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Application of 7-hydroxycoumarin pyrazoline derivatives in the preparation of grp94 inhibitors

An inhibitor, pyrazole technology, applied in the field of coumarin pyrazoline fluorescent compounds

Active Publication Date: 2019-12-13
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] After searching and searching by authoritative organizations, using the above-mentioned coumarin pyrazoline derivative 3-(1,5-diphenyl-4,5-dihydro-1H-pyrazol-3-yl)-7-hydroxyl-2H The research and application of -chromen-2-one as a GRP94 inhibitor has not been reported at home and abroad

Method used

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  • Application of 7-hydroxycoumarin pyrazoline derivatives in the preparation of grp94 inhibitors
  • Application of 7-hydroxycoumarin pyrazoline derivatives in the preparation of grp94 inhibitors
  • Application of 7-hydroxycoumarin pyrazoline derivatives in the preparation of grp94 inhibitors

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Preparation of 3-(1,5-diphenyl-4,5-dihydro-1H-pyrazol-3-yl)-7-hydroxyl-2H-chromene-2-one

[0043] Dissolve 2,4-dihydroxybenzaldehyde (20mmol) and ethyl acetoacetate (24mmol) in 30ml of ethanol solution, and add 1ml of piperidine dropwise as a catalyst. The reaction solution was heated to reflux for 3h to the end of the reaction, and a large amount of grass-green solid was precipitated after cooling to room temperature. The crude product was obtained by suction filtration, and recrystallized with ethanol to obtain the green product 7-hydroxy-3-acetylcoumarin.

[0044] 7-Hydroxy-3-acetylcoumarin, benzaldehyde and a catalytic amount of piperidine (about 1ml) were dissolved in 20ml of absolute ethanol, and the reaction solution was refluxed for 4h and cooled to room temperature. Since the conversion rate of this reaction is low under weak alkaline conditions, 10 equivalents of benzaldehyde is added to react, and the reaction is carried out thoroughly. precipita...

Embodiment 2

[0046] Example 2: HCP1 interacts directly with GRP94

[0047] Surface plasmon resonance analysis by Biacore T200 was used to detect the binding of HCP1 to full-length GRP94 protein. The full-length GRP94 protein was immobilized on the surface of the CM5 chip by amino coupling method. HCP1 was injected at 2.74 μM, 1.83 μM, 1.22 μM, 0.81 μM and 0.54 μM, respectively. The injection time of HCP1 is 120s, the flow rate is 10ul / min, and the dissociation time is 600s. The injection buffer was PBS (10 mM phosphate, 137 mM NaCl, 2.68 mM KCl, 0.1% dimethyl sulfoxide [v / v], pH 8.0), and the temperature was 25°C. Chip regeneration was performed using 10 mM NaOH after each injection. The combined response value was the RU value, and the blank control was deducted from the data processing, and the Biacore T200 evaluation software was used for 1:1 binding simulation analysis.

[0048] The results show that: the response value obtained by Biacore T200 has a concentration-corresponding rel...

Embodiment 3

[0049] Example 3: Colocalization of HCP1 and GRP94

[0050] A549 cells were seeded in glass culture dishes with a diameter of 3.5 cm at 37°C in CO 2 After culturing in the incubator for 24 hours, add HCP1 (10 μM) for 3 hours, remove the culture medium, wash twice with 0.1M PBS buffer, add 1640 stock solution, and place under a laser confocal microscope with different wavelengths of excitation light (405nm, 488nm, 546nm, 633nm and 647nm) to take pictures.

[0051] A549 cells were seeded in 3.5 cm diameter glass-bottom culture dishes at 37°C in CO 2 After culturing in the incubator for 24 hours, add HCP1 (10 μM) to treat for 3 hours, remove the culture medium, wash twice with 0.1M PBS, and fix the cells with 4% paraformaldehyde for 15 minutes. Discard 4% paraformaldehyde, wash 3 times with 0.1M PBS buffer, 5min each time; discard 0.1M PBS buffer, add normal serum blocking solution, block at room temperature for 20min; discard blocking solution, add GRP94 primary antibody (use ...

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Abstract

The invention discloses application of 3-(1,5-diphenyl-4,5-dihydro-1H-pyrazole-3-yl)-7-hydroxy-2H-chroman-2-one in preparing GRP94 inhibitors. The concentration of the 3-(1,5-diphenyl-4,5-dihydro-1H-pyrazole-3-yl)-7-hydroxy-2H-chroman-2-one which effectively inhibits the biological functions of GRP94 is 1-10 micromole. According to the application, a foundation is laid for researching and developing the inhibitors for the GRP94, and the compound can also be used as an effective chemical tool to provide a powerful means for research of the biological functions of the GRP94.

Description

technical field [0001] The present invention relates to the application of a coumarin pyrazoline fluorescent compound, in particular to 7-hydroxycoumarin pyrazoline derivatives namely 3-(1,5-diphenyl-4,5-dihydro-1H- Application of pyrazol-3-yl)-7-hydroxyl-2H-chromen-2-one in the preparation of GRP94 inhibitors. Background technique [0002] GRP94 (Glucose Regulatory Protein 94) is encoded by the HSP90B1 gene, is a subtype of HSP90 (Heat Shock Protein 90) in the endoplasmic reticulum, and is highly conserved with HSP90 in structure. GRP94 is mainly involved in the folding and assembly of endocrine and membrane proteins in cells. Clinical studies have shown that GRP94 is involved in the development and metastasis of a series of cancers such as melanoma, ovarian cancer, multiple myeloma, and lung cancer. GRP94 has become a new target for tumor treatment. In addition, GRP94 is also involved in autoimmunity and inflammation-related diseases. The molecular chaperone function of...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07D405/04A61K31/4155A61P35/00
CPCC07D405/04
Inventor 苗俊英赵宝祥张尚立苏乐魏群
Owner SHANDONG UNIV
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