WO2017076355A1 - 嘧啶类衍生物及其用途 - Google Patents

嘧啶类衍生物及其用途 Download PDF

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WO2017076355A1
WO2017076355A1 PCT/CN2016/104723 CN2016104723W WO2017076355A1 WO 2017076355 A1 WO2017076355 A1 WO 2017076355A1 CN 2016104723 W CN2016104723 W CN 2016104723W WO 2017076355 A1 WO2017076355 A1 WO 2017076355A1
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compound
group
mmol
formula
optionally substituted
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PCT/CN2016/104723
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English (en)
French (fr)
Inventor
王学海
许勇
盛锡军
张晓林
夏汉贵
杨仲文
乐洋
黄璐
肖强
Original Assignee
湖北生物医药产业技术研究院有限公司
人福医药集团股份公司
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Application filed by 湖北生物医药产业技术研究院有限公司, 人福医药集团股份公司 filed Critical 湖北生物医药产业技术研究院有限公司
Priority to EP16861623.3A priority Critical patent/EP3372594B1/en
Priority to US15/773,826 priority patent/US10695347B2/en
Priority to KR1020187015578A priority patent/KR20180073682A/ko
Priority to CA3004372A priority patent/CA3004372C/en
Priority to JP2018522969A priority patent/JP6732906B2/ja
Priority to AU2016349089A priority patent/AU2016349089B2/en
Publication of WO2017076355A1 publication Critical patent/WO2017076355A1/zh
Priority to IL259150A priority patent/IL259150B/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D413/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D413/14—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing three or more hetero 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
    • A61K31/495—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
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • 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
    • 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D409/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms
    • C07D409/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings
    • C07D409/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D409/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms
    • C07D409/14—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing three or more hetero rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2300/00—Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/08—Solutions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/12—Aerosols; Foams
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/20—Pills, tablets, discs, rods
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate

Definitions

  • the present invention belongs to the field of biomedicine, and in particular, the present invention relates to pyrimidine derivatives and uses thereof. More specifically, the present invention relates to a pyrimidine derivative and a process for the preparation thereof, and to the use thereof in the preparation of a medicament.
  • Non-small-cell carcinoma is synonymous with "non-small cell carcinoma.”
  • Non-small cell lung cancer including squamous cell carcinoma, adenocarcinoma, and large cell carcinoma, has slower growth and cell division and longer metastatic spread than small cell carcinoma.
  • Non-small cell lung cancer accounts for about 80-85% of the total number of lung cancers. The data show that the incidence of lung cancer in China is increasing by 26.9% per year. From 2000 to 2005, the number of lung cancer cases in China is estimated to increase by 120,000. Among them, the number of male lung cancer patients increased from 260,000 in 2000 to 330,000 in 2005, and the number of female lung cancer patients increased from 120,000 to 170,000. In addition, lung cancer has become the "first cancer" in many parts of the country.
  • the treatment of advanced NSCLC has entered the era of individualized treatment.
  • the individualized targeted therapy for clinical application is mainly aimed at EGFR mutant and ALK (Anaplastic lymphoma kinase) fusion genotype lung cancer. Both genetic variant lung cancers have clear molecular targets and target detection.
  • the clinical efficacy of the technology and marketed targeted drugs has been significantly improved.
  • Anaplastic lymphoma kinase is a receptor tyrosine protein kinase that was first discovered in a subtype of anaplastic large cell lymphoma (ALCL), hence the name anaplastic lymphoma kinase (Anaplastic). Lymphoma Kinase, ALK) (Science, 1994, 263, 1281-1284; Oncogene, 1994, 9, 1567-1574).
  • the ALK fusion gene is a potent oncogene and is a newly discovered NSCLC-driven gene in which the echinoderms microtubule-associated protein 4 (EML4-ALK) is the most common type.
  • ALK abnormal kinases fused to other genes and is involved in canceration.
  • the ALK fusion gene is mainly found in patients with lung adenocarcinoma who do not smoke or smoke less.
  • ALK-positive non-small cell lung cancer which accounts for 3% to 5% of total NSCLC, is the most recently discovered molecular subtype in 2007.
  • LDK378, a second-generation ALK inhibitor, was developed by Novartis. In early clinical trials, 80% of patients with ALK non-small cell lung cancer treated with Crizotinib had an 80% response rate. And in March 2013, he was awarded the FDA “Breakthrough Treatment” title. The study found that LDK378 inhibits Crizotinib resistance mutations and was approved by the FDA in April 2014 for the treatment of resistance to Crizitinib. A patient with ALK-positive non-small cell lung cancer, named Ceritinib under the trade name Zykadia.
  • LDK378 inhibits Crizotinib resistance mutations, including L1196M, G1269A, I1171T and S1206Y, but the drug does not inhibit G1202R and F1174C, and has certain limitations.
  • Certinib is used in large doses (750mg, P.O qday).
  • ALK inhibitor drugs have great social and economic benefits, and is also a research hotspot of major pharmaceutical companies. Designing a new compound structure by modifying the structure of the compound, improving its drug resistance and drug-forming properties, thereby improving biological activity and bioavailability, and searching for a new class of ALK inhibitors for clinical treatment of diseases caused by ALK mutations. It is of great significance. Therefore, current ALK inhibitors still need to be improved.
  • the present invention is directed to solving at least some of the above technical problems or at least providing a useful commercial choice. To this end, it is an object of the present invention to provide a compound which can be used for the preparation of a medicament for the treatment of cancer.
  • the invention proposes a compound.
  • the compound is a pharmaceutically acceptable salt, hydrate, solvate, metabolite, or prodrug of a compound of Formula I or a compound of Formula I,
  • R 1 is a 5-6 membered cycloalkyl group, a 5-6 membered heterocyclic group, a 5-6 membered aryl group, or a 5-6 membered heteroaryl group
  • the 5-6 membered cycloalkyl group 5-6 membered heterocyclic group, 5-6 membered aryl group and 5-6 membered heteroaryl group are each independently selected from one or more selected from the group consisting of halogen, hydroxy, cyano, nitro, C 1-8 alkyl, C 2-8 alkenyl, C 2-8 alkynyl, C 3-8 cycloalkyl, 3-8 membered heterocyclic, C 5-10 aryl, 5-10 membered heteroaryl, C 1- 6 alkoxy, C 3-8 cycloalkoxy, -S(O)pR 6 , -C(O)R 6 , -C(O)OR 6 , -NR 7 R 8 or -C(O)NR Substit
  • R 2 is hydrogen, halogen, C 1-6 alkyl, C 1-6 alkoxy, halogen substituted C 1-6 alkyl or halogen substituted C 1-6 alkoxy;
  • R 3 is an optionally substituted piperazinyl, an optionally substituted morpholinyl group, an optionally substituted piperidinyl group, an optionally substituted cyclohexylamino group, an optionally substituted C 1-2 alkane Or a substituted 1,2,3,6-tetrahydropyridinyl group, the substituent on the substituted C 1-2 alkyl group being an optionally substituted piperidinyl group;
  • R 4 is hydrogen or C 1-6 alkyl
  • R 5 is hydrogen, chlorine, hydroxy, cyano, C 1-6 alkyl, C 1-6 alkoxy or halogen substituted C 1-6 alkoxy.
  • R 1 is a 5-6 membered heterocyclic group, a 5-6 membered aryl group, or a 5-6 membered heteroaryl group substituted with one or more -S(O)pR 6 .
  • R 2 is hydrogen, halogen, C 1-6 alkyl, C 1-6 alkoxy or halogen substituted C 1-6 alkoxy.
  • R 3 is an optionally substituted piperazinyl, an optionally substituted morpholinyl, an optionally substituted piperidinyl, an optionally substituted methyl or optionally substituted a 1,2,3,6-tetrahydropyridinyl group, the substituent on the optionally substituted methyl group being an optionally substituted piperidinyl group;
  • the optionally substituted piperazinyl group, optionally Substituted morpholinyl, optionally substituted piperidinyl or optionally substituted 1,2,3,6-tetrahydropyridinyl is optionally selected from one or more selected from C1-6 alkyl, hydroxy , C 1-6 alkylamino, C 1-6 alkoxy, oxo, C 1-6 acyl, morpholinyl, C 1-6 alkylmorpholinyl, piperazinyl, C 1-6 alkyl Substituted by a substituent of piperazinyl, C 1-6 acyl pipe
  • R 4 is hydrogen or C 1-6 alkyl.
  • R 5 is chloro or halogen substituted C 1-6 alkoxy.
  • R 6 is C l-4 alkyl.
  • p is 2.
  • R 1 is Any of them.
  • R 2 is hydrogen, chlorine, methyl, methoxy, ethoxy, isopropoxy or difluoromethoxy.
  • R 3 is one of the following:
  • R 3 when R 1 is When R 3 can be one of the following:
  • R 4 is hydrogen or methyl.
  • R 5 is chloro or difluoromethoxy.
  • the compound of the formula I of the present invention may be one of the following compounds, or a pharmaceutically acceptable salt, hydrate, solvate, metabolite, or prodrug thereof:
  • C 1-6 is selected from C 1 , C 2 , C 3 , C 4 , C 5 and C 6 ;
  • C 1-8 is selected from C 1 , C 2 , C 3 , C 4 , C 5 , C 6 , C 7 and C 8 ;
  • C 2-8 is selected from C 2 , C 3 , C 4 , C 5 , C 6 , C 7 and C 8 ;
  • C 3-8 is selected from C 3 , C 4 , C 5 , C 6 , C 7 and C 8 ;
  • C 5-10 are selected from C 5 , C 6 , C 7 , C 8 , C 9 and C 10 .
  • pharmaceutically acceptable as used herein is intended to mean that those compounds, materials, compositions and/or dosage forms are within the scope of sound medical judgment and are suitable for use in contact with human and animal tissues. Without excessive toxicity, irritation, allergic reactions or other problems or complications, commensurate with a reasonable benefit/risk ratio.
  • pharmaceutically acceptable salt refers to a salt of a compound of the invention prepared from a compound having a particular substituent found in the present invention and a relatively non-toxic acid or base.
  • a base addition salt can be obtained by contacting a neutral amount of such a compound with a sufficient amount of a base in a neat solution or a suitable inert solvent.
  • Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic ammonia or magnesium salts or similar salts.
  • an acid addition salt can be obtained by contacting a neutral form of such a compound with a sufficient amount of an acid in a neat solution or a suitable inert solvent.
  • pharmaceutically acceptable acid addition salts include inorganic acid salts including, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, hydrogencarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, Hydrogen sulfate, hydroiodic acid, phosphorous acid, etc.; and an organic acid salt, such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, Similar acids such as fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, and me
  • the salt is contacted with a base or acid in a conventional manner, and the parent compound is separated, thereby regenerating the neutral form of the compound.
  • the parent form of the compound differs from the form of its various salts by certain physical properties, such as differences in solubility in polar solvents.
  • a "pharmaceutically acceptable salt” is a derivative of a compound of the invention wherein the parent compound is modified by salt formation with an acid or with a base.
  • pharmaceutically acceptable salts include, but are not limited to, inorganic or organic acid salts of bases such as amines, alkali metal or organic salts of acid groups such as carboxylic acids, and the like.
  • Pharmaceutically acceptable salts include the conventional non-toxic salts or quaternary ammonium salts of the parent compound, for example salts formed from non-toxic inorganic or organic acids.
  • non-toxic salts include, but are not limited to, those derived from inorganic acids and organic acids selected from the group consisting of 2-acetoxybenzoic acid, 2-hydroxyethanesulfonic acid, acetic acid, ascorbic acid, Benzenesulfonic acid, benzoic acid, hydrogencarbonate, carbonic acid, citric acid, edetic acid, ethane disulfonic acid, ethanesulfonic acid, fumaric acid, glucoheptose, gluconic acid, glutamic acid, glycolic acid, Hydrobromic acid, hydrochloric acid, hydroiodide, hydroxyl, hydroxynaphthalene, isethionethane, lactic acid, lactose, dodecylsulfonic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, nitric acid, oxalic acid, Pamoic acid, pantothenic acid, phenylacetic acid, phen
  • the pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound containing an acid group or a base by conventional chemical methods.
  • such salts are prepared by reacting these compounds in water or an organic solvent or a mixture of the two via a free acid or base form with a stoichiometric amount of a suitable base or acid.
  • a nonaqueous medium such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile is preferred.
  • the compounds provided herein also exist in the form of prodrugs.
  • Prodrugs of the compounds described herein are readily chemically altered under physiological conditions to convert to the compounds of the invention. Any compound that can be converted in vivo to provide a biologically active substance (i.e., a compound of formula I) is a prodrug within the scope and spirit of the invention.
  • a compound containing a carboxyl group can form a physiologically hydrolyzable ester which is prepared by hydrolysis in vivo to give the compound of formula I itself.
  • the prodrug is preferably administered orally because hydrolysis occurs in many cases primarily under the influence of digestive enzymes. Parenteral administration can be used when the ester itself is active or hydrolysis occurs in the blood.
  • prodrugs can be converted to the compounds of the invention by chemical or biochemical methods in an in vivo setting.
  • Certain compounds of the invention may exist in unsolvated or solvated forms, including hydrated forms. In general, the solvated forms are equivalent to the unsolvated forms and are included within the scope of the invention. Certain compounds of the invention may exist in polycrystalline or amorphous form.
  • Certain compounds of the invention may have asymmetric carbon atoms (optical centers) or double bonds. Racemates, diastereomers, geometric isomers and individual isomers are included within the scope of the invention.
  • the compounds of the invention may exist in specific geometric or stereoisomeric forms.
  • the present invention contemplates all such compounds, including the cis and trans isomers, the (-)- and (+)-p-enantiomers, the (R)- and (S)-enantiomers, and the diastereomeric a conformation, a (D)-isomer, a (L)-isomer, and a racemic mixture thereof, and other mixtures, such as enantiomerically or diastereomeric enriched mixtures, all of which belong to It is within the scope of the invention.
  • Additional asymmetric carbon atoms may be present in the substituents such as alkyl groups. All such isomers, as well as mixtures thereof, are included within the scope of the invention.
  • optically active (R)- and (S)-isomers as well as the D and L isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If an enantiomer of a compound of the invention is desired, it can be prepared by asymmetric synthesis or by derivatization with a chiral auxiliary wherein the resulting mixture of diastereomers is separated and the auxiliary group cleaved to provide pure The desired enantiomer.
  • a salt of a diastereomer is formed with a suitable optically active acid or base, followed by stepping as is known in the art.
  • the diastereomeric resolution is carried out by crystallization or chromatography, and then the pure enantiomer is recovered.
  • the separation of enantiomers and diastereomers is generally accomplished by the use of chromatography using a chiral stationary phase, optionally in combination with chemical derivatization (eg, formation of an amino group from an amine). Formate).
  • the compounds of the present invention may contain unnatural proportions of atomic isotopes on one or more of the atoms that make up the compound.
  • radiolabeled compounds can be used, such as tritium (3 H), iodine -125 (125 I) or C-14 (14 C). Alterations of all isotopic compositions of the compounds of the invention, whether radioactive or not, are included within the scope of the invention.
  • excipient generally refers to the carrier, diluent and/or vehicle required to formulate an effective pharmaceutical composition.
  • an "effective amount” or “therapeutically effective amount” with respect to a pharmaceutical or pharmacologically active agent refers to a sufficient amount of a drug or agent that is non-toxic but that achieves the desired effect.
  • an "effective amount” of an active substance in a composition refers to the amount required to achieve the desired effect when used in combination with another active substance in the composition. The determination of the effective amount will vary from person to person, depending on the age and general condition of the recipient, and also on the particular active substance, and a suitable effective amount in a case can be determined by one skilled in the art based on routine experimentation.
  • active ingredient refers to a chemical entity that is effective in treating a target disorder, disease or condition.
  • substituted means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, including variants of heavy hydrogen and hydrogen, as long as the valence of the particular atom is normal and the substituted compound is stable.
  • it means that two hydrogen atoms are substituted.
  • Ketone substitution does not occur on the aryl group.
  • optionally substituted means that it may or may not be substituted, and unless otherwise specified, the kind and number of substituents may be arbitrary on the basis of chemically achievable.
  • any variable eg, R
  • its definition in each case is independent.
  • the group may optionally be substituted with at most two R, and each case has an independent option.
  • combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds.
  • one of the variables is selected from a single bond, it means that the two groups to which it is attached are directly linked. For example, when L represents a single bond in A-L-Z, the structure is actually A-Z.
  • alkyl or “alkane” or “alkyl group” are used interchangeably herein, wherein the alkyl group may optionally be one or more Substituted by the substituents described in the invention.
  • the alkyl group contains from 1 to 8 carbon atoms; in other embodiments, the alkyl group contains from 1 to 6 carbon atoms; in other embodiments, the alkyl group contains from 1 to 4 carbon atom.
  • alkyl groups include, but are not limited to, methyl (Me, -CH 3 ), ethyl (Et, -CH 2 CH 3 ), n-propyl (n-Pr, -CH 2 CH 2 CH 3 ), isopropyl (i-Pr, -CH(CH 3 ) 2 ), n-butyl (n-Bu, -CH 2 CH 2 CH 2 CH 3 ), isobutyl (i-Bu, -CH 2 CH) (CH 3 ) 2 ), sec-butyl (s-Bu, -CH(CH 3 )CH 2 CH 3 ), tert-butyl (t-Bu, -C(CH 3 ) 3 ), n-pentyl (-CH) 2 CH 2 CH 2 CH 2 CH 3 ), 2-pentyl (-CH(CH 3 )CH 2 CH 2 CH 3 ), 3-pentyl (-CH(CH 2 CH 3 ) 2 ), 2-methyl -2-butyl (-C(CHCH
  • alkoxy denotes an alkyl group attached to the remainder of the molecule through an oxygen atom, wherein the alkyl group has the meaning as described herein. Unless otherwise specified, the alkoxy group contains from 1 to 6 carbon atoms.
  • alkoxy groups include, but are not limited to, methoxy (MeO, -OCH 3 ), ethoxy (EtO, -OCH 2 CH 3 ), propoxy (n-PrO, n-propoxy) , -OCH 2 CH 2 CH 3 ), isopropyloxy (i-PrO, i-propoxy, -OCH(CH 3 ) 2 ), n-butoxy (n-BuO, n-butoxy) , -OCH 2 CH 2 CH 2 CH 3 ), 1-methylpropyloxy (s-BuO, s-butoxy, -OCH(CH 3 )CH 2 CH 3 ), 2-methyl-l- Propyloxy (i-BuO, i-butoxy, -OCH 2 CH(CH 3 ) 2 ), tert-butyloxy (t-BuO, t-butoxy, -OC(CH 3 ) 3 ), N-pentyloxy (n-pentyloxy, -
  • cycloalkyl denotes a monovalent or polyvalent saturated monocyclic, bicyclic or tricyclic system containing from 3 to 8 carbon atoms. Wherein the cycloalkyl group can be optionally substituted with one or more substituents described herein. In some embodiments, a cycloalkyl group contains from 3 to 8 carbon atoms. Such examples include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and the like.
  • halogen means fluorine (F), chlorine (Cl), bromine (Br) or iodine (I).
  • alkylamino includes “N-alkylamino” and “N,N-dialkylamino” wherein the amino groups are each independently substituted with one or two alkyl groups. Wherein the alkyl group has the meaning as described in the present invention.
  • Suitable alkylamino groups may be monoalkylamino or dialkylamino, examples of which include, but are not limited to, N-methylamino, N-ethylamino, isopropylamino, propylamino, t-butyl Amino group, n-butylamino group, 1-methylpropylamino group, n-pentylamino group, n-hexylamino group, N,N-dimethylamino group, N,N-diethylamino group and the like.
  • alkylmorpholinyl refers to a morpholinyl group substituted with one or more alkyl groups.
  • C 1-6 acylpiperazinyl refers to piperazinyl substituted by one or more C 1-6 acyl groups.
  • hydroxy C 1-6 alkylpiperazinyl refers to a piperazinyl group substituted with one or more hydroxy substituted C 1-6 alkyl groups.
  • alkylaminopiperidinyl refers to a piperidinyl group substituted with one or more alkyl substituted amino groups.
  • reaction solvent used in each reaction step of the present invention is not particularly limited, and any solvent which can dissolve the starting materials to some extent and does not inhibit the reaction is included in the present invention.
  • solvents, solvent combinations, and solvent combinations described herein are considered to be within the scope of the invention.
  • the invention provides a process for the preparation of a compound of the formula ⁇ .
  • the general synthetic method of this class of compounds is to first synthesize a pyrimidine precursor and then connect it through a carbon chain to a nitrogen-containing structure.
  • a process for the preparation of a compound of the formula which comprises the steps of: substituting a compound of the formula 4A with a compound of the formula 5A, and then according to different definitions of the substituent Alternatively, the deprotecting group is converted to the corresponding compound of formula I to provide a compound of formula ⁇ .
  • R 1 is a 5-6 membered cycloalkyl group, a 5-6 membered heterocyclic group, a 5-6 membered aryl group, or a 5-6 membered heteroaryl group
  • the 5-6 membered cycloalkyl group 5-6 membered heterocyclic group, 5-6 membered aryl group and 5-6 membered heteroaryl group are each independently selected from one or more selected from the group consisting of halogen, hydroxy, cyano, nitro, C 1-8 alkyl, C 2-8 alkenyl, C 2-8 alkynyl, C 3-8 cycloalkyl, 3-8 membered heterocyclic, C 5-10 aryl, 5-10 membered heteroaryl, C 1- 6 alkoxy, C 3-8 cycloalkoxy, -S(O)pR 6 , -C(O)R 6 , -C(O)OR 6 , -NR 7 R 8 or -C(O)NR Substit
  • R 2 is hydrogen, halogen, C 1-6 alkyl, C 1-6 alkoxy, halogen substituted C 1-6 alkyl or halogen substituted C 1-6 alkoxy;
  • R 3 is an optionally substituted piperazinyl, an optionally substituted morpholinyl group, an optionally substituted piperidinyl group, an optionally substituted cyclohexylamino group, an optionally substituted C 1-2 alkane Or a substituted 1,2,3,6-tetrahydropyridinyl group, the substituent on the substituted C 1-2 alkyl group being an optionally substituted piperidinyl group;
  • R 4 is hydrogen or C 1-6 alkyl
  • R 5 is hydrogen, chlorine, hydroxy, cyano, C 1-6 alkyl, C 1-6 alkoxy or halogen substituted C 1-6 alkoxy.
