Translation is an essential cellular process whose deregulation is associated with alterations in cell growth regulation, cell cycle progression, and apoptotic responses. There is much evidence supporting the notion that aberrant control of protein translation contributes to neoplastic transformation. Signaling pathways (e.g. - ras and Akt) that regulate the translational machinery are activated in many human cells, over-expression of certain translation factors can lead to malignant transformation, and several components of the translational apparatus are over-expressed in human cancers. Indeed, elevated expression levels of elF4E (the mRNA cap binding protein involved in ribosome recruitment) leads to transformation in murine cancer model and is implicated in chemoresistance. Rapamycin, an inhibitor of the ribosome recruitment step, shows significant anti-cancer activity and is currently being tested in clinical trials. In this grant application, we propose to develop a series of HTS assays that will form the foundation of a chemical biology program aimed at better understanding the mechanism of translation, as well as the role that deregulation of this process plays in tumor progression. The HTS assays will target translation initiation factors involved in eukaryotic ribosome recruitment and will be used to identify compounds that can specifically inhibit this process.
Our specific aims are to: i) target the interaction between elF4E and the mRNA cap structure for HTS assay design; ii) develop an HTS assay that monitors the helicase activity of elF4A, an ATP-dependent RNA helicase that unwinds mRNA secondary structure in the 5' UTR of mRNAs; iii) develop an HTS assay for inhibitors of elF4B activity, an RNA binding factor that functions in conjunction with elF4A to facilitate mRNA/ribosome binding; and iv) develop an HTS assay to prevent formation of the elF4E/4E-BP inhibitory complex, a heterodimer whose formation is stabilized by rapamycin.
