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Histone deacetylases (HDACs) are key enzymes that modulate chromatin structure and regulate gene expression in cells by deacetylation of conserved lysine residues in histones. Since aberrant expression of genes involved in proliferation, differentiation and apoptosis is an important mechanism of tumorigenesis in various cancers, we examined the effect of the histone deacetylase inhibitor (HDACi), MS-275, on medulloblastoma cell viability and gene expression in vitro. We demonstrate that medulloblastoma cell lines, Daoy and D283, are sensitive to MS-275 treatment and exhibit a rapid accumulation of cells with a sub-G1 DNA content. This is accompanied by an increase in the expression and activity of caspase-8. Activation of caspase-3 as well as an induction of poly ADP-ribose polymerase (PARP) cleavage was also observed as early as 24-48 hours post-treatment with MS-275. To further confirm the involvement of the extrinsic pathway in mediating apoptosis in response to MS-275, we measured the expression of the death receptor FAS/Apo-1 by real-time PCR, western blotting and immunofluorescence assays. These experiments revealed that MS-275 not only increased the expression of FAS receptor, but also promoted a clustering of receptors as evidenced by focal immunostaining of FAS receptor on the surface of MS-275 treated cells. Additionally, FAS ligand expression remained unchanged following drug treatment and did not appear to be necessary for MS-275 induced apoptosis, providing further support for oligomerization of FAS receptors as a possible mechanism by which MS-275 promotes cell death. Finally, the introduction of a dominant negative FADD mutant in Daoy cells interfered with the ability of MS-275 to promote apoptosis. This is the first demonstration that up regulation of FAS receptor and caspase-8 expression by MS-275 plays an important role in mediating apoptosis in medulloblastoma cells.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA