PROJECT SUMMARY/ABSTRACT Pediatric calmodulinopathy is a life-threatening cardiac arrhythmia syndrome resulting from a single heterozygous mutation in any of three calmodulin (CaM) genes. Most (68%) persons with a pathogenic CaM variant will experience a major arrhythmic event such as aborted cardiac arrest or sudden cardiac death by the age of five years. Calmodulinopathies result in heterogenous clinical features with the two predominant clinical phenotypes being long QT syndrome (CALM-LQTS) and catecholaminergic polymorphic ventricular tachycardia (CALM-CPVT). Despite current treatments, 56% of calmodulinopathy patients experience breakthrough cardiac events, underscoring the need for improved therapeutics. A current barrier to effective treatment is our incomplete understanding of how calmodulin genetic variants contribute to pathologic phenotypes. Human induced pluripotent stem cells (hiPSCs) can be differentiated into cardiomyocytes (hiPSC-CMs) while maintaining the patient’s genetic background. Application of hiPSC-CMs in the context of this disease allows us to better understand patient-specific heterogeneity at the cellular level. In this study, we will establish a CALM- LQTS hiPSC-CM model using hiPSCs generated from five persons with a pathogenic CaM variant who present with CALM-LQTS. We hypothesize that we can model a CALM-LQTS phenotype using patient-specific hiPSC- CMs, identify clinically appropriate drugs that correct it, and determine underlying mechanisms that drive this phenotype. We will first determine the cellular CALM-LQTS phenotype using blinded electrophysiological, contractility, and calcium imaging studies and then screen a panel of FDA-approved drugs to identify candidates that alleviate this phenotype. We will then perform experiments to identify underlying mechanisms that drive the cellular CALM-LQTS phenotype. We will first determine the contribution of calcium cycling dysregulation, the primary mechanism of calmodulinopathy, in our CALM-LQTS hiPSC-CM model. We will then leverage this model to investigate CALM wildtype and variant allele expression levels to determine if allelic imbalance is contributing the cellular CALM-LQTS phenotype. We will also assess the pathogenicity of each variant on the cellular phenotype by using genome editing. Phenotyping of these edited lines will allow us to determine if these variants are necessary and sufficient for the cellular CALM-LQTS phenotype. Successful completion of these aims will provide a patient-specific methodology to identify necessary clinical treatments for CALM-LQTS and elucidate the drivers of this rare and severe disease. Through the support of their sponsor and their established training plan, the predoctoral fellowship applicant will develop expert knowledge in the field and the necessary skills in experimental design, analysis, communication, and technical techniques to complete this research strategy and transition to the next stage of their translational me...