Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

A method for preparing tumor antigen-specific CD8+ stem cell-like memory T lymphocytes in vitro

A technology of tumor antigens and tumor cells, which is applied in the field of immunology and tumor therapy, can solve problems such as the complexity of the formation process, and achieve the effects of reducing immune tolerance, increasing production, and better results

Active Publication Date: 2020-06-23
FUDAN UNIV SHANGHAI CANCER CENT
View PDF3 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the formation process of autologous tumor antigen immune memory is very complicated, and it must be processed, processed and presented by antigen-presenting cells. Under the stimulation of an appropriate amount of antigen and an appropriate time, activated T cells undergo an expansion period , T cells enter systolic and memory-forming phases after antigen clearance
Therefore, there is currently no mature method to prepare memory T cells with tumor antigen specificity and stem cell characteristics in vitro

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A method for preparing tumor antigen-specific CD8+ stem cell-like memory T lymphocytes in vitro
  • A method for preparing tumor antigen-specific CD8+ stem cell-like memory T lymphocytes in vitro
  • A method for preparing tumor antigen-specific CD8+ stem cell-like memory T lymphocytes in vitro

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Example 1 The tumor cells are induced by ICD inducer to appear the characteristic CRT translocation phenomenon of ICD.

[0051] In the present invention, tumor cells that induce immunogenic cell death using ICD inducers are used as the source of whole cell antigens, and the specific steps are as follows:

[0052] 1.1 Surgical resection of tumor specimens or crude needle biopsy of tumor tissue, under aseptic conditions, remove necrotic tissue and adjacent non-tumor tissue (using colon cancer CT26 tumor as a model);

[0053] 1.2 Wash 3 times with sterile normal saline; cut the tumor tissue into pieces with sterile tissue scissors, add RPMI 1640 medium, and grind thoroughly;

[0054] 1. Collect the single cell suspension after filtering with a 3200-mesh sterile net;

[0055] 1.4 Resuspend the cells in serum-free medium to 1-2x 10 7 / ml, add ICD inducers such as oxaliplatin or doxorubicin to act for 24 hours, if 8% rose bengal is used, it only takes 30 minutes; then extra...

Embodiment 2D

[0057] The cultivation and identification of embodiment 2DC cells

[0058] Dendritic cells (Dendritic Cells, DC) are the most powerful antigen-presenting cells (Antigen Presenting Cells, APCs) found so far. It has been confirmed that DC is the only APC that can significantly stimulate the proliferation of naive T cells, while other APCs (such as mononuclear macrophages, B cells, etc.) can only stimulate activated or memory T cells. DC is the initiator of the body's adaptive T cell immune response and plays an extremely important role in tumor immunity. If tumor antigen-loaded DCs are co-cultured with sorted CD8+ naive T lymphocytes, under the action of specific cytokines, the body can be stimulated to produce tumor antigen-specific Tscm cells.

[0059] The steps of culturing and identification of DC cells are as follows:

[0060] 2.1 Collect 50ml-60ml of anticoagulated peripheral blood intravenously;

[0061] 2.2 Separation and purification of mononuclear cells (PBMC) by de...

Embodiment 3

[0068] Preparation and identification of embodiment 3CD8+Tscm cells

[0069] 3.1 Culture PBMC with serum-free medium, collect suspended cells, and adjust the cell concentration to 1-2x 10 with serum-free medium 6 / ml;

[0070] 3.2 Positive selection of CD8+ initial T lymphocytes by immunomagnetic bead method;

[0071] 3.3 Antigen-loaded DC cells and CD8+ initial T cells were co-cultured at a ratio of 1:10 (number ratio), and recombinant human IL-2 (300 U / ml) and recombinant human IL-7 (300 U / ml) were added to the serum-free culture medium ) and IL-15 (300U / ml);

[0072] 3.4 After 3 days of co-culture, isolate T lymphocytes and continue to add recombinant human IL-2 (300U / ml) and recombinant human IL-7 (300U / ml) and IL-15 (300U / ml) to culture medium, every 3 days Change medium once in half volume.

[0073] 3.5 Collect cells on the 7th day, the number of cells should reach 1×10 6 more than one;

[0074] 3.6 Identification of CD8+Tscm cells: The expression of CD8, CD62L, CC...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention relates to a method for in-vitro preparation of tumor antigen-specific CD8+ T memory stem cells and application of the tumor antigen-specific CD8+ T memory stem cells in antitumor immune response. The method comprises the following steps: subjecting tumor antigen and immature dendritic cells to co-incubation so as to obtain mature dendritic cells loaded with specific tumor antigen; subjecting the mature dendritic cells and CD8+ initial T cells to co-culture so as to promote generation of tumor antigen-specific CD8+ T memory stem cells; and separating out the CD8+ T memory stem cells capable of specifically recognizing specific tumor cells. The tumor antigen-specific CD8+ T memory stem cells prepared by using the method has good antineoplastic effect, high specificity and few adverse reaction.

Description

technical field [0001] The invention relates to the fields of immunology and tumor therapy. Specifically, the present invention relates to a method for preparing tumor antigen-specific CD8+ memory stem cell-like T lymphocytes (T memory stem cells, Tscm) in vitro. The invention also relates to the use of the prepared tumor-specific Tscm to prevent and treat tumors. [0002] technical background [0003] The current tumor T lymphocyte therapy, that is, by preparing tumor antigen-specific T cells, and then isolating and expanding to a certain number (~10 10 1) and then infuse it back to the patient, which is a very promising tumor treatment strategy. The key to this technology is the quantity, activity, phenotype and survival time of antigen-specific T cells after reinfusion, so many laboratories in the world are trying to find better ways to prepare and reinfuse tumor antigen-specific T cells. T cells in order to produce better anti-tumor efficacy. Tumor antigen-specific me...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0783C12N5/0784A61K39/00A61P35/00
Inventor 吴向华胡爱群邢凯琳
Owner FUDAN UNIV SHANGHAI CANCER CENT
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com