CN110903292A - Multi-target inhibitor acting on QC and GSK-3 β - Google Patents

Multi-target inhibitor acting on QC and GSK-3 β Download PDF

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CN110903292A
CN110903292A CN201911183787.9A CN201911183787A CN110903292A CN 110903292 A CN110903292 A CN 110903292A CN 201911183787 A CN201911183787 A CN 201911183787A CN 110903292 A CN110903292 A CN 110903292A
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吴海强
谢亚洲
余熙
欧阳娜
尚琦
邹浩曼
贺震旦
刘志刚
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Abstract

The invention discloses a multi-target inhibitor acting on QC and GSK-3 β, wherein the structural general formula of the multi-target inhibitor is as follows:
Figure DDA0002291932110000011
the invention integrates various high-purity proteins according to the active center crystal structures of target QC and GSK-3 β enzyme proteinsThe active pharmacophore is used for preparing a multi-target inhibitor which can simultaneously act on QC and GSK-3 β through framework transition and recombination design, the multi-target inhibitor is a molecule with multiple target points and high activity, the molecular structure diversity of a lead drug is remarkably expanded, and the research and development of innovative anti-AD drugs and AD diagnostic kits are actively promoted.

Description

Multi-target inhibitor acting on QC and GSK-3 β
Technical Field
The invention relates to the field of pharmaceutical chemistry, in particular to a multi-targeting inhibitor acting on QC and GSK-3 β.
Background
Alzheimer's Disease (AD) is a common neurodegenerative disease, the major form of senile dementia, with more than 65% of AD patients in total. Clinical symptoms of AD include progressive memory and cognitive dysfunction, which are characterized by irreversible, high morbidity, high mortality, and the like, and have become the third world of sexual health, economic, and social problems. However, the exact pathological mechanism of AD is unknown at present, no specific treatment medicine exists clinically, and research on innovative anti-AD medicines is urgent.
Disclosure of Invention
In view of the defects of the prior art, the invention aims to provide a multi-target inhibitor acting on QC and GSK-3 β, and aims to solve the problems of single action target, insufficient molecular structure diversity, limited molecular activity and low lead compound druggability of the existing anti-AD drug research.
The technical scheme of the invention is as follows:
a multi-targeting inhibitor acting on QC and GSK-3 β, wherein the structural formula is as follows:
Figure BDA0002291932090000011
wherein R is1Is one of hydrogen, alkyl, nitro, amido, halogen, sulfonic acid group or ester group, R1The linked group is one of a benzene ring, a six-membered heteroaromatic ring, a five-membered heteroaromatic ring or a seven-membered heteroaromatic ring; r2Is one of hydrogen, alkoxy, nitro, amido, halogen, sulfonic acid group or ester group, R2The linked group is one of a benzene ring, a six-membered heteroaromatic ring, a five-membered heteroaromatic ring or a seven-membered heteroaromatic ring.
The multi-target inhibitor acting on QC and GSK-3 β, wherein R1And R1May be identical or different.
The multi-target inhibitor acting on QC and GSK-3 β, wherein R1Linked group and R1The linked groups may be identical or different in structure.
The multi-target inhibitor for QC and GSK-3 β, wherein n is 1-3.
The multi-target inhibitor acting on QC and GSK-3 β has a structural formula
Figure BDA0002291932090000021
Figure BDA0002291932090000031
Figure BDA0002291932090000032
But is not limited thereto.
The invention has the beneficial effects that a multi-target inhibitor which can simultaneously act on QC and GSK-3 β is prepared by integrating various high-activity pharmacophores according to the active center crystal structures of target QC and GSK-3 β enzyme proteins and through framework transition and recombination design, the multi-target inhibitor is a molecule with multiple target points and high activity, the diversity of the molecular structure of a lead medicament is remarkably expanded, and the research and development of innovative anti-AD medicaments and AD diagnosis kits are actively promoted.
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FIG. 1 is a schematic diagram of the synthetic route of the multi-targeted inhibitor of the present invention.
Detailed Description
The present invention provides a multi-targeted inhibitor acting on QC and GSK-3 β, which will be described in further detail below for the purpose of making the objects, technical solutions and effects of the present invention clearer and more clear.
The AD is a complex disease caused by multiple factors, and researches show that the regional and specific up-regulation of glutaminyl cyclase (QC) and glycogen synthase kinase-3 (GSK-3) has important inducing and promoting effects in different stages of AD pathogenesis and is an important target point for anti-AD drug research, wherein the QC is an enzyme for catalyzing the intramolecular cyclization reaction of N-terminal glutamine such as polypeptide, protein and the like to generate pyroglutamic acid (pE), has important biological functions of changing the N-terminal chemical structure, regulating activity, enhancing stability and the like, however, in the early stage of the pathogenesis, the AD patient has the characteristics of high specificity and activity enhancement, the QC catalyzes and generates pE-A β (particularly pE-A β/pE-A β) formed by the intramolecular cyclization of N-terminal glutamine residues, the A β variant has stronger neurotoxicity, higher aggregation and precipitation speed, once the variant cannot be degraded and cleared and the like, is more specific than A β, mainly exists in the patient, one of strategic brain (the AD is more than the AD inhibitor of AD in the AD, the AD receptor of AD 2, the clinical pathological factor exists in the early stage of the AD pathogenesis, the AD system, the AD is more than the clinical and the clinical research of anti-AD factor, the AD pathogenesis, the AD factor, the AD also can be more than the clinical and the clinical high expression of the clinical and the promotion of the AD factor of the AD.
Meanwhile, the research finds that GSK-3 (comprising GSK-3 α, GSK-3 β and the like) is a key kinase for regulating glycogen synthase, glycogen synthase activity is reduced by phosphorylation, in-vivo blood sugar concentration is increased, Bax/HKII ratio is regulated, mitochondrial permeability and cytochrome C release are influenced, and the over-activated GSK-3 can act on a plurality of signal proteins, structural proteins and transcription factors to regulate cell proliferation, differentiation, survival, apoptosis and the like, and plays an important role in the pathological process of a plurality of complex diseases such as type II diabetes, neurodegenerative diseases, cancer, chronic inflammation and the like.
As can be seen from the above analysis, the specific high expression of QC and GSK-3 β are important inducing and promoting factors in the pathogenesis of AD, and the research in this direction will inevitably develop and innovate a new field of anti-AD drug research.
Based on the structure, the invention synthesizes a plurality of high-activity pharmacophores according to the active center crystal structures of target QC and GSK-3 β enzyme proteins, prepares a multi-target inhibitor capable of simultaneously acting on QC and GSK-3 β through framework transition and recombination design, and has the following structural general formula:
Figure BDA0002291932090000051
wherein R is1Is one of hydrogen, alkyl, nitro, amido, halogen, sulfonic acid group or ester group, R1The linked group is one of a benzene ring, a six-membered heteroaromatic ring, a five-membered heteroaromatic ring or a seven-membered heteroaromatic ring; r2Is one of hydrogen, alkoxy, nitro, amido, halogen, sulfonic acid group or ester group, R2The linked group is one of a benzene ring, a six-membered heteroaromatic ring, a five-membered heteroaromatic ring or a seven-membered heteroaromatic ring.
The multi-target inhibitor provided by the embodiment is a molecule with multiple target points and high activity, remarkably enlarges the molecular structure diversity of a lead medicament, and actively promotes the research of innovative anti-AD medicaments and AD diagnosis kits.
In some embodiments, the R is1And R1Are the same or different in structure, the R is1Linked group and R1The linked groups may be identical or different in structure.
In some embodiments, n ═ 1 to 3 in the general structural formula of the multi-targeted inhibitor. By way of example, the multi-targeted inhibitor may be
Figure BDA0002291932090000061
Figure BDA0002291932090000062
Figure BDA0002291932090000071
Figure BDA0002291932090000072
But is not limited thereto.
The compound is a QC and GSK-3 β multi-target inhibitor which is disclosed for the first time, and has extremely important scientific significance and research value for researching autonomous innovation AD-resistant lead medicaments in China.
