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Method for preparing memory T cells

A memory and cellular technology, applied in the field of preparation of human memory T cells, can solve the problems of anti-tumor effect in difficult patients, lack of breakthrough in adoptive cellular immunotherapy, affecting T cell activation and metabolism, etc. Survival time, enhanced anti-tumor ability, enhanced lasting and efficient effect

Inactive Publication Date: 2019-03-05
THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, adoptive cellular immunotherapy has not yet achieved a breakthrough in the treatment of solid tumors
The reason is that on the one hand, the existence of the tumor microenvironment affects the activation and metabolism of T cells; on the other hand, because most of the T cells expanded and cultured in vitro are in a terminally differentiated state, it is difficult to exert sustained and long-term anti-tumor effects in patients. effect

Method used

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  • Method for preparing memory T cells
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  • Method for preparing memory T cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Embodiment 1: The preparation method of T cells comprises the following steps

[0033] (1) Collect 20 mL of peripheral blood under sterile conditions and adopt density gradient centrifugation to obtain PBMCs from human peripheral blood mononuclear cells. Both acceleration and deceleration are 9m / s 2 , to obtain serum; B. Transfer the serum obtained in step A to a sterile 50mL centrifuge tube, inactivate at 56°C for 25min, then centrifuge the serum at 4000rpm for 20min, and the acceleration and deceleration accelerations are both 9m / s 2 , using PBS buffer solution to dilute the precipitated blood cells in a ratio of 1:3 to the volume of blood cells to obtain a blood cell dilution; C, then slowly add the blood cell dilution obtained in step B to the lymph separation liquid, the volume of the lymph separation liquid The amount is 1 / 3 of the volume of the blood cell dilution solution. Under the condition of 2500rpm, centrifuge for 25min, and the acceleration and decelerati...

Embodiment 2

[0037] Embodiment 2: The preparation method of T cells comprises the following steps

[0038] (1) Collect 20 mL of peripheral blood under sterile conditions and adopt density gradient centrifugation to obtain PBMCs from human peripheral blood mononuclear cells. Both acceleration and deceleration are 9m / s 2 , to obtain serum; B. Transfer the serum obtained in step A to a sterile 50mL centrifuge tube, inactivate at 56°C for 25min, then centrifuge the serum at 4000rpm for 20min, and the acceleration and deceleration accelerations are both 9m / s 2 , using PBS buffer solution to dilute the precipitated blood cells in a ratio of 1:2 to the volume of blood cells to obtain a blood cell dilution; C, then slowly add the blood cell dilution obtained in step B to the lymph separation liquid, the volume of the lymph separation liquid The amount is 1 / 4 of the volume of the blood cell dilution solution. Under the condition of 2500rpm, centrifuge for 25min, and the acceleration and decelerati...

Embodiment 3

[0042] Embodiment 3: The preparation method of T cell comprises the following steps

[0043] (1) Collect 20 mL of peripheral blood under sterile conditions and adopt density gradient centrifugation to obtain PBMCs from human peripheral blood mononuclear cells. Both the acceleration and deceleration accelerations are 9m / s 2 , to obtain serum; B. Transfer the serum obtained in step A to a sterile 50mL centrifuge tube, inactivate at 56°C for 25min, then centrifuge the serum at 4000rpm for 20min, and the acceleration and deceleration accelerations are both 9m / s 2 , using PBS buffer solution to dilute the precipitated blood cells in a ratio of 1:4 to the volume of blood cells to obtain a blood cell dilution; C, then slowly add the blood cell dilution obtained in step B to the lymph separation liquid, the volume of the lymph separation liquid The volume is 1 / 2 of the volume of the blood cell dilution solution. Under the condition of 2500rpm, centrifuge for 25min, and the accelerati...

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Abstract

The present invention provides a method for efficiently preparing memory T cells by combining TWS119 and cytokines IL-7 and IL-21. The method includes collecting human peripheral blood; performing density gradient centrifugation to obtain human peripheral blood mononuclear cells; sorting CD8<+>T cells with a magnetic separation sorter; adding the CD8<+>T cells into a medium containing a stimulantCD3 / CD28 conjugated magnetic beads, cytokines IL-7and IL-21 and TWS119; and performing amplification so that a high number of CD8<+>T cells having Tcm and Tscm phenotypes can be obtained in the seventh day.

Description

technical field [0001] The invention belongs to the cross technical field of life science and medicine, specifically relates to a preparation method of human memory T cells, and belongs to the field of cellular immunology. Background technique [0002] With the understanding of the human immune system and the in-depth study of the mechanism of tumorigenesis, tumor immunotherapy has developed rapidly in the past few decades. Among them, adoptive cell immunotherapy (ACI), chimeric antigen receptor-modified T cells (CAR-T) and immune checkpoint blockers are research hotspots at home and abroad. Immunotherapy has achieved the most revolutionary progress in the field of cancer research. This treatment method can not only improve the immune status of patients, prolong the survival time of patients, but also point out the direction for future research. ACI mainly refers to the expansion of a large number of functional killer cells in vitro and infusion of them to patients to direc...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783A61P35/00A61P31/00A61P37/00
CPCA61K35/17C12N5/0636C12N2501/2307C12N2501/2321C12N2501/51C12N2501/515C12N2501/727
Inventor 张毅张震申春一
Owner THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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