冷却至室温，减压浓缩，经柱层析纯化得2-氯-4-甲氨基嘧啶(3.4g)，产率：32％，纯度：99％，MS m/z(ESI):144.1[M+H]+
步骤2：N-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)-N-4-甲基嘧啶-2,4-二胺的合成
将1-(4-氨基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺(1.39g,5.57mmol)与三氟乙酸(1.91g,16.72mmol)加入到2-氯-4-甲氨基嘧啶(800mg,5.57mmol)的15ml正丁醇溶液中，130℃下剧烈搅拌2h。反应结束后，体系冷却至室温，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得棕色固体，经柱层析分离纯化得N-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)-N-4-甲基嘧啶-2,4-二胺(1.4g)。产率：24％，纯度：86％，MS m/z(ESI):357.2[M+H]+
步骤3：1-(2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基-3-(3-硝基苯基)脲的合成
将3-硝基苯异氢酸酯(500mg,3mmol)加入到N-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)-N-4-甲基嘧啶-2,4-二胺(700mg,2mmol)的15ml1,2-二氯乙烷(DCE)溶液中，80℃下剧烈搅拌5h。反应结束后，体系冷却至室温，减压浓缩，经柱层析纯化得1-(2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基-3-(3-硝基苯基)脲(100mg)。产率：11％，纯度：74％，MS m/z(ESI):521.2[M+H]+
步骤4：1-(2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基-3-(3-氨基苯基)脲的合成
室温下，将钯碳(10mg,10％wt)加入到1-(2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基-3-(3-硝基苯基)脲(90mg,0.17mmol)的20ml甲醇溶液中，在氢气气氛下室温剧烈搅拌3h。反应结束后，滤去钯碳，将滤液减压浓缩，得到1-(2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基-3-(3-氨基苯基)脲，直接用于下一步反应。产率：92％，纯度：70％，MS m/z(ESI):491.3[M+H]+
步骤5：N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)-3-甲基脲基)苯基)丙烯酰胺(cpd-32)的合成
将三乙胺(60mg,0.55mmol)加入到1-(2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基-3-(3-氨基苯基)脲(90mg,0.18mmol)的2ml二氯甲烷(DCM)溶液中，0℃下剧烈搅拌。将丙烯酰氯(20mg,0.22mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌2h。TLC检测反应进度，待底物完全反应后，加水稀释，用乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经制备液相分离纯化得N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)-3-甲基脲基)苯基)丙烯酰胺(cpd-32)(3.38mg)。产率：4％，纯度：100.0％，MS m/z(ESI):545.0[M+H]+；1H NMR(400MHz,DMSO-d6):δ11.81(s,1H),10.09(s,1H),8.51(s,1H),8.24(s,1H),7.83(s,1H),7.49(s,1H),7.25(s,1H),7.12–7.05(m,1H),6.55–6.24(m,5H),5.75(s,1H),5.32(s,1H),3.71(s,3H),3.56(s,2H),3.29(s,3H),2.72(s,2H),2.19(s,6H),2.00(d,J＝7.3Hz,1H),1.81(s,2H),1.45(s,2H)。
实施例33：3-丙烯酰氨基-N-(2-(2-甲氧基-4-(4-吗啉代-1-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-33)的制备
Figure PCTCN2015073045-appb-000058
步骤1：2-氯-4-异氰酸嘧啶的合成
在0℃下，将4-氨基-2-氯嘧啶(4.8g,37.0mmol)加入到三光气(5.5g,18.5mmol)的200ml THF溶液中，然后缓慢滴加N,N-二异丙基乙胺(6.23g,48.2mmol)。室温下剧烈搅拌4h。TLC检测反应进度，待底物完全反应后，产物直接用于下一步反应。
步骤2：1-((2-氯嘧啶-4-基)氨基甲酰基)哌啶-3-基氨基甲酸叔丁酯的合成
室温下，将三乙胺(10g,97mmol)与3-叔丁氧羰基氨基哌啶(7.1g,35.4mmol)加入到2-氯-4-异氰酸嘧啶(5.0g,32.2mmol)的100ml THF溶液中，室温下剧烈搅拌18h。反应结束后，向反应体系加入150ml乙酸乙酯，以150ml水洗涤有机相，饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经柱层析分离纯化得白色固体化合物(3.5g)。产率：31％，纯度：90％，MS m/z(ESI):356.2[M+H]+。
步骤3：3-氨基-N-(2-氯嘧啶-4-基)哌啶-1-甲酰胺的合成
将1-((2-氯嘧啶-4-基)氨基甲酰基)哌啶-3-基氨基甲酸叔丁酯(3.5g,9.9mmol)溶于50ml盐酸二氧六环溶液中，室温下下剧烈搅拌2h。反应结束后，减压浓缩得白色固体化合物(2.4g)，产物直接用于下一步。产率：94％，纯度：85％，MS m/z(ESI):256.1[M+H]+。
