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1. WO2019214290 - METHOD FOR PRODUCING γδ T CELLS MODIFIED BY CHIMERIC ANTIGEN RECEPTOR

Publication Number WO/2019/214290
Publication Date 14.11.2019
International Application No. PCT/CN2019/072285
International Filing Date 18.01.2019
IPC
C12N 5/078 2010.1
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
5Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
07Animal cells or tissues
071Vertebrate cells or tissues, e.g. human cells or tissues
078Cells from blood or from the immune system
C12N 15/867 2006.1
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
15Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
09Recombinant DNA-technology
63Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
79Vectors or expression systems specially adapted for eukaryotic hosts
85for animal cells
86Viral vectors
867Retroviral vectors
CPC
A61K 35/17
AHUMAN NECESSITIES
61MEDICAL OR VETERINARY SCIENCE; HYGIENE
KPREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES
35Medicinal preparations containing materials or reaction products thereof with undetermined constitution
12Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
14Blood; Artificial blood
17Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes
A61K 39/001113
AHUMAN NECESSITIES
61MEDICAL OR VETERINARY SCIENCE; HYGIENE
KPREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES
39Medicinal preparations containing antigens or antibodies
0005Vertebrate antigens
0011Cancer antigens
001102Receptors, cell surface antigens or cell surface determinants
001111Immunoglobulin superfamily
001113CD22, BL-CAM, siglec-2, sialic acid- binding Ig-related lectin 2
C07K 14/7051
CCHEMISTRY; METALLURGY
07ORGANIC CHEMISTRY
KPEPTIDES
14Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
435from animals; from humans
705Receptors; Cell surface antigens; Cell surface determinants
70503Immunoglobulin superfamily
7051T-cell receptor (TcR)-CD3 complex
C07K 14/70517
CCHEMISTRY; METALLURGY
07ORGANIC CHEMISTRY
KPEPTIDES
14Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
435from animals; from humans
705Receptors; Cell surface antigens; Cell surface determinants
70503Immunoglobulin superfamily
70517CD8
C07K 14/7056
CCHEMISTRY; METALLURGY
07ORGANIC CHEMISTRY
KPEPTIDES
14Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
435from animals; from humans
705Receptors; Cell surface antigens; Cell surface determinants
7056Lectin superfamily, e.g. CD23, CD72
C07K 14/70578
CCHEMISTRY; METALLURGY
07ORGANIC CHEMISTRY
KPEPTIDES
14Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
435from animals; from humans
705Receptors; Cell surface antigens; Cell surface determinants
70578NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95
Applicants
  • 河北森朗生物科技有限公司 HEBEI SENLANG BIOTECHNOLOGY CO., LTD [CN]/[CN]
Inventors
  • 李建强 LI, Jianqiang
  • 王庆龙 WANG, Qinglong
  • 王琳 WANG, Lin
Agents
  • 北京纪凯知识产权代理有限公司 JEEKAI&PARTNERS
Priority Data
201810436500.809.05.2018CN
Publication Language Chinese (zh)
Filing Language Chinese (ZH)
Designated States
Title
(EN) METHOD FOR PRODUCING γδ T CELLS MODIFIED BY CHIMERIC ANTIGEN RECEPTOR
(FR) PROCÉDÉ DE PRODUCTION DE CELLULES γδ T MODIFIÉES PAR RÉCEPTEUR D’ANTIGÈNE CHIMÈRE
(ZH) 一种生产嵌合抗原受体修饰的γδT细胞的方法
Abstract
(EN) A method for producing γδ T cells modified by a chimeric antigen receptor, comprising: transfecting K562 cells with shFPPS targeted to FPP synthase by means of a lentiviral vector, such that the expression of FPPS in the K562 cells is lowered and a K562-shFPPS cell line with reduced FPPS expression is constructed; adding the K562-shFPPS cell line into a γδ T cell culture system for co-culturing with the γδ T cells, wherein it is found that the K562-shFPPS cell line can facilitate in vitro differentiation and proliferation of the γδ T cells; and adding a CAR-expressing lentiviral vector to the γδ T cell culture system comprising the cell line for co-culturing, wherein it is found that the K562-shFPPS cell line can further effectively improve the transfection rate of CAR genes. The provided solution effectively overcomes the technical challenge of the large-scale production of CAR-γδ T cells, and has a good application prospect.
(FR) La présente invention concerne un procédé de production de cellules γδ T modifiées par un récepteur d’antigène chimère, comprenant : la transfection des cellules K562 avec shFPPS ciblé vers la FPP synthase à l’aide d’un vecteur lentiviral, de sorte que l’expression FPPS dans les cellules K562 est réduite et une lignée cellulaire K562-shFPPS avec expression FPPS réduite est construite ; l’addition de la lignée cellulaire K562-shFPPS dans un système de culture de cellules γδ T pour la co-culture avec les cellules γδ T, ayant été découvert que la lignée cellulaire K562-shFPPS peut faciliter la différenciation et la prolifération in vitro des cellules γδ T ; et l’addition d’un vecteur lentiviral exprimant CAR au système de culture de cellules γδ T comprenant la lignée cellulaire pour la co-culture, ayant été découvert que la lignée cellulaire K562-shFPPS peut en outre efficacement améliorer le taux de transfection des gènes CAR. La solution fournie surmonte efficacement le défi technique de la production à grande échelle des cellules CAR-γδ T, et présente une bonne perspective d’application.
(ZH) 一种生产嵌合抗原受体修饰的γδT细胞的方法:将靶向于FPP合成酶的shFPPS通过慢病毒载体转染K562细胞,下调了K562细胞中FPPS的表达量,构建了FPPS表达量降低的K562-shFPPS细胞系。将K562-shFPPS细胞系加入γδT细胞培养体系中与γδT细胞进行共培养,发现K562-shFPPS细胞系能促进γδT细胞的体外分化和扩增。将表达CAR的慢病毒载体加入含有该细胞系的γδT细胞培养体系中进行共培养,发现K562-shFPPS细胞系还可以有效提高CAR基因的转染率。所提供的方案有效解决了大规模生产CAR-γδT细胞的技术瓶颈,具有良好的应用前景。
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