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Preparation method for dendritic cell (DC) tumor vaccine

A technology for dendritic cells and tumor vaccines, which is applied in the field of preparation of dendritic cell tumor vaccines, can solve problems such as difficulty in achieving expected effects, and achieve the effects of low cost, promotion of endocytosis of tumor antigens, and simple preparation process.

Active Publication Date: 2019-09-27
GUANGZHOU UNIVERSITY OF CHINESE MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Neglecting the treatment of abnormal tumor cell membrane glycosylation, the above-mentioned various antigen-loaded dendritic cell vaccine immunotherapy is difficult to achieve the expected effect

Method used

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  • Preparation method for dendritic cell (DC) tumor vaccine
  • Preparation method for dendritic cell (DC) tumor vaccine
  • Preparation method for dendritic cell (DC) tumor vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Embodiment 1 (preparation of tumor vaccine)

[0021] The tumor vaccine described in this example consists of the following steps:

[0022] 1. Add the mouse-derived liver cancer cell body into the DMEM (or RPMI-1640) culture solution containing serum, then add the culture solution with a platycodon saponin D concentration of 5uM, and place it in an incubator with a carbon dioxide concentration of 5%. Incubate at 37°C for more than 2 hours. The purpose of this step is to reduce the expression of sialic acid in tumor cells without causing death of tumor cells, so that antigens covered by sialic acid on the surface of tumor cells can be exposed and recognized by dendritic cells.

[0023] 2. Digest the tumor cells cultured in step (1) with trypsin, then add dendritic cells in the ratio of tumor cell number: dendritic cell number = 0.5: 1 for co-cultivation for more than 2 hours, and add lipopolysaccharide at a concentration of 100 ~300ng / ml of LPS was continued to culture,...

Embodiment 2

[0027] Example 2 (An experiment in which platycodon saponin D inhibits sialic acid in murine breast cancer cells and enhances the effect of dendritic vaccines in preventing breast cancer)

[0028] (1) The effect of the traditional Chinese medicine platycodon saponin D on five highly expressed sialyltransferase genes in breast cancer cells

[0029] The experiments were divided into tumor model group and 5μM Platycodon grandiflora group.

[0030] Breast cancer cells were inoculated into 6cm culture dishes, 5mL per dish, and then cultured in an incubator. Add 5 mL of complete culture solution to the blank control group, add 5 mL of 5 μM PD drug-containing culture solution to the experimental group, and place them in an incubator to continue culturing for 48 hours. When the cells of the two groups grew to a density of 80%, the cells were collected and the total RNA of the cells was extracted. cDNA was reverse transcribed using a reverse transcription kit.

[0031] The cDNA obta...

Embodiment 3

[0047] Example 3 (An experiment of platycodon saponin D inhibiting sialic acid in murine colon cancer cells and enhancing the effect of dendritic vaccines on colon cancer prevention)

[0048] (1) The effect of the traditional Chinese medicine platycodon saponin D on the highly expressed sialyltransferase gene in colon cancer cells

[0049] The experiments were divided into tumor model group and 10μM Platycodon grandiflora group.

[0050] Colon cancer cells were inoculated into 6 cm culture dishes, 5 mL per dish, and then cultured in an incubator. Add 5 mL of complete culture solution to the blank control group, add 5 mL of 10 μM PD drug-containing culture solution to the experimental group, and place them in an incubator to continue culturing for 48 hours. When the cells of the two groups grew to a density of 80%, the cells were collected and the total RNA of the cells was extracted. cDNA was reverse transcribed using a reverse transcription kit.

[0051] The cDNA obtained ...

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Abstract

The invention relates to a preparation method for a DC tumor vaccine. The method comprises the following steps: (1) adding a tumor cell body to a serum-containing culture solution, then adding a platycodin D culture solution, and placing in a carbon dioxide incubator to be cultured for more than 2h; (2) digesting tumor cells cultured in the step (1) with trypsin, performing centrifugation, abandoning the culture solution, resuspending by adding DMEM, then adding DCs to be co-cultured for more than 2h, and adding lipopolysaccharide LPS to be cultured continuously, thereby making DCs suspended; (3) sucking out suspended DCs, carrying out centrifugation, abandoning the culture solution, resuspending by adding DMEM, then performing separation and purification to obtain mature DCs; (4) preparing the DC tumor vaccine by taking mature DCs and adding to isotonic saline or a phosphate buffer solution or serum-free DMEM. The tumor vaccine prepared by the method can promote the immune response of tumor antigens, and the antitumor effect is remarkable.

Description

technical field [0001] The present invention relates to a pharmaceutical preparation containing immune cells, which contains dendritic cells from the immune system and can be used for preventing or treating tumors. Background technique [0002] At present, there are many ways for dendritic cell (DC) tumor vaccines to load antigens, but the goal is to present exogenous tumor antigens to dendritic cells, and then through the endogenous MHC class I of dendritic cells. Molecular processing and presentation pathways to "cross-prime" CD8+ T cells. From the 43 reported clinical trials, the types of antigens used include whole tumor cell antigens (autologous tumor cells: freeze-thawed, irradiated, apoptotic and fusion; apoptotic or necrotic tumor cell lines; accounting for about 34.8%) , tumor polypeptide antigen (about 32.5%), tumor protein antigen (about 16.2%), tumor cell total RNA (electroporation; about 2.3%), mRNA encoding specific tumor antigen (electroporation; about 11.6%)...

Claims

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Application Information

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IPC IPC(8): C12N5/0784A61K39/00A61P35/00
CPCC12N5/0639A61K39/0011A61P35/00C12N2506/115C12N2501/2301C12N2501/22C12N2500/84A61K2039/5154Y02A50/30
Inventor 王剑付远飞李亚飞蓝华全张文静
Owner GUANGZHOU UNIVERSITY OF CHINESE MEDICINE
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