In some embodiments, there is also provided a method for preparing a multi-targeted inhibitor acting on QC and GSK-3 β, comprising the steps of:
to anhydrous CH2Cl2Adding into solvent
Figure BDA0002291932090000073
Anhydrous AlCl3And methoxy oxalyl chloride solution, and reacting to obtain
Figure BDA0002291932090000074
To anhydrous CH2Cl2Adding into solvent
Figure BDA0002291932090000081
Oxalyl chloride solution and DMF, replacing the solvent in the reaction system with anhydrous THF after the reaction is carried out for the first preset time, adding concentrated ammonia water and continuing the reaction for the second preset time to prepare the intermediate product
Figure BDA0002291932090000082
Will be provided with
Figure BDA0002291932090000083
And adding the t-BuOK solution into anhydrous THF, reacting at room temperature for a third preset time, adding concentrated hydrochloric acid, and continuously reacting for a fourth preset time to obtain the compound
Figure BDA0002291932090000084
Adding into 1-butyl-3-methylimidazole hexafluorophosphate solution
Figure BDA0002291932090000085
Figure BDA0002291932090000086
Continuously dropwise adding KOH solution after ice bath cooling, reacting for fifth preset time at 50-100 ℃ to obtain the multi-target inhibitor acting on QC and GSK-3 β, wherein the chemical structural formula of the multi-target inhibitor is shown in the specification
Figure BDA0002291932090000087
Wherein R is1Is one of hydrogen, alkyl, nitro, amido, halogen, sulfonic acid group or ester group, R1The linked group is one of a benzene ring, a six-membered heteroaromatic ring, a five-membered heteroaromatic ring or a seven-membered heteroaromatic ring; r2Is one of hydrogen, alkoxy, nitro, amido, halogen, sulfonic acid group or ester group, R2The linked group is benzene ring, six-membered heteroaromatic ring, five-memberedOne of a heteroaromatic ring or a seven membered heteroaromatic ring.
In some embodiments, the
Figure BDA0002291932090000088
The preparation method comprises the following steps:
will be provided with
Figure BDA0002291932090000091
Dissolving in formic acid, reacting for 2-6h at the temperature of 100-150 ℃ to obtain
Figure BDA0002291932090000092
Adding into anhydrous THF solvent
Figure BDA0002291932090000093
And LiAlH4And stirring for reaction to obtain
Figure BDA0002291932090000094
To SOCl2Adding in
Figure BDA0002291932090000095
Stirring for 1-4h at 50-100 deg.C, and reacting to obtain
Figure BDA0002291932090000096
In some embodiments, the first predetermined time is 3-5 hours; the second preset time is 0.5-2 h; the third preset time is 1-14 h; the fourth preset time is 1-3 h.
In some embodiments, there is also provided a use of a multi-targeted inhibitor, wherein the multi-targeted inhibitor is used in an anti-AD drug.
In some embodiments, there is also provided a use of a multi-targeted inhibitor, wherein the multi-targeted inhibitor is used in an AD diagnostic kit.
In some embodiments, the application of the multi-target inhibitor is further provided, wherein the multi-target inhibitor is applied to the prevention and treatment of QC and/or GSK-3 β specific high expression related diseases.
In some embodiments, the application of the multi-target inhibitor is further provided, wherein the multi-target inhibitor is applied to a diagnostic kit for QC and/or GSK-3 β specific high expression related diseases.
The following examples further illustrate a multi-targeted inhibitor of the invention, capable of acting on QC and GSK-3 β, and the assay of the inhibitory activity of the inhibitor on QC and GSK-3 β enzymes:
example 1
FIG. 1 is a schematic diagram of the synthetic route of the multi-target inhibitor of this example.
