CN108250191A - A kind of disubstituted 2- amino-pyrazinos compound of 3,5- and its preparation process and application - Google Patents
A kind of disubstituted 2- amino-pyrazinos compound of 3,5- and its preparation process and application Download PDFInfo
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- C07D413/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings
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- C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
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Abstract
The invention discloses 3,5 disubstituted 2 Aminopyrazine compounds, the compound has structure shown in formula I：In formula I：NR1R2For n-propylamine base, cyclopropyl amino, cyclopentamine base, cyclohexylamino, morpholinyl, 4 methyl piperazine bases, anilino-, 4 chloroanilinos or 3 chloroanilinos；Ar is 3,5 dimethyl isoxazole bases 4 or 1 methyl 1H pyrazolyls 4；And provide its preparation process and application.Such compound has significant inhibiting effect to tetra- kinds of human cervical carcinoma cell HeLa, human glioma cell U87, human liver cancer cell HepG2 and human colon cancer cell LoVo growth of cancer cells, wherein most compounds are significantly stronger than positive control VX 680 to the inhibiting effect of these cancer cell multiplications, especially chemical compounds I h and I i have stronger activity to these four cancer cells, and apparent inhibitory activity is shown to Aurora kinase A and aurora kinase B, therefore the compound of the present invention can be used for preparing anticancer drug.
Description
Technical field
The present invention relates to a kind of disubstituted 2- amino-pyrazinos compounds of 3,5- and its preparation process and applications.
Background technology
Laser kinases (Aurora kinases) is a kind of novel threonine/serineprotein kinase, is answered in centerbody
It is played extremely during the important mitosis such as system, the formation of the two poles of the earth spindle, chromosomal rearrangement and the monitoring of chromosome examination point
Close important role (Cancer Metastasis Rev.2003,22,451).There are three types of structure and work(for Aurora A family
The highly relevant subgroup of energy:That is Aurora-A, Aurora-B and Aurora-C.There are one their prlmary structure of protein contains
N- ends control region and a C- ends catalytic domain, and enzyme domains sequence similarity is up to the residual of 71%, ATP adenine ring binding sites
Base is also identical；But they have the function of entirely different and non-overlapping copies in cell division.Aurora A are from mitotic S
The end of term to the G phases of next division cycle, influences the detaching of centerbody, ripe and the two poles of the earth spindle formation
(Nat.Rev.Cancer 2005,5,42).Aurora B are located at the centromere region of chromosome, division in mitosis early stage
Stage is moved to micro-pipe from centromere.It is relatively fewer to the research of Aurora C functions at present.1998, American scholar
Bischoff etc. has found that Aurora A over-expresses in a variety of cancer cells for the first time, and unstability with chromosome, canceration,
The processes such as tumor proliferation and chemoresistance are closely related (EMBO is J.1998,17,3052).Due to the unique medicine of Aurora A
Mechanism of action is managed, the drug development using Aurora A as target spot has become one of hot spot of anticancer drug research
(Expert.Opin.Drug Discovery 2011,6,291).There are multiple Aurora A inhibitor carrying out clinic at present
Experiment, such as (pharmacy is in progress by VX-680, PHA-739358, AZD-1152, MLN8054, SNS-314, ENMD-2076, AMG900
2008,32,337；Chinese antibiotic magazine 2010,35,641), but there are no Aurora A inhibitor so far for facing
Bed treatment.
Aminopyrazine is a kind of important drug matrices, and research finds that such compound has kinds of tumors associated kinase
Significant inhibitory activity, as centerbody related protein kinase Nek2 (J.Med.Chem.2010,53,7682；
J.Med.Chem.2012,55,3228), checkpoint kinase 1 (CHK1) (J.Med.Chem.2011,54,8328；
J.Med.Chem.2012,55,10229), phosphatidyl-inositol 3-kinase (PI3K) (J.Med.Chem.2012,55,5467,
Bioorg.Med.Chem.Lett.2017,27,679), MNK1/2 (J.Med.Chem.2016,59,3034) and ATM and Rad3
Associated kinase ART (Nat.Chem.Biol., 2011,7,428；Oncotarget,2014,5,5674；
Bioorg.Med.Chem.Lett.2017,27,2659) etc..But inhibit to live for aurora kinase there is presently no such compound
The report of property.
