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 disintegrin, contortrostatin, from Agkistrodon contortrix contortrix snake 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 fibronectin with 50% inhibitory concentration values of 20, 75, and 220 nM, respectively. Contortrostatin does not significantly inhibit adhesion of M24 met cells to laminin. 125I-labeled contortrostatin binds to M24 met cells in a saturable and displaceable manner. Scatchard analysis indicates that there are two binding sites for 125I-labeled contortrostatin on the surface of these cells. High affinity binding has a Kd of 3 nM with 165,000 sites/cells low affinity binding has a Kd of 60 nM with 500,000 sites/cell. Immobilized contortrostatin can support 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 fibronectin receptor alpha 5 beta 1. The anti-vitronectin receptor (alpha v beta 5) monoclonal antibody which blocks adhesion of M24 met cells 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 micrograms and 100 micrograms inhibits lung colonization of M24 met cells (5 x 10(5)), injected in the tail vein of scid mice, by 51 and 73%, respectively. We conclude that contortrostatin is a potent inhibitor of beta 1 integrin-mediated M24 met cell adhesion in vitro and that it also inhibits lung colonization in vivo.
Trikha
M
M
Department of Biochemistry and Molecular Biology, University of Southern California School of Medicine, Los Angeles 90033.
De Clerck
Y A
YA
Markland
F S
FS
D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects*; D019483:Disintegrins; D006801:Humans; D019012:Integrin beta1; D016023:Integrins / Q000037:antagonists & inhibitors*; D008175:Lung Neoplasms / Q000556:secondary; D008545:Melanoma / Q000378:metabolism / Q000473:pathology / Q000556:secondary*; D051379:Mice; D016513:Mice, SCID; D010455:Peptides / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*; D012910:Snake Venoms / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*
D019483:Disintegrins; D019012:Integrin beta1; D016023:Integrins; D010455:Peptides; D012910:Snake Venoms