The anti-apoptotic protein BCL-2 has a crucial role in the regulation of apoptosis and in the development and progression of numerous malignancies. Overexpression of BCL-2 promotes the survival of cancer cells, making it an attractive therapeutic target. Although effective BCL-2 inhibitors have been developed recently (venetoclax, lisaftoclax, sonrotoclax), their clinical application is limited by adverse effects, reduced efficacy in specific malignancies, and the emergence of acquired resistance resulting from mutations in the BCL-2 protein. Targeted protein degradation has emerged as an alternative therapeutic strategy that exploits the ubiquitin-proteasome system to eliminate proteins of interest. Among these, heterobifunctional PROTAC molecules are currently most developed. They induce the formation of a ternary complex between the target protein, PROTAC and an E3 ligase, thereby promoting ubiquitination of the target protein and its proteasomal degradation. The development of PROTACs is frequently limited by their high molecular weight, poor solubility, limited membrane permeability and the small number of well-characterized E3 ligases. To overcome these limitations, novel targeted protein degradation strategies are being developed. Promising approaches include ByeTAC molecules, which directly recruit target proteins to the proteasome via the Rpn-13 receptor, heterobifunctional degraders that exploit alternative E3 ligases (DCAF11, DCAF16, RBBP7, GID-4 and FBXO22) as well as hydrophobic tags.
Previously synthesized PROTACs based on the BCL-2 inhibitor S55746 and ligands for the E3 ligases CRBN, VHL and IAP failed to induce efficient degradation of BCL-2. Therefore, the aim of this master`s thesis was to design and synthesize heterobifuncional compounds based on the BCL-2 inhibitor S55746 that exploit alternative effector systems for targeted degradation of BCL-2, i.e. novel E3 ligases, Rpn-13 subunit of the proteasome, and hydrophobic tagging approach. We first synthesized a piperazine analogue of S55746, various electrophilic warheads (that bind E3 ligases) and ligands for GID-4 and Rpn-13. The S55746 analogue was coupled to the synthesized ligands and electrophilic warheads via an amide bond formation. Despite multiple synthetic steps, we successfully synthesized nine final heterobifuncional compounds that were evaluated for their ability to induce the degradation of BCL-2 and BCL-XL using a reporter system in HeLa cells. Compound 43, which contains a ligand for the E3 ligase FBXO22, proved to be the most interesting. At a concentration of 1 μM we observed a low level of BCL-2 degradation. Despite suboptimal results, the molecule will serve as a basis for further research into the structure-degradation relationship of molecules targeting BCL-2.
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