Identification and Utilization of a Chemical Probe to Interrogate the Roles of PIKfyve in the Lifecycle of β-Coronaviruses

From a designed library of indolyl pyrimidinamines we identified a highly potent and cell-active chemical probe (analog 17) that inhibits phosphatidylinositol-3-phosphate 5-kinase (PIKfyve). Comprehensive evaluation of inhibitor selectivity confirmed that this PIKfyve probe demonstrates excellent kinome-wide selectivity. A structurally related indolyl pyrimidinamine (analog 30) was characterized as a suitable negative control analog that lacks PIKfyve inhibitory activity and exhibits exquisite selectivity when profiled against the screenable human kinome. Our chemical probe disrupts multiple phases of the life cycle of β-coronaviruses. We observed potent inhibition of viral replication, reduced viral entry, and impacts on a mediator of viral transmission (lysosomes). Our scaffold is a distinct chemotype versus published PIKfyve inhibitors and lacks the canonical morpholine hinge-binder of classical lipid kinase inhibitors. Our chemical probe set can be used by the community to characterize the role of PIKfyve in virology and beyond.