Kamilla Kelemen

Connexin 43 gene therapy prevents persistent atrial fibrillation in a porcine model

AIMS Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia, and effective treatment of AF still remains an unmet medical need. AF is associated with atrial conduction disturbances caused by electrical and/or structural remodelling. We hypothesized that AF suppresses expression of the gap junction protein connexin (Cx) 43 and that Cx43 gene transfer to both atria would prevent persistent AF. The first aim of this study was to assess whether AF is associated with connexin remodelling in a porcine model. A strategy to suppress persistent AF by gene therapy was then developed and evaluated in vivo. METHODS AND RESULTS AF was induced in domestic pigs via atrial burst pacing, causing a 62.4% reduction in atrial Cx43 protein. Adenoviruses encoding for Cx43 (AdCx43) or green fluorescent protein (AdGFP) were injected into both atria, followed by epicardial electroporation to enhance transgene expression. Combining direct injection of adenoviruses with electroporation achieved GFP reporter gene expression in ∼50% of atrial cells in vivo. AdCx43-treated animals exhibited a 2.5-fold increase in atrial Cx43 protein content and did not develop persistent AF during the observation period of 14 days. In contrast, control animals developed persistent AF within 7.4 ± 0.5 days. Rapid ventricular heart rates during AF led to deterioration of cardiac function in control pigs but not in pigs treated with AdCx43. CONCLUSION Our results highlight the contribution of Cx43 to the pathophysiology of AF and demonstrate the viability of gene therapy for prevention of atrial arrhythmias.

Suppression of persistent atrial fibrillation by genetic knockdown of caspase 3: a pre-clinical pilot study

AIMS Atrial fibrillation (AF) is linked to cardiomyocyte apoptosis, leading to atrial remodelling and reduction in electrical conduction velocity. We hypothesized that genetic suppression of an apoptotic key enzyme, caspase 3, would prevent the development of persistent AF by reducing apoptosis which may serve as an arrhythmogenic substrate. METHODS AND RESULTS Atrial fibrillation was induced in domestic pigs by atrial burst pacing via an implanted cardiac pacemaker. Study animals were then assigned to receive either Ad-siRNA-Cas3 gene therapy to inactivate caspase 3 or green fluorescent protein (Ad-GFP) as a control. Adenoviruses were applied using a hybrid technique employing right and left atrial virus injection followed by epicardial electroporation to increase expression of plasmid DNA. In pigs treated with Ad-siRNA-Cas3, the onset of AF was suppressed or significantly delayed compared with controls (10.3 ± 1.2 days vs. 6.0 ± 1.6 days; P= 0.04). Electrical mapping revealed prolonged atrial conduction in the control group that was prevented by Ad-siRNA-Cas3 gene therapy. On the molecular level, Ad-siRNA-Cas3 application resulted in down-regulation of caspase 3 expression and suppression of apoptotic activity. CONCLUSION Knockdown of caspase 3 by atrial Ad-siRNA-Cas3 gene transfer suppresses or delays the onset of persistent AF by reduction in apoptosis and prevention of intra-atrial conduction delay in a porcine model. These results highlight the significance of apoptosis in the pathophysiology of AF and demonstrate short-term efficacy of gene therapy for suppression of AF.

Genetic suppression of atrial fibrillation using a dominant-negative ether-a-go-go–related gene mutant

