2-(2-nitro-1h-imidazol-1-yl)-n-(2-2-3-3-3-pentafluoropropyl)acetamide and Fibrosarcoma

In the framework of the preclinical validation of the hypoxic tracer [(18)F]EF3, a comparison was performed between uptake of [(18)F]EF3 and EF5 adducts detected by immunofluorescence in MCa-4, FSA, FSAII, Sa-NH and NFSA tumour-bearing mice. Mice were allowed to breath carbogen (5% CO(2), 95% O(2)), 21% oxygen or 10% oxygen. A significant correlation (r (2)=0.57; p<0.01) was found between the [(18)F]EF3 tumour-to-muscle ratio and the fluorescence intensity of EF5.

Topics: Animals; Cell Hypoxia; Drug Evaluation, Preclinical; Etanidazole; Fibrosarcoma; Fluorescent Antibody Technique; Hydrocarbons, Fluorinated; Male; Mammary Neoplasms, Experimental; Metabolic Clearance Rate; Mice; Mice, Inbred C3H; Nitroimidazoles; Organ Specificity; Oxygen; Radionuclide Imaging; Radiopharmaceuticals; Tissue Distribution

The primary objective of this study was to establish in vivo the relationship between 2-2-nitro-1H-imidazol-1yl-N-(2,2,3,3,3-pentafluoropropyl)-acetamide (EF5) adduct formation and intratumoral oxygen concentrations measured by electron paramagnetic resonance (EPR) in a tumor model mimicking a clinical situation. The secondary objective was an attempt to calibrate in situ the immunofluorescence (IF) signal with EPR oximetry.. IM syngeneic fibrosarcoma (NFSA) bearing C3H mice were used. Three days after injection of a paramagnetic charcoal into the tumor, the mice were anesthetized, injected with the hypoxic marker EF5, and monitored every 20 min for 3 h with a low-frequency EPR spectrometer. Animals were allowed to breath either under 21 or 100% O(2). Tumors were then harvested, frozen, cut into sections including the charcoal and processed for EF5 adducts detection using monoclonal antibodies. Slices were viewed with a fluorescence microscope and 190x140 micrometer areas surrounding the charcoal were digitized and analyzed with the NIH-Image and Adobe Photoshop software. The fluorescence intensity (FI) was measured in the whole pictures and in strips of 10 micrometer around the charcoal.. EF5 binding increased with decreasing pO(2), most substantially at pO(2) below 5 mm Hg. Baseline (ambient air) pO(2) reached 3.2+/-2.1 mm Hg in NFSA tumors. It increased to 9.8+/-3.2 mm Hg under 100% O(2). A statistically significant correlation was observed on an individual tumor basis between the FI in the first 10 micrometer strip around the charcoal and the pO(2) determined by EPR oximetry (Wilcoxon signed rank test: P<0.001).. The present study confirms the intrinsic relationship between EF5 adduct binding and intratumoral pO(2) in an in vivo environment under biologically-relevant pO(2) values of less than 10 mm Hg.

Topics: Algorithms; Animals; Cell Hypoxia; Charcoal; Computer Graphics; Disease Models, Animal; Electron Spin Resonance Spectroscopy; Etanidazole; Fibrosarcoma; Hydrocarbons, Fluorinated; Male; Mice; Mice, Inbred C3H; Microscopy, Fluorescence; Muscle Neoplasms; Oximetry; Oxygen; Radiation-Sensitizing Agents; Statistics, Nonparametric

Since tissue oxygen tension is a balance between delivery and consumption of oxygen, considerable effort has been directed at increasing the former and/or decreasing the latter. Techniques to decrease the rate of cellular oxygen consumption (increasing the distance oxygen can diffuse into tissues) include increasing glycolysis by administering supra-physiologic levels of glucose. We have examined the effect of hyperglycemia produced by intravenous glucose infusion on the tissue oxygenation and radiation response of subcutaneously implanted murine radiation induced fibrosarcomas (RIF-1). A 0.3 M glucose solution was delivered via tail vein injection according to a protocol that maintained glucose at a plasma concentration of 17+/-1 mM. The effect of this treatment on radiation response (clonogenic and growth delay studies), tumor oxygenation (needle electrode pO2 and 2-[2-nitro-1H-imidazol-1-yl]-N-(2,2,3,3,3-pentafluoropropyl) acetamide (EF5) binding), and tumor bioenergetics and pH (31P NMR spectroscopy) was examined. Systemic measurements included hematocrit and blood glucose and lactate concentrations. The results of these studies suggest that these subcutaneously implanted RIF-1 tumors are both radiobiologically and metabolically hypoxic and that intravenous glucose infusion is not an effective method of modifying this metabolic state.

