Rescue of SERCA2a expression by gene transfer to the endstage failing heart improves its mechanical and electrophysiological (EP) function. However the safety profile of SERCA2a gene therapy as a preventive measure in patients at early stages of remodeling prior to SERCA2a downregulation is unknown. Examination of the EP consequences of SERCA2a gene therapy in non-failing hearts is imperative in determining its potential EP toxicity.
Hypothesis:
SERCA2a upregulation using high titer adeno associated virus type 9 (AAV9) mediated gene transfer to normal rats may promote arrhythmogenic triggers, thereby limiting the therapeutic window for this approach.
Methods:
Using high resolution optical action potential (AP) imaging, we investigated the EP substrate 6 weeks post gene transfer of 5E11 AAV9.SERCA2a (N=13) or AAV9.GFP (N=6) to normal rats. Arrhythmia susceptibility was determined under conditions that promoted Ca overload by burst pacing & ischemia reperfusion (IR) injury.
Results:
AAV9.SERCA2a hearts exhibited a 50% increase in SERCA2a expression at the mRNA and protein levels compared to AAV9.GFP. No differences in heart weight to body weight were found between groups (3.85 vs 3.74 mg/g). During baseline perfusion, AAV9.GFP hearts exhibited significantly prolonged (by 30%, p<0.05) AP durations (APD) and a strong trend (p=0.07) towards greater APD heterogeneity. Challenge with rapid pacing resulted in the onset of sustained VF in 4/6 AAV9.GFP and 0/13 AAV9.SERCA2a hearts (p=0.004). Despite significant SERCA2a upregulation, APD and conduction velocity were identical in AAV9.SERCA2a compared to normal hearts indicating lack of EP toxicity. During prolonged low flow ischemia, AAV9.SERCA2a hearts exhibited a trend towards higher incidence of arrhythmias compared to normal hearts, but differences were not statistically significant (p=0.08). Finally, all AAV9.SERCA2a hearts exhibited rapid recovery of APD and excitability upon reperfusion following 10 min of no flow ischemia.
Conclusions:
High titer gene transfer of AAV9.GFP but not AAV9.SERCA2a elicits adverse EP remodeling in normal rat myocardium. The lack of an EP phenotype of AAV9.SERCA2a despite 50% upregulation in SERCA2a levels is consistent with a wide safety margin for this therapy.
Quantitation of Trace Levels of DNA Released from Disrupted Adeno-Associated Virus Gene Therapy Vectors