Angiostatin generation by cathepsin D secreted by human prostate carcinomacells

Angiostatin, a potent endogenous inhibitor of angiogenesis, is generated by cancer-mediated proteolysis of plasminogen. The culture medium of human pr ostate carcinoma cells, when incubated with plasminogen at a variety of pH values, generated angiostatic peptides and miniplasminogen. The enzyme(s) r esponsible for this reaction was purified and identified as procathepsin D. The purified procathepsin D, as well as cathepsin D, generated two angiost atic peptides having the same NH2-terminal amino acid sequences and compris ing kringles 1-4 of plasminogen in the pH range of 3.0-6.8, most strongly a t pH 4.0 in vitro. This reaction required the concomitant conversion of pro cathepsin D to catalytically active pseudocathepsin D. The conversion of ps eudocathepsin D to the mature cathepsin D was not observed by the prolonged incubation. The affinity-purified angiostatic peptides inhibited angiogene sis both in vitro and in vivo. Importantly, procathepsin D secreted by huma n breast carcinoma cells showed a significantly lower angiostatin-generatin g activity than that by human prostate carcinoma cells. Since deglycosylate d procathepsin D from both prostate and breast carcinoma cells exhibited a similar low angiostatin-generating activity, this discrepancy appeared to b e attributed to the difference in carbohydrate structures of procathepsin D molecules between the two cell types. The seminal vesicle fluid from patie nts with prostate carcinoma contained the mature cathepsin D and procatheps in D, but not pseudocathepsin D, suggesting that pseudocathepsin D is not a normal intermediate of procathepsin D processing in vivo. The present stud y provides evidence for the first time that cathepsin D secreted by human p rostate carcinoma cells is responsible for angiostatin generation, thereby causing the prevention of tumor growth and angiogenesis-dependent growth of metastases.