Abstract Body (Do not enter title and authors here): Background: PRKAG2 encodes the γ2 regulatory subunit of AMP-activated protein kinase (AMPK), which modulates AMPK activity. Chronic AMPK activation has been implicated in hypertrophic cardiomyopathy, myocardial glycogen accumulation, and arrhythmogenesis. Therapeutic silencing of PRKAG2 may offer a disease-modifying approach by mitigating maladaptive AMPK activation.

Aims: To evaluate whether targeted silencing of PRKAG2 mitigates the pathological effects of chronic AMPK activation in a mouse model of pressure overload-induced cardiac dysfunction.

Methods: An antibody-oligonucleotide conjugate (AOC) delivering siRNA targeting PRKAG2 was used to attenuate AMPK activation. The AMPK pathway was activated by subjecting mice to chronic cardiac stress by transverse aortic constriction (TAC). Cardiac morphology and function were assessed by echocardiography and cardiac gene expression was assessed by qPCR.

Results: AOC treatment enabled efficient and homogeneous delivery of siRNA targeting PRKAG2 throughout the myocardium, resulting in a dose-dependent reduction in PRKAG2 expression, achieving over 70% mRNA knockdown at a 3 mg/kg dose. In TAC mice, PRKAG2 expression was elevated by approximately 30% in the vehicle group relative to sham but was reduced by more than 80% following AOC administration. This knockdown was associated with attenuation of AMPK target protein phosphorylation, confirming effective modulation of AMPK signaling. Functionally, AOC-treated TAC mice demonstrated a ~30% improvement in fractional shortening and a ~30% reduction in left ventricular diameter, alongside a ~10% decrease in heart weight compared to vehicle treated TAC mice. Importantly, SERCA2a gene expression was partially preserved, supporting improvements in cardiac remodeling, contractile function, and calcium handling.

Conclusions: AOC-mediated knock-down of PRKAG2 expression can attenuate the pathological consequences of chronic AMPK activation. These results support the development of PRKAG2-targeted therapies for cardiomyopathies associated with maladaptive AMPK signaling, such as PRKAG2 syndrome.