CN103880861B - A kind of 4,6-dimethyl-oxazoles also [5,4-d] pyrimidine-5 acting on FGF receptor, 7(4H, 6H)-derovatives - Google Patents

A kind of 4,6-dimethyl-oxazoles also [5,4-d] pyrimidine-5 acting on FGF receptor, 7(4H, 6H)-derovatives Download PDF

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CN103880861B
CN103880861B CN201410060513.1A CN201410060513A CN103880861B CN 103880861 B CN103880861 B CN 103880861B CN 201410060513 A CN201410060513 A CN 201410060513A CN 103880861 B CN103880861 B CN 103880861B
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dimethyl
pyrimidine
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叶发青
郭平
谢自新
刘剑敏
王跃武
宋晓琴
陈梁芳
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Wenzhou Medical University
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Abstract

The invention discloses a kind of 4,6-dimethyl-oxazoles also [5,4-d] pyrimidine-5,7 (4H acting on FGF receptor, 6H)-derovatives, the present invention is with 1,3-dimethyl pyrimidine-2,4,6 (1H, 3H, 5H)-triketones are starting raw material, use NaNO 2carry out nitrosification, then use Na 2s 2o 4reduction, then carry out Schiff reaction with aromatic aldehyde, finally use SOCl 2cyclization obtains target compound.4,6-dimethyl-oxazoles of the present invention also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives have certain restraining effect to experiment cell, show certain anti-tumor activity.For H460 cell, the IC of compound N 5a, N5g and N5k 50value is all less than positive control drug; For B16F10 cell, the IC of compound N 5f, N5h, N5i and N5k 50value is less than positive control drug; For A549 cell, the IC of positive control 50be worth higher, the IC of compound N 5f, N5g, N5h and N5l 50value is all less than this positive control drug.

Description

A kind of 4,6-dimethyl-oxazoles also [5,4-d] pyrimidine-5 acting on FGF receptor, 7(4H, 6H)-derovatives
Technical field
The present invention relates to technical field of medical chemistry, particularly relate to a kind of 4,6-dimethyl-oxazoles also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives acting on FGF receptor.
Background technology
Malignant tumour is a kind of common and frequently-occurring disease of serious threat human health, and the mortality ratio that the mankind cause because of malignant tumour is only second to cardiovascular and cerebrovascular diseases and is in second.Though its methods for the treatment of has multiple at present, but still main based on chemotherapy.Though traditional antitumor drug effect is strong, lack selectivity, toxic side effect is also larger, limits its clinical application, and therefore people wish to research and develop medicine by the transmission mechanism based on tumor signal always.
Protein kinase is a kind of phosphotransferase, is played a crucial role from the acceptor that ATP transfers to substrate protein by catalytic phosphatase group in adjustment metabolism, genetic expression, Growth of Cells, cell fission and cytodifferentiation etc.In the antitumor research of protein kinase family, the molecular targeted antitumor drug research and development being target spot with receptor tyrosine kinase (RTKs) have become the hot-point and frontier field of current life science and pharmaceutical science.Confirmed by the continuous exploratory development of researchist in recent years, suppress tyrosine protein kinase can reach the object for the treatment of tumour, the research of RTKs inhibitor becomes a focus of antitumor drug research gradually.At present, existing 120 kinds of RTKs inhibitor medicaments nearly go on the market successively or enter clinical experimental stage, wherein most inhibitor is for TYR kinases such as EGF-R ELISA (EGFR), vectors containing human platelet-derived growth acceptor (PDGFR), human vascular endothelial growth factor acceptors (VEGFR), and relatively less for the inhibitor research of FGFR, and such inhibitor mostly is the TYR kinase inhibitor of Mutiple Targets.In view of the effect of increasing report FGFR signal path in induced tumor growth and blood vessel hyperplasia, the research carrying out FGFR inhibitor seems particularly urgent.
