Abstract
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.
Footnotes
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.109.151415.
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ABBREVIATIONS: CK1, casein kinase 1; PF-4800567, 3-(3-chloro-phenoxymethyl)-1-(tetrahydro-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-ylamine; PF-670462, 4-[3-cyclohexyl-5-(4-fluoro-phenyl)-3H-imidazol-4-yl]-pyrimidin-2-ylamine; GFP, green fluorescent protein; PBS, phosphate-buffered saline; D4476, 4-[4-(2,3-dihydro-benzo[1,4]dioxin-6-yl)-5-pyridin-2-yl-1H-imidazol-2-yl] benzamide; TCEP, tris(2-carboxyethyl)phosphine; DMSO, dimethyl sulfoxide.
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↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
- Received January 24, 2009.
- Accepted May 18, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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