Induced amplification method of CIK2 (NK NK-T) cells

A cell and immune cell technology, applied in the field of induction and expansion of CIK2 cells, can solve the problems of low expansion rate, difference in curative effect, and low content, and achieve the effect of high expression and strong efficacy

Active Publication Date: 2021-02-12
上海映天生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] "Cytokine-induced killer cells" (CIK) is an individualized immune cell activation technology. Due to individual differences, many individual lymphocytes are activated and expanded to obtain "Cytokine-induced killer cells" (CIK), its CD16+56+ / CD3+ cell (NK-like T cell) population It does not meet the theoretical or expected requirements, that is, it cannot stably induce a large population of CD16+56+ / CD3+ cells. There are individual differences in efficacy
[0005]And NK cells are a subpopulation of "cytokine-induced killer cells" (CIK), with a low content; if NK cells are simply induced and expanded, their expansion The growth rate is low, and the number of cells is difficult to meet the clinical needs. In order to meet the clinical needs, it is necessary to develop NK (CD56+CD3-) cells and NK-T (CD16 +56+ / CD3+) high cell content and expansion rate, taking into account the induction of new methods to meet clinical needs

Method used

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  • Induced amplification method of CIK2 (NK NK-T) cells
  • Induced amplification method of CIK2 (NK NK-T) cells
  • Induced amplification method of CIK2 (NK NK-T) cells

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Experimental program
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Effect test

Embodiment 1

[0061] (1) Induction and activation of immune cells (CIK2):

[0062] The induction and activation of immune cell CIK2 (NK NK-T) includes the following steps:

[0063] S01: Draw 50ml of anticoagulated peripheral blood Steps: Use 12,500 units of heparin sodium 1ml for anticoagulation, excluding sodium citrate for anticoagulation;

[0064] S02: Coat the T75 culture bottle with 7 ml of "CD16 monoclonal antibody (25-35 μg), Herceptin (trastuzumab 1-2 mg)" inducer for 24 hours, and suck off the coating solution for later use;

[0065] S03: 50ml of peripheral blood is separated to obtain 30ml of plasma (no need to inactivate);

[0066] S04: 3-5×10 lymphocytes isolated from peripheral blood 7 Put it into a coated T75 culture bottle, add 30ml of complete culture solution: AIM-V serum-free medium contains recombinant human γ-interferon (IFN-r) for injection, recombinant human interleukin-2 (IL-2) for injection , recombinant human interleukin-15 (IL-15), recombinant human interleukin-...

Embodiment 2

[0083] (1) Induction and activation of immune cells (CIK2):

[0084] The induction and activation of immune cell CIK2 (NK NK-T) includes the following steps:

[0085] S01: Draw 50ml of peripheral blood for anticoagulation: Use 12,500 units of heparin sodium 1ml for anticoagulation, excluding sodium citrate for anticoagulation;

[0086] S02: Coat the T75 culture bottle with 7 ml of "CD16 monoclonal antibody 25-35 μg, Herceptin (trastuzumab) 1-2 mg" inducer for 24 hours, suck off the coating solution and set aside;

[0087] S03: 50ml of peripheral blood separated and plasma (no inactivation required);

[0088] S04: Lymphocytes (3-5×10 7 Put into the coated T75 culture flask, add

[0089]Complete culture solution 30ml: AIM-V serum-free medium contains recombinant human γ-interferon (IFN-r) for injection, recombinant human interleukin-2 (IL-2) for injection, recombinant human interleukin-15 (IL-15 ), recombinant human interleukin-21 (IL-21), recombinant human stem cell growth ...

Embodiment 3

[0106] (1) Induction and activation of immune cells (CIK2):

[0107] The induction and activation of immune cell CIK2 (NK NK-T) includes the following steps:

[0108] S01: Take 50ml of peripheral blood for anticoagulation: use 12500 units of heparin sodium for anticoagulation, and exclude sodium citrate for anticoagulation;

[0109] S02: Coat the T75 culture bottle with 7 ml of "CD16 monoclonal antibody 25-35 μg, Herceptin (trastuzumab) 1-2 mg" inducer for 24 hours, suck off the coating solution and set aside;

[0110] S03: 50ml of peripheral blood separated and plasma (no inactivation required);

[0111] S04: Lymphocytes (3-5×10 7 Put it into a coated T75 culture flask, add 30ml of complete culture solution: AIM-V serum-free medium contains recombinant human γ-interferon IFN-r for injection, recombinant human interleukin-2 (IL-2) for injection, recombinant Human interleukin-15 (IL-15), recombinant human interleukin-21 (IL-21), recombinant human stem cell growth factor (SCF...

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Abstract

The invention discloses an induction and amplification method of CIK2 (NK NK-T) cells. The method comprises the steps of induction and activation of immune cells CIK2 (NK NK-T) and large-capacity culture and amplification of the immune cells CIK2 (NK NK-T). The method also comprises a step of detecting the immune cell CIK2 (NK NK-T). According to the induced amplification method of the CIK2 (NK NK-T) cells, the percentage of CD16<+> / 56<+> CD3 <-> cells (NK cells) and CD16<+> / 56 <+> CD3<+> cells (NK-like T cells) can be stably increased, and the total number of cells cultured and amplified by the cells can be stably increased. The immune cell provided by the invention retains the characteristic that the NK cell has stronger efficacy of killing tumor cells and virus infected cells than the CIK1 cell; the low amplification rate of the NK cells is compensated, and the new immune cells CIK2 are also named as NK NK-T cells.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for inducing and expanding CIK2 (NK NK-T) cells. Background technique [0002] "Cytokine-induced killer cells" (CIK) are lymphocytes isolated from human peripheral blood that are induced and activated by CD3 monoclonal antibody, γ-interferon, interleukin-1, interleukin-2 and other cytokines, and have the ability to recognize and kill tumor cells, Inhibit the virus and regulate the body's immunity and can cultivate a large number of expanded immune cells. This activated immune cell has a strong proliferation ability, mainly composed of NK-like T cells (CD56 + CD3 + ), T(CD3 + CD8 + ) dominated cell population. It has the advantages of anti-tumor activity of T lymphocytes and non-MHC-limited tumor killing of NK cells. The cells can recognize and kill a variety of tumor cells without harming normal cells in the body. The effect of infusion after surgery or radiotherapy and...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
CPCC12N5/0646C12N2501/24C12N2501/2302C12N2501/2315C12N2501/2321C12N2501/125C12N2500/40C12N2501/515C12N2500/84
Inventor 吴直江戴怀建戴小梅
Owner 上海映天生物科技有限公司
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