(1) Synthesis of intermediate III:
a. with anhydrous CH2Cl2Adding 1mol of raw material I as a solvent, and adding not less than 8mol of anhydrous AlCl3Stirring for 0.5h, dropwise adding not less than 3mol of methoxy oxalyl chloride solution II, stirring at room temperature for reaction for 6-10h, adding excessive saturated NaHCO3Extracting the solution with ethyl acetate for 3 times, combining the organic phases, and adopting anhydrous Na2SO4Drying, concentrating, preparing a product III by column chromatography, calculating the yield, and identifying the structure;
(2) synthesis of intermediate VI:
b. with anhydrous CH2Cl2Cooling the solvent in an ice bath, adding 1mol of the raw material IV, stirring for dissolving, dropwise adding not less than 4mol of oxalyl chloride solution, dropwise adding 2 drops of DMF, stirring to room temperature, reacting for 3-5h, and evaporating to remove the solvent; replacing the solvent of the reaction system with anhydrous THF, cooling in ice bath, dropwise adding concentrated ammonia water of not less than 10 mol, reacting for 0.5-2H, and adding excessive H into the system2Extracting with ethyl acetate for 3 times, mixing organic phases, and extracting with anhydrous Na2SO4Drying, concentrating, preparing a product V by column chromatography, calculating the yield, and identifying the structure;
c. taking anhydrous THF as a solvent, adding 1 quantitative intermediate product III and 1.2 mol of intermediate product V, cooling in an ice bath, stirring and dropwise adding t-BuOK solution with the molar weight not less than 10 mol, reacting at room temperature for 1-14h under the protection of nitrogen,adding concentrated hydrochloric acid, reacting at room temperature for 1-3h, adding 10% NaHCO3Stopping reaction with solution, extracting with ethyl acetate for 3 times, mixing organic phases, and adding anhydrous Na2SO4Drying, concentrating, preparing a product VI by column chromatography, calculating the yield, and identifying the structure;
(3) synthesis of target product XI:
d. dissolving 3, 4-diaminobenzoic acid VII by using excessive formic acid, reacting for 2-6h at 110 ℃, evaporating to remove formic acid after the reaction is finished, adding water, carrying out suction filtration, drying to obtain a solid, namely a product VIII, and calculating the yield.
e. Adding 1mol of intermediate product VIII by taking anhydrous THF as a solvent, cooling in an ice bath, and adding excessive LiAlH4Stirring for 1-3h, returning to room temperature, refluxing at 70 deg.C for 2-6h, pouring into pre-cooled Na2SO4Extracting with ethyl acetate for 3 times, mixing organic phases, and adding anhydrous Na2SO4Drying and concentrating to obtain an intermediate product VIIII, and calculating the yield.
f. In excess of SOCl2Adding a certain amount of intermediate product VIIII, stirring at 80 ℃ for 1-4h, and quickly evaporating excessive SOCl2And obtaining an intermediate product X for the next reaction.
g. Taking 1-butyl-3-methylimidazolium hexafluorophosphate solution as a reaction system, adding 1mol of intermediate product VI and 1.2 mol of intermediate product X, cooling in ice bath, dropwise adding a certain amount of KOH solution while stirring, reacting at 80 ℃ for 1-3h, extracting for 3 times with ethyl acetate, combining organic phases, and adopting anhydrous Na2SO4Drying, concentrating, preparing a target product XI by column chromatography, calculating the yield, and identifying the structure.
