Targeting the inter-monomeric space of TNFR1 pre-ligand dimers: A novel binding pocket for allosteric modulators

Tumor necrosis factor (TNF) receptor 1 (TNFR1) plays a central role in signal transduction mediating inflammation and cell death associated with autoimmune and neurodegenerative disorders. Inhibition of TNFR1 signaling is a highly sought-after strategy to target these diseases. TNFR1 forms pre-ligand dimers held together by the pre-ligand assembly domain (PLAD), which is essential for receptor signaling. TNFR1 dimers form the crucial points of interaction for the entire receptor signaling complex by connecting TNF ligand bound trimeric receptors. While previous studies have shown the feasibility of disrupting TNFR1 dimeric interactions through competitive mechanism that targets the PLAD, our recent studies have demonstrated that small molecules could also bind PLAD to modulate TNFR1 signaling through an allosteric mechanism. Importantly, these allosteric modulators alter receptor dynamics and propagate long-range conformational perturbation that involves reshuffling of the receptors in the cytosolic domains without disrupting receptor-receptor or receptor-ligand interactions. In this study, we perform molecular docking of previously reported allosteric modulators on the extracellular domain of TNFR1 to understand their binding sites and interacting residues. We identify the inter-monomeric space between TNFR1 pre-ligand dimers as a novel binding pocket for allosteric modulators. We further conduct pharmacological analyses to understand the bioactivity of these compounds and their interacting residues and pharmacological properties. We then provide insights into the structure-activity relationship of these allosteric modulators and the feasibility of targeting TNFR1 conformational dynamics. This paves the way for developing new therapeutic strategies and designing chemical scaffolds to target TNFR1 signaling.