Recent studies in cell culture have shown that
isothiocyanates (ITCs) induce apoptosis via activation of
mitogen-activated
protein (MAP)
kinases and p53 pathways, suggesting a potential for ITCs or their conjugates to inhibit
tumorigenesis during the postinitiation phase. To evaluate whether ITC compounds administered after
carcinogen treatment inhibit lung
tumorigenesis, we investigated in A/J mice the effects of the
N-acetylcysteine (NAC) conjugates of benzyl (BITC-NAC) and phenethyl ITC (
PEITC-NAC) in the diet (15 micromol/g) administered after a single dose of 20 micromol
benzo(a)pyrene [B(a)P]. The formation of lung
adenomas was examined 140 days after B(a)P dosing. Both the BITC-NAC and
PEITC-NAC-treated groups showed a significant reduction in lung
tumor multiplicity from 6.1 +/- 3.1
tumors/mouse in the B(a)P group fed the control diet to 3.7 +/- 2.9 and 3.4 +/- 2.7
tumors/mouse (P = 0.018 and 0.006, respectively). To investigate the mechanisms of
tumor inhibition, lung tissues were obtained at 21, 84, and 140 days at interim sacrifices during the bioassay. These tissues showed a significant increase in apoptosis as determined by in situ end-labeling for both ITC-NAC-treated groups. The MAP
kinase pathway was activated in the ITC-NAC-treated groups. The activation of c-Jun NH(2)-terminal
kinase was higher in the BITC-NAC and
PEITC-NAC groups when compared with B(a)P-treated control. The phosphorylation of p38 and
extracellular signal-regulated kinases (ErKs) 1 and 2 was also induced by these treatments. To determine the downstream target of MAP
kinases,
activator protein-1 (AP-1) and
nuclear factor-kappaB activities were evaluated by gel shift assay. The
AP-1 binding activity was remarkably increased in lung tissue from both the BITC-NAC and
PEITC-NAC groups. No change in
nuclear factor-kappaB binding activity was found, however. Phosphorylation of p53 was also higher than the constitutive levels in both ITC-NAC-treated groups, but no induction of p53 expression was detected. This study demonstrates the chemopreventive efficacy of the NAC conjugates of
PEITC and BITC administered in the diet after a single dose of B(a)P for lung
tumorigenesis and provides the first in vivo evidence that activation of MAP
kinases,
AP-1 transcription factors, p53 phosphorylation, and the induction of apoptosis may be involved in the chemopreventive activity of these compounds.