Non-invasive molecular and functional imaging of cytosine deaminase and uracil phosphoribosyltransferase fused with red fluorescence protein Journal Article

Authors: Xing, L.; Deng, X.; Kotedia, K.; Ackerstaff, E.; Ponomarev, V.; Ling, C. C.; Koutcher, J. A.; Li, G. C.
Article Title: Non-invasive molecular and functional imaging of cytosine deaminase and uracil phosphoribosyltransferase fused with red fluorescence protein
Abstract: Introduction. Increased expression of cytosine deaminase (CD) and uracil phosphoribosyltransferase (UPRT) may improve the antitumoral effect of 5-fluorouracil (5-FU) and 5-fluorocytosine (5-FC), and thereby enhance the potential of gene-directed enzyme prodrug therapy. For the applicability of gene-directed enzyme prodrug therapy in a clinical setting, it is essential to be able to monitor the transgene expression and function in vivo. Thus, we developed a preclinical tumor model to investigate the feasibility of using magnetic resonance spectroscopy and optical imaging to measure non-invasively CD and UPRT expression and function. Materials and methods. Expression vectors of CD or CD/UPRT fused to monomeric DsRed (mDsRed) were constructed and rat prostate carcinoma (R3327-AT) cell lines stably expressing either CD/mDsRed or CD/UPRT/mDsRed were generated. The expression of the fusion proteins was evaluated by flow cytometry, fluorescence microscopy, and Western blot analysis. The function of the fusion protein was confirmed in vitro by assessing 5-FC and 5-FU cytotoxicity. In vivo fluorine-19 magnetic resonance spectroscopy (19F MRS) was used to monitor the conversion of 5-FC to 5-FU in mice bearing the R3327-CD/mDsRed and R3327-CD/UPRT/mDsRed tumor xenografts. Results. Sensitivity to 5-FC and 5-FU was higher in cells stably expressing the CD/UPRT/mDsRed fusion gene than in cells stably expressing CD/mDsRed alone or wild-type cells. Whole tumor 19F MRS measurements showed rapid conversion of 5-FC to 5-FU within 20min after 5-FC was administered intravenously in both CD/mDsRed and CD/UPRT/mDsRed tumors with subsequent anabolism to cytotoxic fluoronucleotides (FNucs). CD/UPRT/mDsRed tumor was more efficient in these processes. Conclusion. This study demonstrates the utility of these tumor models stably expressing CD or CD/UPRT to non-invasively evaluate the efficacy of the transgene expression/activity by monitoring drug metabolism in vivo using MRS, with potential applications in preclinical and clinical settings. © 2008 Informa UK Ltd. (Informa Healthcare, Taylor & Francis AS).
Keywords: controlled study; protein expression; unclassified drug; fluorouracil; nonhuman; flow cytometry; sensitivity and specificity; protein function; animal cell; mouse; animals; mice; cell survival; protein protein interaction; animal experiment; animal model; in vivo study; cancer cell culture; cytotoxicity; in vitro study; tumor xenograft; molecular imaging; cell line, tumor; transfection; cell strain cos7; red fluorescent protein; luminescent proteins; prostatic neoplasms; blotting, western; hybrid protein; mice, nude; rat; western blotting; magnetic resonance spectroscopy; nuclear magnetic resonance spectroscopy; fluorescence microscopy; genes, reporter; rats; drug metabolism; transgenes; prostate carcinoma; cytosine deaminase; uracil phosphoribosyltransferase; flucytosine; pentosyltransferases; fluorine radioisotopes; fluoronucleotide derivative; nucleotide derivative
Journal Title: Acta Oncologica
Volume: 47
Issue: 7
ISSN: 0284-186X
Publisher: Informa Healthcare  
Date Published: 2008-01-01
Start Page: 1211
End Page: 1220
Language: English
DOI: 10.1080/02841860802256475
PUBMED: 18661431
PROVIDER: scopus
PMCID: PMC4246416
Notes: --- - "Cited By (since 1996): 5" - "Export Date: 17 November 2011" - "CODEN: ACTOE" - "Source: Scopus"
Altmetric Score
MSK Authors
  1. Gloria C Li
    120 Li
  2. Ligang Xing
    15 Xing
  3. Xuelong Deng
    14 Deng
  4. Jason A Koutcher
    235 Koutcher
  5. Xinzhu Deng
    10 Deng
  6. C Clifton Ling
    306 Ling