Preclinical evaluation of CRISPR-based therapies for Noonan syndrome caused by deep-intronic LZTR1 variants

• Main Authors: Carolin Knauer (Author), Henrike Haltern (Author), Eric Schoger (Author), Sebastian Kügler (Author), Lennart Roos (Author), Laura C. Zelarayán (Author), Gerd Hasenfuss (Author), Wolfram-Hubertus Zimmermann (Author), Bernd Wollnik (Author), Lukas Cyganek (Author)
• Format: Book
• Published: Elsevier, 2024-03-01T00:00:00Z.
• Subjects: article
• Online Access: Connect to this object online.
• Notes: 2162-2531; 10.1016/j.omtn.2024.102123
• Keywords: MT: RNA/DNA Editing; cardiomyocytes; CRISPR-Cas9; gene therapy; genome editing; hypertrophic cardiomyopathy; Therapeutics. Pharmacology; RM1-950
• Source: Molecular Therapy: Nucleic Acids, Vol 35, Iss 1, Pp 102123- (2024)

Gene variants in LZTR1 are implicated to cause Noonan syndrome associated with a severe and early-onset hypertrophic cardiomyopathy. Mechanistically, LZTR1 deficiency results in accumulation of RAS GTPases and, as a consequence, in RAS-MAPK signaling hyperactivity, thereby causing the Noonan syndrome-associated phenotype. Despite its epidemiological relevance, pharmacological as well as invasive therapies remain limited. Here, personalized CRISPR-Cas9 gene therapies might offer a novel alternative for a curative treatment in this patient cohort. In this study, by utilizing a patient-specific screening platform based on iPSC-derived cardiomyocytes from two Noonan syndrome patients, we evaluated different clinically translatable therapeutic approaches using small Cas9 orthologs targeting a deep-intronic LZTR1 variant to cure the disease-associated molecular pathology. Despite high editing efficiencies in cardiomyocyte cultures transduced with lentivirus or all-in-one adeno-associated viruses, we observed crucial differences in editing outcomes in proliferative iPSCs vs. non-proliferative cardiomyocytes. While editing in iPSCs rescued the phenotype, the same editing approaches did not robustly restore LZTR1 function in cardiomyocytes, indicating critical differences in the activity of DNA double-strand break repair mechanisms between proliferative and non-proliferative cell types and highlighting the importance of cell type-specific screens for testing CRISPR-Cas9 gene therapies.