Five proteins with anticoagulant and antimetastatic activities were isolated from the salivary glands of the Amazon leech, Haementeria ghilianil. These proteins, designated ghilantens, were co-purified on DEAE-cellulose and heparin-agarose, and were purified by microbore C-18 reverse-phase HPLC. Each variant had a similar molecular weight (18,000), amino acid composition, and a blocked amino terminus. Ghilantens caused a dose-dependent prolongation of the prothrombin time of normal human plasma and blocked the factor Xa-mediated hydrolysis of methoxycarbonyl-D-cyclohexylglycyl-glycl-arginine-p-nitro anillide acetate. Ghilantens were quantitatively absorbed to bovine factor Xa-AffiGel-15 and were eluted with 0.1 mol/L benzamidine, an active-site reversible inhibitor of factor Xa. These findings show that ghilantens can form a reversible association with the enzyme. When administered intravenously to mice by tall vein injection, ghilantens potently suppressed lung metastases of B16-F10 melanoma cells. These findings suggest that ghilantens may have therapeutic value in the treatment of metastatic disease.