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Application of mir‑125b in antitumor

A tumor cell, purpose technology, applied in the direction of anti-tumor drugs, the introduction of foreign genetic material using a carrier, and the cells modified by the introduction of foreign genetic material, etc., can solve the problem of tumor cell drug resistance to be improved and so on

Active Publication Date: 2017-11-07
FIELD OPERATION BLOOD TRANSFUSION INST OF PLA SCI ACAD OF MILITARY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, the current means of inhibiting tumor metastasis, reducing tumor cell drug resistance, and inducing tumor cell apoptosis, that is, anti-tumor methods, still need to be improved.

Method used

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  • Application of mir‑125b in antitumor
  • Application of mir‑125b in antitumor
  • Application of mir‑125b in antitumor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Expression of miR-125b in human liver cancer

[0045] The experiment was performed according to the following steps to detect the expression of miR-125b in human liver cancer:

[0046] 1. Preparation of liver cancer cell / tissue cDNA template

[0047] 1. Extraction of total cellular RNA

[0048] Before RNA extraction, the tip and Eppendorf tube were treated with 1‰ DEPC water, autoclaved, and dried for later use.

[0049] (1) When the cells cover 80% of the bottom of the bottle, the culture is terminated, the cell culture medium is discarded, and PBS is added to wash once;

[0050] (2) Add 1ml TRIzol to each bottle of cells, and repeatedly pipette to break and dissolve the cells;

[0051] (3) Transfer the liquid to a 1.5ml EP tube and let stand at room temperature for 5-10 minutes;

[0052] (4) Add 0.2ml chloroform / ml TRIzol, vigorously shake and mix for 15s, and let stand at room temperature for 10min;

[0053] (5) 12000rpm, 4°C, centrifuge for 15min;

...

Embodiment 2

[0080] Example 2 The effect of miR-125b on the expression of EMT-related proteins in human liver cancer cells

[0081] The experiment was carried out according to the following steps to study the effect of miR-125b on the expression of EMT-related proteins in human liver cancer cells:

[0082] 1. Construction of pHRS-1cla-miR-125b-CMV-EGFP vector

[0083] 1.1 Use pcDNA3.1(-)-myc-his as a template to amplify the CMV sequence by PCR. The composition and reaction conditions of the PCR amplification system (20 μl) are as follows:

[0084] 10×PCR buffer

2μl

dNTP mix (2.5mM)

2μl

Primers (upstream and downstream)

1μl

rTaq enzyme

0.2μl

template

1μl

wxya 2 o

13.8μl

[0085] Reaction conditions: 95°C for 5min, (95°C for 30s, 60°C for 30s, 72°C for 30s) x 35 cycles, 72°C for 7min, 4°C for maintenance.

[0086] Analysis of PCR products: PCR products were separated by electrophoresis on 1% agarose gel, and analyze...

Embodiment 3

[0167] Example 3 Effect of mir-125b on drug resistance of human liver cancer cells

[0168] Taking doxorubicin as an example, the experiment was carried out according to the following steps to study the effect of mir-125b on the drug resistance of human liver cancer cells:

[0169] 1. The miR-125b inhibitor synthesized by transient transfection reduces the expression of miR-125b in liver cancer cells and its identification

[0170] The sequence of the synthetic miR-125b inhibitor is as follows: 5'-UCACAAGUUAGGGUCUCAGGGA-3' (SEQ ID NO: 10).

[0171] The control sequence is: 5'-CAGUACUUUUGUGUAGUACAA-3' (SEQ ID NO: 11).

[0172] Transient transfection and identification methods are the same as those shown in Example 1.

[0173] 2. Drug resistance test

[0174] First, inoculate wild-type liver cancer cells in a 96-well plate, according to 5×10 3 Inoculate cells / well, add doxorubicin at different concentrations from 0-2 μg / ml to treat the cells 24 hours later, and select the ap...

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Abstract

The present invention relates to the application of miR-125b in anti-tumor. More specifically, the present invention relates to a construct, a method for inhibiting tumor cell metastasis, reducing tumor cell drug resistance or inducing tumor cell apoptosis, the construct or miR-125b in the preparation of medicines, and for the treatment of cancer medicine. Wherein, the construct includes a nucleic acid molecule encoding miR-125b, and the miR-125b has the nucleotide sequence shown in SEQ ID NO:1. Using this construct, nucleic acid molecules encoding miR‑125b can be introduced into tumor cells, and then by overexpressing miR‑125b in tumor cells, it can effectively inhibit tumor cell metastasis, induce tumor cell apoptosis, or reduce tumor Cell resistance to drugs.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the application of miR-125b in anti-tumor. Specifically, it relates to the application of miR-125b in inhibiting tumor cell metastasis, reducing tumor cell drug resistance and inducing tumor cell apoptosis. More specifically, the present invention relates to a construct, a method for inhibiting tumor cell metastasis, reducing tumor cell drug resistance or inducing tumor cell apoptosis, the construct or miR-125b in the preparation of medicines, and for the treatment of cancer Drug. Background technique [0002] Primary liver cancer, more than 90% of which is hepatocellular carcinoma (HCC), is one of the most common malignant tumors in the world, ranking third among the most common causes of cancer deaths worldwide and second in China . At present, the most effective treatment for liver cancer is resection of liver cancer tissue, but the recurrence rate after surgery is very high. T...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/63C12N5/10A61K48/00A61P35/00A61P35/04
Inventor 裴雪涛岳文周军年曾泉李思霆谢小燕姚海雷
Owner FIELD OPERATION BLOOD TRANSFUSION INST OF PLA SCI ACAD OF MILITARY
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