Protein disulfide isomerase (PDI), an endoplasmic reticulum chaperone protein, catalyzes disulfide bond breakage, formation, and rearrangement. The effect of PDI inhibition on ovarian cancer progression is not yet clear, and there is a need for potent, selective, and safe small-molecule inhibitors of PDI. Here, we report a class of propynoic acid carbamoyl methyl amides (PACMAs) that are active against a panel of human ovarian cancer cell lines. Using fluorescent derivatives, 2D gel electrophoresis, and MS, we established that PACMA 31, one of the most active analogs, acts as an irreversible small-molecule inhibitor of PDI, forming a covalent bond with the active site cysteines of PDI. We also showed that PDI activity is essential for the survival and proliferation of human ovarian cancer cells. In vivo, PACMA 31 showed tumor targeting ability and significantly suppressed ovarian tumor growth without causing toxicity to normal tissues. These irreversible small-molecule PDI inhibitors represent an important approach for the development of targeted anticancer agents for ovarian cancer therapy, and they can also serve as useful probes for investigating the biology of PDIimplicated pathways.oral bioavailability | drug resistance | BODIPY-conjugation

BackgroundElucidation of the molecular mechanisms by which cancer cells overcome hypoxia is potentially important for targeted therapy. Complexation of hypoxia-inducible factors (HIFs) with aryl hydrocarbon receptor nuclear translocators can enhance gene expression and initiate cellular responses to hypoxia. However, multiple molecular mechanisms may be required for cancer cells to adapt to diverse microenvironments. We previously demonstrated that a physical interaction between the ubiquitously expressed transcription factor Sp1 and HIF2 is a major cause of FVII gene activation in poor prognostic ovarian clear cell carcinoma (CCC) cells under hypoxia. Furthermore, it was found that FVII activation is synergistically enhanced when serum-starved cells are cultured under hypoxic conditions. In this study, we investigated whether HIFs and transcription factor Sp1 cooperate to activate multiple genes in CCC cells under conditions of serum starvation and hypoxia (SSH) and then contribute to malignant phenotypes.MethodsTo identify genes activated under hypoxic conditions in an Sp1-dependent manner, we first performed cDNA microarray analyses. We further investigated the molecular mechanisms of synergistic gene activations including the associated serum factors by various experiments such as real-time RT-PCR, western blotting and chromatin immunoprecipitation. The study was further extended to animal experiments to investigate how it contributes to CCC progression in vivo.ResultsICAM1 is one such gene dramatically induced by SSH and is highly induced by SSH and its synergistic activation involves both the mTOR and autonomously activated TNFα-NFκB axes. We identified long chain fatty acids (LCFA) as a major class of lipids that is associated with albumin, a serum factor responsible for synergistic gene activation under SSH. Furthermore, we found that ICAM1 can be induced in vivo to promote tumor growth.ConclusionSp1 and HIFs collaborate to activate genes required for the adaptation of CCC cells to severe microenvironments, such as LCFA starvation and hypoxia. This study highlights the importance of transcriptional regulation under lipid starvation and hypoxia in the promotion of CCC tumor growth.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0351-z) contains supplementary material, which is available to authorized users.

Background and objectives We clarified the impact of omentectomy for advanced gastric cancer on patient survival from the surgical results of a high-volume center in Japan.Methods Patients who received curative gastrectomy were divided into two groups based on whether they underwent omentectomy. The propensity score-matching method was used to assemble a well-balanced cohort, and relapse-free survival and the pattern of recurrence were compared. Results For this study, 330 patients who fulfilled the inclusion criteria participated and were divided into two groups: group R, patients who received omentectomy, and group P, patients who received omentum-preserving gastrectomy. After performing score-matching, 196 patients were selected. The 3-and 5-year relapse-free survival rates were 72.9 % (95 % confidence interval, 64.1-81.7) and 66.2 % (56.6-75.8 %) in group R, and 76.7 % (67.9-81.2) and 67.3 % (55.1-79.5) in group P, which were not significantly different (P = 0.750). Regarding sites of relapses, no differences were observed between the groups (P = 0.863). ConclusionsIn this series, omentum-preserving gastrectomy for advanced gastric cancer did not increase the peritoneal relapse rate or affect patient survival compared to conventional gastrectomy. The non-inferiority of the omission of omentectomy should be evaluated by a randomized controlled trial.

Abstract. Claudins, members of a large family of adherent junction proteins, regulate the integrity and function of tight junctions and influence tumorigenesis. Studies have suggested that altered levels of different claudins are related to carcinoma-cell invasion and disease progression. This study examined the relationship between the relative expression of claudin genes and clinicopathological factors, especially invasion and metastasis, in patients with colorectal cancer. We studied surgical specimens of cancer tissue and adjacent normal mucosa from 205 patients with untreated colorectal carcinoma. The relative expression levels of claudin-1, -3, -4 and -7 mRNA in cancer and in normal adjacent mucosa were measured by quantitative real-time, reverse-transcription polymerase chain reaction. The relative expression levels of the claudin-1, -3 and -4 genes were higher in cancer than in normal adjacent mucosa, whereas the relative expression of the claudin-7 gene was similar. An analysis of the relationship between the clinicopathological features and gene expression showed that reduced expression of claudin-7 correlated with venous invasion and liver metastasis. There was also a correlation between claudin-3 and -4 gene expression. Our results suggested that a reduced expression of the claudin-7 gene might lead to venous invasion and liver metastasis in colorectal cancer. Reduced expression of the claudin-7 gene may thus be a useful predictor of liver metastasis in patients with colorectal cancer.

