Conditional agonists of immune responses

An immune cell, structural technology, applied in the direction of immunoglobulin, anti-growth factor immunoglobulin, anti-animal/human immunoglobulin, etc., can solve problems such as no improvement.

Pending Publication Date: 2019-10-22
F STAR THERAPEUTICS LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, the anti-CD137 anticalin protein genetically fused to the Her2-specific monoclonal antibody (mAb) trastuzumab has been reported to provide tumor target-dependent activation of CD137 on lymphocytes at a site where HER2 was overexpressed in a humanized mouse model, but there was no improvement in tumor growth inhibition relative to trastuzumab alone (Hinner et al, "Costimulatory T-cell engagement by PRS-343, a CD137( 4-1BB) / HER2 bispecific, leads to tumor growth inhibition and TIL expansion in humanized mousemodel", poster presentation at the CRI-CIMT-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016Sept 25-28; NewYork,NY.,September 25-26, 2016, New York, NY and PCT publication WO 2016 / 177802A1)

Method used

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  • Conditional agonists of immune responses
  • Conditional agonists of immune responses
  • Conditional agonists of immune responses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0160] Example 1 - Fcab and mAb 2 preparation of

[0161] 1.1 Selection of Fcab molecules

[0162] 1.1.1 Initial selection of mTNFRX Fcab clones

[0163] A library of clones expressing Fcabs on the surface of yeast cells was incubated with biotinylated recombinant murine TNF receptor (mTNFRX) fused to the Fc domain of murine IgG (mTNFRX-mFc) and passed through the magnetic field using streptavidin-coated beads. Bead sorting (magnetic cell sorting, MACS) sorting. Three rounds of fluorescence activated cell sorter (FACS) selection were then performed using decreasing concentrations of biotinylated mTNFRX-mFc in the presence of a five-fold molar excess of mFc. Cells were stained with streptavidin-allophycocyanin (APC) (BD Biosciences, 349024) or anti-biotin-APC (Miltenyi Biotec, 130-090-856) and sorted using a FACSAria (BD Bioscience). Individual clones from enriched populations were screened for antigen binding, positive binders were subcloned and expressed as soluble Fcab...

Embodiment 2

[0181] Example 2 - by mAb 2 Cross-linked conditional receptor agonism (mTNFRX-X direction)

[0182] Activated T cells express the receptor TNFRX on their cell surface. Clustering of TNFRX is known to be required for TNFRX signaling and further T cell activation. T cell activation assays were used to assess TNFRX receptor clustering and signaling in the presence of the antibody molecules detailed below. T cell activation was measured by IL-2 release.

[0183] 2.1 Isolation and activation of T cells.

[0184] For isolation of T cells, spleens were harvested from 4-8 week old female Balb / C mice (Charles River). Mice were sacrificed humanely and spleens were isolated by dissection. Splenocytes were isolated by pushing the spleen through a 70 μm cell strainer (Corning) using a plastic syringe within 5 mL. The cell strainer was washed 10 times with 1 mL of Duchenne's phosphate-buffered saline (DPBS) (Gibco), and the eluate was collected in a 50-ml tube. The erythrocytes pre...

Embodiment 3

[0205] Example 3-mAb 2 Crosslinking works in both directions

[0206] T cell activation assays were used to assess TNFRX receptor clustering and signaling in the presence of the antibodies listed in Table 6 below. T cell activation was measured by IL-2 release.

[0207] 3.1 Generation of cell lines overexpressing human EGFR

[0208] CT26 cells (ATCC, CRL-2638) overexpressing human EGFR were generated using a lentiviral transduction method using Lenti-X HTX Packaging System (Clontech, Cat. No 631249). Lenti-X expression vector (pLVX) (Clontech, Cat.No 631253) containing human EGFR cDNA was co-transfected into Lenti-X 293T cell line (Clontech, Cat.No 632180) with Lenti-X HTX Packaging Mix to produce Virus. The CT26 cell line was transduced using the lentiviral vector produced with the Lenti-X HTXPackaging System.

[0209] 3.2 T cell activation test

[0210] T cells were isolated and activated as described in Example 2.1.

[0211] After overnight culture, the activated T...

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Abstract

The application relates to antibody molecules, which bind to a first and second antigen, the first antigen being a disease antigen, for example a tumour antigen, and the second antigen being a tumournecrosis factor receptor superfamily (TNFRSF) receptor antigen on the surface of an immune cell. The antibody molecules preferably comprise a CDR-based antigen-binding site and an antigen-binding site, which may be located in two or more structural loops of a constant domain of the antibody molecule. The antibody molecules find application, for example, in cancer therapy.

Description

technical field [0001] The present invention relates to antibody molecules that bind a first antigen, which is a disease antigen, such as a tumor antigen, and a second antigen, which is a tumor necrosis factor receptor superfamily (TNFRSF) receptor on the surface of an immune cell. body antigen. The antibody molecule preferably includes a CDR-based antigen binding site, and the antigen binding site may be located in two or more structural loops of the constant domain of the antibody molecule. Antibody molecules of the invention find use, for example, in the treatment of cancer. Background technique [0002] Cell signaling is a fundamental part of life in all organisms from bacteria to higher eukaryotes. It usually contains cell surface receptors that interact with soluble or surface expressed ligands. This interaction results in changes in the receptor or the ligand, or both, which are then transduced by intracellular proteins that interact with the intracellular domain o...

Claims

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Application Information

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IPC IPC(8): C07K16/22C07K16/28C07K16/30
CPCC07K16/22C07K16/2863C07K16/2866C07K16/2878C07K16/2887C07K16/2896C07K16/30C07K16/3007C07K16/3069A61K2039/505C07K2317/31C07K2317/52C07K2317/64C07K2317/75A61P35/00A61K45/06A61K39/3955A61K39/39558C07K16/468C07K2317/40C07K2317/524C07K2317/526C07K2317/565C07K2317/92
Inventor 密里班·图纳约翰·泽伦森·郝瑞木杰奎琳·弗朗索瓦丝·杜迪孙海军米盖尔·加斯帕
Owner F STAR THERAPEUTICS LTD
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