Format of bispecific antibodies (BsAbs)-VH1-VH2-CH1-Fc1(G1) x VL2-VL1-CL-Fc2(G1)


CODV (cross-over dual variable domains) -Ig, contain four polypeptide chains that form two dual variable domains (four antigen binding sites) with a cross-over orientation (Figure 1), which is attained by inverting the alignment of the cognate domains on one chain. In order to adopt the correct VH/VL pairing, linker combinations were designed and optimized using a molecular modeling strategy. The overall CODV structure replicates a circular self-contained architecture (Figure 1A), with binding sites facing up to opposite sites and able to accommodate a large variety of antigen sizes while maintaining parental affinities. The CODV structure was also further developed in a trispecific format in which a single IgG Fab arm is combined with a double arm generated in the CODV structure using the KiH heterodimerization strategy (Figure 1B). A trispecific CODV molecule was successfully engineered to target three distinct epitopes on human immunodeficiency virus HIV-1 envelope, including the CD4 binding site, MPER and the V1V2 glycan site. This innovative molecule exhibited an unprecedented neutralization breadth and potency against HIV when compared to other previously described broadly neutralizing antibodies (BnAbs).

Fig. 1. A schematic illustration of the CODV-Ig formats. (Adopted from: Amaral, M., Hölper, S., Lange, C., Jung, J,, Sjuts, H,, Weil, S,, Fischer, M,, Radoevic, K,, Rao, E. (2020) Engineered technologies and bioanalysis of multispecific antibody formats. J Appl Bioanal 6(1):26-51.)

Formats of bispecific antibodies (BsAbs)

Many formats have been developed for BsAb generation as listed in the following table.

FormatSchematic structureDescriptionExample BsAbTrademark Company
tandem VHHTandem VHH fragment-based BsAbN/A
tandem scFvPicture loading failed.Tandem ScFv fragment-based BsAbAMG330BiTETMAmgen
Dual-affinity re-targeting antibodyPicture loading failed.Tandem domain-exchanged Fv (can also be used to fuse with Fc domain to create whole Abs)FlotetuzumabDARTTMMacrogenics
DiabodyPicture loading failed.dimer of single-chain Fv (scFv) fragmentvixtimotamabReSTORETMAmphivena Therapeutics
(scFv)2-FabPicture loading failed.a Fab domain and two scFv domains bindA-337ITabTMGeneron/EVIVE Biotech
Rat–mouse hybrid IgGPicture loading failed.Full-size IgG-like half antibodies from two different speciesCatumaxomabTriomabTMTrion Pharma
Hetero heavy chain, Common light chainPicture loading failed.Hetero heavy chain, Common light chainEmicizumabART-IgTMGenentech/ Chugai/Roche
Controlled Fab arm exchangePicture loading failed.Recombin the parental half antibodies JNJ-64007957DuobodyTMGenmab/ Janssen
Hetero H, forced HL IgG1Picture loading failed.KIH technology for heterodimerization of 2 distinct H chains, replacing the native disulfide bond in one of the CH1-CL interfaces with an engineered disulfide bond to enhance the cognate of H and L paringMEDI5752DuetMabTMMedImmune/ AstraZeneca
cH IgG1Picture loading failed.Identical heavy chains; 2 different light chains: one kappa (κ) and one lambda (λ)NI-1701κλ bodyTMNovimmune SA
Hetero H, CrossMabPicture loading failed.KIH technology; domain crossover of immunoglobulin domains in the Fab regionVanucizumabCrossMabTMRoche
scFv-Fab IgGPicture loading failed.Fab-Fc; ScFv-FcVibecotamab;
XmabTM (the engineered Fc to enhance the generation of heterodimeric Fc);
Xencor/Amgen; YZYBio
VH1-VH2-CH1-Fc1(G1) x VL2-VL1-CL-Fc2(G1)Picture loading failed.2 binding motif in one half antibodySAR440234CODV-IgTMSanofi
VL1-CL1-VH2-CH2-Fc x VH1-CH1 x VL2-CL2Picture loading failed.2 binding motif in one half antibodyEMB-01FIT-IgTMEPIMAB BIOTHERAPEUTICS
VH-1-TCR Cα x VL-1-TCR Cβ; VH-2-CH-2-Fc x VL-2-CL-2Picture loading failed.KIH technology; TCR Cα/Cβ is used to substitute the CH1 and CL domain in one armWuXibodyTMWuXi Biologics
C-terminal linker of FcPicture loading failed.Link the other molecules at the C-terminal of FcAPVO442ADAPTIR-FLEXTMAptevo Therapeutics
Fc antigen binding sitePicture loading failed.2 natural binding sites; 2 additional binding sites in the Fc loopFS118mAb2F-star Therapeutics