Takao Ide

Selective killing of cancer cells by a small molecule targeting the stress response to ROS

Malignant transformation, driven by gain-of-function mutations in oncogenes and loss-of-function mutations in tumour suppressor genes, results in cell deregulation that is frequently associated with enhanced cellular stress (for example, oxidative, replicative, metabolic and proteotoxic stress, and DNA damage). Adaptation to this stress phenotype is required for cancer cells to survive, and consequently cancer cells may become dependent upon non-oncogenes that do not ordinarily perform such a vital function in normal cells. Thus, targeting these non-oncogene dependencies in the context of a transformed genotype may result in a synthetic lethal interaction and the selective death of cancer cells. Here we used a cell-based small-molecule screening and quantitative proteomics approach that resulted in the unbiased identification of a small molecule that selectively kills cancer cells but not normal cells. Piperlongumine increases the level of reactive oxygen species (ROS) and apoptotic cell death in both cancer cells and normal cells engineered to have a cancer genotype, irrespective of p53 status, but it has little effect on either rapidly or slowly dividing primary normal cells. Significant antitumour effects are observed in piperlongumine-treated mouse xenograft tumour models, with no apparent toxicity in normal mice. Moreover, piperlongumine potently inhibits the growth of spontaneously formed malignant breast tumours and their associated metastases in mice. Our results demonstrate the ability of a small molecule to induce apoptosis selectively in cells that have a cancer genotype, by targeting a non-oncogene co-dependency acquired through the expression of the cancer genotype in response to transformation-induced oxidative stress.

Tumor-stromal cell interaction under hypoxia increases the invasiveness of pancreatic cancer cells through the hepatocyte growth factor/c-Met pathway.

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‐1α 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‐1α 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‐1α 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.

Induction of hepatocyte growth factor activator gene expression under hypoxia activates the hepatocyte growth factor/c-Met system via hypoxia inducible factor-1 in pancreatic cancer

Hepatocyte growth facor activator (HGFA) is a serine protease that converts hepatocyte growth factor (HGF) into its active form. Our previous study demonstrated that tumor–stromal interaction under hypoxia augments the aggressive invasive features of pancreatic cancer line PK8 through activated HGF/c‐Met signaling. The present study investigated whether or not hypoxia increases HGFA expression in PK8 cells and promotes the processing of HGF, and leads to c‐Met activation. Moreover, HGFA promoter assays were performed to define whether hypoxia inducible factor‐1 alpha (HIF‐1α) directly activates the HGFA promoter in a hypoxia‐dependent fashion. As a result, hypoxia induced the HGFA mRNA and protein expression in PK8 and the elevation under hypoxia was inhibited by the transfection of HIF‐1α siRNA, thus indicating HIF‐1α‐dependent induction of HGFA. The transfection of siRNA against HGFA to PK8 cells suppressed the conversion to the active HGF, which is secreted from fibroblast MRC5. Furthermore, the phosphorylation of c‐Met and cancer invasion of PK8 cells were decreased by the transfection of HGFA siRNA under hypoxia. Using the luciferase reporter system, HIF‐1α was shown to transactivate the HGFA promoter under hypoxia. These experiments demonstrated for the first time that HGFA is a novel HIF‐1 target gene. Under hypoxia, HGFA might be overexpressed and secreted from pancreatic cancer cells, which contributes to accelerate processing of HGF from fibroblast, resulting in the activation of the c‐Met pathway. HGF/HGFA/c‐Met recruited between cancer‐stromal fibroblasts is activated under hypoxic conditions and therefore might play a central role in the aggressive invasion of pancreatic cancer. (Cancer Sci 2008; 99: 1341–1347)

The Hypoxic Environment in Tumor-Stromal Cells Accelerates Pancreatic Cancer Progression via the Activation of Paracrine Hepatocyte Growth Factor/c-Met Signaling

