Neutralizing the action of several cytokines with recombinant proteins corresponding to the soluble extracellular receptor chain has proven beneficial in the treatment of various autoimmune diseases in the past years. Interleukin-33 (IL-33) is a cytokine that signals through a receptor complex consisting of ST2 and IL-1RAcP, which initiates multiple pro-inflammatory signalling pathways. Blocking the IL-33/ST2 signalling axis with the soluble (extracellular) form of the ST2 receptor (sST2) has been shown to be beneficial in experimental models of asthma and arthritis. One major drawback of the use of soluble cytokine receptor proteins, however, is their relative low ligand binding affinity. Indeed, recruitment of the IL-1RAcP co-receptor upon IL-33 stimulation has been shown to significantly increase the affinity for IL-33.

Here we report a new approach to interfere with IL-33 function. We have generated a fusion protein consisting of the extracellular domains of mouse ST2 and IL-1RAcP, separated by a flexible linker (referred to as IL33trap). His-tagged recombinant IL-33trap and sST2 were expressed in human HEK293T cells and purified to homogeneity from conditioned medium by metal affinity chromatography. IL-33 neutralizing activity of the purified proteins was compared in an IL-33 bioassay, in which we measure IL-33 induced NF-κB reporter gene activation. Interestingly, the IL-33trap fusion protein is considerably more potent than sST2 in neutralizing mouse IL-33 activity. A similar approach was followed to produce an IL-33trap for human IL-33, and again the IL33trap was found to be much more potent than sST2. Together, these studies demonstrate the improved potential of IL-33trap cytokine blockers for therapeutic use. IL33trap and sST2 proteins are currently further analysed in in vitro binding affinity studies and in vivo mouse disease models.