Isogenic models of hypertrophic cardiomyopathy unveil differential phenotypes and mechanism-driven therapeutics

作者：J. Bhagwan, D. Mosqueira, Karolina Chairez-Cantu, I. Mannhardt, Sara E Bodbin, Mine Bakar, J. G. Smith, C. Denning · 发表于：Journal of Molecular and Cellular Cardiology · 年份：2020 · DOI：10.1016/j.yjmcc.2020.06.003 · 被引用次数：62 · 研究领域：Biology、Medicine

Background Hypertrophic cardiomyopathy (HCM) is a prevalent and complex cardiovascular condition. Despite being strongly associated with genetic alterations, wide variation of disease penetrance, expressivity and hallmarks of progression complicate treatment. We aimed to characterize different human isogenic cellular models of HCM bearing patient-relevant mutations to clarify genetic causation and disease mechanisms, hence facilitating the development of effective therapeutics. Methods We directly compared the p.β-MHC-R453C and p.ACTC1-E99K HCM-associated mutations in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and their healthy isogenic counterparts, generated using CRISPR/Cas9 genome editing technology. By harnessing several state-of-the-art HCM phenotyping techniques, these mutations were investigated to identify similarities and differences in disease progression and hypertrophic signaling pathways, towards establishing potential targets for pharmacological treatment. CRISPR/Cas9 knock-in of the genetically-encoded calcium indicator R-GECO1.0 to the AAVS1 locus into these disease models resulted in calcium reporter lines. Results Confocal line scan analysis identified calcium transient arrhythmias and intracellular calcium overload in both models. The use of optogenetics and 2D/3D contractility assays revealed opposing phenotypes in the two mutations. Gene expression analysis highlighted upregulation of CALM1, CASQ2 and CAMK2D, and downregulatio...