M. Mitrovic, Francesco Greco, Yiliam Cruz Garcia, Aleksandar Lučić, Lasse Hoffmann, Rohit Chander, Julia Schönfeld, Nick Liebisch, Saran Aswathaman Sivashanmugam, Martin P. Schwalm, Markus Egner, Max Lewandowski, Daniel Merk, Viktoria Morasch, Elmar Wolf, Susanne Müller, Thomas Hanke, Ewgenij Proschak, Kerstin Hiesinger, Stefan Knapp
Targeted protein degradation is one of the fastest developing fields in medicinal chemistry and chemical biology. Despite significant development in assay technologies and inhibitor discovery, the development of PROTACs remains a challenging endeavor since rational design approaches remain widely elusive. Our workflow eliminates the rate-limiting step of classic synthesis, namely compound purification, and pairs it with high-throughput, semi-automated plate-based synthesis, and direct cellular assay evaluation. We applied this direct-to-biology approach to four diverse targets, demonstrating the general applicability of this technology. PROTAC synthesis was realized by using the highly efficient copper-catalyzed azide-alkyne cycloaddition reaction. This simplified reaction setup enabled synthesis in the nanomole scale with reaction volumes as low as 5 μL. The high-throughput strategy allows hundreds of PROTACs to be synthesized and evaluated within a few days, facilitating comprehensive assessment of target degradability, rapid hit identification, and selection of the most suitable E3 ligase for degrader development.