BackgroundPancreatic cancer is one of the representative solid tumors, in which the hypoxic microenvironment plays a crucial role in malignant progression. We previously demonstrated that tumor-stromal interaction under hypoxia enhances the invasiveness of pancreatic cancer cells through hepatocyte growth factor (HGF)/c-Met signaling.MethodsWe investigated the immunohistochemical expression of hypoxia inducible factor-1α (HIF-1α) c-Met, and HGF in both cancer and stromal cells using 41 pancreatic cancer tissue specimens, and tried to identify any correlations with the clinical features and survival.ResultsPositive staining for HIF-1α was observed in both pancreatic cancer and the surrounding stromal cells in more than 30% of the cases, and it significantly correlated with lymph node metastasis (P < .05). A significant correlation was observed between the expression of HIF-1α and HGF in stromal cells (P < .05). In addition, the c-Met expression in cancer cells was found to significantly correlate with the HGF expression in not only cancer but also stromal cells. The disease-free survival rates of the patients with HIF-1α in cancer, stromal, c-Met in cancer, and an HGF expression in stromal cells was significantly worse than those without such expressions (P < .05).ConclusionsThese data suggest that the HGF/c-Met signaling via HIF-1α ?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.

Aberrant methylation of p16 predicts candidates for 5-fluorouracil-based adjuvant therapy in gastric cancer patients

BackgroundAberrant methylation of some cancer-related genes has been reported to correlate with sensitivity to chemotherapeutic agents. The present study was designed to determine whether DNA methylation in six cancer-related genes affects recurrence of gastric cancer in patients who received 5-fluorouracil-based adjuvant chemotherapy.MethodsThe methylation status of six genes, MGMT, CHFR, hMLH1, p16INK4a, E-cadherin, and Runx3, was analyzed in 56 surgically resected gastric cancer tissue specimens by methylation-specific polymerase chain reaction. Of the 56 patients who underwent surgical resection, 38 received 5-fluorouracil (5-FU)-based adjuvant chemotherapy postoperatively (adjuvant group), whereas the other 18 (32%) did not (surgery group).ResultsThere were no significant differences between the two groups with respect to sex, cancer differentiation, depth of tumor invasion, lymph node metastasis, lymphatic invasion, vascular invasion and tumor stage. Among the genes, methylation of p16INK4a showed a significant correlation with longer survival in the 38 patients of the adjuvant group, but not in the 18 patients of the surgery group. A multivariate analysis identified p16INK4a methylation to be an independent factor predicting a longer recurrence-free period under 5-FU-based adjuvant chemotherapy.ConclusionsThe present study demonstrated for the first time that gastric cancer patients with p16INK4a methylation specifically benefit from 5-FU-based adjuvant chemotherapy.

Prediction of postoperative complications in elderly patients with hepatocellular carcinoma

BACKGROUND The aim of the present study was to investigate whether advanced age was associated with a higher rate of postoperative complications and identify the predictive factors for postoperative complications in elderly patients with hepatocellular carcinoma (HCC). METHODS Between January 2000 and December 2010, 256 patients who underwent hepatectomy for HCC were investigated. Elderly patients were defined as those aged ≥75 y. The clinicopathologic data and outcomes after hepatectomy for 64 elderly and 192 younger patients were retrospectively collected and compared. RESULTS There were no significant differences in the incidence of postoperative complications (P = 0.936) or the long-term survival after hepatectomy (P = 0.641) between the elderly and younger patients. In multivariate analysis, the estimation of physiological ability and surgical stress-preoperative risk score (PRS) was an independent risk factor for postoperative morbidity in the elderly patients (P < 0.01). Moreover, the patients were analyzed according to the PRS for the assessment of their general preoperative condition and liver damage grade based on the hepatic reserve. The rate of postoperative complications in the patients with a PRS ≥0.5 and liver damage B was significantly higher in the elderly patients (P < 0.01), whereas a PRS and liver damage grade did not affect the incidence of postoperative morbidity in the younger patients (P = 0.516). CONCLUSIONS Hepatectomy for elderly patients with HCC is feasible as well as safe, and the preoperative assessment using the estimation of physiological ability and surgical stress scoring system, combined with the liver damage grade, can help to improve the safety of this procedure for elderly HCC patients.

