Abstract 18689: Building a High-throughput Assay for Arrhythmogenic Cardiomyopathy Modifier Pathways in the Zebrafish

Abstract

Introduction: The arrhythmogenic cardiomyopathies (ACs) are rare inherited heart muscle disorders with a high incidence of ventricular tachyarrhythmias and sudden death. One prominent feature of ACs is variable penetrance that implicates major genetic and epigenetic modifiers of disease expression. We sought to develop a robust model of AC in the zebrafish to enable high-throughput genetic and chemical screens for important modifier pathways in ACs. Methods and Results: We generated a zebrafish AC model using cardiac-specific expression of the 2057del2 mutation in human plakoglobin using the GAL4/UAS transactivation system. Plakoglobin mutant fish exhibit a unique phenotype with mild bradycardia, cardiomegaly, and peripheral edema in early adulthood and substantial mortality by the time maturity is reached (survival 45% vs 77% for WT p<0.01). Using qPCR we confirmed an upregulation of natriuretic peptide b (nppb), in mutant expressing fish compared to non-expressing siblings (2 fold, p<0.01). To adapt this line for high-throughput screening, we mated the founders with an existing nppb:luciferase reporter line that we have previously validated and is compatible with 96-well plates. Native nppb expression was strongly correlated with semi-automated luciferase data in this reporter line (average count/embryo ± SEM 202 ± 15 in mutant vs 117 ± 12 in WT, P<0.001). We have gone on to screen a library of 4800 bioactive compounds for disease modifiers and to date have identified both suppressor compounds (10) and compounds that exacerbate the disease response (14). One suppressor of the disease phenotype (SB2) attenuates nppb expression, bradycardia and contractility in the mutant, while also improving survival at 28 days (survival 1.5 μ M SB2 62% and 74% 3 μ M SB2 vs untreated 46%, p<0.01). Conclusion: These data support the feasibility of high-throughput approaches to the modeling of complex cardiovascular disorders in zebrafish, as well as the use of such models to identify novel chemical probes and candidate drug leads for major disease pathways. One compound identified in the screen has already demonstrated efficacy in long-term use in AC models. Ultimately, screens of type will enable personalized disease modeling for a broad range of human diseases.