Intronic CRISPR Repair in a Preclinical Model of Noonan Syndrome–Associated Cardiomyopathy

Author:

Hanses Ulrich12,Kleinsorge Mandy12,Roos Lennart12,Yigit Gökhan32ORCID,Li Yun3,Barbarics Boris42,El-Battrawy Ibrahim25,Lan Huan5,Tiburcy Malte62ORCID,Hindmarsh Robin12,Lenz Christof78ORCID,Salinas Gabriela3ORCID,Diecke Sebastian2910,Müller Christian3,Adham Ibrahim3,Altmüller Janine11,Nürnberg Peter11,Paul Thomas42,Zimmermann Wolfram-Hubertus6212ORCID,Hasenfuss Gerd1212,Wollnik Bernd3212ORCID,Cyganek Lukas12ORCID

Affiliation:

1. Clinic for Cardiology and Pneumology (U.H., M.K., L.R., R.H., G.H., L.C.)

2. DZHK (German Center for Cardiovascular Research), partner site Göttingen, Mannheim and Berlin, Germany (U.H., M.K., L.R., G.Y., B.B., I.E-B., M.T., R.H., S.D., T.P., W.-H.Z., G.H., B.W., L.C.).

3. Institute of Human Genetics (G.Y., Y.L., G.S., C.M., I.A., B.W.)

4. Clinic for Pediatric Cardiology and Intensive Care Medicine (B.B., T.P.)

5. First Department of Medicine, Medical Faculty Mannheim, University Medical Centre Mannheim, University of Heidelberg, Mannheim, Germany (I.E-B., H.L.)

6. Institute of Pharmacology and Toxicology (M.T., W-H.Z.)

7. Institute for Clinical Chemistry (C.L.), University Medical Center Göttingen, Germany.

8. Bioanalytical Mass Spectrometry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany (C.L.).

9. Stem Cell Core Facility, Max Delbrück Center for Molecular Medicine, Berlin, Germany (S.D.).

10. Berlin Institute of Health, Germany (S.D.).

11. Cologne Center for Genomics, University of Cologne, Germany (J.A., P.N.).

12. Cluster of Excellence “Multiscale Bioimaging: from Molecular Machines to Networks of Excitable Cells” (MBExC), University of Göttingen, Germany (W-H.Z., G.H., B.W.).

Abstract

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–mitogen-activated protein kinase 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 as clinically the most prevalent symptom caused by biallelic mutations within the leucine zipper-like transcription regulator 1 ( LZTR1 ). We generated induced pluripotent stem cell–derived cardiomyocytes of the affected siblings and investigated the patient-specific cardiomyocytes on the molecular and functional level. Results: Patients’ induced pluripotent stem cell–derived cardiomyocytes recapitulated the hypertrophic phenotype and uncovered a so-far-not-described causal link between LZTR1 dysfunction, RAS–mitogen-activated protein kinase 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 (clustered regularly interspaced short palindromic repeats) repair and demonstrated a rescue of the hypertrophic phenotype. Conclusions: Our study revealed the human cardiac pathogenesis in patient-specific induced pluripotent stem cell–derived cardiomyocytes 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 hypertrophic cardiomyopathy.

Publisher

Ovid Technologies (Wolters Kluwer Health)

Subject

Physiology (medical),Cardiology and Cardiovascular Medicine

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