13 can inhibit WNT/β-catenin
signaling activity and act as an antiproliferative agent in COLO 320DM
cells. (a) Representative immunoblots of cytoplasmic TNKS1/2, AXIN1,
AXIN2 and cytoplasmic and nuclear transcriptionally active β-catenin (non-phospho) and β-catenin. Actin or lamin
B1 document equals protein loading, and ∗ indicates that the
same actin immunoblot is used as loading control for both AXIN2 and
β-catenin. For (a) and (b), control = 0.01%
DMSO. (b) Real-time RT-qPCR analyses of WNT/β-catenin signaling target genes (AXIN2, DKK1, NKD1, and APCDD1). For (b) and
(c), ANOVA tests (Holm–Sidak method, versus control) are indicated
by ∗∗∗ (P < 0.001), and ANOVA
on ranks tests (Tukeys test versus control) are indicated by †
(P < 0.05). Mean values ± standard deviations
for combined data from a minimum of three independent experiments
with three replicates each are shown. (c) MTS colorimetric cell growth
assay for various doses of 13 in APCmutated COLO 320DM
(black) and APCwild-type RKO (gray) cells. Control
= 0. 1% DMSO. Dotted lines depict 50% (GI50) and 25% (GI25) growth inhibition levels. Mean values ± standard deviations
for one representative experiment out of three independent experiments
are shown.