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DC-based glioma holoantigen vaccine and preparation method thereof

A technology of glioma and whole tumor antigen, which is applied in the fields of bioengineering and biomedicine, can solve the problems of poor antigen stability, weak ability to stimulate T cells to secrete IFN-γ, and weak specificity, and achieve immunogenicity and Strong specificity, regulating the body's immune mechanism, and improving the effect of the body's immune mechanism

Inactive Publication Date: 2016-01-13
SHEN ZHEN ISTEM REGENERATIVE MEDICINE SCI TECH CO LTD
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Problems solved by technology

[0003] At present, there are two-way regulating therapeutic vaccines for gliomas and their preparation methods in the prior art, but the therapeutic vaccines are found to stimulate T cells to secrete IFN-γ by ELISPOT (solid-phase enzyme-linked immunospot technology). Weak ability, the treatment effect is not significant enough
In addition, those skilled in the art have also developed a therapeutic vaccine made by co-culturing DC and glioma antigen, but the glioma antigen used is to proliferate glioma stem cells through serum culture method. It is obtained by lysing after culture, because the antigen only has the antigen information carried by glioma stem cells, it can only kill tumor-like stem cells and a small amount of tumor cells in glioma, and because there is a certain amount of microbial contamination in the serum culture medium , and animal-derived immunogenicity, so the antigen prepared by the glioma stem cells obtained by serum culture method has poor stability and weak specificity, so the glioma therapeutic vaccine obtained by this antigen has poor stability , weak specificity, low tumor killing rate and weak immunogenicity

Method used

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  • DC-based glioma holoantigen vaccine and preparation method thereof

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preparation example Construction

[0023] A kind of DC-based glioma whole tumor antigen vaccine provided by the present invention, its preparation method comprises the following steps:

[0024] 1. Obtain immature DC;

[0025] 2. Add whole tumor antigen of glioma and co-culture with immature DC for 7-13 days;

[0026] 3. Add tumor necrosis factor-α, interleukin or lipopolysaccharide and continue culturing for 10-24 hours until DCs mature and DCs are loaded with glioma tumor antigens.

[0027] Specifically, in step 1, immature DCs can be derived from human blood, but since immature DCs only account for 0.5-1.0% of the total number of monocytes in blood, the number is very small; therefore, in a preferred embodiment of the present invention, adopt To obtain immature DCs by inducing monocytes, the specific steps are:

[0028] 11. Isolate monocytes from blood;

[0029] 12. Add in vitro induction medium and culture for 72 hours to amplify the monocytes obtained in step 11;

[0030] 13. Use the in vitro induction ...

Embodiment 1

[0064] The preparation method of the DC-based glioma whole tumor antigen vaccine comprises the following steps:

[0065] 1) Take 20ml of human peripheral blood, and use Ficon Lymphocyte Separation Medium to separate CD14 + Monocytes, after washing with PBS solution, press 2.5-5x10 7 / 3ml / well was inoculated in a six-well plate, and incubated with RPMI-1640 medium at 37°C, 5% CO 2 placed in an incubator for 90 minutes to collect adherent cells, namely CD14 + monocytes.

[0066] 2) CD14 + Monocytes were inoculated into RPMI-1640 medium containing 800 U / mL granulo-macrophage factor and 1000 U / mL interleukin-4 and incubated at 37°C, CO 2 After induction in an incubator with a concentration of 5% for 3-5 days, half of the medium was changed, and granule-macrophage factor and interleukin-4 were supplemented to maintain the concentration of granule-macrophage factor and interleukin-4 to obtain immature DC;

[0067] 3) Use PBS solution (PH=7.4) to adjust the concentration of imma...

Embodiment 2

[0075] The difference from Example 1 provided by the present invention is that in this example, the concentration of tumor necrosis factor-α is 10 ng / ml, the concentration of interleukin-1 is 5 ng / ml, and the concentration of glioma whole tumor antigen Concentration is 150ug / uL, immature DC concentration is 2×10 6 individual / mL.

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Abstract

The invention relates to a DC-based glioma holoantigen vaccine and a preparation method thereof. The preparation method includes the steps of 1), acquiring immature DCs; 2), adding in glioma holoantigen and then co-culturing the glioma holoantigen with the immature DCs; and 3), adding in tumor necrosis factor-alpha and interleukin or lipopolysaccharide to continuously culture for 10-24 hours until DCs are matured and loaded with the glioma holoantigen. The DC-based glioma holoantigen vaccine carries complete antigenic information and is high in immunogenicity and specificity and capable of effectively regulating body immune mechanisms for gliomas.

Description

technical field [0001] The invention relates to the fields of bioengineering and biomedicine, in particular to a DC-based glioma whole tumor antigen vaccine and a preparation method thereof. Background technique [0002] Glioma is a tumor derived from the neuroepithelium. It is the most common malignant tumor in the brain, accounting for about 40-50% of all intracranial tumors. It has become the mainstay of tumor treatment due to its high incidence, high recurrence rate, and high mortality rate. problem. Traditional treatment methods are mainly surgery and radiotherapy and chemotherapy to improve the survival rate of patients. However, total resection of gliomas located in important functional areas is difficult. Radiation therapy is the routine treatment for almost all types of glioma, but the efficacy evaluation is different. Except for medulloblastoma, which is highly sensitive to radiotherapy, and ependymoma, which is moderately sensitive, other types are not sensitive...

Claims

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Application Information

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IPC IPC(8): A61K39/00A61P35/00
Inventor 曾宪卓鲁菲
Owner SHEN ZHEN ISTEM REGENERATIVE MEDICINE SCI TECH CO LTD
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