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Small-molecule inhibition of Wnt signaling through activation of casein kinase 1α

Abstract

Wnt/β-catenin signaling is critically involved in metazoan development, stem cell maintenance and human disease. Using Xenopus laevis egg extract to screen for compounds that both stabilize Axin and promote β-catenin turnover, we identified an FDA-approved drug, pyrvinium, as a potent inhibitor of Wnt signaling (EC50 of 10 nM). We show pyrvinium binds all casein kinase 1 (CK1) family members in vitro at low nanomolar concentrations and pyrvinium selectively potentiates casein kinase 1α (CK1α) kinase activity. CK1α knockdown abrogates the effects of pyrvinium on the Wnt pathway. In addition to its effects on Axin and β-catenin levels, pyrvinium promotes degradation of Pygopus, a Wnt transcriptional component. Pyrvinium treatment of colon cancer cells with mutation of the gene for adenomatous polyposis coli (APC) or β-catenin inhibits both Wnt signaling and proliferation. Our findings reveal allosteric activation of CK1α as an effective mechanism to inhibit Wnt signaling and highlight a new strategy for targeted therapeutics directed against the Wnt pathway.

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Figure 1: Xenopus egg extract screen identifies pyrvinium as an inhibitor of Wnt signaling.
Figure 2: Pyrvinium inhibits Wnt signaling in vivo.
Figure 3: CK1α is the critical target of pyrvinium.
Figure 4: Pyrvinium promotes Pygopus degradation.
Figure 5: Pyrvinium selectively decreases cell viability of colon cancer cells with activating mutations in the Wnt pathway.

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Acknowledgements

We thank members of the Lee laboratory, K. Gould, V. Siegel and R. Roberts-Galbraith for critical reading of the manuscript; J. Merkle for help with Drosophila S2 cell culture; members of the Institute of Chemistry and Cell Biology–Longwood for assistance with screening; M. Beinz, K. Basler, J. Nathans, R. Nusse, S. Hiebert, R. Coffey, B. Gumbiner and M. Lenardo for reagents. Nematode strains were obtained from the C. elegans Genetics Center, which is supported by the US National Institutes of Health National Center for Research Resources. This work was supported by American Cancer Society Research Scholar Grant RSG-05-126-01, American Cancer Society Institutional Research Grant IRG-58-009-46, National Cancer Institute Grant GI SPORE P50 CA95103, Mouse Models of Human Cancers Consortium (US National Institutes of Health–National Cancer Institute) 5U01 CA084239, US National Institutes of Health Grant 1 R01 GM081635-01 (E.L.); US National Institutes of Health Grant 1 R01 NS26115 (D.M.M.); American Heart Association Predoctoral Fellowship 0615279B, Molecular Endocrinology Training Grant 5 T 32 DK007563, Training Program in Developmental Biology 5 T32 HD007502 (National Institute of Child Health and Human Development) (C.A.T.); US National Institute of General Medical Studies Medical-Scientist Training Grant 5 T32 GM007347 (C.S.C. and A.J.H.); American Heart Association Predoctoral Fellowships 0615162B, US National Institutes of Health Cancer Biology Training Grant T32 CA09592 (K.K.J.). E.L. is a recipient of a Pew Scholarship in the Biomedical Sciences.

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C.A.T. designed, performed and analyzed biochemical, extract and cell culture experiments. C.A.T. and E.T. performed screen under E.L. and A.S.'s guidance. J.S. designed and performed C. elegans experiments under D.M.M.'s guidance. E.T. and A.J.H. performed Xenopus embryo experiments. B.L. provided bioinformatics support. D.O., A.G.W., K.K., B.M., V.P.G. and G.A.S. designed and performed chemical synthesis. C.S.C., K.K.J., A.J.H., B.I.H. and L.A.L. provided essential reagents and discussions. K.C.M. provided technical assistance. C.A.T., E.L. and L.A.L. wrote the manuscript with advice from all authors. E.L. guided all aspects of study.

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Correspondence to Ethan Lee.

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Competing interests

Ethan Lee is cofounder of StemSynergy Therapeutics Inc., a company that aims to develop inhibitors of major signaling pathways (including the Wnt pathway) as potential chemotherapeutic agents. Darren Orton is an employee of StemSynergy Therapeutics Inc.

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Thorne, C., Hanson, A., Schneider, J. et al. Small-molecule inhibition of Wnt signaling through activation of casein kinase 1α. Nat Chem Biol 6, 829–836 (2010). https://doi.org/10.1038/nchembio.453

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