Ligands that bind il-4 and/or il-13

An IL-4, ligand technology, applied in the direction of anti-animal/human immunoglobulin, antibody, drug combination, etc.

Inactive Publication Date: 2009-11-11
DORMANTIS LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, inhibition of IL-4 or IL-13 alone using conventional drugs may not produce the desi

Method used

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  • Ligands that bind il-4 and/or il-13
  • Ligands that bind il-4 and/or il-13
  • Ligands that bind il-4 and/or il-13

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0303] Preparation of immunoglobulin-based ligands

[0304] Ligands (such as dual-specific ligands, multispecific) according to the present invention can be prepared according to previously established techniques in the field of antibody engineering for the preparation of scFv, "phage" antibodies and other artificially engineered antibody molecules. Techniques for preparing antibodies are described in the following reviews and citations: Winter & Milstein, (1991) Nature 349:293-299; Pluckthun (1992) Immunological Reviews 130:151-188; Wright et al., (1992) ) Crti.Rev.Immunol.12:125-168; Holliger, P. & Winter, G. (1993) Curr.Op.Biotechn.4, 446-449; Carter, et al. (1995) J.Hematother.4, 463-470; Chester, K.A. & Hawkins, R.E. (1995) Trends Biotechn. 13, 294-300; Hoogenboom, H.R. (1997) Nature Biotechnol. 15, 125-126; Fearon, D. (1997) Nature Biotechnol. 15, 618 -619; Plückthun, A. & Pack, P. (1997) Immunotechnology 3, 83-105; Carter, P. & Merchant, A.M. (1997) Curr. Opin. Biotech...

Embodiment 1

[0390] Example 1: General method

[0391] For the primary screen against human IL-13, libraries of VHdAbs and VkdAbs were panned with biotinylated human IL-13 protein (R&D systems, Minneapolis, US). The IL-13 was biotinylated with a 5-fold molar excess of EZ-Link Sulfo-NHS-LC-Biotin reagent (Pierce, Rockford, USA). Round 1 was performed with streptavidin-coated beads (Dynal, Norway) and either 100 nM or 20 nM antigen; round 2 was performed with neutravidin-coated beads and 20 nM or 4 nM antigen (Henderikx et al. al., 2002, Selection of antibodies against biotinylated antigens. Antibody Phage Display: Methods and protocols, Ed. O'Brien and Atkin, Humana Press).

[0392] In the second primary screening, the antigen concentration was kept at 100 nM, and the same VH and Vk dAb phage display libraries were panned in PBS containing 2% Marvel in a final volume of 1 ml. In the first and third rounds, 4 mg of M280 Streptavidin Dynabeads (Dynal, Norway) were used to capture antigen-ph...

Embodiment 2

[0417] Example 2: Ligands that bind IL-4

[0418] IL-4 receptor binding assay (RBA)

[0419] MAXISORP TM Plates (high protein binding ELISA plates, Nunc, Denmark) were coated overnight with 0.5 μg / ml recombinant human IL-4R / Fc (R&D Systems, Minneapolis, USA). The wells were washed three times with PBS containing 0.1% (v / v) Tween 20, then three times with PBS, and blocked with PBS containing 2% (w / v) BSA. The plates were washed again after addition of 10 ng / ml biotinylated-IL-4 (R&D Systems) mixed with anti-IL-4 dAb or IL-4 serial dilutions. Binding to IL-4 was detected with a peroxidase-labeled anti-biotin antibody (Stratech, Soham, UK). Color was then developed with TBM substrate (KPL, Gaithersburg, USA). The reaction was terminated by adding HCl, and the absorbance was read at 450 nm. The activity of the anti-IL-4 dAb reduces the binding of IL-4 to the receptor, thus reducing the uptake values ​​compared to IL-4 only controls.

[0420] IL-4 cell analysis

[0421] The ...

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PUM

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Abstract

Disclosed are ligands that have binding specificity for interleukin-4 (IL-4), for interleukin-13 (IL-13), or for IL-4 and IL-13. Also disclosed are methods of using these ligands. In particular, the use of these ligands for treating allergic asthma is described.

Description

[0001] related application [0002] This application claims the benefit of US Provisional Application No. 60 / 761,708, filed January 24, 2006, the entire teachings of which are incorporated herein by reference. Background of the invention [0003] Interleukin-4 (IL-4) is a pleiotropic cytokine with broad-spectrum biological effects on B cells, T cells and many non-lymphocytes including monocytes, endothelial cells and fibroblasts. For example, IL-4 stimulates the proliferation of several IL-2- and IL-3-dependent cell lines, induces the expression of major histocompatibility complex class II molecules on resting B cells, and increases the secretion of IgG4 and IgE from human B cells. IL-4 is associated with Th2-type immune response, produced by Th2 cells and promotes the differentiation of Th2 cells. IL-4 has been associated with various diseases such as allergy and asthma. [0004] Interleukin-13 (IL-13) is a pleiotropic cytokine that induces immunoglobulin isotype switching ...

Claims

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Application Information

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IPC IPC(8): C07K16/24A61P37/08A61K39/395
CPCC07K16/247C07K2317/31C07K2317/569A61K2039/507C07K16/244A61K2039/505A61P11/06A61P35/00A61P37/00A61P37/02A61P37/06A61P37/08A61P43/00C07K16/24A61K39/395
Inventor I·汤林森A·A·斯图普A·塞普I·德西尔瓦C·J·迪姆奇M·普佩卡R·M·T·德威尔德特P·D·德鲁S·霍尔姆斯
Owner DORMANTIS LTD
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