MEK4 function, genistein treatment, and invasion of human prostate cancer cells.
BACKGROUND
Dietary intake of genistein by patients with prostate cancer has been associated with decreased metastasis and mortality. Genistein blocks activation of p38 mitogen-activated protein kinase and thus inhibits matrix metalloproteinase-2 (MMP-2) expression and cell invasion in cultured cells and inhibits metastasis of human prostate cancer cells in mice. We investigated the target for genistein in prostate cancer cells.


METHODS
Prostate cell lines PC3-M, PC3, 1532NPTX, 1542NPTX, 1532CPTX, and 1542CPTX were used. All cell lines were transiently transfected with a constitutively active mitogen-activated protein kinase kinase 4 (MEK4) expression vector (to increase MEK4 expression), small interfering RNA against MEK4 (to decrease MEK4 expression), or corresponding control constructs. Cell invasion was assessed by a Boyden chamber assay. Gene expression was assessed by a quantitative reverse transcription-polymerase chain reaction. Protein expression was assessed by Western blot analysis. Modeller and AutoDock programs were used for modeling of the structure of MEK4 protein and ligand docking, respectively. MMP-2 transcript levels were assessed in normal prostate epithelial cells from 24 patients with prostate cancer from a phase II randomized trial comparing genistein treatment with no treatment. Statistical significance required a P value of .050 or less. All statistical tests were two-sided.


RESULTS
Overexpression of MEK4 increased MMP-2 expression and cell invasion in all six cell lines. Decreased MEK4 expression had the opposite effects. Modeling showed that genistein bound to the active site of MEK4. Genistein inhibited MEK4 kinase activity with a half maximal inhibitory concentration of 0.40 microM (95% confidence interval [CI] = 0.36 to 0.45 muM). The MMP-2 transcript level in normal prostate epithelial cells was statistically significantly higher in the untreated group (100%) than in the genistein-treated group (24%; difference = 76%, 95% CI = 38% to 115%; P = .045).


CONCLUSIONS
We identified MEK4 as a proinvasion protein in six human prostate cancer cell lines and the target for genistein. We showed, to our knowledge for the first time, that genistein treatment, compared with no treatment, was associated with decreased levels of MMP-2 transcripts in normal prostate cells from prostate cancer-containing tissue.
Xu
Li
L
Department of Medicine, Robert H. Lurie Cancer Center and Center for Drug Discovery and Chemical Biology of Northwestern University, Chicago, IL60610, USA.
Ding
Yongzeng
Y
Catalona
William J
WJ
Yang
Ximing J
XJ
Anderson
Wayne F
WF
Jovanovic
Borko
B
Wellman
Kenji
K
Killmer
Jaqueline
J
Huang
Xiaoke
X
Scheidt
Karl A
KA
Montgomery
R Bruce
RB
Bergan
Raymond C
RC
D000230:Adenocarcinoma / Q000188:drug therapy* / Q000201:enzymology* / Q000473:pathology; D000368:Aged; D000970:Antineoplastic Agents / Q000097:blood / Q000494:pharmacology* / Q000627:therapeutic use; D015153:Blotting, Western; D045744:Cell Line, Tumor; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D019833:Genistein / Q000097:blood / Q000494:pharmacology* / Q000627:therapeutic use; D006801:Humans; D048670:MAP Kinase Kinase 4 / Q000235:genetics / Q000378:metabolism*; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000187:drug effects / Q000378:metabolism; D008875:Middle Aged; D009361:Neoplasm Invasiveness / Q000517:prevention & control; D009367:Neoplasm Staging; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000201:enzymology* / Q000473:pathology; D047428:Protein Kinase Inhibitors / Q000097:blood / Q000494:pharmacology* / Q000627:therapeutic use; D034741:RNA, Small Interfering / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015854:Up-Regulation
D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D019833:Genistein; D048670:MAP Kinase Kinase 4; C482559:MAP2K4 protein, human; D020778:Matrix Metalloproteinase 2