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© PhD Students Symposium 2005. All rights reserved.
Francis J. Markland, PhD
University of Southern California, Los Angeles, California, United States of America
Disintegrins are soluble peptides found in snake venom. They bind to RGD-responsive integrins with high affinity (nM range) and block integrin function. Integrins are heterodimeric proteins located on the surface of cells that are involved in cell-cell and cell-extracellular matrix interactions; they are involved in bidirectional signal transmission from the outside of the cell to the inside and from inside the cell to the outside. Disintegrins are classified according to their size as small, medium, large and dimeric. Contortrostatin (CN), a member of the disintegrin family, was isolated from southern copperhead venom. CN is a homodimeric disintegrin with 65 amino acids in each chain and each chain containing an Arg-Gly-Asp (RGD) motif. CN binds to integrin αIIbβ3 (fibrinogen receptor) on platelets and blocks platelet aggregation with an IC50 of ~65nM. CN also binds with high affinity to integrins αvβ3, αvβ5 and α5β1 on tumor and endothelial cells; it effectively inhibits adhesion, migration and invasion of human breast, ovarian and prostate cancer and glioma cell lines and inhibits human endothelial cell adhesion, invasion and tube formation. In a nude mouse model of human breast cancer, we have shown that CN is an effective inhibitor of cancer growth, angiogenesis and lung metastasis. However, the initial in vivo studies employed daily intratumor injection of CN, which is not a clinically relevant delivery mechanism. Therefore, we developed a liposome encapsulated form of CN (LCN), which can be administered intravenously. Metastatic, human breast cancer cells (MDA-MB-435, 105 cells) were injected into mammary fat pads of nude mice. LCN administered IV twice weekly at 210µg weekly dose, starting two-weeks after injection of the mammary cancer cells, was as effective as daily intratumor administration at the same weekly dose, but represents a much more clinically relevant delivery method. Angiogenesis was inhibited by ~94% (CD31 immunohistochemistry) using this method of delivery. We have recently developed an E. coli expression system to produce a recombinant version of CN (rCN). Interestingly, although rCN is a monomer it possesses in vivo activity equivalent to that of native CN. The availability of recombinant CN will facilitate clinical development of this interesting anti-angiogenic agent with potent anti-tumor activity.