步骤4：3-丙烯酰氨基-N-(2-氯嘧啶-4-基)哌啶-1-甲酰胺的合成
将三乙胺(2.5g,24.6mmol)加入到3-氨基-N-(2-氯嘧啶-4-基)哌啶-1-甲酰胺(2.1g,8.21mmol)的50ml THF溶液中，0℃下剧烈搅拌。将丙烯酰氯(820mg,9.1mmol)，于0℃下，缓慢滴加到反应液中。保持0℃搅拌3h，缓慢升至室温，继续搅拌20h。TLC检测反应进度，反应结束后，向反应体系加入80ml乙酸乙酯，以50ml水洗涤有机相，饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得黄色固体，经柱层析分离纯化得白色固体化合物(640mg)。产率：24％，纯度：92％，MS m/z(ESI):310.1[M+H]+。
步骤5：3-丙烯酰氨基-N-(2-(2-甲氧基-4-(4-吗啉代-1-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-33)的合成
将2-甲氧基-4-(4-吗啉代-1-基)苯胺(141mg,0.48mmol)与三氟乙酸(166mg,1.45mmol)加入到3-丙烯酰氨基-N-(2-氯嘧啶-4-基)哌啶-1-甲酰胺(150mg,0.48mmol)的3ml正丁醇溶液中，120℃下剧烈搅拌2h。反应结束后，体系冷却至室温，减压浓缩得棕色固体。经制备液相纯化得到cpd-33(51.49mg)。产率：20％，纯度：100.0％，MS m/z(ESI):565.3[M+H]+；1H NMR(400MHz,DMSO-d6):δ9.39(d,J＝16.5Hz,1H),8.63–8.28(m,1H),8.02(d,J＝5.9Hz,2H),7.36–7.13(m,1H),6.64(d,J＝2.3Hz,2H),6.50(dd,J＝8.7,2.3Hz,2H),6.07(d,J＝16.9Hz,1H),5.72–5.57(m,1H),4.11–3.65(m,7H),3.64–3.47(m,5H),3.15–2.82(m,1H),2.64(t,J＝12.2Hz,2H),2.48(s,4H),2.24(s,1H),1.86(d,J＝11.2Hz,2H),1.47(d,J＝8.7Hz,5H),1.23(s,2H)。
实施例34：3-丙烯酰氨基-N-(2-(2-甲氧基-4-(4-甲基哌嗪-1-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-34)的制备
Figure PCTCN2015073045-appb-000059
将2-甲氧基-4-(4-甲基哌嗪-1-基)苯胺(107mg,0.48mmol)与三氟乙酸(166mg,1.45mmol)加入到3-丙烯酰氨基-N-(2-氯嘧啶-4-基)哌啶-1-甲酰胺(150mg,0.48mmol)的3ml正丁醇溶液中，120℃下剧烈搅拌2h。反应结束后，体系冷却至室温，减压浓缩得棕色固体。经制备液相纯化得到化合物cpd-34(48.53mg)。产率：21％，纯度：100.0％。MS m/z(ESI):495.2[M+H]+；1H NMR(400MHz,DMSO-d6):δ9.42(d,J＝15.2Hz,1H),8.63–8.31(m,1H),8.27–7.97(m,2H),7.23(s,1H),6.66(dd,J＝14.1,4.0Hz,2H),6.56–6.26(m,2H),6.07(d,J＝16.7Hz,1H),5.73–5.58(m,1H),3.78(d,J＝29.9Hz,5H),3.51(s,1H),3.24–2.81(m,6H),2.44(s,4H),2.21(d,J＝8.1Hz,3H),1.62(d,J＝32.5Hz,2H),1.30(d,J＝52.4Hz,2H)。
实施例35：3-丙烯酰氨基-N-(2-(2-甲氧基-4-(1-甲基哌啶-4-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-35)的制备
Figure PCTCN2015073045-appb-000060
将2-甲氧基-4-(1-甲基哌啶-4-基)苯胺(85mg,0.39mmol)与三氟乙酸(135mg,1.16mmol)加入到3-丙烯酰氨基-N-(2-氯嘧啶-4-基)哌啶-1-甲酰胺(120mg,0.39mmol)的 3ml正丁醇溶液中，130℃下剧烈搅拌2h。反应结束后，体系冷却至室温，减压浓缩得棕色固体。经制备液相纯化得到cpd-35(6.76mg)。产率：4％，纯度：94.53％。MS m/z(ESI):494.0[M+H]+；1H NMR(400MHz,DMSO-d6):δ9.36(d,J＝20.0Hz,0H),8.08(dd,J＝41.4,35.9Hz,1H),7.67(d,J＝37.5Hz,0H),6.93(s,0H),6.84–6.65(m,1H),6.05(d,J＝15.8Hz,0H),5.64(s,0H),3.81(s,1H),3.01(s,0H),2.86(d,J＝9.8Hz,1H),2.67(s,1H),2.30(d,J＝21.1Hz,1H),2.19(s,3H),1.95(t,J＝10.1Hz,1H),1.71(s,2H),1.64–1.54(m,1H)。
实施例36：3-丙烯酰氨基-N-(2-(4-((2-(二甲基氨基)乙基)(甲基)氨基)-2-甲氧基苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺(cpd-36)的制备
Figure PCTCN2015073045-appb-000061
步骤1：N-(2-(二甲基氨基)乙基)-3-甲氧基-4-硝基苯胺的合成
将N,N,-二甲基乙二胺(8.36g，81.9mmol)和碳酸钾(12.1g，122.8mmol)加入到4-氟-2-甲氧基-1-硝基苯(7g,40.9mmol)的200ml的DMF中，50℃下剧烈搅拌24h。TLC检测反应进度，待底物完全反应后，加入200ml水，用乙酸乙酯/水体系萃取三次，分离出有机层，减压浓缩得到粗产品，经Combi-Flash柱层析纯化得N-(2-(二甲基氨基)乙基)-3-甲氧基-4-硝基苯胺(6.5g)。产率：63％，纯度：100％。MS m/z(ESI):254.1[M+H]+。
