3-azidoWA Blocks MMP-2 and Induces Secretory Par-4 Figure 7. The macroscopic appearance of angiogenesis in Matrigel plugs. (A). Preventive study: The effect of dosing with 3-azido WA (30 mg/kg/d, i,p.) for 1–4 days on neovascularisation of Matrigel plugs. After dosing for 1–4 days plugs were removed on day 7 after Matrigel injection for visualisation and quantification of angiogenesis. Representive photographs of plugs from groups of five animals are shown. Quantification of angiogenesis within Matrigel plugs show (n = 5, P,0.05) compared with VEGF +bFGF. (B). Treatment study: The effect of dosing with 3-azido WA (30 mg/kg/d, i,p.) on established Matrigel plug vasculature. Neovascularisation was allowed to develop for 7 days animals were dosed for 1 day or daily for 3 days. Plugs were removed at the end of day 11 for photography and quantification after plugs were removed. Quantification of angiogenesis within Matrigel plugs shows (n = 5, P,0.05) compared with VEGF +bFGF. doi:10.1371/journal.pone.0044039.g007 Matrix metalloproteinases are the lead candidates to facilitate tumor invasion process by triggering a chain of signaling cascades and play crucial role not only in invasion, angiogenesis and metastasis, but also determine cancer cell transformation, growth, apoptosis and signal transduction [28,40]. In the last twenty years majority of the MMP inhibitors showed lack of success in clinical trials because none of these synthetic/semi synthetic drugs were selective for specific MMP’s. Thus, learning lessons from looking back at previous failed attempts necessitate developing selective MMP inhibitors that should not have cross-reaction with other MMP’s [36]. Here we demonstrate that 3-azidoWA only selectively blocked MMP-2 activity and thus motility and invasion of diverse cell line in extracellular Par-4 dependent manner. By degrading the basement membrane, MMP’s are considered to accelerate cell motility in a stationary tumor cell and promote metastasis [41]. Mounting evidences have suggested that MMP’s not only function in cancer progression and metastasis but may also contribute to steps of cancer development [42–44]. These diverse activities may account for the role of MMP-2, a pivotal matrix metalloprotease that control cancer cell motility and invasion [45]. In the present study we have illustrated that non- toxic physiologically relevant doses of 3-azidoWA significantly (P,0.01) inhibited the migration and invasion of HeLa and PC-3 (Supplementary Fig. 1) cells when treated in a dose dependent fashion in Boyden chamber invasion assay. PLOS ONE | www.plosone.org 11 September 2012 | Volume 7 | Issue 9 | e44039