| Abstract: |
Tumor hypoxia is important in the development and treatment of human cancers. We have developed a novel xenograft model for studying and imaging of hypoxia-induced gene expression. A hypoxia-inducible dual reporter herpes simplex virus type 1 thymidine kinase and enhanced green fluorescence protein (HSV1-TKeGFP), under the control of hypoxia response element (9HRE), was stably transfected into human colorectal HT29 cancer cells. Selected clones were further enriched by repeated live cell sorting gated for hypoxia-induced eGFP expression. Fluorescent microscopy, fluorescence-activated cell sorting, and radioactive substrate trapping assays showed strong hypoxia-induced expression of eGFP and HSV1-tk enzyme in the HT29-9HRE cells in vitro. Sequential micropositron emission tomography (PET) imaging of tumor-bearing animals, using the hypoxic cell tracer 18F-FMISO and the reporter substrate 124I-FIAU, yielded similar tumor hypoxia images for the HT29-9HRE xenograft but not in the parental HT29 tumor. Using autoradiography and IHC, detailed spatial distributions in tumor sections were obtained and compared for the following hypoxia-associated biomarkers in the HT29-9HRE xenograft: 124I-FIAU, 18F-FMISO, Hoechst (perfusion), lectin-TRITC ( functional blood vessels), eGFP, pimonidazole, EF5, and CA9. Intratumoral distributions of 124I-FIAU and 18F-FMISO were similar, and eGFP, pimonidazole, EF5, and CA9 colocalized in the same areas but not in well-perfused regions that were positive for Hoechst and lectin-TRITC. In enabling the detection of hypoxia-induced molecular events and mapping their distribution in vivo with serial noninvasive positron emission tomography imaging, and multiple variable analysis with immunohistochemistry and fluorescence microscopy, this human xenograft model provides a valuable tool for studying tumor hypoxia and in validating existing and future exogenous markers for tumor hypoxia. ©2008 American Association for Cancer Research. |
| Keywords: |
immunohistochemistry; controlled study; protein expression; human cell; nonhuman; positron emission tomography; neoplasms; biological markers; mouse; animals; mice; gene expression; animal experiment; tumor xenograft; molecular imaging; cancer model; hypoxia; disease model; genetic transfection; antigens, neoplasm; carbonate dehydratase ix; tissue distribution; mice, nude; positron-emission tomography; arabinofuranosyluracil; transplantation, heterologous; green fluorescent proteins; fluorescence microscopy; 1 fluoro 3 (2 nitro 1 imidazolyl) 2 propanol f 18; fialuridine; iodine 124; pimonidazole; thymidine kinase; cell hypoxia; herpesvirus 1, human; misonidazole; neoplasm transplantation; cell clone; dna responsive element; herpes simplex virus 1; cell strain ht29; ht29 cells; autoradiography; nitroimidazoles; cell selection; 2 (2 nitro 1h imidazol 1 yl) n (2,2,3,3,3 pentafluoropropyl)acetamide; carbonic anhydrases; etanidazole; hydrocarbons, fluorinated; enhanced green fluorescent protein; lectin; tetramethylrhodamine
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