Myocardial kinetics of reporter probe 124I-FIAU in isolated perfused rat hearts after in vivo adenoviral transfer of herpes simplex virus type 1 thymidine kinase reporter gene.
Reporter gene imaging holds promise for noninvasive monitoring of cardiac gene therapy. We recently demonstrated that (124)I-labeled 2'-fluoro-2'-deoxy-5'-iodo-1beta-d-arabinofuranosyluracil ((124)I-FIAU) is suitable for PET of myocardial expression of herpes simplex virus type 1 thymidine kinase reporter gene (HSV1-tk). In contrast to previous studies in tumors, early specific uptake was followed by rapid washout. Myocardial kinetics of (124)I-FIAU are still poorly understood. This study aimed at a further investigation under controlled conditions using an isolated heart perfusion model.


METHODS
Male Wistar rats underwent transthoracic regional injection of replication-defective adenovirus (2.5 x 10(9) plaque-forming units) containing either HSV1-tk (n = 16) or LacZ reporter gene (n = 15) into the inferior wall. Nonmanipulated rats (n = 5) served as further controls. Hearts were excised 2 d later and perfused according to the Langendorff technique with (124)I-FIAU-containing buffer (15 min, followed by 30 min of nonradioactive perfusion). Experiments were performed under baseline conditions and in the presence of thymidine (competitive substrate) or fludarabine (in vitro inhibitor of 5'-nucleotidase). Time-activity curves were acquired by external coincident detectors. The myocardial rate of (124)I-FIAU uptake (K(i)), clearance rate (K(o)), and volume of distribution (V(d) = K(i)/K(o)) were calculated. Subsequently, hearts were subjected to gamma-counting, followed by microtome slicing and autoradiography.


RESULTS
The V(d) from Langendorff perfusion significantly correlated with final whole-heart tracer retention (r = 0.88, P = 0.019) and the autoradiographic area of regional myocardial activity (r = 0.89, P = 0.016). HSV1-tk hearts showed higher K(i) and V(d) of (124)I-FIAU compared with that of controls (P < 0.001) and detectable but slower washout compared with that of the LacZ group (P < 0.01). Addition of thymidine to the perfusate inhibited myocardial uptake of (124)I-FIAU by reducing V(d) and K(i) in HSV1-tk and LacZ hearts compared with the baseline. Addition of fludarabine did not result in the expected reduction of washout in HSV1-tk hearts due to inhibition of 5'-nucleotidases (which may dephosphorylate (124)I-FIAU monophosphate). It acted as an uptake inhibitor similar to thymidine, reducing V(d) in HSV1-tk hearts.


CONCLUSIONS
Assessment of specific reporter probe kinetics after regional in vivo reporter gene transfer is feasible using the isolated perfused rat heart preparation. This model allows one to study the effects of pharmacologic interventions and may refine understanding of the reporter probe signal for in vivo imaging. Different nucleoside analogs significantly inhibit (124)I-FIAU uptake, emphasizing the importance of transporter mechanisms for reporter probe kinetics.
Simões
Marcus V
MV
Nuklearmedizinische Klinik und Poliklinik, Technische Universität München, Ismaninger Strasse 22, 81675 Münich, Germany.
Miyagawa
Masao
M
Reder
Sybille
S
Städele
Christian
C
Haubner
Roland
R
Linke
Wolfgang
W
Lehner
Terry
T
Epple
Philipp
P
Anton
Martina
M
Schwaiger
Markus
M
Bengel
Frank M
FM
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001086:Arabinofuranosyluracil / Q000031:analogs & derivatives* / Q000493:pharmacokinetics*; D003198:Computer Simulation; D005240:Feasibility Studies; D020869:Gene Expression Profiling / Q000379:methods*; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D006321:Heart / Q000000981:diagnostic imaging*; D007457:Iodine Radioisotopes; D007700:Kinetics; D008297:Male; D008657:Metabolic Clearance Rate; D008955:Models, Cardiovascular; D009206:Myocardium / Q000378:metabolism; D011877:Radionuclide Imaging; D019275:Radiopharmaceuticals / Q000493:pharmacokinetics; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000378:metabolism; D013937:Thymidine Kinase / Q000235:genetics* / Q000378:metabolism*; D014162:Transfection / Q000379:methods; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism*
D007457:Iodine Radioisotopes; D019275:Radiopharmaceuticals; D011994:Recombinant Proteins; D014764:Viral Proteins; D001086:Arabinofuranosyluracil; C043457:fialuridine; C099241:thymidine kinase, Canid herpesvirus 1; D013937:Thymidine Kinase