Immune effector cell cryoprotectant and application thereof

A cryopreservation, cell technology, applied in the biological field, can solve the problems of lack of quality, low cell viability, slow proliferation, etc.

Active Publication Date: 2020-04-07
SHANGHAI CELL THERAPY GRP CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition to the need for centrifugal washing in the resuscitation operation of CAR-T cells cryopreserved with the existing serum-free cryopreservation medium, the resuscitated cultured cells often have low viability and slow proliferation, which becomes the limitation of CAR-T

Method used

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  • Immune effector cell cryoprotectant and application thereof
  • Immune effector cell cryoprotectant and application thereof
  • Immune effector cell cryoprotectant and application thereof

Examples

Experimental program
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preparation example Construction

[0091] The preparation method of the cell cryopreservation liquid of the present invention includes the steps: mixing the components of the cell cryopreservation liquid to obtain the cell cryopreservation liquid.

[0092] Cell mixture

[0093] The cell mixture of the present invention includes the following components:

[0094] Cells, and

[0095] The cell cryopreservation solution of the present invention;

[0096] In the cell mixture, the cells are not particularly limited, as long as they can meet the requirements of cell freezing and resuscitation. Preferably, the cells are immune effector cells. More preferably, the cells include (but are not limited to) one or more selected from the group consisting of CAR-T cells, CIK cells, NK cells and cytotoxic T lymphocytes (CTL). Preferably

[0097] In the cell mixture, the cell cryopreservation solution is further defined as described above.

[0098] Freezing method

[0099] The method for freezing cells of the present invention includes th...

Embodiment 1

[0134] Example 1 Preparation of CAR-T cells

[0135] Take blood samples from donors No. 1 and No. 2 (both donors No. 1 and No. 2 are healthy adults), and isolate PBMC cells by FICOL. After adherence treatment, they are electrotransformed and cultured to the desired cells the amount. The specific steps were performed according to the preparation method disclosed in Antitumoractivity of EGFR-specific CART cells against non-small-cell lung cancer cells in vitro and in mice, Li et al. Cell Death and Disease (2018) 9:177. CAR-T cells prepared from blood samples from patient #1 and donor subject #2 were named CAR-T1 and CAR-T2, respectively.

Embodiment 2

[0136] Example 2 Preparation of Immune Effector Cell Cryopreservation Solution and Cell Cryopreservation

[0137] Preparation of Immune Effector Cell Cryopreservation Solution

[0138] Mix the ingredients according to the ratio of the cell cryopreservation solution as shown in Table 1, and resuspend the CAR-T1 and CAR-T2 cells prepared in Example 1. The cell density after resuspension is 5x10 7 / ml. The reference cryopreservation fluid standard is the serum-containing cryopreservation fluid routinely used in the field. The reference cryopreservation fluid CNKym is the cryopreservation fluid used by the known Novartis product Kymriah, the cryopreservation fluid CBBZ1, CB BZ2, CB BZ3 and CB BZ4 These are the cryopreservation solutions used in the present invention. The cryopreservation fluid standard and cryopreservation fluid CNKym resuspend CAR-T1 and CAR-T2 cells, CB BZ1 and CB BZ3 respectively resuspend CAR-T1 cells, and CB BZ2 and CB BZ4 cryopreservation fluids resuspend CAR-T2...

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Abstract

The invention relates to an immune effector cell cryoprotectant and application thereof. Specifically, the invention provides a cell cryoprotectant which comprises human albumin, dimethyl sulfoxide, dextran, glucose, sodium chloride, sodium gluconate, sodium acetate, potassium chloride, magnesium chloride and water. The invention also provides application of the cell cryoprotectant in preparationof immune effector cell suspension. The cell cryoprotectant has the advantages of improving the viability and the proliferation level of immune effector cells, and has excellent application prospects.

Description

Technical field [0001] The invention relates to the field of biotechnology, in particular to a cryopreservation liquid for immune effector cells and its use. Background technique [0002] In recent years, immunotherapy as a new cancer treatment has attracted wide attention from researchers. Among the newly developed cancer immunotherapy technologies, the chimeric antigen receptor modified T cell (CAR-T) technology is very popular. Since the first generation of CAR-T was developed in 1989, CAR-T-based cancer immunotherapy has made significant progress. Dozens of CAR-T clinical trials are being conducted globally. [0003] CAR-T therapy is the use of leukocyte separation to collect patient's T cells, then in vitro T cell activation, transduction of viral vectors containing CAR, expansion of CAR-T cells, and return to the patient's body. CAR is composed of antibody-derived fragments that can recognize, bind and promote T cell expansion costimulatory molecules coupled to tumor antig...

Claims

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Application Information

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IPC IPC(8): A01N1/02
CPCA01N1/0221
Inventor 刘辉孙艳许慧敏梁超沈丹鹏李君程静波鲁高雄张琪钱其军
Owner SHANGHAI CELL THERAPY GRP CO LTD
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