Exisulind and its analogues are inhibitors of
cyclic GMP phosphodiesterases (
PDEs) that have been shown to activate and induce
protein kinase G, resulting in the induction of apoptosis in
colon cancer cells. These drugs also reduce
beta-catenin protein levels and decrease
cyclin D1 mRNA levels in SW480 cells. Herein we report on studies pertaining to
exisulind regulation of
beta-catenin levels and activity in colon
tumor cells.
Exisulind and its higher-affinity PDE analogues, (Z)-5-fluoro-2-methyl-(4-pyridylidene)-3-(N-benzyl)-indenylacetamide hydrochloride (CP461) and (Z)-1H-indene-3-acetamide, 5-fluoro-2-methyl-N-(phenylmethyl)-1-[(3,4,5-trimethoxyphenyl)methylene] (
CP248), reduced
beta-catenin, including the nuclear
beta-catenin in SW480 cells (EC(50) approximately 200 microM, 1 microM, and <1 microM, respectively). The 50% reduction of
beta-catenin was seen in 8-14 hr. There was no change in
beta-catenin mRNA.
Exisulind-induced
beta-catenin reduction was blocked by the proteasomal inhibitor
MG132 (Z-
leu-Leu-Leu-CHO), indicating that the effect of
exisulind involved
ubiquitin-proteasomal degradation. A consequence of reduced
beta-catenin in SW480 cells was that
exisulind, CP461, and
CP248 caused a concentration- and time-dependent decrease in
cyclin D1 levels (EC(50) approximately 300 microM, 1 microM, and <1 microM, respectively) in 4 hr. The effect was via decreased
cyclin D1 mRNA levels.
Exisulind-induced degradation of
beta-catenin was not blocked by the inhibition of
caspase-3 activity and/or apoptosis, and some SW480 cells showed a reduction in
beta-catenin levels before the appearance of early apoptosis indicators. Expression of the N-terminal 170
amino acid fragment of
beta-catenin reduced the effects of
beta-catenin degradation,
cyclin D1 reduction, and the apoptosis response to
exisulind. These results indicate that
exisulind-induced
beta-catenin degradation precedes the induction of apoptosis and that the down-regulation of inappropriate
beta-catenin-activated genes accounts in part for the pro-apoptotic effects of
exisulind and CP461 in colon
tumor cells.