Discovery of a novel selective small molecule interleukin-36 receptor antagonist for inflammation and cancer

Abstract

Interleukin-36 (IL-36) is a pivotal driver of inflammatory responses in autoimmune disorders, including psoriasis, inflammatory bowel disease, and rheumatoid arthritis, and it also contributes to tumor progression. Upon activation, IL-36 triggers the release of pro-inflammatory cytokines such as C-X-C Motif Chemokine Ligand 1 (CXCL1), tumor necrosis factor-alpha (TNF-α), IL-6, and IL-8, exacerbating disease symptoms and fostering a tumor-supportive microenvironment. Despite its critical role, few IL-36 receptor (IL-36R) targeting molecules have been reported, and no specific small-molecule antagonist with confirmed receptor-level competition has been identified. Here, we present IRA10 and its potent derivative, IRA10L, as the first small molecules specifically designed to inhibit IL-36R. Molecular docking suggested strong binding of IRA10L to IL-36R, which molecular dynamics simulations further confirmed by demonstrating stable receptor–ligand interactions. Molecular mechanics poisson-boltzmann surface area (MMPBSA) binding free energy calculations revealed favorable energetics for IRA10L, correlating with its superior antagonistic activity compared with IRA10. Experimentally, IRA10L significantly reduced CXCL1 production across IL-36 isoforms alpha/beta/gamma (IL-36α/β/γ) and downregulated TNF-α, IL-6, and IL-8 expression in a dose-dependent manner. Western blot analyses showed inhibition of IL-36R. p65, Extracellular signal-regulated kinase (ERK), and p38 phosphorylation, blocking IL-36-driven downstream signaling. IRA10L also effectively suppressed IL-36-induced cancer cell metabolic activity, migration, spheroid growth, and colony formation. Biophysical assays, including surface plasmon resonance (SPR), Schild analysis, and competitive ELISA, confirmed direct competition of IRA10L with IL-36 ligands for receptor binding. These findings establish IRA10L as a specific, competitive IL-36R antagonist, highlighting its potential as a targeted therapeutic for IL-36-mediated inflammation and cancer and providing a foundation for future drug development. © 2025 Elsevier Inc.