Both tolerance to nutrient starvation and angiogenesis are essential for cancer progression because of the insufficient supply of nutrients to tumor tissue. Since chronic nutrient starvation seldom occurs in normal tissue, cancer's tolerance to nutrient starvation should provide a novel target for cancer therapy. In this study, we propose an anti-austerity strategy to exploit the ability of agents to eliminate cancer cells' tolerance to nutrient starvation. We established a simple screening method for agents that inhibit cancer cell viability preferentially during nutrient starvation, using PANC-1 cell line cultured in nutrient-rich and nutrientdeprived media. ancers always have deformed, deficient vascular and lymphatic systems due to excessive and unregulated tumor cell growth, as well as improper and insufficient angiogenesis. As a result, perfusion within cancers is inadequate, and the cancers contain regions that are transiently and chronically exposed to nutrient starvation. 1) These microenvironmental inadequacies are present from the earliest stage in the development of cancers, and are fully established while the neoplasm is still microscopic in size. 2) Since nutrient starvation occurs in tumor tissue that is >100-200 µm away from a functional blood supply, 3) tumor survival depends, in part, on the ability to recruit new blood microvessels via angiogenic factors. This affords tumor cells the ability to survive and propagate in a hostile environment, 4) and thus increased angiogenesis by a malignancy is related to a poor prognosis. 5,6) Because of this, antiangiogenic therapy was postulated to be the most promising and specific approach to cancer therapy. Already, extensive studies have been conducted in an attempt to prevent tumor angiogenesis.However, Yu et al. recently suggested that tumors may be able to elude the effect of angiogenesis inhibitors, since antiangiogenic therapy targets genetically stable endothelial cells in the tumor vasculature, and genetic alterations that decrease the vascular dependence of tumor cells can influence the response of tumors to this therapy. 7) The well-known observation that hypovascularity is an outstanding characteristic of pancreatic cancers in diagnostic images suggests that pancreatic cancer tissue has a poor blood supply. 8) However, since pancreatic cancers are known to be among the most aggressive malignancies despite their poor blood supply, we proposed that cancer cells' tolerance of insufficient nutrient might be an important determinant of malignancy. 9) Although the mechanism of tolerance has not yet been fully elucidated, the PI-3K-Akt pathway and AMPK have been found to be involved. 10) Because the blood supply of normal tissue is regulated in a sophisticated manner, angiogenesis is seldom activated in normal healthy adult tissues, except in the female reproductive system. 11) The same is true for nutrient starvation. We therefore speculated that if some compound could abolish cancer cells' tolerance of nutrient starvation, it might be capable of be...

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Regions of hypoxia are present in the majority of solid tumors mainly because of poor and defective vascularization.1,2) Evidence indicating that hypoxic cells exist in the tumors has been obtained from microscopic analysis of histological sections 3,4) and radiobiological studies. 5) Oxygen is required for the cytotoxic effects of many cancer chemotherapeutic drugs and radiation, and hypoxic tumor cells are resistant to the conventional chemotherapy and radiotherapy. 6)Moreover, hypoxia also selects for more aggressive and metastatic cancer phenotypes that are associated with poor prognosis. 7)To conquer the therapeutic resistance induced by hypoxia, several compounds have been developed as prodrugs that are preferentially bioactivated in the hypoxic cells. The most advanced bioreductive drug, tirapazamine, is in phase III clinical trials in combination with cisplatin. 8)To identify a novel hypoxia-selective cytotoxin, we screened 20000 cultured broths of microorganisms and found that rakicidin A showed significant hypoxia-selective cytotoxicity. Rakicidin A was approximately 17.5-fold more cytotoxic under hypoxic than under normoxic conditions. In this paper, we describe the hypoxia-selective cytotoxicity of rakicidin A. MATERIALS AND METHODSCell Culture CHO chinese hamster ovary cells were maintained in minimum essential medium alpha medium (a-MEM) supplemented with 10% fetal bovine serum (ICN Biomedicals, Costa Mesa, CA, U.S.A.) and kanamycin (50 mg/ml). Human colorectal adenocarcinoma cell lines HCT-8 and DLD-1, human glioblastoma cell lines U251 and SF268, human prostate cancer cell lines PC-3 and LNCaP, human pancreatic adenocarcinomas cell line PANC-1 were maintained in RPMI1640 with 10% fetal bovine serum and kanamycin (50 mg/ml). A stable transformant of CHO cells was established by the transfection of the HIF-1-dependent luciferase (5ϫHRE/pGL3/VEGF/E1b) and neomycin resistant genes as previously described.9) Stably transfected cells were maintained in a-MEM supplemented with 10% fetal bovine serum and kanamycin (50 mg/ml). Cells were cultured at 37°C in a humidified incubator containing 5% CO 2 (normoxia).Materials Materials used were as follows: [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] (MTT), CoCl 2 , actinomycin D, bleomycin, camptothecin, chlorambucil, doxorubicin, etoposide, melphalan, mitomycin C, streptonigrin, taxol and tubercidin from Sigma Aldrich, St. Louis, U.S.A.; cisplatin from BioVision, California, U.S.A.; neocarzinostatin from Astellas Pharma, Inc., Japan; glutathione from Cayman Chemical Company, Michigan, U.S.A.; a-tocopherol and N-acetyl-L-cysteine from LKT Laboratories, Inc., Minnesota, U.S.A. All reagents were initially dissolved in dimethyl sulfoxide (DMSO) (Sigma) and diluted with the medium for assay.Screening CHO cells were seeded at a density of 1ϫ10 4 /100 ml medium per well into a pair of 96-well microplates and cultured for 24 h to assure complete adherence of the cells to the plates. The cultured cells were treated with fermentation broths of microor...

Pepstatin is shown to be a specific inhibitor of acid proteases. It also inhibits humangastricsin, but the effect is weaker than against humanpepsin. The content of pepsin and gastricsin in gastric juice of stomach ulcer patients is described and the pepstatin which remains in gastric juice collected at 60 minutes after its administration is also determined. These results and those obtained by the direct measurements of the peptic activity of the gastric juice indicate that a sufficient amount of pepstatin to inhibit pepsin remains in the