步骤2：N1-(2-(二甲基氨基)乙基)-3-甲氧基苯-1,4-二胺的合成
将还原铁粉(256mg,4.74mmol)加入到N-(2-(二甲基氨基)乙基)-3-甲氧基-4-硝基苯胺(150mg,0.59mmol)的3ml乙醇和6滴水溶液中，滴入一滴盐酸，在100℃下剧烈搅拌4h。TLC检测反应进度，待底物完全反应后，过滤，滤液减压浓缩得到N1-(2-(二甲基氨基)乙基)-3-甲氧基苯-1,4-二胺(120mg)。产率：95％，纯度：70％。MS m/z(ESI):224.2[M+H]+。
步骤3：3-丙烯酰氨基-N-(2-(4-((2-(二甲基氨基)乙基)(甲基)氨基)-2-甲氧基苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺的合成
在0℃下，将3-丙烯酰氨基-N-(2-氯嘧啶-4-基)哌啶-1-甲酰胺(100mg,0.32mmol)与TEA(66mg，0.65mmol)加入到N1-(2-(二甲基氨基)乙基)-3-甲氧基苯-1,4-二胺(110mg,0.48mmol)的2ml二氯甲烷溶液中，0℃下剧烈搅拌2h。反应结束后，加水稀释，用二氯甲烷/水体系萃取三次，有机层减压浓缩得到粗产物。经制备液相分离纯化得 cpd-36(41.19mg)。产率：26％，纯度：97％。MS m/z(ESI):497.2[M+H]+；1H NMR(400MHz,DMSO)δ9.40(d,J＝15.8Hz,1H),8.54(d,J＝62.8Hz,1H),8.23(d,J＝30.0Hz,2H),8.03(d,J＝27.3Hz,2H),7.33–7.01(m,1H),6.95–6.49(m,2H),6.52–6.18(m,4H),6.06(dt,J＝17.2,8.5Hz,1H),5.65(d,J＝9.2Hz,1H),4.02–3.58(m,6H),3.47(s,4H),2.91(s,4H),2.62(d,J＝42.9Hz,2H),2.33(s,7H),1.62(d,J＝43.4Hz,2H),1.35(s,1H),1.26–0.84(m,1H)。
实施例37：N-(3-(2-(2-(2-甲氧基-4-(4-甲基哌嗪-1-基)苯基氨基)嘧啶-4-基)-2-甲基丙酰氨基)苯基)丙烯酰胺(cpd-37)的三氟乙酸盐的制备
Figure PCTCN2015073045-appb-000062
步骤1：2-(2-氯嘧啶-4-基)丙二酸二乙酯的合成
将丙二酸二乙酯(6.5.0g,40.0mmol)置于500mL的单口反应瓶中，加入无水甲苯(100mL)混合溶液使其溶解。在室温下，向搅拌的反应瓶中加入氢化钠(4.4g,80.0mmol)，随后将反应体系在室温下持续搅拌0.5h。之后将2,4-二氯嘧啶(5.0g,34.0mmol)加入反应体系中，反应体系用氮气置换空气三次后，加入Pd2(dba)3(600mg,0.68mmol)和P(t-Bu)3(280mg,1.36mmol)，之后再用氮气置换空气三次。将反应体系加热至80℃，并且持续搅拌16h。TLC检测反应进度，待底物完全反应后，过滤除去沉淀，滤饼用乙酸乙酯淋洗三次。滤液减压浓缩得到粗产品，经Combi-Flash[EA:PE＝5:95-50:50]柱层析纯化得到目标化合物黄色油状(2.9g,10.0mmol)。产率：26.6％；纯度：90％。
步骤2：2-(2-氯嘧啶-4-基)乙酸乙酯的合成
将2-(2-氯嘧啶-4-基)丙二酸二乙酯(2.9g,10.0mmol)置于250mL的单口反应瓶中，加入DMSO/H2O(100mL/2mL)混合溶液使底物溶解。在室温下，向搅拌的反应瓶中依次加入氯化钠(600mg,10.0mmol)，随后将反应体系加热至140℃并保持搅拌5h。TLC检测反应进度，待底物完全反应后，乙酸乙酯/水体系萃取三次，分离出有机层，经水和饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到目标化合物(1.4g,7.0mmol)，直接用于下一步反应。
步骤3：2-(2-氯嘧啶-4-基)-2-甲基丙酸乙酯的合成
将2-(2-氯嘧啶-4-基)乙酸乙酯(1.4g,7.0mmol)置于100mL的单口反应瓶中，加入丙酮(50mL)使底物溶解。在室温下，向搅拌的反应瓶中依次加入碳酸钾(2.9g,21.0mmol)和碘甲烷(1.5g,10.5mmol)，随后将反应体系加热至60℃并保持搅拌5h。TLC检测反应进度，待底物完全反应后，过滤除去沉淀，滤饼用乙酸乙酯淋洗三次， 滤液减压浓缩得到粗产品，经Combi-Flash[EA:PE＝5:95-50:50]柱层析纯化得到目标化合物无色油状(900mg,4.0mmol)。产率：39％(两步)；纯度：90％；MS m/z(ESI):229.1[M+H]+。
步骤4：2-(2-氯嘧啶-4-基)-2-甲基丙酸的合成
将2-(2-氯嘧啶-4-基)-2-甲基丙酸乙酯(900mg,4.0mmol)置于50mL的单口反应瓶中，加入甲醇(5mL)使底物部分溶解。之后加入1N氢氧化钠水溶液(5mL)，并保持反应体系在室温下持续搅拌6h。TLC检测反应进度，待底物完全反应后，反应液用乙酸乙酯/水体系萃取一次，分离出水层，2N盐酸水溶液中和pH值至5，用乙酸乙酯/水体系萃取三次，分离出有机层，饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物2-(2-氯嘧啶-4-基)-2-甲基丙酸(450mg,2.25mmol)，直接投入下步反应。产率：56％；纯度：95％。MS m/z(ESI):201.7[M+H]+。
步骤5：N-(3-氨基苯基)-2-(2-氯嘧啶-4-基)-2-甲基丙酰胺的合成
将2-(2-氯嘧啶-4-基)-2-甲基丙酸(450mg，2.25mmol)置于50mL的单口反应瓶中，加入氯化亚砜(5mL)使底物部分溶解。之后将反应体系加热至80℃并持续搅拌1h。待反应结束后，减压蒸馏去除多余氯化亚砜得到酰氯粗产物。在0℃下将酰氯粗产物缓慢地加入苯-1,3-二胺(365mg，3.38mmol)和三乙胺(0.63mL，4.5mmol)的二氯甲烷(10mL)溶液中，保持反应体系在0℃下持续搅拌1h。TLC检测反应进度，待底物完全反应后，反应液用二氯甲烷/水体系萃取三次，分离出有机层，饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物(700mg)，直接投入下步反应。
