A soybean Kunitz trypsin inhibitor suppresses ovarian cancer cell invasion by blocking urokinase upregulation.
We have previously reported in a series of papers that a Kunitz-type protease inhibitor, bikunin, suppresses up-regulation of urokinase-type plasminogen activator (uPA) and its specific receptor (uPAR) expression, phosphorylation of ERK1/2 and cancer cell invasion in vitro and peritoneal disseminated metastasis in vivo. In the present study, we investigated the effects of soy bean trypsin inhibitor (SBTI) on the net enzymatic activity of secreted, extracellular uPA, signal transduction involved in the expression of uPA and invasion in HRA human ovarian cancer cells. SBTI contains a Kunitz trypsin inhibitor (KTI) and a Bowman-Birk inhibitor (BBI). Here, we show 1) that KTI and BBI were purified separately from soybeans; 2) that neither KTI nor BBI effectively inhibits enzymatic activity of uPA; 3) that uPA upregulation observed in HRA cells was inhibited by preincubation of the cells with KTI with an IC50 of approximately 2 microM, whereas BBI failed to repress uPA upregulation, as measured by enzyme-linked immunosorbent assay; 4) that cell invasiveness was inhibited by treatment of the cells with KTI with an IC50 of approximately 3 microM, whereas BBI failed to suppress cell invasion, as measured by an in vitro invasion assay; 5) KTI suppresses HRA cell invasion by blocking uPA up-regulation which may be mediated by a binding protein(s) other than a bikunin binding protein and/or its receptor; and 6) that transforming growth factor-beta 1 (TGF-beta1)-mediated activation of ERK1/2 was significantly reduced by preincubation of the cells with KTI. In conclusion, KTI, but not BBI, could inhibit cell invasiveness at least through suppression of uPA signaling cascade, although the mechanisms of KTI may be different from those of bikunin.
Kobayashi
Hiroshi
H
Department of Obstetrics and Gynecology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan. hirokoba@hama-med.ac.jp
Suzuki
Mika
M
Kanayama
Naohiro
N
Terao
Toshihiko
T
D001667:Binding, Competitive; D045744:Cell Line, Tumor / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D003094:Collagen; D003864:Depression, Chemical; D004338:Drug Combinations; D004790:Enzyme Induction / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005419:Flavonoids / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D007797:Laminin; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008562:Membrane Glycoproteins / Q000378:metabolism; D009361:Neoplasm Invasiveness; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology; D010051:Ovarian Neoplasms / Q000473:pathology*; D010766:Phosphorylation / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011509:Proteoglycans; D013329:Structure-Activity Relationship; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D053773:Transforming Growth Factor beta1; D014358:Trypsin Inhibitor, Bowman-Birk Soybean / Q000494:pharmacology; D014360:Trypsin Inhibitor, Kunitz Soybean / Q000378:metabolism / Q000494:pharmacology*; D014361:Trypsin Inhibitors / Q000494:pharmacology*; D014568:Urokinase-Type Plasminogen Activator / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology
D004338:Drug Combinations; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007797:Laminin; D008562:Membrane Glycoproteins; D009363:Neoplasm Proteins; D011509:Proteoglycans; C109148:SPINT2 protein, human; C508960:TGFB1 protein, human; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D014358:Trypsin Inhibitor, Bowman-Birk Soybean; D014361:Trypsin Inhibitors; C059485:matrigel; D003094:Collagen; D014360:Trypsin Inhibitor, Kunitz Soybean; D019833:Genistein; D014568:Urokinase-Type Plasminogen Activator; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one