Example 2
Preparation of 3- (5- ((1H-benzo [ d ] imidazol-5-yl) methyl) -5H-pyrrolo [3,2-d ] pyrimidin-7-yl) -4-phenyl-1H-pyrrole-2, 5-dione according to the synthetic route of example 1:
a. 20ml of anhydrous CH2Cl2Adding 0.105Mol of 5H-pyrrolo [3,2-d ] into the mixture]Pyrimidine, 1.107Mol of anhydrous AlCl is added3After stirring for 0.5h, 0.439Mol of methoxy oxalyl chloride solution is added dropwise, the mixture is stirred at room temperature for reaction for 6.5h, and 30ml of saturated NaHCO is added3The solution is prepared by mixing a solvent and a solvent,extracting with ethyl acetate 20ml times by 3 times, combining organic phases, and extracting with anhydrous Na2SO4Drying, concentrating, and performing column chromatography to obtain 2-oxo-2 (5H-pyrrolo [3, 2-d)]Pyrimidin-7-yl) acetic acid methyl ester, yield 72%;
b. ice bath cooled 20ml anhydrous CH2Cl2Adding 0.114Mol phenylacetic acid, stirring for dissolving, dropwise adding 0.524Mol oxalyl chloride solution, dropwise adding 2 drops of DMF, stirring to room temperature, reacting for 3h, evaporating to remove solvent, adding 15ml anhydrous THF, cooling in ice bath, dropwise adding 60ml concentrated ammonia water, reacting for 0.5h, adding 20ml H2O, ethyl acetate extraction 30ml times 3 times, organic phase combination, anhydrous Na2SO4Drying, concentrating, and performing column chromatography to obtain 2-phenylacetamide with yield of 82%;
c. 20ml of anhydrous THF was added 0.050Mol of 2-oxo-2 (5H-pyrrolo [3,2-d ]]Pyrimidine-7-yl) methyl acetate and 0.060Mol of 2-phenylacetamide are cooled in an ice bath, 0.065Mol of t-BuOK solution is dropwise added while stirring, nitrogen is protected, after the reaction is carried out for 6h at room temperature, 0.5ml of concentrated hydrochloric acid is added, the reaction is continued for 1h at room temperature, and excess 10 percent NaHCO is added3The reaction was stopped, extracted 20ml X3 times with ethyl acetate, the organic phases were combined and Na anhydrous2SO4Drying, concentrating, and performing column chromatography to obtain 3-phenyl-4- (5H-pyrrolo [3, 2-d)]Pyrimidin-7-yl) -1H-pyrrole-2, 5-dione, yield 63%;
d. adding 0.124Mol of 3, 4-diaminobenzoic acid into 15ml of formic acid, reacting for 4H at 110 ℃, evaporating to remove the formic acid, adding 20ml of water, filtering, and drying to obtain the 1H-benzo [ d ] imidazole-6-carboxylic acid with the yield of 88%.
e. 20ml of anhydrous THF was added 0.102Mol of 1H-benzo [ d ]]Imidazole-6-carboxylic acid, ice-cooled, added with 0.437Mol of LiAlH4Stirring for 2h, returning to room temperature, refluxing at 70 deg.C for 5h, pouring 30ml pre-cooled Na2SO4In solution, extracting 20ml × 3 times with ethyl acetate, combining organic phases, anhydrous Na2SO4Drying and concentrating to obtain (1H-benzo [ d ]]Imidazol-6-yl) methanol, yield 91%.
f、5ml SOCl2To this was added 0.106Mol (1H-benzo [ d ]]Imidazol-6-yl) methanol, stirred at 80 ℃ for 3h, excess SOCl was quickly distilled off2Obtaining 6- (chloromethyl) -1H-benzo [ 2 ]d]Imidazole for the next reaction.
g. 20ml of 1-butyl-3-methylimidazolium hexafluorophosphate solution was added with 0.087Mol of 3-phenyl-4- (5H-pyrrolo [3, 2-d)]Pyrimidin-7-yl) -1H-pyrrole-2, 5-dione and 6- (chloromethyl) -1H-benzo [ d ] prepared in situ in f]Imidazole, ice-bath cooling, stirring and dropping 1ml KOH solution (0.2Mol/L), reacting at 80 deg.C for 2.5h, extracting with ethyl acetate for 20ml times multiplied by 3 times, combining organic phases, anhydrous Na2SO4Drying, concentrating, and performing column chromatography to obtain target product 3- (5- ((1H-benzo [ d)]Imidazol-5-yl) methyl) -5H-pyrrolo [3,2-d]Pyrimidin-7-yl) -4-phenyl-1H-pyrrole-2, 5-dione having the chemical formula:
Figure BDA0002291932090000121
the overall yield was 23%.