BACKGROUND Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia. Gene therapy-dependent modulation of atrial electrophysiology may provide a more specific alternative to pharmacological and ablative treatment strategies. OBJECTIVE We hypothesized that genetic inactivation of atrial repolarizing ether-a-go-go-related gene (ERG) K(+) currents using a dominant-negative mutant would provide rhythm control in AF. METHODS Ten domestic swine underwent pacemaker implantation and were subjected to atrial burst pacing to induce persistent AF. Animals were then randomized to receive either AdCERG-G627S to suppress ERG/I(Kr) currents or green fluorescent protein (AdGFP) as control. Adenoviruses were applied using a novel hybrid technique combining atrial virus injection and epicardial electroporation to increase transgene expression. RESULTS In pigs treated with AdCERG-G627S, the onset of persistent AF was prevented (n = 2) or significantly delayed compared with AdGFP controls (12 ± 2.1 vs. 6.2 ± 1.3 days; P < .001) during 14-day follow-up. Effective refractory periods were prolonged in the AdCERG-G627S group compared with AdGFP animals (221.5 ± 4.7 ms vs. 197.0 ± 4.7 ms; P < .006). Impairment of left ventricular ejection fraction (LVEF) during AF was prevented by AdCERG-G627S application (LVEF(CERG-G627S) = 62.1% ± 4.0% vs. LVEF(GFP) = 30.3% ± 9.1%; P < .001). CONCLUSION Inhibition of ERG function using atrial AdCERG-G627S gene transfer suppresses or delays the onset of persistent AF by prolongation of atrial refractoriness in a porcine model. Targeted gene therapy represents an alternative to pharmacological or ablative treatment of AF.

Tridimensional activation patterns of acquired torsade-de-pointes-tachycardias in dogs with chronic AV-block

Abstract.Background:Dogs with chronic AV block exposed to type-III antiarrhythmic agents develop polymorphic ventricular tachycardias (PVT). Controversy exists regarding PVT mechanism and underlying pathophysiology.Methods and Results:In dogs with acute (n = 10, AAVB) or chronic AV block (n = 14, CAVB, 62 ± 5 days after AV-node ablation) 60 pins (12 mm long, 4 bipolar electrodes) were inserted into both ventricles. QT intervals and effective refractory periods (ERP) at 56 ± 22 randomly selected sites (extrastimulus technique, 800 ms basic cycle length) were determined before and after Almokalant (0.34 μmol/kg). A multiplexer mapping system was used to reconstruct 3D activation patterns. The heart-to-body-weight index (HBWI) was obtained after the experiments. CAVB led to a significant increase in HBWI (11.3 ± 1.5 vs. 9 ± 1.2 g/kg BW, p < 0.001), and a significant increase in ERP (280 ± 28 ms vs. 260 ± 37 ms, p < 0.05) and QT interval (339 ± 16 vs. 288 ±12 ms, p < 0.05). Dispersion (DISP) of ERP was similar for AAVB and CAVB dogs. No AAVB dog, but 9 of 14 CAVB dogs developed PVTs in response to Almokalant. All PVTs originated from an endocardial focus. Consecutive beats continued to reveal centrifugal activation patterns in 8 of 10 episodes. In only 2 episodes was reentrant activation evident.Conclusion:Myocardial hypertrophy associated with CAVB predisposes the canine heart to druginduced PVTs. This seems to be primarily linked to prolonged repolarization. PVTs in this model are not only initiated, but also perpetuated by a centrifugal spread of activation.

Ventricular tachycardia from the healed myocardial infarction scar: Validation of an animal model and utility of gene therapy

Life-threatening ventricular arrhythmias generally occur in the setting of structural heart disease. Current clinical options for patients at risk for these rhythm disturbances are limited. We developed a porcine model of inducible ventricular tachycardia originating in the border region of a healed myocardial infarction scar. After validating the model, we assessed gene transfer techniques, focusing on local modification of border zone tissues. We found that gene transfer of the dominant negative KCNH2-G628S mutation to the anteroseptal infarct border caused localized prolongation of effective refractory period in the target region and eliminated all ventricular arrhythmia inducibility. In this work, we characterize the animal model and review the gene transfer results.

Biological Heart Rate Reduction Through Genetic Suppression of Gαs Protein in the Sinoatrial Node