  • R 1 is a 5-6 membered heterocyclyl, 5-6 membered aryl, or 5-6 membered heteroaryl substituted by one or more -S(O)pR 6 .
  • R 2 is hydrogen, halogen, C 1-6 alkyl, C 1-6 alkoxy or halogen substituted C 1-6 alkoxy.
  • R 3 is an optionally substituted piperazinyl, an optionally substituted morpholinyl, an optionally substituted piperidinyl, an optionally substituted methyl or optionally substituted a 1,2,3,6-tetrahydropyridinyl group, the substituent on the optionally substituted methyl group being an optionally substituted piperidinyl group;
  • the optionally substituted piperazinyl group, optionally Substituted morpholinyl, optionally substituted piperidinyl or optionally substituted 1,2,3,6-tetrahydropyridinyl is optionally selected from one or more selected from C1-6 alkyl, hydroxy , C 1-6 alkylamino, C 1-6 alkoxy, oxo, C 1-6 acyl, morpholinyl, C 1-6 alkylmorpholinyl, piperazinyl, C 1-6 alkyl Substituted by a substituent of piperazinyl, C 1-6 acyl pipe
  • R 4 is hydrogen or C 1-6 alkyl.
  • R 5 is chloro or halogen substituted C 1-6 alkoxy.
  • R 6 is C l-4 alkyl.
  • p is 2.
  • R 1 is Any of them.
  • R 2 is hydrogen, chlorine, methyl, methoxy, ethoxy, isopropoxy or difluoromethoxy.
  • R 3 is one of the following:
  • R 3 when R 1 is When R 3 can be one of the following:
  • R 4 is hydrogen or methyl.
  • R 5 is chloro or difluoromethoxy.
  • the compound of the formula of the present invention is prepared by the following synthetic route:
  • R1, R2, R3, R4, R5 are as defined in the present invention.
  • the reaction of the compound of the formula II with the compound of the formula III may be carried out by contacting the compound of the formula II with the compound of the formula III in the presence of p-toluenesulfonic acid in an organic solvent.
  • a substitution reaction occurs.
  • the organic solvent can be n-butanol.
  • the synthetic route of the compound of Formula I is:
  • the synthetic route of the compound of Formula I-23 is:
  • the synthetic route of the compound of Formulas I-90 is:
  • the synthetic route of the compound of Formulas I-89 is:
  • the inventors have found that the synthesis method of the embodiment of the invention can rapidly and efficiently prepare the compound of the formula I, and the synthesis route is short, the environment is friendly, the yield and purity of the target product are high, the raw materials are easy to obtain, and the operation and post-treatment are simple. Suitable for industrial production.
  • the invention provides a pharmaceutical composition comprising a therapeutically effective amount of the aforementioned compound.
  • “Pharmaceutical composition” means a mixture comprising one or more of the compounds described herein, or a physiologically/pharmaceutically acceptable salt or prodrug thereof, and other chemical components, or other components, such as a physiological/pharmaceutically acceptable carrier, and excipient.
  • the purpose of the pharmaceutical composition is to promote the administration of the organism, which facilitates the absorption of the active ingredient and thereby exerts biological activity.
  • the pharmaceutical composition contains a compound of formula I as described above.
  • the pharmaceutical composition is for inhibiting a kinase; inhibiting anaplastic lymphoma kinase; treating or preventing cancer; and/or inhibiting proliferation of cancer cells.
  • the cancer is lung cancer and anaplastic large cell non-Hodgkin's lymphoma, preferably the lung cancer is non-small cell lung cancer.
  • the pharmaceutical composition further comprises a second agent different from the compound described above and for inhibiting a kinase, inhibiting anaplastic lymphoma kinase, treating or preventing cancer and/or inhibiting Proliferation of cancer cells.
  • the second agent is combined with a compound as described above to make the pharmaceutical composition more effective for inhibiting kinases, inhibiting anaplastic lymphoma kinase, treating or preventing cancer and/or inhibiting cancer cells Proliferation.
  • the pharmaceutical composition further comprises a pharmaceutically acceptable carrier, excipient, diluent, adjuvant, vehicle, or a combination thereof.
  • the pharmaceutical composition is in the form of a tablet, capsule, injection, powder, powder, syrup, solution, suspension or aerosol.
  • the pharmaceutical composition of the above embodiments of the invention may be present in a suitable solid or liquid carrier or diluent and in a suitable sterilizing device for injection or drip.
  • compositions of the present invention can be prepared according to conventional methods of preparation in the pharmaceutical arts.
  • the compounds and pharmaceutical compositions of the present invention can be administered to mammals clinically, including humans and animals, and can be administered by the route of the mouth, nose, skin, lungs or the gastrointestinal tract. Regardless of the method of administration, the optimal dosage for the individual should be based on the specific treatment regimen. Usually starting with a small dose, gradually increase the dose until the most suitable dose is found. The most preferred route of administration is oral.
  • the invention provides the use of a compound as hereinbefore described, a compound prepared by the method described above or a pharmaceutical composition as described above for the preparation of a medicament.
  • the medicament is for use in at least one of the following: as a kinase inhibitor, inhibiting anaplastic lymphoma kinase, treating or preventing cancer and inhibiting proliferation of cancer cells.
  • the compound of the formula I of the present invention has a significant inhibitory effect on ALK and ALKL1196M enzymes, And it has obvious proliferation inhibitory activity on Karpas299 cells.
  • the compound of the formula I of the present invention has significant proliferation inhibitory activity against Ba/F3EML4-ALK cells.
  • the compounds of the invention have good water solubility in the kinetic solubility test and are superior to the positive control (the control samples described herein are crizotinib and LDK378).
  • the compounds of the invention exhibit good metabolic stability in in vitro metabolic stability assays, providing an important basis for further preclinical studies.
  • the compound of the present invention greatly enhances the membrane permeability relative to the control, thereby improving the absorption properties of the compound in the intestinal tract and improving the bioavailability of the drug.
  • the compounds of the present invention are useful as ALK inhibitors for the preparation of anti-cancer therapeutics that inhibit anaplastic lymphoma kinase.
  • a compound of formula I according to the invention for the manufacture of a medicament for the treatment of cancer, wherein the cancer is lung cancer and anaplastic large cell non-Hodgkin's lymphoma, preferably The cancer is non-small cell lung cancer.
  • the agents of the present invention are effective as ALK inhibitors for the treatment of one or more neoplastic diseases associated with ALK activity, including but not limited to lung cancer.
  • the pyrimidine derivative (the compound of the formula I) as an ALK inhibitor according to the present invention has excellent clinical application and medical use as an ALK inhibitor.
  • the invention provides the use of a compound or pharmaceutical composition as hereinbefore described in at least one of the following: inhibiting a kinase; inhibiting an anaplastic lymphoma kinase; treating or preventing cancer; and inhibiting cancer cells proliferation.
  • the invention provides a method of inhibiting a kinase, inhibiting anaplastic lymphoma kinase, treating or preventing cancer, and/or inhibiting proliferation of cancer cells.
  • the method comprises: administering to the subject an effective amount of a compound or pharmaceutical composition as hereinbefore described.
  • the invention provides a method of treating a neoplastic disease associated with ALK activity.
  • the method comprises: administering to the subject an effective amount of a compound or pharmaceutical composition as hereinbefore described.
  • An embodiment of the invention provides a compound of formula I, or a pharmaceutically acceptable salt, hydrate, solvate, metabolite or prodrug thereof, for the preparation of a compound of formula ⁇ or a pharmaceutically acceptable salt or hydrate thereof , methods and intermediates of solvates or prodrugs, pharmaceutical compositions, and the use of the compounds and pharmaceutical compositions of the invention in the manufacture of a medicament.
  • the compound of the formula 4A-2 (30 mg mg, 0.144 mmol), the compound of the formula 5A-2 (50 mg, 0.144 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (24 mg, 0.145 mmol) was added with stirring. The mixture was heated to 100 ° C and stirred for 5 hours. TLC detection showed the reaction was completed and concentrated under reduced pressure. The obtained crude product was purified by column chromatography to give white-white solid product (27 mg, yield 36.1%). Compound shown in 4.
  • the compound of the formula 4A-3 (30 mg, 0.135 mmol), the compound of the formula 5A-2 (47 mg, 0.135 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (23.3 mg, 0.135) was added with stirring. Ment), heating and heating to 100 ° C, stirring the reaction for 5 hours, TLC detection showed that the reaction was completed, concentrated under reduced pressure, and the obtained crude product was purified by column chromatography to give white-white solid product (59 mg, yield 81.8%). This is the compound of formula I-5.
  • the compound of the formula 4A-3 (30 mg, 0.135 mmol), the compound of the formula 5A-1 (47 mg, 0.135 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (23.3 mg, 0.135) was added with stirring. Ment), heating and heating to 100 ° C, stirring the reaction for 5 hours, TLC detection showed that the reaction was completed, concentrated under reduced pressure, and the obtained crude product was purified by column chromatography to give white-white solid product (56 mg, yield 77.8%). -13 compound shown.
  • the compound of the formula 4A-3 (30 mg, 0.135 mmol), the compound of the formula 5A-3 (48 mg, 0.135 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (23 mg, 0.132 mmol) was added with stirring. The mixture was heated to 100 ° C and stirred for 5 hours. TLC detection showed the reaction was completed and concentrated under reduced pressure. The obtained crude product was purified by column chromatography to afford white solid product (25 mg, yield: 34.3%). The compound shown in 2.
  • the compound of the formula 4A-4 (40 mg, 0.137 mmol), the compound of the formula 5A-3 (48 mg, 0.137 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (24 mg, 0.137 mmol) was added with stirring. The mixture was heated to 100 ° C and stirred for 5 hours. TLC detection showed the reaction was completed and concentrated under reduced pressure. The obtained crude product was purified by column chromatography to give white-white solid product (25 mg, yield 30%). The compound shown in 8.
  • the compound of the formula 4A-6 (30 mg, 0.135 mmol), the compound of the formula 5A-2 (47 mg, 0.135 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (24 mg, 0.135 mmol) was added with stirring. The mixture was heated to 100 ° C and stirred for 5 hours. TLC detection showed the reaction was completed and concentrated under reduced pressure. The obtained crude product was purified by column chromatography to give white-white solid product (41 mg, yield 57%). The compound shown in 9.
  • the compound of the formula 4A-6 (30 mg, 0.135 mmol), the compound of the formula 5A-3 (47 mg, 0.135 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (24 mg, 0.135 mmol) was added with stirring. The mixture was heated to 100 ° C and stirred for 5 hours. TLC showed that the reaction was completed and concentrated under reduced pressure. The obtained crude product was purified by column chromatography to afford white-white solid product (28 mg, yield 38.5%). That is, the compound of the formula I-16.
  • the compound of the formula 4A-8 (40 mg, 0.120 mmol), the compound of the formula 5A-2 (42 mg, 0.120 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (21 mg, 0.120 mmol) was added with stirring. The mixture was heated to 100 ° C and stirred for 5 hours. TLC detection showed the reaction was completed and concentrated under reduced pressure. The obtained crude product was purified by column chromatography to give white-white solid product (20 mg, yield: 25.8%). The compound shown in 15.
  • the compound of the formula 4A-8 (40 mg, 0.120 mmol), the compound of the formula 5A-3 (42 mg, 0.120 mmol) was dissolved in n-butanol (2 mL), and p-toluenesulfonic acid (21 mg, 0.120 mmol) was added with stirring. The mixture was heated to 100 ° C and stirred for 5 hours. TLC detection showed the reaction was completed and concentrated under reduced pressure. The obtained crude product was purified by column chromatography to give white solid product (16 mg, yield: 20.5%). The compound shown in 18.
  • the synthetic route and preparation method are as follows:
  • the third step preparation of 1-isopropyl-4-(3-methoxy-4-nitrophenyl)piperazine
  • the fourth step preparation of 4-(4-isopropylpiperazin-1-yl)-2-methoxyaniline
  • the synthetic route and preparation method are as follows:
  • the synthetic route and preparation method are as follows:
  • the synthetic route and preparation method are as follows:
  • Second step Preparation of 4-(4-amino-5-isopropoxy-2-methyl)-3,6-dihydropiperidine-1(2H)-carboxylic acid tert-butyl ester
  • the synthetic route and preparation method are as follows:
  • Second step Preparation of 4-(5-methoxy-2-methyl-4-nitrophenyl)-3,6-dihydropiperidine-1(2H)-carboxylic acid tert-butyl ester
  • the synthetic route and preparation method are as follows:
  • the synthetic route and preparation method are as follows:
  • the synthetic route and preparation method are as follows:
  • the synthetic route and preparation method are as follows:
  • the synthetic route and preparation method are as follows:
  • Example 33 5-Chloro-N 2 -(2-(difluoromethoxy)-4-(4-(dimethylamino)piperidin-1-yl)-5-methylphenyl)-N preparation of (2- (isopropylsulfonyl) phenyl) (compounds of formula I-102) pyrimidine-2,4-diamine 4 -
  • reaction mixture was cooled to room temperature, diluted with EtOAc EtOAc EtOAc EtOAc EtOAcjHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHH
  • Example 37 5-(Difluoromethoxy)-N 2 -(4-(4-(dimethylamino)piperidin-1-yl)-2-methoxyphenyl)-N 4 - ( Preparation of 2-(isopropylsulfonyl)phenyl)pyrimidine-2,4-diamine (compound of formula I-111)
  • the synthetic route and preparation method are as follows:
  • the synthetic route and preparation method are as follows:
  • the third step preparation of 2-chloro-5-(difluoromethoxy)-N-(2-(isopropylsulfonyl)phenyl)pyrimidine-4-amine
  • Example 38 5-(Difluoromethoxy)-N 2 -(4-(4-(dimethylamino)piperidin-1-yl)-2-methoxy-5-methylphenyl) Preparation of -N 4 -(2-(isopropylsulfonyl)phenyl)pyrimidine-2,4-diamine (compound of formula I-112)
  • Example 39 5-(Difluoromethoxy)-N 2 -(4-(4-(dimethylamino)piperidin-1-yl)-2-isopropoxyphenyl)-N 4 - Preparation of (2-(isopropylsulfonyl)phenyl)pyrimidine-2,4-diamine (compound of formula I-113)
  • Example 40 5-Chloro-N 2 -(2-(difluoromethoxy)-4-(4-(dimethylamino)piperidin-1-yl)phenyl)-N 4 -(2 -(Isopropoxysulfonyl)phenyl)pyrimidine-2,4-diamine (Compound of formula I-114)
  • Example 41 5-(Difluoromethoxy)-N 2 -(4-(4-isopropylpiperazin-1-yl)-2-methoxy-5-methylphenyl)-N 4 -(2-(Isopropylsulfonyl)phenyl)pyrimidine-2,4-diamine (Compound of formula I-115)
  • Example 43 Determination of ALK kinase inhibitory activity by a compound of formula I
  • the inhibitory activity of the compound of the present invention against ALK kinase in vitro was measured by the following method, which is expressed by the index of IC50, which is the concentration of the compound when the activity of ALK kinase is inhibited by 50%.
  • Staurosporine (Sigma, Cat. No. S4400-1MG, Lot. No. 046K4080)
  • the compound was mixed into the intermediate plate and shaken for 10 minutes.
  • A1 in a 96-well plate was transferred to A1 and A2 in a 384-well plate.
  • A2 in a 96-well plate was transferred to A3 and A4 in a 384-well plate.
  • the kinase was added to 1X kinase buffer to form a 2.5 fold enzyme solution.
  • the FAM-labeled polypeptide and ATP were added to 1X kinase buffer to form a 2.5-fold substrate solution.
  • test plate already contained 5 ⁇ l of a 10% solution of the compound in DMSO.
  • the conversion rate data is read on Caliper.
  • the biochemical activity of the compounds of the present invention was determined by the above test, and the experimental results of the IC50 values measured by the compounds of the present invention are shown in Table 1 below.
  • the IC50 of the compound designated as "A” for the ALK inhibitory activity is ⁇ 1 nM; the compound having the activity designated “B” provides an IC50 of 1-10 nM; and the compound designated as “C” for the activity provides an IC50 of IC50 ⁇ 10 nM .
  • Compounds designated as "a'” for ALK L1196M inhibitory activity provide IC50 ⁇ 1 nM; compounds designated as “b” for activity provide IC50 of 1-10 nM; compounds designated as "c” for activity provide IC50 of IC50 ⁇ 10 nM .
  • the above test results show that the compound of the formula I of the present invention has a significant inhibitory effect on the activity of ALK and ALKL1196M.
  • the results show that the compounds of the formula I according to the invention all have good ALK kinase inhibitory activity, and the compounds of the invention are useful as ALK inhibitors for the treatment of one or more neoplastic diseases associated with ALK activity, An anti-tumor therapeutic agent for the preparation of an inhibitor of anaplastic lymphoma kinase.
  • Example 44 Determination of Proliferation Inhibitory Activity of Karpas299 Cells by a Compound of Formula I
  • the amount of ATP directly reflects the number of cells and the state of the cells, and the number of viable cells in the culture is detected by quantitatively measuring ATP.
  • the live cell assay kit uses luciferase as a test substance.
  • the kit uses a stable glow-type signal generated by UltraGlow luciferase. During the luminescence process, luciferase requires the participation of ATP, and the respiration of metabolically active cells. And other life activity processes can produce ATP.
  • the assay plate was analyzed using PE company's Envision.
  • Detection reagent live cell detection kit CellTiter-Glo
  • Karpas299 cells were seeded in black 384-well plates at 45 [mu]l cell suspension per well containing 2500 cells per well. The cell plates were placed in a carbon dioxide incubator for overnight culture.
  • test compound was dissolved in a 10 mM DMSO solution and placed in a nitrogen cabinet for long-term storage.
  • Table 2 shows the proliferation inhibitory activity of the compound of the present invention against Karpas299 cells.
  • the compound designated as "D” has an IC50 ⁇ 50 nM; the compound designated as “E” has an IC50 of 50-100 nM; the activity is designated as "F”.
  • the compound provides an IC50 of IC50 ⁇ 100 nM.
  • the above test results indicate that the compound of the formula I of the present invention has significant inhibitory activity against the ALK enzyme and has significant proliferation inhibitory activity against Karpas 299 cells.
  • Example 45 Determination of Proliferation Inhibitory Activity of Ba/F3EML4-ALK Cells by a Compound of Formula I
  • Cell culture RPMI-1640 medium, fetal bovine serum, antibiotic (Penicillin-Streptomycin), IL-3, puromycin
  • the amount of ATP directly reflects the number of cells and the state of the cells, and the number of viable cells in the culture is detected by quantitatively measuring ATP.
  • the live cell assay kit uses luciferase as a test substance.
  • the kit uses a stable glow-type signal generated by UltraGlow luciferase. During the luminescence process, luciferase requires the participation of ATP, and the respiration of metabolically active cells. And other life activity processes can produce ATP.
  • the assay plate was analyzed using PE company's Envision.
  • Ba/F3EML4-ALK cells were separately seeded in 384-well plates containing 300 cells per well. The cell plates were placed in a carbon dioxide incubator for overnight culture.
  • a double-fold experiment was performed using a three-fold dilution of BRAVO and a concentration of 10 compounds.
  • Compounds were transferred to cell plates at a starting concentration of 5 [mu]M.
  • the cell plates were incubated in a carbon dioxide incubator for 3 days.
  • the Promega CellTiter-Glo reagent was added to the cell plate and incubated for 10 minutes at room temperature to stabilize the luminescence signal. Readings were performed using a PerkinElmer Envision multi-label analyzer.
  • Table 3 shows the test results of the compounds of the present invention. Wherein the compound designated as “d” for activity provides an IC50 ⁇ 20 nM; the compound designated as “e” for activity provides an IC50 of 20-100 nM; the compound designated as “f” for activity provides an IC50 of IC50 > 100 nM.
  • test results show that the compound of the present invention has significant proliferation inhibitory activity against Ba/F3EML4-ALK cells.
  • Test method Weighed quantitative compound samples were dissolved in pure DMSO to a final concentration of 10 mM, and the test compound and the control compound (10 mM DMSO mother liquor, 10 ⁇ L per well) were added to a 96-well plate containing 490 ⁇ L of buffer per well. After 2 minutes of vortexing, the sample plates were incubated for 24 hours at room temperature (22 ⁇ 2 °C) on a shaker. 200 ⁇ L of the sample was then transferred to a MultiScreen filter plate (polycarbonate membrane), filtered through a millipore vacuum manifold and the filtrate was collected. The concentration of the compound in the filtrate was determined by HPLC-UV. Three different concentrations of the UV standard solution and the solubility test sample were injected successively. Each sample was inserted twice, and the standard curve was taken to calculate the concentration and averaged.
  • the in vitro metabolic stability assay assesses the clearance of a compound in one-phase metabolism and predicts its intrinsic clearance in hepatocytes and in vivo.
  • Buffer 100 nM potassium phosphate buffer, pH 7.4; 10 mM MgCl 2 ;
  • Stop solution cold acetonitrile (CAN) contains 100 ng/mL tolbutamide (Lolbutamide) and 100 ng/mL Labetalol as internal standard (IS)
  • liver microsome solution (0.625 mg/mL) was dispensed into the corresponding reaction plate in a volume of 80 ⁇ L per well using a liquid workstation. (final concentration of microsomes is 0.5mg/mL)
  • Caco-2 cells are an in vitro model widely used to study intestinal absorption, a human colon cancer cell.
  • the monolayer Caco-2 cell model has been widely used to assess passive and active transport processes during intestinal absorption.
  • GF120918A is a strong efflux transporter inhibitor, and these efflux transporters include P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP).
  • P-gp P-glycoprotein
  • BCRP breast cancer resistance protein
  • control samples were crizotinib and coloritinib (LDK378).
  • the experimental results show that the compound of the present invention greatly enhances the membrane permeability relative to the control, thereby improving the compound in the intestinal tract. Absorbs properties and increases the bioavailability of the drug.

