Design, Synthesis, and Molecular Modeling Studies of Novel Coumarin Carboxamide Derivatives as eEF-2K Inhibitors

Abstract

Eukaryotic elongation factor-2 kinase (eEF-2K) is an unusual alpha kinase commonly upregulated in various human cancers, including breast, pancreatic, lung, and brain tumors. We have demonstrated that eEF-2K is relevant to poor prognosis and shorter patient survival in breast and lung cancers and validated it as a molecular target using genetic methods in related in vivo tumor models. Although several eEF-2K inhibitors have been published, none of them have shown to be potent and specific enough for translation into clinical trials. Therefore, development of highly effective novel inhibitors targeting eEF-2K is needed for clinical applications. However, currently, the crystal structure of eEF-2K is not known, limiting the efforts for designing novel inhibitor compounds. Therefore, using homology modeling of eEF-2K, we designed and synthesized novel coumarin-3-carboxamides including compounds A1, A2, and B1–B4 and evaluated their activity by performing in silico analysis and in vitro biological assays in breast cancer cells. The Molecular Mechanics/Generalized Born Surface Area (MM/GBSA) area results showed that A1 and A2 have interaction energies with eEF-2K better than those of B1–B4 compounds. Our in vitro results indicated that compounds A1 and A2 were highly effective in inhibiting eEF-2K at 1.0 and 2.5 μM concentrations compared to compounds B1–B4, supporting the in silico findings. In conclusion, the results of this study suggest that our homology modeling along with in silico analysis may be effectively used to design inhibitors for eEF-2K. Our newly synthesized compounds A1 and A2 may be used as novel eEF-2K inhibitors with potential therapeutic applications.

Target-driven design of a coumarinyl chalcone scaffold based novel EF2 Kinase inhibitor suppresses breast cancer growthin vivo

“…However, the three-dimensional structures of the catalytic domain of another α-kinase called myosin heavy-chain kinase A (MHCKA) is available and has been studied in considerable detail 35 . Therefore, we developed a predicted model of human eEF-2K used as a myosin heavy-chain kinase A (MHCK-A) template with homology modeling studies 36 . Next, we designed and screened different scaffolds of compounds including coumarin-chalcone scaffold and evaluated their activity by in silico using homology modeled target structure of eEF-2K.…”

“…S1). In order to take into account, the flexibility of binding pocket residues of model protein 36 based on MHCK-A when the ligand approaches to the binding site, induced fit docking (IFD) method was considered. IFD results showed that 2C binds similar region identified by rigid docking, however, its binding pose was slightly different, as expected.…”

“…The efforts for developing potent eEF-2K inhibitors have been hindered by the lack of crystal and a 3-dimensional structure for eEF-2K. However, the development of a homology model of eEF-2K based on the known structures of other members of the α-kinase family such as CMAK has marketly increased rationale drug design and of discovering potent and selective eEF-2K inhibitors 36 . In fact, using the homology modeling of eEF-2K using the homology modeling, recently, we identified coumarin-3-carboxamides (A1 and A2) with morpholine and tert-butyl piperazine-1-carboxylate groups attached to the phenyl ring via the methylene bridge as specific eEF-2K inhibitors at 1 and 2.5 µM doses, respectively 36 .…”

“…However, the development of a homology model of eEF-2K based on the known structures of other members of the α-kinase family such as CMAK has marketly increased rationale drug design and of discovering potent and selective eEF-2K inhibitors 36 . In fact, using the homology modeling of eEF-2K using the homology modeling, recently, we identified coumarin-3-carboxamides (A1 and A2) with morpholine and tert-butyl piperazine-1-carboxylate groups attached to the phenyl ring via the methylene bridge as specific eEF-2K inhibitors at 1 and 2.5 µM doses, respectively 36 . Here, we identified as one of the most effective eEF-2K inhibitors (compound 2C ) that about 100-fold more potent and exerts a higher eEF-2K inhibitors activity compared with the known eEF-2K inhibitors (i.e., A-484954, IC 50 for cell proliferation = 75 µM) 31,32 .…”

“…For MD simulations, the Desmond program was used. Similar MD protocol to our previously reported studies was used 36 . Last 50% of the trajectories from the MD simulations were used in free energy calculations.…”