MMP-9 inhibition provokes premature senescence resulting in inhibition of medulloblastoma tumor growth via p16 INK4A and mitogen-activated protein kinase signaling pathway

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The involvement of matrix metalloproteinases (MMPs)has been suggested in cellular mechanisms leadingto metastatic disease in medulloblastoma (MB), the most commonmalignant brain tumor in children. Matrix metalloproteinases comprise a family of zinc-dependent enzymes, which degrade components of the extracellular matrix (type IV collagen) and play an important role in facilitating neoplastic cell invasion and metastasis. A significantassociation of the expression levels of MMP-9 with survival and M stage suggests that patientswith medulloblastoma metastatic disease at diagnosis may benefit from the anti-MMPtherapy. Here, we have evaluated the tumorigenicity of medulloblastoma cells after infection with an adenovirus containing a 21 bp siRNA sequence of human MMP-9 gene (Ad-MMP-9). Infection of Daoy medulloblastoma cells with Ad-MMP-9 reduced MMP-9 activity and protein levels. Ad-MMP-9 decreased the number of viable Daoy cells in a concentration-dependent manner. The invasive ability of Ad-MMP-9-infected medulloblastoma cells was also reduced in a dose-dependent manner when compared with parental and Ad-SV controls as determined by matrigel invasion assay. Fluorescence-activated cell sorter analyses of cellsinfected with Ad-MMP-9 demonstrated a dose-dependent cell cyclearrest in the G₀/G₁ phase. Ad-MMP-9-induced cell cycle arrest appears to be mediatedby a pathway that requires the ERK mitogen-activated proteinkinase, as well as the cell cycle inhibitors p16^INK4 and is phenotypically indistinguishable from senescence. Ad-MMP-9 treatment inhibited medulloblastoma tumor growth in an intracranial model and was mediated by upregulation of p16 expression. These studies validate the usefulness of targeting MMP-9 and provide a novel perspective in the treatment of pediatric patients with medulloblastoma.