Rat-mouse hybrid IgG


Rat-mouse hybrid IgG is a monoclonal antibody with binding sites for two different antigens, typically CD3 and a tumor antigen, making it a type of bispecific monoclonal antibody. In addition, its intact Fc-part can bind to an fc receptor on accessory cells like conventional monospecific antibodies. The net effect is that this type of drug links T cells (via CD3) and monocytes/macrophages, natural killer cells, dendritic cells or other Fc receptor expressing cells to the tumor cells, leading to their destruction.
At first, monoclonal antibodies against the desired antigens are produced using mouse hybridoma cells. Simultaneously, monoclonal antibodies against another antigen are produced using rat hybridoma cells. These two hybridoma cells are hybridized to produce hybrid-hybridomas or quadromas of hybrid (trifunctional) antibody. The trifunctional antibody is extracted and purified using protein A chromatography.
Trifunctional antibodies eliminate tumor cells by means of antibody-dependent cell-mediated cytoxicity and cytotoxic T cell responses with emphasis on CD8 T cells. These trifunctional antibodies also elicit individual anti-tumor immune responses in cancer patients. The principle of trifunctional antibody activity is shown in Fig. 1. Trifunctional antibody activates T cells, and release TNF and IFN-γ cytokines. In addition, FcγR-positive cells are sent to the tumor foci, where they actively produce proinflammatory cytokines IL-6, IL-12, GM-CSF, and DC-CK1. The interaction of T cells and CD14-positive monocytes, activated via Fc/FcR, leads to increased expression of CD83, CD86 and CD40, providing T cells with an additional costimulatory signal from CD40/CD40L or CD80- CD86/CD28, which enhances the therapeutic effect of the antibodies. In such case tumor cells are effectively destroyed not only by direct attack of T cells, but through other mechanisms such as antibody-dependent cellular cytotoxicity, phagocytosis, perforin/granzyme mediated lysis, and induction of apoptosis by activated immune system accessory cells. The resulting necrotic and apoptotic tumor particles are phagocytized, processed, and presented by professional antigen-presenting cells (macrophages, dendritic cells), which is a prerequisite for long-term anticancer immunization.


Fig. 1. Principle of Triomab® antibody action. TAA: tumor-associated antigen; ADCC: antibody-dependent cellular cytotoxicity. (Adopted from: Vasilenko, E., Mokhonov, V., Gorshkova, E., Astrakhantseva, I. (2018) Bispecific Antibodies: Formats and Areas of Application. Molecular Biology. 52. 323-334.)

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