title: Hypoxia targeted bifunctional suicide gene expression enhances radiotherapy in vitro and in vivo.
abstract: OBJECTIVE
  To investigate whether hypoxia targeted bifunctional suicide gene expression-cytosine deaminase (CD) and uracil phosphoribosyltransferase (UPRT) with 5-FC treatments can enhance radiotherapy.
  
  
  METHODS
  Stable transfectants of R3327-AT cells were established which express a triple-fusion-gene: CD, UPRT and monomoric DsRed (mDsRed) controlled by a hypoxia inducible promoter. Hypoxia-induced expression/function of CDUPRTmDsRed was verified by western blot, flow cytometry, fluorescent microscopy, and cytotoxicity assay of 5-FU and 5-FC. Tumor-bearing mice were treated with 5-FC and local radiation. Tumor volume was monitored and compared with those treated with 5-FC or radiation alone. In addition, the CDUPRTmDsRed distribution in hypoxic regions of tumor sections was visualized with fluorescent microscopy.
  
  
  RESULTS
  Hypoxic induction of CDUPRTmDsRed protein correlated with increased sensitivity to 5-FC and 5-FU. Significant radiosensitization effects were detected after 5-FC treatments under hypoxic conditions. In the tumor xenografts, the distribution of CDUPRTmDsRed expression visualized with fluorescence microscopy was co-localized with the hypoxia marker pimonidazole positive staining cells. Furthermore, administration of 5-FC to mice in combination with local irradiation resulted in significant tumor regression, as in comparison with 5-FC or radiation treatments alone.
  
  
  CONCLUSIONS
  Our data suggest that the hypoxia-inducible CDUPRT/5-FC gene therapy strategy has the ability to specifically target hypoxic cancer cells and significantly improve the tumor control in combination with radiotherapy.
authors.affiliation: Shandong Medical Imaging Research Institute, Shandong University, Jinan, China.
authors.forename: Xiaorong
authors.initials: X
authors.lastname: Sun
authors.forename: Ligang
authors.initials: L
authors.lastname: Xing
authors.forename: Xuelong
authors.initials: X
authors.lastname: Deng
authors.forename: Hung Tsung
authors.initials: HT
authors.lastname: Hsiao
authors.forename: Akiko
authors.initials: A
authors.lastname: Manami
authors.forename: Jason A
authors.initials: JA
authors.lastname: Koutcher
authors.forename: C
authors.initials: C
authors.lastname: Clifton Ling
authors.forename: Gloria C
authors.initials: GC
authors.lastname: Li
mesh_terms: D000818:Animals; D015153:Blotting, Western; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D003114:Colony-Forming Units Assay; D043525:Cytosine Deaminase / Q000235:genetics*; D005434:Flow Cytometry; D005437:Flucytosine / Q000627:therapeutic use*; D015870:Gene Expression; D041722:Genes, Transgenic, Suicide / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods; D066298:In Vitro Techniques; D051379:Mice; D008856:Microscopy, Fluorescence; D009374:Neoplasms, Experimental / Q000532:radiotherapy*; D010430:Pentosyltransferases / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011836:Radiation Tolerance / Q000187:drug effects; D014407:Tumor Cells, Cultured
chemical_list: D005437:Flucytosine; D010430:Pentosyltransferases; C023269:uracil phosphoribosyltransferase; D043525:Cytosine Deaminase