title: Rapid obtention of stable, bioluminescent tumor cell lines using a tCD2-luciferase chimeric construct.
abstract: BACKGROUND
  Bioluminescent tumor cell lines are experimental tools of major importance for cancer investigation, especially imaging of tumors in xenografted animals. Stable expression of exogenous luciferase in tumor cells combined to systemic injection of luciferin provides an excellent signal/background ratio for external optical imaging. Therefore, there is a need to rationalize and speed up the production of luciferase-positive tumor cell lines representative of multiple tumor phenotypes. For this aim we have designed a fusion gene linking the luciferase 2 protein to the c-terminus of a truncated form of the rat CD2 protein (tCD2-luc2). To allow simultaneous assessment of the wild-type luciferase 2 in a context of tCD2 co-expression, we have made a bicistronic construct for concomitant but separate expression of these two proteins (luc2-IRES-tCD2). Both the mono- and bi-cistronic constructs were transduced in lymphoid and epithelial cells using lentiviral vectors.
  
  
  RESULTS
  The tCD2-luc2 chimera behaves as a type I membrane protein with surface presentation of CD2 epitopes. One of these epitopes reacts with the OX34, a widely spread, high affinity monoclonal antibody. Stably transfected cells are sorted by flow cytometry on the basis of OX34 staining. In vitro and, moreover, in xenografted tumors, the tCD2-luc2 chimera retains a substantial and stable luciferase activity, although not as high as the wild-type luciferase expressed from the luc2-IRES-tCD2 construct. Expression of the tCD2-luc2 chimera does not harm cell and tumor growth.
  
  
  CONCLUSIONS
  Lentiviral transduction of the chimeric tCD2-luc2 fusion gene allows selection of cell clones with stable luciferase expression in less than seven days without antibiotic selection. We believe that it will be helpful to increase the number of tumor cell lines available for in vivo imaging and assessment of novel therapeutic modalities. On a longer term, the tCD2-luc2 chimera has the potential to be expressed from multi-cassette vectors in combination with various inserts of interest.
authors: Université Paris-Sud 11, CNRS-UMR 8126 and Institut de Cancérologie Gustave Roussy, Villejuif cedex, France.
authors: Anne-Sophie
authors: AS
authors: Jimenez
authors: Mélanie
authors: M
authors: Gressette
authors: Clément
authors: C
authors: Barjon
authors: Ming
authors: M
authors: Wei
authors: Claire
authors: C
authors: Gourzones
authors: Pierre
authors: P
authors: Busson
mesh_terms: D000818:Animals; D015153:Blotting, Western; D018801:CD2 Antigens / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D005260:Female; D005434:Flow Cytometry; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism*; D008163:Luminescent Measurements / Q000379:methods*; D051379:Mice; D008819:Mice, Nude; D009154:Mutation; D009374:Neoplasms, Experimental / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014183:Transplantation, Heterologous
chemical_list: D018801:CD2 Antigens; D011993:Recombinant Fusion Proteins; D008156:Luciferases