Addressing a Trapped High-Energy Water: Design and Synthesis of Highly Potent Pyrimidoindole-Based Glycogen Synthase Kinase-3β Inhibitors

Abstract

In small molecule binding, water is not a passive bystander but rather takes an active role in the binding site, which may be decisive for the potency of the inhibitor. Here, by addressing a high-energy water, we improved the IC 50 value of our co-crystallized glycogen synthase kinase-3β (GSK-3β) inhibitor by nearly two orders of magnitude. Surprisingly, our results demonstrate that this high-energy water was not displaced by our potent inhibitor (S)-3-(3-((7-ethynyl-9H-pyrimido[4,5-b]indol-4-yl)(methyl)amino)piperidin-1-yl)propanenitrile ((S)-15, IC 50 value of 6 nM). Instead, only a subtle shift in the location of this water molecule resulted in a dramatic decrease in the energy of this high-energy hydration site, as shown by the WaterMap analysis combined with microsecond timescale molecular dynamics simulations. (S)-15 demonstrated both a favorable kinome selectivity profile and target engagement in a cellular environment and reduced GSK-3 autophosphorylation in neuronal SH-SY5Y cells. Overall, our findings highlight that even a slight adjustment in the location of a high-energy water can be decisive for ligand binding.

GSK-3β, FYN, and DYRK1A: Master Regulators in Neurodegenerative Pathways

“…Over the last two decades, the increased interest in GSK-3β led to the discovery of many inhibitors based on chemically different molecular scaffolds and acting with diverse mechanisms of action namely ATP and non-ATP competition, and allosteric modulation [98]. Most inhibitors reported in the literature are ATP competitive agents; some of them have synthetic origin, whereas others have been derived directly or indirectly from small molecules of natural origin (e.g., paullones, maleimides, indirubins, arylindolemaleimides, thiazoles).…”

Discovery of novel α‐carboline derivatives as glycogen synthase kinase‐3β inhibitors for the treatment of Alzheimer's disease

“…Molecular docking was performed to analyze the binding modes of the α‐carboline structure in the ATP binding pocket of GSK‐3β (PDB code: 4PTC and 7B6F, respectively). [ 32,33 ] As shown in Figure 2, there may be two different binding modes between the α‐carboline scaffold and GSK‐3β according to the formation of the classic acceptor–donor–acceptor motif between the azaindole fragment of the carboline structure and the kinase hinge region. Obviously, the pyridine N atom of the α‐carboline structure as a hydrogen‐bond acceptor can interact with Val135 in these two potential binding modes.…”

Frontiers in Medicinal Chemistry 2022 Goes Virtual

“…Investigation of a key compound‘s binding mode by X‐ray crystallography in combination with WaterMap analysis revealed a high energy water molecule trapped behind the tricyclic hinge‐binding motif. Introduction of an ethinyl moiety to reposition this water gave a boost in potency resulting in a single digit nanomolar inhibitor with a clean kinome profile [12] …”

“…Introduction of an ethinyl moiety to reposition this water gave a boost in potency resulting in a single digit nanomolar inhibitor with a clean kinome profile. [12] The next talk was given by Sebastian Pomplun (Leiden University), who recently received an ERC starting grant ("SynTra") to support his work on synthetic transcription factors. He showed how transcriptional activity of the undruggable proto-oncogene MYC could be regulated by protein engineering.…”