Invention content
First purpose of the present invention is to provide for the disubstituted 2- amino-pyrazinos compounds of 3,5- of one kind, cell toxicant
Property experiment find that the compound has a variety of cancer cells stronger In-vitro Inhibitory Effect, while has aurora kinase stronger suppression
It makes and uses.
Second object of the present invention is to provide for the preparation process of above compound.
Third object of the present invention is to provide for the application of above compound.
The disubstituted 2- amino-pyrazinos compounds of 3,5- of one kind, the compound have following structure shown in formula I：
In formula I：
NR1R2For n-propylamine base, cyclopropyl amino, cyclopentamine base, cyclohexylamino, morpholinyl, 4- methyl piperazines base, anilino-,
4- chloroanilinos or 3- chloroanilinos；
Ar is 3,5- dimethyl isoxazoles base -4 or 1- methyl-1 H- pyrazolyls -4.
The preparation process of the above-mentioned disubstituted 2- amino-pyrazinos compounds of 3,5-, the technique are 4-O- (3- amino -6-
Bromo- pyrazinyl -2)-benzoic amide (formula II) and 3,5- dimethyl isoxazole -4- boric acid or (4,4,5,5- tetramethyl -1,3,
2- dioxaborolan -2- bases) the generation Suzuki condensation reaction of -1H- pyrazoles.
Further, 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic amides (formula II) and 3,5- dimethyl-Yi Evil
Azoles -4- boric acid or the amount of substance ratio of (4,4,5,5- tetramethyl -1,3,2- dioxaborolan -2- bases) -1H- pyrazoles reaction are 1:
(1.05-1.20), reaction temperature are 90-110 DEG C；Catalyst is complexed for [bis- (diphenylphosphine) ferrocene of 1,1'-] palladium chloride
Object, [bis- (diphenylphosphine) ethane of 1,2-] palladium chloride complex compound or the complexing of [bis- (diphenylphosphine) propane of 1,3-] palladium chloride
Object；Action solvent is 1,4- dioxane or tetrahydrofuran.
Further, the preparation method of 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic amide (formula II) is with 2-
Bis- bromo- pyrazine (formula III) of amino -3,5- is raw material, propionamide positive with P-hydroxybenzoic acid, P-hydroxybenzoic acid ring propionamide, right
Hydroxybenzoic acid ring pentanamide, P-hydroxybenzoic acid Cyclohexamide, P-hydroxybenzoic acid morpholino amide, P-hydroxybenzoic acid 4- first
Base piperazine amide, P-hydroxybenzoic acid benzamide, P-hydroxybenzoic acid chlorobenzamide between chlorobenzamide or P-hydroxybenzoic acid
Reaction.
Further, the preparation method of 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic amide (formula II) is 3,5-
With para hydroxybenzene formamide substitution reaction, 3,5- bis- bromo- 2- amino-pyrazinos and para hydroxybenzene first occur for two bromo- 2- amino-pyrazinos
The amount of substance ratio of amide reaction is 1:(1.05-1.20), reaction temperature are 60-80 DEG C, and catalyst is potassium carbonate, sodium carbonate, carbon
Sour caesium, action solvent is N-Methyl pyrrolidone, N- ethyl pyrrolidones, N- cyclohexyl pyrrolidones.
Application of the above-mentioned disubstituted 2- amino-pyrazinos compounds of 3,5- as aurora kinase inhibitors.
Application of the above-mentioned disubstituted 2- amino-pyrazinos compounds of 3,5- in anticancer drug.
The disubstituted 2- amino-pyrazinos class compounds of 3,5- provided by the invention have stronger inhibition to aurora kinase
Activity, and also there is preferable proliferation inhibition activity to a variety of cancer cells, there are potentiality of the exploitation for new anticancer drug；It is making
On Preparation Method, the present invention has been efficiently synthesized key intermediate and its target compound by straightforward procedure.
The invention has the advantages that：
The disubstituted 2- amino-pyrazinos compounds of 3,5- provided by the invention, to human cervical carcinoma cell HeLa, human brain colloid
Oncocyte U87, human liver cancer cell HepG2 and tetra- kinds of growth of cancer cells of human colon cancer cell LoVo have significant inhibiting effect,
Middle overwhelming majority compound is significantly stronger than the inhibiting effect of these cancer cell multiplications positive control VX-680, especially chemical compounds I h
There is stronger activity to these four cancer cells with I i, and show that apparent inhibition is lived to Aurora kinase A and aurora kinase B
Property, therefore the compound of the present invention can be used for preparing anticancer drug.