Background Elevated heart rate represents an independent risk factor for cardiovascular outcome in patients with heart disease. In the sinoatrial node, rate increase is mediated by β1 adrenoceptor mediated activation of the Gαs pathway. We hypothesized that genetic inactivation of the stimulatory Gαs protein in the sinoatrial node would provide sinus rate control and would prevent inappropriate heart rate acceleration during β-adrenergic activation. Methods and Results Domestic pigs (n=10) were evenly assigned to receive either Ad-small interfering RNA (siRNA)-Gαs gene therapy to inactivate Gαs or adenovirus encoding for green fluorescent protein (Ad-GFP) as control. Adenoviruses were applied through virus injection into the sinoatrial node followed by epicardial electroporation, and heart rates were evaluated for 7 days. Genetic inhibition of Gαs protein significantly reduced mean heart rates on day 7 by 16.5% compared with control animals (110±8.8 vs 131±9.4 beats per minute; P<0.01). On β-adrenergic stimulation with isoproterenol, we observed a tendency toward diminished rate response in the Ad-siRNA-Gαs group (Ad-siRNA-Gαs, +79.3%; Ad-GFP, +61.7%; n=3 animals per group; P= 0.294). Adverse effects of gene transfer on left ventricular ejection fraction (LVEF) were not detected following treatment (LVEFAd-siRNA-Gαs, 66%; LVEFAd-GFP, 60%). Conclusions In this preclinical proof-of-concept study targeted Ad-siRNA-Gαs gene therapy reduced heart rates during normal sinus rhythm compared with Ad-GFP treatment and prevented inappropriate rate increase after β-adrenergic stimulation. Gene therapy may provide an additional therapeutic option for heart rate reduction in cardiac disease. (J Am Heart Assoc. 2012;1:jah3-e000372 doi: 10.1161/JAHA.111.000372)

The new selective IKs–blocking agent HMR 1556 restores sinus rhythm and prevents heart failure in pigs with persistent atrial fibrillation

AbstractBackgroundAntiarrhythmic drugs for treatment of atrial fibrillation in patients with heart failure are limited by proarrhythmia and low efficacy. Experimental studies indicate that the pure IKs blocking agents chromanol 293b and HMR 1556 prolong repolarization more markedly at fast than at slow heart rates and during β–adrenergic stimulation. These properties may overcome some of the above quoted limitations.Methods and resultsTen domestic swine underwent pacemaker implantation (PM) and atrial burst pacing to induce persistent AF. Four days after onset of persistent AF, pigs were randomized to HMR 1556 (30 mg/kg, p.o., 10 days) or placebo. All animals receiving HMR 1556 converted to SR (5.2 ± 1.9 days), whereas placebo pigs remained in AF. Pigs treated with placebo developed high ventricular rates (297 ± 5 bpm) and severe heart failure, whereas pigs treated with HMR 1556 remained hemodynamically stable. Left ventricular ejection fraction on the day of euthanization was significantly lower in the placebo compared to the HMR 1556 group (30 ± 4% vs. 69 ± 5%, p < 0.005). Similar results were seen with epinephrine levels (placebo 1563 ± 193 pmol/l vs. HMR 613 ± 196 pmol/l, p < 0.05). Right atrial monophasic action potentials were significantly longer in the HMR 1556 compared to the placebo group (230 ± 7 ms vs. 174 ± 13 ms, p < 0.05).ConclusionsThe new IKs blocker HMR 1556 efficiently and safely restores SR and prevents CHF in a model of persistent AF. Restoration of SR is most likely linked to a marked prolongation of atrial repolarization even at high heart rates.

Effizienzpotential in der Nachsorge von Schrittmachern und implantierbaren Kardioverter Defibrillatoren

