CN106188001A - A kind of Anti-angiogenic compounds - Google Patents

A kind of Anti-angiogenic compounds Download PDF

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CN106188001A
CN106188001A CN201610550043.6A CN201610550043A CN106188001A CN 106188001 A CN106188001 A CN 106188001A CN 201610550043 A CN201610550043 A CN 201610550043A CN 106188001 A CN106188001 A CN 106188001A
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formula
reactant mixture
dcm
acid
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侯睿
罗红蓉
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Ningbo Fangchang Pharmaceutical Co Ltd
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Abstract

The invention provides noval chemical compound of a kind of anti-angiogenic rebirth and its production and use.The compounds of this invention has good anti-angiogenic rebirth effect, it was initially believed that this compounds produces activity by suppression VEGFR2 (i.e. KDR).This compounds can be used for the treatment to protein kinase caused by abnormal diseases such as new vessels, VEGFR2, PDGFR β, KIT, such as wet MD, malignant tumor etc..

Description

A kind of Anti-angiogenic compounds
The application is application number 201410125752.0,28 days March 2014 applying date, and denomination of invention is " a kind of anti-angiogenic Xinshenghua compound " divisional application.
Technical field
The present invention relates to a kind of noval chemical compound and purposes.
Background technology
Angiogenesis (angiogenesis), is to germinate from existing blood vessel to generate the process of neovascularity.This process and blood Endothelial cell migrates relevant with propagation.Angiogenesis is relevant, such as malignant tumor with multiple mankind's major disease.Have now been found that, Ocular angiogenesis disease, including age-related macular degeneration (AMD), diabetic retinopathy, neovascular green grass or young crops Light eye etc., the common feature of this kind of disease be all ocular angiogenesis paraplasm (gold dawn, etc., anti vegf agents is at eye Application in section's disease and Recent Advances in Mechanism, China and foreign countries' medical treatment, 2012).
Wherein, degeneration of macula mainly has dryness and moist two kinds, and the feature of wet MD (AMD) is, choroidal New vessels enter under retina and then occur hemorrhage, ooze out and the pathological change such as edema.Moist rapid DE, Even more serious compared with dryness.At present, the most preferably it is in progress in terms of the treatment of wet MD.Laser in early days burns bright hemostasis Replaced by blood vessel endothelial factor antagonist, but because of the latter's poor effect, quickly replaced by photodynamic therapy.Photodynamic therapy Although the curative effect of improve, but still undesirable.Occur in that the most again new blood vessel endothelial factor antagonist Lucentis (Lei Zhudan Anti-), it is the recombinant of a kind of humanized's VEGF hypotype monoclonal antibody fragment, new vessels can be reduced and generate.2006, this medicine Thing is approved by the fda in the United States for treating wet MD, and curative effect is good；Meanwhile, such anti vegf agents pair is currently also found Diabetic retinopathy, neovascular glaucoma have therapeutical effect.But owing to Lucentis is antibody medicine, price is high, It can't be popularized in the whole world.Therefore, the little molecule neovascularization inhibitor medicine that research curative effect is good, cheap is to work as The focus of modern international pharmaceutical industry keen competition.
Summary of the invention
Object of the present invention is to provide a kind of noval chemical compound, and its production and use.
The invention provides the compound as shown in formula A and pharmaceutically acceptable salt thereof or prodrug:
Wherein, Z0For O or S；X be C, Y be N；；Z1、Z2It is respectively and independently selected from N or-CR8；
r7Selected from hydrogen or C1-C6Alkyl；r8Selected from hydrogen or C1-C6Alkyl；r10Selected from H, amino, hydroxyl, sulfydryl, C1-C6Alkane Base or-(CH2)n1NHr1, wherein, n1=1-5, r1Alkyl for H or C1-3；
r9Selected from H, amino, hydroxyl, sulfydryl, (C-r11r12)nNr13r14、(C-r11r12)nOr15、(C-r11r12)nC(O) Er13、(C-r11r12)nS(O)mr17；Or r8And r9Saturated 4-7 unit heterocycle or replacement is formed together with the atom being connected with them Heterocycle, wherein, heterocycle or substituted heterocycle contain 1 or 2 hetero atom selected from N, O or S, and its substituent group is 0,1, or 2, respectively Independently selected from oxo group, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxyl, C1-C6 halogenated alkoxy, C1-C6 Alkanoyl, C1-C6 alkylamino radical, C3-C7 cycloalkyl, halogen, hydroxyl, amino, carbonyl, amino carbonyl, C1-C6 aminoalkyl carbonyl, C1-C4 acyl group, C1-C6 alkoxy carbonyl, C1-C6 sulfonyl, amino-sulfonyl；
Ar2Selected from phenyl, naphthyl, monocycle or bicyclic heteroaryl, bicyclo-or tricyclic heterocyclic base, the most each heteroaryl or miscellaneous Ring group has 1,2,3 or 4 hetero atom selected from N, O or S；Described phenyl, naphthyl, heteroaryl or heterocyclic group be unsubstituted or Substituted phenyl, naphthyl, heteroaryl or heterocyclic group, its substituent group is 1,2, or 3, separately selected from C1-C8 alkyl, C1-C8 haloalkyl, hydroxyl C1-C6 alkyl, C1-C8 alkoxyl, C1-C8 halogenated alkoxy, halogen, hydroxyl, amino, amino C- C6 alkyl, C1-C6 alkyl amino, phenyl, C3-C7 cycloalkyl, the C3-C7 cycloalkyl of volution, amino-sulfonyl, C1-C6 alkyl Amino-sulfonyl；
M is 0,1, or 2；N is 0,1,2, or 3；E is empty, oxygen or-Nr18；
r11,r12And r18It is identical or different, is respectively and independently selected from hydrogen or C1-C4 alkyl；r13,r14,r15,r16And r17 Be independently selected from hydrogen, C1-C6 alkyl, C1-C6 acyl group, phenyl and heterocycle, or substituted C1-C6 alkyl, C1-C6 acyl group, phenyl and Heterocycle, its substituent group for for 0,1 or 2, is respectively and independently selected from C1-C4 alkoxyl, C1-C4 alkyl, halogen, hydroxyl, amino, C1-C6 aminoalkyl.
Further, described structural formula of compound is as follows:
Wherein, X be C, Y be N；
R1Selected from H, amino, hydroxyl or sulfydryl；R2Selected from H, amino, hydroxyl, sulfydryl or-(CH2)nNHR8, wherein, n=1- 5, R8Alkyl for H or C1-3；R3Selected from H or C1-6 alkyl；R4、R5、R6It is respectively and independently selected from H, halogen, the alkyl of C1-6 or halogen Element replaces alkyl；R7Alkyl or halogen selected from H, C1-6；
Or, R2、R3Coupled carbon atom collectively form substituted or non-substituted containing 1-2 heteroatomic five yuan or Hexatomic ring, described hetero atom is N, O or S, and its substituent group is the alkyl of C1-6.
Further, X, Y differ；R1Selected from H or amino；R2Selected from H, amino or-(CH2)nNHR8, wherein, n=1-3, R8Alkyl for H or C1-2；R3Selected from H or C1-2 alkyl；R4、R5、R6It is respectively and independently selected from halogen, the alkyl of C1-2 or halogen to take Substituted alkyl；R7Selected from H or halogen；
Or, R2、R3Coupled carbon atom collectively forms substituted or non-substituted five yuan containing 1 N or hexatomic ring, Substituent group is the alkyl of C1-3.
Further, R2Selected from amino or-(CH2)nNHR8；Or, R2、R3Coupled carbon atom collectively forms and takes Generation or non-substituted five yuan containing 1 N or hexatomic ring, substituent group is the alkyl of C1-3.
Further, described structural formula of compound is as follows:
When compound is Formula II, W1, W2 are respectively and independently selected from C or hetero atom, n=1 or 2；R9 is H, C1-C4 alkyl or halogen Element；R10 is halogen；Wherein, during n=1, it is C during W1, W2 difference；Preferably, during n=1, W1, W2 are hetero atom simultaneously, miscellaneous former Son is preferably N；During n=2, W1, W2 are C simultaneously；
When compound is formula III, W3 is selected from C or hetero atom, and hetero atom is preferably N；R10 is halogen.
Further, described halogen is F or Cl.
Preferably, described compound is:
Present invention also offers above-claimed cpd and pharmaceutically acceptable salt thereof or prodrug and prepare angiogenesis Purposes in inhibitor.
Further, described angiogenesis inhibitors is VEGF R2 inhibitor.
Further, described inhibitor is the medicine of anti-ocular angiogenesis.
Further, described medicine is choroid neovascularization inhibitors.
Further, described medicine is treatment or prevention wet MD, diabetic retinopathy or new hemopoietic The medicine of pipe glaucoma.
Present invention also offers above-claimed cpd and pharmaceutically acceptable salt thereof or prodrug preparation VEGFR2, PDGFR-β is or/and purposes in KIT inhibitor class medicine.
Present invention also offers a kind of pharmaceutical composition, it is containing above-claimed cpd and pharmaceutically acceptable salt thereof Ophthalmic preparation.In addition to the above-claimed cpd that the present invention provides, preparation can also comprise other and known there is similar therapeutic The medicine of purposes.
Further, described ophthalmic preparation is eye drop, Eye ointments or ophthalmically acceptable injection.
The method being known in the art can prepare the salt of the compounds of this invention.With acid treatment, or with suitable anion Exchanger, can become salt with above-claimed cpd.The pharmaceutically acceptable salt of the compound of the present invention, can have from above-claimed cpd There is the organic or inorganic acid-addition salts of basic nitrogen atom.
Preferably, suitable mineral acid includes but not limited to, halogen acids, example hydrochloric acid, sulphuric acid, or phosphoric acid.
