Intronic CRISPR repair in a preclinical model of Noonan syndrome-associated cardiomyopathy

BACKGROUND: Noonan syndrome (NS) is a multisystemic developmental disorder characterized by common, clinically variable symptoms, such as typical facial dysmorphisms, short stature, developmental delay, intellectual disability as well as cardiac hypertrophy. The underlying mechanism is a gain-of-function of the RAS-MAP kinase (MAPK) signaling pathway. However, our understanding of the pathophysiological alterations and mechanisms, especially of the associated cardiomyopathy, remains limited and effective therapeutic options are lacking. METHODS: Here, we present a family with two siblings displaying an autosomal recessive form of NS with massive hypertrophic cardiomyopathy (HCM) as the clinically most prevalent symptom caused by biallelic mutations within the leucine zipper like transcription regulator 1 (LZTR1). We generated induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) of the affected siblings and investigated the patient-specific CMs on the molecular and functional level. RESULTS: The patients' iPSC-CMs recapitulated the hypertrophic phenotype and uncovered a so far not described causal link between LZTR1 dysfunction, RAS-MAPK signaling hyperactivity, hypertrophic gene response and cellular hypertrophy. Calcium channel blockade and MEK inhibition could prevent some of the disease characteristics, providing a molecular underpinning for the clinical use of these drugs in patients with NS, but might not be a sustainable therapeutic option. In a proof-of-concept approach, we explored a clinically translatable intronic CRISPR repair and demonstrated a rescue of the hypertrophic phenotype. CONCLUSIONS: Our study revealed the human cardiac pathogenesis in patient-specific iPSC-CMs from NS patients carrying biallelic variants in LZTR1 and identified a unique disease-specific proteome signature. In addition, we identified the intronic CRISPR repair as a personalized and in our view clinically translatable therapeutic strategy to treat NS-associated HCM.