Hepatoid carcinoma of the pancreas penetrating into the gastric cavity: A case report and literature review

A 79‐year‐old Japanese woman was admitted to our hospital for treatment of a pancreatic tumor measuring approximately 7 × 5 cm. The tumor had invaded the left adrenal gland and gastric wall and had penetrated into the gastric cavity. Surgical resection was performed. The tumor was composed of a brown to whitish solid area and a zone of hemorrhage, necrosis, and cystic degeneration resembling the gross features of solid pseudopapillary tumor (SPT). Histologically, the tumor showed a heterogeneous growth pattern with a combination of seat‐like, trabecular, papillary and hemorrhagic‐necrotic areas in various proportions. The differential diagnoses first considered were acinar cell carcinoma, neuroendocrine carcinoma and SPT with malignant transformation. Immunohistochemistry showed tumor cells were negative for pancreatic exocrine enzymes and endocrine markers. Tumor cells diffusely expressed cytokeratin 19, alpha‐fetoprotein, carcinoembryonic antigen and glypican‐3, but lacked vimentin or β‐catenin expression. Small proportions of tumor cells expressed hepatocyte paraffin‐1. Although typical morphological features of well‐differentiated hepatocellular carcinoma (HCC) were not distinctly apparent, the tumor morphology partly resembled poorly differentiated HCC. Given these findings and considerations, the tumor was finally diagnosed as poorly differentiated hepatoid carcinoma of the pancreas.

Clinicopathologic features of advanced gallbladder cancer associated with adenomyomatosis

Adenomyomatosis of the gallbladder has not been considered to have malignant potential, but gross features of adenomyomatosis are sometimes encountered in gallbladders resected under a diagnosis of gallbladder carcinoma. The purpose of this study was to determine the clinicopathologic features and survival rates in cases of gallbladder cancer with gross features of adenomyomatosis. The study subjects were 97 surgically treated gallbladder carcinoma patients. Only gallbladder showing typical gross features of adenomyomatosis was judged as adenomyomatosis-positive gallbladder cancer. In terms of gross findings, 25 cases (25.8%) were classified as adenomyomatosis-positive. The status of adenomyomatosis was significantly associated with the T stage (P = 0.0004), lymph node (LN) metastasis (P < 0.0001), distant metastasis (P = 0.008), and stage (P = 0.0005). In the adenomyomatosis-positive group, 16 of the 25 cases (64.0%) were classified as segmental type and 9 cases (36.0%) were classified as fundal type. No diffuse-type cases were present in this series. The status of adenomyomatosis correlated significantly with survival (P = 0.0007). However, the multivariate analysis of significant variables identified from the univariate analysis identified only T stage (P = 0.0178) and LN metastasis (P = 0.0048) as independent prognostic factors. Subset analysis with T stage according to the status of adenomyomatosis showed no significant impact on survival. These results indicate that adenomyomatosis-positive gallbladder cancer is more often diagnosed clinically in the advanced stages. Since preceding adenomyomatosis may prevent the early detection of gallbladder cancer, the usefulness of preventive cholecystectomy in cases of asymptomatic adenomyomatosis should be considered.

Loss of trefoil factor 1 is regulated by DNA methylation and is an independent predictive factor for poor survival in advanced gastric cancer

Trefoil factor 1 (TFF1) is considered to be a tumor suppressor gene in gastric cancer. However, the role of TFF1 expression and its regulation in gastric cancer patients remain unclear. The aims of this study were to clarify the clinical significance of TFF1 and to determine its regulatory mechanisms. We assessed the immunohistochemical expression of TFF1 in 182 gastric cancer patients and examined whether or not TFF1 is associated with the clinicopathological factors and patient survival. In vitro study using TFF1 knockdown gastric cancer cells evaluated the role of TFF1 in cancer invasion. Bisulfite sequencing was performed to assess DNA methylation of TFF1 in cells and resected tissues. Patients with low expression of TFF1 showed a significantly deeper invasion of the tumor than those with high expression (p=0.037). Low expression of TFF1 was also associated with a poor survival (p=0.029) in 108 patients who were treated by surgery alone. Both TFF1 expression and lymph node metastasis are independent predictive factors for disease-specific survival in a multivariate analysis. In an in vitro study, invasive power of the cells was significantly increased in the TFF1‑deficient cells compared with the control cells. Bisulfate sequencing showed that TFF1 expression is strongly dependent on DNA methylation in both gastric cancer cells and tissues. Interestingly, methylation status of two specific CpG sites, which are located close to a TATA box and hypoxia response element (HRE), determined the TFF1 expression in the resected tissues. TFF1 expression is silenced by DNA methylation and is associated with tumor invasion and a poor survival in gastric cancer patients. The expression and̸or methylation status of TFF1 may, therefore, serve as a useful biomarker for predicting survival in patients with advanced gastric cancer.

Corrigendum: Selective killing of cancer cells by a small molecule targeting the stress response to ROS.

This corrects the article DOI: 10.1038/nature10167