Abstract

本发明提出了嘧啶类衍生物及其用途。该嘧啶类衍生物为式I所示化合物或式I所示化合物的药学上可接受的盐、水合物、溶剂化物、代谢产物、或前药，其中，R1、R2、R3、R4和R5如说明书所定义。该化合物能够用作ALK抑制剂，用于制备成抑制间变性淋巴瘤激酶的抗肿瘤治疗药物。

Description

嘧啶类衍生物及其用途 技术领域
本发明属于生物医药领域，具体的，本发明涉及嘧啶类衍生物及其用途，更具体的，本发明涉及一种嘧啶类衍生物及其制备方法，以及其在制备药物中的用途。
背景技术
非小细胞肺癌(Non-small-cell carcinoma,NSCLC)与“非小细胞癌”同义。非小细胞型肺癌，包括鳞癌、腺癌、大细胞癌，与小细胞癌相比其癌细胞生长***较慢，扩散转移相对较晚。非小细胞肺癌约占肺癌总数的80-85％。数据显示，目前我国肺癌发病率每年增长26.9％，自2000年至2005年间，中国肺癌的发病人数估计增加12万人。其中，男性肺癌病人从2000年的26万人增加到2005年的33万人，同期女性肺癌患者从12万人增加到17万人。此外，肺癌也成为全国多地区的“众癌之首”。北京市肺癌发病率自2001年至2010年增长56％。十年间，北京新发癌症患者中五分之一为肺癌患者；浙江省肿瘤医院发布的2011年浙江省“癌谱”中，肺癌依然是排名第一的癌症；广州地区与30年前相比，肺癌发病率增长了7倍。
随着分子医学进展和靶向药物的不断涌现，晚期NSCLC的治疗已进入到个体化治疗的时代。目前临床应用的个体化靶向治疗主要针对EGFR突变型和ALK(Anaplastic lymphoma kinase，间变性淋巴瘤激酶)融合基因型肺癌，这两种基因变异型肺癌均具有明确的分子靶点、靶点检测技术及上市的靶向药物，临床疗效得到明显提高。
间变性淋巴瘤激酶(ALK)是一种受体酪氨酸蛋白激酶，最早是在间变性大细胞淋巴瘤(ALCL)的一个亚型中被发现的，因此定名为间变性淋巴瘤激酶(Anaplastic Lymphoma Kinase，ALK)(Science,1994,263,1281-1284；Oncogene,1994,9,1567-1574)。ALK融合基因是一种强致癌基因，是新发现的NSCLC驱动基因，其中棘皮动物微管相关类蛋白4(EML4-ALK)为最常见类型。ALK的基因异常(易位、点基因突变和基因扩增)的结果，会导致产生与其他基因相融合的异常激酶并且参与癌变。ALK融合基因主要出现在不吸烟或少吸烟的肺腺癌患者。占NSCLC总数3％～5％的ALK阳性非小胞肺癌是2007年最新发现的分子亚型。
虽然人们已经研究了大量的对蛋白激酶有抑制活性的化合物，且一些蛋白激酶抑制剂如克唑替尼等已经上市用于NSCLC的治疗，但是会产生耐药性，存在一定程度的缺陷。EML4-ALK作为一个新的肿瘤癌症融合基因的概念已被Pfizer第一代ALK抑制剂Crizotinib验证，基于临床试验所表现出的约60％的客观反应率和约10个月的无进展生存期的良好效果，导致了FDA的快速审批，于2011年11月成为第一个用于ALK阳性NSCLC的一线治疗药物。但是治疗9到12个月时，患者会发生耐药。目前认为ALK激酶自身次级突变谱是继发耐药的原因之一，常见有L1196M，G1269A，S1206Y，G1202R，1151Tins，L11152R和C1156Y，临床上被检出频率最高的次级突变是L1196M和G1269A。研究发现约有1/3的Crizotinib耐药患者产生ALK次级突变，并且约有40％ALK阳性患者在继续给予Crizotinib治疗时无明显进展。而且，两项克唑替尼临床研究观察到的最常见的不良反应为视力障碍、恶心、腹泻、呕吐、水肿和便秘，不良反应发生率≥25％。
作为第二代ALK抑制的LDK378由诺华公司开发，在早期的临床试验中对88位以前应用Crizotinib治疗过的ALK非小细胞肺癌患者仍然有80％的反应率。并且于2013年3月获得FDA“突破性治疗”称号。研究发现LDK378抑制Crizotinib耐药突变，并与2014年4月获得FDA批准用于治疗对Crizitinib产生耐药性 的ALK阳性非小细胞肺癌患者，命名为Ceritinib，商品名为Zykadia。研究发现LDK378抑制Crizotinib耐药突变，包括L1196M，G1269A，I1171T和S1206Y，但是该药物没有抑制G1202R和F1174C，还有一定局限性。同时Certinib临床使用剂量大(750mg,P.O qday)。
因而，开发出更安全、高效的治疗癌症的新型ALK抑制剂药物具有巨大的社会价值和经济效益，也是目前各大医药企的研究热点。通过对化合物结构的修饰设计新的化合物结构，改善其耐药性和成药性，从而提高生物活性和生物利用度，来寻找一类新的ALK抑制剂，对于临床上因ALK突变引起的疾病治疗具有重要意义。因此，目前的ALK抑制剂仍有待改进。
发明内容
本发明旨在至少在一定程度上解决上述技术问题之一或至少提供一种有用的商业选择。为此，本发明的一个目的在于提出一种能够用于制备治疗癌症的药物的化合物。
根据本发明的一个方面，本发明提出了一种化合物。根据本发明的实施例，所述化合物为式I所示化合物或式I所示化合物的药学上可接受的盐、水合物、溶剂化物、代谢产物、或前药，
Figure PCTCN2016104723-appb-000001
其中，R1为5-6元环烷基、5-6元杂环基、5-6元芳基、或5-6元杂芳基，任选地，所述5-6元环烷基、5-6元杂环基、5-6元芳基和5-6元杂芳基各自独立地被一个或多个选自卤素、羟基、氰基、硝基、C1-8烷基、C2-8链烯基、C2-8链炔基、C3-8环烷基、3-8元杂环基、C5-10芳基、5-10元杂芳基、C1-6烷氧基、C3-8环烷氧基、-S(O)pR6、-C(O)R6、-C(O)OR6、-NR7R8或-C(O)NR8的取代基取代，其中，R6、R7、R8各自独立地为氢或Cl-4烷基，p为0、1或2；
R2为氢、卤素、C1-6烷基、C1-6烷氧基、卤素取代的C1-6烷基或卤素取代的C1-6烷氧基；
R3为任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基、任选被取代的环己胺基、任选被取代的C1-2烷基或任选被取代的1,2,3,6-四氢吡啶基，所述被取代的C1-2烷基上的取代基为任选被取代的哌啶基；
R4为氢或C1-6烷基；
R5为氢、氯、羟基、氰基、C1-6烷基、C1-6烷氧基或卤素取代的C1-6烷氧基。
根据本发明的实施例，R1为地被一个或多个-S(O)pR6取代的5-6元杂环基、5-6元芳基、或5-6元杂芳基。
根据本发明的实施例，R2为氢、卤素、C1-6烷基、C1-6烷氧基或卤素取代的C1-6烷氧基。
根据本发明的实施例，R3为任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基、任选被取代的甲基或任选被取代的1,2,3,6-四氢吡啶基，所述任选被取代的甲基上的取代基为任选被取代的哌啶基；所述任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基或任选被取代的1,2,3,6-四氢吡啶基任选地被一个或多个选自C1-6烷基、羟基、C1-6烷基氨基、C1-6烷氧基、氧代、C1-6酰基、吗啉基、C1-6烷基吗啉基、哌嗪基、C1-6烷基哌嗪基、C1-6酰基哌嗪基；羟基C1-6烷基哌嗪基、哌啶基和C1-6烷基氨基哌啶基的取代基取代。
根据本发明的实施例，R4为氢或C1-6烷基。
根据本发明的实施例，R5为氯或卤素取代的C1-6烷氧基。
根据本发明的实施例，R6为Cl-4烷基。
根据本发明的实施例，p为2。
根据本发明的实施例，R1为
Figure PCTCN2016104723-appb-000002
中的任意一种。
根据本发明的实施例，R2为氢、氯、甲基、甲氧基、乙氧基、异丙氧基或二氟甲氧基。
根据本发明的实施例，R3为下列之一：
Figure PCTCN2016104723-appb-000003
在本发明的一些实施例中，当R1为
Figure PCTCN2016104723-appb-000004
时，R3可以为下列之一：
Figure PCTCN2016104723-appb-000005
根据本发明的实施例，R4为氢或甲基。
根据本发明的实施例，R5为氯或二氟甲氧基。
本领域技术人员可以理解，根据本领域中使用的惯例，在本申请的结构式中，
Figure PCTCN2016104723-appb-000006
用于描绘化学键，所述化学键为部分或取代基与核心结构或骨架结构相连的点。
由此，在本说明书通篇中，本领域技术人员可对式I所示化合物中所述R1～R5的基团及其取代基、以及p进行选择，以提供本发明的实施例中所述的、稳定的式I所示化合物或其可药用盐、水合物、溶剂化物、代谢产物或前药。
根据本发明的实施例，本发明所述式I所示化合物可以为下列化合物之一，或其药学上可接受的盐、水合物、溶剂化物、代谢产物、或前药：
Figure PCTCN2016104723-appb-000007
Figure PCTCN2016104723-appb-000008
Figure PCTCN2016104723-appb-000009
Figure PCTCN2016104723-appb-000010
Figure PCTCN2016104723-appb-000011
在本发明中所使用的术语，C1-6选自C1、C2、C3、C4、C5和C6；C1-8选自C1、C2、C3、C4、C5、C6、C7和C8；C2-8选自C2、C3、C4、C5、C6、C7和C8；C3-8选自C3、C4、C5、C6、C7和C8；C5-10选自C5、C6、C7、C8、C9和C10。这里所采用的术语“药学上可接受的”，是针对那些化合物、材料、组合物和/或剂型而言，它们在可靠的医学判断的范围之内，适用于与人类和动物的组织接触使用，而没有过多的毒性、刺激性、过敏性反应或其它问题或并发症，与合理的利益/风险比相称。
术语“药学上可接受的盐”是指本发明化合物的盐，由本发明发现的具有特定取代基的化合物与相对无毒的酸或碱制备。当本发明的化合物中含有相对酸性的功能团时，可以通过在纯的溶液或合适的惰性溶剂中用足够量的碱与这类化合物的中性形式接触的方式获得碱加成盐。药学上可接受的碱加成盐包括钠、钾、钙、铵、有机氨或镁盐或类似的盐。当本发明的化合物中含有相对碱性的官能团时，可以通过在纯的溶液或合适的惰性溶剂中用足够量的酸与这类化合物的中性形式接触的方式获得酸加成盐。药学上可接受的酸加成盐的实例包括无机酸盐，所述无机酸包括例如盐酸、氢溴酸、硝酸、碳酸、碳酸氢根、磷酸、磷酸一氢根、磷酸二氢根、硫酸、硫酸氢根、氢碘酸、亚磷酸等；以及有机酸盐，所述有机酸包括如乙酸、丙酸、异丁酸、马来酸、丙二酸、苯甲酸、琥珀酸、辛二酸、反丁烯二酸、乳酸、扁桃酸、邻苯二甲酸、苯磺酸、对甲苯磺酸、柠檬酸、酒石酸和甲磺酸等类似的酸；还包括氨基酸(如精氨酸等)的盐，以及如葡糖醛酸等有机酸的盐(参见Berge et al.,"Pharmaceutical Salts",Journal of Pharmaceutical Science 66:1-19(1977))。本发明的某些特定的化合物含有碱性和酸性的官能团，从而可以被转换成任一碱或酸加成盐。
优选地，以常规方式使盐与碱或酸接触，再分离母体化合物，由此再生化合物的中性形式。化合物的母体形式与其各种盐的形式的不同之处在于某些物理性质，例如在极性溶剂中的溶解度不同。
本文所用的“药学上可接受的盐”属于本发明化合物的衍生物，其中，通过与酸成盐或与碱成盐的方式修饰所述母体化合物。药学上可接受的盐的实例包括但不限于：碱基比如胺的无机酸或有机酸盐、酸根比如羧酸的碱金属或有机盐等等。药学上可接受的盐包括常规的无毒性的盐或母体化合物的季铵盐，例如无毒的无机酸或有机酸所形成的盐。常规的无毒性的盐包括但不限于那些衍生自无机酸和有机酸的盐，所述的无机酸或有机酸选自2-乙酰氧基苯甲酸、2-羟基乙磺酸、乙酸、抗坏血酸、苯磺酸、苯甲酸、碳酸氢根、碳酸、柠檬酸、依地酸、乙烷二磺酸、乙烷磺酸、富马酸、葡庚糖、葡糖酸、谷氨酸、乙醇酸、氢溴酸、盐酸、氢碘酸盐、羟基、羟萘、羟乙磺酸、乳酸、乳糖、十二烷基磺酸、马来酸、苹果酸、扁桃酸、甲烷磺酸、硝酸、草酸、双羟萘酸、泛酸、苯乙酸、磷酸、多聚半乳糖醛、丙酸、水杨酸、硬脂酸、亚乙酸、琥珀酸、氨基磺酸、对氨基苯磺酸、硫酸、单宁、酒石酸和对甲苯磺酸。
本发明的药学上可接受的盐可由含有酸根或碱基的母体化合物通过常规化学方法合成。一般情况下，这样的盐的制备方法是：在水或有机溶剂或两者的混合物中，经由游离酸或碱形式的这些化合物与化学计量的适当的碱或酸反应来制备。一般地，优选醚、乙酸乙酯、乙醇、异丙醇或乙腈等非水介质。
除了盐的形式，本发明所提供的化合物还存在前药形式。本文所描述的化合物的前药容易地在生理条件下发生化学变化从而转化成本发明的化合物。可在体内转化以提供生物活性物质(即式I所示化合物)的任何化合物是在本发明的范围和主旨内的前药。例如，含有羧基的化合物可形成生理上可水解的酯，其通过在体内水解以得到式I所示化合物本身而充当前药。所述前药优选口服给药，这是因为水解在许多情况下主要在消化酶的影响下发生。当酯本身具有活性或水解发生在血液中时，可使用肠胃外给药。此外，前体药物可以在体内环境中通过化学或生化方法被转换到本发明的化合物。
本发明的某些化合物可以以非溶剂化形式或者溶剂化形式存在，包括水合物形式。一般而言，溶剂化形式与非溶剂化的形式相当，都包含在本发明的范围之内。本发明的某些化合物可以以多晶或无定形形式存在。
本发明的某些化合物可以具有不对称碳原子(光学中心)或双键。外消旋体、非对映异构体、几何异构体和单个的异构体都包括在本发明的范围之内。
本文中消旋体、ambiscalemic and scalemic或者对映体纯的化合物的图示法来自Maehr,J.Chem.Ed.1985,62:114-120。除非另有说明，用楔形键和虚线键表示一个立体中心的绝对构型。当本文所述化合物含有烯属双键或其它几何不对称中心，除非另有规定，它们包括E、Z几何异构体。同样地，所有的 互变异构形式均包括在本发明的范围之内。
本发明的化合物可以存在特定的几何或立体异构体形式。本发明设想所有的这类化合物，包括顺式和反式异构体、(-)-和(+)-对对映体、(R)-和(S)-对映体、非对映异构体、(D)-异构体、(L)-异构体，及其外消旋混合物和其他混合物，例如对映异构体或非对映体富集的混合物，所有这些混合物都属于本发明的范围之内。烷基等取代基中可存在另外的不对称碳原子。所有这些异构体以及它们的混合物，均包括在本发明的范围之内。
可以通过手性合成或手性试剂或者其他常规技术制备光学活性的(R)-和(S)-异构体以及D和L异构体。如果想得到本发明某化合物的一种对映体，可以通过不对称合成或者具有手性助剂的衍生作用来制备，其中将所得非对映体混合物分离，并且辅助基团裂开以提供纯的所需对映异构体。或者，当分子中含有碱性官能团(如氨基)或酸性官能团(如羧基)时，与适当的光学活性的酸或碱形成非对映异构体的盐，然后通过本领域所公知的分步结晶法或色谱法进行非对映异构体拆分，然后回收得到纯的对映体。此外，对映异构体和非对映异构体的分离通常是通过使用色谱法完成的，所述色谱法采用手性固定相，并任选地与化学衍生法相结合(例如由胺生成氨基甲酸盐)。
本发明的化合物可以在一个或多个构成该化合物的原子上包含非天然比例的原子同位素。例如，可用放射性同位素标记化合物，比如氚(3H)，碘-125(125I)或C-14(14C)。本发明的化合物的所有同位素组成的变换，无论放射性与否，都包括在本发明的范围之内。
术语“赋形剂”通常是指配制有效的药物组合物所需要载体、稀释剂和/或介质。
针对药物或药理学活性剂而言，术语“有效量”或“治疗有效量”是指无毒的但能达到预期效果的药物或药剂的足够用量。对于本发明中的口服剂型，组合物中一种活性物质的“有效量”是指与该组合物中另一种活性物质联用时为了达到预期效果所需要的用量。有效量的确定因人而异，取决于受体的年龄和一般情况，也取决于具体的活性物质，个案中合适的有效量可以由本领域技术人员根据常规试验确定。
术语“活性成分”、“治疗剂”，“活性物质”或“活性剂”是指一种化学实体，它可以有效地治疗目标紊乱、疾病或病症。
术语“被取代的”是指特定原子上的任意一个或多个氢原子被取代基取代，包括重氢和氢的变体，只要特定原子的价态是正常的并且取代后的化合物是稳定的。当取代基为酮基(即＝O)时，意味着两个氢原子被取代。酮取代不会发生在芳香基上。术语“任选被取代的”是指可以被取代，也可以不被取代，除非另有规定，取代基的种类和数目在化学上可以实现的基础上可以是任意的。
当任何变量(例如R)在化合物的组成或结构中出现一次以上时，其在每一种情况下的定义都是独立的。因此，例如，如果一个基团被0-2个R所取代，则所述基团可以任选地至多被两个R所取代，并且每种情况下的R都有独立的选项。此外，取代基和/或其变体的组合只有在这样的组合会产生稳定的化合物的情况下才是被允许的。当其中一个变量选自单键时，表示其连接的两个基团直接相连，比如A-L-Z中L代表单键时表示该结构实际上是A-Z。
在本发明的上下文中，所有涉及到的术语“烷基”或“烷”或“烷基基团”在此处可交换使用，其中，所述烷基可以任选地被一个或多个本发明描述的取代基所取代。一些实施方案中，烷基基团含有1-8个碳原子；另一些实施方案中，烷基基团含有1-6个碳原子；另一些实施方案中，烷基基团含有1-4个碳原子。烷基基团的实例包含，但并不限于：甲基(Me、-CH3)、乙基(Et、-CH2CH3)、正丙基(n-Pr、-CH2CH2CH3)、异丙基(i-Pr、-CH(CH3)2)、正丁基(n-Bu、-CH2CH2CH2CH3)、异丁基(i-Bu、-CH2CH(CH3)2)、仲丁基(s-Bu、-CH(CH3)CH2CH3)、叔丁基(t-Bu、-C(CH3)3)、正戊基(-CH2CH2CH2CH2CH3)、2-戊基(-CH(CH3)CH2CH2CH3)、3-戊基(-CH(CH2CH3)2)、2-甲基-2-丁基(-C(CH3)2CH2CH3)、3-甲基-2-丁基(-CH(CH3)CH(CH3)2)、3-甲基-1-丁基(-CH2CH2CH(CH3)2)、2-甲基-1- 丁基(-CH2CH(CH3)CH2CH3)、正己基(-CH2CH2CH2CH2CH2CH3)、2-己基(-CH(CH3)CH2CH2CH2CH3)、3-己基(-CH(CH2CH3)(CH2CH2CH3))、2-甲基-2-戊基(-C(CH3)2CH2CH2CH3)、3-甲基-2-戊基(-CH(CH3)CH(CH3)CH2CH3)、4-甲基-2-戊基(-CH(CH3)CH2CH(CH3)2)、3-甲基-3-戊基(-C(CH3)(CH2CH3)2)、2-甲基-3-戊基(-CH(CH2CH3)CH(CH3)2)、2,3-二甲基-2-丁基(-C(CH3)2CH(CH3)2)、3,3-二甲基-2-丁基(-CH(CH3)C(CH3)3)、正庚基、正辛基等。
术语“烷氧基”表示烷基基团通过氧原子与分子其余部分相连，其中烷基基团具有如本发明所述的含义。除非另外详细说明，所述的烷氧基基团含有1-6个碳原子。烷氧基基团的实例包括，但并不限于：甲氧基(MeO、-OCH3)、乙氧基(EtO、-OCH2CH3)、丙氧基(n-PrO、n-丙氧基、-OCH2CH2CH3)、异丙基氧基(i-PrO、i-丙氧基、-OCH(CH3)2)、正丁氧基(n-BuO、n-丁氧基、-OCH2CH2CH2CH3)、1-甲基丙基氧基(s-BuO、s-丁氧基、-OCH(CH3)CH2CH3)、2-甲基-l-丙氧基(i-BuO、i-丁氧基、-OCH2CH(CH3)2)、叔丁基氧基(t-BuO、t-丁氧基、-OC(CH3)3)、正戊氧基(n-戊氧基、-OCH2CH2CH2CH2CH3)、2-戊氧基(-OCH(CH3)CH2CH2CH3)、3-戊氧基(-OCH(CH2CH3)2)、2-甲基-2-丁氧基(-OC(CH3)2CH2CH3)、3-甲基-2-丁氧基(-OCH(CH3)CH(CH3)2)、3-甲基-l-丁氧基(-OCH2CH2CH(CH3)2)、2-甲基-l-丁氧基(-OCH2CH(CH3)CH2CH3)、正己氧基(n-己氧基、-OCH2CH2CH2CH2CH2CH3)等等。
术语“环烷基”表示含有3-8个碳原子的，单价或多价的饱和单环、双环或三环体系。其中，所述环烷基基团可以任选地被一个或多个本发明描述的取代基所取代。在一些实施方案中，环烷基包含3-8个碳原子。这样的实例包括，但并不限于：环丙基、环丁基、环戊基、环己基、环庚基、环辛基等。
术语“卤素”是指氟(F)、氯(Cl)、溴(Br)或碘(I)。
术语“烷基氨基”包括“N-烷基氨基”和“N,N-二烷基氨基”，其中，氨基基团分别独立地被一个或两个烷基基团所取代。其中，烷基基团具有如本发明所述的含义。合适的烷基氨基基团可以是单烷基氨基或二烷基氨基，这样的实例包括，但并不限于：N-甲氨基、N-乙氨基、异丙基氨基、丙氨基、叔丁基氨基、正丁氨基、1-甲基丙基氨基、正戊基氨基、正己基氨基、N,N-二甲氨基、N,N-二乙氨基等。
术语“C1-6酰基”是指R-C(＝O)-，其中R为C1-6烷基，C1-6烷基具有如本发明所述的含义。
术语“烷基吗啉基”是指被一个或多个烷基取代的吗啉基。
术语“C1-6酰基哌嗪基”是指被一个或多个C1-6酰基取代的哌嗪基。
术语“羟基C1-6烷基哌嗪基”是指被一个或多个羟基取代的C1-6烷基取代的哌嗪基。
术语“烷基氨基哌啶基”是指被一个或多个烷基取代的氨基取代的哌啶基。
术语“包含”为开放式表达，即包括本发明所指明的内容，但并不排除其他方面的内容。
本发明所述的各反应步骤所使用的反应溶剂没有特别限制，任何在一定程度上能溶解起始原料并且不抑制反应的溶剂均包含在本发明中。另外，本领域的许多类似改动，等同替换，或等同于本发明所描述的溶剂，溶剂组合，及溶剂组合的不同比例，均视为本发明的包含范围。
根据本发明的第二方面，本发明提出了一种制备式Ι所示化合物的方法。本类化合物的通用合成方法是先合成嘧啶类母体，然后通过一个碳链与含氮类结构连接而成。
根据本发明的实施例，本发明提供一种制备式Ι所示化合物的方法，该制备方法包括如下步骤：式4A所示化合物与式5A所示化合物发生取代反应，然后根据取代基的不同定义或脱保护基转换成相应的式I化合物，得到式Ι所示化合物。
Figure PCTCN2016104723-appb-000012
其中，R1为5-6元环烷基、5-6元杂环基、5-6元芳基、或5-6元杂芳基，任选地，所述5-6元环烷基、5-6元杂环基、5-6元芳基和5-6元杂芳基各自独立地被一个或多个选自卤素、羟基、氰基、硝基、C1-8烷基、C2-8链烯基、C2-8链炔基、C3-8环烷基、3-8元杂环基、C5-10芳基、5-10元杂芳基、C1-6烷氧基、C3-8环烷氧基、-S(O)pR6、-C(O)R6、-C(O)OR6、-NR7R8或-C(O)NR8的取代基取代，其中，R6、R7、R8各自独立地为氢或Cl-4烷基，p为0、1或2；
R2为氢、卤素、C1-6烷基、C1-6烷氧基、卤素取代的C1-6烷基或卤素取代的C1-6烷氧基；
R3为任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基、任选被取代的环己胺基、任选被取代的C1-2烷基或任选被取代的1,2,3,6-四氢吡啶基，所述被取代的C1-2烷基上的取代基为任选被取代的哌啶基；
R4为氢或C1-6烷基；
