Features emblematic to glioblastoma multiforme (GBM) include diffuse tumor cell invasion into normal brain and hyperplastic tumor-associated endothelium. Previously we reported the over-expression of MMP-1 in human GBM and demonstrated the importance of MMP-1 over-expression in GBM cell motility. Herein we report on our current investigations into the potential consequences of MMP-1 expression in GBM cells as related to: proteomic shifts with changes in MMP-1, tumor take in a non-permissive environment in vivo, and endothelial proliferation and differentiation in vitro. METHODS: The T98G cell line produces a detectable level of MMP-1. These cells were stably transduced with a MMP-1 specific shRNA lentivirus. Conversely U251 cells do not produce MMP-1 and these were stably transfected with a cDNA vector containing MMP-1. Cell lysates were applied to proteomic arrays containing antibodies for angiogenesis related proteins. To assess tumor cell growth in a non-permissive setting, athymic nude mice were inoculated in the hind flanks with these cells. Palpable tumor growth was observed for up to 41 days using external caliper measurement. In vitro endothelial assays consisted of microcarrier beads coated with endothelial cells and immobilized in a three-dimensional fibrin gel culture system. The gels were overlaid with either soluble factors or glioma cells. After 2-5 days of incubation, beads were randomly assessed for endothelial sprout number as a sign of proliferation and sprout length as differentiation. RESULTS: Angiogenesis antibody arrays revealed a balance between MMP-1 expression and anti-angiogenic proteins. With the over-expression of MMP-1 the detection of endostatin and CXCL4 decreased. Yet most notable is an induction of tissue inhibitor of metalloproteinases (TIMP)-4 in the absence of MMP-1. This was confirmed with immunoblot analysis. Tumor growth data indicated a significant increase in tumor volume with MMP-1 over-expression and decrease with knockdown (p<0.05). This was during the initial growth delay phase of either tumor type indicating that tumor take in a non-permissive environment was impacted. There was a 62.5% reduction in tumor take with MMP-1 knockdown, and a 44.4% increase with over-expression (p<0.05). Three-dimensional culture data indicated increased endothelial proliferation and differentiation with either recombinant MMP-1 or co-cultured over-expressers, and the converse with MMP-1 knockdowns (p<0.05). Altogether these and prior data implicate MMP-1 as an important factor in the hallmark GBM pathologies. MMP-1 positively influences the propagation of small population tumor foci in a non-permissive host environment, and it is integral to angiogenic induction. A key mechanism of MMP-1 mediated induction of angiogenesis may involve a coordinated inhibition of TIMP-4 and other anti-angiogenic proteins.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2327.