Fibroblast growth factor acceptor (FGFRs) is the Tyrosylprotein kinase receptor of a class cross-film, it and fibroblast growth factor (FGFs) combine the FGF/FGFR signal transmission formed can mediate many barss Signal Transduction Pathways, all plays important regulating effect to the aspect such as growth and transfer of the propagation of normal cell and tumour cell, differentiation and tumour.Many research reports are pointed out, the generation of tumour, growth and transfer all activate closely related with FGF/FGFR abnormal expression and signal path thereof, be embodied in: 1. in the kinds of tumor cells such as bladder cancer, cancer of the stomach, colorectal carcinoma, carcinoma of endometrium, prostate cancer, fibroma, rhabdosarcoma, neurospongioma and melanoma, all find the sudden change of FGF/FGFR or high-caliber expression, the highlyest can exceed normal value more than 100 times.2.FGFR path is short proliferation signal, can promote the abnormality proliferation of tumour cell.The activation of 3.FGFR signal path can also stimulate angiogenic growth.Angiogenesis not only provides nutrient and oxygen for tumour cell, promotes that it grows, and is also that invasion and attack, the metastasis and extension of tumour cell provides basic substance.(4) FGFR also can mediate the antitumor drug resistance of wide spectrum, and tumour cell to the resistance of chemotherapeutics and in metastatic tumo(u)r treatment the limited efficacy of chemotherapy be topmost two challenges clinically.Therefore, FGFR is regarded as one of oncotherapy target and receives much concern.
FGF is as extracellular stimulus signal, RTK extracellular region is attached to by HSPG, receptor protein is made to form dimer, there is autophosphorylation in the RTK active zone activating intracellular region, then brings out PLC-γ approach, P-I-3K-AKT-PKB approach, the effect of Ras2/Raf/MAPK approach performance series of physiological and pathological.The map kinase (comprising ERKs, p38 and JNKs) wherein activated in Ras2/Raf/MAPK approach enters nucleus, makes several genes Function protein phosphorylation, thus affect gene synthesis, the process such as to transcribe.The genetic transcription of cyclin is mainly made to increase for tumour cell, cyclin dependent protein kinase is activated, cyclin dependent protein kinase reactivation cell cycle advance gene after activation, causes the cell cycle to shorten, breeds and accelerate, thus the formation of induced tumor.
The inhibitor being target spot based on FGFR can be divided into two classes substantially: namely born of the same parents block inhibitor and the intracellular signal transduction inhibitor of FGF and FGFR combination outward.Wherein Extracellular inhibitor is mainly divided into polyanionic inhibitor, carbohydrate inhibitor and peptide inhibitor.In born of the same parents inhibitor research in, based on active region ATP phosphorylation target spot research particularly active.At present existing multiple important small molecules FGFR-1 inhibitor: as indoles as SU4984 and SU5402; Pyridines is as the Sunitinib of PD173074 and indole ketone.SU4984 and SU5402 suppresses the IC of FGFR1 kinase activity 50value is 10-20 μM.PD173074 is the selectivity FGFR inhibitor of unique synthesis at present, its IC suppressed FGFR1 50value is nmole level.In addition, the indole ketone FGFR inhibitor Sunitinib of Pfizer's research and development is to obtain FDA approval listing, and this medicine is also a kind of TYR kinase inhibitor of Mutiple Targets.
FGFR-1 micromolecular inhibitor
The research of such target spot in the recent period has new breakthrough again, Shanghai in 2011 institute of materia medica Ding Jian etc. and East China University of Science Xu Yu virtue waits and has found acenaphthene also [1 by Rational drug design and optimization, 2-b] pyroles FGFR-1 selectivity micromolecular inhibitor, and structure activity relationship is good, molecular docking shows that this inhibit activities is relevant with the ATP-binding site of compound and FGFR-1.Cell levels display compound is to the IC of FGFR-1 high expression tumour cell 50be worth even low at micromolar levels.
This kind of target spot inhibitor mainly inhibitor and competition suppresses kinases ATP binding domain, FGFR-1 kinases ATP binding domain is the catalytic core region of a high conservative, be folded into " double leaf " structure connected by " hinge " by 12 subunits, the VITAMIN B4 ring of ATP is combined with kinases " hinge " region by forming hydrogen bond.We have collected the FGFR-1 Tyrosylprotein kinase crystalline structure reported at present, find that current FGFR-1 inhibitor all has larger conjugated structure by computer simulation analysis, and the ATP-binding site in its inhibit activities region is only a very little part for FGFR-1 albumen pocket, be positioned at the inside left of whole pocket cavity, and ATP pocket right part also has larger space not to be utilized.Thus design can utilize unemployed space, right side can increase compound undoubtedly to the selectivity of FGFR-1 and inhibition strength.
