Development of a bicistronic vector for multimodality imaging of estrogen receptor activity in a breast cancer model: preliminary application.
OBJECTIVE
The aim of this study was to develop a cellular model for the concurrent imaging of reporter genes expression by using positron emission tomography (PET) and bioluminescence imaging (BLI) for the assessment of estrogen receptor activity in vivo in a breast cancer model.


METHODS
Two reporters were chosen: a mutated form of the dopaminergic D2 receptor (D(2)R80A) for PET imaging, and the Firefly Luciferase for BLI. The presence of an IRES sequence between the two reporters ensured the coordinated expression driven by the same regulatory sequence containing an estrogen responsive element (ERE). To prevent chromatin effects on reporter expression, the construct was flanked by insulator sequences (Matrix Attachment Region, MAR).


RESULTS
In vitro studies showed that the vector was efficient in coordinating the expression of the two genes. Moreover, stably transfected cells implanted in recipient animals maintained their capacity to express the reporters and react to systemic treatments permitting the in vivo study of ERs activity by PET and BLI imaging. In vitro expression analysis after long-term treatments showed different behaviour of the two reporter proteins in monitoring estrogen-dependent transcription outlining the importance of multi-reporter systems. With this model, PET and BLI can be applied to the concurrent evaluation of gene expression induced by estrogen and its analogues by using a bicistronic construct.


CONCLUSIONS
The combined features of rapid, sensitive, sequential BLI and tomographic and quantitative PET imaging will allow the use of this strategy for the in vivo evaluation of molecular processes also for pharmacodynamic studies.
Ottobrini
Luisa
L
Institute of Radiological Sciences, University of Milan, Unit of Nuclear Medicine, Hospital San Paolo, Milan, Italy.
Ciana
Paolo
P
Moresco
Rosamaria
R
Lecchi
Michela
M
Belloli
Sara
S
Martelli
Cristina
C
Todde
Sergio
S
Fazio
Ferruccio
F
Gambhir
Sanjiv Sam
SS
Maggi
Adriana
A
Lucignani
Giovanni
G
D000818:Animals; D001943:Breast Neoplasms / Q000000981:diagnostic imaging* / Q000378:metabolism*; D045744:Cell Line, Tumor; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D007089:Image Enhancement / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D015336:Molecular Probe Techniques; D010865:Pilot Projects; D011877:Radionuclide Imaging; D011960:Receptors, Estrogen / Q000235:genetics / Q000378:metabolism*; D013382:Subtraction Technique; D014162:Transfection / Q000379:methods*
D011960:Receptors, Estrogen