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

Recombinant CAR gene and carrier, CAR-T cell and application thereof

A gene carrier and gene technology, applied in the field of CAR-T cells and applications, recombinant CAR genes and their vectors, which can solve the problems of T cell activation and insufficient proliferation ability.

Active Publication Date: 2017-03-22
IMMUNE CELL BIOTECH CO LTD +3
View PDF5 Cites 6 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The purpose of the present invention is to provide a recombinant CAR gene and its carrier, CAR-T cell

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
  • Recombinant CAR gene and carrier, CAR-T cell and application thereof
  • Recombinant CAR gene and carrier, CAR-T cell and application thereof
  • Recombinant CAR gene and carrier, CAR-T cell and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Construction of embodiment 1CART-19scFv plasmid

[0046] The method for obtaining the CART-19scFv plasmid is as follows: the complete sequence of the recombinant CAR gene (SEQ ID NO: 7) is synthesized, and then ligated into the lentiviral plasmid vector pLent-EF1a by AsisI / NsiI double enzyme digestion. For the plasmid map of the constructed CART-19scFv plasmid and pLent-EF1a, see Figure 5 and Figure 6 .

Embodiment 2

[0047] Embodiment 2 T lymphocyte transfection

[0048] The constructed CART-19scFv plasmid was subjected to lentiviral packaging and purification, and then the lentiviral vector technology was used (references: Tumaini B, Lee DW, Lin T, Castiello L, et al. Simplified process for the production of anti-CD19-CAR engineered T cells.Cytotherapy.2013; 15(11):1406-1415.) Express the recombinant CAR (SEQ ID NO:7) gene in the patient's T cells, and use flow cytometry to detect the positive rate of green fluorescent protein (GFP), see results Figure 4 ,Depend on Figure 4 It can be seen that the transfection efficiency of lentivirus infection of T lymphocytes is about 30%.

Embodiment 3

[0049] Example 3 In vitro proliferative ability detection of T lymphocytes after transfection

[0050] The transfected T lymphocytes (CAR-T cells) were expanded in vitro for 2 weeks, and the results are shown in figure 1 . And use flow cytometry to measure the ratio of CD3 positive T cells and CD8 positive T cells before and after expansion, the results are shown in figure 2 and image 3 . Depend on figure 2 It can be seen that the proportion of CD3 positive T cells before expansion is 55.8%+16.8%, about 72.7% in total; the proportion of CD3 positive T cells after expansion is 30.4%+56.2%, about 86.9% in total, compared with before expansion, After 2 weeks of in vitro expansion, CD3 positive T cells increased by 14.2%. Depend on image 3 It can be seen that the proportion of CD8-positive T cells before expansion was 13.8%; the proportion of CD8-positive T cells after expansion was 42.9%. Compared with before expansion, after 2 weeks of in vitro expansion, CD8-positive...

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 belongs to the field of tumor cell immunotherapy, and particularly relates to a recombinant CAR gene and carrier, CAR-T cell and application thereof. The recombinant CAR gene includes a nucleotide sequence coded the following parts of an antigen-binding part of a CD19 antibody, a transmembrane part and a CD28 endochylema functional domain, a 41BB endochylema functional domain and a CD3zeta endochylema functional domain connected in a random sequence. The efficiency of using the recombination CAR gene and the carrier containing the recombinant CAR gene to transfect T cell is high, and the positive T cell through transfection has good multiplication capacity. Besides, the recombination CAR gene is high in specificity to B cell tumor antigen, and prompts CAR-T to kill and wound B cell tumor specifically.

Description

technical field [0001] The invention belongs to the field of tumor cell immunotherapy, and specifically relates to a recombinant CAR gene and its carrier, CAR-T cells and applications. Background technique [0002] B lymphocytic neoplasms include a variety of malignancies, such as most non-Hodgkin's lymphomas, acute lymphoblastic leukemia, and chronic lymphocytic leukemia. Current treatments for B lymphocyte malignancies include chemotherapy, radiation therapy, bone marrow transplantation, and peripheral blood stem cell transplantation. Despite a variety of treatments, there are still many patients who cannot be cured. For example, B-lineage acute leukemia is sensitive to chemotherapy but cannot be eradicated, with relapse occurring in about 65% of adults and 20% of children. So far, people can only improve the sensitivity of chemotherapeutic drugs by enhancing their cytotoxicity, but it also increases the mortality rate. Although adoptive immunotherapy using leukocytes f...

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 Applications(China)
IPC IPC(8): C12N15/62C12N15/85C12N5/10A61K35/17A61P35/00
CPCA61K35/17C07K14/7051C07K14/70517C07K14/70521C07K14/70578C07K16/2803C07K2319/03C07K2319/30C12N5/0636C12N2510/00
Inventor 孙秀莲
Owner IMMUNE CELL BIOTECH CO LTD
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