Generation and characterization of small single domain antibodies inhibiting human TNF receptor 1 [Immunology]

December 23rd, 2014 by Steeland, S., Puimege, L., Vandenbroucke, R. E., Van Hauwermeiren, F., Haustraete, J., Devoogdt, N., Hulpiau, P., Leroux–Roels, G., Laukens, D., Meuleman, P., De Vos, M., Libert, C.

The cytokine TNF is a well-known drug target for several inflammatory diseases such as Crohn's disease (CD). Despite the great success of TNF blockers, therapy could be improved because of high costs and side-effects. Selective inhibition of TNFR1 signaling holds the potential to greatly reduce the pro-inflammatory activity of TNF, thereby preserving the advantageous immunomodulatory signals mediated by TNFR2. We generated a selective human TNFR1 inhibitor based on Nanobody (Nb) technology. Two anti-hTNFR1 Nbs were linked with an anti-albumin Nb to generate Nb Alb-70-96 named 'TNF Receptor-One Silencer' (TROS). TROS selectively binds and inhibits TNF/TNFR1 and LTa/TNFR1 signaling with good affinity and IC50, both of which are in the nanomolar range. Surface Plasmon Resonance analysis reveals that TROS competes with TNF for binding to hTNFR1. In HEK293T-cells, TROS strongly reduces TNF-induced gene expression, like IL8 and TNF in a dose-dependent manner, and in ex vivo cultured colon biopsies of CD patients, TROS inhibits inflammation. Finally, in liver chimeric humanized mice, TROS antagonizes inflammation in a model of acute TNF-induced liver inflammation, reflected in reduced human IL8 expression in liver and reduced IL6 levels in serum. These results demonstrate the considerable potential of TROS and justify the evaluation of TROS in relevant disease animal models of both acute and chronic inflammation, and eventually in patients.