Preferably, suitable organic acid includes, but not limited to carboxylic acid, phosphoric acid, sulfonic acid or amino carboxylic acid, such as acetic acid, and third Acid, octanoic acid, capric acid, dodecylic acid, hydroxyacetic acid, lactic acid, fumaric acid, succinic acid, adipic acid, 1,5-pentanedicarboxylic acid., suberic acid, nonyl two Acid, malic acid, tartaric acid, citric acid, aminoacid, such as glutamic acid or aspartic acid, maleic acid, hydroxy acid, citraconic acid, ring Cyclohexane carboxylic-acid, adamantanecarboxylic acid, benzic acid, salicylic acid, 4 aminosallcylic acids, phthalic acid, phenylacetic acid, mandelic acid, Cortex Cinnamomi Acid, methane or ethane sulfonic acid sulfonic acid, 2-ethylenehydrinsulfonic acid, ethane-1,2-disulfonic acid, benzenesulfonic acid, 2-LOMAR PWA EINECS 246-676-2,1,5-naphthalene two sulphur Acid, 2-toluene sulfonic acide, p-methyl benzenesulfonic acid, ethyl sulfuric acid, the acid of lauryl sulphate acid, N Cyclohexylamino acetic acid, N-first Base-N-ethyl-N-propyl-amino sulfonic acid, or other organic acid, such as ascorbic acid.
It addition, it can also be used in isolated or purified by the most unacceptable salt, such as picrate or perchloric acid Salt.But, for therapeutic use, can only be pharmaceutically acceptable salt or free cpds, with applicable pharmaceutical preparation Form.
Pharmaceutically acceptable prodrug of the present invention, refers to that described compound obtains after modifying for chemical structure Conversion through enzyme or non-enzymatic in vivo discharge active component and play the compound of drug effect.One embodiment of the present invention In, further comprises isotope-labeled above-claimed cpd or its pharmaceutically acceptable salt, described compound isotopically labelled is Refer to herein listed by compound identical, but one or more atom is replaced by another atom, this atom former Protonatomic mass or mass number are different from atomic mass common in nature or mass number.The isotope bag in compound can be introduced Include hydrogen, carbon, nitrogen, oxygen, sulfur, i.e. 2H, 3H, 13C, 14C, 15N, 17O, 18O, 35S.Containing above-mentioned isotope and/or other atom Isotopic compound and stereoisomer thereof, and the pharmaceutically useful salt of this compound, stereoisomer should be included in this Within invention scope.
Key intermediate and compound in the present invention carry out separating and purification, and the mode used is normal in organic chemistry Isolation and purification method and the example of described method include filtering, extract, be dried, be spin-dried for and various types of chromatographs.Can Selectively, can make intermediate the most purified i.e. carry out next step reaction.
In some embodiments, one or more compounds of the present invention can be used in conjunction with one another.Also may select will The compound of the present invention is used in combination with other active agent any, for preparing regulating cell function or the medicine for the treatment of disease Thing or pharmaceutical composition.If using one group of compound, then can by these compounds simultaneously, respectively or in an orderly manner to tested Object is carried out.
The compounds of this invention has good anti-angiogenic rebirth effect, it was initially believed that this compounds is by suppression VEGFR2 (i.e. KDR) produces activity.This compounds can be used for the protein kinase caused by abnormal disease such as new vessels, VEGFR2 Sick treatment, such as wet MD, malignant tumor etc..
Accompanying drawing explanation
Fig. 1 the compounds of this invention inhibitory action to Brachydanio rerio vascular development
Fig. 2 negative control result figure, wherein, Brachydanio rerio vascular development is normal
Fig. 3 medicine group result figure, wherein, Brachydanio rerio vascular development position is suppressed by the compounds of this invention part
Fig. 4 medicine group result figure, wherein, Brachydanio rerio vascular development position is completely inhibited by the compounds of this invention
Fig. 5 the compounds of this invention is to VEGFR2 In-vitro Inhibitory Effect
The inhibitory action of Fig. 6 the compounds of this invention KDR4 choroidal neovascularization, wherein, A: negative control, B:KDR4
The inhibitory action of Fig. 7 the compounds of this invention V01 choroidal neovascularization, wherein, A: negative control, B:V01
The nuclear-magnetism figure of Fig. 8 compound V01
The mass spectrum of Fig. 9 compound V3
The mass spectrum of Figure 10 compound V4
The mass spectrum of Figure 11 compound V5
The nuclear-magnetism figure of Figure 12 compound V5
The mass spectrum of Figure 13 compound V6
The mass spectrum of Figure 14 compound K DR3
Figure 15 KDR4 compound mass spectrum
The mass spectrum of Figure 16 KDR6
The mass spectrum of Figure 17 KDR6A
The mass spectrum of Figure 18 KDR8
The nuclear-magnetism figure of Figure 19 KDR8A
The mass spectrum of Figure 20 KDR8A
The mass spectrum of Figure 21 KDR8B
The dose effect curve of Figure 22 compound V01
Figure 23 compound V01 is on mice eye new vessels and hemorrhage impact, A: right eye V01 process, B: left eye PBS pair According to
Detailed description of the invention
The preparation of embodiment 1 intermediate of the present invention
Step is as follows:
1st step: mixing Michaelis acid (75.3g, 0.522mol) and triethyl orthoformate (92mL, 0.55mol), is heated to 55 DEG C, maintain 90 minutes, be then cooled to 45 DEG C.Compound 1 (92g, 0.5mol) is dissolved in methanol (200mL), and joins reaction Mixture reacts 45 minutes, keeps the temperature of reactant mixture less than 50 DEG C simultaneously, be then stirred at room temperature overnight, thin layer Chromatograph (PE/EA=3:1) monitoring is to having reacted.Reactant mixture is cooled to 0 DEG C, and precipitate filters, and obtains white after drying Color Purify compound 2 (155.9g, yield: 96%).
2nd step: compound 2 (155.9g, 0.441mol) (1.1L) in Dowtherm A heat pipe is heated to 100 DEG C, then it is slowly added to connect in the flask having Dowtherm A heat pipe in (500mL is preheating to 210 DEG C), keeps simultaneously Temperature is higher than 207 DEG C.Reactant stirs reaction one hour at 210 DEG C, is then cooled to room temperature.The precipitation being collected by filtration, After washing with ether, acetone, obtain the Purify compound 3 (67g, yield: 61%) of brown after drying.
3rd step: at room temperature, with stirring and toward phosphorus oxychloride P DEG C l3 (35mL) adds compound 3 (10g, 0.0398mol).After interpolation, reactant mixture return stirring 3 hours, thin layer chromatography (dichloromethane DCM/ methanol=10:1) Tracking reaction completes, then reactant mixture injects frozen water, and pH=8 is adjusted in sodium carbonate alkalization, then extracts with ethyl acetate, collects Organic facies, washs with saline, and dried with Na2SO4, crude product is again by silica gel column chromatography (petroleum ether PE/ ethyl acetate EA= 20:1,1:1), obtain faint yellow solid pure compound 4 (6.0g, yield: 56%).
4th step: by compound 4 (10.0g, 37mmol), zinc powder (0.36g, 5.55mmol), Zn (CN) 2 (2.69g, 22.9Zn), dppf (8.82g, 15.9mmol) and Pd2 (dba) 3 (7.8g, 8.51mmol) is mixed in DMA (50mL), 80 DEG C It is stirred overnight.Thin layer chromatography (PE/EA=5:1) tracks to almost all of compound 4 and is consumed.Reactant mixture is cooled to During room temperature is fallen back, filtering, filtrate is extracted with ethyl acetate, collects organic facies, washs with saline, is dried dense on na 2 so 4 Contracting obtains crude product, and silica gel chromatography (PE/EA=50:1,10:1) obtains faint yellow solid pure compound 5, and (6.0g produces Amount: 62.3%).
5th step: mixing cpd 5 (6.0g, 23.08mmol), sodium hydroxide (10g, 250mmol) are at ethylene glycol (70mL) Stir 3 hours at cohobation device with water (10mL).Thin layer chromatography (DCM/ methanol=10:1) tracks to reaction and completes.Will reaction Mixture is cooled to room temperature, adds aqueous citric acid solution and is acidified to pH=4, filters, and precipitation obtains faint yellow solid after drying Pure compound 6 (6.0g, yield: 93.2%).
6th step: stirring mixing compound 6 (6.0g, 21.5mmol), in methanol (100mL), is added dropwise at 0 DEG C SOCl2(4.7mL,64.5mmol).After interpolation, reactant mixture is stirred at room temperature 1 hour, then heated overnight at reflux. Thin layer chromatography (DCM/ methanol=10:1) tracks to reaction and completes.Reactant mixture evaporation is removed major part solvent, residue Inject frozen water, add solid sodium carbonate regulation pH=8, and extract by ethyl acetate, collect organic facies, after washing with saline, Na2SO4 is upper to be dried, and concentrates and obtains bright-yellow solid compound 7 (3.9g, yield: 61.9%).
7th step: mixing compound 7 (3.9g, 13.3mmol) and 10%Pd/C (1.0g) are in methanol (150mL), in room The lower H2 gas of temperature stirs 2 hours.Thin layer chromatography (PE/EA=2:1) tracks to reaction and completes.Reactant mixture is filtered, filtrate The solid chemical compound 8 (2.6g, yield: 96.3%) of brown is obtained after concentration.
8th step: stirring mixing compound 8 (0.5g, 2.46mmol), in methanol (5mL), adds 2N NaOH under room temperature (2.5mL,5mmol.After interpolation, reactant mixture is stirred at room temperature night.Thin layer chromatography (DCM/ methanol=10:1) Indicator Reaction Complete.Being evaporated by reactant mixture, residue diluted with water, and extract by ethyl acetate, discard ethyl acetate layer, aqueous phase adds Aqueous citric acid solution also regulates pH=2, filters, and precipitation obtains pure compound 9 (0.4g, yield: 86%) after drying.
9th step: mixing compound 9 (0.4g, 2.11mmol), 3-5-trifluoromethylaniline (375mg, 2.32mmol), HATU (960mg, 2.53mmol) and DIPEA (820mg, 6.33mmol) are stirred overnight under DMF (5mL) room temperature in N2.Thin layer color Spectrum (DCM/ methanol=10:1) display reaction completes.Ethyl acetate after reactant mixture dilute with water is extracted, collects organic facies, Wash with saline, be dried on na 2 so 4 after concentrating and obtain crude product, pure by the solid obtaining yellow after silica gel chromatography Compound 10 (0.4g, yield: 57%).
