ABSTRACT
Cytokine release syndrome (CRS), commonly known as cytokine storm, is an acute systemic inflammatory response that is a significant global health threat. Interleukin-6 (IL-6) and interleukin-1 (IL-1) are key pro-inflammatory cytokines involved in CRS and are hence critical therapeutic targets. Current antagonists, such as tocilizumab and anakinra, target IL-6R/IL-1R but have limitations due to their long half-life and systemic anti-inflammatory effects, making them less suitable for acute or localized treatments. Here we present the de novo design of small protein antagonists that prevent IL-1 and IL-6 from interacting with their receptors to activate signaling. The designed proteins bind to the IL-6R, GP130 (an IL-6 co-receptor), and IL-1R1 receptor subunits with binding affinities in the picomolar to low-nanomolar range. X-ray crystallography studies reveal that the structures of these antagonists closely match their computational design models. In a human cardiac organoid disease model, the IL-1R antagonists demonstrated protective effects against inflammation and cardiac damage induced by IL-1ß. These minibinders show promise for administration via subcutaneous injection or intranasal/inhaled routes to mitigate acute cytokine storm effects.
Subject(s)
Cytokine Release Syndrome , Interleukin-6 , Humans , Cytokine Release Syndrome/drug therapy , Interleukin-6/metabolism , Interleukin-6/antagonists & inhibitors , Crystallography, X-Ray , Receptors, Interleukin-6/antagonists & inhibitors , Receptors, Interleukin-6/metabolism , Interleukin-1/metabolism , Interleukin-1/antagonists & inhibitors , Interleukin 1 Receptor Antagonist Protein/pharmacology , Interleukin 1 Receptor Antagonist Protein/chemistry , Interleukin 1 Receptor Antagonist Protein/metabolism , Drug Design , Cytokine Receptor gp130/metabolism , Cytokine Receptor gp130/antagonists & inhibitors , Cytokine Receptor gp130/chemistry , Protein Binding , Signal Transduction/drug effects , Receptors, Interleukin-1 Type I/antagonists & inhibitors , Receptors, Interleukin-1 Type I/metabolismABSTRACT
While there has been progress in the de novo design of small globular miniproteins (50-65 residues) to bind to primarily concave regions of a target protein surface, computational design of minibinders to convex binding sites remains an outstanding challenge due to low level of overall shape complementarity. Here, we describe a general approach to generate computationally designed proteins which bind to convex target sites that employ geometrically matching concave scaffolds. We used this approach to design proteins binding to TGFßRII, CTLA-4 and PD-L1 which following experimental optimization have low nanomolar to picomolar affinities and potent biological activity. Co-crystal structures of the TGFßRII and CTLA-4 binders in complex with the receptors are in close agreement with the design models. Our approach provides a general route to generating very high affinity binders to convex protein target sites.