Summary of the invention
By the structural research to these active FGFR-1 micromolecular inhibitors preferably, we find that this compounds often has following character: 1. parent nucleus is generally two dimensional structure, but the structural specificity of parent nucleus is not very strong, and this has just pointed out the binding site of inhibitor and the FGFR-1ATP binding domain specificity that also tool is too not large.2. compound generally has good conjugacy.We early stage synthesized by 42 xanthine derivatives possess These characteristics completely, external FGFR-1 kinases ATP competitive experimental result display section compound suppresses FGFR-1 really better, but this compounds shortcoming is exactly that space structure is too single, substituted radical just extends a horizontal direction, the FGFR-1ATP pocket right part found in conjunction with computer simulation analysis also has the unemployed fact of larger space, we propose the xanthine analogue that meter can act on this region, in view of this conception, we adopt the drug design methods such as isostere, the N on xanthine 9 is replaced with O, by shortening horizontal direction substituent length and increase the substituting group of vertical direction gradually with it, to expect to act on mutually with FGFR-1ATP pocket right part larger space, devise a class and retain parent nucleus and substituting group conjugation, and space performance more changeable 4, 6-dimethyl-oxazole also [5, 4-d] pyrimidine-5, 7 (4H, 6H)-derovatives, to inquire into this compounds to the inhibiting impact of FGFR-1.Wherein the test of external FGFR-1 inhibit activities adopts its vitro kinase activity to suppress screening method, with mtt assay cell levels test compounds to FGFR-1 high expressing cell anti-tumor activity, finally by molecular biology method detection of active compound to the inhibit activities of FGFR downstream signal passage, obtain active definite FGFR micromolecular inhibitor.
The present invention adopts following technical scheme:
The chemical structure of 4,6-dimethyl-oxazoles acting on FGF receptor of the present invention also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives is as follows:
Wherein, described derivative is 2-[4-(benzyloxy)-phenyl]-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone, and chemical structure is as follows:
Wherein, described derivative is 2-(3-1H-indyl)-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone, and chemical structure is as follows:
Wherein, described derivative is 2-[1-(2-hydroxyl) naphthyl)]-4,6-dimethyl-oxazoles also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone, and chemical structure is as follows:
One prepares the method for 2-[4-(benzyloxy)-phenyl]-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone, and the concrete steps of described method are as follows:
(1) add the ratio of the mixed solution of 140ml acetic acid and water with every 5g1,3-dimethyl pyrimidine-2,4,6 (1H, 3H, 5H)-triketone, at 75 DEG C, heat 30min homogeneous to reaction mixture, temperature is reduced to 50 DEG C, adds NaNO in batches 24.0g, after adding, is cooled to room temperature, and stirs 1h, be precipitated thing, filters, washing, and suction filtration is dry, obtains 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone;
(2) every 0.01mol1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone is added 40ml14.5%NH 4the ratio of OH, stirs 30min at 70 DEG C, until mixture is homogeneous, by near for temperature 50 DEG C, adds the Na of 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone amount of substance 2 times in batches 2s 2o 4, and continue to stir 30min, rotary evaporation screws out ammoniacal liquor, separates out solid, and refrigerator cools, and drains, obtains 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketone; (3) by 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketones and 4-benzoxybenzaldehyde are with the ratio of 1:1, every mmole adds in 15ml dehydrated alcohol, cools, suction filtration at 80 DEG C after stirring reaction 3h in refrigerator, washing, is drying to obtain intermediate 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones;
(4) every mmole 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones add 1.5ml and heavily steam rear thionyl chloride, stir 1h in 60 DEG C, rotary evaporation screws out thionyl chloride, adds little water and dissolves, adjust PH to 7 with ammoniacal liquor, leave standstill, suction filtration, obtain 2-[4-(benzyloxy)-phenyl]-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone.
In step (1), in the mixed solution of acetic acid and water, the volume ratio of acetic acid and water is 1:1.
