During the past two decades we have seen a phenomenal evolution of bispecific antibodies for therapeutic applications. The ‘zoo’ of bispecific antibodies is populated by many different species, comprising around 100 different formats, including small molecules composed solely of the antigen-binding sites of two antibodies, molecules with an IgG structure, and large complex molecules composed of different antigen-binding moieties often combined with dimerization modules. The application of sophisticated molecular design and genetic engineering has solved many of the technical problems associated with the formation of bispecific antibodies such as stability, solubility and other parameters that confer drug properties. These parameters may be summarized under the term ‘developability’. In addition, different ‘target product profiles’, i.e., desired features of the bispecific antibody to be generated, mandates the need for access to a diverse panel of formats. These may vary in size, arrangement, vale...

https://www.tandfonline.com/doi/pdf/10.1080/19420862.2016.1268307

The making of bispecific antibodies

Alternative molecular formats and therapeutic applications for bispecific antibodies.

The development of bispecific antibodies has attracted substantial interest, and many different formats have been described Those specifically containing an Fc part are mostly tetravalent, such as stabilized IgG-scFv fusions or dual-variable domain (DVD) IgGs However, although they exhibit IgG-like properties and technical developability, these formats differ in size and geometry from classical IgG antibodies Thus, considerable efforts focus on bispecific heterodimeric IgG antibodies that more closely mimic natural IgG molecules The inherent chain association problem encountered when producing bispecific heterodimeric IgG antibodies can be overcome by several methods While technologies like knobs-into-holes (KiH) combined with a common light chain or the CrossMab technology enforce the correct chain association, other approaches, eg, the dual-acting Fab (DAF) IgGs, do not rely on a heterodimeric Fc part This review discusses the state of the art in bispecific heterodimeric IgG antibodies, with an emphasis on recent progress

https://www.tandfonline.com/doi/pdf/10.4161/mabs.21379

Progress in overcoming the chain association issue in bispecific heterodimeric IgG antibodies

Therapeutic antibodies: market considerations, disease targets and bioprocessing.

The state-of-play and future of antibody therapeutics ☆

The Triomab family of trifunctional, bispecific antibodies that maintain an IgG-like shape are novel tumor targeting agents. These chimeras consist of two half antibodies, each with one light and one heavy chain, that originate from parental mouse IgG2a and rat IgG2b isotypes. This combination allows cost-effective biopharmaceutical manufacturing at an industrial scale since this specific mouse/rat isotype combination favors matching of corresponding antibody halves during production by means of quadroma technology. Whereas every Triomab family member is composed of an anti-CD3 rat IgG2b half antibody for T cell recognition, the antigen binding site presented by the mouse IgG2a isotype is exchangeable. Several Triomab antibodies have been generated that bind to tumor-associated antigens, e.g., EpCAM (catumaxomab), HER2/neu (ertumaxomab), CD20 (FBTA05), gangliosides GD2/GD3 (Ektomun), on appropriate tumor target cells associated with carcinomas, lymphomas or melanomas. Catumaxomab (Removab) was launched in Europe for treatment of malignant ascites in April 2009. Here, we report the structural and functional characterization of this product. Mass spectrometry revealed an intact mass of 150511 Dalton (Da) and 23717 Da, 24716 Da, 51957 Da and 52019 Da of the reduced and alkylated rat light chain, mouse light chain, rat heavy chain, mouse heavy chain chains, respectively. The observed masses were in agreement with the expected masses based on the amino acid sequence obtained from cDNA sequencing. The glycosylation profile was similar to other human IgG consisting of biantennary oligosaccharides with different numbers of terminal galactose. CD spectroscopy showed mainly beta-sheets secondary structure that is typical for IgG antibodies. Binding measurement revealed the unique trifunctional features of catumaxomab. Other analytical tools were used to evaluate characteristics of catumaxomab preparations, including the presence of isoforms and aggregates.

https://www.tandfonline.com/doi/pdf/10.4161/mabs.2.3.11791

Patrick Gruber

Papers

Structural and functional characterization of the trifunctional antibody catumaxomab.

Identification and characterization of oxidation and deamidation sites in monoclonal rat/mouse hybrid antibodies.