Cardiovascular diseases remain the greatest cause of death in the US and gene therapy has the potential to be an effective
therapy. In this study, we demonstrated MMP-9 based protease-activatable virus (PAV) for selective infection of
myocardial infarct (MI) that is associated with active MMP-9 expression. To test the specificity of PAV, we used
expression of a far-red fluorescence protein (iRFP) delivered by the PAV together with a dual PET/NIRF imaging agent
specific for active MMP-9 activity at the site of MI in a murine model. Calibrated fluorescence imaging employed a
highly-sensitive intensified camera, laser diode excitation sources, and filtration schemes based upon the spectra of iRFP
and the NIRF agent. One to two days after ligation of the left anterior descending artery, the PAV or WT AAV9 virus
encoding for iRFP (5x1010 genomic particles) and radiolabeled MMP-9 imaging agent (3 nmol) were injected
intravenously (i.v.). PET imaging showed MMP activity was associated with adverse tissue remodeling at the site of the
MI. One week after, animals were again injected i.v. with the MMP-9 agent (3 nmol) and 18-24 h later, the animals
were euthanized and the hearts were harvested, sliced, and imaged for congruent iRFP transgene expression and NIRF
signals associated with MMP-9 tissue activity. The fluorescent margins of iRFP and NIRF contrasted tissues were
quantified in terms Standard International units of mW/cm2/sr. The sensitivity, specificity, and accuracy of PAV and
WT targeting to sites of MI was determined from these calibrated fluorescence measurements. The PAV demonstrated
significantly higher delivery performance than that of the WT AAV9 virus.
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