eristostatin and Neoplasm-Metastasis

Peptides containing the integrin recognition sequence, RGD, can inhibit experimental metastasis of mouse melanoma cells, but the integrin(s) affected in these experiments is unknown. Besides "classical" RGD-binding integrins such as alpha 5 beta 1 and alpha v beta 3, RGD has been reported to bind alpha 4 beta 1, and mAbs to alpha 4 beta 1 can inhibit melanoma metastasis. We investigated the mode of action of the disintegrin eristostatin, an RGD-containing peptide isolated from snake venom, in a human melanoma experimental metastasis model. Lung colonization following i.v. injection of MV3 cells in nude mice was strongly inhibited by eristostatin. MV3 cells bound FITC-eristostatin and adhered to eristostatin-coated wells. This adhesion was partially inhibited by a GRGDSP peptide and by alpha 4 mAb. Binding of FITC-eristostatin to Jurkat cells and adhesion of Jurkat (but not K562) cells to eristostatin-coated wells further suggested that eristostatin binds alpha 4 beta 1, even though, again, alpha 4 mAb only partially inhibited adhesion. Expression of alpha 4 beta 1 was enhanced in metastatic melanoma cells compared to normal melanocytes and nonmetastatic melanoma cells. Finally, eristostatin inhibited adhesion of both MV3 and CHO alpha 4 cells to the alpha 4 beta 1-ligand VCAM-1, while adhesion to other ligands via other integrins was not affected. These findings demonstrate that inhibition of melanoma cell metastasis by RGD-containing peptides such as eristostatin, may be due to interference with alpha 4 beta 1-VCAM binding, in addition to inhibition of the classical RGD-binding integrins.

Topics: Animals; Binding Sites; Humans; Infant, Newborn; Integrin alpha4beta1; Integrins; Intercellular Signaling Peptides and Proteins; Male; Melanocytes; Melanoma; Mice; Mice, Nude; Neoplasm Metastasis; Oligopeptides; Peptides; Platelet Aggregation Inhibitors; Receptors, Lymphocyte Homing; Skin; Skin Neoplasms; Snake Venoms; Viper Venoms

Adhesion molecules, including integrins, are important for interactions of cancer cells with vessel walls, a step leading to cancer metastasis. Disintegrins block the action of integrins by binding to them. We tested the hypothesis that the disintegrin eristostatin would block metastasis by interfering with cancer cell adhesion to vessel walls, thus preventing extravasation. Experimental metastasis assays, in which B16F1 melanoma cells (controls vs eristostatin-treated, 25 micrograms/ml) were injected via mesenteric veins of anesthetized C57BL/6 mice, showed that eristostatin reduced (P < 0.05) the mean number of liver metastases from 14.4 to 0.6 at 11 days postinjection (p.i.). We examined three different steps in metastasis at which eristostatin could have exerted its effect, namely, cell arrest, extravasation, and migration. Control and eristostatin-treated B16F1 cells were fluorescently labeled and examined by videomicroscopy in liver microcirculation in vivo at various times up to 14 days p.i. Measurements of vessel size in which cell arrest occurred and length/width ratio of arrested cells showed only small differences between control and eristostatin-treated cells. Eristostatin treatment did not prevent extravasation, and the timing and process of extravasation were similar for both treated and control cells; by 3-4 days p.i. more than 90% of the cells had extravasated or were in the process. Eristostatin also did not affect the ability of extravasated cells to migrate through the extracellular matrix to the subcapsular region where tumors later form. Therefore, we conclude that eristostatin exerted its primary effect by regulating the number of individual cancer cells that grow after extravasation.

Topics: Animals; Blood Vessels; Cell Adhesion; Cell Movement; Integrins; Liver Neoplasms; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Microscopy, Video; Neoplasm Metastasis; Peptides; Viper Venoms