Single domain antibodies against tnfr1 and methods of use therefor

A tumor necrosis factor and receptor technology, applied in the direction of antibody, anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, anti-animal/human immunoglobulin, etc., can solve problems such as decreased stability

Inactive Publication Date: 2007-12-05
DORMANTIS LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Furthermore, in this case, the heavy chain variable region will not associate with the complementary light chain va

Method used

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  • Single domain antibodies against tnfr1 and methods of use therefor
  • Single domain antibodies against tnfr1 and methods of use therefor
  • Single domain antibodies against tnfr1 and methods of use therefor

Examples

Experimental program
Comparison scheme
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preparation example Construction

[0347] Preparation of immunoglobulin-based multispecific ligands

[0348] The dual-specific ligand of the present invention, whether open or closed in the conformation required by the present invention, can be prepared according to previously established techniques used in the field of antibody engineering, and used to prepare scFv and "phage." Antibodies and other engineered antibody molecules. For the preparation techniques of antibodies, especially bispecific antibodies, see, for example, the following review and references cited therein: Winter and Milstein, (1991) Nature 349:293-299; Pluckthun (1992) Immunological Reviews 130:151-188; Wright (1992) Crti. Rev. Immunol., 12: 125-168; Holliger, P. and Winter, G. (1993) Curr. Op. Biotechn., 4, 446-449; Carter et al. (1995) J. Hernatother ., 4, 463-470; Chester, KA and Hawkins, RE (1995) Trends Biotechn., 13, 294-300; Hoogenboom, HR (1997) Nature Biotechnol., 15, 125-126; Fearon, D. (1997) ) Nature Biotechnol., 15, 618-619; Pluckt...

Embodiment 1

[0527] Example 1. Selection of bispecific scFv antibody (K8) against human serum albumin (HSA) and β-galactosidase (β-gal)

[0528] This example illustrates the method of preparing bispecific antibodies against β-gal and HAS, in which a combination with the germline (dummy) V H The Vκ variable region library connected to the region is used to bind β-gal, and the Vκ region connected to the dummy Vκ region is selected. H Variable region library for binding HAS. Then change the selected variable V HHSA and Vκ β-gal regions bind together, and antibodies that bind β-gal and HSA are selected. HAS is a half-life extending protein that exists in human blood.

[0529] Four human phage antibody libraries were used in this experiment.

[0530] Library 1: Germline Vκ / DVT V H 8.46×10 7

[0531] Library 2: Germline Vκ / NNK V H 9.64×10 7

[0532] Library 3: Germline V H / DVT Vκ 1.47×10 8

[0533] Library 4: Germline V H / NNK Vκ 1.45×10 8

[0534] All libraries are based on V H (V3-23 / DP...

Embodiment 2

[0540] Example 2. Characterization of K8 antibody binding properties

[0541] First, the binding properties of the K8 antibody were characterized by monoclonal phage ELISA. Use 100μl of 10μg / ml HSA and β-gal as well as alkaline phosphatase (APS), bovine serum albumin (BSA), peanut agglutinin, lysozyme and cytochrome c (to check cross-reactivity) of PBS in a 96-well plate at 4 ℃ coated overnight. According to Harrison et al. (1996), the phagemid from the K8 clone was rescued with KM13, and the supernatant containing the phage (50 μl) was directly assayed. According to the standard ELISA protocol (Hoogenboom et al., 1991), the bound phage was detected with an anti-M13-HRP conjugate. When displayed on the surface of phage, the bispecific K8 antibody was found to bind HSA and β-gal with an uptake signal greater than 1.0 (Figure 4). Strong binding to BSA was also observed (Figure 4). Because HSA and BSA share 76% homology at the amino acid level, it is not surprising that the K8 antibo...

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Abstract

The invention provides methods for treating inflammatory diseases (e.g., chronic inflammatory diseases) comprising administering an antagonist of Tumor Necrosis Factor Receptor 1. The invention also provides ligands that contain an immunoglobulin single variable domain (domain antibody, dAb) monomer that binds Tumor Necrosis Factor Receptor 1, and methods of using the ligands. Also provided are nucleic acids encoding the ligands, recombinant host cells and methods for preparing the ligands.

Description

[0001] Related application [0002] This application is a partial continuation application of U.S. Patent Application No. 10 / 985,847 filed on November 10, 2004. It is: [0003] 1) Part of the continued application of the International Patent Application No. PCT / GB2004 / 004253 (the designated country is the United States) filed on October 8, 2004; and [0004] 2) Part of the international patent application number PCT / GB2003 / 005646 (the designated country is the United States) filed on December 24, 2003 continues to apply, and the British application number GB 0230202.4 filed on December 27, 2002 is required to be applied in 2003. Priority of the British application number GB 0327706.8 filed on November 28, [0005] Part of the international patent application number PCT / GB2003 / 002804 (the designated country is the United States) filed on June 30, 2003 continues to apply, and the priority of the British application GB 0230202.4 filed on December 27, 2002 is claimed, which is [0006] ...

Claims

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Application Information

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IPC IPC(8): A61K39/395C07K16/28
Inventor N・D・布鲁易斯B・P・伍尔文S・霍尔姆斯I・M・汤林森J・李C・埃尼弗A・巴斯兰K・琼斯R・德维尔德特S・布莱恩
Owner DORMANTIS LTD
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