BackgroundThe tumor suppressor Programmed Cell Death 4 (PDCD4) has been found to be under-expressed in several cancers and associated with disease progression and metastasis. There are no current studies characterizing PDCD4 expression and its clinical relevance in Oral Squamous Cell Carcinoma (OSCC). Since nodal metastasis is a major prognostic factor in OSCC, we focused on determining whether PDCD4 under-expression was associated with patient nodal status and had functional relevance in OSCC invasion. We also examined PDCD4 regulation by microRNA 21 (miR-21) in OSCC.ResultsPDCD4 mRNA expression levels were assessed in 50 OSCCs and 25 normal oral tissues. PDCD4 was under-expressed in 43/50 (86%) OSCCs, with significantly reduced mRNA levels in patients with nodal metastasis (p = 0.0027), and marginally associated with T3-T4 tumor stage (p = 0.054). PDCD4 protein expression was assessed, by immunohistochemistry (IHC), in 28/50 OSCCs and adjacent normal tissues; PDCD4 protein was absent/under-expressed in 25/28 (89%) OSCCs, and marginally associated with nodal metastasis (p = 0.059). A matrigel invasion assay showed that PDCD4 expression suppressed invasion, and siRNA-mediated PDCD4 loss was associated with increased invasive potential of oral carcinoma cells. Furthermore, we showed that miR-21 levels were increased in PDCD4-negative tumors, and that PDCD4 expression may be down-regulated in OSCC by direct binding of miR-21 to the 3'UTR PDCD4 mRNA.ConclusionsOur data show an association between the loss of PDCD4 expression, tumorigenesis and invasion in OSCC, and also identify a mechanism of PDCD4 down-regulation by microRNA-21 in oral carcinoma. PDCD4 association with nodal metastasis and invasion suggests that PDCD4 may be a clinically relevant biomarker with prognostic value in OSCC.

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Histone deacetylase inhibitors (HDACi) have recently emerged as promising anticancer agents. However, the mechanisms by which HDAC inhibitors arrest proliferation and induce apoptosis in tumor cells is far from clear. Activation of the stress MAP kinases and induction of DNA damage by different HDACi have been reported, however, the potential role of the MAP kinase p38 in the antitumor activity of these drugs has not been described. P38 is known to be activated independently or downstream of DNA damage. The purpose of this study was to elucidate the mechanisms by which the HDACi vorinostat (Zolinza®) triggers apoptosis in haematological malignant cells. We show that DNA damage as well as activation of p38 occurs relatively early in acute myeloid leukemia (AML) cell lines after treatment with vorinostat. Using comet assays, we detected direct evidence of early DNA damage and western blotting revealed induction of the DNA damage response proteins ATM and Chk2. We performed cell cycle analysis, and observed within the vorinostat-treated AML cell population, cells exiting G1 and accumulating in the G2-M phase of the cell cycle, where they subsequently underwent apoptosis. Notably, downregulation of p38 by shRNA or inhibition of p38 and β activity by the inhibitor SB203580 significantly decreased both G2-M accumulation and apoptosis induced by vorinostat, indicating a pro-apoptotic p38 function. Interestingly, several other HDACi tested all induced p38 activation but, depending on the HDACi, this activation was found to be either pro or antiapoptotic. The short-chain fatty acid sodium butyrate (NaB) requires p38 for induction of apoptosis, like vorinostat. On the other hand, LBH589, from the structural class encompassing vorinostat, does not depend on p38 for induction of apoptosis. Furthermore, p38 serves as a pro-survival signal when induced by the benzimide MGCD0103. In conclusion, we have shown that vorinostat-induced apoptosis in AML cells is preceded by generation of DNA damage and accumulation of cells in the G2-M phase of the cell cycle. Further, G2-M arrest and apoptosis induction (but not DNA damage) by vorinostat requires activation of the p38 MAP kinase, which is not the case for all HDACi. Therefore, a better understanding of the role of p38 MAPK in the action of specific HDACi may help in the development of rational combination regimes including these targeted agents. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A188.