Example 3
Preparation of 3- (1- ((1H-benzo [ d ] imidazol-5-yl) methyl) -5-nitro-1H-indol-3-yl) -4- (4-methoxyphenyl) -1H-pyrrole-2, 5-dione according to the synthetic route of example 1:
a. 20ml of anhydrous CH2Cl2Adding 0.112Mol of 5-nitro-1H-indole, adding 1.326Mol of anhydrous AlCl3Stirring for 0.5h, then dropwise adding 0.513Mol of methoxy oxalyl chloride solution, stirring at room temperature for reaction for 8h, adding 30ml of saturated NaHCO3The solution was extracted 20ml X3 times with ethyl acetate, the organic phases were combined and Na anhydrous2SO4Drying, concentrating, and performing column chromatography to obtain 2- (5-nitro-1H-indol-3-yl) -2-oxoacetic acid methyl ester with yield of 53%;
b. ice bath cooled 20ml anhydrous CH2Cl2Adding 0.108Mol of p-methoxyphenylacetic acid, stirring for dissolving, dropwise adding 0.493Mol of oxalyl chloride solution, dropwise adding 2 drops of DMF, stirring to room temperature, reacting for 2.5h, evaporating to remove the solvent, adding 15ml of anhydrous THF, cooling in ice bath, dropwise adding 60ml of concentrated ammonia water, reacting for 1h, adding 20ml of H2O, ethyl acetate extraction 30ml times 3 times, organic phase combination, anhydrous Na2SO4Drying, concentrating, and performing column chromatography to obtain 2- (4-methoxyphenyl) acetamido with yield of 80%;
c. 20ml of anhydrous THF was added 0.050Mol of 2- (5-nitro-1H-indole-3-yl) -2-oxoacetic acid methyl ester and 0.060Mol of 2- (4-methoxyphenyl) acetamide, cooling in ice bath, stirring and dripping 0.065Mol of t-BuOK solution, reacting for 8h at room temperature under the protection of nitrogen, adding 0.5ml of concentrated hydrochloric acid, continuing to react for 1h at room temperature, and adding excess 10% NaHCO3The reaction was stopped, extracted 20ml X3 times with ethyl acetate, the organic phases were combined and Na anhydrous2SO4Drying, concentrating, and performing column chromatography to obtain 3- (4-methoxyphenyl) -4- (5-nitro-1H-indol-3-yl) -1H-pyrrole-2, 5-dione with yield of 67%;
d. adding 0.131Mol of 3, 4-diaminobenzoic acid into 15ml of formic acid, reacting for 4H at 110 ℃, evaporating to remove the formic acid, adding 20ml of water, filtering, and drying to obtain the 1H-benzo [ d ] imidazole-6-carboxylic acid with the yield of 86%.
e. To 20ml of anhydrous THF was added 0.122Mol of 1H-benzo [ d ]]Imidazole-6-carboxylic acid, cooled in an ice bath, 0.514Mol of LiAlH was added4Stirring for 2h, returning to room temperature, refluxing at 70 deg.C for 5h, pouring 30ml pre-cooled Na2SO4In solution, extracting 20ml × 3 times with ethyl acetate, combining organic phases, anhydrous Na2SO4Drying and concentrating to obtain (1H-benzo [ d ]]Imidazol-6-yl) methanol, yield 93%.
f、5ml SOCl2To which was added 0.127Mol (1H-benzo [ d ]]Imidazol-6-yl) methanol, stirred at 80 ℃ for 3h, excess SOCl was quickly distilled off2To obtain 6- (chloromethyl) -1H-benzo [ d]Imidazole for the next reaction.
g. To 20ml of 1-butyl-3-methylimidazolium hexafluorophosphate solution were added 0.101Mol of 3- (4-methoxyphenyl) -4- (5-nitro-1H-indol-3-yl) -1H-pyrrole-2, 5-dione and the now prepared 6- (chloromethyl) -1H-benzo [ d ] in f]Imidazole, ice-bath cooling, stirring and dropping 1ml KOH solution (0.2Mol/L), reacting for 2h at 80 ℃, extracting 20ml multiplied by 3 times with ethyl acetate, combining organic phases and anhydrous Na2SO4Drying, concentrating and carrying out column chromatography to prepare a target product, wherein the chemical structural formula of the target product is as follows:
Figure BDA0002291932090000141
the overall yield was 28%.
