title: Structure-activity relationships and mechanism of action of Eph-ephrin antagonists: interaction of cholanic acid with the EphA2 receptor.
abstract: The Eph-ephrin system, including the EphA2 receptor and the ephrinA1 ligand, plays a critical role in tumor and vascular functions during carcinogenesis. We previously identified (3α,5β)-3-hydroxycholan-24-oic acid (lithocholic acid) as an Eph-ephrin antagonist that is able to inhibit EphA2 receptor activation; it is therefore potentially useful as a novel EphA2 receptor-targeting agent. Herein we explore the structure-activity relationships of a focused set of lithocholic acid derivatives based on molecular modeling investigations and displacement binding assays. Our exploration shows that while the 3-α-hydroxy group of lithocholic acid has a negligible role in recognition of the EphA2 receptor, its carboxylate group is critical for disrupting the binding of ephrinA1 to EphA2. As a result of our investigation, we identified (5β)-cholan-24-oic acid (cholanic acid) as a novel compound that competitively inhibits the EphA2-ephrinA1 interaction with higher potency than lithocholic acid. Surface plasmon resonance analysis indicates that cholanic acid binds specifically and reversibly to the ligand binding domain of EphA2, with a steady-state dissociation constant (K(D) ) in the low micromolar range. Furthermore, cholanic acid blocks the phosphorylation of EphA2 as well as cell retraction and rounding in PC3 prostate cancer cells, two effects that depend on EphA2 activation by the ephrinA1 ligand. These findings suggest that cholanic acid can be used as a template structure for the design of effective EphA2 antagonists, and may have potential impact in the elucidation of the role played by this receptor in pathological conditions.
authors.affiliation: Dipartimento di Scienze Farmacologiche, Biologiche e Chimiche applicate Università degli Studi di Parma, Viale delle Scienze 27/A, 43124 Parma, Italy.
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Tognolini
authors.forename: Matteo
authors.initials: M
authors.lastname: Incerti
authors.forename: Iftiin
authors.initials: I
authors.lastname: Hassan-Mohamed
authors.forename: Carmine
authors.initials: C
authors.lastname: Giorgio
authors.forename: Simonetta
authors.initials: S
authors.lastname: Russo
authors.forename: Renato
authors.initials: R
authors.lastname: Bruni
authors.forename: Barbara
authors.initials: B
authors.lastname: Lelli
authors.forename: Luisa
authors.initials: L
authors.lastname: Bracci
authors.forename: Roberta
authors.initials: R
authors.lastname: Noberini
authors.forename: Elena B
authors.initials: EB
authors.lastname: Pasquale
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Barocelli
authors.forename: Paola
authors.initials: P
authors.lastname: Vicini
authors.forename: Marco
authors.initials: M
authors.lastname: Mor
authors.forename: Alessio
authors.initials: A
authors.lastname: Lodola
mesh_terms: D001665:Binding Sites; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002793:Cholic Acids / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D003198:Computer Simulation; D015195:Drug Design; D036382:Ephrin-A1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D036104:Receptor, EphA2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013329:Structure-Activity Relationship; D020349:Surface Plasmon Resonance
chemical_list: D002793:Cholic Acids; D036382:Ephrin-A1; D036104:Receptor, EphA2; C007036:cholanic acid