Application of bacillus subtilis protein AMEP412 in inhibition of tumor cell proliferation

A technology for the proliferation of Bacillus subtilis and tumor cells, which is applied in the direction of anti-tumor drugs, desipeptides, peptides, etc., and can solve problems such as complex drug coupling methods, high difficulty in purification and characterization, and slow diffusion of monoclonal antibodies

Active Publication Date: 2021-12-14
HEILONGJIANG BAYI AGRICULTURAL UNIVERSITY +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Monoclonal antibodies have the disadvantages of slow diffusion in tumor tissue, complex drug conjugation methods, and high difficulty in purification and characterization

Method used

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  • Application of bacillus subtilis protein AMEP412 in inhibition of tumor cell proliferation
  • Application of bacillus subtilis protein AMEP412 in inhibition of tumor cell proliferation
  • Application of bacillus subtilis protein AMEP412 in inhibition of tumor cell proliferation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047]Inhibition of tumor cell activity by Bacillus subtilis AMEP412 protein.

[0048] method:

[0049] (1) Cell recovery: wearing sterile gloves, take out the cell cryopreservation tube from the liquid nitrogen tank. Put it into a 37°C water bath quickly, and shake it from time to time to make it completely melt within 1 minute, and then remove the cells under aseptic conditions. After adding 5mL of 1640 culture solution, inoculate it into a culture bottle, place it in a 37°C incubator for static culture, replace the culture solution once the next day, continue the culture, and observe the growth situation.

[0050] (2) Cell subculture: when the cells are fused to 90%, discard 25cm 2 For the culture medium in the culture flask, wash the cells twice with PBS; add 1.5ml of 0.25% trypsin digestion solution, and incubate in a 37°C incubator for a certain period of time. After the cells shrink back and become round, add complete medium to stop digestion. Gently blow the cells w...

Embodiment 2

[0073] Transcriptomic analysis of AMEP412 protein in breast cancer cell line 4T1.2.

[0074] (1) Cell sample preparation

[0075] Resuscitate 4T1.2 cells for expansion culture. When the cells reached a certain amount, they were subcultured, and the cells were washed twice with PBS; digested with 0.25% trypsin digestion solution, incubated in a 37°C incubator for a certain period of time, and complete medium was added after the cells shrank and rounded to stop digestion. Gently blow the cells with a pipette to make them fall off, transfer the cell suspension to a 15ml centrifuge tube, and centrifuge at 1000rpm for 5min; discard the supernatant, resuspend the cell pellet with fresh medium, adjust the cell concentration, and prepare multiple batch samples, randomly Divided into control group and treatment group. After 24 hours of cell attachment, the administration group was replaced with the medium containing 50ug / mLAMEP412 protein, and the control group was replaced with the ...

Embodiment 3

[0090] Toxicity test of AMEP412 protein to normal cells.

[0091] In order to detect whether the AMEP412 protein is toxic to normal cells, the method in Example 1 was used to determine the IC50 value of normal human liver cells HL-7702 and prostate epithelial cells WPMY-1, using the CCK8 method. The results showed that the IC50 value of HL-7702 was 312 μg / mL, while the IC50 value of WPMY-1 was 357 μg / mL, both of which were significantly higher than those of each tumor cell, indicating that the appropriate concentration of AMEP412 protein can be used without affecting the activity of normal cells. It has an inhibitory effect on the proliferation of tumor cells and has the potential to be developed as a drug for treating tumors.

[0092] The present invention has discovered the inhibitory function of AMEP412 protein on tumor cells; the protein can inhibit 10 types of tumor cells, and has selective specificity for the inhibitory effect on different types of tumor cells, among whi...

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Abstract

The invention relates to an application of a bacillus subtilis protein AMEP412 with an amino acid sequence as shown in SEQ ID NO: 1 in inhibition of tumor cell proliferation. The tumor cells are any one of liver cancer cells, cervical cancer cells, pancreatic cancer cells, colon cancer cells, prostate cancer cells, lung cancer cells, uterine squamous cell carcinoma cells, gastric adenocarcinoma cells, malignant glioblastoma cells and breast cancer cells. According to the invention, the inhibition function of the protein AMEP412 on tumor cells is found; the protein can generate an inhibition effect on 10 kinds of tumor cells, has selective specificity on inhibition effects on different kinds of tumor cells, and has the best effect on breast cancer cells; transcriptomics research finds that the protein AMEP412 can cause the change of a plurality of pathways related to apoptosis and death in tumor cells, and a new material is accumulated for inhibiting the activity of the tumor cells.

Description

technical field [0002] The invention belongs to the field of biology and medicine, and relates to the application of a bacillus subtilis AMEP412 protein in inhibiting tumor cell proliferation. Background technique [0003] Since the beginning of the 21st century, tumors have become a serious threat to human health, and have also become a worldwide problem affecting public health. In China, factors such as population aging, environmental changes, and lifestyle changes have led to an increasing trend in the incidence of malignant tumors in my country year by year. At present, the traditional tumor treatment methods mainly include surgical resection, chemotherapy and radiotherapy. Among them, chemotherapy is a method of suppressing and killing tumor cells through chemically synthesized or isolated drugs, and plays an important role in tumor treatment. However, traditional chemotherapy drugs have problems such as poor water solubility, poor tumor selectivity, strong side effec...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/32A61P35/00
CPCC07K14/32A61P35/00
Inventor 刘权
Owner HEILONGJIANG BAYI AGRICULTURAL UNIVERSITY
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