Example 4
Preparation of 3- (1- ((1H-benzo [ d ] imidazol-5-yl) methyl) -5-fluoro-1H-indol-3-yl) -4- (1H-imidazol-5-yl) -1H-pyrrole-2, 5-dione according to the synthetic route of example 1:
a. 20ml of anhydrous CH2Cl2Adding 0.131Mol of 5-fluoro-1H-indole, and adding 1.403Mol of anhydrous AlCl3Stirring for 0.5h, dropwise adding 0.627Mol of methoxy oxalyl chloride solution, stirring at room temperature for reaction for 7h, adding 30ml of saturated NaHCO3The solution was extracted 20ml X3 times with ethyl acetate, the organic phases were combined and Na anhydrous2SO4Drying, concentrating, and performing column chromatography to obtain 2- (5-fluoro-1H-indol-3-yl) -2-oxoacetic acid methyl ester with yield of 53%;
b. ice bath cooled 20ml anhydrous CH2Cl2Adding 0.134Mol 2- (1H-imidazole-5-yl) acetic acid, stirring, dripping 0.704Mol oxalyl chloride solution, dripping 10 drops of DMF, stirring to room temperature, reacting for 4H, evaporating to remove solvent, adding 15ml anhydrous THF, cooling in ice bath, dripping 60ml concentrated ammonia water, reacting for 1.5H, adding 20ml H2O, ethyl acetate extraction 30ml times 3 times, organic phase combination, anhydrous Na2SO4Drying, concentrating, and performing column chromatography to obtain 2- (1H-imidazole-5-yl) acetamido with yield of 56%;
c. adding 0.050Mol of methyl 2- (5-fluoro-1H-indol-3-yl) -2-oxoacetate and 0.060Mol of 2- (1H-imidazol-5-yl) acetamido into 20ml of anhydrous THF, cooling in an ice bath, dropwise adding 0.080Mol of t-BuOK solution while stirring, reacting at room temperature for 11H under the protection of nitrogen, adding 0.7ml of concentrated hydrochloric acid, continuing to react at room temperature for 2H, and adding excess 10% NaHCO3The reaction was stopped, extracted 20ml X3 times with ethyl acetate, the organic phases were combined and Na anhydrous2SO4Drying, concentrating, and performing column chromatography to obtain 3- (5-fluoro-1H-indol-3-yl) -4- (1H-imidazole-5-yl) -1H-pyrrole-2, 5-dione with yield of 72%;
d. adding 0.152Mol of 3, 4-diaminobenzoic acid into 15ml of formic acid, reacting for 4 hours at 110 ℃, evaporating to remove the formic acid, adding 20ml of water, filtering, and drying to obtain the 1H-benzo [ d ] imidazole-6-carboxylic acid with the yield of 89%.
e. 20ml of anhydrous THF was added 0.119Mol of 1H-benzo [ d ]]Imidazole-6-carboxylic acid, cooled in an ice bath, 0.561Mol of LiAlH was added4Stirring for 2h, returning to room temperature, refluxing at 70 deg.C for 5h, and reactingThe reaction was poured into 30ml of pre-cooled Na2SO4In solution, extracting 20ml × 3 times with ethyl acetate, combining organic phases, anhydrous Na2SO4Drying and concentrating to obtain (1H-benzo [ d ]]Imidazol-6-yl) methanol, yield 91%.
f、5ml SOCl2To this was added 0.105Mol (1H-benzo [ d ]]Imidazol-6-yl) methanol, stirred at 80 ℃ for 3h, excess SOCl was quickly distilled off2To obtain 6- (chloromethyl) -1H-benzo [ d]Imidazole for the next reaction.
g. To 20ml of 1-butyl-3-methylimidazolium hexafluorophosphate solution were added 0.089Mol of 3- (5-fluoro-1H-indol-3-yl) -4- (1H-imidazol-5-yl) -1H-pyrrole-2, 5-dione and the now prepared 6- (chloromethyl) -1H-benzo [ d ] in f]Imidazole, ice-bath cooling, stirring and dropping 1ml KOH solution (0.2Mol/L), reacting for 3h at 80 ℃, extracting 20ml multiplied by 3 times with ethyl acetate, combining organic phases and anhydrous Na2SO4Drying, concentrating and carrying out column chromatography to prepare a target product, wherein the chemical structural formula of the target product is as follows:
Figure BDA0002291932090000151
the total yield thereof was 21%.