SummaryThe aim of the present study was to elucidate whether the duration of a technical follow-up (FU) of a pacemaker (PM)/implantable cardioverter defibrillator (ICD) has an impact on cost-effectiveness in the outpatient clinic. We determined the time required for a complete FU of devices from three different manufacturers.In 130 patients (70 VVI/DDD-PM, 60 VVI/DDD-ICD) with either a PM (Phylos, Chorum/Talent, Kappa, EnPulse) or an ICD (Belos, Alto or GEM) the time was recorded for a complete FU including determination of lead impedance, sensing and pacing threshold. The time for activation of individual menue buttons was excluded. On the basis of time required for FU, cost-units (CU) were calculated for 2000 FU/year and for a presumed device longevity (PM 7 years, ICD 5 years).For VVI-PM, the duration of FU was almost identical for devices from different manufacturers (105±11 s to 125±8 s; p=n.s.). However, analysis of DDD-PM revealed marked differences (140±25 s vs 282±23 s, p<0.05). Time for FU of ICDs varied between 108±5 s and 207±21 s (p<0.05) in VVI-ICDs and between 129±8 ms and 225±23 s (p<0.05) in DDD-ICDs. The total savings could be 55 000 CU in VVI- and 53 333 CU in DDD-ICDs. For full automatic DDD-pacemakers (EnPulse) time for FU could be reduced to 58±3 s (p<0.05). Differences in FU times were caused by problems with telemetry, delay during booting of the programmer, interrogation at the beginning and at the end of FU and for sensing tests.Improving not only programmers and devices but also test automaticity could significantly increase cost-efficiency in the outpatient clinic.ZusammenfassungDie vorliegende Studie untersucht geräteabhängige Kostensenkungspotentiale im Prozess der Nachsorge beim Einsatz verschiedener Schrittmacher (SM) und implantierbarer Kardioverter-Defibrillatoren (ICD).Bei 130 Patienten (n=60 VVI-SM/ICD, n=70 DDD-SM/ICD) mit implantiertem SM (Phylos, Chorum/Talent, Kappa, EnPulse) oder ICD (Belos, Alto oder GEM) erfolgte eine Zeiterfassung für die Nachsorge der Aggregate (inklusive Erst- und Endabfrage, Testung der Impedanz, Wahrnehmung intrinsischer Signale und Stimulationsreizschwelle). Das Aufsuchen der einzelnen Funktionen am Programmiergerät wurde in den Analysen nicht berücksichtigt. Um eine wirtschaftliche Evaluierung der Ergebnisse zu ermöglichen, wurden die Nachsorgezeiten mit einem fiktiven Referenzkostensatz von 200 Kosteneinheiten pro Aktivitätsstunde bewertet.Die ermittelte Zeit pro Nachsorgevorgang variiert herstellerabhängig bei Einkammer-SM-Aggregaten von 105±11 s (Phylos) bis 125±8 s (Kappa, p>0,05). Bei DDD-SM-Aggregaten wurde eine Nachsorgezeit von 140±25 s (Phylos) bis 282±23 s (Talent, p<0,05) ermittelt. Bei ICD-Aggregaten wurden bei Einkammer-Geräten zwischen 108±5 s (Belos) und 207±21 s (Alto, p<0,05), bei Zweikammer-Aggregaten zwischen 129±8 s (Belos) und 225±23 s (Alto, p<0,05) benötigt. Die unterschiedliche Dauer der Nachsorge beruhte überwiegend auf Problemen in der Telemetrie, Verzögerungen beim Systemstart, bei Erstabfrage und Endabfrage mit Ausdruck sowie der Bestimmung der Wahrnehmung. Durch Einsatz vollautomatischer DDD-SM (EnPulse) ergibt sich eine minimale Nachsorgezeit von 58±3 s. Verglichen mit dem Wert von 282±23 s, p<0,05 beim ungünstigsten SM bedeutet dies eine Verringerung der Nachsorgekosten um nahezu 80%.Der unterschiedliche Einfluss der Gerätetypen verschiedener Hersteller auf die Nachsorgezeit und auf die Wirtschaftlichkeit des Nachsorgeprozesses wird bestätigt. Die Leistungsfähigkeit der Programmiergeräte und die Automatisierung der Gerätetestungen bestimmen dabei die Wirtschaftlichkeit in der Nachsorge von SM/ICD.

[Efficiency potential in the pacemaker/implantable cardioverter defibrillator outpatient clinic].