One prepares the method for 2-(3-1H-indyl)-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone, and the concrete steps of described method are as follows:
(1) add the ratio of the mixed solution of 140ml acetic acid and water with every 5g1,3-dimethyl pyrimidine-2,4,6 (1H, 3H, 5H)-triketone, at 75 DEG C, heat 30min homogeneous to reaction mixture, temperature is reduced to 50 DEG C, adds NaNO in batches 24.0g, after adding, is cooled to room temperature, and stirs 1h, be precipitated thing, filters, washing, and suction filtration is dry, obtains 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone;
(2) every 0.01mol1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone is added 40ml14.5%NH 4the ratio of OH, stirs 30min at 70 DEG C, until mixture is homogeneous, by near for temperature 50 DEG C, adds 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone amount of substance, 2 times of Na in batches 2s 2o 4, and continue to stir 30min, rotary evaporation screws out ammoniacal liquor, separates out solid, and refrigerator cools, and drains, obtains 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketone;
(3) by 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketones and 4-benzoxybenzaldehyde are with the ratio of 1:1, every mmole adds in 15ml dehydrated alcohol, cools, suction filtration at 80 DEG C after stirring reaction 3h in refrigerator, washing, is drying to obtain intermediate 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones;
(4) every mmole 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones add 1.5ml and heavily steam rear thionyl chloride, stir 1h in 60 DEG C, rotary evaporation screws out thionyl chloride, adds little water and dissolves, adjust PH to 7 with ammoniacal liquor, leave standstill, suction filtration, obtain 2-[4-(benzyloxy)-phenyl]-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone.
In step (1), in the mixed solution of acetic acid and water, the volume ratio of acetic acid and water is 1:1.
4,6-dimethyl-oxazoles of the present invention also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives may be used for treating tumour.
4,6-dimethyl-oxazoles of the present invention also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives have certain restraining effect to experiment cell, show certain anti-tumor activity.For H460 cell, the IC of compound N 5a, N5g and N5k 50value is all less than positive control drug; For B16F10 cell, the IC of compound N 5f, N5h, N5i and N5k 50value is less than positive control drug; For A549 cell, the IC of positive control 50be worth higher, the IC of compound N 5f, N5g, N5h and N5l 50value is all less than this positive control drug.
Accompanying drawing explanation
Fig. 1 is the schematic diagram of the mean OD value of compound of the present invention 50 μMs.
Fig. 2 is the schematic diagram of the percent inhibition of compound of the present invention 50 μMs.
Fig. 3 is the schematic diagram of compound of the present invention to the IC50 value of H460 cell.
Fig. 4 is the schematic diagram of compound of the present invention to the IC50 value of B16F10 cell.
Fig. 5 is the schematic diagram of the compounds of this invention to the IC50 value of A549 cell.
Embodiment
The following examples describe in further detail of the present invention.
The synthesis of embodiment 1 compound
The synthetic route of 1.1 compounds
Concrete synthetic route is as follows:
i:NaNO 2,CH 3COOH:H 2O＝1:1；ii:Na 2S 2O 4,NH 4OH；iii:R-CHO,CH 3CH 2OH；iv:SOCl 2
1.2 synthesis step
1.2.11, the preparation of 3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone (2)
By 5g1,3-dimethyl pyrimidine-2,4,6 (1H, 3H, 5H)-triketones (1) add in the mixed solution (acetic acid: water=1:1) of 140ml acetic acid and water, at 75 DEG C, heat 30min homogeneous to reaction mixture, temperature is reduced to 50 DEG C, adds NaNO in batches 24.0g, after adding, is cooled to room temperature, and stirs 1h, be precipitated thing, filters, washing, and suction filtration is dry, obtains 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone (2), yield: 90%.
1.2.21, the preparation of 3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketone (3)
1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone is added 40ml14.5%NH 4in OH, at 70 DEG C, stir 30min, until mixture is homogeneous, by near for temperature 50 DEG C, add 3.13gNa in batches 2s 2o 4, and continue to stir 30min, rotary evaporation screws out ammoniacal liquor, separates out solid, and refrigerator cools, and drains, obtains 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketone (3), yield: 85%.
1.2.31, the preparation of 3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketone (N4a-l).
By 0.001mol1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketones and 0.001mol aromatic aldehyde add in 15ml dehydrated alcohol, cool in refrigerator after stirring reaction 3h at 80 DEG C, suction filtration, washing, be drying to obtain intermediate 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketone (N4a-4l).1.2.44, the preparation of 6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives (N5d-5l)
By 0.001mol1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketone (N4a-4l) adds 1.5ml and heavily steams rear thionyl chloride, stirs 1h in 60 DEG C, rotary evaporation screws out thionyl chloride, add little water to dissolve, adjust PH to 7 with ammoniacal liquor, leave standstill, suction filtration, obtains target product.For compound N 5b, N5d, N5f, N5h, N5i, N5j, N5l, with ethanol and twice, DMSO mixed solution recrystallization and get final product, for compound N 5c, N5e, N5g, N5j, N5k, gained crude product methyl alcohol: methylene dichloride/1:30 crosses post and can obtain, fusing point and yield are in table 1.