Specific embodiment
The preferred embodiment of the present invention is illustrated below, it should be understood that preferred embodiment described herein is only used
In the description and interpretation present invention, it is not intended to limit the present invention.
Embodiment 1：The preparation of the positive propionamide of P-hydroxybenzoic acid
By P-hydroxybenzoic acid (690mg, 5.0mmol) stirring and dissolving in 15mL acetone, n-propylamine is then added in
(6.0mmol) adds in 1- (3- dimethylamino-propyls) -3- ethyl-carbodiimide hydrochlorides until completely dissolved, in 70 DEG C of reflux
React 12h.Acetone is evaporated off after reaction, residue is diluted with ethyl acetate, then respectively with 1mol/L hydrochloric acid and
10wt%Na2CO3Solvent is evaporated off after organic phase drying in washing, and column chromatography obtains white powder.Product detection data are as follows：
Yield:71%；1H NMR(600MHz,DMSO-d6) δ 10.08 (s, 1H), 9.97 (s, 1H), 7.85 (d, J=
9.0Hz, 2H), 7.75 (d, J=7.2Hz, 2H), 7.32 (t, J=7.8Hz, 2H), 7.07-7.06 (m, 1H), 6.86 (d, J=
9.0Hz,2H)。
With cyclopropylamine, cyclopenta amine, cyclo-hexylamine, morpholine, 4- methyl piperazines, aniline, parachloroanilinum or m-chloroaniline
Instead of the n-propylamine in above-mentioned reaction, corresponding intermediate is prepared with method.
Embodiment 2：The preparation of 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide
The logical method of synthesis：Two bromo- pyrazines (253mg, 1.0mmol) of 2- amino -3,5- are dissolved in 10mL N- crassitudes
In ketone (or N- ethyl pyrrolidones or N- cyclohexyl pyrrolidones), P-hydroxybenzoic acid prepared above positive third is then added in
Amide (1.2mmol), adds in K until completely dissolved2CO3(276mg, 2.0mmol), in N2Under protection 6h is reacted in 100 DEG C.Instead
Should after, diluted with 30mL ethyl acetate, organic layer washs with saturated brine, dried, be evaporated off column chromatography after solvent obtain it is intermediate
Body 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide.Bright brown liquid.Product detection data are as follows：
Yield:90%；1H NMR(600MHz,DMSO-d6) δ 9.94 (s, 1H), 8.20 (s, 1H), 7.70 (d, J=
8.4Hz, 2H), 6.78 (d, J=8.4Hz, 2H), 3.19-3.15 (m, 2H), 1.52-1.48 (m, 2H), 0.87 (t, J=
7.2Hz,3H)。
With 4-HBA cyclopropyl amide, 4-HBA cyclopentyl amide, 4-HBA cyclohexyl acyl
Amine, 4-HBA morpholinyl amide, 4-HBA 4- methyl piperazine bases amide, 4-HBA benzamide, 4-
Hydroxybenzoic acid chlorobenzamide between chlorobenzamide, 4-HBA replaces the positive propionyl of P-hydroxybenzoic acid in above-mentioned reaction
Amine prepares corresponding intermediate with method.
Embodiment 3：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoic acid n-propyl acyl
The preparation of amine
4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide (0.5mmol) and 3 of above-mentioned gained is taken,
5- dimethyl isoxazole -4- boric acid (84.6mg, 0.6mmol) is dissolved in 10mL Isosorbide-5-Nitraes-dioxane, then adds in catalyst
Pd(dppf)Cl2·CH2Cl2(40.8mg, 0.05mmol), 2mol/L Na2CO3Solution (0.05mL, 0.1mmol), in N2Protection
Under be back to reaction in 100 DEG C and terminate, remove solvent under reduced pressure, residue is diluted with 15mL ethyl acetate, and is washed with saturated salt
It washs, dry, column chromatography obtains white solid after solvent is evaporated off.Product detection data are as follows：
Yield：72%；Purity:95% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=7.60min)；1H
NMR(600MHz,DMSO-d6) δ 8.44 (t, J=5.4,1H), 7.89 (d, J=7.2Hz, 2H), 7.79 (s, 1H), 7.31 (d, J
=7.2Hz, 2H), 6.78 (s, 2H), 3.21-3.18 (m, 2H), 2.29 (s, 3H), 2.07 (s, 3H), 1.53-1.49 (m, 2H),
0.87 (t, J=7.2Hz, 3H)；13C NMR(150MHz,DMSO-d6)δ165.4,165.3,158.0,155.1,145.9,
145.2,134.6,131.2,128.6(2C),127.8,121.4(2C),112.5,40.9,22.3,11.8,11.4,
11.0.ESI-MS:m/z 368.2 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I a.
