A toolbox of promiscuous immobilisation chemistries to identify the targets of natural compounds by chemoproteomics (PromisChemProt)

• Applicant: Professor Dr. Bernhard Küster, since 10/2022
• Subject Area: Analytical Chemistry ; Biological and Biomimetic Chemistry
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 452256511

Natural products have evolved to benefit their producing organisms: contrary to synthetic man-made compounds, they have been selected and optimized by evolution to interact with biological targets for example as attack or defense mechanisms. Natural products thus constitute a promising pool of innovative chemical biology tools and potential drugs that would benefit from systematic exploration. Target proteins can be involved in essential cellular pathways of other organisms, and are many times not yet validated drug targets for human medicine. Phenotypic screenings have uncovered many natural compounds with anti-tumor activity but it is often unclear which cellular targets are engaged and lead to the observed phenotypic outcome. Yet, acquiring this information, i.e. target deconvolution, is of utmost importance to convert observable phenotypes into actionable novel chemotypes and/or novel biological targets. Target deconvolution of those screening hits is often deterred by the lack of generic experimental approaches in the context of complex cellular systems. In particular, chemical synthesis of an ad-hoc linkable analogue is often used to interrogate the target space of a molecule. However natural products are often difficult to synthesize which complicates e.g. activity-based or affinity-based target deconvolution approaches. With this research proposal, we ambition to develop a robust chemoproteomics pipeline that allows to perform target deconvolution of any chosen unmodified natural product: More precisely, we will build a toolbox of chemistries able to immobilize small amounts of natural compounds on beads. We will investigate diazirine for carbene insertion, Mitsunobu reaction, gold catalyzed cycloaddition and alkoxylation as well as enyne metathesis as means to promiscuously react natural products with reagent-loaded beads. The obtained affinity matrices will enable the affinity purification of targets of natural compounds from cellular lysates and their characterization by quantitative mass spectrometry, building on our chemical proteomics expertise. Because of the wealth of alkenes and alcohol functionalities present in natural products, we anticipate that at least one of the utilized chemistries will produce a linkage compatible with the molecule:target binding event. Since such propitious linkage site cannot be predicted for unknown targets, our concept relies on the parallel agnostic use of a set of chemistries, which will be applied to forty natural compounds confirmed to act on cancer cell lines. Any confirmed causality link between the engagement of the newly identified target by the natural product and he phenotype will constitute an actionable starting point for pharmaceutical research, where the lack of validated targets is pressing for alternative drug discovery paradigms.

• DFG Programme: Research Grants
• Ehemaliger Antragsteller: Dr. Guillaume Médard, until 9/2022