Identification, validation and functional characterization of targets of myxobacterial compounds with potential for pharmacological cancer treatment

• Applicant: Professor Dr. Stephan A. Sieber
• Subject Area: Pharmacy ; Biological and Biomimetic Chemistry
• Term: from 2010 to 2014
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 159663715

This project focuses on target identification and validation via chemical proteomic based methods. In the first funding period we utilized functionalized pretubulysin derivatives for target analysis. While activity based protein profiling (ABPP) with cell permeable photoprobes revealed tubulin as target protein, affinity chromatography with immobilized pretubulisin provided evidence for the specific binding to proteasomal subunits. Especially the latter finding was unexpected and will be further validated during the next funding period. In the new funding period we will use our established chemical proteomic platform for the target characterization of soraphen A and archazolid. The design of both functionalized molecules will be carried out in close collaboration with P2 and P9. In order to gain quantitative data we will use stable isotope labeling in cell culture (SILAC). This method will allow to immediately rank the identified protein hits according to their binding specificity and thus increase the confidence in the obtained results. In addition, we will further refine the method of ABPP and implement a photolinker free native enrichment strategy that would allow to reduce the extent of structural modifications on the target compound. All novel hits will be validated by in depth biological characterization. Putative targets for archazolid have been predicted based on computational and biochemical data and will be also validated within these studies. With this comprehensive target identification/validation strategy we will provide important insights into the mechanism of action of the FOR compounds and thus help to generate hypotheses on the biological activity and mode of action.

• DFG Programme: Research Units
• Subproject of: FOR 1406: Exploiting the Potential of Natural Compounds: Myxobacteria as Source for Leads, Tools and Therapeutics in Cancer Research