R5为氢、氯、羟基、氰基、C1-6烷基、C1-6烷氧基或卤素取代的C1-6烷氧基。
根据本发明的实施例，R1为被一个或多个-S(O)pR6取代的5-6元杂环基、5-6元芳基、或5-6元杂芳基。
根据本发明的实施例，R2为氢、卤素、C1-6烷基、C1-6烷氧基或卤素取代的C1-6烷氧基。
根据本发明的实施例，R3为任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基、任选被取代的甲基或任选被取代的1,2,3,6-四氢吡啶基，所述任选被取代的甲基上的取代基为任选被取代的哌啶基；所述任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基或任选被取代的1,2,3,6-四氢吡啶基任选地被一个或多个选自C1-6烷基、羟基、C1-6烷基氨基、C1-6烷氧基、氧代、C1-6酰基、吗啉基、C1-6烷基吗啉基、哌嗪基、C1-6烷基哌嗪基、C1-6酰基哌嗪基；羟基C1-6烷基哌嗪基、哌啶基和C1-6烷基氨基哌啶基的取代基取代。
根据本发明的实施例，R4为氢或C1-6烷基。
根据本发明的实施例，R5为氯或卤素取代的C1-6烷氧基。
根据本发明的实施例，R6为Cl-4烷基。
根据本发明的实施例，p为2。
根据本发明的实施例，R1为
Figure PCTCN2016104723-appb-000013
中的任意一种。
根据本发明的实施例，R2为氢、氯、甲基、甲氧基、乙氧基、异丙氧基或二氟甲氧基。
根据本发明的实施例，R3为下列之一：
Figure PCTCN2016104723-appb-000014
在本发明的一些实施例中，当R1为
Figure PCTCN2016104723-appb-000015
时，R3可以为下列之一：
Figure PCTCN2016104723-appb-000016
根据本发明的实施例，R4为氢或甲基。
根据本发明的实施例，R5为氯或二氟甲氧基。
根据本发明的实施例，本发明所述式Ι所示化合物，是通过如下的合成路线制备得到的：
Figure PCTCN2016104723-appb-000017
其中，R1、R2、R3、R4、R5如本发明所定义。
根据本发明的一些实施例，使式II所示化合物与式III所示化合物发生的反应可以是在有机溶剂中、存在对甲苯磺酸条件下、式II所示化合物与式III所示化合物接触发生取代反应。在本发明的一些实施例中，有机溶剂可以为正丁醇。由此，可以为式II所示化合物与式III所示化合物提供合适的反应条件与环境，有利于提高目标产物的收率和反应效率，减少副反应发生。
在本发明的一些实施例中，式I所示化合物的合成路线为：
Figure PCTCN2016104723-appb-000018
在本发明的一些实施例中，式I-23所示化合物的合成路线为：
Figure PCTCN2016104723-appb-000019
在本发明的一些实施例中，式I-90所示化合物的合成路线为：
Figure PCTCN2016104723-appb-000020
在本发明的一些实施例中，式I-89所示化合物的合成路线为：
Figure PCTCN2016104723-appb-000021
发明人发现，利用本发明实施例的合成方法能够快速有效地制备式I所示化合物，且合成路线短、环境友好、目标产物的收率和纯度较高，原料易得、操作及后处理简单、适合工业化生产。
根据本发明的第三方面，本发明提出了一种药物组合物，所述药物组合物含有治疗有效剂量的前述化合物。
“药物组合物”表示含有一种或多种本文所述化合物或其生理学上/可药用盐或前体药物与其他化学组分的混合物，或其他组分例如生理学/可药用的载体和赋形剂。药物组合物的目的是促进对生物体的给药，利于活性成分的吸收进而发挥生物活性。
根据本发明的具体实施例，该药物组合物含有前面所述的式I所示化合物。根据本发明的实施例，所述药物组合物用于抑制激酶；抑制间变性淋巴瘤激酶；治疗或者预防癌症；和/或抑制癌细胞的增殖。所述癌症为肺癌和间变性大细胞非霍奇金淋巴瘤，优选地，所述肺癌为非小细胞肺癌。
根据本发明的实施例，所述药物组合物进一步包括第二药剂，所述第二药剂不同于前面所述化合物并且用于抑制激酶、抑制间变性淋巴瘤激酶、治疗或者预防癌症和/或抑制癌细胞的增殖。根据本发明的实施例，第二药剂与前面所述化合物的联用，使得所述药物组合物更加有效地用于抑制激酶、抑制间变性淋巴瘤激酶、治疗或者预防癌症和/或抑制癌细胞的增殖。
根据本发明的具体示例，药物组合物进一步包含药学上可接受的载体、赋形剂、稀释剂、辅剂、媒介物或其组合。
根据本发明的具体实施例，药物组合物呈片剂、胶囊、注射剂、粉针剂、粉剂、糖浆、溶液状、悬浮液或气雾剂。由此可以显著提高该药物组合物的适用性。并且本发明上述实施例的药物组合物可以存在于适宜的固体或液体的载体或稀释液中和适宜的用于注射或滴注的消毒器具中。
本发明的药物组合物的各种剂型可按照药学领域的常规制备方法制备。本发明的化合物和药物组合物可对哺乳动物临床使用，包括人和动物，可以通过口、鼻、皮肤、肺或者胃肠道等的途径给药。不管采用何种服用方法，个人的最佳剂量应依据具体的治疗方案而定。通常情况下是从小剂量开始，逐渐增加剂量一直到找到最适合的剂量。最优选的给药途径为口服。
根据本发明的第四方面，本发明提出了前面所述的化合物、前面所述的方法制备得到的化合物或前面所述的药物组合物在制备药物中的用途。
根据本发明的具体实施例，所述药物用于下列至少之一：用作激酶抑制剂，抑制间变性淋巴瘤激酶、治疗或者预防癌症和抑制癌细胞的增殖。
根据本发明的具体示例，本发明所述式I所示化合物对ALK及ALKL1196M酶有明显的抑制作用， 并且对Karpas299细胞具有明显的增殖抑制活性。
根据本发明的具体示例，本发明所述式I所示化合物对Ba/F3EML4-ALK细胞具有明显的增殖抑制活性。
根据本发明的具体示例，在动力学溶解度测试中，本发明所述化合物具有良好的水溶性质，且优于阳性对照样(本发明所述的对照样为克唑替尼和LDK378)。
根据本发明的具体示例，在体外代谢稳定性测试中，本发明所述化合物显示出了良好的代谢稳定性，为进一步的临床前研究提供了重要依据。
根据本发明的具体示例，在Caco-2实验中，本发明所述化合物相对于对照样大大提高了透膜性，从而改善了化合物在肠道中的吸收性质，并能提高药物的生物利用度。
本发明所述化合物可用作ALK抑制剂，用于制备成抑制间变性淋巴瘤激酶的抗癌症治疗药物。
根据本发明的具体实施例，本发明所述的式I所示化合物在制备治疗癌症的药物中的应用，其中所述癌症为肺癌和间变性大细胞非霍奇金淋巴瘤，优选地，所述癌症为非小细胞肺癌。
因此，本发明所述药物能够有效作为ALK抑制剂，用于治疗一种或一种以上与ALK活性有关的肿瘤疾病，所述肿瘤疾病包括但不限于肺癌。本发明所述的作为ALK抑制剂的嘧啶衍生物(式I所示化合物)，其作为ALK抑制剂，具有良好的临床应用和医药用途。
在本发明的第五方面，本发明提供了前面所述的化合物或药物组合物在下列至少之一中的用途：抑制激酶；抑制间变性淋巴瘤激酶；治疗或者预防癌症；以及抑制癌细胞的增殖。
在本发明的第六方面，本发明提供了一种抑制激酶、抑制间变性淋巴瘤激酶、治疗或者预防癌症和/或抑制癌细胞的增殖的方法。根据本发明的实施例，该方法包括：给予受试对象有效剂量的前面所述的化合物或药物组合物。
在本发明的第七方面，本发明提供了一种治疗与ALK活性有关的肿瘤疾病的方法。根据本发明的实施例，该方法包括：给予受试对象有效剂量的前面所述的化合物或药物组合物。
本发明的附加方面和优点将在下面的描述中部分给出，部分将从下面的描述中变得明显，或通过本发明的实践了解到。
具体实施方式
下面将结合实施例对本发明的方案进行解释。本领域技术人员将会理解，下面的实施例仅用于说明本发明，而不应视为限定本发明的范围。实施例中未注明具体技术或条件的，按照本领域内的文献所描述的技术或条件或者按照产品说明书进行。所用试剂或仪器未注明生产厂商者，均为可以通过市购获得的常规产品。
本发明的实施例提供了式I所示化合物或其药学上可接受的盐、水合物、溶剂化物、代谢物或前药，制备式Ι所示化合物或其药学上可接受的盐、水合物、溶剂化物或前药的方法和中间体、药物组合物、以及本发明的化合物和药物组合物在制备药物中的用途。
Figure PCTCN2016104723-appb-000022
实施例1式I-17所示化合物的制备流程
Figure PCTCN2016104723-appb-000023
1、式3A-1所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-1所示化合物(0.5g,2.52mmol)和碳酸钾(807mg，5.84mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-1所示化合物粗品(0.8g，产率78.4％)无需纯化，可直接用于下一步反应。
2、式4A-1所示化合物的制备
式3A-1所示化合物(0.8g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g)，室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析分离得式4A-1所示化合物(600mg，产率82.3％)。
LCMS:t＝0.079min，292.3(M+H+)。
3、式I-17所示化合物的制备
将式4A-1所示化合物(42mg，0.131mmol)，式5A-1所示化合物(45mg，0.131mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(23mg，0.132mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法分离得到白色固体产物(30mg，产率41.1％)，即为式I-17所示化合物。
1H NMR(400MHz,cd3od)δ8.51(d,J＝8.3Hz,1H),8.08(s,1H),7.87(d,J＝8.0Hz,1H),7.63(dd,J＝16.9,8.0Hz,2H),7.31(t,J＝7.7Hz,2H),6.67(d,J＝2.2Hz,1H),6.51(dd,J＝8.7,2.1Hz,1H),3.84(s,3H),3.78-3.65(m,4H),3.20(m,2H),2.73(dd,J＝22.8,11.1Hz,2H),2.30-2.14(m,2H),2.13-2.07(m,2H),1.85–1.52(m,4H),1.22(dd,J＝12.8,6.5Hz,12H).
LCMS:t＝0.730min,629.3(M),630.3(M+1).
实施例2式I-12所示化合物的制备流程
Figure PCTCN2016104723-appb-000024
将式4A-1所示化合物(41.85mg，0.131mmol)，式5A-2所示化合物(45.48mg，0.131mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(23mg，0.132mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(35mg，产率42.4％)， 即为式I-12所示化合物。
1H NMR(400MHz,cd3od)δ9.11(d,J＝8.6Hz,1H),8.34(dd,J＝4.4,1.3Hz,1H),8.10(s,1H),7.49(dd,J＝12.8,6.5Hz,2H),6.68(d,J＝2.4Hz,1H),6.56(dd,J＝8.7,2.5Hz,1H),3.86–3.79(m,3H),3.79–3.72(m,2H),3.72–3.62(m,2H),3.46(d,J＝7.2Hz,1H),2.94(d,J＝11.0Hz,2H),2.72(t,J＝11.5Hz,2H),2.37(d,J＝11.4Hz,2H),2.04(d,J＝12.2Hz,2H),1.94(t,J＝10.9Hz,2H),1.66(ddd,J＝24.2,12.2,3.8Hz,3H),1.30(t,J＝4.8Hz,6H),1.16(d,J＝6.3Hz,6H).
LCMS:t＝0.718min,630.3(M).
实施例3式I-4所示化合物的制备流程
Figure PCTCN2016104723-appb-000025
1、式3A-2所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-2所示化合物(254.5mg,2.92.mmol)和碳酸钾(1.21g，8.77mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-2所示化合物粗品(566mg，产率81.3％)无需纯化，可直接用于下一步反应。
2、式4A-1所示化合物的制备
式3A-2所示化合物(0.54g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g),室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析得式4A-2所示化合物(420mg，产率88.97％)。
3、式I-4所示化合物的制备
将式4A-2所示化合物(30mgmg，0.144mmol)，式5A-2所示化合物(50mg，0.144mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(24mg，0.145mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(27mg，产率36.1％)，即为式I-4所示化合物。
1H NMR(400MHz,cd3od)δ9.10(d,J＝8.6Hz,1H),8.34(dd,J＝4.4,1.3Hz,1H),8.10(s,1H),7.55–7.45(m,2H),6.67(s,1H),6.55(d,J＝7.3Hz,1H),3.85(dd,J＝8.7,3.9Hz,4H),3.83–3.74(m,4H),3.16(s,4H),1.30(d,J＝6.9Hz,6H).
LCMS:t＝0.927min,518.15(M),519.2(M+1)
实施例4式I-1所示化合物的制备流程
Figure PCTCN2016104723-appb-000026
将式4A-2所示化合物(30mg，0.144mmol)，式5A-3所示化合物(50.7mg，0.144mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(24.8mg，0.144mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(25mg，产率33.1％)，即为式I-1所示化合物。
1H NMR(400MHz,cd3od)δ8.10(s,1H),8.06(s,1H),7.81(d,J＝5.5Hz,1H),7.54(d,J＝8.6Hz,1H),6.67(d,J＝2.4Hz,1H),6.56(dd,J＝8.7,2.2Hz,1H),3.86(dd,J＝13.1,8.4Hz,4H),3.82(s,3H),3.36(dt,J＝13.6,6.8Hz,1H),3.20–3.08(m,4H),1.30(t,J＝7.8Hz,6H).
LCMS:t＝0.799min,523.1(M),524.2(M+1)
实施例5式I-5所示化合物的制备流程
Figure PCTCN2016104723-appb-000027
1、式3A-3所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-3所示化合物(0.30g,2.92mmol)和碳酸钾(1.21g，8.77mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-3所示化合物粗品(0.61g，产率83.3％)无需纯化，可直接用于下一步反应。
2、式4A-3所示化合物的制备
式3A-3所示化合物(0.57g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g),室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析得式4A-3所示化合物(406mg，产率80.8％)。
3、式I-4所示化合物的制备
将式4A-3所示化合物(30mg，0.135mmol)，式5A-2所示化合物(47mg，0.135mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(23.3mg，0.135mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(59mg，产率81.8％)， 即为式I-5所示化合物。
1H NMR(400MHz,cd3od)δ9.09(d,J＝8.2Hz,1H),8.33(dd,J＝4.3,1.2Hz,1H),8.09(s,1H),7.54(d,J＝8.7Hz,1H),7.48(dd,J＝8.7,4.3Hz,1H),6.67(d,J＝2.2Hz,1H),6.54(dd,J＝8.7,2.4Hz,1H),3.89–3.73(m,4H),3.27(dd,J＝9.3,4.9Hz,4H),2.89–2.73(m,4H),2.49(s,3H),1.30(d,J＝6.9Hz,6H).
LCMS:t＝0.662min,532.3(M),534.3(M+1).
实施例6式I-13所示化合物的制备流程
Figure PCTCN2016104723-appb-000028
将式4A-3所示化合物(30mg，0.135mmol)，式5A-1所示化合物(47mg，0.135mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(23.3mg，0.135mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(56mg，产率77.8％)，即为式I-13所示化合物。
1H NMR(400MHz,cd3od)δ8.49(d,J＝8.4Hz,1H),8.07(s,1H),7.86(dd,J＝8.0,1.3Hz,1H),7.67(d,J＝8.7Hz,1H),7.60(dd,J＝11.5,4.3Hz,1H),7.30(t,J＝7.6Hz,1H),6.67(s,1H),6.49(dd,J＝8.7,2.1Hz,1H),3.84(s,3H),3.29–3.20(m,4H),2.97–2.80(m,4H),2.55(s,3H),1.23(d,J＝6.8Hz,6H).
LCMS:t＝0.685min,531.3(M),532.3(M+1)
实施例7式I-2所示化合物的制备流程
Figure PCTCN2016104723-appb-000029
将式4A-3所示化合物(30mg，0.135mmol)，式5A-3所示化合物(48mg，0.135mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(23mg，0.132mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(25mg，产率34.3％)，即为式I-2所示化合物。
1H NMR(400MHz,cd3od)δ8.11(s,1H),8.05(s,1H),7.81(d,J＝5.5Hz,1H),7.57(d,J＝8.6Hz,1H),6.68(d,J＝2.3Hz,1H),6.57(dd,J＝8.7,2.4Hz,1H),3.83(s,3H),3.42–3.34(m,1H),3.29(s,4H),2.89(s,4H),2.55(s,3H),1.30(d,J＝6.7Hz,6H).
LCMS:t＝0.690min,537.2(M),538.2(M+1).
实施例8式I-8所示化合物的制备流程
Figure PCTCN2016104723-appb-000030
1、式3A-4所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-4所示化合物(0.497,2.92mmol)和碳酸钾(1.21g，8.77mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-4所示化合物粗品(0.756g，产率80.5％)无需纯化，可直接用于下一步反应。
2、式4A-4所示化合物的制备
式3A-4所示化合物(0.73g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g),室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析得式4A-4所示化合物(536mg，产率80.98％)。
3、式I-4所示化合物的制备
将式4A-4所示化合物(40mg，0.137mmol)，式5A-3所示化合物(48mg，0.137mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(24mg，0.137mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(25mg，产率30％)，即为式I-8所示化合物。
1H NMR(400MHz,cd3od)δ8.20(s,1H),8.04(d,J＝4.9Hz,2H),7.48(d,J＝8.3Hz,1H),7.14(s,1H),6.99(d,J＝8.2Hz,1H),4.11(d,J＝13.3Hz,2H),3.97(t,J＝12.4Hz,2H),3.94–3.81(m,5H),3.60(d,J＝11.9Hz,3H),3.45(dt,J＝13.6,6.8Hz,1H),3.28–3.21(m,2H),2.43(d,J＝12.7Hz,2H),2.17(d,J＝11.8Hz,2H),1.30(dd,J＝15.3,5.5Hz,6H).
LCMS:t＝0.693min,607.3(M),608.3(M+1),609.3(M+2).
实施例9式I-7所示化合物的制备流程
Figure PCTCN2016104723-appb-000031
将式4A-4所示化合物(40mg，0.137mmol)，式5A-2所示化合物(47mg，0.137mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(24mg，0.137mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(23mg，产率27.8％)， 即为式I-7所示化合物。
1H NMR(400MHz,cd3od)δ9.11(d,J＝8.0Hz,1H),8.34(dd,J＝4.4,1.4Hz,1H),8.10(s,1H),7.49(dd,J＝12.8,6.5Hz,2H),6.68(d,J＝2.5Hz,1H),6.56(dd,J＝8.7,2.5Hz,1H),3.88–3.80(m,4H),3.75(dd,J＝13.4,9.0Hz,6H),2.83–2.60(m,6H),2.42(t,J＝11.4Hz,1H),2.07(d,J＝12.2Hz,2H),1.66(ddd,J＝24.2,12.4,4.0Hz,2H),1.29(t,J＝7.0Hz,6H).
