Selective Inhibition of Casein Kinase 1ϵ Minimally Alters Circadian Clock Period

The circadian clock links our daily cycles of sleep and activity to the external environment. Deregulation of the clock is implicated in a number of human disorders, including depression, seasonal affective disorder, and metabolic disorders. Casein kinase 1 epsilon (CK1ϵ) and casein kinase 1 delta (CK1δ) are closely related Ser-Thr protein kinases that serve as key clock regulators as demonstrated by mammalian mutations in each that dramatically alter the circadian period. Therefore, inhibitors of CK1δ/ϵ may have utility in treating circadian disorders. Although we previously demonstrated that a pan-CK1δ/ϵ inhibitor, 4-[3-cyclohexyl-5-(4-fluoro-phenyl)-3H-imidazol-4-yl]-pyrimidin-2-ylamine (PF-670462), causes a significant phase delay in animal models of circadian rhythm, it remains unclear whether one of the kinases has a predominant role in regulating the circadian clock. To test this, we have characterized 3-(3-chloro-phenoxymethyl)-1-(tetrahydro-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-ylamine (PF-4800567), a novel and potent inhibitor of CK1ϵ (IC₅₀ = 32 nM) with greater than 20-fold selectivity over CK1δ. PF-4800567 completely blocks CK1ϵ-mediated PER3 nuclear localization and PER2 degradation. In cycling Rat1 fibroblasts and a mouse model of circadian rhythm, however, PF-4800567 has only a minimal effect on the circadian clock at concentrations substantially over its CK1ϵ IC₅₀. This is in contrast to the pan-CK1δ/ϵ inhibitor PF-670462 that robustly alters the circadian clock under similar conditions. These data indicate that CK1ϵ is not the predominant mediator of circadian timing relative to CK1δ. PF-4800567 should prove useful in probing unique roles between these two kinases in multiple signaling pathways.