FIG 2.

The novel Tnks inhibitor chemotype IWR-8 directly inhibits Tnks enzymes and disrupts β-catenin-dependent transcriptional activity induced by loss of APC function. (A) IWR-3, -6, and -8 directly target Tnks. Representative IWRs from each chemotype were evaluated for the ability to inhibit purified recombinant Tnks1 protein (Tnks1 SAM-PARP protein). Parsylation of immobilized histone protein was determined by colorimetric detection of incorporated biotinylated NAD⁺ in a 96-well format. (B) IWR-8 inhibits β-catenin activity induced by loss of APC function. The 50% effective concentrations (EC₅₀s) of IWR-6, as well as IWR-1, IWR-3, and IWR-8, were measured in DLD-1 cells using a Wnt/β-catenin-specific luciferase reporter. (C) IWR-8 inhibits cell growth in a cancer cell line with compromised APC function. SW403 cells previously shown to exhibit Wnt/β-catenin pathway-dependent cell growth in culture (58) were treated with either IWR-3, IWR-8, or IWP-2 (an inhibitor of the Wnt acyltransferase Porcupine [16]; a negative control).