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Inhibition application of respiratory chain compound V in colorectal cancer stem cell treatment

A technology of colorectal cancer and compound, applied in the direction of drug combination, medical preparations containing active ingredients, organic active ingredients, etc., can solve the problems of inhibiting electron transfer, increasing electrochemical gradient, affecting the function of proton pump, etc.

Inactive Publication Date: 2016-05-11
YUNNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The inability of protons to flow in leads to an increase in the electrochemical gradient on both sides of the membrane, which affects the function of the proton pump and inhibits electron transfer.

Method used

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  • Inhibition application of respiratory chain compound V in colorectal cancer stem cell treatment
  • Inhibition application of respiratory chain compound V in colorectal cancer stem cell treatment
  • Inhibition application of respiratory chain compound V in colorectal cancer stem cell treatment

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1: Expression of tumor stem cell marker CD44 in colorectal cancer cells and isolation and culture of CD44 positive cells.

[0019] Both colorectal cancer cell lines HT29 and HCT116 in the present invention were purchased from the Cell Bank of Kunming Institute of Zoology, Chinese Academy of Sciences. Colorectal cancer cells HT29 and HCT116 in the present invention were prepared in DMEM (Gibco Company, Code No. 10566- 016) culture medium, in a 37°C incubator (ThermoScientific Company, CodeNo.370), CO 2 Cultured at a concentration of 5%.

[0020] The expression rate of tumor stem cell marker CD44 was detected by flow cytometry. Two colorectal cancer cell lines were digested into single cells by trypsin (Gibico Company, Code No. 25200-056), transferred to a 15ml centrifuge tube filled with phosphate buffer, centrifuged at 300g for 7 minutes to collect 2×10 5 Cells were resuspended in 160 microliters of phosphate buffered saline, and the cells were divided into t...

Embodiment 2

[0023] Example 2: Differences in mitochondrial morphology and function between colorectal cancer stem cells (CD44 positive) and non-stem cells (CD44 negative).

[0024] The cultured colorectal cancer stem cells HT29 / CD44+, HCT116 / CD44+ and non-stem cells HT29 / CD44-, HCT116 / CD44- were fixed with glutaraldehyde, and the mitochondria were observed with a transmission electron microscope (assisted by the microscope observation laboratory of Kunming Medical University) The microstructure, take pictures of 10000 times and 50000 times, see Figure 4 indicated by the arrow. It was found that colorectal cancer stem cells HT29 and HCT116 had larger mitochondrial diameters than non-stem cells, and there was no significant boundary between mitochondrial inner cristae structures.

[0025] In addition, colorectal cancer stem cells HT29 / CD44+, HCT116 / CD44+ and non-stem cells HT29 were treated with mitochondrial inner membrane dyes Mito-TrackerGreen (ThermoScientific Company, Code No. / CD44...

Embodiment 3

[0028] Example 3: The effect of complex V inhibition on the proliferative activity of colorectal cancer stem cells (CD44 positive).

[0029] Colorectal cancer stem cells HT29 / CD44+, HCT116 / CD44+ and non-stem cells HT29 / CD44-, HCT116 / CD44- were seeded in 96-well cell culture plates at a density of 1000 cells / well, and three different concentrations (0.1 μg / ml, 0.5μg / ml, 1.0μg / ml) respiratory chain complex V inhibitor oligomycin A (TOCRIS company, CodeNo.4110) to treat the cells, and use the cell proliferation activity detection kit (Promega company, Code No.G3581) to detect cell activity on days 0, 2, 4, 6, and 8. Compared with cells not treated with uncoupling agent, the cell proliferation activity of colorectal cancer stem cells HT29 / CD44+ and HCT116 / CD44+ after treatment with oligomycin A was significantly inhibited on the second day, see Figure 8 As shown in A and C, the proliferative activity of non-stem cells HT29 / CD44- and HCT116 / CD44- showed a slight increase, see F...

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Abstract

In the invention, the activity of a respiratory chain protein compound V in colorectal cancer stem cells is higher than that of non-stem cells; according to traditional ideas, the colorectal cancer cells get a growth energy source mainly by a glycolysis way; however, the colorectal cancer cells are proved to get the growth energy source mainly by an oxidative phosphorylation way. Moreover, the oxidative phosphorylation respiratory chain protein compound V is proved to play a remarkably important role in the growth and proliferation processes of colorectal cancer stem cells while the activity in inhibiting the compound V can effectively inhibit the proliferation of the compound V. In the invention, a therapeutic molecular target is provided for clinical intervention of the colorectal cancer stem cells.

Description

technical field [0001] The present invention relates to the inhibitory effect of oligomycin A (Oligomycin A, a function inhibitor of respiratory chain complex V) on the proliferation of colorectal cancer stem cells (Colon Cancer Stem Cells, cCSCs), and explores the application of complex V in the treatment of colorectal cancer stem cells. Background technique [0002] Cancer stem cells (Cancer Stem Cells, CSCs) belong to a subgroup of cancer cells, which have similar self-renewal and multi-differentiation potentials to adult stem cells. Common cancer stem cell markers include CD133, CD44, CD2ni4, EpCAM, THY1, ATP-bindingcassetteB5, and CD200. CD44 is a cell adhesion molecule that regulates cell signaling, migration and homing and is widely used to mark colorectal cancer stem cells. Normal adult stem cells can encapsulate chemotherapeutic drugs with vesicles and pump them out of the cell, thus exhibiting natural tolerance to chemotherapeutic drugs. Cancer stem cells are als...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/365A61P35/00A61P35/04
CPCA61K31/365
Inventor 李梅章王文学丁磊
Owner YUNNAN UNIV
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