1.3 experimental result
The physical data of table 1 target compound N5a-5l
The IR of table 2 target compound N5d-5l and 1h-NMR data
Target compound synthesized by the present invention is white, yellow or brown powder or flocculence solid.Compound N 5a-5l is all slightly soluble in methyl alcohol, and ethanol dissolves in ethyl acetate, is soluble in methylene dichloride, chloroform, ether, DMSO.Water insoluble.
Compound synthesized by the present invention, carries out mass spectrometric measurement, and positive pole all shows [M+1] +, and signal is stronger.Part of compounds also show [M+Na] +, as compound N 5a, N5f, N5h, its [M+Na] +be respectively 270.0,330.1,319.0.Also part of compounds is had to occur [2M+1] +peak, such as compound N 5e, N5f, its [2M+1] +be respectively 681.2,595.2.
Compound synthesized by the present invention, carries out infrared test, and target compound, all containing carbonyl, IR collection of illustrative plates collection of illustrative plates can obviously see that it exists, as compound N 5a, N5h, and its ν c=O: 1717.24,1682.77, ν c=O: 1710.75,1674.39, compound N 5h ν n-H: 3227.94.
Compound synthesized by the present invention, carries out 1h-NMR tests, from 1h-NMR collection of illustrative plates can be clearly seen that the hydrogen signal of compound and residing chemical shift thereof.As compound N 5h, its proton can be divided into two classes: 6 non-aromatic protons on basic parent nucleus, and 6 aromatic on substituting group indoles, and the proton wherein on parent nucleus affects its chemical shift to low field displacement by uride, shows as 3.38ppm, 3.55ppm; Proton on substituting group indole ring is entirely in and deshields, and chemical shift is also to low field displacement, and 7.25ppm place is 2 protons, shows multiplet; 7.53ppm place is a proton, shows bimodal, even summation constant J=7.2Hz; 8.14ppm place is also a proton, shows bimodal, even summation constant J=7.2Hz; 8.20ppm place is that a proton is unimodal; 11.97ppm place is proton on nitrogen.
Embodiment 2 antitumor activity of compound
MTT is the hydrionic yellow compound of a kind of acceptance, the respiratory chain in viable cell plastosome can be acted on, succinodehydrogenase in viable cell plastosome makes it reduce, generate a kind of water-insoluble bluish voilet crystallization first a ceremonial jade-ladle, used in libation (Formazan) and be deposited in cell, and dead cell is without this function.First a ceremonial jade-ladle, used in libation in DMSO energy dissolved cell, locates to measure its absorbance value with enzyme-linked immunosorbent assay instrument at a particular wavelength, and within the scope of certain cell count, the amount that first a ceremonial jade-ladle, used in libation is formed is directly proportional to cell count, therefore can indirectly reflect viable cell quantity.
2.1 experimentation
2.1.1 cell cultures
By A549 cell, B16F10 cell and the H460 cell 5%CO at 37 DEG C 2under condition, respectively with the RPMI1640 nutrient solution Secondary Culture containing 10% foetal calf serum, experiment cell is all in logarithmic phase.
2.1.2 the preparation of compound sample
With the RPMI-1640 dissolved compound of 10% foetal calf serum, be made into 50 μMs, 10 μMs, 4 μMs, the liquid sample of 0.2 μM, and preserved under 4 DEG C of conditions.