The preparation of embodiment 2 compound V01 (also can be designated as V01 in the present invention)
Step 1:
Compound 11A (20g, 0.15mol), DMAP (1.9g, 15.4mmol) and (BoC)2O (75g, 0.34mol), four Mixing in hydrogen furan (750mL), reactant mixture is stirred at room temperature overnight.Thin layer chromatography (PE/EA=3:1) display has been reacted Become.It is suspended in PE/EA (10:1,200mL) after being concentrated by reactant mixture, after filtration, obtains white solid pure compound 11 (50g, 100%).
Step 2:
Mixing cpd 10 (50mg, 0.15mmol) and Cs2CO3 (150mg, 0.45mmol) are in DMSO (1mL), at N2 In half an hour, is stirred at room temperature, be subsequently adding compound 11 (60mg).Consequent reactant mixture stirring half an hour, use Thin layer chromatography (DCM/ methanol=10:1) detection is to having reacted.After reactant mixture dilute, extracting with EA, collection has Machine phase, washs with saline, dried with Na2SO4, prepares the Purify compound 12 (30mg, 32%) of yellow after chromatogram purification.
Under room temperature, N2 stirs mixing cpd 12 (20mg, 0.032mmol) and TFA (0.1mL) half an hour.Thin layer color Spectrum (DCM/ methanol=10:1) display is to having reacted.After being alkalized by Na2CO3 by reactant mixture, extracting with DCM, collection has Machine phase, washs with saline, to be dried on Na2SO4, obtains crude product after concentration, by obtaining yellow after silica gel chromatography Purify compound V01 (4.5mg, 33%), Structural Identification sees Fig. 8.
The preparation of embodiment 3 compound V3 (also can be designated as V03 in the present invention)
Step 1:
Stirring mixing compound 13 (5.0g, 45mmol) and pyridine (200mL), by (Boc) at 65 DEG C2O(14.7g, 67.5mmol) it is added drop-wise in said mixture.After interpolation, 85 DEG C of stirring reactions 4 hours.Thin layer chromatography (DCM/ methanol=10: 1) track to after reaction completes, reactant mixture be cooled to 0 DEG C, adds dense HCl (100mL) and H2O (50mL), EA and extract After, collect organic facies, use NaHCO3 solution washing, dried with Na2SO4, obtain the grease of yellow, be suspended in Et2O, filters, it is thus achieved that compound as white solid 14 (4.3g, yield: 45.2%).
Stirring mixing compound 14 (2.4g, 11.4mmol) and N, N dimethyl aniline (6.6mL) in DCM (84mL), 0 In DEG C N2, drip POCl3(3.2mL,34.2mmol).After interpolation, reactant mixture is stirred at room temperature 2 hours.Thin layer chromatography (PE/EA=2:1) track to after reaction completes, reactant mixture be injected frozen water.Organic facies salt is washed, and does on na 2 so 4 Crude product is obtained, by silica gel chromatography, it is thus achieved that white solid pure compound 15 (1.8g, 70%) after dry concentration.
Stirring mixing compound 15 (100mg, 0.47mmol) and DMAP (12mg, 0.09mmol), room in THF (1mL) Temperature is lower adds (BOC)2O (124mg, 0.57mmol), is then stirred overnight under room temperature.Thin layer chromatography (PE/EA=2:1) shows instead After should completing, by reactant mixture dilute with water, EA extracts, and collects organic facies, washes with salt, is dried on na 2 so 4 after concentrating Obtain crude product, by silica gel chromatography, it is thus achieved that white solid pure compound 16 (70mg, 44.8%).
Under room temperature, at N2Stirring mixing compound 10 (50mg, 0.15mmol) and Cs in middle DMSO (1mL)2CO3(150mg, 0.45mmol) half an hour, it is subsequently adding compound 16 (60mg, 0.18mmol), is stirred for half an hour.Thin layer chromatography (PE/EA= After 2:1) showing that reaction completes, by reactant mixture dilute with water, EA extracts, and collects organic facies, washes with salt, on na 2 so 4 It is dried after concentrating and obtains crude product, by silica gel chromatography, it is thus achieved that yellow solid pure compound 17 (50mg, 53.3%).
Step 2:
Under room temperature, in N2, in DMSO (1mL), stirring mixes compound 17 (50mg, 0.08mmol) and TFA (0.2mL) half Hour.After thin layer chromatography (DCM/ methanol=10:1) shows that reaction completes, after this reactant mixture is alkalized by Na2CO3, use DCM extracts.Organic facies is with brine wash, is dried on na 2 so 4 after concentrating and obtains crude product, is obtained by after silica gel chromatography The Purify compound V3 (15mg, 44%) of yellow, its Structural Identification data see Fig. 9.
The preparation of embodiment 3 compound V4 (also can be designated as V04 in the present invention)
Step 1:
Stirring mixing compound 18 (50g, 0.47mol), in DCM (600mL), adds TEA (94g, 0.93mol), then At 0 DEG C, in N2, dropping bromoacetate (94g, 0.56mol).Reactant mixture is stirred at room temperature overnight.Thin layer chromatography (PE/EA=1:1) after showing that reaction completes, reactant mixture salt is washed, and is dried on na 2 so 4 after concentrating and obtains crude product, By silica gel chromatography (using PE/EA=20:1to10:1to5:1 eluting), it is thus achieved that yellow oily pure compound 19 (46g, 51%).
At 95 DEG C, toluene (800mL) stirring mixing compound 19 (38g, 196.65mmol) and TEA (29.9g, 295mmol), 4-bromobutyrate (72.9g, 373.65mmol) it is added dropwise over. add rear heated at reflux overnight.Thin layer chromatography (PE/EA=5:1), after showing that reaction completes, reactant mixture concentrates, by silica gel chromatography (with PE/EA=20:1to5: Eluting), it is thus achieved that yellow oily pure compound 20 (32g, yield:53%).
Stirring mixing compound 20 (32g, 104.3mmol) in toluene (800mL), at 0 DEG C, adds t-BuOK (51.2g,456.3mmol).After interpolation, reactant mixture at room temperature reacts 1 hour.Thin layer chromatography (PE/EA=5:1) shows After having reacted, adding 2N HCl and regulate pH=6, then EA extracts, and collects organic facies, washes with salt, is dried on na 2 so 4 Obtain crude product 21 (17g, yield:62.5%) after concentration, for dark yellow oil, can be directly used for next step reaction, be not required to into one Step purification.
((11g, 161.65mmol) is dissolved in methanol (280mL) to MeONa, and reactant mixture is cooled to 5 DEG C, adds second Acid carbonamidine (3.0g, 29.15mmol).Stirring reactant mixture half an hour, add compound 21 (17g, 65.1mmol).Will Reactant mixture 40 DEG C is stirred overnight.After thin layer chromatography (DCM/ methanol=10:1) shows that reaction completes, reactant mixture is cold But arriving room temperature, evaporation removes most solvent.Residue water processes and EA extracts, and organic facies salt is washed, on na 2 so 4 Be dried after concentrating and obtain crude product, by silica gel chromatography obtain faint yellow solid pure compound 22 (2.5g, yield: 16%).
It is 0.5MPa H at pressure2In, it is stirred overnight mixing compound 22 (2.5g, 10.4mmol), 10%Pd/C (0.5g) with (B DEG C)2O (2.7g, 12.4mmol) is in MeOH (40mL).Thin layer chromatography (DCM/ methanol=10:1) shows reaction After completing, reactant mixture filtering and concentrating is obtained yellow solid pure compound 23 (2.6g, 100%).
Mixing compound 23 (1.6g, 6.3mmol), PPh in 1,2-DCE (64mL)3(3.33g, 12.6mmol) and CCl4 (2.93g, 18.9mmol), 70 DEG C are heated 1 hour.After thin layer chromatography (DCM/ methanol=10:1) shows that reaction completes, will reaction After mixture concentrates, silica gel chromatography obtain faint yellow solid pure compound 24 (1.38g, 81%).
In room temperature N2, stirring mixing compound 10 (100mg, 0.3mmol) and Cs in DMSO (2mL)2CO3(300mg, 0.9mmol) half an hour, add compound 24 (97mg, 0.36mmol).Reactant mixture stirring mixes half an hour.Thin layer After chromatograph (DCM/MeOH=10:1) shows that reaction completes, reactant dilute with water, then EA extracts.Organic facies salt is washed, It is dried on na 2 so 4 after concentrating and obtains crude product, after being concentrated by reactant mixture, obtained yellow solid by silica gel chromatography Pure compound 25 (21mg, 12.4%).
Step 2:
In room temperature N2, middle stirring mixing compound 25 (21mg, 0.037mmol) and TFA (0.2mL) half an hour.Thin layer Chromatograph (DCM/MeOH=10:1) shows that reaction completes.After reactant mixture is alkalized by Na2CO3, extract with DCM.Organic facies Wash with salt, be dried on na 2 so 4 after concentrating and obtain crude product, by the Purify obtaining yellow after silica gel chromatography Compound V4 (10mg, 57.9%), its Structural Identification data see Figure 10.
The preparation of embodiment 4 compound V5 (also can be designated as V05 in the present invention)
Step 1:
(S)-1-phenylethylamine (10g, 8.26mmol) and the 1,2-DCE of ethyl propanoic acid (11.9g, 8.26mmol) (200mL), in solution, it is dividedly in some parts NaBH (OAc)3(35g,16.52mmol).After adding, reactant mixture is at room temperature stirred Mix 4 hours.Then reactant mixture is added NaHCO3Aqueous solution alkalizes, and extracts with DCM, separates organic facies, and with saturated Brine It, is dried on na 2 so 4, and is concentrated to give thick compound 34 (20.5g, yield: 100%), can be without entering one Step purification is directly used in next step.
