Abstract 4344136: The Genetic Basis of Early Mortality in Neonates with Single Ventricle Disease: An NC-DEFINE Prospective Observational Cohort Study

Background: Single ventricle disease (SVD) is the most severe form of congenital heart disease. Despite surgical advances improving survival, heart failure (HF) remains a key contributor to early morbidity and mortality, especially in the first months of life. Defining genetic drivers of HF in SVD could enable early risk stratification and guide precision therapies to improve outcomes. Hypothesis: Ultra-rare variants in genes associated with dilated cardiomyopathy (DCM) increase HF risk in neonates with SVD. Approach: Neonates ≤21 days old with SVD were prospectively enrolled and followed at Duke (n=40). An additional 69 individuals (0.2–65y) from Duke and UNC formed an ambispective cohort. Chromosomal abnormalities were excluded. Ultra-rare (MAF<e⁻⁴) variants in DCM genes were manually classified as likely pathogenic/pathogenic (LP/P) or of uncertain significance (VUS) per ACMG criteria. Multimodal deep clinical phenotyping was performed. HF was defined as severe (VAD, transplant, or death) or medically managed (MM; EF ≤40% and/or HF therapy escalation). Findings were validated in a blinded analysis of an independent SVD cohort from Nationwide Children’s Hospital (NCH; n=36) and compared to the AllofUs population. Results: In the prospective cohort, 8 (20%) neonates developed severe HF, 11 (28%) developed MM HF, and 21 (52%) remained HF free, with mean follow-up of 2 years. Hosting a DCM-associated LP/P variant was linked to an 11-fold increased risk of severe HF ( P =0.0009), while VUSs increased MM HF risk 6-fold ( P =0.01). Most HF occurred within the first month. Findings were independently validated in the NCH cohort, where LP/P variants reduced freedom from severe HF ( P =0.0007). Associations were attenuated in the ambispective cohort, suggesting survivor bias and underscoring the importance of early detection and risk stratification. Compared with the AllofUs cohort, the prospective cohort had a higher prevalence of LP/P variants ( P <0.0001) but similar VUS burden, supporting a model in which LP/P variants drive primary disease risk, while low-penetrant variants may modify susceptibility in the context of SVD. Conclusion: This study provides the first prospective evidence linking DCM-associated variants to significant risk for early-onset HF in SVD. These findings, independently validated in external cohorts, underscore the potential for genetic screening to inform early risk stratification and family counseling in this high-risk population.