Suramin, a polysulfonated naphthylurea, has anti-reverse transcriptase and anti-proliferative activities and inhibits the binding of various growth factors to their cell surface receptors. This drug is used in the treatment of acquired immunodeficiency syndrome and several types of cancers. Increased levels of circulating glycosaminoglycans have been observed in suramin-treated cancer patients, suggesting that it may inhibit glycosaminoglycan catabolism. Melanoma-derived heparanase, a heparan sulfate-specific endo-beta-D-glucuronidase that plays an important role in metastatic melanoma cell invasion through basement membranes, is inhibited by suramin in a dose-dependent manner: 100% inhibition was observed at a concentration of approximately 100 microM. Structurally related polysulfonated compounds, such as trypan blue and Evans blue, had lower heparanase inhibitory activities: the concentrations required for 50% heparanase inhibition (ID50) were 310-320 microM and six times higher than for suramin (ID50 = 46 microM). Oversulfated heparin tetrasaccharide, whose average molecular size is similar to suramin, had also much lower heparanase inhibitory activity than suramin. The inhibition constants (Ki) for suramin and oversulfated heparin tetrasaccharide were 48 and 290 microM, respectively. Suramin had a remarkable inhibitory activity against B16 melanoma cell invasion through reconstituted basement membranes (ID50 less than 10 microM). The inhibitory effects of suramin on melanoma heparanase and cell invasion appeared to be completely independent of its antiproliferative activity, because significant effects on melanoma cell growth were not observed at the concentrations of suramin used in this study. The results suggest that the antimetastatic effects of suramin may be due to its antiinvasive rather than antiproliferative activities.