cyclin-d1 and phenethyl-isothiocyanate

Cruciferous vegetable-derived isothiocyanates (ITCs) display potent cancer chemopreventive activity, but also markedly stimulate oncogenic activator protein 1 (AP-1). AP-1 is well known to promote cell survival and proliferation. We examined the impact of AP-1 activation on antiproliferative activity of ITCs, using bladder cancer cells and phenethyl isothiocyanate (PEITC) as models. AP-1 transactivation induced by PEITC was almost completely suppressed by a dominant-negative c-jun (TAM67). However, suppression of AP-1 transactivation did not affect PEITC-induced apoptosis or cell-cycle arrest. Moreover, we previously showed that in response to ITC treatment c-jun was predominantly stimulated among AP-1 family members largely by c-jun N-terminal kinase (JNK) [Food Chem Toxicol 2005; 43: 1373-1380], but neither JNK inhibition nor forced expression of c-jun altered the antiproliferative activity of PEITC. In addition, cyclin D1, which is considered as an AP-1 target gene and promotes cell proliferation, was markedly elevated in PEITC-treated cells. Unexpectedly, neither TAM67 or JNK inhibition, nor forced c-jun expression had a significant impact on cyclin D1 induction by PEITC, indicating that c-jun/AP-1 does not play an important role in cyclin D1 induction by PEITC. In conclusion, despite the known role of c-jun/AP-1 as a stimulator of cell growth and proliferation, our data show that its activation does not diminish the antiproliferative activity of PEITC and is not responsible for cyclin D1 induction by PEITC.

Topics: Anticarcinogenic Agents; Apoptosis; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Humans; Isothiocyanates; JNK Mitogen-Activated Protein Kinases; Peptide Fragments; Proto-Oncogene Proteins c-jun; Replication Protein C; Signal Transduction; Transcription Factor AP-1; Tumor Cells, Cultured; Urinary Bladder Neoplasms

Recent studies indicate that natural isothiocyanates, such as sulforaphane (SFN) and phenethyl isothiocyanate (PEITC) possess strong antitumor activities in vitro and in vivo. The nuclear factor kappa B (NF-kappaB) is believed to play an important role in cancer chemoprevention due to its involvement in tumor cell growth, proliferation, angiogenesis, invasion, apoptosis, and survival. In this study, we investigated the effects and the molecular mechanisms of SFN and PEITC on NF-kappaB transcriptional activation and NF-kappaB-regulated gene expression in human prostate cancer PC-3 C4 cells. Treatment with SFN (20 and 30 microM) and PEITC (5 and 7.5 microM) significantly inhibited NF-kappaB transcriptional activity, nuclear transloction of p65, and gene expression of NF-kappaB-regulated VEGF, cylcin D1, and Bcl-X(L) in PC-3 C4 cells. To further elucidate the mechanism, we utilized the dominant-negative mutant of inhibitor of NF-kappaB alpha (IkappaBalpha) (SR-IkappaBalpha). Analogous to treatments with SFN and PEITC, SR-IkappaBalpha also strongly inhibited NF-kappaB transcriptional activity as well as VEGF, cylcin D1, and Bcl-X(L) expression. Furthermore, SFN and PEITC also inhibited the basal and UVC-induced phosphorylation of IkappaBalpha and blocked UVC-induced IkappaBalpha degradation in PC-3 C4 cells. In examining the upstream signaling, we found that the dominant-negative mutant of IKKbeta (dnIKKbeta) possessed inhibitory effects similar to SFN and PEITC on NF-kappaB, VEGF, cylcin D1, Bcl-X(L) as well as IkappaBalpha phosphorylation. In addition, treatment with SFN and PEITC potently inhibited phosphorylation of both IKKbeta and IKKalpha and significantly inhibited the in vitro phosphorylation of IkappaBalpha mediated by IKKbeta. Taken together, these results suggest that the inhibition of SFN and PEITC on NF-kappaB transcriptional activation as well as NF-kappaB-regulated VEGF, cyclin D1, and Bcl-X(L) gene expression is mainly mediated through the inhibition of IKK phosphorylation, particularly IKKbeta, and the inhibition of IkappaBalpha phosphorylation and degradation, as well as the decrease of nuclear translocation of p65 in PC-3 cells.

Topics: Anticarcinogenic Agents; bcl-X Protein; Cell Line, Tumor; Cell Survival; Cyclin D1; Enzyme Inhibitors; Gene Expression Regulation, Neoplastic; Humans; I-kappa B Kinase; I-kappa B Proteins; Isothiocyanates; Male; NF-kappa B; NF-KappaB Inhibitor alpha; Phosphorylation; Prostatic Neoplasms; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-bcl-2; Sulfoxides; Thiocyanates; Transcription, Genetic; Vascular Endothelial Growth Factor A

Male strain A mice were fed a diet containing chemopreventive agents. After 1 and 3 weeks on the diets, lung nuclear fractions were examined for expression of cyclin D1/2 with western blot analysis. In animals fed a diet containing a mixture of myoinositol and dexamethasone, a treatment found previously to be effective in preventing the development of tobacco smoke-induced lung tumors in A/J mice, cyclin D1/2 expression was reduced to 30-40% of control levels. A similar decrease in cyclin D1/2 expression was found when animals were fed either myoinositol or dexamethasone alone. Paradoxically, tobacco smoke by itself had a similar effect on cyclin D1/2 expression. On the other hand, several agents that had been previously found not to be effective against tobacco smoke carcinogenesis [phenethyl isothiocyanate, 1,4-phenylenebis(methylene)selenoisocyanate, N-acetylcysteine, acetylsalicylic acid, D-limonene and beta carotene] did not decrease cyclin D1/2 expression after 1 or 3 weeks of feeding. It was concluded that expression of cyclin D1/2 might be a potentially useful marker in the identification of chemopreventive agents for tobacco smoke and could be of some help in the evaluation of their effects.

Topics: Acetylcysteine; Animals; Anticarcinogenic Agents; Aspirin; Blotting, Western; Cell Nucleus; Cyclin D1; Cyclin D2; Cyclins; Cyclohexenes; Dexamethasone; Free Radical Scavengers; Immunohistochemistry; Inositol; Isothiocyanates; Limonene; Lung; Lung Neoplasms; Male; Mice; Nicotiana; Organoselenium Compounds; Tea; Terpenes; Time Factors