Lithocholic acid is an Eph-ephrin ligand interfering with Eph-kinase activation

• Published in: PloS one 2011-03, Vol.6 (3), p.e18128-e18128
• Main Authors: Giorgio, Carmine, Hassan Mohamed, Iftiin, Flammini, Lisa, Barocelli, Elisabetta, Incerti, Matteo, Lodola, Alessio, Tognolini, Massimiliano
• Format: Article
• Language: English
• Subjects: Acids; Activation; Adenocarcinoma; Bile; Bile acids; Binding; Binding, Competitive - drug effects; Biology; Cell Adhesion - drug effects; Cell Death - drug effects; Cell Line, Tumor; Cell Shape - drug effects; Chemistry; Colon; Colorectal cancer; Enzyme Activation - drug effects; Enzyme-linked immunosorbent assay; Eph protein; EphA2 protein; Ephrins - metabolism; Epidermal growth factor receptors; Genetic transformation; Homeostasis; Humans; Inhibition; Intestine; Kinases; Ligands; Lithocholic Acid - metabolism; Lithocholic Acid - pharmacology; Medicine; Metabolites; Metastases; Phenotypes; Phosphorylation; Phosphorylation - drug effects; Prostate; Prostate cancer; Protein Binding - drug effects; Protein interaction; Protein-protein interactions; Receptor, Epidermal Growth Factor - metabolism; Receptor, IGF Type 1 - metabolism; Receptors; Receptors, Eph Family - metabolism; Receptors, Vascular Endothelial Growth Factor - metabolism; Rounding; Signaling; Transformation; Tumor cell lines; Tumorigenesis; Tumors; Tyrosine; Vascular endothelial growth factor; Vascular endothelial growth factor receptors
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0018128

Eph-ephrin system plays a central role in a large variety of human cancers. In fact, alterated expression and/or de-regulated function of Eph-ephrin system promotes tumorigenesis and development of a more aggressive and metastatic tumour phenotype. In particular EphA2 upregulation is correlated with tumour stage and progression and the expression of EphA2 in non-transformed cells induces malignant transformation and confers tumorigenic potential. Based on these evidences our aim was to identify small molecules able to modulate EphA2-ephrinA1 activity through an ELISA-based binding screening. We identified lithocholic acid (LCA) as a competitive and reversible ligand inhibiting EphA2-ephrinA1 interaction (Ki =  49 µM). Since each ephrin binds many Eph receptors, also LCA does not discriminate between different Eph-ephrin binding suggesting an interaction with a highly conserved region of Eph receptor family. Structurally related bile acids neither inhibited Eph-ephrin binding nor affected Eph phosphorylation. Conversely, LCA inhibited EphA2 phosphorylation induced by ephrinA1-Fc in PC3 and HT29 human prostate and colon adenocarcinoma cell lines (IC(50)  = 48 and 66 µM, respectively) without affecting cell viability or other receptor tyrosine-kinase (EGFR, VEGFR, IGFR1β, IRKβ) activity. LCA did not inhibit the enzymatic kinase activity of EphA2 at 100 µM (LANCE method) confirming to target the Eph-ephrin protein-protein interaction. Finally, LCA inhibited cell rounding and retraction induced by EphA2 activation in PC3 cells. In conclusion, our findings identified a hit compound useful for the development of molecules targeting ephrin system. Moreover, as ephrin signalling is a key player in the intestinal cell renewal, our work could provide an interesting starting point for further investigations about the role of LCA in the intestinal homeostasis.