步骤6：N-(3-(2-(2-氯嘧啶-4-基)-2-甲基丙酰氨基)苯基)丙烯酰胺的合成
将N-(3-氨基苯基)-2-(2-氯嘧啶-4-基)-2-甲基丙酰胺(700mg)置于50mL的单口反应瓶中，加入二氯甲烷(10mL)使底物溶解。之后在0℃下依次加入三乙胺(1mL)和丙烯酰氯(0.5mL)，并保持反应体系在0℃下持续搅拌1h。TLC检测反应进度，反应液用二氯甲烷/水体系萃取三次，分离出有机层，饱和食盐水洗涤，无水硫酸钠干燥后，减压浓缩得到粗产物(710mg，0.91mmol)，直接投入下步反应。产率：40％(两步)；纯度：44％；MS m/z(ESI):345.7[M+H]+。
步骤7：N-(3-(2-(2-(2-甲氧基-4-(4-甲基哌嗪-1-基)苯基氨基)嘧啶-4-基)-2-甲基丙酰氨基)苯基)丙烯酰胺的三氟乙酸盐合成
将N-(3-(2-(2-氯嘧啶-4-基)-2-甲基丙酰氨基)苯基)丙烯酰胺(710mg，0.91mmol)和2-甲氧基-4-(4-甲基哌嗪-1-基)苯胺(402mg，1.82mmol)置于50mL的单口反应瓶中，加入正丁醇(10mL)使底物部分溶解。之后加入三氟乙酸(620mg，5.46mmol)，并保持反应体系在130℃下加热16h。待反应结束后，冷却至室温，减压浓缩得到粗产物。经Prep-HPLC柱层析[H2O(0.05％TFA):CH3CN＝65:35～5:95]纯化得到目标产品cpd-37的三氟乙酸盐(10.38mg，0.016mmol)。产率：1.8％；纯度：95％；MS m/z(ESI):530.7[M+H]+；1H NMR(400MHz，CDCl3):δ9.18(s,1H),8.47–8.34(m,2H),8.03(d,J＝8.7Hz,1H),7.50(s,1H),7.32(d,J＝9.9Hz,2H),7.25(s,1H),7.17(t,J＝8.1Hz,1H),6.80(d,J＝5.2Hz,1H),6.52–6.37(m,3H),6.16(d,J＝8.7Hz,1H),5.73(t,J＝5.8Hz,1H),3.87(s,3H),3.23(s,4H),3.12(s,4H),2.74(s,3H),1.68(s,6H)。
实施例38：1-(3-丙烯酰基酰胺基)-3-(2-(2-甲氧基-4-(4-丙基哌嗪-1-基)苯基氨基) 嘧啶-4-基)脲(cpd-38)的制备
Figure PCTCN2015073045-appb-000063
制备方法同实施例23，不同的是将步骤中的4-(4-乙基哌嗪-1-基)-2-甲氧基苯胺换成4-(4-丙基哌嗪-1-基)-2-甲氧基苯胺(制备方法类似中间体11)。MS m/z(ESI):531[M+H]+；1H NMR(400MHz,DMSO)δ10.47(s,1H),10.12(s,1H),9.57(s,1H),8.26(s,1H),8.10(d,J＝5.5Hz,1H),7.93(s,1H),7.47(d,J＝7.7Hz,1H),7.35(d,J＝8.4Hz,1H),7.13(t,J＝8.0Hz,1H),6.59(d,J＝6.8Hz,2H),6.48–6.41(m,2H),6.25(d,J＝18.8Hz,1H),5.74(d,J＝11.6Hz,1H),3.75(s,3H),3.29(s,4H),3.11(s,4H),2.32–2.25(m,2H),1.48(dd,J＝14.7,7.4Hz,2H),0.89(t,J＝7.4Hz,3H).
实施例39：3-(3-丙烯酰基酰胺基)-1-(2-(4-((2-(二甲基氨基)乙基)(甲基)氨基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基脲(cpd-39)的制备
Figure PCTCN2015073045-appb-000064
制备方法同实施例32，不同的是将步骤2中的1-(4-氨基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺换成N1-(2-(二甲基氨基)乙基)-3-甲氧基-N1甲基苯-1,4-二胺(制备方法类似中间体4)。MS m/z(ESI):519[M+H]+；1H NMR(400MHz,DMSO)δ12.02(s,1H),10.11(s,1H),8.52(s,1H),8.23(d,J＝5.7Hz,1H),7.84(s,1H),7.48(d,J＝7.8Hz,1H),7.13(d,J＝8.6Hz,1H),7.01(s,1H),6.45(dd,J＝16.4,10.6Hz,2H),6.40–6.17(m,3H),6.13(d,J＝9.0Hz,1H),5.80–5.68(m,1H),3.70(s,3H),3.33(d,J＝6.9Hz,5H),2.86(s,3H),2.27(t,J＝6.8Hz,2H),2.14(s,6H).
实施例40：1-(3-丙烯酰基酰胺基)-3-(5-氯-2-(2-甲氧基-4-(4-吗啉代哌啶-1-基)苯基氨基)嘧啶-4-基)脲(cpd-40)的制备
Figure PCTCN2015073045-appb-000065
制备方法同实施例9，不同的是将起始原料1-(2-氯嘧啶-4-基)-3-(3- 硝基苯基)脲换成1-(2,5-二氯吡啶-4-基)-3-(3-硝基苯基)脲。MS m/z(ESI):607[M+H]+；1H NMR(400MHz,CDCl3)δ10.82(s,1H),10.13(s,1H),8.76(s,1H),8.64(s,1H),8.21(s,1H),7.85(s,1H),7.53(d,J＝8.0Hz,1H),7.24(d,J＝8.6Hz,1H),7.12(s,1H),6.65–6.31(m,4H),6.24(dd,J＝17.0,1.9Hz,1H),5.76–5.71(m,1H),3.71(s,3H),3.62(s,1H),3.60–3.56(m,5H),2.59(t,J＝11.6Hz,2H),2.47(s,4H),2.22(d,J＝10.9Hz,1H),1.83(d,J＝11.7Hz,2H),1.50–1.41(m,2H).
