Nonlysine-analog plasminogen modulators promote autoproteolytic generation of plasmin(ogen) fragments with angiostatin-like activity.
We recently discovered several nonlysine-analog conformational modulators for plasminogen. These include SMTP-6, thioplabin B and complestatin that are low molecular mass compounds of microbial origin. Unlike lysine-analog modulators, which increase plasminogen activation but inhibit its binding to fibrin, the nonlysine-analog modulators enhance both activation and fibrin binding of plasminogen. Here we show that some nonlysine-analog modulators promote autoproteolytic generation of plasmin(ogen) derivatives with its catalytic domain undergoing extensive fragmentation (PMDs), which have angiostatin-like anti-endothelial activity. The enhancement of urokinase-catalyzed plasminogen activation by SMTP-6 was followed by rapid inactivation of plasmin due to its degradation mainly in the catalytic domain, yielding PMD with a molecular mass ranging from 68 to 77 kDa. PMD generation was observed when plasmin alone was treated with SMTP-6 and was inhibited by the plasmin inhibitor aprotinin, indicating an autoproteolytic mechanism in PMD generation. Thioplabin B and complestatin, two other nonlysine-analog modulators, were also active in producing similar PMDs, whereas the lysine analog 6-aminohexanoic acid was inactive while it enhanced plasminogen activation. Peptide sequencing and mass spectrometric analyses suggested that plasmin fragmentation was due to cleavage at Lys615-Val616, Lys651-Leu652, Lys661-Val662, Lys698-Glu699, Lys708-Val709 and several other sites mostly in the catalytic domain. PMD was inhibitory to proliferation, migration and tube formation of endothelial cells at concentrations of 0.3-10 microg.mL(-1). These results suggest a possible application of nonlysine-analog modulators in the treatment of cancer through the enhancement of endogenous plasmin(ogen) fragment formation.
Ohyama
Shigeki
S
Department of Applied Biological Science, Tokyo Noko University, Saiwaicho, Fuchu-shi, Tokyo, Japan.
Harada
Tomotaka
T
Chikanishi
Toshihiro
T
Miura
Yutaka
Y
Hasumi
Keiji
K
D000596:Amino Acids / Q000032:analysis; D015119:Aminocaproic Acid / Q000494:pharmacology; D043168:Angiostatins / Q000737:chemistry* / Q000494:pharmacology; D000818:Animals; D001578:Benzopyrans / Q000737:chemistry / Q000494:pharmacology; D001665:Binding Sites; D016466:CHO Cells; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D002733:Chlorophenols / Q000737:chemistry / Q000494:pharmacology; D006224:Cricetinae; D004730:Endothelium, Vascular / Q000166:cytology; D005341:Fibrinolysin / Q000378:metabolism; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D051379:Mice; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000737:chemistry / Q000494:pharmacology; D010958:Plasminogen / Q000737:chemistry / Q000378:metabolism*; D010960:Plasminogen Activators / Q000737:chemistry / Q000494:pharmacology*; D011760:Pyrrolidinones / Q000737:chemistry / Q000494:pharmacology; D020539:Sequence Analysis, Protein / Q000379:methods; D014471:Umbilical Veins
D000596:Amino Acids; D001578:Benzopyrans; D002733:Chlorophenols; D010446:Peptide Fragments; D010456:Peptides, Cyclic; D011760:Pyrrolidinones; C117810:SMTP 6; C023490:complestatin; D043168:Angiostatins; D010958:Plasminogen; D010960:Plasminogen Activators; D005341:Fibrinolysin; D015119:Aminocaproic Acid