Example 5
QC enzyme inhibitory activity was tested using the compounds in table 1:
the enzyme activity test is carried out in a 96-well enzyme label plate, 200 mul of a pH8.0 Tris buffer system, 0.3mM NADH, 2.0mM freshly prepared Gln-Gln, 14mM α -ketoglutaric acid, 30U/ml glutamate dehydrogenase, 50mM Tris, pH8.0 buffer solution are adopted, finally, a mixed solution of 0.28 MuM recombinant human QC protein and 1 MuM inhibitor is added, an enzyme label is used for dynamically detecting the absorption value change of the NADH at the wavelength of 340nm within 15min after oscillating for 30 seconds, data acquisition is carried out every 30 seconds, the inhibition rate (IR and percent) of the inhibitor on the activity of QC enzyme is calculated according to the test result, the larger IR value indicates that the activity of the compound is higher, and the test result is shown in Table 1.
Example 6
GSK-3 β enzyme inhibitory activity was tested using the compounds in table 1:
the enzyme activity test is carried out in a 96-well enzyme label plate, 200 mul of pH7.5 buffer system of 25mM HEPES, 10mM MgAc2, 1mM dithiothreitol, 0.01% BSA, 40ng GSK-3 β, 100ng peptide substrate and 0.1 mul inhibitor are adopted, finally 500nM ATP solution is added to start the reaction, the reaction is incubated for 90min at 37 ℃, Kinase-Glo reagent (EDTA, 50 mul/well) is added to stop the reaction, the fluorescence value of the reaction system is detected by a fluorescence enzyme label analyzer after 10min, according to the test result, the inhibition rate (IR, percent) of the inhibitor on the GSK-3 β enzyme activity is calculated by taking 5% DMSO as a contrast, the larger IR value indicates that the activity of the compound is higher, and the test result is shown in Table 1.
QC and GSK-3 β enzyme inhibitory Activity test results for the compounds of Table 1
Figure BDA0002291932090000161
Figure BDA0002291932090000171
Figure BDA0002291932090000181
Figure BDA0002291932090000191
Figure BDA0002291932090000201
In conclusion, according to the active center crystal structures of target QC and GSK-3 β enzyme proteins, multiple high-activity pharmacophores are integrated, a multi-target inhibitor capable of acting on QC and GSK-3 β simultaneously is prepared through framework transition and recombination design, the multi-target inhibitor is a molecule with multiple target points and high activity, the diversity of the molecular structure of a lead medicament is remarkably expanded, and research and development of innovative anti-AD medicaments and AD diagnosis kits are actively promoted.
It is to be understood that the invention is not limited to the examples described above, but that modifications and variations may be effected thereto by those of ordinary skill in the art in light of the foregoing description, and that all such modifications and variations are intended to be within the scope of the invention as defined by the appended claims.

Claims (5)

1. A multi-targeting inhibitor acting on QC and GSK-3 β, characterized by the general structural formula:
Figure FDA0002291932080000011
wherein R is1Is one of hydrogen, alkyl, nitro, amido, halogen, sulfonic acid group or ester group, R1The linked group is one of a benzene ring, a six-membered heteroaromatic ring, a five-membered heteroaromatic ring or a seven-membered heteroaromatic ring; r2Is one of hydrogen, alkoxy, nitro, amido, halogen, sulfonic acid group or ester group, R2The linked group is one of a benzene ring, a six-membered heteroaromatic ring, a five-membered heteroaromatic ring or a seven-membered heteroaromatic ring.
2. The multi-targeted inhibitor for QC and GSK-3 β according to claim 1, wherein R is1And R1May be identical or different.
3. The multi-targeted inhibitor for QC and GSK-3 β according to claim 1, wherein R is1Linked group and R1The linked groups may be identical or different in structure.
4. The multi-targeted inhibitor for QC and GSK-3 β according to claim 1, wherein n is 1-3.
5. The multi-targeted inhibitor for QC and GSK-3 β according to claim 1, having the formula
Figure FDA0002291932080000012
Figure FDA0002291932080000021
Figure FDA0002291932080000031
Figure FDA0002291932080000032
One kind of (1).
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