实施例41：酶平台(z-lyte)药物筛选
以下z-lyte测试方法中所用试剂均购自Invitrogen。
利用z-lyte方法测定待测物对双突变型EGFR激酶T790M/L858R激酶(Invitrogen，PV4879)、野生型EGFR激酶(EGFR WT)(Invitrogen，PV3872)激酶活性的抑制作用。
10μL T790M/L858R激酶反应中各组分的工作浓度为：25μM ATP,0.08ng/μL T790M/L858R激酶,2μM Tyr04底物(Invitrogen，PV3193)。加入本发明上述实施例制备的化合物(即待测物)后DMSO的浓度为2％。
10μL EGFR WT激酶反应中各组分的工作浓度为：10μM ATP,0.8ng/μL T790M WT激酶,2μM Tyr04底物(Invitrogen，PV3193)。加入待测物后DMSO的浓度为2％。
室温溶解10mM的药物储存液经4％DMSO的水梯度稀释至终浓度10-0.005μM。每孔中加入2.5μL的待测物溶液以及5μL经反应缓冲液稀释的T790M/L858R激酶(或EGFR WT激酶)与Tyr04底物的混合物，再加入2.5μL的ATP启动反应。C1孔用反应缓冲液代替ATP，C2孔不加入任何药物，C3孔按说明书描述加入磷酸化的底物。在室温摇床反应60min后。加入5μL Development Reagent B(Invitrogen，用TR-FRET稀释缓冲液进行稀释)，于室温摇床反应60min。在VictorX5荧光酶标仪(PerkinElmer)上读板，测定激发波长为405nm，发射波长为450nm和520nm的光吸收。(例如，C3520nm表示C3孔在520nm的读值)。抑制率计算方法如下：
1、ER＝Coumarin Emission(450nm)/Fluorescein Emission(520nm)
2、磷酸化率＝(1-((ER×C3520nm-C3450nm)/((C1450nm-C3450nm)+ER×(C3520nm-C1520nm))))×100％
3、抑制率(IR)＝(1-(测试化合物的磷酸化率)/(C2的磷酸化率))×100％
用XLFIT 5.0软件(英国IDBS公司)拟合计算半数抑制浓度IC50。示例化合物对酶的抑制活性或选择抑制活性见表1-表2。
表1-酶抑制活性
化合物编号 T790M/L858R(IC50/μM) EGFR WT(IC50/μM)
cpd-1 0.017 0.017
cpd-2 0.048 0.143
cpd-5 0.005 0.080
cpd-7 0.058 0.348
cpd-9 0.004 0.086
cpd-10 0.002 0.045
cpd-14 0.015 0.225
cpd-19 0.028 0.273
cpd-20 0.022 0.158
cpd-21 0.012 0.183
cpd-22 0.002 0.036
cpd-23 0.013 0.039
cpd-31 0.087 0.644
cpd-32 0.028 0.054
cpd-38 0.010 0.042
cpd-39 0.019 0.049
cpd-40 0.037 0.641
BIBW2992 0.005 0.001
CO1686 0.008 0.074
表2-酶的选择抑制活性
化合物编号 对酶的选择抑制活性[IC50(EGFR WT)/IC50(T790M/L858R)]
cpd-5 16
cpd-9 21.5
cpd-10 22.5
cpd-14 15
cpd-15 >7.8
cpd-17 >10
cpd-19 9.8
cpd-21 15.3
cpd-22 18
cpd-40 17.3
BIBW2992 0.2
CO1686 9.25
从表1、表2可以看出，本发明的示例化合物对EGFR突变型酶(T790M/L858R和L858R)表现出较强的抑制活性，而对EGFR野生型酶(EGFR WT)抑制活性较弱，与阳性对照物BIBW2992(Afatinib)相比，本发明的化合物对EGFR突变型酶具有明显的选择抑制活性。本发明的部分示例化合物的选择抑制活性甚至超过了阳性对照物CO1686(具体参见文献Cancer Discovery,doi:10.1158/2159-8290.CD-13-0314)。
实施例42：MTT(3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐)方法检测细胞活性
MTT测试方法步骤采用本领域技术人员熟知的方法进行，方法中所用试剂均可市购得到。
首先，移除培养基并加入0.25％的胰酶/EDTA(Gibco,25200-056)。
洗一次后，再加入1.5mL胰酶/EDTA消化贴壁细胞，至细胞分离，然后加入3.5mL培养基终止消化。将消化完的细胞悬浮液移至15mL离心管，1300rpm离心3min后弃上清，并用新鲜的培养基悬浮细胞。
然后细胞计数，并稀释细胞至以下浓度：A431和H1975细胞每ml2.78万，NIH3T3每ml3.33万。将细胞种入96孔板(BD 3072)，每孔90μL，培养过夜。
A431细胞培养基为：10％FBS(Gibco，10099-141)DMEM(Hyclone SH30243.01B)；
NIH3T3细胞培养基为：10％FBS(Gibco，10099-141)DMEM(Hyclone SH30243.01B)；
H1975细胞培养基为：10％FBS(Gibco，10099-141)RPMI-1640(Hyclone  SH30809.01B)；
取20μL10mM待测化合物，按照如下浓度梯度(2000，666.67，222.22，74.07，24.69，8.23，2.74，0.91μM)稀释10X药品，并加入每孔10μl药品到细胞培养板内，再加入无血清培养基(终浓度为：10，3.333，1.111，0.370，0.123，0.041，0.014，0.005μM)，其中DMSO终浓度为0.5％。
加药后将细胞放入培养箱，培养72h后，每孔加入10μL的5mg/ml的MTT(Sigma,M5655)溶液，然后将96孔板放入37℃5％CO2培养箱孵育4h。
再在2000rpm，5min的条件下离心平板，移除上清后，每孔加入150μL DMSO，并在摇床中震荡平板至所有结晶紫溶解(约10-20min)。最后使用荧光酶标仪测定492nm光吸收，使用XLFIT 5.0软件(英国IDBS公司)计算IC50。示例化合物对细胞的抑制活性或选择抑制活性见表3至表6。
表3-化合物对细胞活性的抑制结果(H1975细胞)
化合物编号 H1975细胞(IC50/μM)
cpd-9 0.037
cpd-19 0.075
cpd-21 0.086
cpd-22 0.031
cpd-23 0.070
BIBW2992 0.088
CO1686 0.029
表4-化合物对细胞活性的抑制结果(A431细胞)
化合物编号 A431细胞(IC50/μM)
cpd-9 >10
cpd-10 3.379
