E Mastantuono (1), R Berutti (2,3), T Meitinger (2,4), CJ Kupatt (1,4), P Gruber(5,6), M Krane (4,5,7,8), KL Laugwitz (1,4), A Moretti (4,9)

1:First Department of Medicine, TUM Klinikum Rechts der Isar, Technical University of Munich, 81675 Munich, Germany; 2:Institute of Human Genetics, TUM Klinikum Rechts der Isar, Technical University of Munich, 81675 Munich, Germany; 3:CRS4, Center for Advanced Studies, Research, and Development in Sardinia, Pixinamanna Ed.1, 09050 Pula, Italy; 4: German Centre for Cardiovascular Research (DZHK), Munich Heart Alliance, 80336 Munich, Germany; 5:Department of Surgery, Yale School of Medicine, New Haven, CT 06510, USA; 6:Department of Genetics, Harvard Medical School, Boston, MA 02115, USA; 7:Department of Cardiovascular Surgery, German Heart Center Munich, School of Medicine and Health, TUM University Hospital, Technical University of Munich, 80636 Munich, Germany; 8:Institute Insure, German Heart Center Munich, School of Medicine and Health, TUM University Hospital, Technical University of Munich, 80636 Munich, Germany; 9:Regenerative Medicine in Cardiovascular Diseases, First Department of Medicine, TUM Klinikum Rechts der Isar, Technical University of Munich, 81675 Munich, Germany

Background:
Hypoplastic left heart syndrome (HLHS) is a leading cause of neonatal mortality, accounting for approximately 25% of cardiac-related deaths within the first week of life. Its etiology remains largely unknown and it involves complex genetic mechanisms. Recent large-scale genomic studies have broadened our understanding of the genetic basis of congenital heart defects (CHDs), highlighting distinct gene functions in dominant and recessive inheritance patterns.

Methods:
We analysed whole exome sequencing (WES) or whole genome sequencing (WGS) data from 100 HLHS patients to investigate damaging recessive (including compound heterozygous) and dominant (de novo and rare inherited heterozygous) genotypes. Gene set enrichment and protein network analyses were performed to identify biological pathways contributing to HLHS pathogenesis.

Results:
In the dominant model analysis, we identified 17,686 variants across 9,397 genes, including 1,271 loss-of-function (LOF) and 12,924 damaging missense variants; 93 were de novo mutations. Gene enrichment analysis revealed significant involvement in pathways related to chromatin remodelling, heart development, cilia and cell projection.
In the recessive model, 1,769 variants were detected in 724 genes, comprising 118 LOF and 1,044 damaging missense variants. Enrichment analysis revealed significant associations with the dynein complex, cell adhesion, and Wnt signalling pathways.

Conclusions:
This study demonstrates that genomic analysis in HLHS patients can uncover distinct biological pathways depending on the mode of inheritance. Our findings highlight mechanisms involved in cardiac development and provide new insights into the complex and poorly understood genetic basis of HLHS.