Isothiocyanate sulforaphane inhibits protooncogenic ornithine decarboxylase activity in colorectal cancer cells via induction of the TGF-β/Smad signaling pathway.
METHODS
The objective of this study was to elucidate molecular mechanisms behind the antitumor activities of the isothiocyanate sulforaphane (SFN) in colorectal cancer cells.


RESULTS
Cell growth was determined by BrdU incorporation and crystal violet staining. Protein levels were examined by Western blot analysis. Ornithine decarboxylase (ODC) activity was assayed radiometrically. Reverse transcriptase-PCR was used for measuring mRNA expression. For reporter gene assays plasmids were transfected into cells via lipofection and luciferase activity was measured luminometrically. Acetyl-histone H3 and H4 chromatin immunoprecipitation (ChIP) assays were performed followed by PCR with TGF-β-receptor II promoter specific primers. We could show that SFN-mediated cell growth inhibition closely correlates with a dose-dependent reduction of protein expression and enzymatic activity of ODC. This effect seems to be due to reduced protein levels and transactivation activity of transcription factor c-myc, a direct regulator of ODC expression, as a consequence of SFN-induced TGF-β/Smad signaling. The coherency of these results was further confirmed by using TGF-β receptor kinase inhibitor SB431542, which largely abolishes inhibitory effects of SFN on both, ODC activity and cell growth.


CONCLUSIONS
Since elevated ODC enzyme activity is associated with enhanced tumor development, SFN may be a dietary phytochemical with potential to prevent carcinogenesis.
Kaminski
Bettina M
BM
Institute of Pharmaceutical Chemistry, Biozentrum, Goethe University, Frankfurt am Main, Germany.
Loitsch
Stefan M
SM
Ochs
Meike J
MJ
Reuter
Kerstin C
KC
Steinhilber
Dieter
D
Stein
Jürgen
J
Ulrich
Sandra
S
D016588:Anticarcinogenic Agents / Q000494:pharmacology; D018938:Caco-2 Cells; D049109:Cell Proliferation / Q000187:drug effects; D015179:Colorectal Neoplasms / Q000188:drug therapy*; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D017879:Isothiocyanates / Q000494:pharmacology*; D009955:Ornithine Decarboxylase / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D000077293:Receptor, Transforming Growth Factor-beta Type I; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D051785:Smad Proteins / Q000235:genetics / Q000378:metabolism*; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism; D051901:Smad4 Protein / Q000235:genetics / Q000378:metabolism; D013454:Sulfoxides; D013861:Thiocyanates / Q000494:pharmacology*; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism*
D016588:Anticarcinogenic Agents; D004268:DNA-Binding Proteins; D017879:Isothiocyanates; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D018125:Receptors, Transforming Growth Factor beta; C497790:SMAD3 protein, human; C497796:SMAD4 protein, human; D051785:Smad Proteins; D051900:Smad3 Protein; D051901:Smad4 Protein; D013454:Sulfoxides; D013861:Thiocyanates; D016212:Transforming Growth Factor beta; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; D000077294:Receptor, Transforming Growth Factor-beta Type II; D009955:Ornithine Decarboxylase; C016766:sulforaphane