Human immunocompetent cell DC-CIK cytomedicine and effective preparation method thereof

A technology of DC-CIK and immune activity, applied in the field of human immunocompetent cell DC-CIK cell preparations and its effective preparation, can solve the problems of long culture time, hindering the promotion and application of DC-CIK cells, and low killing activity of DC-CIK cells problems, to achieve the effect of clearing residual lesions, enhancing specific anti-tumor immune response, and improving anti-cancer activity

Inactive Publication Date: 2015-09-16
奥思达干细胞有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the long culture time of DC-CIK cells and the low killing activity of DC-CIK cells obtained by t...

Method used

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  • Human immunocompetent cell DC-CIK cytomedicine and effective preparation method thereof
  • Human immunocompetent cell DC-CIK cytomedicine and effective preparation method thereof
  • Human immunocompetent cell DC-CIK cytomedicine and effective preparation method thereof

Examples

Experimental program
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Embodiment 1

[0032] Example 1 Preparation of DC-CIK cells

[0033] 1.1 Preparation of DC-CIK cells optimized by the present invention (experimental group)

[0034] The specific preparation steps are as follows: the peripheral blood of the patient is collected, and mononuclear cells are separated by density gradient centrifugation of human lymphocyte separation fluid. Inoculate mononuclear cells into serum-free medium and adjust the cell concentration to 2x10 6 / m1, add IFN-γ (2000U / m1), IL-4 (500U / m1), GM-CSF (800U / m1) at 37℃ 5%C0 2 Culture under conditions, add IL-2 (1000U / m1), IL-15 (20ng / m1), anti-CD3 monoclonal antibody (50ng / m1) to continue culture after 1 day, add TNFα (500U / m1) to promote DC after 3 days Cells mature. After 4 days, IL-2 (1000 U / m1) and IL-15 (20 ng / m1) were added and cultured for 8 days to obtain DC-CIK cells.

[0035] 1.2 Preparation of routine protocol for DC-CIK cells before optimization (control group)

[0036] The specific preparation steps are as follows:...

Embodiment 2

[0037]Example 2 Determination of cell number, cell phenotype and killing activity to cancer cells

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Abstract

The invention discloses a human immunocompetent cell DC-CIK cell preparation and an effective preparation method of the human immunocompetent cell DC-CIK cell preparation. The method includes the following steps: (1) isolating single mononuclear cell from the human peripheral blood; (2) inoculating the single mononuclear cell into the serum-free medium, adding IFN gamma, GM-CSF and IL-4 and culturing for 1 day; (3) adding IL-2, IL-15 and anti-CD3 monoclonal antibody and culturing for 2 days; (4) adding TNF alpha, continuously culturing for 1day and promoting the DC cell maturation; (5) adding IL-2 and IL-15, continuously culturing for 8 days and obtaining the DC-CIK cell; and (6) collecting the DC-CIK cell and preparing the cell preparation. Through the culturing program of the DC-CIK cell, the human immunocompetent cell DC-CIK cell preparation not only can shorten the cell culturing time, improve the multiplication rate and strengthen the killing activity of the DC-CIK cell on the cancer cell, but also can simplify the operating steps and facilitate the clinic popularization and application.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a DC-CIK cell preparation of human immunocompetent cells and an effective preparation method thereof. Background technique [0002] In recent years, with the development of biotechnology, studies have found that non-specific immune cells can kill the stem cells of cancer cells and prevent the recurrence and metastasis of cancer cells (R. Tallerico et al, The Journal of Immunology, 2013, 190(5): 2381 -90). Currently, the most widely used non-specific immune cell therapy is CIK cell and DC-CIK cell therapy. [0003] Dendritic cell (DC) is the professional antigen presenting cell (antigen presenting, APC) with the strongest function found in the human body. It can recognize antigen and activate the acquired immune system; and cytokine-induced killer cells (cytokine-induced killers, CIK cells) are immune cells with strong proliferation ability and non-specific killing of ca...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
Inventor 周萱王云娟吴海敏
Owner 奥思达干细胞有限公司
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