Isothiocyanate sulforaphane inhibits protooncogenic ornithine decarboxylase activity in colorectal cancer cells via induction of the TGF-β/Smad signaling pathway

TY - JOUR

T1 - Isothiocyanate sulforaphane inhibits protooncogenic ornithine decarboxylase activity in colorectal cancer cells via induction of the TGF-β/Smad signaling pathway

AU - Kaminski, Bettina M

AU - Loitsch, Stefan M

AU - Ochs, Meike J

AU - Reuter, Kerstin C

AU - Steinhilber, Dieter

AU - Stein, Jürgen

AU - Ulrich, Sandra

PY - 2010/10

Y1 - 2010/10

N2 - SCOPE: The objective of this study was to elucidate molecular mechanisms behind the antitumor activities of the isothiocyanate sulforaphane (SFN) in colorectal cancer cells.METHODS AND 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.CONCLUSION: Since elevated ODC enzyme activity is associated with enhanced tumor development, SFN may be a dietary phytochemical with potential to prevent carcinogenesis.

AB - SCOPE: The objective of this study was to elucidate molecular mechanisms behind the antitumor activities of the isothiocyanate sulforaphane (SFN) in colorectal cancer cells.METHODS AND 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.CONCLUSION: Since elevated ODC enzyme activity is associated with enhanced tumor development, SFN may be a dietary phytochemical with potential to prevent carcinogenesis.

KW - Anticarcinogenic Agents/pharmacology

KW - Caco-2 Cells

KW - Cell Proliferation/drug effects

KW - Colorectal Neoplasms/drug therapy

KW - DNA-Binding Proteins/antagonists & inhibitors

KW - Gene Expression Regulation, Neoplastic/drug effects

KW - Humans

KW - Isothiocyanates/pharmacology

KW - Ornithine Decarboxylase/metabolism

KW - Protein Kinase Inhibitors/pharmacology

KW - Protein Serine-Threonine Kinases/antagonists & inhibitors

KW - RNA, Messenger/metabolism

KW - Receptor, Transforming Growth Factor-beta Type I

KW - Receptor, Transforming Growth Factor-beta Type II

KW - Receptors, Transforming Growth Factor beta/antagonists & inhibitors

KW - Signal Transduction/drug effects

KW - Smad Proteins/genetics

KW - Smad3 Protein/genetics

KW - Smad4 Protein/genetics

KW - Sulfoxides

KW - Thiocyanates/pharmacology

KW - Transforming Growth Factor beta/genetics

U2 - 10.1002/mnfr.201000105

DO - 10.1002/mnfr.201000105

M3 - SCORING: Journal article

C2 - 20603835

VL - 54

SP - 1486

EP - 1496

JO - MOL NUTR FOOD RES

JF - MOL NUTR FOOD RES

SN - 1613-4125

IS - 10

ER -