In the 1,2-DCE (200mL) stirring mixing compound 34 (20.5g, 82mmol) and glyoxylic acid ethyl ester ((33mL, 165mmol, the toluene solution of 50%), NaBH (OAc)3(35g, 165mmol), is stirred at room temperature 4 hours.Then will reaction Mixture NaHCO3Aqueous solution alkalizes, and extracts with DCM.Separate organic facies and wash with saturated aqueous common salt, doing at Na2SO4 Dry and be concentrated to give thick compound 35 (26g, 100%), can be directly used in without further purification in next step.
Compound 35 (26g, 78mmol) and the t-BuOK (22g, 156mmol) mixing in toluene (600mL), stir back Flow through night.Then by reactant mixture NaHCO3Aqueous solution alkalization also extracts with DCM.Organic phases washed with brine, Na2SO4 is dried and concentrates, and obtains crude compound 36.
Take crude compound 36 and pass through silica gel chromatography (PE/EA=20:1 eluting) purification, obtain pure compound 37 (3g).
Mixing cpd 37 (3.0g, 0.01mol) in ethanol (50mL) and formamidine acetate (3.24g, 0.03mol), mixture adds NaOEt (2.38g, 0.035mol) in solution.Stir this reactant mixture in 90 DEG C overnight, Then evaporate to remove most solvent.Residue H2O dilutes, and being acidified to pH with 2N HCl is 6, then extracts with DCM. Organic phases washed with brine, is dried at Na2SO4 and concentrates, obtain crude product, passed through Silica gel chromatography, obtains pure Compound 38 (1.0g, yield: 35.8%), for yellow solid.
By compound 38 (1.0g, 10.4mmol), the Pd/C (0.1g) of 10% and (B DEG C)2O (2.7g, 12.4mmol) mixes Together in MeOH (40mL), it is stirred overnight in the hydrogen that pressure is 0.5 MPa.TLC (DCM/ methanol=10:1) shows reaction Complete.Reactant mixture is filtered, filtrate is concentrated, obtains crude product, passed through Silica gel chromatography, obtain pure change Compound 39 (0.7g, productivity: 71%), for yellow solid.
Take compound 39 (0.7g, 6.3mmol) and triphenylphosphine PPh3(3.33g, 12.6mmol) is at carbon tetrachloride (10mL) mixing in, in 80 DEG C of stirred overnight.TLC (DCM/ methanol=15:1) shows that reaction completes.Reactant mixture is concentrated, And by residue by Silica gel chromatography, obtain pure compound 40 (0.4g, 53.4%), for light yellow oil.
Under a nitrogen, compound 10 (150mg, 0.45mmol) and Cs2CO3(440mg, 1.35mmol) is in DMSO (2ml) Mixing, is stirred at room temperature half an hour, is subsequently adding compound 40 (153mg, 0.54mmol).The reactant mixture of gained is stirred Mixing half an hour, TLC (DCM/ methanol=10:1) shows that reaction completes.By reactant mixture dilute with water and extract with EA.Organic Wash with saline, at Na2SO4It is dried and concentrates, obtain crude product, passed through the TLC purification of preparation, obtain pure chemical combination Thing 41 (60mg, 23%), for yellow solid.
Step 2:
Take the mixture of compound 41 (60mg, 0.104mmol) and TFA (0.3mL) under a nitrogen, be stirred at room temperature half little Time.TLC (DCM/ methanol=10:1) shows that reaction completes.By reactant mixture NaHCO3Aqueous solution alkalization also extracts with DCM.Have Machine saline washs, at Na2SO4Upper being dried concentrates, and obtains crude product, is passed through the TLC purification of preparation, obtains purification and closes Thing V5 (20mg, 40.3%), for yellow solid, its Structural Identification data see Figure 11,12.
The preparation of embodiment 5 compound V6 (also can be designated as V06 in the present invention)
Step 1:
In flask equipped with mechanical agitator and the 3L of calcium chloride tube, load compound 26 (100g, 0.716mol) and Ethanol (1.0L).This mixture is stirred 20 minutes, then drips triethylamine (72.5g, 0.716mol).By the mixture of gained Stir 10 minutes, then, add ethyl acrylate (61.6g, 0.716mol).Reactant mixture is stirred at room temperature 17 little Time.(BOC)2O (234.5g, 1.08mol) is at room temperature added dropwise over.After completion of dropwise addition, reactant mixture is at room temperature stirred Mix overnight.Concentrate reactant mixture to remove in major part EtOH.Residue is dissolved in water (3L), and with Et 2O (1L × 3) extraction.The organic facies merged ammonium chloride (500L × 3) aqueous solution and saline (500L × 3) wash, and anhydrous Na 2SO4 is done Dry, and be concentrated to give thick compound 28 (300g, 92%), for yellow oil, its for next step without extra Purification.
Sodium (27.6g, 0.765mol) adds in dehydrated alcohol (1.5L).When solid sodium is wholly absent, by compound 28 (300g, 1.04mol) adds in hot solution.Reactant mixture refluxes overnight.Thin layer chromatography (petrol ether/ethyl acetate=4: 1) show that initiation material is consumed completely.Reactant mixture is evaporated, residue is dissolved in water (1L), then citric acid water It is 6 that solution is acidified to pH.Then mixture EtOAc (1 liter × 3) is extracted.Extract is merged, and with saline (1L × 3) Washing, at Na2SO4It is dried and evaporates, obtaining compound 29 (169g, 63.4%), for brown oil.This crude product is without entering One step purification is used for next step.
Feldalat NM (2.6g, 48.58mmol) is dissolved in MeOH (50mL), and this solution is cooled to 5 DEG C.Add acetic acid Carbonamidine (3.0g, 29.15mmol), stirs half an hour by reactant mixture.Add compound 29 (5.0g, 19.43mmol), will Reactant mixture is stirred at reflux overnight.TLC (DCM/ methanol=10:1) shows that reaction completes.Reactant mixture is cooled to room Temperature, and evaporate to remove major part solvent.Residue with water is processed and extracts with EA.Organic phases washed with brine, at Na2SO4 Be dried and concentrate, obtain crude product, passed through Silica gel chromatography, obtain pure compound 30 (680mg, yield: 14.7%), for light yellow solid.
Under 0 DEG C of nitrogen, phosphorus oxychloride (P DEG C of l3,174mg, 1.26mmol) joins stirring and is mixed with compound 30 In the DCM (4mL) of (100mg, 0.42mmol) and DMA (0.28mL), after adding, reactant mixture is poured into In frozen water, alkalize by adding solid sodium carbonate, then extract with DCM.Organic phases washed with brine, at Na2SO4It is dried and dense Contracting, obtains crude product, by it by preparation TLC purification, obtains pure compound 31 (60mg, 55.6%), for white solid.
Under a nitrogen, compound 10 (50mg, 0.15mmol) and Cs2CO3(150mg, 0.45mmol) is mixed in DMSO (1mL) in, mixture is stirred at room temperature half an hour, is subsequently adding compound 31 (46mg, 0.18mmol).By the reaction of gained Mixture stirs half an hour, and TLC (DCM/ methanol=10:1) shows that reaction completes.By reactant mixture dilute with water and extract with EA Take.Organic phases washed with brine, is dried at Na2SO4 and concentrates, obtain crude product, is passed through the TLC purification of preparation, obtains Pure compound 32 (13mg, 15.7%), for yellow solid.
Step 2:
Under nitrogen, stir half an hour under the mixture room temperature of compound 32 (13mg, 0.038mmol) and TFA (0.1mL). TLC (DCM/ methanol=10:1) shows that reaction completes.Reaction the mixture alkalization of Na2CO3 aqueous solution and DCM are extracted.Have Machine saline washs, and is dried at Na2SO4 and concentrates, obtains crude product, by it by preparation TLC purification, obtains pure chemical combination Thing, V6 (6mg, yield: 56.3%), for yellow solid, its Structural Identification data see Figure 13.
The preparation method of embodiment 6 KDR3
1st step: by compound 1 (1.4 grams, 0.01mmol), 4-(benzyloxymethyl)-6-chloropyrimide (4.7 grams, 0.02) and K2CO3 (4.2 grams, 0.03 mole) is mixing in the DMF (50mL), 100 DEG C, under N2 stirring be 5 hours.TLC (PE/EA=4: 1) show that reaction completes.Reactant mixture is cooled to room temperature dilute with water, extracts with EA.Collect organic facies, wash with salt Wash, be dried with Na2SO4 and concentrate, obtain crude product, passed through column chromatography purification, obtain Rate: 90%), for yellow solid.
Step 2: under N2 atmosphere, compound 2 (1.2 grams, 3.56 mMs), hydrazine hydrate (0.36 gram, 7.12 mMs) With the mixture return stirring 1 hour in the THF (20mL) of Raney Ni (0.2g).TLC (PE/EA=2:1) shows to have reacted Become.Reactant mixture is filtered, filtering and concentrating, obtain crude product, passed through column chromatography purification, obtain pure compound 3 (0.8 gram, productivity: 73%), for yellow solid.
Step 3: at 0 DEG C under a nitrogen, exists TCDI (0.78 gram, 4.4mmol) and compound 3 (0.9 gram, 2.93mmol) The DCM (10mL) being dried is slowly mixed together.After adding, reactant mixture is stirred at room temperature half an hour.TLC (PE/EA=1: 1) show reactant mixture completes.Reactant mixture is concentrated, obtains crude product, purify with flash chromatography, obtain Crude compound 4 (1.0 grams, productivity: 100%), for pale solid.
Step 4: by compound 4 (1.1 grams, 3.15 mMs), 4-methyl-5-(trifluoromethyl) benzene 1,2-diamidogen (0.6 Gram, 3.15 mMs) and carbodiimide (1.2 grams, 6.30 mMs) THF (50 milliliters) under a nitrogen, stirred at reflux 2 is little Time.TLC (DCM/ methanol=10:1) shows, completes in reactant mixture.Dilute this reactant mixture with EA, and filter precipitation Thing.Filtrate is evaporated, obtains crude product, used column chromatography (PE/EA=10:1 to 4:1) purification, obtain pure compound 5 (0.8 gram, productivity: 50%), for brown oil.