Embodiment 4：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoic acid cyclopropyl acyl
The preparation of amine
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid cyclopropyl acyl
Amine replaces 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, obtains white solid.Product detection data are such as
Under：
Yield：74%；Purity:95% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=7.00min)；1H
NMR(600MHz,DMSO-d6) δ 8.44 (d, J=4.2Hz, 1H), 7.89 (d, J=8.4Hz, 2H), 7.81 (s, 1H), 7.32
(d, J=8.4Hz, 2H), 6.79 (s, 2H), 2.85-2.84 (m, 1H), 2.29 (s, 3H), 2.07 (s, 3H), 0.71-0.68 (m,
2H),0.59-0.56(m,2H)；13C NMR(150MHz,DMSO-d6)δ166.6,165.3,158.0,155.1,145.9,
145.2,134.7,130.9,129.0,128.6(2C),121.3(2C),114.6,23.0,11.8,11.0,5.68(2C)
.ESI-MS m/z 366.2 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I b.
Embodiment 5：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoic acid cyclopenta acyl
The preparation of amine
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid cyclopenta acyl
Amine replaces 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, obtains white solid.Product detection data are such as
Under：
Yield：66%；Purity:98% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=8.46min)；1H
NMR(600MHz,DMSO-d6) δ 8.27 (d, J=7.8Hz, 1H), 7.90 (d, J=8.4Hz, 2H), 7.78 (s, 1H), 7.30
(d, J=8.4Hz, 2H), 6.78 (s, 2H), 4.22-4.18 (m, 1H), 2.29 (s, 3H), 2.07 (s, 3H), 1.87-1.85 (m,
2H),1.67-1.65(m,2H),1.49-1.45(m,4H)；13C NMR(150MHz,DMSO-d6)δ165.3,165.0,158.0,
155.0,145.9,145.2,134.6,131.3,128.7(2C),127.7,121.3(2C),112.5,50.9,32.0(2C),
23.6(2C),11.8,11.0.ESI-MS m/z 394.2 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I c.
Embodiment 6：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoate base acyl
The preparation of amine
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoate base acyl
Amine replaces 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, obtains white solid, product detection data are such as
Under：
Yield：58%；Purity:95% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=9.05min)；1H
NMR(600MHz,DMSO-d6) δ 8.18 (d, J=8.4Hz, 1H), 7.91 (d, J=8.4Hz, 2H), 7.79 (s, 1H), 7.30
(d, J=9.0Hz, 2H), 6.77 (s, 2H), 3.74-3.72 (m, 1H), 2.29 (s, 3H), 2.07 (s, 3H), 1.80 (s, 2H),
1.72(s,2H),1.61-1.58(m,1H),1.31-1.28(m,4H),1.13-1.10(m,1H)；13C NMR(150MHz,
DMSO-d6)δ165.3,164.5,158.0,155.0,145.9,145.2,134.7,131.3,128.7(2C),127.7,
121.3(2C),112.6,48.3,32.4(2C),25.2,24.9(2C),11.8,11.0.ESI-MS m/z 408.2 for[M+
H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I d.
