Molecular Imaging of the Kinetics of Vascular Endothelial Growth Factor Gene Expression in Ischemic Myocardium

Abstract

Background— Angiogenic gene therapy is a promising treatment paradigm for patients with ischemic heart disease. In this study, we used micro-positron emission tomography (microPET) to monitor the transgene expression, function, and effects in a whole-body system. Methods and Results— Adenovirus with cytomegalovirus promoter driving an angiogenic gene (vascular endothelial growth factor [VEGF]) linked to a PET reporter gene (herpes simplex virus type 1 mutant thymidine kinase; Ad-CMV-VEGF 121 -CMV-HSV1-sr39tk) was used to transfect rat embryonic cardiomyoblasts in vitro. Expression of both genes correlated strongly ( r =0.98; P <0.001). Afterward, rats underwent ligation of the left anterior descending artery followed by injection of 1×10 10 pfu of Ad-CMV-VEGF 121 -CMV-HSV1-sr39tk (study; n=35) or Ad-null (control; n=15) at the peri-infarct region. Noninvasive microPET imaging was used to assess the uptake of 9-(4-[ 18 F]-fluoro-hydroxymethylbutyl)guanine ([ 18 F]-FHBG) PET reporter probe by cells expressing the HSV1-sr39tk PET reporter gene. Cardiac transgene expression peaked at day 1 and declined over the next 2 weeks. Repeat adenoviral injections at day 60 yielded no detectable signal. The in vivo reporter gene expression (% injected dose/g of [ 18 F]-FHBG) correlated well with ex vivo gamma counting ( r =0.92), myocardial tissue HSV1-sr39TK enzyme activity ( r =0.95), and myocardial tissue VEGF level ( r =0.94; P <0.001 for all). The VEGF 121 isoform induced significant increases in capillaries and small blood vessels. However, the level of neovasculature did not translate into significant improvements in functional parameters such as myocardial contractility by echocardiography, perfusion by nitrogen-13 ammonia imaging, and metabolism by [ 18 F]-fluorodeoxyglucose imaging. Conclusions— Taken together, these findings establish the feasibility of molecular imaging for monitoring angiogenic gene expression with a PET reporter gene and probe noninvasively, quantitatively, and repetitively. The principles demonstrated here can be used to evaluate other therapeutic genes of interest in animal models before future clinical trials are initiated.

Noninvasive Characterization of Myocardial Molecular Interventions by Integrated Positron Emission Tomography and Computed Tomography

“…The CT morphology helped in accurately identifying the area of intervention and in localization of perfusion increase. It is tempting to speculate that lack of morphologic information to accurately identify injection sites may have resulted in a lack of observation of physiologic in vivo effects of gene transfer in a previous study with stand-alone microPET (15). Interestingly, expression of ␣ v ␤ 3 integrins did not seem to play a role in the VEGF-induced increase of perfusion early after gene transfer in our study.…”

Molecular cardiovascular imaging using scintigraphic methods

“…Successful co-expression of the two reporter gene/probe systems HSV1-sr39tk and D2R [66] as well as coexpression of a therapeutic gene for VEGF expression and the reporter gene HSV1-sr39tk [67] have established the feasibility of monitoring "angiogenic gene expression with a PET reporter gene and probe noninvasively, quantitatively, and repetitively". [67].…”

Molecular imaging of cardiac stem cell transplantation

“…First, a reporter gene construct is created that links a promoter or regulatory gene of interest to the reporter gene. Depending on the nature of the promoter (eg, constitutive versus inducible), the pattern of reporter gene expression may vary [45]. Second, the reporter gene construct is introduced into cells by viral or nonviral vectors.…”

“…As more probes accumulate within the cell, signal strength increases and can be detected by PET. This imaging approach has been successfully used in animal models [45] and human trials [46]. In the case of optical bioluminescence imaging, the reporter gene produces a firefly luciferase enzyme, which oxidizes the probe (D-luciferin) to produce light as the signal.…”