αv‐Integrin antagonist EMD 121974 induces apoptosis in brain tumor cells growing on vitronectin and tenascin

Abstract

Orthotopic brain tumor growth is inhibited in athymic mice by the daily systemic administration of the ␣v-integrin antagonist EMD 121974. This compound, a cyclic RGDpenta-peptide, is a potent inhibitor of angiogenesis, which induces apoptosis of growing endothelial cells through inhibition of their ␣v-integrin interaction with the matrix proteins vitronectin and tenascin. Here we show that EMD 121974 also induces apoptosis in the ␣v-integrin-expressing tumor cell lines U87 MG and DAOY by detaching them from vitronectin and tenascin, matrix proteins known to be essential for brain tumor growth and invasion. These matrix proteins are shown to be produced by the brain tumor cells in vitro and in vivo. Key words: brain tumor; xenograft; anti-angiogenesis; ␣v-integrins; apoptosisAngiogenesis, the recruitment of new blood vessels from existing capillaries, is essential for tumor growth and progression. 1 This complex process is characterized by proliferation, migration and invasion of capillary endothelial cells, as well as functional features that depend on their interactions with the surrounding extracellular matrix components. The expression of the integrin adhesion molecules ␣v␤3 and ␣v␤5 on sprouting capillary cells and their interaction with specific matrix ligands has been shown to play a key role in angiogenesis. [2][3][4] Although vitronectin is the only matrix ligand for ␣v␤5, the promiscuous ␣v␤3 receptor interacts with various matrix proteins including vitronectin, tenascin, fibronectin, fibrinogen, etc. 5 Ligand binding to these receptors through specific RGD motifs leads to complex intracellular signaling resulting in endothelial cell survival. 3,6,7 Consequently, blocking of these receptors with specific antibodies or RGD peptides results in endothelial cell apoptosis in vitro and suppression of angiogenesis in vivo, with subsequent suppression of tumor growth. 2,4 We have recently shown that the cyclic RGD pentapeptide EMD 121974, a specific ␣v-integrin antagonist, inhibited growth of brain tumor cell lines xenotransplanted into the forebrain of nu/nu mice, resulting in long-term survival. 8 In the present study, we investigated in more detail the mechanisms responsible for this growthinhibitory effect. Our data demonstrate that EMD 121974 detaches both the ␣v-integrin-expressing brain capillary and brain tumor cells from the matrix proteins vitronectin and tenascin, resulting in significant apoptosis of both cell types. We also show that the brain tumor cells produce these matrix proteins in vitro and in vivo. Furthermore, we found that only ␣v-integrin-expressing tumor cells responded in vivo to the treatment with EMD 121974. Taken together, these data suggest that EMD 121974 not only inhibits angiogenesis but is also cytotoxic for brain tumor cells. MATERIAL AND METHODS Cell linesThe human brain tumor cell lines DAOY (medulloblastoma) and U87MG (glioblastoma) were purchased from ATCC (Manassas, VA) and grown in RPMI-1640 medium with 10% FBS in 5% CO 2 at 37°C. Primary human and mouse brain capillar...

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