Introduction: Antisense locked nucleic acids (LNA) GapmeRs are stable small oligonucleotides able to bind their mRNA target with high affinity and induce gene silencing. Small conductance calcium-activated potassium (SK) channels represent a novel target for rhythm control therapy in atrial fibrillation (AF). We designed specific SK3-LNA-GapmeRs, which in vitro demonstrated specific SK3-knockdown. Here, we show the effect of transient SK3 gene silencing in vivo to investigate the potential of LNA-GapmeRs in the treatment of AF.
Material and methods: 22 male Wistar rats were randomly assigned to receive either 50 mg/kg SK3-GapmeR or vehicle subcutaneously once a week for two weeks. Seven days after the last treatment, rats were euthanized by a IP lethal injection of sodium pentobarbital, organs were removed and Langendorff experiments were performed to investigate electrophysiological parameters, such as action potential duration (APD), effective refractory period (ERP) and AF propensity. SK3 channel activity was evaluated using the SK channel blocker, N-(pyridin-2-yl)-4-(pyridine-2-yl)thiazol-2-amine (ICA). SK3 protein expression level was assessed by Western Blot. The experiments were performed under the animal license (2017-15-0201-01231) authorized by the Danish Animal Inspectorate and in accordance with the EU legislations for animal protection and care.
Results: The designed LNA-GapmeR effectively downregulated the SK3 protein expression level in the heart (p<0.01). We found a 78% reduction in average duration of AF episodes elicited by burst pacing in the hearts of rats treated with SK3-GapmeR compared to controls (3.7 vs. 16.8 s, p<0.05). These AF events were also significantly shorter in duration (p<0.05). Refractoriness was not altered at the baseline. However, ICA did not prolong ERP in the SK3-GapmeR group.
Conclusion: The designed SK3-LNA-GapmeR silenced SK3 channels, preventing acutely induced AF in rats. Thus, GapmeR technology can be applied as an experimental platform for downregulating cardiac proteins and offers a potential modality for the treatment of cardiac arrhythmias.
