Method for efficiently producing DC-CIK cells through induction of polyinosinic: polycytidylic acid copolymer

A technology of DC-CIK and polymyocytes, applied in animal cells, vertebrate cells, cell culture active agents, etc., can solve the problems of ineffective identification and killing of tumor/cancer cells, low immune system, etc., and achieve growth The possibility of great cure, the effect of improving the quality of life and prolonging the survival time

Pending Publication Date: 2020-10-02
成都源泉生物科技有限公司 +1
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Problems solved by technology

However, for cancer patients, the immune system generally has a low immune system, which cannot effectively identify and kill tumor/cancer cells; There are many mechanisms to escape the recognition and killing of immune cells. The immunotherapy of tumor/cancer is to improve the immunogenicity of tumor/cancer by means of molecular biology technology and cell engineering technology, ...

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  • Method for efficiently producing DC-CIK cells through induction of polyinosinic: polycytidylic acid copolymer
  • Method for efficiently producing DC-CIK cells through induction of polyinosinic: polycytidylic acid copolymer
  • Method for efficiently producing DC-CIK cells through induction of polyinosinic: polycytidylic acid copolymer

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1, the selection of the optimal concentration of polymyocytes

[0039] Material:

[0040] 1. Polymyocytes (Place of origin: Shanghai Yuanye Biotechnology Co., Ltd.; Cas: 24939-03-5)

[0041] 2.Hyclone RPMI 1640

[0042] 3. Peripheral anticoagulation of healthy volunteers

[0043] 4. Conditioned medium: containing IL-2 100000IU / ml, anti-human CD3 humanized chimeric antibody 1000μg / ml, IL-1α

[0044] 10000ng / ml, GMCSF10000ng / ml, gamma interferon 10000ng / ml.

[0045] 5. CCK8 detection kit (Beiren Chemical Technology (Beijing) Co., Ltd., model: CK04)

[0046] 6. Others: plate microplate reader (Shenzhen Redu Life Science Co., Ltd., model: RT-6100), carbon dioxide incubator (Thermo SCIENTIFIC, model: 3111), clean bench (Thermo SCIENTIFIC, model: 1379), etc.

[0047] Prepare:

[0048] 1. Preparation of DC-CIK medium: Take the plasma-containing medium in "method step 2" respectively, and add 500 μl of DC-CIK conditioned medium according to the ratio of DC-CIK ...

Embodiment 2

[0067] Example 2 Effects of Polymyocytes on the Proliferation of DC-CIK

[0068] Material:

[0069] 1. Polymyocytes (Shanghai Yuanye Biotechnology Co., Ltd.; Cas: 24939-03-5)

[0070] 2.Hyclone RPMI 1640

[0071] 3. Peripheral anticoagulation of healthy volunteers

[0072] 4. Conditioned medium: containing IL-2 100000IU / ml, anti-human CD3 humanized chimeric antibody 1000μg / ml, IL-1α10000ng / ml, GMCSF10000ng / ml, gamma interferon 10000ng / ml.

[0073] 5. Plate microplate reader (Shenzhen Redu Life Science Co., Ltd., model: RT-6100), carbon dioxide incubator (Thermo SCIENTIFIC, model: 3111), clean bench (Thermo SCIENTIFIC, model: 1379), etc.

[0074] Prepare:

[0075] 1. Production of DC-CIK medium for the control group: Take 250ml of the HycloneRPMI 1640 medium containing 2% autologous plasma in "Method Step 2" and add 2.5ml of DC-CIK conditioned medium to form Control DC-CIK- Hyclone RPMI 1640 medium.

[0076] 2. Production of DC-CIK medium in the experimental group: Take 2...

Embodiment 3

[0092] Example 3 Identification of highly efficient DC-CIK phenotype induced by polymyocytes

[0093] Material:

[0094] 1. Polymyocytes (Shanghai Yuanye Biotechnology Co., Ltd.; Cas: 24939-03-5)

[0095] 2.Hyclone RPMI 1640

[0096] 3. Peripheral anticoagulation of healthy volunteers

[0097] 4. Conditioned medium: containing IL-2 100000IU / ml, anti-human CD3 humanized chimeric antibody 1000μg / ml, IL-1α10000ng / ml, GMCSF10000ng / ml, gamma interferon 10000ng / ml.

[0098] 5. Flow cytometry antibody CD3: BV510, CD4: BUV496, CD8: BV786

[0099] 6. Others: flow cytometer (BD, model: FACSymphony), carbon dioxide incubator (ThermoSCIENTIFIC, model: 3111), clean bench (Thermo SCIENTIFIC, model: 1379), etc.

[0100] Prepare:

[0101] 1. Production of DC-CIK medium for the control group: Take 250ml of Hyclone RPMI1640 medium containing 2% autologous plasma in "method step 2", add 2.5ml of DC-CIK conditioned medium to form DC-CIK conditioned medium, namely into ControlDC-CIK-Hyclone ...

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Abstract

The invention relates to the technical field of immunization, in particular to a polyinosinic: polycytidylic acid copolymer (Poly I: C). The polyinosinic: polycytidylic acid copolymer is a ligand of atype-3 Toll-like receptor in an animal body, can mediate a series of immune reactions of the body after TLR-3 is activated, and has a good promoting effect on specific immunity and non-specific immunity of the body. Compared with a control group, DC-CIK induced by the polyinosinic: polycytidylic acid copolymer has higher proliferation, high differentiation (especially CD3+CD4-CD8+, CTL) and hightumor killing activity. The autologous efficient DC-CIK induced by the polyinosinic: polycytidylic acid copolymer and other immune cells are applied to treatment of clinical tumor patients. The polyinosinic: polycytidylic acid copolymer aims to recover or improve the immune function of the tumor patients, and the immune system of the body is improved to kill and inhibit proliferation of tumor cells. The tumor cell load is reduced, tiny residual lesions are removed, or the proliferation mode of residual tumor/cancer cells is obviously inhibited, factors such as relapse and metastasis are eliminated, the cure possibility is increased, the survival time is prolonged, and the life quality is improved. Therefore, the purpose of treating tumors/cancers is achieved.

Description

technical field [0001] The present invention relates to the technical field of immunization, in particular to the use of polyinosinic acid and polycytidylic acid copolymer-polyinosinic acid (polyinosinic: polycytidylic acid copolymer, Poly I:C), polyinosinic is a type 3 in animals Ligands of Toll-like receptors, after activating TLR-3, can mediate a series of immune responses in the body, such as inducing the secretion of cytokines such as interferon, interleukin, and tumor necrosis factor, and promoting the immune response of monocytes, macrophages, and lymphocytes. It can promote the specific immunity and non-specific immunity of the body. [0002] Polymyocytes were added to the induction medium to synergistically induce the proliferation and differentiation of DC-CIK (macrophage-cytokine-activated tumor killer cells) cells, so that the tumoricidal activity of DC-CIK was greatly improved. In order to obtain a kind of autologous high-efficiency DC-CIK cells induced by polymy...

Claims

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Application Information

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IPC IPC(8): C12N5/0783C12N5/0784
CPCC12N5/0638C12N5/0636C12N5/0646C12N5/0639C12N2500/40C12N2501/50
Inventor 刘才明刘杰
Owner 成都源泉生物科技有限公司
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