title: Lithocholic acid is an Eph-ephrin ligand interfering with Eph-kinase activation.
abstract: 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.
authors.affiliation: Dipartimento di Scienze Farmacologiche, Biologiche e Chimiche Applicate, Università di Parma, Parma, Italy.
authors.forename: Carmine
authors.initials: C
authors.lastname: Giorgio
authors.forename: Iftiin
authors.initials: I
authors.lastname: Hassan Mohamed
authors.forename: Lisa
authors.initials: L
authors.lastname: Flammini
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Barocelli
authors.forename: Matteo
authors.initials: M
authors.lastname: Incerti
authors.forename: Alessio
authors.initials: A
authors.lastname: Lodola
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Tognolini
mesh_terms: D001667:Binding, Competitive / Q000187:drug effects; D002448:Cell Adhesion / Q000187:drug effects; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D048430:Cell Shape / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D036342:Ephrins / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D008024:Ligands; D008095:Lithocholic Acid / Q000378:metabolism* / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D017526:Receptor, IGF Type 1 / Q000378:metabolism; D036081:Receptors, Eph Family / Q000378:metabolism*; D040262:Receptors, Vascular Endothelial Growth Factor / Q000378:metabolism
chemical_list: D036342:Ephrins; D008024:Ligands; D008095:Lithocholic Acid; D066246:ErbB Receptors; D017526:Receptor, IGF Type 1; D036081:Receptors, Eph Family; D040262:Receptors, Vascular Endothelial Growth Factor