Development of selective kinase inhibitors that target the ATP binding site continues to be a challenge largely due to similar binding pockets. Palbociclib is a cyclin-dependent kinase inhibitor that targets the ATP binding site of CDK4 and CDK6 with similar potency. The enzymatic function associated with the kinase can be effectively probed using kinase inhibitors however the kinase independent functions cannot. Herein, we report a palbociclib based PROTAC that selectively degrades CDK6 while sparing the homolog CDK4. We used competition studies to characterize the binding and mechanism of CDK6 degradation.

Design, synthesis, and evaluation of α-methylene-γ-butyrolactone analogues and their evaluation as anticancer agents is described. SAR identified a spirocyclic analogue 19 that inhibited TNFα-induced NF-κB activity, cancer cell growth and tumor growth in an ovarian cancer model. A second iteration of synthesis and screening identified 29 which inhibited cancer cell growth with low-μM potency. Our data suggest that an isatin-derived spirocyclic α-methylene-γ-butyrolactone is a suitable core for optimization to identify novel anticancer agents.

The carboxy terminal BRCT domains of the breast cancer susceptibility gene 1 (BRCA1) bind to a plethora of phosphorylated proteins through a pSXXF consensus recognition motif. BRCT-protein binding regulates key cellular functions such as lipogenesis, cell-cycle checkpoint control and DNA damage response. Identification of the minimal binding sequence and defining the key interactions responsible for biological activity are critical steps in the peptidomimetic design process. Here, we report a systematic structure activity relationship study that maps the BRCT(BRCA1)-pSXXF binding interface. The study has led to the identification of peptides with nanomolar binding affinities comparable to the previously reported 13-mer peptides. The results also provide a clear description of the pSXXF-BRCT interface, which is essential for developing small molecule inhibitors via the peptidomimetic approach.

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The C-terminal domain of BRCA1 (BRCT) is involved in the DNA repair pathway by recognizing the pSXXF motif in interacting proteins. It has been reported that short peptides containing this motif bind to BRCA1(BRCT) in the micromolar range with high specificity. In this work, the binding of pSXXF peptides has been studied computationally and experimentally in order to characterize their interaction with BRCA1(BRCT). Elucidation of the contacts that drive the protein-ligand interactions is critical for the development of high affinity small-molecule BRCA1 inhibitors. Molecular dynamics simulations revealed the key role of threonine at the peptide P+2 position in providing structural rigidity to the ligand in the bound state. The mutation at P+1 had minor effects. Peptide extension at the N-terminal position with the naphthyl amino acid exhibited a modest increase in binding affinity, what could be explained by the dispersion interaction of the naphthyl side-chain with a hydrophobic patch. Three in silico end-point methods were considered for the calculation of binding free energy. The Molecular Mechanics Poisson-Boltzmann Surface Area (MM/PB-SA) and the Solvated Interaction Energy (SIE) gave reasonable agreement with experimental data, exhibiting a Pearlman predictive index of 0.71 and 0.78, respectively. The MM-Quantum Mechanics-Surface Area (MM-QMSA) method yielded improved results, which was characterized by a Pearlman index of 0.78. The correlation coefficients were 0.59, 0.61 and 0.69, respectively. The ability to apply a QM level of theory within an end-point binding free energy protocol may provide a way for a consistent improvement of accuracy in computer-aided drug design.