Step 5: the mixture in compound 5 (0.7 gram, 1.38 mMs) and TFA (15ml) is in 60 DEG C of stirred overnight.Will Reactant mixture is warming up to 70 DEG C, and stirring 1 hour at this temperature.TLC (DCM/ methanol=10:1) shows, reactant mixture In complete.Reactant mixture being cooled to room temperature and is poured in frozen water, alkalizing to pH with the NaOH of 2N is 10, then extracts with EA Take.Organic phases washed with brine, is dried with Na2SO4 and concentrates, and obtains the crude product (0.4g, yield: 70%) of compound 6, for Yellow oil, it is directly used in next step without further purification.
Step 6: in the DCM (5ml) of the compound 6 (60mg, every day 0.1445mmol) that stirred, add TEA (30 millis Gram, 0.289mmol) and MSCL (25 milligrams, 0.2168mmol) under the protection of 0 DEG C of nitrogen.After completion of dropwise addition, reaction terminates.Will This mixture DCM dilutes, and washs with saline, is dried at Na2SO4 and concentrates, obtain the crude product of compound 7, and it is without entering One step purification is directly used in next step.
7th step: compound 7 (60 milligrams, 0.12 mM) and MeNH2 (0.6 milliliter, 1.2 mMs, in MeOH The mixture in anhydrous THF (5ml) 2N) is in stirred overnight at room temperature.TLC (DCM/ methanol=10:1) shows, reactant mixture In complete.Reactant mixture is concentrated and purification residue, TLC purification three times, obtains pure KDR3 (6 milligrams, productivity: 12%), For white solid, its Structural Identification data see Figure 14.
Prepared by embodiment 7 KDR4 compound
The first step: by compound 1 (1.0 grams, 4.5 mMs) mixing 2-chloro-5-nitropyridine 2-in ACN (20 milliliters) Chloro-5-nitropyridine (0.8 gram, 5 mMs) and cesium carbonate (3.3 grams, 10 mMs), be stirred at room temperature overnight.TLC(PE/ EA=1:1) show reactant mixture completes.By reactant mixture dilute with water, extract with EA.Organic phases washed with brine, Be dried at Na2SO4 and concentrate, obtain crude product, passed through purification by flash chromatography, obtain pure compound 2 (0.4 gram, receive Rate: 24.5%), for yellow solid.
Second step: add in Ruan in the stirring mixture in the THF (5mL) of compound 2 (0.3 gram, 0.83 mM) Nickel (0.1 gram), is then heated to backflow.Solution in the THF (5mL) of H2N-NH2H2O (1.66 mMs) is added dropwise over.Will The reactant mixture of gained stirs 10 minutes.TLC (DCM/ methanol=15:1) shows, completes in reactant mixture.Reaction is mixed . compound filters, and filtering and concentrating obtains crude product, pass through purification by flash chromatography, obtain pure compound 3 (0.16 gram, product Rate: 50%), for yellow solid.
3rd step: in the agitating solution of compound 3 (0.16 gram, 0.483mmol), under 0 DEG C of nitrogen protection, at DCM (10mL) TCDI (95mg, 0.531mmol) is added in.After completion of dropwise addition, reactant mixture is stirred at room temperature.Overnight.TLC (DCM/ methanol=10:1) shows, completes in reactant mixture.Reactant mixture is concentrated, obtains crude product, passed through fast Speed chromatogram purification, obtaining pure compound 4 (50 milligrams, productivity: 28%) is pale solid.
4th step: by compound 4 (50 milligrams, 0.134mmol), 4-methyl-5-(trifluoromethyl)-benzene-1,2-diamidogen , in anhydrous THF (10mL), refluxed under nitrogen is stirred for (25.5mg, 0.134 mM) and EDCI (51 Micks, 0.268 mM) Mix 1 hour.TLC (DCM/MEOH=15:1) shows, completes in reactant mixture.Diluted reaction mixture is heavy with EA and filtration Shallow lake thing, obtains crude product, by it by pre-TLC purification, obtains pure compound 5 (40 milligrams, yield: 56%), for white solid.
5th step: compound 5 (20 milligrams, 0.038 mM) and the mixture of trifluoroacetic acid (0.2 milliliter), in room temperature Lower stirring half an hour.TLC (DCM/ methanol=5:1) shows to complete in reactant mixture.By reactant mixture evaporation to remove TFA, is dissolved in residue in THF, solid K2CO3 alkalize, and filters and concentrated filtrate, obtains pure compound KDR4 (10 millis Gram, productivity: 62.5%), for white solid, its Structural Identification data see Figure 15.
Prepared by embodiment 10 KDR6 compound
The first step: 7-methoxy quinoline-4-alcohol (10.0 grams, 57.1mmol) is dissolved in 50ml phosphorus oxychloride, then will be anti- Mixture is answered to heat 3 hours at 120 DEG C.Then mixture is cooled to room temperature, and pours frozen water (50 milliliters) into.Extract with EtOAc Take this mixture (100mLx 3), organic layer saturated aqueous common salt is washed, is dried with anhydrous sodium sulfate and concentrates, slightly being produced Thing, is used flash-chromatography (with PE/EA=30:1 eluting) purification, and the product obtained (8.5 grams, yield: 76.9%), in vain Color solid.
Second step: 4-chloro-7-methoxy quinoline (5.5G, 28.4mmol), zinc (276.9mg, 4.26mmol), Zn (CN) 2 (2.07 grams, 17.6mmol), DPPF (6.65 grams, 12.21mmol), PD2 (DBA) 3 (5.98 grams, 6.53mmol) is dissolved in In the DMAc of 100mL, mixture argon covered and heats at 160 DEG C overnight, then mixture being cooled to room temperature, and leads to Cross kieselguhr to filter, by filtrate with H2O process, then with EA extraction, organic layer saturated aqueous common salt is washed, use anhydrous slufuric acid Sodium is dried, and is concentrated under vacuum.By flash-chromatography (with PE/EA=10:1 eluting) purification, obtain product 7-methoxyl group Quinoline-4-formonitrile HCN (4.38 grams, yield: 83.7%), for brown solid.
3rd step: 7-methoxy quinoline-4-formonitrile HCN (4.38 grams, 23.78mmol) is dissolved in the H2O of 30mL, adds hydrogen Sodium oxide (2.38 grams, 59.45mmol).Then 100 DEG C are heated the mixture to overnight.Reactant mixture is cooled to room temperature, And extract with EA, to remove impurity.Aqueous phase is acidified, is 2 by adding 6N hydrochloric acid to pH.By the precipitate of yellow by filtering Collecting and be dried, obtain the 7-quininic acid of 2.5g, productivity 52%, for brown solid.
4th step: 7-quininic acid (2.0 grams, 9.84mmol) and DIPEA (3.82 grams, 29.52mmol) are molten Solution is in DMF.Then reactant mixture is cooled to 0 DEG C, adds HATU (4.49 grams, 11.81mmol) and 3-(trifluoromethyl) Aniline (1.74 grams, 10.82mmol).This mixture argon covers, and is then stirred at room temperature overnight.This mixture water is dilute Release and use EtOAc (3 × 50 milliliters) to extract.Washing organic layer, washs with saline, is dried with anhydrous sodium sulfate, and under vacuo Concentrate, obtain crude product, passed through purification by flash chromatography, and PE/EA=1:1 eluting, obtain product (2.5g, yield: 79.6%) it is light yellow solid.
5th step: 7-methoxyl group-N-(3-(trifluoromethyl) phenyl) quinoline-4-Methanamide (0.6 gram, 1.733mmol) in In DCM (50 milliliters) solution, 10 DEG C of drop wise under nitrogen add BBr3 (1.73mL, 6.93 mMs.After adding, reaction is mixed Thing is stirred at room temperature 100 hours.TLC (DCM/ methanol=15:1) shows that reaction completes.Reactant mixture is poured in ice-water And extract with DCM.Washing organic layer, with saline wash, anhydrous Na 2SO4 is dried, the product of concentration, for brown solid (0.5 gram, Productivity: 87%), use it for next step, it is not necessary to be further purified.
6th step: by 7-hydroxy-n-(3-(trifluoromethyl) phenyl) quinoline-4-Methanamide (50 milligrams, 0.151mmol), The tert-butyl group (6-chloropyrimide-4-base) methyl (methyl) t-butyl carbamate (46.6mg, 0.181mmol) and K2CO3 (41.7mg, 0.302mmol) mixes in DMSO (2ml) solution, is stirred overnight at 90 DEG C.TLC (PE/EA=1:2) shows instead Should complete.By reactant mixture dilute with water and extract with EA.Washing organic layer, washs with saline, and anhydrous Na 2SO4 is dried, dense Contracting, crude product, this is by pre--TLC purification, obtains pure product (25 milligrams, productivity: 30%), for white solid.
7th step: ((6-(4-(3-(trifluoromethyl) phenylcarbamoyl) quinoline-7-base epoxide) is phonetic for tertbutyl methyl Pyridine-4-base) methyl) t-butyl carbamate (25 milligrams, 0.045mmol) and the mixture of TFA (0.25 milliliter), at 0 DEG C Stir half an hour.TLC (DCM/ methanol=5:1) shows that reaction completes.Evaporation TFA, and residue is alkalized by aqueous solution. Na2CO3 and extracting with DCM.Washing organic facies, washs with saline, is dried with Na2SO4 and concentrates, obtaining crude product, being led to Crossing pre--TLC purification, obtain pure compound 06 (7 milligrams, yield: 21%), for white solid, its Structural Identification data see Figure 16.
