Clin Res Cardiol (2022). https://doi.org/10.1007/s00392-022-02002-5

Preclinical CRISPR/Cas9 gene therapy in dominant and recessive LZTR1-caused Noonan syndrome patients

A. Busley1, L. B. Böhmer1, M. Kleinsorge2, B. Wollnik3, E. Hammer4, L. Cyganek5, für die Studiengruppe: DZHK

1Kardiologie & Pneumologie, Universitätsmedizin Göttingen, Göttingen; 2Herzzentrum, Klinik für Kardiologie und Pneumologie, Universitätsmedizin Göttingen, Göttingen; 3Instiut für Humangenetik, Göttingen; 4Center for Functional Genomics of Microbes, Greifswald; 5Herzzentrum Göttingen - Stem Cell Unit, Universitätsmedizin Göttingen, Göttingen;

Noonan syndrome (NS) is a multisystem disorder with a broad spectrum of clinical symptoms and varying degrees of disease severity. Common symptoms are low-set ears, dysmorphic facial appearance, a webbed neck, chest deformities and in many cases a severe form of congenital heart disease, e.g. hypertrophic cardiomyopathy (HCM). NS belongs to the spectrum of RASopathies and is causally linked to mutations in a number of genes including PTPN11, SOS1, RAF1 and RIT1, leading to an increased RAS/MAPK signaling pathway activity. In addition, dominant as well as recessive mutations in a newly NS-associated gene called LZTR1 were discovered. As the pathomechanisms behind the role of LZTR1 in the manifestation of HCM are not entirely understood, we investigated patient-derived induced pluripotent stem cells (iPSCs) from NS patients with autosomal dominant and recessive variants in LZTR1. CRISPR/Cas9 genome editing was applied to correct the respective variants. On the one hand, patients’ iPSC-derived cardiomyocytes displayed an increased cell size that could be rescued upon genetic correction. On the other hand, proteomic analysis revealed increased levels of RAS/MAPK- and HCM-associated candidates, such as accumulation of LZTR1-associated interaction partners, indicating a non-functional degradation of RAS isoforms by the LZTR1-CUL3 complex. Correction of the gene variants was able to normalize the signaling activity. Summarizing, our iPSC-CM model consisting of NS patients carrying dominant and recessive variants in LZTR1 could recapitulate major hallmarks of the disease by uncovering patient-specific disease pathomechanisms. Results obtained by these experiments could set the basis for a personalized and clinically translatable CRISPR/Cas9 gene therapy for NS and NS-associated HCM.

https://dgk.org/kongress_programme/jt2022/aP1879.html