Tumor necrosis factor-α (TNF-α) is a key proinflammatory cytokine that drives rheumatoid arthritis (RA) pathogenesis by sustaining synovial inflammation and joint destruction via binding to its receptors (TNFR1 and TNFR2), making this interaction a core therapeutic target. Ellagic acid (EA), a dietary polyphenol naturally abundant in pomegranates, berries, and nuts, is well known for its anti-inflammatory and antioxidant properties. In this study, we investigated whether EA targets the TNF-α pathway using a combination of molecular interaction analyses and cellular assays. Surface plasmon resonance (SPR) revealed that EA binds directly to TNF-α (K_D = 3.588 × 10^-6 M), TNFR1 (K_D = 6.488 × 10^-6 M), and TNFR2 (K_D = 7.952 × 10^-6 M) with high affinity. Molecular dynamics simulations and competitive SPR assays demonstrated that EA disrupts the TNF-α–TNFR interaction. Functionally, EA (0.625–2.5 μM) suppressed TNF-α-induced apoptosis in L929 cells and inhibited TNF-α-triggered NF-κB activation in 293-TNF-α Res (NF-κB) cells. In RA-relevant cell models using human synovial fibroblasts (HFLS and MH7A), EA attenuated TNF-α-stimulated NF-κB signaling by reducing phosphorylation of IKKα/β, IκBα, and p65, and by blocking p65 nuclear translocation. Notably, EA did not inhibit LPS-induced NF-κB activation, indicating pathway selectivity. These findings demonstrate that EA inhibits NF-κB activation by directly targeting the TNF-α–TNFR interaction, highlighting its potential as a food-derived functional ingredient for managing RA and related inflammatory conditions.

Rheumatoid arthritis (RA) is a chronic systemic autoimmune disorder that has long been hampered by limited treatment efficacy and significant side effects. Tumor necrosis factor-α (TNF-α) plays a pivotal role in RA pathogenesis by binding to its receptor (TNFR) and activating the downstream nuclear factor-kappa B (NF-κB) signaling pathway, which promotes the transcription of pro-inflammatory genes and perpetuates disease progression. Blocking the TNF-α–TNFR interaction thus represents a promising therapeutic strategy for RA. In this study, we identified a natural compound, 1-norbetulonic acid (DOCA), that disrupts the binding between TNF-α and TNFR, leading to therapeutic benefits in RA. Our results show that DOCA inhibits TNF-α-induced activation of the NF-κB pathway in human fibroblast-like synoviocytes and MH7A cells, and prevents nuclear translocation of the p65 subunit. Notably, DOCA demonstrated significant therapeutic efficacy in a mouse model of RA. Together, these findings support the hypothesis that DOCA alleviates RA by blocking TNF-α–TNFR signaling, underscoring its potential as a natural product-derived inhibitor of this interaction and highlighting a viable approach for the discovery of TNF-α/TNFR-targeted natural therapeutics.