LCMS:t＝0.682min,602.2(M),603.2(M+1).
实施例10式I-6所示化合物的制备流程
Figure PCTCN2016104723-appb-000032
1、式3A-5所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-5所示化合物(0.375,2.92mmol)和碳酸钾(1.21g，8.77mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-5所示化合物粗品(0.684g，产率83.8％)无需纯化，可直接用于下一步反应。
2、式4A-5所示化合物的制备
式3A-5所示化合物(0.64g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g)，室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析得式4A-5所示化合物(450mg，产率78.7％)。
3、式I-6所示化合物的制备
将式4A-5所示化合物(50mg，0.200mmol)，式5A-2所示化合物(70mg，0.200mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(35mg，0.200mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(78mg，产率69.4％)，即为式I-6所示化合物。
1H NMR(400MHz,cd3od)δ9.11(d,J＝6.9Hz,1H),8.36–8.31(m,1H),8.09(d,J＝3.6Hz,1H),7.58–7.45(m,2H),6.67(t,J＝2.8Hz,1H),6.57–6.50(m,1H),3.92–3.76(m,6H),3.40–3.31(m,1H),2.90(s,6H),2.79(m,2H),2.18(m,2H),1.87(m,2H),1.29(t,J＝6.6Hz,6H).
LCMS:t＝0.807min,559.2(M),560.2(M+1)
实施例11式I-14所示化合物的制备流程
Figure PCTCN2016104723-appb-000033
将式4A-5所示化合物(50mg，0.200mmol)，式5A-1所示化合物(69mg，0.200mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(35mg，0.200mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(58mg，产率51.7％)，即为式I-14所示化合物。
1H NMR(400MHz,cd3od)δ8.52(d,J＝8.4Hz,1H),8.08(s,1H),7.87(dd,J＝8.0,1.5Hz,1H),7.66(d,J＝8.7Hz,1H),7.62(t,J＝7.2Hz,1H),7.31(t,J＝7.2Hz,1H),6.68(s,1H),6.51(dd,J＝8.7,2.5Hz,1H),3.82-3.79(m,5H),3.27–3.17(m,1H),2.85(s,6H),2.78-2.72(m,3H),2.16-2.11(m,2H),1.84-1.79(m,2H),1.24(d,J＝6.8Hz,6H).
LCMS:t＝0.692min,559.3(M).
实施例12式I-3所示化合物的制备流程
将式4A-5所示化合物(30mg，0.120mmol)，式5A-3所示化合物(42mg，0.120mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(21mg，0.121mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(20mg，产率29.4％)，即为式I-3所示化合物。
1H NMR(400MHz,cd3od)δ8.31(s,1H),8.22(d,J＝10.3Hz,1H),8.04(d,J＝4.8Hz,1H),7.59–7.41(m,1H),7.19(s,1H),7.03(d,J＝7.7Hz,1H),4.00–3.86(m,5H),3.74–3.54(m,1H),3.44(m,3H),2.95(s,6H),2.53–2.29(m,2H),2.18(m,2H),1.31(t,J＝10.4Hz,6H).
LCMS:t＝0.689min,565.3(M),566.3(M+1).
实施例13式I-9所示化合物的制备流程
Figure PCTCN2016104723-appb-000035
1、式3A-6所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-6所示化合物(0.295g,2.92mmol)和碳酸钾(1.21g，8.77mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-6所示化合物粗品(0.6g，产率81.4％)无需纯化，可直接用于下一步反应。
2、式4A-6所示化合物的制备
式3A-6所示化合物(0.58g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g)室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析得式4A-6所示化合物(411mg，产率80.4％)。
3、式I-9所示化合物的制备
将式4A-6所示化合物(30mg，0.135mmol)，式5A-2所示化合物(47mg，0.135mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(24mg，0.135mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(41mg，产率57％)，即为式I-9所示化合物。
1H NMR(400MHz,cd3od)δ8.65(dd,J＝5.4,4.3Hz,2H),8.32(s,1H),7.82–7.68(m,2H),7.47(d,J＝2.4Hz,1H),7.21(dd,J＝8.7,2.3Hz,1H),4.12(m,1H),4.04–3.91(m,4H),3.84(m,2H),3.65(m,2H),2.39–2.23(m,2H),2.12–2.03(m,2H),1.36–1.22(m,6H).
LCMS:t＝0.679min,533.2(M),534.3(M+1).
实施例14式I-16所示化合物的制备流程
Figure PCTCN2016104723-appb-000036
将式4A-6所示化合物(30mg，0.135mmol)，式5A-3所示化合物(47mg，0.135mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(24mg，0.135mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(28mg，产率38.5％)， 即为式I-16所示化合物。
1H NMR(400MHz,cd3od)δ8.10(s,1H),8.05(s,1H),7.80(d,J＝5.5Hz,1H),7.51(d,J＝8.7Hz,1H),6.69(d,J＝2.5Hz,1H),6.59(dd,J＝8.7,2.5Hz,1H),3.82(s,3H),3.80–3.72(m,1H),3.64–3.50(m,2H),3.35(dd,J＝13.6,6.8Hz,1H),2.98–2.83(m,2H),2.05–1.91(m,2H),1.75–1.58(m,2H),1.31(d,J＝6.8Hz,6H).
LCMS:t＝0.734min,538.2(M),539.2(M+1)
实施例15式I-10所示化合物的制备流程
Figure PCTCN2016104723-appb-000037
1、式3A-7所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-7所示化合物(0.53,2.92mmol)和碳酸钾(1.21g，8.77mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-7所示化合物粗品(0.826g，产率84.5％)无需纯化，可直接用于下一步反应。
2、式4A-7所示化合物的制备
式3A-7所示化合物(0.76g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g)室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析得式4A-7所示化合物(566mg，产率81.8％)。
3、式I-10所示化合物的制备
将式4A-7所示化合物(40mg，0.131mmol)，式5A-2所示化合物(45.6mg，0.131mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(23mg，0.132mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(30mg，产率37.11％)，即为式I-10所示化合物。
1H NMR(400MHz,cd3od)δ9.10(d,J＝8.3Hz,1H),8.33(d,J＝4.3Hz,1H),8.09(d,J＝2.3Hz,1H),7.57–7.41(m,2H),6.66(d,J＝2.3Hz,1H),6.54(dd,J＝8.7,2.5Hz,1H),3.92–3.62(m,6H),3.21–2.81(m,7H),2.73(dd,J＝24.5,12.5Hz,3H),2.65(s,3H),2.04(d,J＝11.9Hz,2H),1.71(dt,J＝11.7,8.6Hz,2H),1.29(t,J＝6.8Hz,6H).
LCMS:t＝0.665min,615.3(M),616.2(M+1)
实施例16式I-11所示化合物的制备流程
Figure PCTCN2016104723-appb-000038
将式4A-7所示化合物(40mg，0.131mmol)，式5A-3所示化合物(46mg，0.131mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(23mg，0.132mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(18mg，产率22％)，即为式I-11所示化合物。
实施例17式I-15所示化合物的制备流程
Figure PCTCN2016104723-appb-000039
1、式3A-8所示化合物的制备
将式1A-1所示化合物(0.5g，2.92mmol)，式2A-8所示化合物(0.617,2.92mmol)和碳酸钾(1.21g，8.77mmol)溶入无水DMF(10mL)中，加热升温到80℃，搅拌反应5小时，TLC监测反应完毕，加入乙酸乙酯100毫升萃取，用饱和食盐水洗三次，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，所得式3A-8所示化合物粗品(0.856g，产率80.8％)无需纯化，可直接用于下一步反应。
2、式4A-8所示化合物的制备
式3A-8所示化合物(0.83g，2.28mmol)溶于THF/MeOH(v/v＝1:1,共20ml)和10ml饱和氯化铵水溶液，搅拌10min，分批加入锌粉(1.6g)，室温搅拌1小时。TLC显示反应完全，过滤，滤液减压浓缩，所得粗品经柱层析得式4A-8所示化合物(650mg，产率85.4％)。
3、式I-15所示化合物的制备
将式4A-8所示化合物(40mg，0.120mmol)，式5A-2所示化合物(42mg，0.120mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(21mg，0.120mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(20mg，产率25.8％)，即为式I-15所示化合物。
实施例18式I-18所示化合物的制备流程
Figure PCTCN2016104723-appb-000040
将式4A-8所示化合物(40mg，0.120mmol)，式5A-3所示化合物(42mg，0.120mmol)溶于正丁醇(2mL)中,搅拌下加入对甲苯磺酸(21mg，0.120mmol)，加热升温至100℃搅拌反应5小时，TLC检测显示反应完毕，减压浓缩，所得粗品用柱层析法纯化分离得到类白色固体产物(16mg，产率20.5％)，即为式I-18所示化合物。
实施例19 5-氯-N2-(4-(4-(异丙基哌嗪-1-基)-2-甲氧基苯基)-N4-(2-(异丙基磺酰基)吡啶-3-基)嘧啶-2,4-二胺(式I-23所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000041
将2,5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(183mg,0.527mmol)和4-(4-异丙基哌嗪-1-基)-2-甲氧基苯胺(131mg,0.527mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.527mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品经柱层析分离纯化得黄色固体159mg，产率53.8％，即为式I-23所示化合物。
1H NMR(400MHz,CDCl3):δ10.06(s,1H),9.18(d,J＝8.4Hz,1H),8.36(s,1H),8.14(s,1H),7.88(d,J＝8.0Hz,1H),7.43(s,1H),7.19(s,1H),6.56(s,1H),6.50(d,J＝8.0Hz,1H),3.86(s,4H),3.23(s,4H),2.76(s,5H),1.37(d,J＝6.4Hz,6H),1.14(d,J＝4.4Hz,6H).
LCMS:t＝3.17min,560.1(M+H+).
其中，本实施例中所述4-(4-异丙基哌嗪-1-基)-2-甲氧基苯胺(式1所示化合物)
Figure PCTCN2016104723-appb-000042
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000043
第一步：4-(3-甲氧基-4-硝基苯基)哌嗪-1-羧酸叔丁酯的制备
Figure PCTCN2016104723-appb-000044
将化合物4-氟-2-甲氧基-1硝基苯(3g,17.53mmol)溶于10毫升DMF,在搅拌下加入叔丁基哌嗪-1-羧酸酯(3.59g,19.28mmol)和碳酸铯(17.14g,52.6mmol)，加热至80℃反应过夜。监测反应完后，冷却至室温，加入100mL水稀释并用DCM/i-PrOH(3:1)萃取(50mL×3)，合并的有机相经饱和食盐水洗，无水硫酸钠干燥后，过滤蒸除溶剂后得到黄色油状的产品5.4g，产率91％。
LCMS:t＝3.88min,282.0(M-55).
第二步：1-(3-甲氧基-4-硝基苯基)哌嗪的制备
Figure PCTCN2016104723-appb-000045
将化合物4-(3-甲氧基-4-硝基苯基)哌嗪-1-羧酸叔丁酯(5.4g,16.01mmol)溶于二氯甲烷(15mL)。在室温搅拌下慢慢加入三氟乙酸(5mL)，反应在室温搅拌过夜，旋干溶剂得到黑色油状产品3.8g，产率100％。
第三步：1-异丙基-4-(3-甲氧基-4-硝基苯基)哌嗪的制备
Figure PCTCN2016104723-appb-000046
将化合物1-(3-甲氧基-4-硝基苯基)哌嗪三氟乙酸盐(3.8g,16mmol)溶于15毫升DMF后，在搅拌下依次加入碳酸钾(6.64g,48.0mmol)和异丙基溴(4.53ml,32.0mmol)。加热至80℃搅拌4小时。TLC监测完毕后，冷却至室温，加水稀释并用EtOAc萃取。有机相经水洗，无水硫酸钠干燥后，过滤旋干，经柱层析纯化分离得到绿色油状产品4.2g，产率52％。
LCMS:t＝1.57min,280.1(M+H+).
第四步：4-(4-异丙基哌嗪-1-基)-2-甲氧基苯胺的制备
Figure PCTCN2016104723-appb-000047
将化合物1-异丙基-4-(3-甲氧基-4-硝基苯基)哌嗪(4.2g,11.73mmol)溶在50毫升甲醇中，氮气置换后，加入钯碳(125mg)。氢气置换，在氢气氛围室温搅拌过夜。TLC监测完毕后，过滤除去碳钯，母液减压浓缩得到黑色油状的产品0.4g，产率13％。
LCMS:t＝0.43min,250.1(M+H+).
实施例20：5-氯-N2-(4-(4-(二甲基氨基)哌啶-1-基)-2-异丙氧苯基-N4-(2-(异丙基磺酰)吡啶-3-基)嘧啶-2,4-二胺(式I-91所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000048
将2,5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(150mg,0.43mmol)和1-(4-胺基-3-异丙氧苯基)-N,N-二甲基氨基哌啶-4-胺(式9所示化合物)(120mg,0.43mmol)溶于正丁醇(1.5mL)，并向反应液中加入对甲苯磺酸(74mg,0.43mmol)，升温至115摄氏度，搅拌反应5小时。TLC监测反应完毕，冷却至室温，减压蒸除溶剂后，将粗品溶于二氯己烷，经饱和碳酸氢钠水溶液及饱和食盐水洗涤后，无水硫酸钠干燥，过滤浓缩得粗品，经柱层析分离纯化得到浅绿色的固体产物100mg，产率39.4％，即为式I-91所示化合物。
1H NMR(400MHz,CDCl3)：δ10.04(s,1H),9.17(d,J＝8.8Hz,1H),8.38(d,J＝4.4Hz,1H),8.15(s,1H),7.96(d,J＝8.8Hz,1H),7.45(dd,J＝8.8,4.4Hz,1H),7.31(s,1H),6.53(s,1H),6.46(d,J＝9.2Hz,1H),4.75-4.44(m,2H),3.69(d,J＝12.0Hz,3H),3.15(s,2H),2.78(s,6H),2.26(d,J＝9.6Hz,2H),2.00-1.86(m,2H),1.37(t,J＝5.6Hz,12H).
LCMS：t＝3.39min，588.2(M+H+),294.7(M/2+H+).
其中，本实施例中所述1-(4-氨基-3-异丙氧基苯基)-N,N-二甲基哌啶-4-胺(式9所示化合物)
Figure PCTCN2016104723-appb-000049
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000050
第一步：2-异丙氧基-4-氟-1-硝基苯的制备
Figure PCTCN2016104723-appb-000051
将化合物2,4-二氟-1-硝基苯(5.0g,31.43mmol)溶于异丙醇(60mL)并加入碳酸铯(30.64g,94.29mmol)，加热至80℃搅拌过夜。TLC监测反应完成后，加水稀释并用DCM萃取，合并的有机相用无水硫酸钠干燥，过滤旋干，经层析柱分离纯化得到白色固体产品5.2g，产率83％.
第二步：1-(3-异丙氧基-4-硝基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000052
将化合物4-氟-2-异丙氧基-1-硝基苯(5g,25.1mmol)溶于DMF(60mL)，加入N,N-二甲基派啶-4-胺盐酸盐(3.54g,27.61mmol)和碳酸钾(10.41g,75.31mmol)，加热至82℃搅拌过夜。TLC监测反应完成后，反应冷却至室温，用水稀释，乙酸乙酯萃取。有机层用饱和食盐水洗，无水硫酸钠干燥，过滤浓缩得到白色的产品6.2g，产率80％。
LCMS:t＝2.476min,308.1(M+H+).
第三步：1-(4-氨基-3-异丙氧基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000053
将化合物1-(3-异丙氧基-4-硝基苯基)-N,N-二甲基哌啶-4-胺(1.7g,5.53mmol)溶于15毫升甲醇并加入钯碳(580mg)。氢气置换，氢气氛围下室温搅拌过夜。TLC监测反应完成后，过滤，蒸除甲醇得到黑色油状产品1.45g，产率90％。
LCMS:t＝0.43min,278.1(M+H+).
实施例21：5-氯-N2-(4-(4-(二甲基氨基)哌啶-1-基)-2-乙氧苯基-N4-(2-(异丙基磺酰)吡啶-3-基)嘧啶-2,4-二胺(式I-92所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000054
将2,5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(150mg,0.43mmol)和1-(4-氨基-3-乙氧基苯基)-N,N-二甲基哌啶-4-胺(式8所示化合物)(114mg,0.43mmol)溶于正丁醇(1.5mL)，并向反应液中加入对甲苯磺酸(74mg,0.43mmol)，升温至115摄氏度，搅拌反应5小时。TLC监测反应完毕，冷却至室温，减压蒸除溶剂后，将粗品溶于二氯己烷，经饱和碳酸氢钠水溶液及饱和食盐水洗涤后，无水硫酸钠干燥，过滤浓缩得粗品，粗品经柱层析分离纯化得到浅绿色的固体90mg，产率36.3％，即为式I-92所示化合物。
1H NMR(400MHz,cd3od)δ9.08(d,J＝8.4Hz,1H),8.34(d,J＝4.1Hz,1H),8.10(s,1H),7.69(d,J＝8.1Hz,1H),7.49(dd,J＝8.6,4.3Hz,1H),7.22(d,J＝8.0Hz,1H),6.66(d,J＝2.2Hz,1H),4.05(dd,J＝13.9,6.9Hz,2H),3.81(dd,J＝13.5,6.8Hz,2H),3.22(m,1H),2.84(s,6H),2.78(m,2H),2.17(m,2H),1.83(m,2H),1.38–1.27(m,6H).
LCMS：t＝3.37min，574.1(M+H+).
其中，本实施例中所述1-(4-氨基-3-乙氧基苯基)-N,N-二甲基哌啶-4-胺(式8所示化合物)
Figure PCTCN2016104723-appb-000055
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000056
第一步：2-乙氧基-4-氟-1-硝基苯的制备
Figure PCTCN2016104723-appb-000057
将化合物2,4-二氟-1-硝基苯(5.0g,31.44mmol)溶于乙醇(70mL)，加入碳酸铯(30.66g,94.34mmol)，加热搅拌过夜。TLC监测反应完成后，溶剂旋干，加水稀释，用乙酸乙酯萃取，合并的有机相用无水硫酸钠干燥，过滤旋干，经层析柱分离纯化得到白色固体产品5.5g，产率95％。
第二步：1-(3-乙氧基-4-硝基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000058
将化合物4-氟-2-乙氧基-1-硝基苯(5g,27mmol)溶于DMF(60mL)，加入N,N-二甲基派啶-4-胺盐酸盐(3.81g,29.7mmol)和碳酸钾(11.2g,781.01mmol)，加热82℃搅拌过夜。TLC监测反应结束后，加水稀释，用乙酸乙酯萃取两次。合并的有机层用饱和食盐水洗涤，无水硫酸钠干燥后，过滤蒸除溶剂得到白色的产品6.2g，产率78％。
LCMS:t＝2.602min,294.1(M+H+).
第三步：1-(4-氨基-3-乙氧基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000059
将化合物1-(3-乙氧基-4-硝基苯)-N,N-二甲基哌啶-4-胺(2.22g,7.57mmol)溶于15毫升甲醇，加入钯碳(300mg)。氢气置换三次，氢气氛围室温搅拌过夜。TLC监测反应完成后，过滤除去碳钯，母液减压浓缩得到黑色油状产品1.54g，产率75％。
LCMS:t＝0.430min,264.1(M+H+).
实施例22：5-氯-N2-(2-异丙氧基-5-甲基-4-(1，2，3，6-四氢哌啶-4-基)苯基)-N4-(2-异丙磺酰)吡啶-3-基)嘧啶-2,4-二胺(式I-90所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000060
第一步：4-(4-((5-氯-4-((2-(异丙基磺酰)吡啶-3-基)氨基)嘧啶-2-基)氨基)-5-异丙氧基-2-甲基苯)-3,6-二氢哌啶-1(2H)-羰酸叔丁酯的制备
将2，5-二氯-N-(2-异丙磺酰)吡啶-3-基)嘧啶-4-胺(0.37g,1.07mmol)，4-(4-氨基-5-异丙氧基-2-甲基苯基)-3，6-二氢哌啶-1(2H)-羧酸叔丁酯(0.41g,1.17mmol)，碳酸铯(1.04g,3.20mmol)，Pd2(dba)3(49.0mg,0.053mmol)，Xantphos(62.0mg,0.107mmol)置于10毫升的微波管里，加入3毫升的1,4-二氧六环，氮气置换三次，在130摄氏度微波反应40分钟。TLC监测反应完成后，冷却至室温，反应液用水稀释，乙酸乙酯萃取两次，有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤浓缩，得到粗品经柱层析分离纯化(PE：EtOAc＝3:1)得到白色的固体产物260mg，产率37％。
LCMS:t＝7.97min,657.2(M+H+).
第二步：5-氯-N2-(2-异丙氧基-5-甲基-4-(1，2，3，6-四氢哌啶-4-基)苯基)-N4-(2-(异丙氧磺酰)吡啶-3-基)嘧啶-2,4-二胺的制备
将4-(4-((5-氯-4-((2-异丙磺酰)吡啶-3-基)氨基)嘧啶-2基)氨基)-5-异丙氧基-2甲基苯基)-3，6-二氢哌啶-1(2H)-羧酸叔丁酯(260mg,0.396mmol)加入盐酸乙酸乙酯(4M)，室温过夜。LCMS监测反应完成后，减压浓缩，得到粗品用乙酸乙酯打浆得到浅黄色的固体产物100mg，产率43％，即为式I-90所示化合物。
1H NMR(400MHz,CDCl3)：δ10.77(s,2H),10.09(s,1H),9.56(s,1H),8.98(s,1H),8.48(s,1H),8.15(s,1H),7.58(s,1H),7.41(s,1H),6.73(s,1H),5.65(s,1H),4.58(s,1H),4.11-3.78(m,3H),3.50(s,2H),2.75(s,2H),2.19(s,3H),1.52-1.25(m,12H).
