The hypoxic environment in tumor is reported to play an important role in pancreatic cancer progression. The interaction between stromal and cancer cells also contributes to the malignant behavior of pancreatic cancer. In the present study, we investigated whether hypoxic stimulation affects stromal as well as pancreatic cancer cells. Our findings demonstrated that hypoxia remarkably elevated the HIF-1a expression in both pancreatic cancer (PK8) and fibroblast cells (MRC5). Hypoxic stimulation accelerated the invasive activity of PK8 cells, and invasiveness was thus further accelerated when the hypoxic PK8 cells were cultured with conditioned medium prepared from hypoxic MRC5 cells (hypoxic conditioned medium). MMP-2, MMP-7, MT1-MMP and c-Met expressions were increased in PK8 cells under hypoxia. Hypoxic stimulation also increased the hepatocyte growth factor (HGF) secretion from MRC5 cells, which led to an elevation of c-Met phosphorylation in PK8 cells. Conversely, the elevated cancer invasion, MMP activity and c-Met phosphorylation of PK8 cells were reduced by the removal of HGF from hypoxic conditioned medium. In immunohistochemical study, the HIF-1a expression was observed in surrounding stromal as well as pancreatic cancer cells, thus indicating hypoxia exists in both of cancer and stromal cells. Moreover, the stromal HGF expression was found to significantly correlate with not only the stromal HIF-1a expression but also the c-Met expression in cancer cells. These results indicate that the hypoxic environment within stromal as well as cancer cells activates the HGF/c-Met system, thereby contributing to the aggressive invasive features of pancreatic cancer. ' 2006 Wiley-Liss, Inc.Key words: hypoxia; tumor-stromal interaction; pancreatic cancer; hepatocyte growth factor; c-Met Most aggressive tumors have acquired the ability to develop their own blood vessels as they grow. However, the tumor cells grow faster than the endothelial cells, so the structure and function of the tumor vasculature is highly disorganized in comparison to normal tissues. 1,2 In this microenvironment, within tumor tissues, cancer cells are exposed to hypoxia. 3 Recent studies have revealed that hypoxia-inducible factor-1 (HIF-1) plays a key role in tumor progression under hypoxia. 4,5 HIF-1 is a heterodimeric basic helix-loop-helix PER/ARNT/SIM (HLH-PAS) transcription factor that consists of HIF-1a and a constitutively expressed aryl hydrocarbon receptor nuclear translocator (ARNT) known as HIF-1b. Under normoxic condition, HIF-1a is bound to the tumor suppressor Von Hippel-Lindau (VHL) protein. This protein complex causes HIF-1a to be targeted by proteasomes, thus leading to rapid protein degradation. Conversely, under hypoxic conditions, HIF-1a is stabilized and dimerized with HIF-1b, translocates to the nucleus and transactivates various gene expressions. HIF-1 is reported to regulate various gene expressions by binding to the hypoxia-responsive element on the target genes. More than 40 genes that are important for glucose transpo...

These data suggest that the HGF/c-Met signaling via HIF-1alpha ?may therefore negatively affect the prognosis in patients with pancreatic cancer, and targeting tumor stroma under hypoxia might thus be potentially useful as a novel therapy for this cancer.

Runx3, a member of the human runt-related transcription factor family, is known as a possible tumor suppressor gene for gastric cancer. Runx 3 expression is frequently suppressed by the promoter hypermethylation in gastric cancer cell lines and tissues. However, the precise mechanism of the induction of Runx3 methylation, which is considered to be a critical step in gastric carcinogenesis, remains to be elucidated. In the present study, we evaluated runx3 gene methylation in 57 resected early gastric cancer specimens. Then, we correlated Runx3 methylation in the cancer tissue specimens with clinicopathological factors as well as the mucosal backgrounds, such as intestinal metaplasia surrounding the cancer cells and Helicobacter pylori (H. pylori) infection. Runx3 methylation was observed in 30 of the 57 (52.6%) cancer specimens, whereas methylation was detected in 10 of the 57 (17.5%) corresponding noncancerous mucosae. In comparison to the clinicopathological factors, Runx3 methylation was significantly correlated with both age and tumor location. A multivariate analysis demonstrated that age and tumor location as well as H. pylori infection were independent risk factors for Runx3 methylation. We demonstrated for the first time that H. pylori infection contributes to Runx3 methylation in gastric cancer tissues. When a persistent infection by H. pylori continues in the middle/lower stomach for a long period, Runx3 methylation may be induced and the subsequent loss of Runx3 expression may therefore affect gastric carcinogenesis.

Abstract. Tumor hypoxia has been reported to induce tumor progression in several carcinomas. Current studies have shown that hypoxia inducible factor-1· (HIF-1·) is stabilized under hypoxic conditions and transactivates various genes related to cancer aggressiveness. In the present study, we examined whether hypoxia affects cancer invasion in hepatocellular carcinoma. We aimed to solve the molecular mechanism of tumor invasion under the hypoxic condition. We showed that tumor hypoxia accelerated cancer invasion in two hepatoma cell lines. Using Western blot and RT-PCR analyses we demonstrated striking evidence that the expression of HIF-1·, ETS-1, MMP-7 and MT1-MMP was strongly upregulated by hypoxic stimulation. To examine whether these invasionrelated genes are regulated by HIF-1·, we treated hepatoma cells with TX-402, which was reported to repress HIF-1· expression. HIF-1· expression was strongly repressed by the TX-402 treatment. In contrast, the expression of ETS-1, MMP-7 and MT1-MMP mRNA was not affected by TX-402 treatment. We further established stable transfectants in which HIF-1· dominant negative vector was introduced into Hep3B cells (pHIF-1·DN). In the pHIF-1·DN cells, the expression of ETS-1, MMP-7 and MT1-MMP was not repressed. Moreover, the invasion activity of pHIF-1·DN was not altered, compared with that of the mock. In hepatoma cells, we provided evidence that hypoxic stress accelerates cancer invasion by upregulating ETS-1 and the MMP family by an HIF-1·-independent pathway.

The present study demonstrated for the first time that gastric cancer patients with p16INK4a methylation specifically benefit from 5-FU-based adjuvant chemotherapy.