Method for culturing cord blood lymphocyte DC-CIK

A technology of DC-CIK and lymphocytes, which is applied in the direction of cell culture active agents, blood/immune system cells, tissue culture, etc., can solve the problem of CIK cell proliferation multiples and effective cell content, increased expression, and failure to take into account The effect of T regulatory cell content on tumor immune escape and other issues can be achieved to improve the activation efficiency and expansion efficiency, increase the expansion level, and increase the effect of killing activity

Pending Publication Date: 2016-09-28
山东省齐鲁细胞治疗工程技术有限公司
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Problems solved by technology

[0007] Deficiencies of the current DC-CIK technology: (1) The general CIK preparation method is to treat isolated umbilical cord blood mononuclear cells (PBMC) with IFN-γ for 24 hours, and then add CD3mAb, IL-1α and IL-2 Finally, a certain number of CIK cells can be obtained, but the multiplication factor and effective cell content of the finally obtained CIK cells are not ideal; (2) The expression levels of CTLA4 and PD-1 on the surface of CIK cells in the tumor microenvironment Elevated, binds to the corresponding ligand on DC, thereby mediating the immune negative regulatory mechanism and inhibiting the killing function of CIK cells
Existing DC-CIK techniques do not take into account the effect of T regulatory cell content on tumor immune escape

Method used

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  • Method for culturing cord blood lymphocyte DC-CIK
  • Method for culturing cord blood lymphocyte DC-CIK
  • Method for culturing cord blood lymphocyte DC-CIK

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Embodiment

[0055] The reagents used in this embodiment are as follows:

[0056] Umbilical cord blood was purchased from Shandong Qilu Stem Cell Engineering Co., Ltd.

[0057] Ficoll, purchased from GE company;

[0058] IL-2 was purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0059] CD3mAb was purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0060] GM-CSF was purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0061] IL-4 was purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0062] rhTNF-a was purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0063] IFN-γ, purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0064] CTLA-4, purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0065] PD-1, purchased from Beijing Tongli Haiyuan Biotechnology Co., Ltd.;

[0066] GT-T551H3 medium was purchased from TAKARA Company.

[0067] The method used in this embodiment for amplifying DC-CIK:

[006...

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Abstract

The invention belongs to the technical field of immune cells in-vitro culture, and discloses a method for culturing cord blood lymphocyte DC-CIK. According to the invention, cord blood is taken as a processing object, and is subjected to separation of cord blood mononuclear cells, isolated culture of DC cell, induction culture of CIK cell, and co-culture of DC and CIK to prepare the DO-DIK cell, CTLA-4mAb and PD-1mAb are added, and the maturated DO-DIK cell is subjected to centrifugal collection to obtain the product. By using CD3mAb coating culture bottle, continuous stimulation is carried out at an early stage of culture, cytokine usage is reduced, effector cell population activation efficiency and amplification efficiency are increased, and the culture cost is reduced.

Description

(1) Technical field [0001] The invention belongs to the technical field of in vitro culture of immune cells, in particular to a method for culturing umbilical cord blood lymphocyte DC-CIK. (2) Background technology [0002] Adoptive immunotherapy is to infuse sensitized lymphocytes (with specific immunity) or their products to patients with low cellular immunity (such as tumor patients) to improve the anti-tumor immunity of patients, so as to achieve the purpose of treatment and prevention of recurrence. [0003] Dendritic cells (Dendritic cells) are the most powerful professional antigen-presenting cells (Antigen presenting cells, APCs) in the body. Mature DCs can effectively activate naive T cells, and are at the center of initiating, regulating, and maintaining immune responses. It can present antigens to T lymphocytes, thereby exerting the killing effect of T lymphocytes. [0004] Cytokine-induced killer cells (CIK) is the combination of human peripheral blood or umbili...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783C12N5/0784
CPCC12N5/0639C12N5/0646C12N2501/22C12N2501/2302C12N2501/2304C12N2501/24C12N2501/25C12N2501/998C12N2502/1121C12N2502/1164
Inventor 张慧慧王芝辉武建明张剑慧谭毅
Owner 山东省齐鲁细胞治疗工程技术有限公司
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