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PDK1 inhibitor, composition containing inhibitor and application

A technology of composition and inhibitor, which is applied in the field of medicine, can solve the problems of peripheral nerve toxicity and other problems, and achieve obvious therapeutic effect and reasonable compatibility effect

Inactive Publication Date: 2017-08-18
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, people gradually discovered that long-term exposure to the drug can easily cause peripheral neurotoxicity

Method used

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  • PDK1 inhibitor, composition containing inhibitor and application
  • PDK1 inhibitor, composition containing inhibitor and application
  • PDK1 inhibitor, composition containing inhibitor and application

Examples

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preparation example Construction

[0044] The preparation method of the pharmaceutical composition of the present invention comprises the following steps:

[0045] Mix Laoniuhui, motherwort root, ginseng, pinellia chinensis, Magnolia officinalis and Citrus aurantium, and decoct in water with a mass 3-5 times the total amount of mixed medicinal materials for 0.8-1.5 hours to obtain decoction and herbal medicine. dregs, concentrating the decoction into an extract, drying the extract and pulverizing to obtain a first medicinal powder, drying and pulverizing the medicinal dregs to a 60-80 mesh medicinal dregs powder;

[0046] After mixing turmeric, velvety indigo, chuanxiong and white wax tree, pulverize to 60-80 mesh, and mix it with the medicine dregs powder and add until the quality is the mixed powder (specifically, the medicine dregs powder and turmeric, velvety wood indigo , Ligusticum chuanxiong and white wax tree seed) 6-10 times and the concentration is 95% ethanol, reflux extraction at 80-90 ℃ to obtain t...

Embodiment 1

[0051] Detection of the Effect of Dicoumarin on PDK1 Activity

[0052] S11: Purchase p-PDHA1 ELISA kit and PDK1 active enzyme from abcam.

[0053] S12: Prepare reagents (incubation, wash buffer) according to the instructions, collect, lyse and quantify SKOV3 cells.

[0054] S13: Add the quantified protein to the 96-well plate, 250mg / 50μl / well, cover the lid, incubate at room temperature for 2h (96-well plate centrifuge, 300rpm), suck off the liquid, wash 2 times (300μl 1*wash buffer ), after the last suction of the liquid, invert it on a clean paper to remove residual liquid.

[0055] S14: preparing a phosphorylation solution and adding PDK1;

[0056] Add 22ml1*wash buffer, MgCl2 (1mM), ATP (2mM) into a clean 50ml centrifuge tube. PDK1 was added according to the design of the experimental group. (incubate PDK1 2μg / well in EP tube containing 200μl phosphorylation solution, DCA50mM+PDK1 2μg / well, DMSO 2%+PDK1 2μg / well, dicoumarol 100μM+PDK1 2μg / well, dicoumarol 200μM+ PDK1 ...

Embodiment 2

[0062] The effect of dicoumarin on the activity of ovarian cancer cell skov3 detected by MTT method

[0063] S21: Digest the skov3 cells in the logarithmic growth phase, and count the cells.

[0064] S22: Take a 96-well cell culture plate, add 100 μl containing 8×10 3 For the cell suspension of cells, only 100 μl of cell-free culture solution was added to each well of the blank and zero group. 37°C, 5% CO 2 cultured in an incubator. When the cell confluency was 80%, the cells were treated with drugs.

[0065] S23: drug treatment of cells;

[0066] Control group: 100 μl / well of 1640 cell culture medium containing 10% FBS;

[0067] Experimental group: single dicoumarin group - drug concentration 800μM, 400μM, 200μM, 100μM, 50μM, 25μM, 12.5μM, 6.25μM, 3.125μM, 1.5625μM, doubling dilution; final volume is 100μl / well; Each group was set up with 5 duplicate wells, and cultured in a 37°C, 5% CO2 incubator for 24 hours.

[0068] S24: Add 10μl / well of MTT liquid with a concentra...

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Abstract

The invention provides a PDK1 inhibitor, a composition containing the inhibitor and application, belonging to the technical field of medicine. Specifically speaking, bishydroxycoumarin enables cell p-PDH expression to be reduced through inhibiting PDK1 activity in ovarian cancer cells, promotes cellular tricarboxylic acid cycle, increases reactive oxygen species in cells, promotes mitochondrial membrane potential to be reduced, and enables mitochondrial membrane permeability to be lowered and cytochrome C to be increased, thereby causing a cancer cell apoptosis mechanism to realize application in preparation of medicines for treating and / or preventing ovarian cancer. In addition, a pharmaceutical composition which takes the bishydroxycoumarin as an active ingredient has an obvious inhibiting effect on the tumor volume size of the ovarian cancer. The bishydroxycoumarin and bishydroxycoumarin compounds provided by the application not only have an ovarian cancer treating effect through inhibiting PDK1, but also cannot cause peripheral nervous toxicity, provide new thoughts and ways for treating the ovarian cancer, and have broad application prospects and clinical values.

Description

technical field [0001] The present invention belongs to the field of medical technology, in particular, it relates to a PDK1 inhibitor, a composition containing the inhibitor and its application. Background technique [0002] During the Great Depression of the 1920s, farmers discovered that large numbers of cattle and sheep had died from abnormal bleeding. Veterinarians and farmers worked together to find the cause, and finally locked on a plant that was used as fodder, sweet rhinoceros. Carl Link of the Wisconsin Agricultural University and his students spent six years purifying the plant, and finally found the substance that prevents blood from coagulating normally—coumarin. [0003] In 1941, the Wisconsin Alumni Research Foundation supported the synthesis of dicoumarin, which has an antagonistic effect on vitamin K, by oxidizing coumarin. Almost at the same time, Link used coumarin and its derivatives as rodenticides, while forward-thinking New York Hospital / Cornell Med...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/37A61K36/9066A61P35/00A61K35/413A61K31/36A61K31/192A61K31/121
CPCA61K31/37A61K31/121A61K31/192A61K31/36A61K35/413A61K36/236A61K36/258A61K36/34A61K36/48A61K36/575A61K36/63A61K36/704A61K36/752A61K36/88A61K36/8888A61K36/889A61K36/896A61K36/9066A61K2300/00
Inventor 苏静孙连坤张文嘉周晓丽许华丹
Owner JILIN UNIV
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