CRISPR/Cas9‐mediated Genome Editing in Patient‐Derived iPSC‐Cardiomyocytes Recapitulates an MYH6‐R443P Phenotype in a HLHS Family

Abstract

Hypoplastic Left Heart Syndrome (HLHS) is a clinically and anatomically severe form of Congenital Heart Disease (CHD). Although prior studies suggest that HLHS has a complex genetic inheritance, its etiology remains largely unknown. We previously demonstrated that rare variants in the α‐myosin heavy chain (MYH6) gene are significantly enriched in HLHS (p<1×10−5, observed in 10.5% or 20/190 HLHS subjects). Analysis of clinical outcomes showed that survival without cardiac transplant was reduced in HLHS subjects with MYH6 variants (p=6×10−3) as compared to those without MYH6 variants. Transcriptome and protein expression analyses from cardiac tissue revealed differential expression of cardiac contractility genes, notably upregulation of the β‐myosin heavy chain (MYH7) gene in subjects with MYH6 variants. Visualizing α actinin in atria showed intrinsic disrupted sarcomere structure in HLHS patients with MYH6 variants.We observed that patient‐specific induced pluripotent stem cells (iPSCs) were defective in cardiomyogenic differentiation in vitro. Additionally, sarcomere structure in cardiomyocytes derived from iPSCs produced from a family trio (unaffected mother‐affected father‐patient) carrying a variant in MYH6 (MYH6‐R443P) was dysmorphic.We introduced the MYH6‐R443P variant into iPSCs derived from the unaffected parent using CRISPR/cas9 technology and observed that the MYH6‐R443P phenotype in HLHS‐iPSCs was recapitulated; specifically, gene‐edited MYH6‐R443P+/− and MYH6‐R443P−/− iPSCs displayed defective cardiomyogenic differentiation, and sarcomere structure in these cardiomyocytes was dysmorphic. We conclude that the etiology of MYH6‐associated HLHS may be informed utilizing iPSCs.Support or Funding InformationThe Advancing a Healthier Wisconsin Endowment (AHW)Wolfe Family FoundationGreat Milwaukee FoundationChildren's Research InstituteLittle Heart for LifeThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.