Embodiment 7：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoic acid morpholino amide
Preparation
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid morpholinyl acyl
Amine replaces 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, obtains light yellow solid, product detection data are such as
Under：
Yield：48%；Purity:97% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=6.72min)；1H
NMR(600MHz,acetone-d6) δ 7.82 (s, 1H), 7.54 (d, J=8.4Hz, 2H), 7.34 (d, J=9.0Hz, 2H),
6.16 (d, J=6.6Hz, 1H), 3.67 (s, 4H), 3.58 (s, 4H), 2.34 (s, 3H), 2.14 (s, 3H)；13C NMR
(150MHz,DMSO-d6)δ168.5,165.3,157.9,153.8,146.1,145.1,134.4,132.1,128.6(2C),
127.8,121.9(2C),112.4,65.9(2C),46.1(2C),11.8,11.0.ESI-MS m/z 394.2 for[M-H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I e.
Embodiment 8：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoic acid 4- methyl piperazines
The preparation of carboxamide dihydrochloride
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid 4- methyl piperazines
Carboxamide dihydrochloride replaces 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, obtains yellow solid, product detection data
It is as follows：
Yield：55%；Purity:98% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=6.88min)；1H
NMR(600MHz,DMSO-d6) δ 7.79 (s, 1H), 7.43 (d, J=9.0Hz, 2H), 7.29 (d, J=8.4Hz, 2H), 6.77
(s,2H),3.32(brs,8H),2.27(s,3H),2.18(s,3H),2.05(s,3H)；13C NMR(150MHz,DMSO-d6)δ
168.4,165.3,157.9,153.7,146.1,145.2,134.4,132.5,128.4(2C),127.8,121.9(2C),
120.9,114.8,112.4,54.4,45.4(2C),11.8,11.0.ESI-MS m/z 409.2 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I f.
Embodiment 9：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-phenylamino benzoic acid amide
It prepares
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-phenylamino benzoic acid amide generation
For 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, white solid is obtained, product detection data are as follows：
Yield：72%；Purity:95% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=9.29min)；1H
NMR(600MHz,DMSO-d6) δ 10.25 (s, 1H), 8.05 (d, J=9.0Hz, 2H), 7.83 (s, 1H), 7.78 (d, J=
7.8Hz, 2H), 7.41 (d, J=8.4Hz, 2H), 7.36 (d, J=7.2Hz, 2H), 7.10 (s, 1H), 6.83 (s, 2H), 2.33
(s,3H),2.12(s,3H)；13C NMR(150MHz,DMSO-d6)δ165.3,164.5,157.9,155.5,145.8,145.2,
139.0,134.8,131.2,129.1(2C),128.4(2C),127.8,123.5,121.3(2C),120.3(2C),92.5,
11.7,10.9.ESI-MS m/z 402.2 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I g.
Embodiment 10：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoic acid 4- chlorphenyls
The preparation of amide
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid 4- chlorphenyls
Amide replaces 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, obtains white solid, product detection data are such as
Under：
Yield：77%；Purity:95% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=12.40min)；1H
NMR(600MHz,DMSO-d6) δ 10.35 (s, 1H), 8.05 (d, J=9.0Hz, 2H), 7.83 (s, 1H), 7.82 (d, J=
8.4Hz, 2H), 7.42 (d, J=8.4Hz, 4H), 6.80 (s, 2H), 2.33 (s, 3H), 2.11 (s, 3H)；13C NMR(150MHz,
DMSO-d6)δ165.4,164.7,158.0,155.7,145.8,145.2,138.1,134.8,131.0,129.2(2C),
128.4(2C),127.8,127.2,121.8(2C),121.5(2C),112.5,11.8,11.0.ESI-MS m/z 436.2
for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I h.
Embodiment 11：4-O- (3- amino -6- (3,5- dimethyl isoxazoles base -4) pyrazinyl -2)-benzoic acid 3- chlorphenyls
The preparation of amide
Operating process is same with case study on implementation 3, only with 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid 3- chlorphenyls
Amide replaces 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic acid n-propyl amide, obtains white solid, product detection data are such as
Under：
Yield：66%；Purity:96% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=12.63min)；1H
NMR(600MHz,DMSO-d6) δ 10.40 (s, 1H), 8.06 (d, J=8.4Hz, 2H), 7.98 (s, 1H), 7.84 (s, 1H),
7.72 (d, J=8.4Hz, 1H), 7.43 (d, J=9.0Hz, 2H), 7.39 (t, J=8.4Hz, 1H), 7.17 (d, J=8.4Hz,
1H),6.82(s,2H),2.33(s,3H),2.12(s,3H)；13C NMR(150MHz,DMSO-d6)δ165.4,164.9,
158.0,155.8,145.8,145.2,140.6,134.9,132.8,130.8,130.2,129.3(2C),127.8,123.3,
121.5(2C),119.7,118.6,112.5,11.8,11.0.ESI-MS m/z 436.2 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I i.