Viable and stable human cancer cell lines and animal models combined with adequate clinical information are essential for future advances in cancer research and patient care. Conventional in vitro cancer cell lines are commonly available; however, they lack detailed information on the patient from which they originate, including disease phenotype and drug sensitivity. Patient-derived xenografts (PDX) with clinical information (so-called ‘cancer xenopatients’) are a promising advance that may accelerate the development of anticancer therapies. We established 61 PDX lines from 116 surgically removed tumor tissues inoculated subcutaneously into NOG mice (53% success rate). PDX lines were established from various types of epithelial tumors and also from sarcomas, including gastrointestinal stromal tumors and Ewing/PNET sarcomas. The metastatic tumors yielded PDX lines more effectively (65%) than the primary tumors (27%, P<0.001). In our PDX models, morphological characteristics, gene expression profiles, and genetic alteration patterns were all well preserved. In eight cases (7%), the transplantable xenografts for several generations were composed of large monotonous nonepithelial cells of human origin, revealed to be Epstein-Barr virus infection-associated lymphoproliferative lesions. Despite this, PDX linked with clinical information offer many advantages for preclinical studies investigating new anticancer drugs. The fast and efficient establishment of individual PDX may also contribute to future personalized anticancer therapies.

Ovarian clear cell adenocarcinoma (CCC) is the second most common subtype of ovarian cancer after high-grade serous adenocarcinomas. CCC tends to develop resistance to the standard platinum-based chemotherapy, and has a poor prognosis when diagnosed in advanced stages. The ANXA4 gene, along with its product, a Ca++-binding annexin A4 (ANXA4) protein, has been identified as the CCC signature gene. We reported two subtypes of ANXA4 with different isoelectric points (IEPs) that are upregulated in CCC cell lines. Although several in vitro investigations have shown ANXA4 to be involved in cancer cell proliferation, chemoresistance, and migration, these studies were generally based on its overexpression in cells other than CCC. To elucidate the function of the ANXA4 in CCC cells, we established CCC cell lines whose ANXA4 expressions are stably knocked down. Two parental cells were used: OVTOKO contains almost exclusively an acidic subtype of ANXA4, and OVISE contains predominantly a basic subtype but also a detectable acidic subtype. ANXA4 knockdown (KO) resulted in significant growth retardation and greater sensitivity to carboplatin in OVTOKO cells. ANXA4-KO caused significant loss of migration and invasion capability in OVISE cells, but this effect was not seen in OVTOKO cells. We failed to find the cause of the different IEPs of ANXA4, but confirmed that the two subtypes are found in clinical CCC samples in ratios that vary by patient. Further investigation to clarify the mechanism that produces the subtypes is needed to clarify the function of ANXA4 in CCC, and might allow stratification and improved treatment strategies for patients with CCC.

Abstract. Expression of the fibroblast growth factor (FGF)-1, FGF-2, fibroblast growth factor receptor (FGFR)-1, andFGFR-2 genes has been reported in various cancers and is associated with poor outcomes in patients with solid tumors. This study examined the relations between the relative expression of the FGF genes and clinicopathological factors, especially invasion and metastasis, in patients with colorectal cancer. We studied surgical specimens of cancer tissue and adjacent normal mucosa obtained from 202 patients with untreated colorectal carcinoma. The relative expression levels of FGF-1, FGF-2, FGFR-1, and FGFR-2 mRNA in cancer and in normal adjacent mucosa were measured by quantitative real-time, reverse-transcription polymerase chain reaction. The relative expression level of the FGFR-2 gene was higher in normal adjacent mucosa than in cancer, whereas the relative expression levels of the FGF-1, FGF-2, and FGFR-1 genes were similar. FGFR-1 gene expression levels were higher in the presence than in the absence of liver metastasis. An analysis of the relation between clinicopathological features and gene expression showed that overexpression of FGFR-1 correlated with liver metastasis. Our results suggested that overexpression of the FGFR-1 gene might lead to liver metastasis in colorectal cancer. Overexpression of the FGFR-1 gene may thus be a useful predictor of liver metastasis in patients with colorectal cancer. IntroductionFibroblast growth factors (FGFs) are a family of heparinbinding growth factors. FGFs promote angiogenesis by interacting with various endothelial cell-surface receptors, including tyrosine kinase receptors, heparin-sulfate proteoglycans, and integrins. The relation between angiogenesis and tumor growth is well established, and numerous inducers of angiogenesis have been identified (1). Gospodarowicz (2) discovered FGF-2 in 1974. This protein was found to strongly promote the proliferation of fibroblasts. Since then, 22 structurally-related members of the FGF family and 4 FGFhomologous factors have been identified. FGFs exert their biological activities by binding to high-affinity tyrosine kinase FGF receptors (FGFRs) on the surface of target cells. Angiogenic potential has been assessed for only a limited number of the 22 members of the FGF family in vitro and in vivo. Most experimental studies have focused on the prototypes FGF-1 and FGF-2 (3).FGFs exert their biologic activity by interacting with highaffinity FGFRs. Four members of the FGFR family (FGFR-1, FGFR-2, FGFR-3, and FGFR-4) are encoded by distinct genes, and their structural variability is increased by alternative splicing. FGFR-1 is expressed by endothelial cells in vivo and in vitro. Some cultured endothelial cells can express FGFR-2 (3,4).Expression levels of the FGF-1, FGF-2, FGFR-1, and FGFR-2 genes have been examined in various cancers, including breast cancer (5-8), brain tumors (9-12), hepatocellular carcinoma (13-15), cervical and esophageal cancers (16-20), and pancreatic cancer (21-23). Correlations between FGF...

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