2.1.3MTT method measures each sample antitumor action
Nutrient solution in culturing bottle is carefully inhaled and abandons, add appropriate PBS liquid nutrient solution is cleaned, then with 0.25% trypsin solution digestion adherent growth cell 3min, add 10% serum free culture system liquid stop digestion, piping and druming cell make cell all come off and be dispersed in nutrient solution.Add RPMI-1640 piping and druming mixing, adjust cell density to the every hole of 3000-5000, (the aseptic PBS of marginal pore fills).Get 96 well culture plates, if experimental group and control group, using 5-FU as positive controls.Experimental group is respectively organized as a kind of compound, often kind of compound four concentration, and each concentration establishes three multiple holes, and separately establish a blank group, 100 μ L cell suspensions are inoculated in every hole, 5%CO 2, 24h cultivated by 37 DEG C of incubators, treats cell attachment.Supernatant liquor is abandoned in suction, and experimental group often organizes 50 μMs that add respectively and configured, 10 μMs, 4 μMs, and each 100 μ L of solution of the compound of 0.2 μM of four concentration, control group adds nutrient solution 100 μ L, cultivates 24 hours.Add 5mgmLMTT20 μ L, after continuing to hatch 4 hours, inhale and abandon supernatant solution, every hole adds 150 μ L dimethyl sulfoxide (DMSO), puts low-speed oscillation 10min on shaking table, crystallisate is fully dissolved.The light absorption value in each hole is measured, record result at enzyme-linked immunosorbent assay instrument OD570nm place.And by its inhibiting rate of following formulae discovery:
Inhibiting rate (%)=(control wells OD value-experimental port OD value)/control wells OD value × 100%.
2.2 experimental result
2.2.1MTT primary dcreening operation result
Mean OD value during table 3 compound 50 μMs
The inhibiting rate of table 4 compound 50 μMs time
Table 5 compound is to the IC50 value of three kinds of cells
As can be seen from table 3-5 and Fig. 1-5:
1) this experiment employing three kinds of tumour cells: A549 cell, B16F10 cell and H460 cell, set four concentration: 50 μMs, 10 μMs, 4 μMs, 0.2 μM take Fluracil as positive control drug.
2) under 50 μMs of concentration, for H460 cell, the inhibiting rate of compound N 5f and N5g reaches 76.51% and 80.62% respectively, higher than the inhibiting rate of positive control drug 63.95%; For B16F10 cell, the inhibiting rate of compound N 5e, N5f and N5h reaches 56.05%, 85.26% and 64.65% respectively, higher than the inhibiting rate of positive control drug 54.24%; For A549 cell, the inhibiting rate of compound N 5f and N5h reaches 72.25% and 56.88% respectively, higher than the inhibiting rate of positive control drug 45.69%.
3) by this concentration gradient gained IC 50value shows: part of compounds has good restraining effect to experiment tumour cell.For H460 cell, the IC of compound N 5a, N5g and N5k 50value is all less than positive control drug; For B16F10 cell, the IC of compound N 5e, N5f, N5h, N5i and N5k 50value is less than positive control drug; For A549 cell, the IC of positive control 50be worth higher, the IC of compound N 5f, N5g, N5h and N5l 50value is all less than this positive control drug.
4) can be found by comparison object compound and positive control drug: compound N 5f and N5g is all better to the restraining effect of three kinds of cells, and other compound to the antitumor action of each cell without evident regularity.2 of compound N 5f and N5g are lipophilic substituted phenylethylene base, styryl-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5 that preliminary deduction 2-replaces, the research and development of 7 (4H, 6H)-cyclohexadione compounds to antitumor drug have certain potentiality.
Although illustrate and describe embodiments of the invention, for the ordinary skill in the art, be appreciated that and can carry out multiple change, amendment, replacement and modification to these embodiments without departing from the principles and spirit of the present invention, scope of the present invention is by claims and equivalents thereof.