Embodiment 12：4-O- (3- amino -6- (1- methyl-1 H- pyrazolyls -4) pyrazinyl -2)-benzoic acid phenyl amide
It prepares
Operating process and case study on implementation 9 are same, only with (4,4,5,5- tetramethyls -1,3,2- dioxaborolan -2- bases) -
1H- pyrazoles replaces 3,5- dimethyl isoxazole -4- boric acid, obtains white solid, product detection data are as follows：
Yield：76%；Purity:99% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=7.81min)；1H
NMR(600MHz,DMSO-d6) δ 10.25 (s, 1H), 8.05 (d, J=9.0Hz, 2H), 8.04 (s, 1H), 7.84 (s, 1H),
7.79 (d, J=7.2Hz, 2H), 7.67 (s, 1H), 7.41 (d, J=7.8Hz, 2H), 7.37 (t, J=7.2Hz, 2H), 7.10
(s,1H),6.54(s,2H),3.79(s,3H)；13C NMR(150MHz,DMSO-d6)δ164.7,156.0,145.0,144.9,
139.1,135.5,132.2,131.1,130.6,129.1(2C),128.4(2C),127.3,123.5,120.3(2C),120.2
(2C),119.8,38.4.ESI-MS m/z 387.2 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I j.
Embodiment 13：4-O- (3- amino -6- (1- methyl-1 H- pyrazolyls -4) pyrazinyl -2)-benzoic acid 4- chlorphenyl acyls
The preparation of amine
Operating process is same with case study on implementation 10, only uses (4,4,5,5- tetramethyls -1,3,2- dioxaborolans -2-
Base) -1H- pyrazoles replace 3,5- dimethyl isoxazole -4- boric acid, obtain white solid, product detection data are as follows：
Yield：72%；Purity:95% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=10.58min)；1H
NMR(600MHz,DMSO-d6) δ 10.38 (s, 1H), 8.05 (d, J=9.0Hz, 2H), 8.04 (s, 1H), 7.84 (s, 1H),
(7.83 d, J=9.0Hz, 2H), 7.67 (s, 1H), 7.42 (d, J=7.8Hz, 4H), 6.53 (s, 2H), 3.79 (s, 3H)；13C
NMR(150MHz,DMSO-d6)δ164.9,156.2,145.0,138.1,135.5,131.1,130.4,129.7,129.3,
128.5(2C),128.4(2C),127.3,121.8(2C),120.3(2C),119.8,114.9,38.5.ESI-MS m/z
421.1 for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I k.
Embodiment 14：4-O- (3- amino -6- (1- methyl-1 H- pyrazolyls -4) pyrazinyl -2)-benzoic acid 3- chlorphenyl acyls
The preparation of amine
Operating process is same with case study on implementation 11, only uses (4,4,5,5- tetramethyls -1,3,2- dioxaborolans -2-
Base) -1H- pyrazoles replace 3,5- dimethyl isoxazole -4- boric acid, obtain white solid.Product detection data are as follows：
Yield：71%；Purity:99% (HPLC:MeOH:H2O=70:30,0.5mL/min,tR=10.05min)；1H-
NMR(600MHz,DMSO-d6) δ 10.40 (s, 1H), 8.06 (d, J=9.6Hz, 2H), 8.04 (s, 1H), 7.98 (t, J=
1.8Hz, 1H), 7.84 (s, 1H), 7.72 (d, J=8.4Hz, 1H), 7.67 (s, 1H), 7.42 (d, J=9.0Hz, 2H), 7.39
(t, J=8.4Hz, 1H), 7.38 (s, 1H), 7.17 (d, J=8.4Hz, 1H), 6.53 (s, 2H), 3.80 (s, 3H)；13C-NMR
(150MHz,DMSO-d6)δ165.0,156.3,145.0,140.6,135.5,132.9,132.3,131.1,130.3,130.2,
129.3(2C),127.3,123.2,120.3(2C),119.8,119.6,118.6,114.9,38.5.ESI-MS m/z 421.2
for[M+H].
