In vivo detection and treatment of ischemia-induced cardiac apoptosis using an MRI-detectable molecular probe and an alpha-adrenergic receptor agonist

Summary A novel anti-apoptotic alpha-adrenergic agonist preserves left ventricular ejection fraction in mice following myocardial infarction. This therapeutic effect is able to be detected and quantified non-invasively, using T2* signal loss assessment of an Annexin-SPIO molecular apoptosis probe. Background Myocardial infarction (MI) leads to cardiomyopathy through a combination of programmed cell death (re: apoptosis) and necrotic cell death. The relative contribution of apoptosis to ischemic cardiomyopathy and the effect of targeting apoptosis specifically to prevent MIinduced damage is unknown. Our laboratory previously developed and validated an in vivo, MRI-detectable apoptosis probe. Annexin-V (ANX), which binds to cells in the earliest stages of apoptosis, was conjugated to superparamagnetic iron oxide (SPIO) nanoparticles, allowing for the non-invasive detection of early apoptotic cell populations (ANX-SPIO r1: 8.6 ± 0.61 mM-1 s-1 and r2: 326 ± 16 mM-1 s-1). To test the effect of apoptosis reversal in an MI model, we employed A61603 (A6), an a1-adrenergic receptor agonist, which was previously shown to rescue cardiac cells from Doxorubicininduced cardiac apoptosis through activation of the cardio-protective ERK pathway. Our hypothesis is that A6 therapy will protect against MI-induced cardiomyopathy, and that cardiac MRI of systemic ANX-SPIO will detect and longitudinally monitor this therapeutic effect in vivo.

In vivo detection and treatment of ischemiainduced cardiac apoptosis using an MRIdetectable molecular probe and an alphaadrenergic receptor agonist Justin Lam 1 , Ildiko Toma 1 , Yongquan Gong 2 , Paul C Simpson 3,4 , Robert C Robbins 2 , Phillip C Yang 1* , Rajesh Dash 1 From 15th Annual SCMR Scientific Sessions Orlando, FL, USA. 2-5 February 2012 Summary A novel anti-apoptotic alpha-adrenergic agonist preserves left ventricular ejection fraction in mice following myocardial infarction. This therapeutic effect is able to be detected and quantified non-invasively, using T2* signal loss assessment of an Annexin-SPIO molecular apoptosis probe.

Background
Myocardial infarction (MI) leads to cardiomyopathy through a combination of programmed cell death (re: apoptosis) and necrotic cell death. The relative contribution of apoptosis to ischemic cardiomyopathy and the effect of targeting apoptosis specifically to prevent MIinduced damage is unknown. Our laboratory previously developed and validated an in vivo, MRI-detectable apoptosis probe. Annexin-V (ANX), which binds to cells in the earliest stages of apoptosis, was conjugated to superparamagnetic iron oxide (SPIO) nanoparticles, allowing for the non-invasive detection of early apoptotic cell populations (ANX-SPIO r1: 8.6 ± 0.61 mM-1 s-1 and r2: 326 ± 16 mM-1 s-1). To test the effect of apoptosis reversal in an MI model, we employed A61603 (A6), an α1-adrenergic receptor agonist, which was previously shown to rescue cardiac cells from Doxorubicininduced cardiac apoptosis through activation of the cardio-protective ERK pathway. Our hypothesis is that A6 therapy will protect against MI-induced cardiomyopathy, and that cardiac MRI of systemic ANX-SPIO will detect and longitudinally monitor this therapeutic effect in vivo.

Methods
Mice underwent MI (via LAD ligation) along with a subcutaneous pump implant that delivered A6 or vehicle (VEH) solution at a rate of 10 ng/kg/day over a course of two weeks. Cardiac MRI (CMR) was performed at 2 days, 1 week, and 2 weeks post MI. ANX-SPIO was delivered via tail vein one day prior to CMR to simultaneously assess left ventricular function and apoptosis (using T2* signal loss as a marker of apoptotic activity) in vivo.

Conclusions
These results suggest that cardiomyocyte apoptosis is a prominent contributor to the functional impairment of ischemic cardiomyopathy and that A6-mediated 1 Figure 1 Graph of EFs for A6-treated vs VEH-treated mice over a two week period. Note the significant preservation of EFs in A6-treated hearts at 1 and 2 weeks.
Figure 2 a) CMR images of an infarcted mouse heart showing T2* signal loss from ANX-SPIO; b) CMR of A6-treated, infarcted mouse heart showing a visibly reduced degree of T2* signal loss, compared to VEH control hearts; c) T2* signal loss curves for control animals, as well as MI +A6-treated and MI+VEH hearts. The steeper decay curve of MI+VEH hearts reflects the increased apoptosis and subsequent ANX-SPIO uptake, compared to A6-treated hearts. cardioprotection from MI-induced apoptosis preserves cardiac function. Moreover, ANX-SPIO can non-invasively detect and monitor A6's therapeutic effect longitudinally.