CDK1, cyclin-dependent kinase 1, is the key regulator of the cell cyle. In complex with cyclin B, it plays a crucial role in the cell transition from the G2 phase to the M phase and M phase progression. CDK1 is often overexpressed and overactive in cancer cells, which is associated with uncontrolled cell proliferation and the development of various cancers, including breast, lung, pancreatic and ovarian cancer. Therefore, CDK1 represents an important and promising target for the development of new cancer therapies. Among the known CDK1 inhibitors, the molecule RO-3306 stands out due to its high selectivity making it a good candidate for further chemical modifications. Targeted protein degradation has emerged as a promising medicinal chemistry approach with high therapeutic potencial. Proteolysis-targeting chimeras (PROTACs) have several advantages compared to traditional inhibitors, such as increased efficacy, reduced toxicity, and improved selectivity.
As a part of this master's thesis, we focused on investigating RO-3306 derivatives that could be suitable for incorporation into heterobifunctional molecules, specifically PROTAC molecules targeting CDK1. Since the crystal structure of the CDK1-RO-3306 complex is not yet known, it is necessary to use alternative approaches to identify the appropriate modification site on the inhibitor. Namely, linker attachment should not negatively affect binding to the target. Therefore, we synthesized several RO-3306 derivatives and after enzymatic evaluation determined which modification retains the affinity for CDK1. During the course of our research, we synthesized five RO-3306 derivatives, which were differed in the structure of their linker and the type of attachment point. Our main goal was to explore how the linker structure or different functional groups attached at the specific position of the inhibitor affect inhibitory activity against CDK1. The activity of the final compounds was evaluated with commercially available CDK1 inhibition assay. The results showed varying levels of inhibitory activity among our compounds, revealing that some structural modifications lead to a loss of inhibitory activity against CDK1, whereas with others we can prepare RO-3306 analogs, which partially retain inhibitory activity and are therefore appropriate for incorporation into heterobifunctional molecules.
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