It carries out in aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I l.
Embodiment 15：2- amino -3,5- two replaces in vitro cytotoxic effect of the pyrazine compound to cancer cell
In order to study the ability that target compound synthesized in the present invention inhibits tumor cell proliferation, compound is determined
To four kinds of human tumor cells (human glioma cells U87, human cervical carcinoma cell HeLa, human liver cancer cell HepG2 and human colon carcinoma
Cell LoVo) in vitro cytotoxic effect, and using VX-680 as positive control.The detection method that experiment uses is standard
Mtt assay.Experimental method is：
It takes the logarithm the cell in growth period, cell suspension is made with 1640 culture mediums of RPMI containing 10% fetal calf serum, per hole
In 6000 cell inoculations to 96 orifice plates, tablet is put into 37 DEG C, containing 5%CO2The incubator culture of air and 100% humidity is for 24 hours
Make its adherent, rear substitution contains 1640 culture mediums of RPMI (200 μ L/ holes) containing 10% fetal calf serum of various concentration drug, medicine
Object concentration is respectively 10-4, 2 × 10-5, 4 × 10-6, 8 × 10-7, 1.6 × 10-7,3.2×10-8Mol/L, and zeroing hole is set, blank
Group, positive controls VX-680, every group of three wells are taken out after being incubated culture 72 hours, and 20 μ L MTT (5mg/mL) are added in per hole,
It is incubated culture 4h again, MTT is made to be reduced to Jia Za, supernatant is sucked out, 150 μ L DMSO are added in per hole, concussion Shi Jia Za crystal is molten
Solution measures OD value of the cell liquid at 570nm with microplate reader, calculates the inhibiting rate of each concentration of compound.GI is calculated by inhibiting rate50
Value, and take the average value tested three times.
Chemical compounds I a-l are to human glioma cells U87, human cervical carcinoma cell HeLa, human liver cancer cell HepG2 and people's colon
The in-vitro pharmacological experiments that tetra- kinds of cancer cell multiplications of cancer cell LoVo inhibit the results are shown in Table 1.
The Aurora kinase inhibitory activity and tumor cell proliferation inhibition activity of table 1 chemical compounds I a-l and VX-680
Note：(1) experimental result is the statistical result of parallel laboratory test three times；(2) action time：72 hours
Experiment in vitro proves, 12 compounds of synthesis are to U87, HeLa, HepG2 and LoVo all than positive control VX-680
There is preferable external inhibitory activity；Especially chemical compounds I h and I i shows HeLa and LoVo cells optimal inhibition and lives
Property.
Embodiment 16：Chemical compounds I a-l are to the inhibitory activity of laser kinases
In order to probe into the mechanism of the antitumor proliferation functions of chemical compounds I a-l, while confirm it whether can be to Aurora A
Inhibitory activity, using Kinase-Glo luminescent kinase assay (Assay Drug Dev.Technol.,
2004,2,153-160) the external inhibitory activity of Aurora A is studied.
Specific method：In each aperture of 96 orifice plates, 10ng kinases to be measured (Aurora-A, B), 2 μ g substrates are taken
The untested compound of Kemptide and gradient concentration (0.01nmol/L, 0.1nmol/L, 1nmol/L, 10nmol/L,
100nmol/L) it is blended in phosphorylation reaction buffer solution (40mmol/L Tris, the 10mmol/L MgCl that 35 μ L are prepared in advance2,
1mmol/L DTT, 0.1mg/mL BSA, 10 μm of ol/L ATP, pH7.4) in, 37 DEG C of culture 0.5h, every group of concentration is respectively provided with three
A secondary orifices.Then 50 μ L Kinase-Glo reagents are added in be uniformly mixed, 10min are placed at room temperature, by the phase for recording reactant
Remaining ATP contents are detected to Fluorescence Unit values (Relative Luminescence Unit, RLU).Finally use
Origin v7.0 softwares come calculate target compound inhibit Aurora A IC50Value.
Experimental result is shown in Table 1.
By table 1 as it can be seen that chemical compounds I a-l have stronger inhibitory activity to Aurora A and Aurora B, but to two kinds
Kinases is without apparent selective trend.
