In vitro activity: JW 55 is a potent and selective small molecule inhibitor of β-catenin signaling pathway, it functions via inhibition of the PARP domain of tankyrase 1 and tankyrase 2 (TNKS1/2) which are regulators of the β-catenin destruction complex. JW 55 decreases auto-PARsylation of TNKS1/2 in vitro with IC50s of 1.9 μM and 830 nM respectively. Inhibition of TNKS1/2 poly(ADP-ribosyl)ation activity by JW55 led to stabilization of AXIN2, a member of the β-catenin destruction complex, followed by increased degradation of β-catenin. JW55 inhibited canonical Wnt signaling in a dose-dependent manner in colon carcinoma cells that contained mutations in either the APC (adenomatous polyposis coli) locus or in an allele of β-catenin. In addition, JW55 reduced XWnt8-induced axis duplication in Xenopus embryos and tamoxifen-induced polyposis formation in conditional APC mutant mice.

Kinase Assay: JW 55 (JW55) is a potent and selective inhibitor of the canonical Wnt pathway. Wnt3a-induced HEK293 cells containing a transiently transfected ST-Luc (SuperTop-luciferase) reporter show inhibition by JW55 with an IC50 value of 470 nM. JW55 is effective in the range of 1 to 5 μM in SW480 cells and 0.01 to 5 μM in HCT-15 cells. JW55 is effective in the range of 1 to 5 μM in SW480 cells and 0.01 to 5 μM in HCT-15 cells

Cell Assay: A total of 1,000 SW480 or RKO cells are seeded in 96-well plates. The day after, the cell culture medium is exchanged to solutions that contained 0.1% DMSO or 10 μM JW55 for RKO cells and 0.1% DMSO or 10, 5, or 1 μM JW55 for SW480 cells. All samples consist of a minimum of 6 replicates. The plate is incubated in an IncuCyte inside a cell culture incubator. Images are captured every second hour to monitor proliferation.