cpd-19 >10
cpd-21 >10
cpd-23 >10
cpd-31 >10
cpd-32 3.577
BIBW2992 0.029
CO1686 2.892
表5-化合物对细胞的选择性
化合物编号 对细胞的选择抑制活性[IC50(A431细胞)/IC50(H1975细胞)]
cpd-9 >270
cpd-19 >133
cpd-21 >116
cpd-23 >143
BIBW2992 0.33
CO1686 99.7
表6-化合物对NIH3T3细胞的毒性测试结果
化合物编号 NIH3T3细胞MTT测试(IC50/μM)
cpd-1 4.199
cpd-2 7.261
cpd-5 4.246
cpd-7 >10
cpd-9 >10
cpd-10 >10
cpd-14 >10
cpd-15 6.291
cpd-19 >10
cpd-21 >10
cpd-22 4.263
cpd-25 >10
cpd-31 9.689
cpd-32 3.103
BIBW2992 2.750
CO1686 2.295
从表3-5可以看出，本发明的化合物对EGFR突变型细胞(H1975细胞)表现出较强的抑制活性，而对EGFR野生型细胞(A431细胞)表现出较弱抑制活性，与阳性对照物BIBW2992相比，本发明的化合物对EGFR突变型细胞具有明显的选择抑制活性。并且本发明的部分示例化合物对EGFR突变型细胞的选择抑制活性甚至超过了阳性对照物CO1686。
从表6可以看出，本发明的化合物对NIH3T3细胞具有较高的IC50值，因此显示出较小的毒性。
实施例43：EGFR T790M抑制剂细胞活性ELISA法测定
以下方法中的试剂、溶液的配置方法以及细胞处理和裂解液制备步骤、ELISA检测步骤均按照R&D DYC3570，R&D DYC1095E以及R&D DYC1095BE的说明书进行操作。
一、试剂和溶液
细胞裂解缓冲液：1％NP-40，20mM Tris(pH 8.0)，137mM NaCl，10％glycerol，1mM NaVO3，2mM EDTA。
细胞裂解液：细胞裂解缓冲液+10μg/mL抑肽酶(Aprotinin)(Sigma)，10μg/mL亮抑蛋白肽酶(Leupeptin)(Sigma)，现配现用。
1x PBS缓冲液：NaCl:0.137M，KCl:0.0027M，Na2PO4-12H2O:0.01M，KH2PO4:0.0015M，pH7.4。
洗涤缓冲液：含有0.05％Tween-20的PBS缓冲液。
检测抗体稀释液：20mM Tris，137mM NaCl，0.05％Tween-20，0.1％BSA，pH7.2-7.4。
封闭液：含有1％BSA的PBS缓冲液。
ELISA试剂盒：R&D DYC3570，R&D DYC1095E和R&D DYC1095BE。
二、H1975细胞
2.1H1975细胞处理和裂解液制备
(1)将H1975细胞以1×104/孔的密度种到96孔板中，每孔90微升10％FBS,1640培养基，37℃、5％CO2培养过夜。
(2)将待测化合物按照MTT实验中药物稀释方法稀释，将10μL稀释后的化合物或稀释后的DMSO加入到细胞培板的对应孔中，DMSO终浓度为0.5％，37℃、5％CO2 培养1小时。以纯DMSO处理的细胞培养体系作为细胞对照。
(3)吸掉培养基后加入100μL细胞裂解液，封板模封置于-80℃冰箱中过夜。以细胞裂解缓冲液作为空白对照。
2.2ELISA检测步骤
按照R&D DYC1095E或R&D DYC1095BE给定说明书进行操作。
(1)R&D捕获抗体((DYC1095BE或DYC1095E))用PBS 1:180稀释，稀释好的抗体100μL/孔加入ELISA反应板(Corning costar 42592)，25℃摇床包被过夜；
(2)360μL洗涤缓冲液洗3次；
(3)加入300μL封闭液，25℃摇床孵育2小时；
(4)360μL洗涤缓冲液洗3次；
(5)加入40μL细胞裂解缓冲液和60μL细胞裂解液，25℃摇床孵育2小时；
(6)360μL洗涤缓冲液洗3次；
(7)检测抗体用检测抗体稀释液以试剂盒说明规定比例稀释，每孔加入100μL，25℃摇床避光孵育1小时；
(8)360μL洗涤缓冲液洗3次；
(9)将TMB底物(R&D DY999)中的A试剂和B试剂以1:1进行混合，每孔100μL，25℃摇床避光孵育20分钟；
(10)2N H2SO4每孔加入50μL；
(11)用酶标仪读板(Thermo Multiskan K3)分别测定细胞对照、空白对照以及药物处理情况下的OD 450值和OD570值，并用相同孔的OD 450值减去相应OD570值分别得到OD细胞、OD空白和OD药物处理。
2.3数据分析
抑制率(％)＝100％×(OD细胞-OD药物处理)/(OD细胞-OD空白)
2.4将计算得到的抑制率用XLFIT 5.0软件计算出IC50值，参见表7。
三、A431细胞
3.1A431细胞的处理和测试步骤
(1)将A431细胞以1×104/孔的密度种到96孔板中，每孔90微升含有10％FBS的DMEM培养基37℃、5％CO2培养过夜。
(2)将A431细胞培养基更换为90微升无血清DMEM培养基，继续培养过夜。
(3)将待测化合物按照MTT实验中药物稀释方法稀释，将10μL稀释后的化合物或稀释后的DMSO加入到细胞培板的对应孔中，DMSO终浓度为0.5％，37℃、5％CO2培养1小时。然后在除细胞对照孔外的每孔中加入10微升2μg/L的EGF，在细胞孔加入10微升无血清DMEM培养45分钟；以不加入EGF与药物处理的细胞作为细胞对照，以不加入药物的只加入EGF处理的细胞作为EGF对照。
(4)吸掉培养基后加入100μL细胞裂解液，封板模封置于-80℃冰箱中过夜。
3.2ELISA检测步骤
参照R&D DYC3570E说明书进行操作。
(1)R&D捕获抗体(DYC3570E)用PBS 1:180稀释，稀释好的抗体100μL/孔加入ELISA反应板(Corning costar 42592)，25℃摇床包被过夜；
(2)360μL洗涤缓冲液洗3次；
(3)加入200μL封闭液，25℃摇床孵育2小时；
(4)360μL洗涤缓冲液洗3次；
(5)加入40μL细胞裂解缓冲液和60μL细胞裂解液，25℃摇床孵育2小时；
(6)360μL洗涤缓冲液洗3次；
(7)检测抗体用检测抗体稀释液以试剂盒说明规定比例稀释，每孔加入100μL，25℃摇床避光孵育1小时；
(8)360μL洗涤缓冲液洗3次；
(9)将TMB底物(R&D DY999)中的A试剂和B试剂以1:1进行混合，每孔100μL，25℃摇床避光孵育20分钟；
(10)2N H2SO4每孔加入50μL；
(11)用酶标仪读板(Thermo Multiskan K3)分别测定细胞对照、空白对照以及药物处理情况下的OD 450值和OD570值，并用相同孔的OD 450值减去相应OD570值分别得到OD EGF、OD药物、OD细胞。
3.3数据分析
抑制率(％)＝100％×(OD EGF-OD药物)/(OD EGF-OD细胞)
