Prevalence of rare missense TTN variants in a cohort of patients with cardiomyopathy

Cardiomyopathies refer to myocardial disorders in which the heart muscle has an abnormal structure and function, in absence of hypertension and, coronary artery, valvular and congenital heart disease. Based on morpho-functional phenotypes, cardiomyopathies are classified into five categories: hypertrophic (HCM), dilated (DCM), restrictive (RCM), non-dilated left ventricular (NDLVC) and arrhythmogenic right ventricular (ARVC). Their most relevant hallmarks are genetic and clinical heterogeneity, variable expressivity and incomplete penetrance. Among the genes involved in cardiomyopathies, TTN plays a central role due to its structural function within the striated muscle sarcomeres. Recessive and dominant variants of the TTN gene are associated with skeletal and cardiac myopathies, respectively. Specifically, dominant titin truncation variants (TTNtv) are the most frequent genetic cause of human DCM, and are predisposition alleles in HCM, NDLVC, ARVC and RCM. Moreover, heterozygous TTNtv are also encountered in ∼1 % of the general population, possibly non-affected carriers of recessive myopathies. At the moment, there is emerging understanding of the functional consequences of TTNtv, however, for TTN missense variants (TTNmv) only a potential biological role has been suggested and, in the clinical practice, their interpretation still represents a challenge. To date, only three TTNmv (namely the p.A178D, p.W976R, and p.C3575S) have definitively been associated to DCM, mainly based on the robust cosegregation with this disease. To get more insight into the role of non-truncating TTN variants, this study was aimed to define the prevalence of rare TTNmv, predicted pathogenic by in silico algorithms, in a large cohort of individuals affected by cardiomyopathy. The study involved the participation of 16 Italian medical genetics laboratories that perform the DNA analyses by target massive parallel sequencing as part of their routine clinical service. The capturing of the genomic regions of interest was predominantly performed through a probes-based approach, with only about 1 % of the sequences' enrichment performed by PCR-amplicons. The involved laboratories retrospectively reviewed the TTN gene sequencing data obtained from all the cases collected in each center's cohort, including both patients with a cardiomyopathy phenotype (classified as Cardio) and cases not affected by cardiomyopathy and analyzed by exome sequencing for other clinical indications (classified as non-Cardio). The data collection period covered a time from 2015 to middle of 2023 and concerned the DNA sequencing of 84,525 individuals. Out of those, 9038 (11 %) were Cardio, while the remaining 75,487 (89 %) were non-Cardio cases. In total, 161 TTNmv that were rare (absent in the GnomAD 2.1 database) and predicted pathogenic (Revel score > 0.644) were identified. Those selected variants were labelled as TTNmv* (Table 1). The data were summarized using descriptive statistics (counts and percentages). We calculated the prevalence of TTNmv* and 95 % confidence intervals with the Wilson's exact method. Between group Cardio and non-Cardio, to evaluate the differences of TTNmv* localization and Revel score, we performed the chi square and the Mann-Whitney U tests. Statistical significance was set at 0.05. All the analyses were performed using the statistical software STATA v.17 (Supplemental material).