In vivo characterization of a reporter gene system for imaging hypoxia-induced gene expression

Purpose: 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 F-18-fluoromisonidazole (F-18-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 I-124-2'-fluoro-2'-deoxy-1-beta-D-arabinofuranosyl-5-iodouracil (I-124-FIAU) 3 h prior to sacrifice. Statistical analysis of the spatial association between I-124-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 F-18-FMISO. Two rats, each bearing a single R3327-AT/HRE tumor, were injected with I-124-FIAU (3 h, before sacrifice) and F-18-FMISO (2 h before sacrifice). Statistical analysis of the spatial association between F-18-FMISO and I-124-FIAU on a pixel-by-pixel basis was performed. Results: Correlation coefficients between I-124-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 F-18-FMISO and I-124-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. (C) 2009 Elsevier Inc. All rights reserved.