3.4将计算得到的抑制率用XLFIT 5.0软件计算出IC50值，参见表7。
表7细胞活性ELISA法测定结果
Figure PCTCN2015073045-appb-000066
从表7可以看出，与阳性对照物BIBW2992相比，本发明的示例化合物对细胞水平靶点具有明显的选择抑制活性。并且部分示例化合物的选择抑制活性甚至超过了阳性对照物CO1686，最高提高了6倍。
从体外酶、细胞生长抑制实验显示，本发明化合物对EGFR突变型酶、细胞表现出较强的抑制活性，而对EGFR野生型酶、细胞表现出较弱抑制活性，因此此类化合物对T790M突变的EGFR有较好的选择抑制活性和较低的细胞毒性。
在本发明提及的所有文献都在本申请中引用作为参考，就如同每一篇文献被单独引用作为参考那样。此外应理解，在阅读了本发明的上述讲授内容之后，本领域技术人员可以对本发明作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。

Claims (10)

  1. 一种芳氨基嘧啶类化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药，所述化合物的结构如式Ⅰ所示：
    Figure PCTCN2015073045-appb-100001
    式中，
    A选自：取代或未取代的C6-10芳烃基、取代或未取代的C5-8杂芳基、取代或未取代的C3-8环烷基、取代或未取代的C3-8杂环烷基、取代或未取代的8-14元稠环基团；
    Z1选自NR9或CR10R11；其中,R9选自氢或C1-6烷基；R10、R11各自独立地选自氢、卤素、C1-6烷基；或R10和R11与相连的碳原子共同形成C3-8环烷基或C3-8杂环烷基；
    Z2、Z3各自独立地选自CR12或N；其中,R12选自氢、卤素、硝基、氨基、C1-6烷基取代的氨基、C1-6酰基取代的氨基、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、卤代的C1-6烷氧基；
    R1选自氢或C1-6烷基；或者，R1与A及它们相连的氮原子一起组成选自下列的基团：取代或未取代的C5-8杂芳基、取代或未取代的C3-8杂环烷基、或者取代或未取代的8-14元稠环基团；
    上述基团中，所述的“取代”是指基团中的一个或多个氢被选自下组的取代基所取代：硝基、卤素、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、卤代的C1-6烷氧基；
    R2选自氢、卤素、C1-6烷基、卤代C1-6烷基、或C1-6烷氧基；
    R3、R4、R5各自独立地选自：氢、卤素、硝基、氨基、C1-6烷基取代的氨基、C1-6酰基取代的氨基、C1-6烷基取代的二氨基、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、C1-6烷氧基取代的C1-6烷氧基、C6-10芳烃基取代的C1-6烷氧基、卤代的C1-6烷氧基、-O-C6-10芳烃基、-CO-C3-6杂环烷基、-CO-N R13R14、C3-6杂芳基、C3-6杂环烷基、C3-6杂环烷基-CO-C1-6烷基，C3-6杂环烷基-N-C1-6烷基，C3-6杂环烷基-C1-6烷基，C3-6杂环烷基-C3-6杂环烷基；任选地，所述C6-10芳烃基、C3-6杂芳基、或者C3-6杂环烷基可被一个或多个选自下组的取代基所取代：硝基、卤素、氰基、C1-6烷基、卤代的C1-6烷基、C1-6烷氧基、卤代的C1-6烷氧基、C1-6烷基取代的氨基；其中，R13、R14各自独立地选自氢、或C1-6烷基；
    R7、R8各自独立地选自：氢、卤素、C1-6烷基、C3-8环烷基、C3-8杂环烷基、C6-10芳烃基或C5-8杂芳基。
  2. 如权利要求1所述的化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药，其中，所述化合物的结构如式(II)所示：
    Figure PCTCN2015073045-appb-100002
    式中，A、Z2、Z3、R1、R2、R3、R4、R5、R7、R8和R9如权利要求1中所定义。
  3. 如权利要求1所述的化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药，其中，所述化合物的结构如式(III)所示：
    Figure PCTCN2015073045-appb-100003
    式中，A、Z2、Z3、R1、R2、R3、R4、R5、R7、R8、R10和R11如权利要求1中所定义。
  4. 如权利要求1-3任一项所述的化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药，其中，(ⅰ)A为苯环，或(ⅱ)A与R1及它们相连的氮原子一起组成哌啶。
  5. 如权利要求1所述的化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药，其中，所述化合物选自下组：
    N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(4-甲基哌嗪-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((5-甲氧基吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((3-氯-4-((3-氟苄基)氧)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2-甲氧基-4-(4-(4-甲基哌嗪-1-基)哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((6-吗啉代吡啶-3-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2,4-二甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((3-氯-4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2-甲氧基-4-(4-吗啉哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(二甲氨基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((5-吗啉代吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2-甲氧基-4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2-甲氧基-4-(吗啉-4-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2-甲氧基-4-(4-甲基哌嗪-1-羰基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    4-((4-(3-(3-丙烯酰基酰胺苯基)脲基)嘧啶-2-基)氨基)-N,N-二乙基-3-甲氧基苯甲酰胺；
