Heparanase (HPSE) and fibroblast growth factor-2 (FGF2) are critical regulators of melanoma angio-genesis and metastasis. Elevated HPSE expression contributes to melanoma progression; however, further augmentation of HPSE presence can inhibit tumori-genicity. HPSE enzymatically cleaves heparan sulfate glycosaminoglycan chains (HS) from proteoglycans. HS act as both low-affinity FGF2 receptors and co-receptors in the formation of high-affinity FGF2 receptors. We have investigated HPSE's ability to modulate FGF2 activity through HS remodeling. Extensive HPSE degradation of human metastatic melanoma cells (70W) inhibited FGF2 binding. Unexpectedly, treatment of 70W cells with low HPSE concentrations enhanced FGF2 binding. In addition, HPSE-unexposed cells did not phosphorylate extracellular signal-related kinase (ERK) or focal adhesion kinase (FAK) in response to FGF2. Conversely, in cells treated with HPSE, FGF2 stimulated ERK and FAK phosphorylation. Secondly, the presence of soluble HPSE-degraded HS enhanced FGF2 binding and ERK phosphorylation at low HS concentrations. Higher concentrations of soluble HS inhibited FGF2 binding, but FGF2 signaling through ERK remained enhanced. Soluble HS were unable to support FGF2-stimulated FAK phosphorylation irrespective of HPSE treatment. Finally, cell exposure to HPSE or to HPSE-degraded HS modulated FGF2-induced angiogenesis in melanoma. In conclusion, these effects suggest relevant mechanisms for the HPSE modulation of melanoma growth factor responsiveness and tumorigenicity. Neoplasia (2006) 8, 596-606

Cyclooxygenase-2 (COX-2) is important in the progression of epithelial tumors. Evidence indicates that -6 PUFAs such as arachidonic acid (AA) promote the growth of tumor cells; however, -3 fatty acids [eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] inhibit tumor cell proliferation. We investigated the effects of -3 PUFA on the expression and function of COX-2 in 70W, a human melanoma cell line that metastasizes to the brain in nude mice. We show that 1 ) tumor necrosis factor-␣ upregulates the expression of both COX-2 mRNA and prostaglandin E 2 (PGE 2 ) production, and 2 ) -3 and -6 PUFA regulate COX-2 mRNA expression and PGE 2 production. AA increased COX-2 mRNA expression and prostaglandin production in -6-stimulated 70W cells. Conversely, COX-2 mRNA expression decreased in cells incubated with EPA or DHA. AA increased Matrigel™ invasion 2.4-fold, whereas EPA or DHA did not. Additionally, PGE 2 increased in vitro invasion 2.5-fold, whereas exposure to PGE 3 significantly decreased invasion. Our results demonstrate that incubation of 70W cells with either AA or PGE 2 increased invasiveness, whereas incubation with EPA or DHA downregulated both COX-2 mRNA and protein expression, with a subsequent decrease in Matrigel™ invasion. Taken together, these results indicate that -3 PUFA regulate COX-2-mediated invasion in brain-metastatic melanoma.

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