The Mechanism of Cancer-Mediated Conversion of Plasminogen to the Angiogenesis Inhibitor Angiostatin.

Angiostatin,a potent naturally occurring inhibitor of angiogenesis and growth of tumor metastases,is generated by cancer-mediated proteolysis of plasminogen. Human prostate carcinoma cells (PC-3) release enzymatic activity that converts plasminogen to angiostatin. We have now identified two components released by PC-3 cells,urokinase (uPA) and free sulfhydryl donors (FSDs),that are sufficient for angiostatin generation. Furthermore,in a defined cell-free system,plasminogen activators [uPA,tissue-type plasminogen activator (tPA),or streptokinase],in combination with one of a series of FSDs (N-acetyl-l-cysteine,d-penicillamine,captopril,l-cysteine,or reduced glutathione] generate angiostatin from plasminogen. An essential role of plasmin catalytic activity for angiostatin generation was identified by using recombinant mutant plasminogens as substrates. The wild-type recombinant plasminogen was converted to angiostatin in the setting of uPA/FSD; however,a plasminogen activation site mutant and a catalytically inactive mutant failed to generate angiostatin. Cell-free derived angiostatin inhibited angiogenesis in vitro and in vivo and suppressed the growth of Lewis lung carcinoma metastases. These findings define a direct mechanism for cancer-cell-mediated angiostatin generation and permit large-scale production of bioactive angiostatin for investigation and potential therapeutic application.