Anti-hCD20-hIgG4 (S228P)

Engineered monoclonal human IgG4 antibody against human CD20

Anti-hCD20-hIgG4 (S228P) features the variable region of rituximab and a mutated constant region of the human IgG4 isotype reported to reduce Fab-arm exchange ^[1].

Rituximab is a mouse/human chimeric monoclonal antibody that targets the CD20 antigen found on the surface of malignant and normal B lymphocytes.

Binding of rituximab to CD20 results in cell destruction through different mechanisms including direct signaling of apoptosis, complement activation and cell-mediated cytotoxicity.

Rituximab has been approved by the FDA for the treatment of various lymphoid malignancies, including B-cell non-Hodgkin's lymphoma and B-cell chronic lymphocytic leukemia.

Anti-hCD20-hIgG4 (S228P) is an engineered human IgG4 isotype. IgG4 antibodies display low antibody-dependent cell-mediated cytotoxicity (ADCC) and no complement-dependent cytotoxicity (CDC).

IgG4s are dynamic molecules that exchange Fab arms by swapping a heavy chain and attached light chain (half molecule) with a heavy-light chain pair from another molecule, resulting in bispecific antibodies ^{[1, 2]}.

IgG4 molecules thereby lose their ability to cross-link antigen and to form immune complexes under most conditions ^[2]. Thus, mutations that prevent Fab-arm exchange in vivo should be considered when designing therapeutic IgG4.

The constant region of Anti-hCD20-hIgG4 (S228P) contains the S228P mutation which reduces Fab-arm exchange by stabilizing the disulfides in the core-hinge of the IgG4 molecules ^[1].

Anti-hCD20-hIgG4 (S228P) was generated by recombinant DNA technology. It has been produced in CHO cells and purified by affinity chromatography with protein G.

References:

1. Labrijn AF. et al., 2009. Therapeutic IgG4 antibodies engage in Fab-arm exchange with endogenous human IgG4 in vivo. Nat Biotechnol. 27(8):767-71.
2. van der Neut Kolfschoten M. et al., 2007. Anti-inflammatory activity of human IgG4 antibodies by dynamic Fab arm exchange. Science. 317(5844):1554-7.

Figures

Comparison of ADCC potency for native and engineered anti-human CD20 antibody isotypes: Raji-Null cells were incubated with gradient concentrations of Anti-hCD20 or Anti-β-galactosidase (β-gal) mAbs for 1 hour. Jurkat-Lucia™ NFAT-CD16 effector cells were then co-incubated with targets cells for 6 hours. NFAT activation, reflecting the induced ADCC response, was assessed by determining Lucia luciferase activity in the supernatant using QUANTI-Luc™. Percentages of the maximal response normalized to the IgG1 isotype are shown

Specifications

Clonality: Engineered monoclonal antibody

Specificity: Targets cells expressing human CD20

Isotype: Human IgG4, kappa

Application :
Anti-hCD20-hIgG4 (S228P) can be compared with anti-hCD20-hIgG4 to study the impact of Fab-arm exchange.

Quality control

• The absence of bacterial contamination (e.g. endotoxins and peptidoglycans) is controlled using HEK-Blue™ TLR2 and HEK-Blue™ TLR4 cells.
• The recognition of human CD20 with this antibody is tested using flow cytometry.

Contents

• 100 µg purified anti-hCD20-hIgG4 (S228P) antibody, provided azide-free and lyophilized.

Product is shipped at room temperature.

Store lyophilized antibody at -20°C.

Lyophilized product is stable for 1 year.