Prepared by embodiment 11 KDR6A compound
The first step: 7-methoxy quinoline-4-alcohol (10.0 grams, 57.1mmol) is dissolved in 50ml phosphorus oxychloride, then will be anti- Mixture is answered to heat 3 hours at 120 DEG C.Then mixture is cooled to room temperature, and pours frozen water (50 milliliters) into.Extract with EtOAc Take this mixture (100mLx 3), organic layer saturated aqueous common salt is washed, is dried with anhydrous sodium sulfate and concentrates, slightly being produced Thing, is used flash-chromatography (with PE/EA=30:1 eluting) purification, and the product obtained (8.5 grams, yield: 76.9%), in vain Color solid.
Second step: 4-chloro-7-methoxy quinoline (5.5G, 28.4mmol), zinc (276.9mg, 4.26mmol), zinc (CN) 2 (2.07 grams, 17.6mmol), DPPF (6.65 grams, 12.21mmol), PD2 (DBA) 3 (5.98 grams, 6.53mmol) is dissolved in In the DMAc of 100mL, mixture argon covered and heats at 160 DEG C overnight, then mixture being cooled to room temperature, and leads to Cross kieselguhr to filter, by filtrate with H2O process, then with EA extraction, organic layer saturated aqueous common salt is washed, use anhydrous slufuric acid Sodium is dried, and is concentrated under vacuum.By flash-chromatography (with PE/EA=10:1 eluting) purification, obtain product 7-methoxyl group Quinoline-4-formonitrile HCN (4.38 grams, yield: 83.7%), for brown solid.
3rd step: 7-methoxy quinoline-4-formonitrile HCN (4.38 grams, 23.78mmol) is dissolved in the H2O of 30mL, adds hydrogen Sodium oxide (2.38 grams, 59.45mmol).Then 100 DEG C are heated the mixture to overnight.Reactant mixture is cooled to room temperature, And extract with EA, to remove impurity.Aqueous phase is acidified, is 2 by adding 6N hydrochloric acid to pH.By the precipitate of yellow by filtering Collecting and be dried, obtain the 7-quininic acid of 2.5g, productivity 52%, for brown solid.
4th step: 7-quininic acid (406mg, 2 mMs) and DIPEA (775mg, 6 mMs) are dissolved in In DMF (5mL).Reactant mixture is cooled to 5 DEG C, is subsequently adding HATU (912mg, 2.4mmol) and 4-fluoro-3-(fluoroform Base) aniline (394mg, 2.2 mMs).This mixture uses argon and covering at ambient temperature, overnight.TLC (PE/EA=1: 2) show that reaction completes.By reactant mixture dilute with water and extract with EA.Washing organic layer, washs with saline, anhydrous Na2SO4 is dried, and concentrates, crude product, by it by CCTO purification, obtains pure product (220 milligrams, yield: 30%), for palm fibre Color solid.
5th step: N-(4-fluoro-3-(trifluoromethyl) phenyl)-7-methoxy quinoline-4-Methanamide (0.2 gram, In DCM (10ml) solution 0.549mmol), at 0 DEG C, N2 stirred under argon adds 4N BBr3's (0.68 milliliter, 2.75mmol) DCM solution.After adding, being stirred at room temperature by reactant mixture is 20 hours.Then reactant mixture is stirred 4 at-30 DEG C Hour.TLC (DCM/ methanol=10:1) shows that initiation material is still.Reactant mixture is poured in ice-water and extracts with DCM. Organic layer saturated aqueous common salt is washed, is dried with anhydrous sodium sulfate and concentrates, obtaining crude product, by it by pre-TLC purification, The raw material (120 milligrams) arrived and pure product N-(the fluoro-3-of 4-(trifluoromethyl base) phenyl)-7-hydroxyquinoline-4-Methanamide (60 Milligram, 78%), for white solid.
6th step: by N-(4-fluoro-3-(trifluoromethyl) phenyl)-7-hydroxyquinoline-4-Methanamide (30 milligrams, 0.086mmol), the tert-butyl group (6-chloropyrimide-4-base) methyl (methyl) t-butyl carbamate (26.5mg, 0.103mmol) and K2CO3 (23.8mg, 0.172mmol), in DMF (1mL), stirs 15 hours at 50 DEG C.TLC (DCM/ methanol=10:1) shows Reaction completes.By reactant mixture dilute with water.And extract EA.Organic layer saturated aqueous common salt is washed, uses anhydrous sodium sulfate It is dried and concentrates, obtain crude product, passed through pre--TLC purification, obtain pure product (6-(4-(4-fluoro-3-(trifluoromethyl The tert-butyl group) phenylcarbamoyl) quinoline-7-base epoxide) pyrimidine-4-yl) methyl (methyl) t-butyl carbamate (20 millis Gram, yield: 41%), for yellow solid.
7th step: mixture (6-(4-(4-fluoro-3-(trifluoromethyl) phenylcarbamoyl) quinoline-7-of the first tert-butyl ester Base epoxide) pyrimidine-4-yl) methyl (methyl) t-butyl carbamate (20 milligrams, 0.035mmol) and trifluoroacetic acid (0.3 milli Rise) it is stirred at room temperature one hour.TLC (DCM/MEOH=5:1) shows that reaction completes.Evaporation TFA, and by residue by Na2CO3 alkalization also extracts with DCM.Washing organic facies, washs with saline, is dried with Na2SO4 and concentrates, obtaining crude product, will It, by pre--TLC purification, obtains pure compound 06A (5 milligrams, yield: 30%), for yellow solid, its Structural Identification data See Figure 17.
Prepared by embodiment 12 KDR8 compound
1st step: the mixture of ethyl oxalate (70 milliliters) is heated to 120 DEG C, adds compound 1 (50 grams, 0.4mol), And heat the mixture to 180 DEG C, maintain 5 minutes.By mixture refrigerated overnight, white depositions is collected by filtration, is dried, from And obtain the compound 2 (75.0 grams, yield: 59.11%) of white powder.
Step 2: compound 2 (25 grams, 0.11 mole) is dissolved in the dimethylbenzene (1L) of boiling, is slowly added into P2S5 (9.5 grams, 0.0385 mole), backflow, until having reacted (about 5 hours), it is further continued for backflow.Amide TLC (PE/EA=5/1) After showing that this reaction completes, being cooled down by reactant mixture, and extract with 1N NaOH, aqueous phase hydrochloric acid is acidified, and collects yellow mercury oxide Thing, is then dissolved in EtOAc, washs with H2O, is dried with Na2SO4, and concentrates, Rate: 70.9%) it is a kind of red oil.
Step 3: compound 3 (30 grams, 0.142 mole is dissolved in 1N sodium hydroxide (500mL), with the potassium ferricyanide (135 grams, 0.416mol, in 340ml water) oxidation, carry out to ferricyanide solution by being slowly added to thioamides, make temperature keep Less than 10 DEG C.After 1h, TLC (in DCM/MeOH=3/1) shows that reaction is completely.Reactant mixture is being diluted in water, Acidifying (being 6 by the HCl to pH of 2N), then filters, the compound 4 (15g, yield: 50.48%) obtained.
Step 4: compound 4 (1 gram, 4.34 mMs) stirring and dissolving in DMF (20mL), adds 1-methyl-5- (trifluoromethyl)-1H-pyrazol-3-amine (0.717g, 4.35mmol), is subsequently added and adds in room temperature under nitrogen Enter HATU (2.47g, 6.5mmol) and DIEA (1.12g, 8.6mmol).After being stirred overnight under at a temperature of identical, TLC (PE/EA=2/1) show that this reaction completes.This reactant mixture is poured in water, and extracts with EtOAc.Washing will organically Phase, washs with saline, is dried on na 2 so 4 and concentrates, and obtains thick product, by silica gel chromatography (PE/EA eluting=20/ 1-5/1) it is purified, obtain pure compound 5 (900 milligrams, yield: 75.5%).
In step 5:-10 DEG C nitrogen, in DCM (10mL) solution of compound 5 (0.4g, 1.12mmol), add BBr 3 (10mL,4N in DCM).After 4 hours, TLC (PE/EA=1/1) shows that reaction completes.Then reaction add ice, filters Remove undissolved material.Separating filtrate, extracts with DCM.By organic phases washed with brine, it is dried with Na2SO4 and concentrates, To thick product, this is purified, TLC obtain the compound (100mg, yield: 26.02%) of pure compound 6.
Step 6: in DMF (10mL) solution of the solution (50mg, 0.146mmol) of compound 6 add K2CO3 (70mg, 0.500mmol) with tert-butyl (6-chloropyrimidin-4-yl) methyl (methyl) carbamate (41.41mg,0.161mmol).By reactant mixture at 50 DEG C of stirred under nitrogen overnight.TLC (PE/EA=1/1) shows reaction Complete.Reactant mixture is washed with water, extracts with EA.Organic facies salt washs 4 times, is dried with Na2SO4 and concentrates, obtaining Thick product, is obtained pure compound 7 (40mg, yield: 48.59%) by TLC purification.7th step: compound 7 (40mg, 0.071mmol) with TFA (0.4mL) mixed at room temperature under a nitrogen, stir half an hour.TLC (DCM/ methanol=10/1) shows reaction Complete.Reactant mixture is evaporated to remove TFA.Residue is alkalized, uses Na2CO3Alkalization, to pH9, then extracts with DCM.Will Organic phases washed with brine, sodium sulfate is dried, and concentrates, obtains crude product, passed through TLC purification, obtain pure compound 8 (20mg, yield: 44%).Its Structural Identification data see Figure 18.
Prepared by embodiment 13 KDR8A compound
1st step: ethyl oxalate (70 milliliters) is heated to 120 DEG C, adds compound 1 (50 grams, 0.4mol), and will mixing Thing is heated to 180 DEG C, and heat time heating time is: 5 minutes.After cooling, mixture, in refrigerator overnight, adds 50 milliliters of water, then shapes Become white depositions.Collect this solid, be dried, obtain 2 (75.0 grams, yield: 59.11%) of the compound of white powder.
