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Covalent dimerization of camelidae anti‐human TNF‐alpha single domain antibodies by the constant kappa light chain domain improves neutralizing activity

2010; Wiley; Volume: 106; Issue: 1 Linguagem: Inglês

10.1002/bit.22653

1097-0290

Martin Giersberg, Doreen M. Floß, Sergey M. Kipriyanov, Udo Conrad, Jürgen Scheller,

Toxin Mechanisms and Immunotoxins

The tumor necrosis factor-alpha (TNFalpha) plays an important role in a number of chronic inflammatory disorders. Monoclonal camelidae variable heavy chain domain-only antibodies (V(H)H) have been developed to antagonize the action of human TNFalpha (anti-TNF-V(H)H). Here we describe a strategy to obtain functional dimeric anti-TNF-V(H)H molecules, based on the C-terminal fusion of a kappa light chain domain to the anti-TNF-V(H)H. The resulting fusion protein was transiently expressed by use of viral vectors in Nicotiana benthamiana((Nb)) leaves and purified. Competitive ELISA and cell cytotoxicity assays revealed that the dimerized anti-(Nb)TNF-V(H)H(Ckappa) proteins blocked TNFalpha-activity more effectively than either the monomeric Escherichia coli((Ec)) produced anti-(Ec)TNF-V(H)H or the monomeric anti-(Nb)TNF-V(H)H(Ckappa). We suggest that enhanced inhibition shown by dimeric anti-(Nb)TNF-V(H)H(Ckappa) proteins is achieved by an increase in avidity.