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Chimeric antigen receptor and application thereof

An antibody and antigen technology, applied in the field of medicine and biology, can solve the problem of unsatisfactory effect of immunotherapy

Pending Publication Date: 2020-07-28
CHENGDU KELUN PRECISION BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Immunotherapies currently under development for BCMA are very unsatisfactory

Method used

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  • Chimeric antigen receptor and application thereof
  • Chimeric antigen receptor and application thereof
  • Chimeric antigen receptor and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0557] Example 1: Production of anti-BCMA murine antibody

[0558] 1) Antigen preparation

[0559] The CDS sequence of human BCMA (NM_001192.2) was retrieved from NCBI data. After the whole gene was synthesized, it was cloned into the vector; CHO-K1 (ATCC) was revived and cultured to keep the cells in the logarithmic growth phase. The cells were infected with lentivirus to overexpress BCMA, and the CHO-K1-BCMA recombinant cell line with high expression of BCMA protein was constructed. A K562 (ATCC)-BCMA cell line with high expression of BCMA was screened by a similar method.

[0560] 2) immunity

[0561] Resuscitate the CHO-K1-BCMA cell line expressing the target antigen, take 5-8 Balb / C mice, inject the CHO-K1-BCMA cell suspension expressing the target antigen into the abdominal cavity of the mouse with a disposable syringe, and repeat the immunization three times ; Carry out booster immunization and hybridoma preparation.

[0562] 3) SP2 / 0-B cell fusion

[0563] The mye...

Embodiment 2

[0566] Example 2: Affinity Determination of Anti-BCMA Murine Antibody

[0567] Surface plasmon resonance (SPR) can dynamically reflect the association / dissociation rate and affinity constant of antibody-antigen interaction in real time. If the sample to be assayed is an antibody, it can be directly carried out with the cultured supernatant of hybridoma cells. Its operation process is in the surface plasmon resonance (SPR) biosensor ( T200 (GE HealthCare)), in short, firstly, the antigen (human BCMA) recognized by the antibody to be tested (the monoclonal antibody secreted by the hybridoma obtained in Example 1) was coupled to the surface of the sensor chip CM5, Then the antibody to be tested is injected, and the antibody-antigen binding process is directly monitored through the chip surface, and the kinetic parameters of the fast semi-quantitative antibody to be tested can be measured. The results showed that the binding rate constant of the mouse monoclonal antibody was ka...

Embodiment 3

[0568] Example 3: Humanization and affinity determination of anti-BCMA murine antibody

[0569] 1) Determination of the sequence of the variable region of the murine antibody

[0570] Take 1×10 6 The hybridoma monoclonal cells obtained in Example 1 were subjected to RNA extraction, and cDNA was prepared, and VH and VL were cloned and sequenced to obtain the heavy chain variable region sequence SEQ ID NO: 1 and the light chain variable region sequence of the murine antibody. The sequence of the variable region is SEQ ID NO: 2; the heavy chain CDRs (HCDR1, HCDR2 and HCDR3) of the antibody are SEQ ID NOs: 3-5 respectively, and the light chain CDRs (LCDR1, LCDR2 and LCDR3) of the antibody are SEQ ID NOs: 6-8. The above CDRs region sequences are defined by the Chothia numbering system, and any other method for determining the CDR region sequence known in the art can also be used to identify the amino acid residues of the CDR region in the variable region.

[0571] 2) Design and p...

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Abstract

The invention relates to the medical biology field, in particular to an anti-BCMA (B-cell maturation antigen) antibody, an antigen binding fragment thereof, and a chimeric antigen receptor (CAR) containing the anti-BCMA antibody. The invention also relates to a nucleic acid molecule capable of encoding the CAR, a modified immune cell containing the CAR, and a method for preparing the modified immune cell. The invention also relates to the purposes of the CARs and the modified immune cell for preventing and / or treating B-cell related illness states (such as B-cell related and plasma cell related malignant tumors and autoimmune diseases) and a method for preventing and / or treating the B-cell related illness states.

Description

technical field [0001] The invention relates to the field of medicine and biology, in particular, the invention relates to an anti-BCMA antibody and an antigen-binding fragment thereof and a chimeric antigen receptor (CAR) comprising the same. The present invention also relates to nucleic acid molecules encoding such CARs, engineered immune cells comprising such CARs, and methods of preparing such engineered immune cells. The present invention also relates to the use of these CARs and engineered immune cells for the prevention and / or treatment of B cell related conditions (such as B cell and plasma cell related malignancies, autoimmune diseases) and the prevention and / or treatment of said B cell Methods for cell-associated conditions. Background technique [0002] B-cell maturation antigen (BCMA) is a member of B cell maturation antigen (CD269) tumor necrosisfactor (TNF) receptor superfamily, which contains 184 amino acids and is a type I transmembrane protein. BCMA is mai...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28C07K19/00C12N15/13C12N15/62C12N5/10C12N5/20G01N33/574G01N33/577A61K39/395A61K35/17A61K47/68A61P35/00A61P37/02
CPCC07K16/2878C07K14/7051C12N5/0636G01N33/57407G01N33/57492G01N33/577A61K35/17A61K47/6849A61P35/00A61P37/02C07K2317/565C07K2317/56C07K2317/622C07K2317/92C07K2317/24C07K2317/76C07K2319/00C07K2319/02C07K2319/03C07K2319/33C07K2319/74C12N2510/00G01N2333/70578Y02A50/30
Inventor 荆光军常建辉李成欧晓红岳庆朱雁林喻海旻肖亮薛彤彤王晶翼
Owner CHENGDU KELUN PRECISION BIOTECHNOLOGY CO LTD
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