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High affinity antibodies to pd-1 and lag-3 and bispecific binding proteins made therefrom

A LAG-3, PD-1 technology, applied in the direction of anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, anti-animal/human immunoglobulin, antibody, etc., can solve the limitation of CD8+ T cells Anti-tumor response and other issues to achieve the effect of reducing tumor-infiltrating Treg cell population and overcoming anti-tumor immunosuppression

Pending Publication Date: 2020-12-11
EPIMAB BIOTHERAPEUTICS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Recent studies have also shown that high LAG-3 expression on depleted lymphocytic choriomeningitis virus (LCMV)-specific CD8+ T cells contributes to their anergic state and limits the antitumor response of CD8+ T cells

Method used

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  • High affinity antibodies to pd-1 and lag-3 and bispecific binding proteins made therefrom
  • High affinity antibodies to pd-1 and lag-3 and bispecific binding proteins made therefrom
  • High affinity antibodies to pd-1 and lag-3 and bispecific binding proteins made therefrom

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0164] Example 1: Production of anti-human PD-1 monoclonal antibody

[0165] Anti-human PD-1 monoclonal antibodies were generated as follows:

Embodiment 11

[0166] Example 1.1: Immunization with human PD-1 antigen

[0167] On day 1, 50 μg of recombinant purified human PD-1 extracellular domain (ECD) polypeptide mixed with complete Freund's adjuvant was injected intraperitoneally into five 6-8 week old Balb / C and five SJL mice middle. On days 16 and 26, 25 μg of recombinant purified human PD-1 ECD immunogen mixed with incomplete Freund's adjuvant was injected intraperitoneally into the same mice. 3-4 days before fusion, a final booster immunization was performed with 25 μg of immunogen.

Embodiment 12

[0168] Example 1.2: Generation of hybridomas

[0169] According to the determination method described in Kohler and Milstein, Nature, 256:495-497 (1975), the splenocytes obtained from the immunized mice described in Example 1.1 were mixed with SP2 / 0-Ag-14 cells at a ratio of 5:1 The proportion of fusion, resulting in hybridoma. Fusion products were mixed at 1×10 per well 5 Splenocytes were plated at a density of 96-well plates in selective medium containing hypoxanthine-aminopterin-thymidine (HAT). Seven to ten days after fusion, macroscopic hybridoma colonies were observed.

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Abstract

High-affinity antibodies recognizing Programmed Death Ligand-1 (PD-1) and Lymphocyte Activation Gene 3 protein (LAG-3) are disclosed. Binding sites from humanized anti-PD-1 and anti-LAG-3 antibodies are incorporated into a Fabs-in-Tandem Immunoglobulin format without significant loss of binding affinity, and the resultant bispecific, multivalent binding proteins are able to bind to both PD-1 and LAG-3 simultaneously. Such bispecific FIT-Ig binding proteins are useful for treatment of cancer.

Description

technical field [0001] The present invention relates to novel antibodies recognizing programmed cell death protein 1 (PD-1), novel antibodies recognizing lymphocyte activation gene 3 protein (LAG-3), and bispecific PD-1 / LAG-3 prepared from those antibodies Binding proteins, such as FIT-Ig binding proteins. Antibodies and bispecific binding proteins can be used to treat immune diseases and blood cancers. Background technique [0002] Programmed cell death protein 1 (PD-1) [0003] Programmed cell death protein 1 (PD-1, CD279) is a member of the CD28 receptor family, which includes CD28, CTLA-4, ICOS, PD-1 and BTLA. Expression of PD-1 is frequently found in immune cells such as T cells, B cells, monocytes and natural killer (NK) cells. PD-1 and similar family members are type I transmembrane glycoproteins comprising an immunoglobulin-like domain similar to the Ig variable domain responsible for ligand binding, and a cytoplasmic tail responsible for binding of signaling mol...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28C12N5/10A61K39/395A61P35/00
CPCA61P35/00C07K16/2818C07K2317/31C07K16/2803C07K2317/92C07K2317/24C07K2317/76C07K2317/55C07K2317/33C07K2317/90A61K2039/505C07K2317/64C07K2317/565C07K2317/526C07K2319/30C07K2317/515C07K2317/522
Inventor 巫玄宫世勇吴辰冰
Owner EPIMAB BIOTHERAPEUTICS INC
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