title: Development of biochemical assays for the identification of eIF4E-specific inhibitors.
abstract: Control of mRNA translation plays a critical role in cell growth, proliferation, and differentiation and is tightly regulated by AKT and RAS oncogenic pathways. A key player in the regulation of this process is the mRNA 5' cap-binding protein, eukaryotic translation initiation factor 4E (eIF4E). eIF4E contributes to malignancy by selectively enabling the translation of a limited pool of mRNAs that generally encode key proteins involved in cell cycle progression, angiogenesis, and metastasis. Several data indicate that the inhibition of eIF4E in tumor cell lines and xenograft models impairs tumor growth and induces apoptosis; eIF4E, therefore, can be considered a valuable target for cancer therapy. Targeting the cap-binding pocket of eIF4E should represent a way to inhibit all the eIF4E cellular functions. We present here the development and validation of different biochemical assays based on fluorescence polarization and surface plasmon resonance techniques. These assays could support high-throughput screening, further refinement, and characterization of eIF4E inhibitors, as well as selectivity assessment against CBP80/CBP20, the other major cap-binding complex of eukaryotic cells, overall providing a robust roadmap for development of eIF4E-specific inhibitors.
authors.affiliation: Biotechnology Department, Nerviano Medical Sciences, Nerviano (MI), Italy.
authors.forename: Carlo
authors.initials: C
authors.lastname: Visco
authors.forename: Claudia
authors.initials: C
authors.lastname: Perrera
authors.forename: Sandrine
authors.initials: S
authors.lastname: Thieffine
authors.forename: Federico Riccardi
authors.initials: FR
authors.lastname: Sirtori
authors.forename: Roberto
authors.initials: R
authors.lastname: D'Alessio
authors.forename: Paola
authors.initials: P
authors.lastname: Magnaghi
mesh_terms: D055808:Drug Discovery / Q000379:methods; D039561:Eukaryotic Initiation Factor-4E / Q000037:antagonists & inhibitors* / Q000302:isolation & purification / Q000378:metabolism; D005454:Fluorescence Polarization / Q000379:methods*; D006801:Humans; D007700:Kinetics; D039502:Nuclear Cap-Binding Protein Complex / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D012315:RNA Caps / Q000187:drug effects; D015203:Reproducibility of Results; D020349:Surface Plasmon Resonance / Q000379:methods*
chemical_list: D039561:Eukaryotic Initiation Factor-4E; D039502:Nuclear Cap-Binding Protein Complex; D012315:RNA Caps