The
anticancer agent 2-cyano-3,12-dioxooleana-1,9-dien-28-oic
acid (
CDDO) and its methyl
ester (
CDDO-Me) typically induce a broad spectrum of growth-inhibitory, proapoptotic, and antiangiogenic responses. Treatment of Panc1, Panc28, and L3.6pL
pancreatic cancer cells with low micromolar concentrations of
CDDO or
CDDO-Me resulted in growth inhibition, induction of apoptosis, and down-regulation of
cyclin D1,
survivin,
vascular endothelial growth factor (
VEGF), and its receptor (VEGFR2). RNA interference studies indicate that these repressed genes are regulated by specificity
protein (
Sp) transcription factors Sp1, Sp3, and Sp4, and Western blot analysis of lysates from
pancreatic cancer cells treated with
CDDO and
CDDO-Me shows for the first time that both compounds decreased the expression of Sp1, Sp3, and Sp4. Moreover,
CDDO-Me (7.5 mg/kg/day) also inhibited pancreatic human L3.6pL
tumor growth and down-regulated Sp1, Sp3, and Sp4 in
tumors using an orthotopic
pancreatic cancer model.
CDDO-Me also induced
reactive oxygen species (ROS) and decreased mitochondrial membrane potential (
MMP) in Panc1 and L3.6pL cells, and cotreatment with
antioxidants (
glutathione and
dithiothreitol) blocked the formation of ROS, reversed the loss of
MMP, and inhibited down-regulation of Sp1, Sp3, and Sp4. Repression of Sp and Sp-dependent genes by
CDDO-Me was due to the down-regulation of
microRNA-27a and induction of zinc finger and BTB domain containing 10 (ZBTB10), an Sp repressor, and these responses were also reversed by
antioxidants. Thus, the anticancer activity of
CDDO-Me is due, in part, to activation of ROS, which in turn targets the
microRNA-27a:ZBTB10-Sp
transcription factor axis. This results in decreased expression of Sp-regulated genes, growth inhibition, induction of apoptosis, and antiangiogenic responses.