Histone deacetylase inhibitors VPA and TSA induce apoptosis and autophagy in pancreatic cancer cells.

PURPOSE: Histone deacetylase inhibitors (HDACi) are anti-neoplastic agents that are known to affect the growth of different cancer types, but their underlying mechanisms are still incompletely understood. Here, we compared the effects of two HDACi, i.e., Trichostatin A (TSA) and Valproic Acid (VPA), on the induction of cell death and autophagy in pancreatic cancer-derived cells that exhibit a high metastatic capacity and carry KRAS/p53 double mutations. METHODS: Cell viability and proliferation tests were carried out using Trypan blue dye exclusion, MTT and BrdU assays. FACS analyses were carried out to assess cell cycle progression, apoptosis, reactive oxygen species (ROS) production and mitochondrial depolarization, while Western blot and immunoprecipitation analyses were employed to detect proteins involved in apoptosis and autophagy. RESULTS: We found that both VPA and TSA can induce apoptosis in Panc1 and PaCa44 pancreatic cancer-derived cells by triggering mitochondrial membrane depolarization, Cytochrome c release and Caspase 3 activation, although VPA was more effective than TSA, especially in Panc1 cells. As underlying molecular events, we found that ERK1/2 was de-phosphorylated and that the c-Myc and mutant p53 protein levels were reduced after VPA and, to a lesser extent, after TSA treatment. Up-regulation of p21 and Puma was also observed, concomitantly with mutant p53 degradation. In addition, we found that in both cell lines VPA increased the pro-apoptotic Bim level, reduced the anti-apoptotic Mcl-1 level and increased ROS production and autophagy, while TSA was able to induce these effects only in PaCA44 cells. CONCLUSIONS: From our results we conclude that both VPA and TSA can induce pancreatic cancer cell apoptosis and autophagy. VPA appears have a stronger and broader cytotoxic effect than TSA and, thus, may represent a better choice for anti-pancreatic cancer therapy.

Sodium butyrate sensitises human pancreatic cancer cells to both the intrinsic and the extrinsic apoptotic pathways.

Pancreatic cancer is characterised by a highly malignant phenotype with a marked resistance to conventional therapies and to apoptotic activators. Here, we demonstrate that sodium butyrate (NaBt), an inhibitor of histone deacetylases, sensitises human pancreatic cancer cell lines to both mitochondria- and Fas-mediated apoptosis. The analysis of anti-apoptotic and pro-apoptotic members of the Bcl-2 family in untreated pancreatic cancer cell lines shows a generalised low expression of Bcl-2 and a strong expression of Bcl-xL. NaBt treatment results in a marked down-regulation of Bcl-xL expression, mitochondrial membrane depolarization, cytochrome c release from mitochondria, activation of caspase-9 and -3 and apoptosis induction. Furthermore, NaBt sensitises pancreatic cancer cells to Fas-mediated apoptosis as well. In fact, the combined treatment with NaBt and the agonistic antibody anti-Fas (CH11) is able to induce apoptosis at an early time, in which neither NaBt nor CH11 alone induce apoptosis. Down-regulation of FLIP and activation of caspase-8 allow apoptosis to occur. These findings suggest that sodium butyrate could represent a good candidate for the development of new therapeutic strategies aimed at improving chemotherapy and immunotherapy in pancreatic cancer.