Claims (9)

1. one kind acts on 4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives of FGF receptor, it is characterized in that: the chemical structure of described derivative is as follows:
2. as claimed in claim 14,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives, it is characterized in that: described derivative is 2-[4-(benzyloxy)-phenyl]-4,6-dimethyl-oxazole is [5,4-d] pyrimidine-5,7 (4H also, 6H)-diketone, chemical structure is as follows:
3. as claimed in claim 14,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives, it is characterized in that: described derivative is 2-(3-1H-indyl)-4,6-dimethyl-oxazole is [5,4-d] pyrimidine-5,7 (4H also, 6H)-diketone, chemical structure is as follows:
4. as claimed in claim 14,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives, it is characterized in that: described derivative is 2-[1-(2-hydroxyl) naphthyl)]-4,6-dimethyl-oxazole is [5,4-d] pyrimidine-5,7 (4H also, 6H)-diketone, chemical structure is as follows:
5. prepare the method for 4,6-dimethyl as claimed in claim 2-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives for one kind, it is characterized in that: the concrete steps of described method are as follows:
(1) add the ratio of the mixed solution of 140ml acetic acid and water with every 5g1,3-dimethyl pyrimidine-2,4,6 (1H, 3H, 5H)-triketone, at 75 DEG C, heat 30min homogeneous to reaction mixture, temperature is reduced to 50 DEG C, adds NaNO in batches 24.0g, after adding, is cooled to room temperature, and stirs 1h, be precipitated thing, filters, washing, and suction filtration is dry, obtains 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone;
(2) every 0.01mol1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone is added 40ml14.5%NH 4the ratio of OH, stirs 30min at 70 DEG C, until mixture is homogeneous, cools the temperature to 50 DEG C, adds the Na of 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone amount of substance 2 times in batches 2s 2o 4, and continue to stir 30min, rotary evaporation screws out ammoniacal liquor, separates out solid, and refrigerator cools, and drains, obtains 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketone;
(3) by 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketones and 4-benzoxybenzaldehyde are with the ratio of 1:1, every mmole adds in 15ml dehydrated alcohol, cools, suction filtration at 80 DEG C after stirring reaction 3h in refrigerator, washing, is drying to obtain intermediate 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones;
(4) every mmole 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones add 1.5ml and heavily steam rear thionyl chloride, stir 1h in 60 DEG C, rotary evaporation screws out thionyl chloride, adds little water and dissolves, adjust PH to 7 with ammoniacal liquor, leave standstill, suction filtration, obtain 2-[4-(benzyloxy)-phenyl]-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone.
6. preparation method as claimed in claim 5, it is characterized in that: in step (1), in the mixed solution of acetic acid and water, the volume ratio of acetic acid and water is 1:1.
7. prepare the method for 4,6-dimethyl as claimed in claim 3-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives for one kind, it is characterized in that: the concrete steps of described method are as follows:
(1) add the ratio of the mixed solution of 140ml acetic acid and water with every 5g1,3-dimethyl pyrimidine-2,4,6 (1H, 3H, 5H)-triketone, at 75 DEG C, heat 30min homogeneous to reaction mixture, temperature is reduced to 50 DEG C, adds NaNO in batches 24.0g, after adding, is cooled to room temperature, and stirs 1h, be precipitated thing, filters, washing, and suction filtration is dry, obtains 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone;
(2) every 0.01mol1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone is added 40ml14.5%NH 4the ratio of OH, stirs 30min at 70 DEG C, until mixture is homogeneous, cools the temperature to 50 DEG C, adds 1,3-dimethyl-5-nitroso-group pyrimidine-2,4,6 (1H, 3H, 5H)-triketone amount of substance, 2 times of Na in batches 2s 2o 4, and continue to stir 30min, rotary evaporation screws out ammoniacal liquor, separates out solid, and refrigerator cools, and drains, obtains 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketone;
(3) by 1,3-dimethyl-5-aminopyrimidine-2,4,6 (1H, 3H, 5H)-triketones and 4-benzoxybenzaldehyde are with the ratio of 1:1, every mmole adds in 15ml dehydrated alcohol, cools, suction filtration at 80 DEG C after stirring reaction 3h in refrigerator, washing, is drying to obtain intermediate 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones;
(4) every mmole 1,3-dimethyl-5-imido grpup pyrimidine-2,4,6 (1H, 3H, 5H)-triketones add 1.5ml and heavily steam rear thionyl chloride, stir 1h in 60 DEG C, rotary evaporation screws out thionyl chloride, adds little water and dissolves, adjust PH to 7 with ammoniacal liquor, leave standstill, suction filtration, obtain 2-[4-(benzyloxy)-phenyl]-4,6-dimethyl-oxazole also [5,4-d] pyrimidine-5,7 (4H, 6H)-diketone.
8. preparation method as claimed in claim 7, it is characterized in that: in step (1), in the mixed solution of acetic acid and water, the volume ratio of acetic acid and water is 1:1.
9. 4,6-dimethyl-oxazoles as described in any one of claim 1-4 are the purposes of [5,4-d] pyrimidine-5,7 (4H, 6H)-derovatives in the medicine for the preparation for the treatment of tumour also.
CN201410060513.1A 2014-02-21 2014-02-21 A kind of 4,6-dimethyl-oxazoles also [5,4-d] pyrimidine-5 acting on FGF receptor, 7(4H, 6H)-derovatives Active CN103880861B (en)

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