Suppression of invasion and peritoneal carcinomatosis of ovarian cancer cell line by overexpression of bikunin.
Bikunin (bik), a Kunitz-type protease inhibitor, also known as urinary trypsin inhibitor, is proposed as a main participant in the inhibition of tumor cell invasion and metastasis, possibly through the direct inhibition of cell-associated plasmin activity and suppression of urokinase-type plasminogen activator (uPA) mRNA expression. In the present study, we transfected the human ovarian carcinoma cell line HRA, highly invasive cells, with an expression vector harboring a cDNA encoding for human bik. Our study was designed to investigate the effect of bik overexpression and changes in tumor cell phenotype and invasiveness in the stably transfected clones. Bik gene transfection of HRA gave the following results: 1) transfection of HRA with the bik cDNA resulted in 5 variants stably expressing functional bik; 2) bik(+) clones exhibited a significantly reduced uPA mRNA expression as compared to the parental cells; 3) bikunin negatively regulates the ERK1/2 activity; 4) secretion-blocking treatments of bik(+) clones abrogated bik-mediated suppression of ERK1/2 activation and uPA expression; 5) the regulation of invasion seen in the HRA cells is mainly mediated by the uPA-plasmin-MMP-2 system; 6) transfection of HRA with the bik gene significantly reduced invasion, but not proliferation, adhesion, or migration relative to the parental cells; and 7) animals with bik(+) clones induced reduced peritoneal dissemination and long term survival. We conclude that transfection of HRA cells with the bik cDNA constitutively suppresses ERK1/2 activation, which results in inhibition of uPA expression and subsequently reduces dissemination of bik(+) clones.
Suzuki
Mika
M
Department of Obstetrics and Gynecology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan.
Kobayashi
Hiroshi
H
Tanaka
Yoshiko
Y
Hirashima
Yasuyuki
Y
Kanayama
Naohiro
N
Takei
Yuji
Y
Saga
Yasushi
Y
Suzuki
Mitsuaki
M
Itoh
Hiroshi
H
Terao
Toshihiko
T
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002277:Carcinoma / Q000235:genetics / Q000517:prevention & control* / Q000556:secondary; D017931:DNA Primers / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005341:Fibrinolysin / Q000037:antagonists & inhibitors / Q000378:metabolism; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D006801:Humans; D007124:Immunoenzyme Techniques; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009361:Neoplasm Invasiveness / Q000517:prevention & control; D010051:Ovarian Neoplasms / Q000235:genetics / Q000473:pathology / Q000517:prevention & control*; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors / Q000378:metabolism; D010534:Peritoneal Neoplasms / Q000235:genetics / Q000517:prevention & control* / Q000556:secondary; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015842:Serine Proteinase Inhibitors / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014357:Trypsin / Q000378:metabolism; D014360:Trypsin Inhibitor, Kunitz Soybean; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; C109148:SPINT2 protein, human; D015842:Serine Proteinase Inhibitors; D014360:Trypsin Inhibitor, Kunitz Soybean; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D010196:Pancreatic Elastase; D014357:Trypsin; D005341:Fibrinolysin