Evaluation of (76)Br-FBAU as a PET reporter probe for HSV1-tk gene expression imaging using mouse models of human glioma.
The utility of 5-(76)Br-bromo-2'-fluoro-2'-deoxyuridine ((76)Br-FBAU), a uracil analog, as a PET reporter probe for use with the herpes simplex virus type 1 thymidine kinase (HSV1-tk) reporter gene system for gene expression imaging was evaluated in vivo and in vitro using human and rat glioma cells.


METHODS
Human glioma cell lines U87 and U251 were transduced with replication-defective adenovirus constitutively expressing HSV1-tk (Ad.TK) or a control expressing green fluorescent protein (Ad.GFP). These cells were incubated with (76)Br-FBAU for 20-120 min to determine the percentage of total dose uptake. In vitro uptake of equimolar concentrations (1.8 x 10(-8) mol/L) of (76)Br-FBAU and 2'-fluoro-2'-deoxy-5-iodouracil-beta-d-arabinofuranoside ((14)C-FIAU) was also determined in RG2-TK rat glioma cells stably expressing HSV1-tk and in control RG2 cells at 30-120 min. In vivo uptake of (76)Br-FBAU was determined in subcutaneous U87 tumor intratumorally transduced with Ad.TK by ex vivo biodistribution. Uptake in intracranial U87 tumors transduced with Ad.TK expressing HSV1-tk was measured by brain autoradiography. In vivo PET was performed on subcutaneous and intracranial U87 tumors transduced with Ad.TK and on subcutaneous and intracranial stably expressing RG2-TK tumors.


RESULTS
U87 and U251 cells transduced with Ad.TK had significantly increased uptake of (76)Br-FBAU compared with cells transduced with Ad.GFP over 20-120 min. In stably expressing cells at 120 min, (14)C-FIAU uptake in RG2-TK tumor cells was 11.3 %ID (percentage injected dose) and in RG2 control cells was 1.7 %ID, and (76)Br-FBAU uptake in RG2-TK tumor cells was 14.2 %ID and in RG2 control cells was 1.5 %ID. Ex vivo biodistribution of subcutaneous U87 tumors transduced with Ad.TK accumulated (76)Br-FBAU significantly more than in the control Ad.GFP transduced tumor and normal tissue, with the lowest uptake in brain. Autoradiography showed localized uptake in intracranial U87 and U251 cells transduced with Ad.TK. PET image analyses of mice with RG2-TK tumors resulted in an increased tumor-to-background ratio of 13 and 26 from 2 to 6 h after injection, respectively, in intracranial tumors.


CONCLUSIONS
(76)Br-FBAU accumulates in glioma cells constitutively expressing HSV1-tk by either adenoviral transduction or in stably expressing cell lines both in vitro and in vivo. (76)Br-FBAU shows promise as a PET reporter probe for use with the HSV1-tk in vivo gene expression imaging system.
Cho
Steve Y
SY
NeuroOncology Branch, National Cancer Institute and National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892, USA.
Ravasi
Laura
L
Szajek
Lawrence P
LP
Seidel
Jurgen
J
Green
Michael V
MV
Fine
Howard A
HA
Eckelman
William C
WC
D000818:Animals; D001976:Bromouracil / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D004195:Disease Models, Animal; D004353:Drug Evaluation, Preclinical; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D005910:Glioma / Q000000981:diagnostic imaging*; D006801:Humans; D008657:Metabolic Clearance Rate; D051379:Mice; D008819:Mice, Nude; D015336:Molecular Probe Techniques; D009928:Organ Specificity; D011877:Radionuclide Imaging; D019275:Radiopharmaceuticals / Q000493:pharmacokinetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013937:Thymidine Kinase / Q000235:genetics* / Q000378:metabolism*; D014018:Tissue Distribution; D014162:Transfection / Q000379:methods*; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism*
C470854:1-(2-fluoro-2-deoxyarabinofuranosyl)-5-bromouracil; D019275:Radiopharmaceuticals; D011994:Recombinant Proteins; D014764:Viral Proteins; D001976:Bromouracil; C099241:thymidine kinase, Canid herpesvirus 1; D013937:Thymidine Kinase