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Protective agent for CAR-T cell cryopreservation and cryopreservation method

A cryopreservation method and protective agent technology, applied in the field of immune cells, can solve problems such as poor safety, inability to meet the needs of use, limited autologous plasma materials, etc., to maintain stability, prolong cryopreservation time, and reduce cell damage Effect

Pending Publication Date: 2021-12-28
长春金鸟生物医药发展有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, dimethyl sulfoxide and autologous plasma are added to the cryoprotectant formulations that are widely used. Although this cryoprotectant can maintain the activity of cells, it is safe because dimethyl sulfoxide has certain cytotoxicity. The performance is not good, and the autologous plasma is limited. When a large number of cells are expanded and frozen, it cannot meet the needs of use.

Method used

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  • Protective agent for CAR-T cell cryopreservation and cryopreservation method

Examples

Experimental program
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Effect test

Embodiment 1

[0023] A protective agent for cryopreservation of CAR-T cells, consisting of RPMI1640 medium and the following components added to the medium: Paulownin, inositol nicotinate, wax gourd seed extract, hydroxytyrosol, β-glucan , insulin, glutathione; the final concentration of each component in the medium is: Paulownin 3ng / mL, inositol nicotinate 5μg / mL, wax gourd seed extract 2μg / mL, hydroxytyrosol 14ng / mL, β - Dextran 12 μg / mL, Insulin 7 μg / mL, Glutathione 8 μg / mL.

[0024] A cryopreservation method for CAR-T cells, mixing CAR-T cells and a protective agent to obtain a cell suspension, the density of CAR-T cells in the protective agent is 7×10 6 pcs / mL, divided into sterile cryopreservation tubes, 1.5mL per tube, the cryopreservation tubes were cooled to -80°C by 3°C / min program, and then transferred to liquid nitrogen for cryopreservation.

Embodiment 2

[0026] A protective agent for cryopreservation of CAR-T cells, consisting of RPMI1640 medium and the following components added to the medium: Paulownin, inositol nicotinate, wax gourd seed extract, hydroxytyrosol, β-glucan , insulin, glutathione; the final concentration of each component in the culture medium is: Paulownin 1ng / mL, inositol nicotinate 2μg / mL, wax gourd seed extract 1μg / mL, hydroxytyrosol 12ng / mL, β - Dextran 10 μg / mL, Insulin 4 μg / mL, Glutathione 5 μg / mL.

[0027] A method for cryopreserving CAR-T cells, mixing CAR-T cells and a protective agent to obtain a cell suspension, and the density of CAR-T cells in the protective agent is 5×10 6 pcs / mL, divided into sterile cryopreservation tubes, 1.5mL per tube, the cryopreservation tubes were cooled to -80°C by 3°C / min program, and then transferred to liquid nitrogen for cryopreservation.

Embodiment 3

[0029] A protective agent for cryopreservation of CAR-T cells, consisting of RPMI1640 medium and the following components added to the medium: Paulownin, inositol nicotinate, wax gourd seed extract, hydroxytyrosol, β-glucan , insulin, glutathione; the final concentration of each component in the medium is: Paulownin 5ng / mL, inositol nicotinate 8μg / mL, wax gourd seed extract 3μg / mL, hydroxytyrosol 16ng / mL, β - Dextran 14 μg / mL, Insulin 9 μg / mL, Glutathione 10 μg / mL.

[0030] A cryopreservation method for CAR-T cells, mixing CAR-T cells and a protective agent to obtain a cell suspension, the density of CAR-T cells in the protective agent is 9×10 6 pcs / mL, divided into sterile cryopreservation tubes, 1.5mL per tube, the cryopreservation tubes were cooled to -80°C by 3°C / min program, and then transferred to liquid nitrogen for cryopreservation.

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Abstract

The invention discloses a protective agent for CAR-T cell cryopreservation. The protective agent comprises a culture medium and the following components added in the culture medium: paulownin, inositol nicotinate, a wax gourd seed extract, hydroxytyrosol, beta-glucan, insulin and glutathione. The protective agent provided by the invention is composed of the culture medium, and the paulownin, the inositol nicotinate, the wax gourd seed extract and the like which are added into the culture medium, wherein the paulownin is beneficial to reducing the size of ice crystals in a cryopreservation process and reducing damage to cells. The inositol nicotinate and the wax gourd seed extract are beneficial to keeping stability of the CAR-T cells in the cryopreservation process and reducing cell apoptosis. The components, hydroxytyrosol, beta-glucan, insulin and glutathione together provide a protection effect in the cryopreservation process of the CAR-T cells, so that the viability of the cryopreserved CAR-T cells is high, and the cell viability is hardly influenced. The invention further provides a cryopreservation method of the CAR-T cells, the protective agent is adopted, the operation is easy and convenient, and the cryopreservation cost of the CAR-T cells is effectively reduced.

Description

technical field [0001] The invention relates to the field of immune cells, in particular to a protective agent for cryopreservation of CAR-T cells and a cryopreservation method. Background technique [0002] Solid tumors are the main cause of death for cancer patients. Traditional therapies such as surgery, radiotherapy, and chemotherapy have made great progress in recent years, but this therapy has a great burden on the body, and after malignant metastasis occurs, no matter what It is difficult to completely cure the disease with traditional therapies. However, with the rapid development of molecular biology techniques, immune cell therapy has become a new means of treating tumors. [0003] CAR-T therapy is chimeric antigen receptor T cell immunotherapy, which is a new type of precise targeted therapy for tumor treatment. In recent years, it has achieved good results in clinical tumor treatment through optimization and improvement. It is a very effective A promising new t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01N1/02
CPCA01N1/0221A01N1/0284
Inventor 李广超石连兵
Owner 长春金鸟生物医药发展有限公司
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