Step 2: compound 2 (25 grams, 0.11 mole) is dissolved in the dimethylbenzene (1L) of boiling, is slowly added to P2S5 (9.5 grams, 0.0385 mole), backflow, until having reacted (about 5 hours), it is further continued for backflow.TLC (PE/EA=5/1) represents After having reacted, being cooled down by reactant mixture, and extract with 1N NaOH, aqueous phase hydrochloric acid is acidified, and collects yellow mercury oxide, so After be dissolved in EtOAc, wash with H2O, be dried with Na2SO4, and concentrate, the compound that obtains 3 (19 grams, yield: 70.9%), for red oil.
Step 3: crude compound 3 (30g, 0.142mol) is dissolved in 1N sodium hydroxide (500mL), with the potassium ferricyanide (135 Gram, 0.416mol, in 340ml water) oxidation, carry out to ferricyanide solution by being slowly added to thioamides, make temperature Keep below 10 DEG C.After 1h, TLC (in DCM/MeOH=3/1) shows that reaction is completely.By reactant mixture in water dilute Release, the HCl of 2N regulating to pH is 6, then filters, the compound 4 (15g, yield: 50.48%) obtained
Step 4: compound 4 (1 gram, 4.78 mMs) adds 3-(trifluoromethyl) aniline in DMF (40 milliliters) solution, It is subsequently added HATU (2.73 grams, 7.17 mMs) and DIEA (1.24 grams, 9.56 mMs), stirs under nitrogen at room temperature.? It is stirred overnight after rear TLC (PE/EA=2/1) shows that reaction completes at a temperature of identical, reactant mixture is poured in water, and uses EtOAc extracts.Washing organic facies, washs with saline, is dried at Na2SO4 and concentrates, obtain crude product, passed through silica gel color Spectrum (with PE/EA=20/1-5/1 eluting) purification, obtains pure compound 5 (1.68 grams, yield: 99.0%).
Step 5:-10 DEG C under a nitrogen, adds 4N in DCM (10mL) solution of compound 5 (0.1 gram, 0.283 mM) BBr3(0.5mL).After 4 hours, after TLC (PE/EA=1/1) shows that reaction completes, add ice, then filter to remove described Undissolved material.Filtrate extracts with DCM.Wash organic facies with saline, be dried with Na2SO4 and concentrate, obtain crude product, will Its logical TLC purification, obtains pure compound 6 (64mg, yield: 66.65%).
Step 6: in compound 6 (64 milligrams, 0.189 mM) DMF (10 milliliters) solution, addition K2CO3 (78 milligrams, 0.567 mM) and the tert-butyl group (6-chloropyrimide-4-base) methyl (methyl) t-butyl carbamate (tert-butyl (6- Chloropyrimidin-4-yl) methyl (methyl) carbamate, 53 milligrams, 0.208 mM).By reactant mixture 50 DEG C are stirred overnight under a nitrogen, and TLC (PE/EA=1/1) shows that reaction completes.Being washed with water by reactant mixture, EA extracts. Washing organic facies, washs four times with saline, is dried with Na2SO4 and concentrates, obtaining crude product, is passed through pre--TLC purification, To pure compound 7 (20mg, yield: 19.68%).
7th step: compound 7 (20 milligrams, 0.035 mM) and the mixture of TFA (0.2mL), room temperature is stirred under a nitrogen Mix half an hour.TLC (in DCM/MeOH=10/1) show that reaction completes.Reactant mixture evaporation is removed TFA.By residue by SNa2CO3 regulation pH is 9, then extracts with DCM.Washing organic facies, washs with saline, is dried with Na2SO4 and concentrates, obtaining Crude product, is passed through pre--TLC purification, is obtained pure compound K DR08A (12mg, yield: 73.07%), its Structural Identification Data see Figure 19,22.
Prepared by embodiment 14 KDR8B compound
1st step: the mixture of ethyl oxalate (70 milliliters) is heated to 120 DEG C, adds compound 1 (50 grams, 0.4mol), And heat the mixture to 180 DEG C, maintain 5 minutes.By refrigerated overnight after mixture, form white depositions.Collect this solid In by filtering, and it is dried, thus obtains 2 (75.0 grams, yield: 59.11%) of the compound for white powder.
Step 2: compound 2 (25 grams, 0.11 mole) is dissolved in the dimethylbenzene (1L) of boiling, is slowly added into P2S5 (9.5 grams, 0.0385 mole), backflow, show to have reacted until reactant completes (about 5 hours) .TLC (PE/EA=5/1) Become.This reactant mixture cools down, and with the extraction of 1N NaOH, aqueous phase hydrochloric acid acidifying, collects yellow mercury oxide, then by it Being dissolved in EtOAc, wash with H2O, be dried with Na2SO4, and concentrate, the compound 3 (19 grams, yield: 70.9%) obtained is Red oil.
Step 3: compound 3 (30 grams, 0.142mol) is dissolved in 1N sodium hydroxide (500 milliliters), uses the potassium ferricyanide (135 grams, 0.416mol, in 340ml water) aoxidize, and carry out to ferricyanide solution by being slowly added to thioamides, make Temperature keeps below 10 DEG C.After 1h, TLC (in DCM/MeOH=3/1) shows that reaction completes.By reactant mixture at water Middle dilution, is acidified (being 6 by 2N HCl to pH), then filters, the compound 4 (15g, yield: 50.48%) obtained.
Step 4: in DMF (20mL) solution of compound 4 (1 gram, 4.78 mMs), under room temperature under nitrogen, stirring adds 4- Fluoro-3-(trifluoromethyl) anilinechloride (4-fluoro-3-(trifluoromethyl) aniline Hydrochloride, 1.24 grams, 5.74 mMs), it is subsequently added HATU (2.73 grams, 7.17 mMs) and DIEA (1.85 grams, 14.34 mMs). After being stirred overnight at a temperature of identical, TLC (PE/EA=2/1) shows that reaction completes.Reactant mixture is poured in water, And extract with EtOAc.Wash organic facies with saline, be dried at Na2SO4 and concentrate, obtain crude product, passed through silica gel chromatography (with Eluting) purification, obtain pure compound 5 (1.3 grams, yield: 73.4%).
Step 5: in compound 5 (0.5 gram, 1.35 mMs) DCM (10mL) solution, under-10 DEG C of nitrogen, adds 4N BBr3 (2.5mL), after reacting 4 hours, TLC (PE/EA=1/1) show that reaction completes.In reaction add ice, then filter with Remove undissolved material.Separating filtrate, extracts with DCM.Saline washing organic facies, is dried with Na2SO4 and concentrates, obtaining Crude product, is passed through pre--TLC purification, is obtained pure compound 6 (250mg, yield: 51.97%).
Step 6: in DMF (10mL) solution of compound 6 (70 milligrams, 0.196 mM), adds K2CO3 (81.46 millis Gram, 0.589 mM) and the tert-butyl group (6-chloropyrimide-4-base) methyl (methyl) t-butyl carbamate (tert-butyl (6- Chloropyrimidin-4-yl) methyl (methyl) carbamate, 55.70 milligrams, 0.216 mM), reaction is mixed Compound is at 50 DEG C of stirred under nitrogen overnight.TLC (PE/EA=1/1) shows that reaction completes.Reactant mixture is washed with water, with EA extracts.Washing organic facies, washs 4 times with saline, is dried at Na2SO4 and concentrates, obtain crude product, passed through TLC pure Change, obtain pure compound 7 (40 milligrams, 39.51%).
7th step: compound 7 (38 milligrams, 0.071 mM) and the mixture of TFA (0.4 milliliter), under nitrogen room temperature Stir half an hour.TLC (in DCM/MeOH=10/1) show that reaction completes.Reactant mixture evaporation is removed TFA.By residue Being regulated to pH by Na2CO3 is 9, then extracts with DCM.Washing organic facies, washs with saline, is dried with Na2SO4 and concentrates, To crude product, being passed through TLC purification, obtain pure compound K DR8B (15 milligrams, 47.75%), its Structural Identification data are joined See Figure 21.
Beneficial effects of the present invention is illustrated below by way of test example.
Test example 1 Brachydanio rerio vascular development inhibition test
Brachydanio rerio (Danio rerio) is a kind of bony fish that Cyprinidae (Cyprinidae) short hundredweight Buddhist nun fish belongs to (Danio).Its Gene is up to 85% with the similarity of human gene, and raun once can lay eggs 200～300 pieces, its fertilization and embryo development procedure Carry out the most in vitro, just can grow shaping in 24 hours, and embryo's entire body is transparent, it is simple to observe the change of intracorporeal organ tissue. Various features becomes one of 5 kinds of Fish as Laboratory Animals of Liao Shi International Organization for Standardization accreditation.At present, Brachydanio rerio is the mankind Being used widely in disease research, the research in terms of cardiovascular system is the most.
Experimental technique: this experiment uses the Brachydanio rerio usually used as examination compound on vascular formation function influence to be animal Model.After selected for postnatal zebrafish embryo, it be put in culture dish and be placed in incubator fostering 3-5 days, the present invention Compound is pressed variable concentrations (1 μM, 10 μMs, 30 μMs, 100 μMs) after Brachydanio rerio is born and is directly added in culture fluid.After 24 hours Check the developmental state of vertebra blood vessel and take a picture with fluorescence microscope.130B is pazopanib (Pazopanib), right as the positive According to, DMSO is as negative control.
Result of the test sees Fig. 1-4, table 1a, 1b.
The table 1a:KDR series little molecule inhibitory action to FLK-1 transgenic zebrafish vascular development
Concentration (uM) KDR3 KDR4 KDR6 KDR8 KDR8A
1 0% 0% 0% 0% 0%
10 0% 0% 0% 0% 0%
30 0% 10% 0% 0% 0%
100 50% 100% 10% 5% 10%
The table 1b KDR series little molecule inhibitory action to FLK-1 transgenic zebrafish vascular development
AI: vascular study rate
From Fig. 1-4, the compounds of this invention KDR4 and V01-v6 all can suppress Brachydanio rerio vascular development, especially V01, V3, v6, its drug activity is notable.
From table 1b, the activity of compound DKR4 is substantially better than KDR3, KDR6, KDR8 and KDR8A, compound V01, The activity of V03 is substantially better than KDR4.