LCMS：t＝3.44min，557.1(M+H+).
其中，本实施例中所述4-(4-氨基-5-异丙氧基-2-甲基苯基)-3，6-二氢哌啶-1(2H)-羧酸叔丁酯(式6所示化合物)
Figure PCTCN2016104723-appb-000061
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000062
第一步：4-(5-异丙氧基-2-甲基-4-硝基苯)-3,6-二氢哌啶-1(2H)-羧酸叔丁酯的制备
Figure PCTCN2016104723-appb-000063
将化合物1-氯-5-异丙氧基-2-甲基-4-硝基苯(1g,4.35mmol)，N-Boc-1,2,5,6-四氢吡啶-4-硼酸频哪醇酯(1.481g,4.79mmol)，碳酸钾(1.805g,13.06mmol)，PdCl2(dppf).CH2Cl2(356mg,0.43mmol)加入50毫升的单口瓶里，加入10毫升的1,4-二氧六环。氮气置换，在氮气氛围，加热至120℃搅拌过夜。冷却至室温，加水稀释，用乙酸乙酯萃取。有机相用饱和食盐水洗涤，无水硫酸钠干燥后，过滤，蒸除溶剂得到油状的产品0.81g，产率49％。
LCMS:t＝4.83min,321.1(M-55).
第二步：4-(4-氨基-5-异丙氧基-2-甲基)-3,6-二氢哌啶-1(2H)-羧酸叔丁酯的制备
Figure PCTCN2016104723-appb-000064
将化合物4-(5-异丙氧基-2-甲基-4-硝基苯基)-3,6-二氢哌啶-1(2氢)-羧酸叔丁酯(0.81g,2.152mmol)溶于15毫升甲醇中，并加入氯化铵(1.151g,21.52mmol)。在室温下分批加入锌粉(1.40，21.538mmol)，加热至回流，搅拌过夜。冷却至室温，过滤，旋干。粗品用二氯甲烷溶解，用饱和的碳酸氢钠洗，无水硫酸钠干燥，过滤浓缩得到固体产品0.26g，产率97％。
LCMS:t＝3.97min,347.1(M+H+).
实施例23：5-氯-N4-(2-异丙磺酰)吡啶-3-基)-N2-(2-甲氧基-5-甲基-4-(1,2,3,6-四氢哌啶-4-基)苯基)嘧啶-2,4-二胺(式I-89所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000065
第一步：4-(4-((5-氯-4-((2-(异丙基磺酰)吡啶-3-基)氨基)嘧啶-2-基)氨基)-5-甲氧基-2-甲基苯基)-3,6-二氢哌啶-1(2H)-羰酸叔丁酯的制备
将化合物4-(4-氨基-5-甲氧基-2-甲基苯基)-3,6-二氢哌啶-1(2H)-羧酸叔丁酯(0.726g,2.281mmol)，2,5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(0.72g,2.074mmol)，碳酸铯(2.027g,6.22mmol)，Pd2(dba)3(0.095g,0.104mmol)，Xantphos(0.120g,0.207mmol)置于10毫升的微波管里，加入10毫升1,4-二氧六环。氮气置换，微波加热130℃反应40分钟。冷却至室温后，反应加入EtOAc稀释，饱和食盐水洗涤，无水硫酸钠干燥，过滤浓缩，所得到的粗品经柱层析分离纯化得到白色固体产200mg，产率14％。
第二步：5-氯-N4-(2-(异丙基磺酰)吡啶-3-基)-N2-(2-甲氧基-5-甲基-4-(1,2,3,6-四氢哌啶-4-基)苯基)嘧啶-2,4-二胺的制备
将化合物4-(4-((5-氯-4-((2-(异丙基磺酰)吡啶-3-基)氨基)嘧啶-2-基)氨基)-5-甲氧基-2-甲基苯基)-3,6-二氢哌啶-1(2H)-羰酸叔丁酯(0.2g,0.318mmol)溶于1毫升EtOAc，在室温下慢慢加入4毫升的HCl/EtOAc(4M)，搅拌反应过夜。反应溶剂浓缩，用乙酸乙酯打浆，再经过HPLC制备分离得到浅黄色固体产品100mg，产率51.6％，即为(式I-89所示化合物。
1H NMR(400MHz,cd3od)δ9.11(d,J＝8.5Hz,1H),8.38(d,J＝4.3Hz,1H),8.16(s,1H),7.75(s,1H),7.58(dd,J＝8.7,4.4Hz,1H),6.75(s,1H),5.65(s,1H),3.90–3.77(m,6H),3.46(t,J＝6.0Hz,2H),2.63(d,J＝1.8Hz,2H),2.18(s,3H),1.31(d,J＝6.9Hz,6H).
LCMS：t＝3.29min，531.1(M+H+),265.2(M/2+H+).
其中，本实施例中所述4-(4-氨基-5-甲氧基-2-甲基苯基)-3,6-二氢哌啶-1(2H)-羧酸叔丁酯(式7所示化合物)
Figure PCTCN2016104723-appb-000066
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000067
第一步：1-氯-5-甲氧基-2-甲基-4-硝基苯的制备
Figure PCTCN2016104723-appb-000068
将化合物1,5-二氯-2-甲基硝基苯(2g,9.71mmol)溶于甲醇(20mL)并加入碳酸铯(9.49g,29.12mmol)。反应在室温反应3小时。TLC监测反应结束后，加水(80mL)稀释，DCM(100mL×2)萃取，合并的有机相用饱和食盐(50mL)水洗，无水硫酸钠干燥，过滤蒸除溶剂得到白色固体产品1.76g，产率90％。
第二步：4-(5-甲氧基-2-甲基-4-硝基苯基)-3,6-二氢哌啶-1(2H)-羧酸叔丁酯的制备
Figure PCTCN2016104723-appb-000069
将化合物1-氯-5-甲氧基-2-甲基-4-硝基苯(3.5g,17.36mmol),N-Boc-1,2,5,6-四氢吡啶-4-硼酸频哪醇酯(5.90g,19.10mmol)，碳酸钾(7.20g,52.1mmol)，Pd(Ph3P)4(1.003g,0.868mmol)加入100毫升的单口瓶里，加入20毫升1，4-二氧六环。氮气置换，在氮气氛围加热120℃搅拌过夜。TLC监测反应完成后，溶剂旋干，加水稀释，用EtOAc萃取。有机相用饱和食盐水洗，无水硫酸钠干燥，过滤旋干，经柱层析分离得到白色固体产品3g，产率49％。
LCMS:t＝4.507min,293.0(M-55).
第三步：4-(4-氨基-5-甲氧基-2-甲基苯基)-3,6-二氢哌啶-1(2H)-羧酸叔丁酯的制备
Figure PCTCN2016104723-appb-000070
将化合物4-(5-甲氧基-2-甲基-4-硝基苯基)-3,6-二氢哌啶-1(2H)-羰酸叔丁酯(0.8g,2.296mmol)溶于 10毫升甲醇，在室温下加入氯化铵(0.614g,11.48mmol)并分批加入锌粉(0.751g,11.48mmol)，加热回流2小时。TLC监测反应结束，冷却到室温，旋干溶剂，所得到的粗品用EtOAc溶解，用碳酸氢钠溶液洗，无水硫酸钠干燥，过滤旋干得到白色固体产品0.72g，产率98.6％。
LCMS:t＝3.489min,319.1(M+H+).
实施例24：5-氯-N2-(2-(二氟甲氧基)-4-(4-(二甲基氨基)哌啶-1-基)苯基)-N4-(2-(异丙基磺酰)吡啶-3-基)嘧啶-2,4-二胺(式I-100所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000071
将2，5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(150mg,0.432mmol)和1-(4-氨基-3-(二氟甲氧基)苯基)-N,N-二甲基哌啶-4-胺(式3所示化合物)(123mg,0.432mmol)溶于正丁醇(1.5mL)，并向反应液中加入对甲苯磺酸(74mg,0.432mmol)，升温至115摄氏度，搅拌反应5小时。TLC监测反应完毕，冷却至室温，减压蒸除溶剂后，将粗品溶于二氯己烷，经饱和碳酸氢钠水溶液及饱和食盐水洗涤后，无水硫酸钠干燥，过滤浓缩得粗品，粗品经柱层析分离纯化得棕红色的固体110mg，产率43％，即为式I-100所示化合物。
1H NMR(400MHz,CDCl3)：δ10.11(s,1H),9.11(d,J＝8.8Hz,1H),8.37(dd,J＝4.4,1.3Hz,1H),8.15(s,1H),7.86(d,J＝8.8Hz,1H),7.41(dd,J＝8.8,4.4Hz,1H),6.95(s,1H),6.78(dd,J＝8.8,2.6Hz,1H),6.75(s,1H),6.50(t,J＝73.6Hz,1H),3.90(dq,J＝13.6,7.2Hz,1H),3.67(t,J＝12.4Hz,2H),2.75(dd,J＝1.8,12.2Hz,2H),2.42(s,6H),2.03(d,J＝12.8Hz,2H),1.76-1.66(m,2H),1.38(d,J＝6.8Hz,6H).
LCMS:t＝3.50min,596.2(M+H+).
其中，本实施例中所述1-(4-氨基-3-(二氟甲氧基)苯基)-N,N-二甲基哌啶-4-胺(式3所示化合物)
Figure PCTCN2016104723-appb-000072
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000073
第一步：4-氯-2-(二氟甲氧基)-1-硝基苯的制备
Figure PCTCN2016104723-appb-000074
将化合物2-氟-2-硝基苯酚(4g,25.5mmol)溶于DMF(50ml)/水(10ml)，加入2-氯-2，2-二氟乙酸钠(11.65g,46mmol)和碳酸钾(10.56g,76mmol)。氮气置换，在氮气氛围，加热至100℃搅拌反应5小时。TLC监测反应完成后，反应冷却至室温，加水稀释并用乙酸乙酯萃取。有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤旋干，经层析柱(石油醚)得到黄色的油状产品2.5g，产率47％。
第二步：1-(3-(二氟甲氧基)-4-硝基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000075
将化合物2-(二氟甲氧基)-4-氯-1-硝基苯(2.5g,12.07mmol)溶于DMF(20ml)，搅拌下加入N,N-二甲基哌啶-4-胺盐酸盐(2.428g,12.07mmol)和碳酸钾(5.00g,36.2mmol)。混合物加热80℃搅拌反应过夜。TLC监测反应完成后，反应冷却至室温，加水稀释并用乙酸乙酯萃取。有机相用饱和食盐水洗，无水硫酸钠干燥后，过滤浓缩，经层析柱(DCM:MeOH＝30:1)得到黄色油状的产品1.4g，产率34％。
LCMS:t＝2.68min,316.0(M+H+).
第三步：1-(4-氨基-3-(二氟甲氧基)苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000076
将化合物1-(3-(二氟甲氧基)-4-硝基苯基)-N,N-二甲基哌啶-4-胺(0.7g,2.220mmol)溶于甲醇(10ml)，搅拌下加入氯化铵(1.188g,22.20mmol)，分批加入锌粉(1.451g,22.20mmol)后，加热回流3小时。TLC监测反应完成后，冷却至室温，过滤浓缩，所得到的粗品加DCM溶解，用碳酸氢钠溶液洗，无水硫酸钠干燥，过滤浓缩得到棕色的油状的产品0.5g，产率77％，即为本实施例所述中间体3。
LCMS:t＝0.48min,286.1(M+H+).
实施例25：5-氯-N2-(4-(1-异丙基-1,2,3,6-四氢吡啶-4-基)-2-甲氧基-5-甲基苯基)-N4-(2-(异丙基磺酰基)吡啶-3-基)嘧啶-2,4-二胺(式I-80所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000077
将2,5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(183mg,0.527mmol)和4-(1-异丙基-1,2,3,6-四氢吡啶-4-基)-2-甲氧基-5-甲基苯胺(137mg,0.527mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.527mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得黄色固体186mg，产率61.8％，即为式I-80所示化合物。
1H NMR(400MHz,CDCl3):δ10.08(s,1H),9.18(d,J＝8.3Hz,1H),8.37(d,J＝3.4Hz,1H),8.18(s,1H),7.96(s,1H),7.53–7.42(m,2H),6.76(s,1H),5.59(s,1H),3.99–3.86(m,2H),3.83(s,3H),3.74-3.69(m3H),3.34(s,3H),2.17(s,3H),1.47(d,J＝6.6Hz,6H),1.38(d,J＝6.9Hz,6H).
LCMS:t＝3.42min,571.1(M+H+).
实施例26:5-氯-N2-(4-(4-(二甲基氨基)哌啶-1-基)-2-异丙氧基苯基)-N4-(2-(异丙基磺酰基)噻吩-3-基)嘧啶-2,4-二胺(式I-82所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000078
将2,5-二氯-N-(2-(异丙基磺酰)噻吩-3-基)嘧啶-4-胺(186mg,0.528mmol)和中间体9(147mg,0.530mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.528mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得黄色固体220mg，产率70.2％.
1H NMR(400MHz,CDCl3):δ9.40(s,1H),8.27(d,J＝5.2Hz,1H),8.11(s,1H),7.99(d,J＝8.4Hz,1H),7.78(d,J＝8.0Hz,1H),7.64(d,J＝5.6Hz,1H),7.18(d,J＝7.6Hz,1H),6.55(s,1H),6.52(d,J＝9.2Hz,1H),4.56(dt,J＝11.6,6.0Hz,1H),3.66(d,J＝12.0Hz,2H),3.32(dt,J＝13.6,6.8Hz,1H),2.73(t,J＝11.2Hz,3H),2.58(s,6H),2.17–2.01(m,2H),1.90–1.73(m,2H),1.37(t,J＝6.8Hz,12H).
LCMS:t＝3.38min,593.1(M+H+).
实施例27:5-氯-N4-(2-(异丙基磺酰基)吡啶-3-基)-N2-(2-甲氧基-4-(1-甲基-1,2,3,6-四氢吡啶-4-基)苯基) 嘧啶-2,4-二胺(式I-84所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000079
将2,5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(183mg,0.527mmol)和2-甲氧基-4-(1-甲基-1,2,3,6-四氢哌啶-4-基)苯胺(式4所示化合物)(115mg,0.527mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.527mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得黄色固体115mg，产率41.2％，即为式I-84所示化合物。
1H NMR(400MHz,CDCl3):δ10.06(s,1H),9.17(d,J＝8.8Hz,1H),9.11(dd,J＝18.8,6.0Hz,1H),8.52-8.45(m,1H),8.39(d,J＝4.4Hz,1H),8.28–8.19(m,1H),8.08(d,J＝8.8Hz,1H),7.48(d,J＝3.6Hz,1H),6.97–6.90(m,1H),6.03(s,1H),3.91(s,3H),3.13(d,J＝2.8Hz,2H),2.69(t,J＝5.6Hz,3H),2.60(s,2H),2.42(s,3H),1.39(t,J＝6.2Hz,6H).
LCMS:t＝3.25min,529.1(M+H+).
其中，本实施例中所述2-甲氧基-4-(1-甲基-1,2,3,6-四氢哌啶-4-基)苯胺(式4所示化合物)
Figure PCTCN2016104723-appb-000080
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000081
第一步：4-(3-甲氧基-4-硝基苯基)-3,6-二氢哌啶-1(2H)-羧酸叔丁酯的制备
Figure PCTCN2016104723-appb-000082
将化合物4-溴-2-甲氧基-1-硝基苯(1.5g,6.46mmol)，N-Boc-1,2,5,6-四氢吡啶-4-硼酸频哪醇酯(2.199g,7.11mmol)，碳酸钾(2.68g,19.39mmol)，Pd(Ph3P)4(0.374g,0.323mmol)置于100毫升的单口烧瓶里，往里加入1，4-二氧六环。氮气置换，在氮气氛围加热至90℃搅拌过夜。TLC监测反应完成后，旋干溶剂，加水稀释并用二氯甲烷萃取。有机相用无水硫酸钠，过滤用无水硫酸钠干燥，经柱层析分离纯化得到白色固体产品1.72g，产率65％。
LCMS:t＝4.30min,279.0(M-55).
第二步：4-(3-甲氧基-4-硝基苯基)-1,2,3,6-四氢哌啶三氟乙酸盐的制备
Figure PCTCN2016104723-appb-000083
将化合物4-(3-甲氧在-4-硝基苯基)-5,6-二氢哌啶-1(2H)-羧酸叔丁酯(1.72g,5.14mmol)溶在10毫升二氯甲烷里。在室温下滴入TFA(4mL)，搅拌过夜，反应完毕后旋去三氟乙酸得到油状的产品1.79g，产率100％。
第三步：4-(3-甲氧基-4-硝基苯基)-1-甲基-1,2,3,6-四氢哌啶的制备
Figure PCTCN2016104723-appb-000084
将化合物4-(3-甲氧基-4-硝基苯基)-1,2,3,6-四氢哌啶三氟乙酸盐(0.6g,1.723mmol)溶于10毫升的甲醇中，加入多聚甲醛(0.155g,5.17mmol)，在室温搅拌半个小时。分批加入氰基硼氢化钠(0.325g,5.17mmol)，室温下搅拌反应过夜。TLC监测反应结束后，加入50mL水并用EtOAc萃取(30mL×2)，合并有机相后用无水硫酸钠干燥，过滤，蒸除溶剂，所得粗品经柱层析得油状产物0.2g，产率47％。
LCMS:t＝1.57min,249.0(M+H+).
第四步：2-甲氧基-4-(1-甲基-1,2,3,6-四氢哌啶-4-基)苯胺的制备
Figure PCTCN2016104723-appb-000085
将化合物4-(3-甲氧基-4-硝基苯基)-1-甲基-1,2,3,6-四氢哌啶(0.2g,0.806mmol)溶在15毫升甲醇里，并加入氯化铵(0.215g,4.03mmol)。在室温搅拌下分批加入锌粉(0.263g,4.03mmol)，加热回流过夜。TLC监测反应完成后，减压过滤，旋干溶剂。所得的粗品用DCM溶解，并用碳酸氢钠水溶液洗涤，无水硫酸钠干燥，过滤浓缩得到固状产品157mg，产率90％。
LCMS:t＝0.45min,219.0(M+H+).
实施例28:5-氯-N2-(4-(1-异丙基-1,2,3,6-四氢吡啶-4-基)-2-甲氧基苯基)-N4-(2-(异丙基磺酰基)吡啶-3- 基)嘧啶-2,4-二胺(式I-85所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000086
将2,5-二氯-N-(2-(异丙基磺酰)吡啶-3-基)嘧啶-4-胺(183mg,0.527mmol)和4-(1-异丙基-1,2,3,6-四氢哌啶-4-基)-2-甲氧基苯胺(式11所示化合物)(130mg,0.527mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.527mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品经柱层析分离纯化得黄色固体52mg，产率17.7％，即为式I-85所示化合物。
1H NMR(400MHz,CDCl3):δ10.06(s,1H),9.12(s,1H),8.41(s,1H),8.16(d,J＝21.2Hz,1H),7.54(s,1H),6.91(s,1H),5.97(s,1H),5.29-5.16(m,1H),3.91(s,3H),3.72-3.58(m,3H),3.25-3.02(m,5H),1.36(t,J＝39.6Hz,12H).
LCMS:t＝3.43min,557.2(M+H+).
其中，本实施例中所述4-(1-异丙基-1,2,3,6-四氢哌啶-4-基)-2-甲氧基苯胺(式11所示化合物)
Figure PCTCN2016104723-appb-000087
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000088
第一步：1-异丙基-4-(3-甲氧基-4-硝基苯基)-1,2,3,6-四氢哌啶的制备
Figure PCTCN2016104723-appb-000089
将化合物4-(3-甲氧基-4-硝基苯基)-1,2,3,6-四氢哌啶三氟乙酸盐(1.2g,3.45mmol)溶于15毫升DMF中，依次加入碳酸钾(1.905g,13.78mmol)和异丙基溴(0.642ml,6.89mmol)。加热至80℃搅拌过夜。TLC监测反应完毕后，加水稀释并用EtOAc萃取。有机相用饱和食盐水水洗，无水硫酸钠干燥，过滤旋干后，经柱层析分离纯化得到油状产品0.3g，产率27％。
LCMS:t＝2.40min,277.0(M+H+).
第二步：4-(1-异丙基-1,2,3,6-四氢哌啶-4-基)-2-甲氧基苯胺的制备
Figure PCTCN2016104723-appb-000090
将化合物1-异丙基-4-(3-甲氧基-4-硝基苯基)-1,2,3,6-四氢哌啶(0.3g,1.086mmol)溶在甲醇(15ml)，加入氯化铵(0.410g,7.67mmol)，在室温下分批加入锌粉(15.21g,233mmol)。加热回流3小时。TLC监测反应完成。反应冷却至室温，过滤，旋干。粗品用DCM溶解，碳酸氢钠溶水洗，无水硫酸钠干燥，过滤浓缩得到黄色油状的产品0.26g，产率97％。
LCMS:t＝0.55min,247.1(M+H+).
实施例29:5-氯-N2-(4-(1-异丙基-1,2,3,6-四氢吡啶-4-基)-2-甲氧基苯基)-N4-(2-(异丙基磺酰基)噻吩-3-基)嘧啶-2,4-二胺(式I-86所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000091
将2,5-二氯-N-(2-(异丙基磺酰基)噻吩-3-基)嘧啶-4-胺(185mg,0.525mmol)和式11所示化合物(130mg,0.528mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.528mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品经柱层析分离纯化得黄色固体65mg，产率22％，即为式I-86所示化合物。
1H NMR(400MHz,CDCl3):δ9.42(s,1H),8.15(s,1H),7.73(s,1H),7.57(s,1H),7.07–6.83(m,1H),5.97(s,1H),3.90(s,3H),3.76(s,2H),3.58(s,1H),3.33-3.30(m,3H),2.04(s,1H),1.70(s,1H),1.58–1.27(m,6H).
LCMS:t＝3.63min,562.1(M+H+).
