The rare bile acid isoallolithocholic acid (IALCA) is an EphA2 antagonist sparing FXR and TGR5 receptors

The role of the Eph/ephrin system is well recognized in various physiological and pathological processes, including acute inflammation and cancer. We previously discovered that the secondary bile acid lithocholic acid (LCA) is a competitive antagonist of Eph receptors. The utility of LCA as a pharmacological tool for investigating Eph/ephrin biology was hampered by its primary activity at the FXR and TGR5 receptors. A recent study of centenarians’ gut microbiomes revealed that a rare bile acid closely related to LCA, isoallolithocholic acid (IALCA), exerts marked protective effects on the intestinal epithelium, but its specific molecular target was unidentified. Considering the well-documented involvement of EphA2 in regulating intestinal epithelial/endothelial permeability, we asked whether IALCA could act through this receptor. Molecular docking and dynamics simulations predicted that IALCA binds within the ephrin-A1–binding pocket of EphA2. Our findings were validated through wet experiments, and IALCA emerged as a selective EphA2 inhibitor, blocking ephrin-A1 binding with low-micromolar potency. In functional studies, IALCA inhibited ephrin-A1–induced EphA2 phosphorylation, cell retraction, and rounding, confirming its antagonistic activity. Moreover, IALCA showed no detectable activity at the classical bile-acid receptors FXR, PXR, LXRα, or TGR5, thereby potentially linking its cellular and phenotypic effects to modulation of the Eph–ephrin system. As a final step, we demonstrated that IALCA also provides an attractive template for synthesizing new Eph antagonists. Overall, this work underscores the potential of the human gut microbiome as a reservoir of privileged chemical scaffolds for both fundamental pharmacology and therapeutic drug development.