Longdaysin inhibits Wnt/β-catenin signaling and exhibits antitumor activity against breast cancer

BACKGROUND: CK1 is involved in regulating Wnt/β-catenin signaling and represents a promising target for the treatment of breast cancer. A purine derivative longdaysin has recently been identified as a novel modulator of cellular circadian rhythms through targeting the protein kinases CK1δ, CK1α, and ERK2. However, the antitumor activity of longdaysin and its underlying mechanisms remain unclear. METHODS: The inhibitory effect of longdaysin on Wnt/β-catenin signaling was investigated using the SuperTOPFlash reporter system. The levels of phosphorylated LRP6, total LRP6, DVL2, active β-catenin, and total β-catenin were examined by Western blot. The expression of Wnt target genes was determined using real-time PCR. The ability of colony formation of breast cancer cells was measured by colony formation assay. The effects of longdaysin on cancer cell migration and invasion were assessed using transwell assays. The effect of longdaysin on cancer stem cells was tested by sphere formation assay. The in vivo antitumor effect of longdaysin was evaluated using MDA-MB-231 breast cancer xenografts. RESULTS: Longdaysin suppressed Wnt/β-catenin signaling through inhibition of CK1δ and CK1ε in HEK293T cells. In breast cancer Hs578T and MDA-MB-231 cells, micromolar concentrations of longdaysin attenuated the phosphorylation of LRP6 and DVL2 and reduced the expression of active β-catenin and total β-catenin, leading to the downregulation of Wnt target genes Axin2, DKK1, LEF1, and Survivin. Furthermore, longdaysin inhibited the colony formation, migration, invasion, and sphere formation of breast cancer cells. In MDA-MB-231 breast cancer xenografts, treatment with longdaysin suppressed tumor growth in association with inhibition of Wnt/β-catenin signaling. CONCLUSION: Longdaysin is a novel inhibitor of the Wnt/β-catenin signaling pathway. It exerts antitumor effect through blocking CK1δ/ε-dependent Wnt signaling.