IZI-06.1 is a humanized anti-TNFR1 single-chain fragment variable (scFv) that selectively inhibits binding of tumor necrosis factor (TNF) and lymphotoxin alpha to tumor necrosis factor receptor 1 (TNFR1) but not TNFR2. Recently, IZI-06.1 was converted into a fully human IgG1 antibody (ATROSAB) for the treatment of inflammatory diseases. Here, we compare the bivalent ATROSAB with a monovalent scFv-human serum albumin (HSA) fusion protein lacking any antibody-associated effector functions and possessing approximately only half the molecular mass of an IgG, which should facilitate accumulation in inflamed tissues. Furthermore, the half-life of the scFv should be strongly extended while maintaining monovalent binding, avoiding a possible signal transduction by receptor cross-linking in the absence of TNF. The scFv-HSA fusion protein was produced by stably transfected Chinese hamster ovary cells and purified by affinity chromatography. The fusion protein bound specifically to TNFR1 in enzyme-linked immunosorbent assay and TNFR1-transfected mouse embryonic fibroblasts. Affinity determined by quartz crystal microbalance was reduced compared with ATROSAB, which resulted also in a reduced inhibitory activity. Compared with the scFv fragment, the half-life of the fusion protein was significantly increased, although not reaching the long half-life of ATROSAB. In summary, the scFv-HSA may provide an alternative to the full-length IgG1 with the ability to selectively inhibit TNFR1 and exploiting the pharmacokinetic properties of albumin.

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http://dx.doi.org/10.1093/protein/gzt044DOI Listing

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