BBB seminar: Wolfgang Link

Wolfgang Link
Experimental Therapeutics Program, Spanish National Cancer Research Centre, Madrid, Spain
 

High-content cellular imaging has increasingly met the challenges of high-throughput needs and facilitates the integration of disease relevant screens at early stages of the drug discovery process. In cancer biology the FOXO (Forkhead box, class O) subfamily of Forkhead transcription factors is of particular interest as activation of these proteins is associated with cell cycle arrest and apoptosis and they have been suggested to function as tumor suppressors. In order to identify chemical probes to study the role of FOXO proteins as well as compounds with therapeutic potential we developed a high-throughput cellular imaging assay that follows the intracellular location of the FOXO3a protein in tumor cells. Nuclear accumulation of fluorescent signals (GFP) upon treatment with the known PI3K inhibitors LY294002, Wortmannin and PI-103 was dose dependent and in good agreement with the IC₅₀ values reported previously for PI3K inhibition in vitro. In order to investigate the FOXO shuttling system we screened a panel of 73 small chemical compounds with known biological activity using three different concentrations per compound. We identified 17 compounds capable of inducing nuclear accumulation of GFP-FOXO, including compounds known to interfere with known biochemical components of the PI3K/Akt signaling, nuclear export and Ca²⁺/calmodulin (CaM)-dependent signaling events. Our data illustrate the potential of chemical genetics when combined with robust and sensitive high-content technology to study the complex network of signaling pathways.

Host: Jim Lorens <jim.lorens@biomed.uib.no>, Department of Biomedicine