Addressing a Trapped High-Energy Water: Design and Synthesis of Highly Potent Pyrimidoindole-Based Glycogen Synthase Kinase-3β Inhibitors
Stanislav Andreev, Tatu Pantsar, Roberta Tesch, Niclas Kahlke, Ahmed El‐Gokha, Francesco Ansideri, Lukas Grätz, Jenny Romasco, Giulia Sita, Christian Geibel, Michael Lämmerhofer, Andrea Tarozzi, Stefan Knapp, Stefan Laufer, Pierre Koch
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 IC50 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, IC50 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.