We reported previously a significant increase in survival of nude rats harboring orthotopic A549 human
non-small cell lung cancer tumors after treatment with a combination of
exisulind (
Sulindac Sulfone) and
docetaxel (D. C. Chan, Clin.
Cancer Res., 8: 904-912, 2002). The purpose of the current study was to determine the biochemical mechanisms responsible for the increased survival by an analysis of the effects of both drugs on A549 orthotopic lung
tumors and A549 cells in culture. Orthotopic A549 rat lung tissue sections from
drug-treated rats and A549 cell culture responses to
exisulind and
docetaxel were compared using multiple apoptosis and proliferation analyses [i.e.,
terminal deoxynucleotidyl transferase-mediated nick end labeling, active
caspase 3, the
caspase cleavage products
cytokeratin 18 and p85
poly(ADP-ribose) polymerase, and Ki-67]. Immunohistochemistry was used to determine
cyclic GMP (cGMP)
phosphodiesterase (PDE) expression in
tumors. The cGMP PDE composition of cultured A549 cells was resolved by
DEAE-Trisacryl M chromatography and the pharmacological sensitivity to
exisulind, and additional known PDE inhibitors were determined by
enzyme activity assays.
Exisulind inhibited A549 cell cGMP hydrolysis and induced apoptosis of A549 cells grown in culture. PDE5 and 1 cGMP PDE gene family
isoforms identified in cultured cells were highly expressed in orthotopic
tumors. The in vivo apoptosis rates within the orthotopic
tumors increased 7-8-fold in animals treated with the combination of
exisulind and
docetaxel.
Exisulind increased the in vivo apoptosis rates as a single agent.
Docetaxel, but not
exisulind, decreased proliferative rates within the
tumors. The data indicate that
exisulind-induced apoptosis contributed significantly to the increased survival in rats treated with
exisulind/
docetaxel. The mechanism of
exisulind-induced apoptosis involves inhibition of cGMP
PDEs, and these results are consistent with a cGMP-regulated apoptosis pathway.