In vivo characterization of a reporter gene system for imaging hypoxia-induced gene expression.
OBJECTIVE
To characterize a tumor model containing a hypoxia-inducible reporter gene and to demonstrate utility by comparison of reporter gene expression to the uptake and distribution of the hypoxia tracer (18)F-fluoromisonidazole ((18)F-FMISO).


METHODS
Three tumors derived from the rat prostate cancer cell line R3327-AT were grown in each of two rats as follows: (1) parental R3327-AT, (2) positive control R3327-AT/PC in which the HSV1-tkeGFP fusion reporter gene was expressed constitutively, (3) R3327-AT/HRE in which the reporter gene was placed under the control of a hypoxia-inducible factor-responsive promoter sequence (HRE). Animals were coadministered a hypoxia-specific marker (pimonidazole) and the reporter gene probe (124)I-2'-fluoro-2'-deoxy-1-beta-d-arabinofuranosyl-5-iodouracil ((124)I-FIAU) 3 h prior to sacrifice. Statistical analysis of the spatial association between (124)I-FIAU uptake and pimonidazole fluorescent staining intensity was then performed on a pixel-by-pixel basis. Utility of this system was demonstrated by assessment of reporter gene expression versus the exogenous hypoxia probe (18)F-FMISO. Two rats, each bearing a single R3327-AT/HRE tumor, were injected with (124)I-FIAU (3 h before sacrifice) and (18)F-FMISO (2 h before sacrifice). Statistical analysis of the spatial association between (18)F-FMISO and (124)I-FIAU on a pixel-by-pixel basis was performed.


RESULTS
Correlation coefficients between (124)I-FIAU uptake and pimonidazole staining intensity were: 0.11 in R3327-AT tumors, -0.66 in R3327-AT/PC and 0.76 in R3327-AT/HRE, confirming that only in the R3327-AT/HRE tumor was HSV1-tkeGFP gene expression associated with hypoxia. Correlation coefficients between (18)F-FMISO and (124)I-FIAU uptakes in R3327-AT/HRE tumors were r=0.56, demonstrating good spatial correspondence between the two tracers.


CONCLUSIONS
We have confirmed hypoxia-specific expression of the HSV1-tkeGFP fusion gene in the R3327-AT/HRE tumor model and demonstrated the utility of this model for the evaluation of radiolabeled hypoxia tracers.
Carlin
Sean
S
Department of Medical Physics, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Pugachev
Andrei
A
Sun
Xiaorong
X
Burke
Sean
S
Claus
Filip
F
O'Donoghue
Joseph
J
Ling
C Clifton
CC
Humm
John L
JL
D000818:Animals; D015687:Cell Hypoxia / Q000235:genetics*; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D018259:Herpesvirus 1, Human / Q000201:enzymology; D006801:Humans; D007553:Isotope Labeling; D008297:Male; D011849:Radioactive Tracers; D051381:Rats; D020218:Response Elements / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics; D047368:Tumor Burden / Q000235:genetics
D011849:Radioactive Tracers; D013937:Thymidine Kinase