SummaryThe aim of the present study was to elucidate whether the duration of a technical follow-up (FU) of a pacemaker (PM)/implantable cardioverter defibrillator (ICD) has an impact on cost-effectiveness in the outpatient clinic. We determined the time required for a complete FU of devices from three different manufacturers.In 130 patients (70 VVI/DDD-PM, 60 VVI/DDD-ICD) with either a PM (Phylos, Chorum/Talent, Kappa, EnPulse) or an ICD (Belos, Alto or GEM) the time was recorded for a complete FU including determination of lead impedance, sensing and pacing threshold. The time for activation of individual menue buttons was excluded. On the basis of time required for FU, cost-units (CU) were calculated for 2000 FU/year and for a presumed device longevity (PM 7 years, ICD 5 years).For VVI-PM, the duration of FU was almost identical for devices from different manufacturers (105±11 s to 125±8 s; p=n.s.). However, analysis of DDD-PM revealed marked differences (140±25 s vs 282±23 s, p<0.05). Time for FU of ICDs varied between 108±5 s and 207±21 s (p<0.05) in VVI-ICDs and between 129±8 ms and 225±23 s (p<0.05) in DDD-ICDs. The total savings could be 55 000 CU in VVI- and 53 333 CU in DDD-ICDs. For full automatic DDD-pacemakers (EnPulse) time for FU could be reduced to 58±3 s (p<0.05). Differences in FU times were caused by problems with telemetry, delay during booting of the programmer, interrogation at the beginning and at the end of FU and for sensing tests.Improving not only programmers and devices but also test automaticity could significantly increase cost-efficiency in the outpatient clinic.ZusammenfassungDie vorliegende Studie untersucht geräteabhängige Kostensenkungspotentiale im Prozess der Nachsorge beim Einsatz verschiedener Schrittmacher (SM) und implantierbarer Kardioverter-Defibrillatoren (ICD).Bei 130 Patienten (n=60 VVI-SM/ICD, n=70 DDD-SM/ICD) mit implantiertem SM (Phylos, Chorum/Talent, Kappa, EnPulse) oder ICD (Belos, Alto oder GEM) erfolgte eine Zeiterfassung für die Nachsorge der Aggregate (inklusive Erst- und Endabfrage, Testung der Impedanz, Wahrnehmung intrinsischer Signale und Stimulationsreizschwelle). Das Aufsuchen der einzelnen Funktionen am Programmiergerät wurde in den Analysen nicht berücksichtigt. Um eine wirtschaftliche Evaluierung der Ergebnisse zu ermöglichen, wurden die Nachsorgezeiten mit einem fiktiven Referenzkostensatz von 200 Kosteneinheiten pro Aktivitätsstunde bewertet.Die ermittelte Zeit pro Nachsorgevorgang variiert herstellerabhängig bei Einkammer-SM-Aggregaten von 105±11 s (Phylos) bis 125±8 s (Kappa, p>0,05). Bei DDD-SM-Aggregaten wurde eine Nachsorgezeit von 140±25 s (Phylos) bis 282±23 s (Talent, p<0,05) ermittelt. Bei ICD-Aggregaten wurden bei Einkammer-Geräten zwischen 108±5 s (Belos) und 207±21 s (Alto, p<0,05), bei Zweikammer-Aggregaten zwischen 129±8 s (Belos) und 225±23 s (Alto, p<0,05) benötigt. Die unterschiedliche Dauer der Nachsorge beruhte überwiegend auf Problemen in der Telemetrie, Verzögerungen beim Systemstart, bei Erstabfrage und Endabfrage mit Ausdruck sowie der Bestimmung der Wahrnehmung. Durch Einsatz vollautomatischer DDD-SM (EnPulse) ergibt sich eine minimale Nachsorgezeit von 58±3 s. Verglichen mit dem Wert von 282±23 s, p<0,05 beim ungünstigsten SM bedeutet dies eine Verringerung der Nachsorgekosten um nahezu 80%.Der unterschiedliche Einfluss der Gerätetypen verschiedener Hersteller auf die Nachsorgezeit und auf die Wirtschaftlichkeit des Nachsorgeprozesses wird bestätigt. Die Leistungsfähigkeit der Programmiergeräte und die Automatisierung der Gerätetestungen bestimmen dabei die Wirtschaftlichkeit in der Nachsorge von SM/ICD.

Is the determination of the defibrillation threshold in patients with an implantable cardioverter–defibrillator still required?

Hintergrund Die intraoperative Testung von implantierbaren Kardioverter/Defibrillatoren (ICD) ist personalintensiv und mit Komplikationen vergesellschaftet. In der vorliegenden Studie untersuchten wir, ob bei ICD-Implantationen mit unterschiedlicher maximaler Energieabgabe eine intraoperative Testung inklusive der Induktion von Kammerflimmern (DFT) notwendig gewesen ware.