实施例30:5-氯-N2-(4-(4-(异丙基哌嗪-1-基)-2-甲氧基-5-甲基苯基)-N4-(2-(异丙基磺酰基)吡啶-3-基)嘧啶-2,4-二胺(式I-88所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000092
将2,5-二氯-N-(2-(异丙基磺酰基)吡啶-3-基)嘧啶-4-胺(183mg,0.527mmol)和4-(4-异丙基哌嗪-1-基)-2-甲氧基-5-甲基苯胺(式10所示化合物)(139mg,0.527mmol)溶于n-BuOH(2.5ml)，并向其中 加入对甲苯磺酸(91mg,0.527mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品经柱层析分离纯化得黄色固体171mg，产率56.5％，即为式I-88所示化合物。
1H NMR(400MHz,CDCl3):δ10.07(s,1H),9.20(d,J＝8.4Hz,1H),8.36(d,J＝4.0Hz,1H),8.16(s,1H),7.88(s,1H),7.44(dd,J＝8.4,4.4Hz,1H),7.30(s,1H),6.70(s,1H),3.96–3.87(m,1H),3.85(s,3H),3.04(s,4H),2.82(s,5H),2.20(s,3H),1.38(d,J＝6.8Hz,6H),1.20(s,6H).
LCMS:t＝3.39min,574.1(M+H+).
其中，本实施例中所述4-(4-异丙基哌嗪-1-基)-2-甲氧基-5-甲基苯胺(式10所示化合物)
Figure PCTCN2016104723-appb-000093
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000094
第一步：4-(5-甲氧基-2-甲基-4-硝基苯基)哌嗪-1-羧酸叔丁酯的制备
Figure PCTCN2016104723-appb-000095
将化合物1-氯-5-甲氧基-2-甲基-4-硝基苯(2g,9.92mmol)，N-Boc哌嗪(2.032g,10.91mmol)，碳酸铯(9.70g,29.8mmol)，Pd2(dba)3(0.454g,0.496mmol)，Xantphos(0.574g,0.992mmol)一起加入100毫升的单口瓶里，加入20毫升的1,4-二氧六环。氮气置换，在氮气氛围加热至100℃搅拌过夜。TLC监测反应完成后，加入100mL水并用EtOAc(50mL×2)萃取，合并的有机相经饱和食盐水洗，无水硫酸钠干燥，过滤，蒸除溶剂，经柱层析分离纯化得到黄色固体产品2g，产率57％。
LCMS:t＝4.361min,296.0(M-55).
第二步：1-(5-甲氧基-2-甲基-4-硝基苯基)哌嗪的制备
Figure PCTCN2016104723-appb-000096
将化合物4-(5-甲氧基-2-甲基-4-硝基苯基)哌嗪-1-羧酸叔丁酯(2g,5.69mmol)溶于10毫升的二氯甲烷。室温下滴加TFA(4mL)，搅拌4个小时。TLC监测反应完成后，旋干得到黑色油状产品1.9g，产率100％。
第三步：1-异丙基-4-(5-甲氧基-2-甲基-4-硝基苯基)哌嗪的的制备
Figure PCTCN2016104723-appb-000097
将化合物1-(5-甲氧基-2-甲基-4-硝基苯基)哌嗪(1.9g,5.69mmol)溶于20毫升DMF，依次加入碳酸钾(2.36g,17.05mmol)和异丙基溴(1.4g,11.37mmol)。加热至80℃搅拌3小时。TLC监测反应完毕后，加水稀释并用EtOAc萃取。有机相经饱和食盐水洗，无水硫酸钠干燥，过滤旋干后，经柱层析分离得到黄色油状产品0.68g，产率41％。
LCMS:t＝2.676min,294.1(M+H+).
第四步：4-(4-异丙基哌嗪-1-基)-2-甲氧基-5-甲基苯胺的制备
Figure PCTCN2016104723-appb-000098
将化合物1-异丙基-(5-甲氧基-2-甲基-4-硝基苯基)哌嗪(0.68g,2.32mmol)溶在10毫升甲醇中，加入钯碳(100mg)。氢气置换，在氢气氛围室温搅拌过夜。TLC监测反应完成后，过滤除去碳钯，母液减压浓缩得到产品0.4g，产率65％。
LCMS:t＝4.36min,296.1(M-55).
实施例31:5-氯-N2-(2-二氟甲氧基)-4-(4-(二甲基氨基)哌啶-1-基)苯基)-N4-(2-(异丙基磺酰基)苯基)嘧啶-2,4-二胺(式I-95所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000099
将2,5-二氯-N-(2-(异丙基磺酰基)苯基)嘧啶-4-胺(180mg,0.520mmol)和式3所示化合物(150mg,0.526mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(90mg,0.523mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和 食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品经柱层析分离纯化得黄色固体115mg，产率37.2％，即为式I-95所示化合物。
1H NMR(400MHz,CDCl3):δ9.60(s,1H),8.53(d,J＝8.4Hz,1H),8.12(s,1H),8.00(d,J＝8.8Hz,1H),7.90(dd,J＝8.0,1.2Hz,1H),7.58(t,J＝7.2Hz,1H),7.28–7.20(m,1H),7.00(s,1H),6.73(s,1H),6.48(t,J＝73.6Hz,2H),3.65(d,J＝12.8Hz,2H),3.25-3.19(m,1H),2.99–2.85(m,1H),2.72(dd,J＝24.4,13.2Hz,2H),2.50(s,6H),2.10-2.03(m,2H),1.80-1.72(m,2H),1.29(t,J＝6.8Hz,6H).
LCMS:t＝3.75min,595.14(M+H+).
实施例32:5-氯-N2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基-5-甲基苯基)-N4-(2-(异丙基磺酰基)吡啶-3-基)嘧啶-2,4-二胺(式I-98所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000100
将2,5-二氯-N-(2-(异丙基磺酰基)吡啶-3-基)嘧啶-4-胺(183mg,0.527mmol)和1-(4-氨基-5-甲氧基-2-甲基苯基)-N,N-二甲基哌啶-4-胺(式2所示化合物)(139mg,0.527mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.527mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品经柱层析分离纯化得白色固体50mg，产率16.5％，即为式I-98所示化合物。
1H NMR(400MHz,CDCl3):δ10.08(s,1H),9.19(dd,J＝8.4,1.2Hz,1H),8.37(dd,J＝4.4,1.2Hz,1H),8.16(s,1H),7.91(s,1H),7.55–7.43(m,1H),7.32(s,1H),6.60(s,1H),3.95-3.91(m,1H),3.86(s,3H),3.23(t,J＝12.2Hz,2H),3.16(t,J＝12.0Hz,1H),2.79(s,6H),2.74(t,J＝11.2Hz,2H),2.28(t,J＝12.0Hz,2H),2.18(s,3H),1.97-1.94(m,2H),1.39(d,J＝6.8Hz,6H).
LCMS:t＝3.52min,574.3(M+H+).
其中，本实施例中所述1-(4-氨基-5-甲氧基-2-甲基苯基)-N，N-二甲基哌啶-4-胺(式2所示化合物)
Figure PCTCN2016104723-appb-000101
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000102
第一步：1-(5-甲氧基-2-甲基-4-硝基苯基)-N，N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000103
将化合物1-氯-5-甲氧基-2-甲基-4-硝基苯(2g,9.92mmol)，N，N-二甲基哌啶盐酸盐(2.195g,10.91mmol)，碳酸铯(16.16g,49.6mmol)，Pd2(dba)3(0.454g,0.496mmol)，Xantphos(0.574g,0.992mmol)置于10毫升的微波管，加入DMF(20ml)。往液下鼓氮气2分钟，微波下在130度搅拌反应40分钟。TLC监测完毕后，反应冷却至室温，往反应混合物加水稀释，用乙酸乙酯萃取。有机相用饱和食盐水，无水硫酸钠干燥，过滤浓缩，经柱层析(DCM：MeOH 20:1)得到棕色的固体产品0.45g，产率15.46％。
LCMS:t＝2.69min,294.1(M+H+).
第二步：1-(4-氨基-5-甲氧基-2-甲基苯基)-N，N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000104
将化合物1-(5-甲氧基-2-甲基-4-硝基苯基)-N,N-二甲基哌啶-4-胺(0.45g,1.534mmol)和氯化铵(0.410g,7.67mmol)置于100毫升的单口烧瓶中，加入10毫升甲醇，室温搅拌下分批加入锌粉(0.501g,7.67mmol)。加热回流3小时。TLC监测反应完成后，冷却至室温，减压过滤，母液旋干，加入DCM溶解，用碳酸氢钠溶液洗涤，无水硫酸钠干燥，过滤浓缩得到白色的产品0.38g，产率80％。
LCMS:t＝0.43min,264.1(M+H+).
实施例33:5-氯-N2-(2-(二氟甲氧基)-4-(4-(二甲基氨基)哌啶-1-基)-5-甲基苯基)-N4-(2-(异丙基磺酰基)苯基)嘧啶-2,4-二胺(式I-102所示化合物)的制备
制备路线：
Figure PCTCN2016104723-appb-000105
将2,5-二氯-N-(2-(异丙基磺酰)苯基)嘧啶-4-胺(183mg,0.529mmol)和1-(4-氨基-5-(二氟甲氧基)-2-甲基苯基)-N,N-二甲基吡啶-4-胺(158mg,0.529mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.529mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，粗品经柱层析分离纯化得黄色固体81mg，产率25.2％，即为式I-102所示化合物。
实施例34：式I-103所示化合物的制备
Figure PCTCN2016104723-appb-000106
将
式a所示化合物(200mg,0.529mmol)和式b所示化合物(140mg,0.529mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.529mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得浅黄色固体产物136mg，收率42.5％，即为式I-103所示化合物。
实施例35：式I-107所示化合物的制备
Figure PCTCN2016104723-appb-000107
将式c所示化合物(150mg,0.43mmol)和式b所示化合物(114mg,0.43mmol)溶于正丁醇(1.5mL),并向反应液中加入对甲苯磺酸(74.40mg,0.43mmol)，升温至115摄氏度，搅拌反应5小时。TLC监测反应完毕，冷却至室温，减压蒸除溶剂后，将粗品溶于二氯己烷，经饱和碳酸氢钠水溶液及饱和食盐水洗涤后，无水硫酸钠干燥，过滤浓缩得粗品，粗品经柱层析分离纯化得到浅绿色的固体101mg，产率41％，即为式I-107所示化合物。
实施例36：式I-109所示化合物的制备
合成路线及制备方法：
Figure PCTCN2016104723-appb-000108
将式d所示化合物(137mg,0.397mmol)和式b所示化合物(105mg,0.397mmol)溶于正丁醇(1.5mL)，并向反应液中加入对甲苯磺酸(68mg,397mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得浅绿色的固体81mg，产率35.6％，即为式I-109所示化合物。
实施例37：5-(二氟甲氧基)-N2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基苯基)-N4-(2-(异丙基磺酰基)苯基)嘧啶-2,4-二胺(式I-111所示化合物)的制备
合成路线及制备方法：
Figure PCTCN2016104723-appb-000109
将2-氯-5-(2-氟甲氧基)-N-(2-(异丙基磺酰基)苯基)嘧啶-4-胺(式12所示化合物)(200mg,0.529mmol)和1-(4-胺基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺(式5所示化合物)(132mg,0.529mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,0.529mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得浅黄色固体产物121mg，收率38.3％，即为式I-111所示化合物。
1H NMR(400MHz,CDCl3)δ9.56(s,1H),8.70(d,J＝8.4Hz,1H),8.07-8.04(m,2H),7.87(dd,J＝7.9,1.2Hz,1H),7.60(t,J＝7.3Hz,1H),7.29(s,1H),7.21(t,J＝7.6Hz,1H),6.62(d,J＝144Hz,1H),6.60–6.54(m,1H),6.50(dd,J＝8.8,2.4Hz,1H),3.87(s,3H),3.65(d,J＝12.3Hz,2H),3.22(dt,J＝13.6,6.8Hz,1H),2.71(dd,J＝12.0,10.4Hz,2H),2.36(d,J＝12.8Hz,7H),1.99(d,J＝12.3Hz,2H),1.72(tt,J＝12.0,6.1Hz,2H),1.29(d,J＝6.8Hz,6H).
LCMS:t＝3.08min,591.2(M+H+).
其中，本实施例中所述1-(4-胺基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺(式5所示化合物)
Figure PCTCN2016104723-appb-000110
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000111
第一步：1-(3-甲氧基-4-硝基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000112
将化合物4-氟-2-甲氧基-1-硝基苯(5g,29.2mmol),N,N-二甲基哌啶-4-胺盐酸盐(5.88g,29.2mmol)和碳酸钾(12.11g,88mmol)加入100毫升的单口瓶里，加入60毫升DMF。加热至80℃搅拌过夜。TLC监测反应完成后，冷却至室温，加水稀释，用乙酸乙酯萃取两次。合并的有机层经饱和食盐水洗涤，无水硫酸钠干燥后，浓缩溶剂得到棕色油状产品5g，产率80％。
LCMS:t＝0.48min,280.1(M+H+).
第二步：1-(4-氨基-3-甲氧基苯基)-N,N-二甲基哌啶-4-胺的制备
Figure PCTCN2016104723-appb-000113
将化合物1-(3-甲氧基-4-硝基苯基)-N,N-二甲基哌啶-4-胺(6.5g,23.27mmol)溶在甲醇(10ml)，并加入氯化铵(0.410g,7.67mmol)，在室温搅拌下分批加入锌粉(15.21g,233mmol)。加热回流3个小时。TLC监测反应完成后，冷却至室温，过滤，旋干，用DCM溶解，饱和的碳酸氢钠溶液洗涤，无水硫酸钠干燥，过滤浓缩得到棕色油状的产品，即为式5所示化合物，得量5g，产率86％。
1H NMR(400MHz,dmso)：δ6.47(d,J＝8.4Hz,1H),6.45(d,J＝2.4Hz,1H),6.26(dd,J＝8.4,2.4Hz,1H),4.17(s,2H),3.70(s,3H),3.38(d,J＝12.4Hz,4H),2.16(s,6H),2.08(s,1H),1.78(d,J＝12.4Hz,2H),1.45(dd,J＝11.6,3.6Hz,2H).
LCMS:t＝0.43min,250.1(M+H+).
其中，本实施例中所述2-氯-5-(2-氟甲氧基)-N-(2-(异丙基磺酰基)苯基)嘧啶-4-胺(式12所示化合物)
Figure PCTCN2016104723-appb-000114
其合成路线及制备方法如下：
Figure PCTCN2016104723-appb-000115
第一步:2-氯-N-(2-(异丙基磺酰基)苯基)-5-甲氧基嘧啶-4-胺的制备
将2-异丙基磺酰基苯胺(2g,10.04mmol)溶于无水DMF(25ml)，冰水浴冷却到0℃，并在氮气保护下，向反应体系中分批加入NaH(1.806g,60.2mmol,80％)，继续在0℃下搅拌反应1小时，向反应体系中滴加2，4-二氯-5-甲氧基嘧啶(10.78g,60.2mmol)的DMF溶液(10mL)。反应体系自然升温至室温条件下继续搅拌反应10小时，冰水浴冷却，滴加冰水淬灭反应，并用乙酸乙酯萃取，用水及饱和食盐水洗涤后，经无水硫酸钠干燥，过滤浓缩得粗产品，经柱层析分离纯化(PE/EA＝10:1～5:1)后得白色固体产 物3.18g，收率90％。
LCMS:t＝3.85min,342.0(M+H+).
第二步：2-氯-4-((2-(异丙基磺酰基)苯基)氨基)嘧啶-5-酚的制备
将2-氯-4-((2-(异丙基磺酰基)苯基)-5-甲氧基嘧啶-4-胺(2.5g,7.31mmol)溶于无水二氯甲烷(25mL)中，冰水浴冷却到0℃，向反应体系中滴加三溴化硼(2.78ml,29.3mmol)的二氯甲烷(10mL)溶液。滴加完毕后，反应液继续搅拌反应5小时，用冰水淬灭反应，经饱和碳酸氢钠和饱和食盐水洗涤，无水硫酸钠干燥后，过滤浓缩得粗产品，经柱层析分离纯化后PE/EA(V/V＝5:1～3:1)得白色固体2.06g，收率77％。
LCMS:t＝3.60min，328.0(M+H+).
第三步：2-氯-5-(二氟甲氧基)-N-(2-(异丙基磺酰基)苯基)嘧啶-4-胺的制备
将2-氯-4-((2-(异丙基磺酰基)苯基)氨基)嘧啶-5-酚(500mg,1.342mmol)溶于混合溶剂DMF/H2O(5mL/1.5mL)中，并向其中加入2-氯-2,2-二氟醋酸钠(614mg,4.03mmol)及K2CO3(557mg,4.03mmol)，氮气保护，加热至120℃搅拌反应15小时，反应完毕，冷却到室温，用乙酸乙酯(50mL)稀释，经水及饱和食盐水洗涤，无水硫酸钠干燥，过滤浓缩得粗产品，经柱层析分离PE/EA(V/V＝5:1～2:1)得白色固体产物235mg，收率45％。
LCMS:t＝4.00min，378.0(M+H+).
实施例38：5-(二氟甲氧基)-N2-(4-(4-(二甲基氨基)哌啶-1-基)-2-甲氧基-5-甲基苯基)-N4-(2-(异丙基磺酰基)苯基)嘧啶-2,4-二胺(式I-112所示化合物)的制备
合成路线及制备方法：
将中间体12(200mg,529mmol)和1-(4-氨基-5-甲氧基-2-甲基苯基)-N，N-二甲基哌啶-4-胺(式2所示化合物)(139mg,529mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(91mg,529mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得灰白色固体64mg，产率20％，即为式I-112所示化合物。
1H NMR(400MHz,CDCl3):δ9.56(s,1H),8.67(d,J＝8.4Hz,1H),8.06(d,J＝12.8Hz,2H),7.87(d,J＝7.2Hz,1H),7.60(t,J＝7.2Hz,1H),7.38(s,1H),7.21(t,J＝7.6Hz,1H),6.58(d,J＝144Hz,1H),6.58(s,2H),3.85(s,3H),3.23-3.20(m,3H),2.82–2.59(m,8H),2.21(d,J＝11.2Hz,2H),2.17(s,3H),1.90(dd,J＝21.2,9.2Hz,3H),1.28(d,J＝6.8Hz,6H).
LCMS:t＝3.05min,605.2(M+H+).
实施例39：5-(二氟甲氧基)-N2-(4-(4-(二甲基氨基)哌啶-1-基)-2-异丙氧基苯基)-N4-(2-(异丙基磺酰基)苯基)嘧啶-2,4-二胺(式I-113所示化合物)的制备
合成路线及制备方法：
Figure PCTCN2016104723-appb-000117
将中间体12(150mg,397mmol)和1-(4-胺基-3-异丙氧苯基)-N,N-二甲基氨基哌啶-4-胺(式9所示化合物)(110mg,397mmol)溶于正丁醇(1.5mL)，并向反应液中加入对甲苯磺酸(68mg,397mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得浅绿色的固体54mg，产率22％，即为式I-113所示化合物。
1H NMR(400MHz,CDCl3)：δ9.57(s,1H),8.70(d,J＝8.4Hz,1H),8.11(d,J＝8.8Hz,1H),8.07(s,1H),7.87(d,J＝8.0Hz,1H),7.60(s,1H),7.38(s,1H),7.22(t,J＝7.6Hz,1H),6.67(d,J＝72.2Hz,1H),6.54(s,1H),6.48(d,J＝8.8Hz,1H),4.75-4.40(m,2H),3.66(d,J＝12.0Hz,2H),3.21(d,J＝6.8Hz,1H),2.73(t,J＝12.0Hz,2H),2.65(s,6H),2.18(d,J＝12.0Hz,2H),1.87(d,J＝9.6Hz,2H),1.37(d,J＝8.0Hz,6H),1.29(d,J＝6.8Hz,6H).
LCMS:t＝3.94min,619.0(M+H+).
实施例40：5-氯-N2-(2-(二氟甲氧基)-4-(4-(二甲基胺基)哌啶-1-基)苯基)-N4-(2-(异丙氧基磺酰基)苯基)嘧啶-2,4-二胺(式I-114所示化合物)的制备
合成路线及制备方法：
Figure PCTCN2016104723-appb-000118
将中间体12(150mg,397mmol)和1-(4-氨基-3-(二氟甲氧基)苯基)-N,N-二甲基哌啶-4-胺(式3所示化合物)(113mg,397mmol)溶于正丁醇(1.5mL)，并向反应液中加入对甲苯磺酸(68mg,397mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得白色的固体64mg，产率25.7％，即为式I-114所示化合物。
1H NMR(400MHz,CDCl3):δ9.63(s,1H),8.63(d,J＝8.4Hz,1H),8.07(s,1H),8.03(d,J＝9.2Hz,1H),7.86(d,J＝8.0Hz,1H),7.57(t,J＝8.0Hz,1H),7.21(t,J＝7.6Hz,1H),7.01(s,1H),6.77(dd,J＝8.8,2.8Hz,1H),6.73(s,1H),6.58-6.31(m,2H),6.54(d,J＝37.6Hz,1H),6.35(d,J＝39.2Hz,1H),3.67(d,J＝12.4Hz,2H),3.20(dd,J＝13.6,6.8Hz,1H),2.73(t,J＝11.6Hz,2H),2.42(s,6H),2.03(d,J＝11.6Hz,2H),1.71(dd,J＝20.8,11.6Hz,2H),1.29(d,J＝6.8Hz,6H).
LCMS:t＝3.62min,627.2(M+H+).
实施例41：5-(二氟甲氧基)-N2-(4-(4-异丙基哌嗪-1-基)-2-甲氧基-5-甲基苯基)-N4-(2-(异丙基磺酰基)苯基) 嘧啶-2,4-二胺(式I-115所示化合物)的制备
合成路线及制备方法：
Figure PCTCN2016104723-appb-000119
将式12所示化合物(150mg,397mmol)和4-(4-异丙基哌嗪-1-基)-2-甲氧基-5-甲基苯胺(式10所示化合物)(105mg,397mmol)溶于n-BuOH(2.5ml)，并向其中加入对甲苯磺酸(68mg,397mmol)，加热至120℃，搅拌反应3小时。LCMS检测反应完毕。将反应混合液冷却至室温，用乙酸乙酯稀释(25mL)，经水洗及饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得粗产品，再经柱层析分离(DCM/MeOH(v/v＝40:1～20:1))得白色固体42mg，产率17.5％，即为式I-115所示化合物。
1H NMR(400MHz,CDCl3):δ9.57(s,1H),8.68(d,J＝8.0Hz,1H),8.10(s,2H),7.89(dd,J＝8.0,1.6Hz,1H),7.69–7.59(m,1H),7.43(s,1H),7.24–7.13(m,1H),6.80–6.37(m,3H),3.88(s,3H),3.57-2.75(m,10H),2.19(s,3H),1.30(d,J＝6.8Hz,6H).
