Composite tumor antibody vaccine using bacterial nano-magnetosome as carrier, and preparation method thereof

A nano-magnetic and bacterial technology, applied in the direction of antibodies, anti-tumor drugs, chemical instruments and methods, etc., to achieve a strong anti-tumor effect

Active Publication Date: 2018-12-18
THE FIRST AFFILIATED HOSPITAL OF CHONGQING MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although there are research reports at home and abroad that it can be connected to membrane antibodies, only the membrane antibodies are used to detect the corresponding bacteria
At present, it has not been reported that after the bacterial magnetosome is linked to the antibody, the compound antibody vaccine can be used to treat tumors, and the targeting and precision of antibody therapy can be improved.

Method used

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  • Composite tumor antibody vaccine using bacterial nano-magnetosome as carrier, and preparation method thereof
  • Composite tumor antibody vaccine using bacterial nano-magnetosome as carrier, and preparation method thereof
  • Composite tumor antibody vaccine using bacterial nano-magnetosome as carrier, and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Preparation of compound tumor antibody vaccine (BMP-anti-4-1BB):

[0037] (1) For pure bacterial nano-magnetosomes (preparation methods: Sun JB, Zhao F, Tang T, Jiang W, TianJS, Li Y Li JL. High-yield growth and magnetosome formation by Magnetospirillum gryphiswaldense MSR-1 in an oxygen-controlled fermentor supplied solely with air.2008; 79(3):389-397) is sterilized by γ-ray irradiation, and the irradiation dose is 15kGy; according to the method of Nakamura, N. et al. Absorbance value (Nakamura, N., Mutsunaga, T. Highly sensitive detection of allergen using bacterial magnetic particles: Biosensors. Anal Chem Acta, 1993, 281, 585-589) and quantification according to 1OD660 = 172 μg of bacterial nano magnetosomes.

[0038] (2) Anti-4-1BB monoclonal antibody was prepared according to Zhou H et al. (Zhou H, Zhang GB, Dong QM, Yu GH, Xu Y, Zhang XG. Preparation of anti-human 4-1BB monoclonal antibody and characterization of its Biological activities, Xi Bao Yu Fen Zi Mian ...

Embodiment 2

[0041] Selection of Optimal Dose of Compound Tumor Antibody Vaccine Stimulating T Cells

[0042] (1) Preparation of compound tumor antibody vaccine: (see the corresponding part of Example 1 for specific operation method)

[0043] (2) Fresh spleen cells of mice were obtained from C57BL / 6 mice. After removing red blood cells with Tris solution (Beijing Zhongshan Jinqiao Biological Co., Ltd.), the splenocyte solution was passed through a nylon mesh to make a single splenocyte solution. 25 μg, 50 μg, 100 μg and 200 μg of compound tumor antibody vaccine were used as stimulators, and the concentration was 2×10 5Splenocytes / ml were used as reactive T cells. After the reactive T cells and stimulators were added to each well of the 96-well incubation plate, a 600mT magnet was placed under each well for 10 minutes. The 96-well plate to which the mixture was added was then placed in a chamber containing 5% CO 2 37°C incubator. After 72 hours, detect with CCK8 reagent, read the OD va...

Embodiment 3

[0045] Effect of compound tumor antibody vaccine on stimulating CD8+ T cells

[0046] (1) Isolation and purification of CD8+ T cells from mice

[0047] CD8+ T cells were isolated from the spleen of C57BL / 6 mice using CD8+ T cell MACS isolation and purification kit (Miltenyi, Biotec). The biotin-labeled monoclonal antibody mixture was combined with non-CD8+ T cells, and then the non-CD8+ T cells were removed by indirect labeling magnetic beads method, and the purity of the remaining CD8+ T cells was detected by CD8-PE antibody in flow cytometry.

[0048] (2) Detection of cytokines

[0049] Add the CD8+T cells isolated in the above steps into a 96-well plate, add 1×105CD8+T cells to each well, and then add 100 μg of compound tumor antibody vaccine, and then use a 600mT magnet to act on each well for 10 minutes, and incubate in carbon dioxide. After 48 hours of co-cultivation in the box, the supernatant was collected, and the concentrations of IFN-γ and TNF-α were determined wi...

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Abstract

The invention relates to a composite tumor antibody vaccine using a bacterial nano-magnetosome as a carrier, and a preparation method thereof. The composite tumor antibody vaccine is established withthe bacterial nano-magnetosome as the carrier, a novel simple and efficient antibody treatment mode is established, and a new immunization method is provided for biotherapy.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a composite tumor antibody vaccine with bacterial nano-magnetosomes as a carrier and its preparation [0002] preparation method. Background technique [0003] Cell receptor 4-1BB (CD137), as an important regulatory element for T cell activation and functional maintenance, can directly activate cytotoxic CD8+ T cells. Studies have shown that in mouse models, anti-4-1BB antibodies alone or in combination with other antibodies can eliminate primary tumors. However, when using antibodies for treatment, intraperitoneal injection is often used. Due to poor targeting, the amount of antibodies used is relatively large, and the therapeutic effect is limited. Therefore, it is extremely important to establish a safe, simple and efficient antibody therapy. Bacterial nanomagnetosomes (BMPs) are new biological nanomaterials, the main component of which is Fe 3 o 4 , with a diameter between 25n...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/395A61K47/52A61P35/00
CPCA61K47/52A61P35/00C07K16/2878A61K2039/505
Inventor 唐翌姝张叔人王冬梅周春霞马文波
Owner THE FIRST AFFILIATED HOSPITAL OF CHONGQING MEDICAL UNIVERSITY
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