Topics: Animals; Cell Division; Energy Metabolism; Etanidazole; Female; Fibrosarcoma; Flow Cytometry; Glucose; Hematocrit; Hydrocarbons, Fluorinated; Hyperglycemia; Magnetic Resonance Spectroscopy; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Neoplasms, Radiation-Induced; Oxygen Consumption; Radiation Tolerance; Radiation-Sensitizing Agents; Sarcoma, Experimental; Survival Rate

To examine the effects of hydralazine on vascular perfusion and hypoxia in spontaneous vs. first generation and long-term transplanted murine tumor models.. Total anatomic blood vessels were quantified using image analysis of CD31 stained frozen sections, perfused vessels by i.v. injection of fluorescent DiOC(7), and tumor hypoxia was measured using the EF5 hypoxia marker. KHT sarcomas, spontaneous mammary carcinomas, and first generation transplants of the spontaneous tumors were evaluated before and after i.p. administration of 5 mg/kg hydralazine.. Although anatomic and perfused vessel spacings were similar among untreated tumors, response to hydralazine varied widely among the three tumor models. In KHT tumors, perfused vessel numbers decreased significantly at 30 min post-hydralazine, then recovered somewhat by 60 min. First-generation transplants showed a less substantial decrease in perfused vessels following hydralazine, which tapered off slightly by 60 min. Finally, spontaneous tumors had only a modest decrease in perfused vessel numbers, with complete recovery at 60 min. Although response of individual tumors varied widely, overall hypoxic marker uptake was significantly increased in both KHT and first generation tumors, and slightly reduced in the spontaneous tumors.. Response to hydralazine varies substantially between transplanted and spontaneous tumor models. Results suggest that increased tumor pressure may be a critical factor in tumor response to hydralazine, possibly explaining tumor volume dependent variations.

Topics: Animals; Carbocyanines; Cell Hypoxia; Etanidazole; Female; Fibrosarcoma; Fluorescent Dyes; Hydralazine; Hydrocarbons, Fluorinated; Indicators and Reagents; Mammary Neoplasms, Experimental; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Neoplasms; Oxygen; Radiobiology; Tumor Cells, Cultured; Vasodilator Agents

Numerous experimental and clinical studies have been completed regarding the effects of carbogen and nicotinamide on tumor oxygenation and radiosensitivity. The current study incorporates three physiological measurement techniques to further define spatial variations in oxygen availability and development of hypoxia after single- and multifraction irradiation in KHT murine fibrosarcomas. Distances to anatomical and perfused blood vessels were measured using immunohistochemical and fluorescent staining, intravascular oxygen levels were determined cryospectrophotometrically, and tumor hypoxia was quantified using uptake of EF5, a marker of hypoxia. Carbogen, nicotinamide, and the combination of both all increased intravascular oxygen availability compared to controls. While nicotinamide had no effect on the number of perfused blood vessels in nonirradiated tumors, carbogen produced a substantial closing of vessels. After a single dose of 4 Gy, only the combination of nicotinamide and carbogen produced significant improvements in oxygen availability, while numbers of perfused vessels were significantly increased for nicotinamide, unchanged for the combination of nicotinamide and carbogen, and significantly decreased for carbogen. After 4 x 4-Gy fractions, oxygen availability was increased substantially with the combination of nicotinamide and carbogen, somewhat with carbogen, and not at all with nicotinamide. Tumor oxygenation changes were estimated by EF5/Cy3 intensity distributions, which demonstrated that manipulative agents could produce disparate effects on tumor hypoxia when combined with either single- or multifraction irradiation.

Topics: Animals; Carbocyanines; Carbon Dioxide; Cell Hypoxia; Combined Modality Therapy; Dose Fractionation, Radiation; Etanidazole; Female; Fibrosarcoma; Fluorescent Dyes; Hydrocarbons, Fluorinated; Indicators and Reagents; Mice; Mice, Inbred C3H; Niacinamide; Oxygen; Oxygen Consumption; Radiation-Sensitizing Agents; Tumor Cells, Cultured