LCMS:t＝3.42min,605.3(M+H+).
实施例42：
在该实施例中，按照与上述实施例36类似的制备过程，合成了式I-110所示化合物。
实施例43：式I所示化合物对ALK激酶抑制活性的测定
采用以下方法来测定本发明的化合物在体外对ALK激酶的抑制活性，该抑制活性采用IC50这一指标来表示，IC50即ALK激酶的活性被抑制50％时的化合物的浓度。
简写及定义
mg    毫克
mL    毫升
μg   微克
μl   微升
mM    毫摩尔每升
EDTA  乙二胺四乙酸
DMSO  二甲亚砜
SD    标准偏差
SOP   标准操作程序
实验材料：
ALK(Carna,Cat.No 08-105,Lot.No.08CBS-0112)
ALK L1196M(Carna,Cat.No 08-529,Lot.No.11CBS-1134)
Peptide FAM-P22(GL Biochem,Cat.No.112393,Lot.No.P080401-XY112393)
ATP(Sigma,Cat.No.A7699-1G,CAS No.987-65-5)
DMSO(Sigma,Cat.No.D2650,Lot.No.474382)
EDTA(Sigma,Cat.No.E5134,CAS No.60-00-4)
96孔板(Corning,Cat.No.3365,Lot.No.22008026)
384孔板(Corning,Cat.No.3573,Lot.No.12608008)
Staurosporine(Sigma,Cat.No.S4400-1MG,Lot.No.046K4080)
实验方法：
1.配制1倍的激酶缓冲液和终止液
1)1倍激酶缓冲液
50mM HEPES,pH 7.5
0.0015％Brij-35
10mM MgCl2
2mM DTT
2)终止液
100mM HEPES,pH 7.5
0.015％Brij-35
0.2％Coating Reagent#3
50mM EDTA
2.化合物配制
1)将化合物稀释50倍最终所需的最高抑制浓度的100％DMSO溶液。转移100μL这个化合物稀释液在96孔板中。例如，如果需要最高的抑制剂浓度为1μM，在这一步就制备50μM的DMSO溶液。
2)将化合物按3倍序列稀释成10个浓度。
3)添加100μl 100％DMSO到2个空的没有化合物控制和无酶控制的相同的96孔板中。标记源板。
4)中间板的制备
从源板中转移10μL化合物到一个新的96孔板中作为中间板。
添加90μL1倍激酶缓冲液到每一个中间板的孔中。
混合化合物到中间板中并且振荡10分钟。
3.准备试验板
从96孔中间板中每一个孔中转移5μl到384孔板中作为重复。例如，96孔板中的A1转移到384孔板中的A1和A2。96孔板中的A2转移到384孔板的A3和A4。
4.激酶反应
1)配制2.5倍酶溶液
将激酶加入1倍激酶缓冲液，形成2.5倍酶溶液。
2)配制2.5倍的底物溶液
将FAM标记的多肽和ATP加入1倍激酶缓冲液，形成2.5倍底物溶液。
3)试验板已含有5μl化合物的10％的DMSO溶液。
4)转移2.5倍酶溶液到试验板。
5)室温下孵育10分钟
6)转移2.5倍肽溶液到试验板。
7)激酶的反应和停止
28℃下孵育20分钟。加入25μl停止液终止反应。
5.Caliper读取数据
Caliper上读取转化率数据。
6.曲线拟合
1)从Caliper上复制转化率数据。
2)把转化率转化成抑制率数据。其中max是指DMSO对照的转化率，min是指无酶活对照的转化率。
Percent inhibition＝(max-conversion)/(max-min)*100.
3)将数据导入MS Excel并使用XLFit excel add-in version 4.3.1进行曲线拟合。
使用的公式是:
Y＝Bottom+(Top-Bottom)/(1+10^((LogIC50-X)*HillSlope))
本发明所述化合物的生化学活性通过以上试验进行测定，本发明所述化合物测得的IC50值实验结果见下表1。
其中，对ALK抑制活性指定为“A”的化合物提供的IC50≤1nM；活性指定为“B”的化合物提供的IC50为1-10nM；活性指定为“C”的化合物提供的IC50为IC50≥10nM。对ALK L1196M抑制活性指定为“a’”的化合物提供的IC50≤1nM；活性指定为“b”的化合物提供的IC50为1-10nM；活性指定为“c”的化合物提供的IC50为IC50≥10nM。
表1
Figure PCTCN2016104723-appb-000120
Figure PCTCN2016104723-appb-000121
由此，上述测试结果表明：本发明式I所示化合物对ALK及ALKL1196M酶活性均有明显的抑制作用。结果显示，本发明所述式I所示化合物均具有良好的ALK激酶抑制活性，本发明所述化合物可用作ALK抑制剂，用于治疗一种或一种以上与ALK活性有关的肿瘤疾病，用于制备成抑制间变性淋巴瘤激酶的抗肿瘤治疗药物。
实施例44：式I所示化合物对Karpas299细胞增殖抑制活性的测定
1实验原理
ATP的含量直接反应了细胞的数量及细胞状态，通过对ATP进行定量测定来检测培养物中活细胞数目。活细胞检测试剂盒采用萤光素酶作检测物，试剂盒中使用UltraGlow萤光素酶生成的稳定辉光型信号，发光过程中萤光素酶需要ATP的参与，有代谢活性细胞的呼吸作用和其他生命活动过程可以产生ATP。向细胞培养基中加入CellTiter-GloTM试剂，测量发光值，光信号和体系中ATP量成正比，而ATP又和活细胞数正相关，从而可以检测出细胞的增殖情况。检测板使用PE公司的Envision进行分析。
2材料
细胞培养：RPMI-1640培养基、胎牛血清、抗生素(Penicillin-Streptomycin)
细胞系：Karpas299
检测试剂：活细胞检测试剂盒CellTiter-Glo
其他主要耗材及试剂：化合物稀释板，中间板，检测板，DMSO
3实验步骤
3.1制备细胞板
将Karpas299细胞种于黑色384孔板中，每孔45μl细胞悬液，包含2500个细胞每孔。细胞板置于二氧化碳培养箱中过夜培养。
3.2准备化合物
1)将待测化合物溶解成10mM DMSO溶液，并放置于氮气柜中进行长期保存。
2)取10μl 10mM待测化合物贮存溶液，稀释成为2.5mM的工作溶液，用ECHO进行3倍稀释至第10个浓度，即从5μM稀释至0.25nM，设置双复孔实验。
3.3化合物处理细胞
向中间板中加入49μl培养基，再按照对应位置，每孔转移1μl梯度稀释的化合物至中间板，混匀后转移5μl每孔到细胞板中。细胞板置于二氧化碳培养箱中培养3天。
3.4检测
向细胞板中加入每孔25μl的Promega CellTiter-Glo试剂，室温孵育10分钟使发光信号稳定。采用PerkinElmer Envision多标记分析仪读数。
表2显示了本发明化合物对Karpas299细胞增殖抑制活性。其中，在对Karpas299细胞增殖抑制活性实验数据中，活性指定为“D”的化合物提供的IC50≤50nM；活性指定为“E”的化合物提供的IC50为50-100nM；活性指定为“F”的化合物提供的IC50为IC50≥100nM。
表2
Figure PCTCN2016104723-appb-000122
由此，上述测试结果表明：本发明式I所示化合物对ALK酶具有明显的抑制活性，并且对Karpas 299细胞具有明显的增殖抑制活性。
实施例45：式I所示化合物对Ba/F3EML4-ALK细胞增殖抑制活性的测定
1、试剂和耗材
1)细胞培养：RPMI-1640培养基、胎牛血清、抗生素(Penicillin-Streptomycin),IL-3,嘌呤霉素
2)细胞系：Ba/F3EML4-ALK
3)检测试剂：活细胞检测试剂盒CellTiter-Glo
4)其他主要耗材及试剂：化合物稀释板，中间板，检测板，DMSO
2、实验原理
ATP的含量直接反应了细胞的数量及细胞状态，通过对ATP进行定量测定来检测培养物中活细胞数目。活细胞检测试剂盒采用萤光素酶作检测物，试剂盒中使用UltraGlow萤光素酶生成的稳定辉光型信号，发光过程中萤光素酶需要ATP的参与，有代谢活性细胞的呼吸作用和其他生命活动过程可以产生ATP。向细胞培养基中加入CellTiter-GloTM试剂，测量发光值，光信号和体系中ATP量成正比，而ATP又和活细胞数正相关，从而可以检测出细胞的增殖情况。检测板使用PE公司的Envision进行分析。
3、实验方法
1)制备细胞板
将Ba/F3EML4-ALK细胞分别种于384孔板中，每孔包含300个细胞。细胞板置于二氧化碳培养箱中过夜培养。
2)准备化合物
用BRAVO进行3倍稀释，10个化合物浓度，设置双复孔实验。
3)化合物处理细胞
将化合物转移到细胞板中，化合物起始浓度为5μM。细胞板置于二氧化碳培养箱中培养3天。
4、检测
向细胞板中加入Promega CellTiter-Glo试剂，室温孵育10分钟使发光信号稳定。采用PerkinElmer Envision多标记分析仪读数。
表3显示了本发明化合物的测试结果。其中，活性指定为“d”的化合物提供的IC50≤20nM；活性指定为“e”的化合物提供的IC50为20-100nM；活性指定为“f”的化合物提供的IC50为IC50≥100nM。
表3
化合物编号 Ba/F3EML4-ALK(IC50nM)
I-1 d
I-2 d
I-3 d
I-4 d
I-5 d
I-6 d
I-7 d
I-8 d
I-9 e
I-10 d
I-11 d
I-12 d
I-13 d
I-14 d
I-15 d
I-16 d
I-17 d
I-18 d
I-23 d
I-80 e
I-82 e
I-84 e
I-85 e
I-86 d
I-88 e
I-89 d
I-90 d
I-91 d
I-92 d
I-95 e
I-98 e
I-100 e
I-102 d
I-103 d
I-107 e
I-109 d
I-110 d
I-111 d
I-112 d
I-113 d
I-114 d
I-115 d
试验结果表明，本发明所述化合物对Ba/F3EML4-ALK细胞具有明显的增殖抑制活性。
实施例45动力学溶解度测试：
动力学溶解度的测试在药物发现阶段通常用于药物的高通量筛选。在动力学分析中，一个良好的溶解度应该有助于产生可靠的在体外和体内的数据。由于动力学溶解度是pH依赖性，水相的pH值总是指定的，通常是测量在pH值为7.4(体液的生理pH值)。
测试方法：称量定量化合物样品溶解在纯DMSO中，终浓度为10mM，将受试化合物与对照化合物(10mM DMSO母液，每孔10μL)加入含有每孔490μL缓冲液的96孔板中。涡旋2分钟后，样品板在振荡器上室温下(22±2℃)孵育24小时。然后转移200μL样品到MultiScreen过滤板(聚碳酸酯膜)，以微孔真空歧管(millipore vacuum manifold)过滤并收集滤液。以HPLC-UV测定滤液中化合物的浓度。3个不同浓度的UV标准品溶液与溶解度测试样品先后进样。每个样品进针2次，带入标准曲线计算浓度，求平均值。
实验结果显示，本发明所述化合物具有良好的水溶性质，且优于对照样(其中对照样为克唑替尼和色瑞替尼(LDK378))。
实施例46体外代谢稳定性测试：
体外代谢稳定性实验评定化合物在一相代谢中的清除率，并能预测其在肝细胞和体内的固有清除率。我们通过体外代谢稳定实验评价了本发明所述部分化合物在人和大鼠肝微粒体的代谢稳定性。其中 对照样为克唑替尼和色瑞替尼(LDK378)。
材料和方法：
1.待测化合物；
2.缓冲液，100nM的磷酸钾缓冲液，PH 7.4；10mM MgCl2；
化合物稀释：
1)中间液：将5μL待测化合物或对照样的储存溶液(10mM)用45μL DMSO和450μL 1:1甲醇/水(Conc.:100μM,45％甲醇)进行稀释；
2)工作液：将50μL中间液用450μL 100mM的磷酸钾缓冲液(Conc.:10μM,4.5％甲醇)稀释。
停止液：冷的乙腈(CAN)包含100ng/mL甲苯磺丁脲(Tolbutamide)和100ng/mL拉贝洛尔(Labetalol)作为内标(IS)
实验操作：
1.使用液体工作站将工作液(10μM)以每孔10μL体积分到相应反应板上。
2.使用液体工作站将肝微粒体溶液(0.625mg/mL)以每孔80μL体积分到相应反应板。(微粒体终浓度为0.5mg/mL)
3.在37℃下预热化合物与微粒体的混合液10分钟。
4.在不含有辅助因子的60分钟反应板中，每孔添加10μL浓度为100mM的磷酸缓冲液，并开始计时60分钟。
5.在其他反应板中，每孔添加10μL NADPH辅助因子溶液(组分终浓度为1mM NADP,1mM氯化镁,6.5mM异柠檬酸和1unit/mL异柠檬酸脱氢酶)，启动反应并计时。对于0分钟反应板，先加入乙腈终止液终止反应后再加入微粒体溶液和NADPH溶液。
6.在5、10、20、30和60分钟，分别添加入300μL乙腈终止液(含100ng/mL甲苯磺丁脲)，终止反应。
7.充分摇板后，在4000rpm下离心20分钟。
8.转移100μL上清液与300μL纯水混匀。
9.使用LC-MS/MS分析样品。
使用一级动力学方程计算半衰期，清除率和固有清除率。
实验结果显示，本发明所述化合物显示出了良好的代谢稳定性，为进一步的临床前研究提供了重要依据。
实施例47Caco-2实验：
Caco-2细胞是一种广泛用于研究小肠吸收的体外模型，它是一种人的结肠癌细胞。单层Caco-2细胞模型已广泛地应用于评估在小肠吸收过程中的被动和主动转运过程。GF120918A为一个强外排转运体抑制剂，这些外排转运体包含P糖蛋白(P-gp)和乳腺癌抗性蛋白(BCRP)等。发明人通过此实验评估了本发明所述部分化合物的透膜性，从而预估了化合物在小肠中吸收的性质。
其中对照样为克唑替尼和色瑞替尼(LDK378)。
实验操作：实验标准条件如下：1.测试浓度:2μM(DMSO≤1％)；2.重复：n＝2；3.方向:双向转运，包括A→B和B→A两个方向；4.孵育时间：单个时间点，2小时；5.转运缓冲液：HBSS，pH7.4；6.孵育条件:37℃,5％CO2。7.孵育结束后，取给药孔和接收孔内的样品溶液立即与含有内标的冷乙腈溶液混合。8.采用LC/MS/MS方法分析待测化合物在所有样品(包括起始给药液，给药孔上清液，接收液)中的浓度。并计算表观渗透系数，外排比等参数。
实验结果显示，本发明所述化合物相对于对照样大大提高了透膜性，从而改善了化合物在肠道中的 吸收性质，并能提高药物的生物利用度。
在本说明书的描述中，参考术语“一个实施例”、“一些实施例”、“示例”、“具体示例”或“一些示例”等的描述意指结合该实施例或示例描述的具体特征、结构、材料或者特点包含于本发明的至少一个实施例或示例中。在本说明书中，对上述术语的示意性表述不一定指的是相同的实施例或示例。而且，描述的具体特征、结构、材料或者特点可以在任何的一个或多个实施例或示例中以合适的方式结合。
尽管上面已经示出和描述了本发明的实施例，可以理解的是，上述实施例是示例性的，不能理解为对本发明的限制，本领域的普通技术人员在不脱离本发明的原理和宗旨的情况下在本发明的范围内可以对上述实施例进行变化、修改、替换和变型。

Claims (17)

  1. 一种化合物，其特征在于，为式I所示化合物或式I所示化合物的药学上可接受的盐、水合物、溶剂化物、代谢产物、或前药，
    Figure PCTCN2016104723-appb-100001
    其中，R1为5-6元环烷基、5-6元杂环基、5-6元芳基、或5-6元杂芳基，任选地，所述5-6元环烷基、5-6元杂环基、5-6元芳基和5-6元杂芳基各自独立地被一个或多个选自卤素、羟基、氰基、硝基、C1-8烷基、C2-8链烯基、C2-8链炔基、C3-8环烷基、3-8元杂环基、C5-10芳基、5-10元杂芳基、C1-6烷氧基、C3-8环烷氧基、-S(O)pR6、-C(O)R6、-C(O)OR6、-NR7R8或-C(O)NR8的取代基取代，其中，R6、R7、R8各自独立地为氢或Cl-4烷基，p为0、1或2；
    R2为氢、卤素、C1-6烷基、C1-6烷氧基、卤素取代的C1-6烷基或卤素取代的C1-6烷氧基；
    R3为任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基、任选被取代的环己胺基、任选被取代的C1-2烷基或任选被取代的1,2,3,6-四氢吡啶基，所述被取代的C1-2烷基上的取代基为任选被取代的哌啶基；
    R4为氢或C1-6烷基；
    R5为氢、氯、羟基、氰基、C1-6烷基、C1-6烷氧基或卤素取代的C1-6烷氧基。
  2. 根据权利要求1所述的化合物，其特征在于，
    R1为被一个或多个-S(O)pR6取代的5-6元杂环基、5-6元芳基、或5-6元杂芳基；
    R2为氢、卤素、C1-6烷基、C1-6烷氧基或卤素取代的C1-6烷氧基；
    R3为任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基、任选被取代的甲基或任选被取代的1,2,3,6-四氢吡啶基，所述任选被取代的甲基上的取代基为任选被取代的哌啶基；所述任选被取代的哌嗪基、任选被取代的吗啉基、任选被取代的哌啶基或任选被取代的1,2,3,6-四氢吡啶基任选地被一个或多个选自C1-6烷基、羟基、C1-6烷基氨基、C1-6烷氧基、氧代(＝O)、C1-6酰基、吗啉基、C1-6烷基吗啉基、哌嗪基、C1-6烷基哌嗪基、C1-6酰基哌嗪基；羟基C1-6烷基哌嗪基、哌啶基和C1-6烷基氨基哌啶基的取代基取代；
    R4为氢或C1-6烷基；
    R5为氯或卤素取代的C1-6烷氧基；
    R6为Cl-4烷基；
    p为2。
  3. 根据权利要求2所述的化合物，其特征在于，
    R1为
    Figure PCTCN2016104723-appb-100002
    中的任意一种；
    R2为氢、氯、甲基、甲氧基、乙氧基、异丙氧基或二氟甲氧基；
    R3为下列之一：
    Figure PCTCN2016104723-appb-100003
    R4为氢或甲基；
    R5为氯或二氟甲氧基。
  4. 根据权利要求1所述的化合物，其特征在于，所述化合物为下列化合物之一，或其药学上可接受的盐、水合物、溶剂化物、代谢产物、或前药：
    Figure PCTCN2016104723-appb-100004
    Figure PCTCN2016104723-appb-100005
    Figure PCTCN2016104723-appb-100006
    Figure PCTCN2016104723-appb-100007
  5. 一种药物组合物，其特征在于，包含权利要求1～4中任一项所述的化合物。
  6. 根据权利要求5所述的药物组合物，其特征在于，所述药物组合物用于下列至少之一：
    抑制激酶；
    抑制间变性淋巴瘤激酶；
    治疗或者预防癌症；以及
    抑制癌细胞的增殖。
  7. 根据权利要求6所述的药物组合物，其特征在于，所述癌症为肺癌和间变性大细胞非霍奇金淋巴瘤。
  8. 根据权利要求7所述的药物组合物，其特征在于，所述肺癌为非小细胞肺癌。
  9. 根据权利要求7或8所述的药物组合物，其特征在于，所述药物组合物进一步包括第二药剂，所述第二药剂不同于权利要求1～4中任一项所述化合物并且用于抑制激酶、抑制间变性淋巴瘤激酶、治疗或者预防癌症和抑制癌细胞的增殖的至少之一。
  10. 根据权利要求5所述的药物组合物，其特征在于，所述药物组合物进一步包括药学上可接受的载体、赋形剂、稀释剂、辅剂、媒介物或其组合。
  11. 根据权利要求10所述的药物组合物，其特征在于，所述药物组合物呈片剂、胶囊、注射剂、粉针剂、粉剂、糖浆、溶液状、悬浮液或气雾剂。
  12. 权利要求1～4中任一项所述化合物或权利要求5-11中任一项所述的药物组合物在制备药物中的用途，所述药物用于下列至少之一：
    抑制激酶；
    抑制间变性淋巴瘤激酶；
    治疗或者预防癌症；以及
    抑制癌细胞的增殖。
  13. 根据权利要求12所述的用途，其特征在于，所述癌症为肺癌和间变性大细胞非霍奇金淋巴瘤。
  14. 根据权利要求13所述的用途，其特征在于，所述肺癌为非小细胞肺癌。
  15. 权利要求1～4中任一项所述化合物或权利要求5-11中任一项所述的药物组合物在下列至少之一中的用途：
    抑制激酶；
    抑制间变性淋巴瘤激酶；
    治疗或者预防癌症；以及
    抑制癌细胞的增殖。
  16. 一种抑制激酶、抑制间变性淋巴瘤激酶、治疗或者预防癌症和/或抑制癌细胞的增殖的方法，其特征在于，包括：给予受试对象有效剂量的权利要求1～4中任一项所述化合物或权利要求5-11中任一项所述的药物组合物。
  17. 一种治疗与ALK活性有关的肿瘤疾病的方法，其特征在于，包括：给予受试对象有效剂量的权利要求1～4中任一项所述化合物或权利要求5-11中任一项所述的药物组合物。
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