    N-(3-(3-(2-((4-(2-甲氧基乙氧基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2-甲氧基-4-(哌啶-1-基)苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(4-(2-氟乙基)哌嗪-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(二乙基氨基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(4-(二乙基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(4-乙基哌嗪-1-基)-2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-(4-甲苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((5-(4-甲基哌嗪-1-基)吡啶-2-基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-氟苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-乙氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((2-甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-氯苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((3,4,5-三甲氧基苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-吗啉代苯基)氨基)嘧啶-4-基)脲基)苯基)丙烯酰胺；
    N-(3-(3-(2-((4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧苯基)氨基)嘧啶-4-基)-3-甲基脲基)苯基)丙烯酰胺；
    3-丙烯酰氨基-N-(2-(2-甲氧基-4-(4-吗啉代-1-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺；
    3-丙烯酰氨基-N-(2-(2-甲氧基-4-(4-甲基哌嗪-1-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺；
    3-丙烯酰氨基-N-(2-(2-甲氧基-4-(1-甲基哌啶-4-基)苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺；
    3-丙烯酰氨基-N-(2-(4-((2-(二甲基氨基)乙基)(甲基)氨基)-2-甲氧基苯基氨基)嘧啶-4-基)哌啶-1-甲酰胺；
    N-(3-(2-(2-(2-甲氧基-4-(4-甲基哌嗪-1-基)苯基氨基)嘧啶-4-基)-2-甲基丙酰氨基)苯基)丙烯酰胺；
    1-(3-丙烯酰基酰胺基)-3-(2-(2-甲氧基-4-(4-丙基哌嗪-1-基)苯基氨基)嘧啶-4-基) 脲；
    3-(3-丙烯酰基酰胺基)-1-(2-(4-((2-(二甲基氨基)乙基)(甲基)氨基)-2-甲氧基苯基氨基)嘧啶-4-基)-1-甲基脲；或
    1-(3-丙烯酰基酰胺基)-3-(5-氯-2-(2-甲氧基-4-(4-吗啉代哌啶-1-基)苯基氨基)嘧啶-4-基)脲。
  6. 一种药物组合物，所述组合物包括：权利要求1至5任一项所述的化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药；以及药学可接受的载体。
  7. 权利要求1至5任一项所述的化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药在制备调控EGFR酪氨酸激酶活性或者治疗EGFR相关疾病的药物中的应用。
  8. 如权利要求7所述的应用，其中，所述的EGFR相关疾病选自：癌症、糖尿病、免疫系统疾病、神经退行性疾病、心血管疾病、使用EGFR调节剂治疗期间具有获得性耐药性的疾病。
  9. 如权利要求8所述的应用，其中，所述获得性耐药性的疾病是由EGFR外显子20编码的T790突变所引起的，或者是包含EGFR外显子20编码的T790突变所引起的。
  10. 一种药用组合物，所述药用组合物包括：权利要求1-5任一项所述的化合物、或其药学上可接受的盐、立体异构体、溶剂化合物或其前药；以及选自下组药物中的一种或多种：吉非替尼、厄洛替尼、埃克替尼、拉帕替尼、XL647、NVP-AEE-788、ARRY-334543、EKB-569、BIBW2992、HKI272、BMS-690514、CI-1033、凡德他尼、PF00299804、WZ4002、西妥昔单抗、曲妥珠单抗、帕尼突单抗、马妥珠单抗、尼妥珠单抗、扎鲁木单抗、帕妥珠单抗、MDX-214、CDX-110、IMC-11F8、Zemab、Her2疫苗PX 1041、HSP90抑制剂、CNF2024、坦螺旋霉素、阿螺旋霉素、IPI-504、SNX-5422、NVP-AUY922、或其组合。
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US10377747B2 (en) 2015-08-31 2019-08-13 Wuxi Shuangliang Biotechnology Co., Ltd. 2-arylamino pyridine, pyrimidine or triazine derivatives, preparation method and use thereof
JP2019519512A (ja) * 2016-05-20 2019-07-11 浙江海正薬業股▲ふん▼有限公司Ｚｈｅｊｉａｎｇ Ｈｉｓｕｎ Ｐｈａｒｍａｃｅｕｔｉｃａｌ ＣＯ．，ＬＴＤ． ピリミジン誘導体、その調製方法および医療での使用
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CN113402469A (zh) * 2021-07-07 2021-09-17 沈阳信康药物研究有限公司 一种6-（3-氯丙基）氨基-1,3-二甲基脲嘧啶的制备方法

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