Inhibition of the EphA2-Sam/Ship2-Sam Association through Peptide Ligands: Studying the Combined Effect of Charge and Aromatic Character

The Sam (sterile alpha motif) domain from the lipid phosphatase Ship2 binds the Sam domain from the EphA2 receptor to negatively regulate receptor endocytosis and degradation. This interaction is primarily linked to pro-oncogenic effects. We report on the design and evaluation of EphA2-Sam/Ship2-Sam peptide inhibitors provided with positive charges and different aromatic characters. Starting from the sequence of previously identified Ship2-Sam targeting peptides, an in silico approach was set up to predict higher affinity peptide ligands. A few peptides were experimentally tested through an interdisciplinary approach. Interaction studies were performed by nuclear magnetic resonance spectroscopy and biolayer interferometry. 3D models of Ship2-Sam/peptide complexes were predicted by AlphaFold2. Cell-based assays were carried out to investigate whether such peptide sequences might have an influence on EphA2 signaling. The approach led to the identification of novel Ship2-Sam ligands and shed further light on original approaches to design inhibitors of the Ship2-Sam/EphA2-Sam interaction.