The disubstituted 2- amino-pyrazinos compound synthesis methods of 3,5- of the present invention are simple, and raw material is cheap and easy to get, and pharmacology is lived
Property it is notable, be expected to become potentially possess China's independent intellectual property right a kind for the treatment of cancer newtype drug.
Comparative example
Comparison of therapeutic
It is compared with VX-680, the compound of the present invention is to human glioma cells U87, human cervical carcinoma cell HeLa, human liver cancer
Tetra- kinds of cancer cell multiplication inhibitory activity of cell HepG2 and human colon cancer cell LoVo are better than positive reference compound VX-680.
Wherein chemical compounds I h be respectively to the inhibitory activity of tetra- kinds of cancer cells of U87, HeLa, HepG2 and LoVo VX-680 5.6,19.5,
3.9 and 25.6 times；Chemical compounds I i is VX-680 respectively to the inhibitory activity of tetra- kinds of cancer cells of U87, HeLa, HepG2 and LoVo
4.2nd, 32,7.8 and 14.4 times.And it is respectively 57 times and 10 times of VX 680 to the inhibitory activity of Aurora B.
The foregoing is only a preferred embodiment of the present invention, is not intended to restrict the invention, although with reference to aforementioned reality
Example is applied the present invention is described in detail, it for those skilled in the art, still can be to aforementioned each implementation
Technical solution recorded in example modifies or carries out equivalent replacement to which part technical characteristic.All essences in the present invention
With within principle, any modification, equivalent replacement, improvement and so on should all be included in the protection scope of the present invention god.
Claims (7)
1. one kind 3, the disubstituted 2- amino-pyrazinos compounds of 5-, which is characterized in that the compound has following structure shown in formula I：
In formula I：
NR1R2For n-propylamine base, cyclopropyl amino, cyclopentamine base, cyclohexylamino, morpholinyl, 4- methyl piperazines base, anilino-, 4- chlorine
Anilino- or 3- chloroanilinos；
Ar is 3,5- dimethyl isoxazoles base -4 or 1- methyl-1 H- pyrazolyls -4.
2. the preparation process of the disubstituted 2- amino-pyrazinos compounds of 3,5- described in claim 1, which is characterized in that described
Technique for 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic amides and 3,5- dimethyl isoxazole -4- boric acid or (4,4,5,
5- tetramethyl -1,3,2- dioxaborolan -2- bases) the generation Suzuki condensation reaction of -1H- pyrazoles.
3. the preparation process of the disubstituted 2- amino-pyrazinos compounds of 3,5- according to claim 2, which is characterized in that
In the technique, catalyst is [1,1'- bis- (diphenylphosphine) ferrocene] palladium chloride complex compound, [1,2- bis- (diphenylphosphines)
Ethane] palladium chloride complex compound or [bis- (diphenylphosphine) propane of 1,3-] palladium chloride complex compound.
4. the preparation process of the disubstituted 2- amino-pyrazinos compounds of 3,5- according to claim 2, which is characterized in that
The preparation method of 4-O- (the bromo- pyrazinyls -2 of 3- amino -6-)-benzoic amide is using two bromo- pyrazines of 2- amino -3,5- as original
Material and the positive propionamide of P-hydroxybenzoic acid, P-hydroxybenzoic acid ring propionamide, P-hydroxybenzoic acid ring pentanamide, para hydroxybenzene
Acid cyclohexyl amide, P-hydroxybenzoic acid morpholino amide, P-hydroxybenzoic acid 4- methyl piperazines amide, P-hydroxybenzoic acid benzoyl
Amine, the P-hydroxybenzoic acid chlorobenzamide between chlorobenzamide or P-hydroxybenzoic acid react.
5. the preparation process of the disubstituted 2- amino-pyrazinos compounds of 3,5- according to claim 4, which is characterized in that
Action solvent is N-Methyl pyrrolidone, N- ethyl pyrrolidones or N- cyclohexyl pyrrolidones.
6. application of the disubstituted 2- amino-pyrazinos compounds of 3,5- described in claim 1 as aurora kinase inhibitors.
7. application of the disubstituted 2- amino-pyrazinos compounds of 3,5- described in claim 1 in anticancer drug.
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| NZ562953A (en) * | 2002-06-20 | 2009-06-26 | Vertex Pharma | Processes for preparing substituted pyrimidines and pyrimidine derivatives as inhibitors of protein kinases |
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