Connecting Conformational Landscapes with Bioactivity in Macrocyclic Kinase Inhibitors

Abstract

Macrocyclization can improve the selectivity and pharmacokinetic properties of kinase inhibitors, yet the conformational basis of these effects remains incompletely understood. Here, we characterized the structures and conformational ensembles of the clinically relevant macrocyclic kinase inhibitors pacritinib and zotiraciclib using microcrystal electron diffraction (MicroED), NMR spectroscopy, and molecular docking. Both compounds populate a set of related macrocyclic core conformations, with additional conformational adaptability arising from changes in aromatic torsion angles and substituent orientations. These conformational adaptations correlate with physicochemical properties relevant to aqueous solubility and membrane permeability. Pacritinib exhibits chameleonic behavior through environment-dependent modulation of its exposed polar surface area, whereas protonation of zotiraciclib generates distinct conformational states stabilized by intramolecular cation–π interactions. Molecular docking indicates that the experimentally determined conformations are compatible with productive binding across multiple kinase targets. Collectively, these findings establish a link between conformational adaptability, physicochemical properties, and polypharmacology in macrocyclic kinase inhibitors, providing insights that may guide the rational design of future macrocyclic therapeutics.