Dual-targeting WEE1/AURK-A kinase proteolysis targeting chimera (PROTAC) for cancer therapy

• Applicants: Professor Dr. Stefan Hüttelmaier; Professor Dr. Wolfgang Sippl
• Subject Area: Pharmacy
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 553776460

The project aims to develop a novel anti-cancer strategy by dual targeting of two important kinases, WEE1 and Aurora A kinase (AURKA), using a proteolysis targeting chimera (PROTAC) approach. The rationale of this approach relies on the role of these kinases in DNA replication, cell cycle regulation and their significance for the survival of cancer cells. Chemotherapeutic agents and high cell division rates cause replication stress and DNA damage. This activates control kinases such as WEE1 and CHK1 to enable DNA repair and maintain cell viability. Inhibition of WEE1 proved promising in the induction of replication fork collapse and cell death. Inhibition of AURKA also leads to mitotic, spindle and replication defects, but also activates tumor suppressors and limits various oncogene activations. However, single-target inhibition is generally not therapeutically sufficient due to compensation mechanisms and the development of resistance. Accordingly, various dual inhibition strategies of WEE1/AURKA are currently evaluated for cancer therapy. PROTACs are novel molecules which degrade their target proteins via the ubiquitin-proteasome system. These frequently provide superior selectivity and efficacy in comparison to classical, small molecule inhibitors. In preliminary work, we have synthesized lead PROTACs based on novel potent dual inhibitors of WEE1/AURKA settling on Von-Hippel-Lindau (VHL) and Cereblon (CRBN) E3 ligase moieties. Both PROTACs promote the proteasomal degradation of both kinases in cancer cells, leading to significant reduction of tumor cell and organoid viability in the nanomolar range. We aim to optimize our initial PROTACs through structure-based design and activity testing in vitro, in tumor cell and organoid models. This includes the use of different E3 ligase ligands as well as optimization of the structure of the linkers and attachment points. The PROTACs and negative controls will be characterized in pharmacological selectivity studies, proteomic and genetic analyses as well as initial pharmacokinetic tests up to evaluation in organoids. The specificity and possibly further targets of developed PROTACs will be investigated via complementary proteome analyses. Current clinical data suggest that the resistance of tumor cells to certain PROTACs is broken with other E3 ubiquitin ligase elements, including DCAFs, KLHDC2 or RNF126. Accordingly, in addition to CRBN/VHL-based PROTACs, further proteolysis concepts will be evaluated for the two kinases. The antiproliferative effect of the dual WEE1/AURKA PROTACs will be characterized in selected cancer cells (neuroblastoma, ovarian carcinoma, and pancreatic carcinoma) at the molecular level and their clinical prospects will be explored in organoids.

• DFG Programme: Research Grants
• International Connection: France
• Cooperation Partner: Privatdozent Dr. Christophe Romier
• Co-Investigators: Professorin Dr. Simone Hettmer; Professor Dr. Jörg Kleeff