CONTORTROSTATIN, A SNAKE-VENOM DISINTEGRIN, INHIBITS BETA-1 INTEGRIN-MEDIATED HUMAN METASTATIC MELANOMA CELL-ADHESION AND BLOCKS EXPERIMENTAL METASTASIS

Disintegrins are Arg-Gly-Asp-containing proteins that inhibit integrin -mediated cell-cell and cell-matrix interactions. We have purified a d isintegrin, contortrostatin, from Agkistrodon contortrix contortrix sn ake venom that is a potent inhibitor of human metastatic melanoma (M24 met) cell adhesion to extracellular matrix proteins. Contortrostatin inhibits M24 met cell adhesion to type I collagen, vitronectin, and fi bronectin with 50% inhibitory concentration values of 20, 75, and 220 nM, respectively. Contortrostatin does not significantly inhibit adhes ion of M24 met cells to laminin. I-125-labeled contortrostatin binds t o M24 met cells in a saturable and displaceable manner. Scatchard anal ysis indicates that there are two binding sites for I-125-labeled cont ortrostatin on the surface of these cells. High affinity binding has a K-d of 3 nM with 165,000 sites/cell; low affinity binding has a K-d o f 60 nM with 500,000 sites/cell. Immobilized contortrostatin can suppo rt adhesion of M24 met cells; this binding is blocked by a monoclonal antibody to the beta(1) integrin subunit and by an antibody to the fib ronectin receptor alpha(5) beta(1). The anti-vitronectin receptor (alp ha(v) beta(5)) monoclonal antibody which blocks adhesion of M24 met ce lls to immobilized vitronectin does not block binding of M24 met cells to immobilized contortrostatin. In an in vivo experimental metastasis model system, contortrostatin at 20 mu g and 100 mu g inhibits lung c olonization of M24 met cells (5 x 10(5)), injected in the tail vein of scid mice, by 51 and 73%, respectively. We conclude that contortrosta tin is a potent inhibitor of beta(1) integrin-mediated M24 met cell ad hesion in vitro and that it also inhibits lung colonization in vivo.