Abbreviations: CK1, casein kinase 1; GSK-3β, glycogen synthase kinase-3β; PAUF, pancreatic adenocarcinoma up-regulated factor; PKA, protein kinase A; sh-RNA, short hairpin-RNA; TCF/LEF, T cell factor/lymphocyte enhancer factor

Many oncolytic viruses are currently being tested as potential cancer therapeutic agents. To be effective, these viruses must replicate and propagate efficiently through the tumor mass. However, it is possible that the hypoxia that characterizes many tumors may be an obstacle to viral therapy because of its inhibition of viral replication and propagation. We, therefore, decided to test how oncolytic reovirus and its target cells respond to hypoxia. We found that reovirus infection suppresses hypoxia inducible factor (HIF)-1a protein levels (but not transcript abundance) in colon cancer HCT116 cells under CoCl 2 or hypoxia. Reovirus infection was able to reduce HIF-1a levels in both von Hippel Lindau (VHL)À/À renal carcinoma A498 and p53À/À HCT116 cells, indicating that the decrease of HIF-1a mediated by reovirus requires neither VHL nor p53 proteins. However, treatment with the inhibitor MG132 restored HIF-1a levels, suggesting that reovirus-induced HIF-1a decrease needs proteosomal activity. A498 VHLÀ/À cells with constitutive expression of HIF-1a were relatively resistant to reovirus-induced apoptosis when compared with A498 VHL þ / þ cells. However, we found that the use of YC-1 to target HIF-1a promoted reovirus-induced apoptosis in A498 VHLÀ/À cells. Accordingly, we propose that reovirus may be used together with YC-1 as a potential therapeutic agent against chemoresistant or radioresistant tumors that are hypoxic and show increased levels of HIF-1a.

Pancreatic cancer is a notorious disease with a poor prognosis and low survival rates, which is due to limited advances in understanding of the molecular mechanism and inadequate development of effective treatment options over the decades. In previous studies, we demonstrated that a novel soluble protein named pancreatic adenocarcinoma up-regulated factor (PAUF) acts on tumor and immune cells and plays an important role in metastasis and progression of pancreatic cancer. Here we show that PAUF promotes adhesiveness of pancreatic cancer cells to various extracellular matrix (ECM). Our results further support a positive correlation of activation and expression of focal adhesion kinase (FAK), a key player in tumor cell metastasis and survival, with PAUF expression. PAUF-mediated adhesiveness was significantly attenuated upon blockade of the FAK pathway. Moreover, PAUF appeared to enhance resistance of pancreatic cancer cells to anoikis via modulation of FAK. Our results suggest that PAUF-mediated FAK activation plays an important role in pancreatic cancer progression.

Abstract. The hepatitis B virus X (HBX) protein, a regulatory protein of the hepatitis B virus (HBV), has been shown to generate reactive oxygen species (ROS) in human liver cell lines; however, the mechanism by which cells protect themselves under this oxidative stress is poorly understood. Here, we show that HBX induces the up-regulation of Forkhead box class O 4 (Foxo4) not only in chang cells stably expressing HBX (chang-HBX) but also in primary hepatic tissues from HBX-transgenic mice. HBX also increased ROS, but reduction of the abundance of ROS using N-acetylcystein (NAc) diminished the levels of Foxo4. Elevated Foxo4 was also detected in nuclei of chang-HBX cells but not in chang cells stably expressing the vector (chang-Vec), suggesting that HBX activates the transcriptional activity of Foxo4. When we examined whether HBX bypasses JNK signaling that targets Foxo4, we found that the activity of JNK but not of ERK is required for the up-regulation of Foxo4 even in the presence of HBX. Furthermore, the reduction of Foxo4 levels using siRNA or a JNK inhibitor rendered chang-HBX cells sensitive to apoptosis under oxidative stress, suggesting that up-regulation of Foxo4 mediated by HBX enhances resistances to oxidative stress-induced cell death. Accordingly, we propose that Foxo4 may be a useful target for suppression in the treatment of HBV-associated hepatocellular carcinoma cells.

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