The effect experiment of test example 2 the compounds of this invention
1. pair VEGFR2 body outer suppressioning test
The experimental technique of detection the compounds of this invention suppression VEGFR2 tyrosine phosphorylation is as follows:
1) cultivation to P3-P5HUVEC cell is transferred to 6 orifice plates, every hole about 2*105Cell
2) treat that cell grows to 70-80%, add candidate small molecule inhibitor (10nM, 100nM or 1 μM), hatch 30min
3) add VEGF 50ng/ml to stimulate, continue 10min
4) add the termination of non denatured lysate to react, collect cell pyrolysis liquid, carry out protein quantification
5) SDS-PAGE electrophoresis, transferring film, with the TTBS buffer (Tris-of 5% defatted milk preparation after cutting film Buffered saline/0.01%Tween 20) close 2 hours
6) wash film, close overnight with anti-phosphorylation VEGFR2 antibody (1:1000 dilution) 4 DEG C
7) 1 hour is hatched with the goat antirabbit two of horseradish peroxidase-labeled is anti-altogether with film after within the 2nd day, washing film
8) develop the color with chemical luminescence reagent kit (Millipore) after washing film, photograph.
Result of the test sees Fig. 5.
As shown in Figure 5, the compounds of this invention KDR4 and V01, V3 all can suppress VEGFR2, and wherein, compound V01, V3 are imitated Fruit is more preferable compared with KDR4.
Test example 3 the compounds of this invention inhibitory action to choroidal neovascularization
Mice is c57/BL, uses the choroidal neovascularization (Choroidal of induced with laser Neovascularization, CNV) animal model tests.The one that this model is currently also most widely used is used to grind Study carefully medicine and CNV is formed the animal model of impact.Use 532 laser C57/bl6 mice (4 laser spots of each retina) OK Laser retinal light coagulates the CNV induced and is formed.Laser is injected after implementing immediately: a mice right eye injection concentration The KDR4 of 150uM, the V01 of another mice right eye injection concentration 1uM, two mice left eye injection all injections and medicine same volume PBS as comparison.Inject latter 5 days and animal is put to death and obtains eyeball.After removing anterior ocular segment, vitreous body and retina, make Choroid tile Parallel I solectin dyes.Zeiss fluorescence microscope system is used to carry out taking pictures and the measurement of CNV area.Knot Fruit sees Fig. 6, Fig. 7.
All can significantly inhibit choroidal neovascularization according to Fig. 6, compound K DR4 (150uM), V01 (1uM) to be formed, Can effectively treat or alleviate wet MD.
Test example 4 the compounds of this invention is on kinase whose impact
Protein kinase (protein kinases) is also known as protein phosphorylation enzyme (protein phosphakinase).One The enzyme of class catalytic proteins phosphorylation reaction.It can transfer to protein molecule the γ-phosphoric acid on adenosine triphosphate (ATP) On amino acid residue on the hydroxyl of some serine, threonine or tyrosine residue, thus change protein, the conformation of enzyme and work Property.The phosphorylation of protein is the important step of multi-signal pathway, and the important life of intracellular major part lives through journey all It is unable to do without protein phosphorylation.
Protein kinase falls into 5 types: Protein Serine/threonine kinase, protein tyrosine kinase, protein groups histidine kinase, Albumen tryptophan kinases and albumen aspartyl/glutamyl kinases.Protein kinase is in the regulation of cell processes and maintains Having arrived important function, all observed abnormal kinase in numerous disease state, described morbid state includes: pernicious swollen Tumor, immune disease, cardiovascular disease, diabetes, infectious disease, arthritis and other immunologic derangement, nervous system are as old Year dementia, A Mohaici disease AD etc., it has now been found that to relevant with protein kinase more than 400 kinds of human diseasess.
Wherein, VEGFR (vascular endothelial growth factor receptor) family member is receptor tyrosine kinase, as VEGFR1, VEGFR2 etc., this receptoroid malignant tumor growth, transfer in and vascular proliferative disease (as degeneration of macula, Tumor) etc. have a major impact during advancing of disease.
PDGFR (platelet derived growth factor receptor) family member is receptor tyrosine kinase, such as PDGFR α and PDGFR β and colony-stimulating factor 1 receptor, stem cell factor receptor KIT etc., currently also find sending out of such kinases and tumor Raw, development has close ties.As PDGFR unconventionality expression melanoma, meningioma, neuroendocrine tumor, ovarian cancer, Being found in carcinoma of prostate, pulmonary carcinoma and cancer of pancreas, the abnormal activation of KIT is then the direct inducement of many tumorigenesis.
1) suppression dose response studies
Compound V01 is dissolved in 100%DMSO, is diluted to 3 groups of series concentration, and DMSO all keeps in last test 1%.Test maximum concentration is 50uM.With the activity inhibitor D-82041 DEISENHOFEN of non-selective protein kinase (Staurosporine) as reference, maximum concentration is 1uM.The results are shown in Table 3, Figure 22.
Table 2
Target Supplier [enzyme], nM [ATP],μM Incubation time, hr
KDR Invitrogen 0.25 80 3
The table 3 inhibitory activity to KDR
Compound IC50(μM) 95% confidence rate Hill
Staurosporine 0.00942 0.00121 1.002392
V01 0.0207 0.00287 1.007953
2) suppression specificity research
To compound VO1, testing 22 kinds of kinase activity inhibition tests, test concentrations is 5mM, is repeated twice, at ATP Km tests.First V01 is dissolved in 100%DMSO, and concentration of ordinary dissolution is 100 times of final test concentration, during all final tests DMSO is 1%.The activity inhibitor D-82041 DEISENHOFEN (Staurosporine) of non-selective protein kinase, as comparison, is surveyed Cmax 10mM. is used during examination
Concrete test method and reagent are as shown in the table:
Table 4
Kinases is tested average inhibition such as following table for twice by V01:
Table 5
Knowable to result of the test, V01 is up to 100% to the suppression ratio of protein kinase K DR, relevant to tumor to other two kinds Protein kinase also have certain inhibitory action, wherein, PDGFR-β suppression ratio is 52.7%, and KIT suppression ratio is 68%, but V01 pair Remaining protein kinase inhibiting activity overwhelming majority is less than 5%.As can be seen here, and the small molecule kinase inhibitors phase of existing research and development Ratio, the compounds of this invention has high specific and a high efficiency inhibitory action to KDR, and to PDGFR-β and KIT both with swollen Tumor has the protein kinase of close association also to have certain inhibitory action, and this indicates that compound V01 is to KDR, PDGFR-β and KIT The diseases such as various tumors that these three protein kinase abnormal activation is relevant and old wet MD all have potential therapeutic activity.
3) research of new vessels suppression
Test 5 Mus altogether. in cornea central authorities, burn with 75% silver nitrate solution and 25% potassium nitrate rod induced chemical Modeling.Immediately at conjunctiva of right eye hemostasis 0.2ml compound V01 after chemical burn, then drip 0.2ml compound V01 every day 2 times.After 7 days, cornea is taken pictures and is compared.
Shown in Figure 23, compound V01 can significantly reduce new vessels and hemorrhage.
In sum, the compounds of this invention has good anti-angiogenic rebirth effect, it was initially believed that this compounds is logical Cross suppression VEGFR2 (i.e. KDR) and produce activity.This compounds can be used for new vessels, VEGFR2, PDGFR-β, KIT etc. The treatment of protein kinase caused by abnormal disease, such as wet MD, malignant tumor etc..

Claims (11)

1. compound as shown in formula B and pharmaceutically acceptable salt thereof or prodrug:
Wherein, R represents C1-C6Alkyl, X indicate without or on phenyl ring the substituted halogen atom in optional position.
Compound the most according to claim 1, its feature is being: R is selected from methyl.
Compound the most according to claim 1 and 2, it is characterised in that: described halogen atom is fluorine atom.
4., according to the compound described in claim 1-3 any one and pharmaceutically acceptable salt thereof or prodrug, it is special Levy and be: described compound is:
5. prepare the method for compound described in claim 1-4 any one for one kind, it is characterised in that: it comprises the following steps:
Wherein, Pg represents amino protecting group；
(1) with formula (I b) compound as raw material, formula (II b) compound is prepared with phosphorus oxychloride reaction；
(2) as raw material, react with cyanylation agent with formula (II b) compound, prepare formula (III b) compound；
(3) in the presence of base, by formula (III b) compound hydrolysis, formula (IV b) compound is prepared；
(4) as raw material, carry out condensation reaction with formula (IV b) compound and formula (V b) compound, prepare formula (VI b) chemical combination Thing；
(5) in the presence of Boron tribromide, the methyl ether structure on the aromatic ring of formula (VI b) compound is hydrolyzed, prepares formula (VII B) compound；
(6) as raw material, carry out condensation reaction with formula (VII b) compound and formula (VIII b) compound, prepare formula (Ⅸ b) chemical combination Thing；
(7) in acid condition, slough the amino protecting group of formula (Ⅸ b) compound, prepare formula (B) compound.
6. compound described in claim 1-4 any one and pharmaceutically acceptable salt thereof or prodrug are to prepare blood vessel new Purposes in raw inhibitor class medicine.
Purposes the most according to claim 6, it is characterised in that: described angiogenesis inhibitors is VEGF Receptor inhibitor 2；Preferably, described inhibitor is the medicine of anti-ocular angiogenesis.
Purposes the most according to claim 7, it is characterised in that: described medicine is choroid neovascularization inhibitors.
9. according to the purposes described in claim 6-8 any one, it is characterised in that: described medicine is treatment or pre-moisture resistance Huang The medicine of speckle degeneration, diabetic retinopathy or neovascular glaucoma.
10. compound described in claim 1-4 any one and pharmaceutically acceptable salt or prodrug thereof are in preparation VEGFR2, PDGFR-β is or/and purposes in KIT inhibitor class medicine.
11. 1 kinds of pharmaceutical compositions, it is characterised in that: it is containing compound and medicine thereof described in claim 1-4 any one Acceptable salt or the ophthalmic preparation of prodrug on.
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Application publication date: 20161207