Dual Variable Region Antibody-Like Binding Proteins Having Cross-Over Binding Region Orientation

Abstract

The invention provides antibody-like binding proteins comprising four polypeptide chains that form four antigen binding sites, wherein each pair of polypeptides forming an antibody-like binding protein possesses dual variable domains having a cross-over orientation. The invention also provides methods for making such antigen-like binding proteins.

Description

CROSS REFERENCE TO RELATED APPLICATIONS[0001]This application claims the benefit of priority from U.S. Provisional Application No. 61 / 468,276, filed Mar. 28, 2011, and French Patent Application No. 1160311, filed Nov. 14, 2011, the disclosures of which are incorporated by reference herein in their entirety.FIELD OF THE INVENTION[0002]The invention relates to antibody-like binding proteins comprising four polypeptide chains that form four antigen binding sites, wherein each pair of polypeptides forming the antibody-like binding protein possesses dual variable domains having a cross-over orientation. The invention also relates to methods for making such antigen-like binding proteins.BACKGROUND OF THE INVENTION[0003]Naturally occurring IgG antibodies are bivalent and monospecific. Bispecific antibodies having binding specificities for two different antigens can be produced using recombinant technologies and are projected to have broad clinical applications. It is well known that comple...

AI Technical Summary

Benefits of technology

[0058]Specific embodiments of the invention will become evident from the

Problems solved by technology

A disadvantage associated with the use of such fragments is that they have only one antigen binding site, leading to reduced avidity.

In addition, due to their small size, they are cleared very fast from the serum, and hence display a short half-life.

Bispecific antibodies were originally made by fusing two hybridomas, each capable of producing a different immunoglobulin (Milstein et al., 1983, Nature 305(5934): 537-40), but the complexity of species (up to ten different species) produced in cell culture made purification difficult and expensive (George et al., 1997, THE ANTIBODIES 4: 99-141 (Capra et al., ed., Harwood Academic Publishers)).

The resulting bispecific heterodimer is fully non-human, hence highly immunogenic, which could have deleterious side effects (e.g., “HAMA” or “HARA” reactions), and / or neutralize the therapeutic.

Despite the promising results obtained using heteroconjugates or bispecific antibodies produced from cell fusions as cited above, several factors made them impractical for large scale therapeutic applications.

Such factors include: rapid clearance of heteroconjugates in vivo, the laboratory intensive techniques required for generating either type of molecule, the need for extensive purification of heteroconjugates away from homoconjugates or mono-specific antibodies, and the generally low yields obtained.

The predictability of the final structure of diabody molecules is very poor.

Although sc-BsAb and diabody-based constructs display interesting clinical potential, it was shown that such non-covalently associated molecules are not sufficiently stable under physiological conditions.

Insufficient stability of the VH-VL interface of scFv fragments has often been suggested as a main cause of irreversible scFv inactivation, since transient opening of the interface, which would be allowed by the peptide linker, exposes hydrophobic patches that favor aggregation and therefore instability and poor production yield (Worn et al., 2001, J. Mol. Biol. 305(5): 989-1010).

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