AAV-mediated gene therapy over-expressing wild-type PLN significantly reduced stress-induced arrhythmia in a preclinical mouse model of PLN-R14del cardiomyopathy.
Does AAV9-wtPLN gene therapy reduce stress-induced arrhythmia in a humanized PLN-R14del mouse model of cardiomyopathy?
AAV-mediated over-expression of wild-type PLN modestly restores PLN expression and significantly reduces arrhythmogenicity in a preclinical mouse model of PLN-R14del cardiomyopathy.
Abstract Background/ Objectives The PLN-R14del mutation is a founder mutation originating in the Netherlands approximately 600-800 years ago and is identified in 10-15% of patients with dilated cardiomyopathy (DCM) or arrhythmogenic cardiomyopathy (ACM) (1). PLN regulates cardiomyocyte calcium handling by acting as a primary inhibitor of sarcoplasmic/endoplasmic reticulum Ca2+ATPase 2a (SERCA2a). The PLN-R14del mutation disrupts the phosphorylation site at Ser 16 of PLN, leading to impaired SERCA2a activity, abnormal calcium handling, and arrhythmia (2). Current therapy strategies are limited to symptom management and prevention of sudden cardiac death highlighting an urgent need for novel therapeutic strategies. The purpose of this study is to evaluate a novel Adeno-associated virus (AAV) -mediated treatment over-expressing the wild-type (WT) PLN, and to investigate how over-expression of the WT gene impacts cardiac function in a humanized PLN-R14del mouse model. Methods Three-month-old heterozygous (HET) hPLN-R14del mice were injected with 1 x 10¹2 vg/mouse dose of AAV9-wtPLN vector, or saline control. Five weeks post-treatment, efficacy was assessed using an in-vivo stress induced arrhythmia protocol. A baseline ECG was recorded for 5 minutes, followed by a simultaneous intraperitoneal (IP) injection of epinephrine (2mg/Kg) and caffeine (120 mg/Kg), with an additional 10-15 minutes of post-injection ECG recording. The mice were subsequently sacrificed, and their hearts and other organs were collected for downstream analysis to determine the effectiveness of the AAV-mediated therapy. The ECGs were analyzed using the beat classifier module in LabChart8 to quantify the percentage of arrhythmic beats, while viral genome and PLN gene expression were assessed by qPCR and digital droplet PCR. Results Arrhythmia could be efficiently induced by the simultaneous IP injection of epinephrine and caffeine in PLN-R14del HET mice. HET mice show significantly more arrhythmia compared to WT mice. The efficiency of the AAV-mediated treatment was determined by the viral genome expression. Treated mice show sufficient expression of the viral genome compared to saline injected mice. The effectiveness of the AAV9-hPLN vector to over-express the WT PLN was assessed in HET treated mice, resulting in a 15% increase in WT PLN gene expression compared to saline injected mice, along with a corresponding increase in PLN protein levels. Functionally, this resulted in a significant reduction in arrhythmia in AAV-treated mice compared to controls. Conclusion AAV-mediated gene therapy has demonstrated sufficient efficiency in promoting the over-expression of the target gene in a preclinical model for PLN-R14del cardiomyopathy. A 15% increase in WT-PLN led to a significant improvement of the arrhythmogenic phenotype of HET mice, suggesting that even a modest restoration of PLN expression can reduce disease-related arrhythmogenicity and serve as a potential therapeutic strategy.
Gaar-Humphreys et al. (Sat,) conducted a other in PLN-R14del induced cardiomyopathy. AAV9-wtPLN vector vs. saline control was evaluated on Percentage of arrhythmic beats induced by epinephrine and caffeine. AAV-mediated gene therapy over-expressing wild-type PLN significantly reduced stress-induced arrhythmia in a preclinical mouse model of PLN-R14del cardiomyopathy.