All consecutive patients with early-onset cardiomyopathy (≤ 18 years) who were evaluated at Hospital Sant Joan de Déu Barcelona, Spain, between 2014 and 2024 were examined. Only pediatric patients clinically diagnosed with NSCM (HCM, DCM, and ACM), were included. All diagnosed cardiomyopathies were reevaluated according to the ESC 2023 guideline recommendations.1 NSCM was defined as the absence of extracardiac features suggesting syndromic, metabolic, neuromuscular, or storage diseases, based on appropriate negative investigations when indicated. Patients with cardiomyopathy secondary to syndromic disorders were excluded from the analyses. Pediatric patients carrying P or LP variants in NSCM-associated genes who did not yet meet the criteria for a definitive clinical diagnosis, including those with borderline phenotypes, were excluded. The study was approved by the institutional ethics committee review board (PIC-37-22). Written informed consent for genetic testing and for the use of clinical and genetic data for research was obtained from the patients’ legal representatives, and assent was obtained from minors when appropriate according to age and local regulations.

At diagnosis, echocardiographic parameters were extracted from clinical reports. In HCM, maximal left ventricular (LV) wall thickness (interventricular septum/left ventricular posterior wall) and z-scores (based on Pediatric Heart Network reference standards)21 were recorded. In DCM, LV end-diastolic diameter with z-scores and left ventricular ejection fraction (LVEF), assessed by the Simpson method, were collected. In ACM, available structural criteria were documented. Cardiac magnetic resonance imaging was performed based on clinical indication, and myocardial fibrosis was assessed by late gadolinium enhancement (LGE), recorded as present or absent when available.

Genomic deoxyribonucleic acid was extracted from all patients. Index cases underwent comprehensive next-generation sequencing panel testing, while relatives initially received Sanger testing for the familial P or LP variant, with panel testing reserved for those with early-onset severe phenotypes. Ultimately, all individuals were analyzed using the same next-generation sequencing approach, namely a cardiomyopathy gene panel covering all genes currently implicated in familial cardiomyopathies (Methods of the supplementary data).

Identified variants or alterations, including single-nucleotide variants, small insertions or deletions (indels), and copy-number variants, were reported according to the nomenclature recommended by the Human Genome Variation Society, using transcripts reported in RefSeq. Variants were classified following the recommendations of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology.22 Only rare variants classified as P or LP were considered clinically actionable.1,14 Variants classified as having an ambiguous role, namely variants of uncertain significance (VUS), were also included in our study due to their potential deleterious or modulatory role, despite not being clinically actionable. All available data for each rare genetic variant were updated until the submission date (February 2025).

Demographic, clinical, and genetic data were extracted from electronic medical records following ethical and legal protocols approved by our institution that ensure strict data anonymization. A descriptive analysis of the total cohort was performed. Categorical variables are described using frequency tables, and numerical variables are described as mean± standard deviation or median [interquartile range (IQR)], depending on the normality of their distribution as evaluated by the Kolmogorov-Smirnov test. Kaplan-Meier analysis with the log rank test (Mantel-Cox) and Cox models were used to assess differences in survival curves between 2 or more groups.

Clinical and genetic variables were compared between patients with and without CE using Cox proportional hazards models, reporting hazard ratios (HR). Statistical significance was defined as a 2-sided P-value <.05, and the confidence interval was set at 95% (95%CI). In cases in which a patient experienced multiple CEs, only the age at the first CE was considered for analysis, and subsequent events were censored.

To assess whether baseline phenotype improved prediction of CE beyond genetic testing alone, incremental discrimination was evaluated using the integrated discrimination improvement. This analysis was performed in subgroups in which phenotypic variables were significant in Cox models. A genetic model including genetic burden alone was compared with an integrated model combining genetic burden and baseline phenotypic features. Predicted risks were derived from Cox models at a prespecified 5-year time horizon, and integrated discrimination improvement estimates with 95%CI were obtained using bootstrap resampling.

CEs were defined as ventricular tachycardia or fibrillation, appropriate implantable cardioverter-defibrillator shocks, heart failure stages C and D,23 heart transplantation, SCD, and sudden cardiac arrest. Descriptive and statistical analyses were performed using IBM SPSS Statistics, version 30 (IBM Corp., United States).

At baseline, HCM was the most frequent diagnosis with a median maximal left ventricular wall thickness z-score at diagnosis of 2.1 [IQR 1.09-3.77]; overall, maximal left ventricular wall thickness was not significantly associated with CEs (P=.145). In DCM, patients with CEs showed a more severe baseline phenotype, characterized by greater LV dilation and lower systolic function: LV end-diastolic diameter z-score (HR, 1.22; 95%CI, 1.05-1.43; P <.010) and LVEF (HR, 0.91; 95%CI, 0.87-0.96; P <.001). Finally, in ACM, myocardial fibrosis/LGE was frequent; however, the presence of fibrosis was not significantly associated with CEs (P=.710). Overall, CE patients displayed a more severe baseline phenotype in DCM (greater LV dilation and lower LVEF), whereas maximal left ventricular wall thickness in HCM was not significantly different between CE and non-CE groups (table 1). In DCM, adding baseline phenotype (LV end-diastolic diameter z-score and LVEF) to a genetics-only model significantly improved discrimination for CEs (integrated discrimination improvement, 0.528; 95%CI, 0.312-0.683; P <.001).

Cardiac magnetic resonance imaging was performed in 71.4% of patients in the cohort (60 of 84). Because magnetic resonance imaging data were extracted from clinical reports, only LGE-positive cases, indicating fibrosis, are detailed in table S1, whereas the prevalence estimates below include all patients with available LGE status. Overall, myocardial fibrosis assessed by LGE was present in 31 of 84 patients. By phenotype, LGE was detected in 22 of 53 patients with HCM (41.5%), 2 of 21 patients with DCM (9.5%), and 7 of 10 patients with ACM (70%). Among all patients with eoNSCM, LGE was observed more frequently in those with CEs compared with those without CEs, although this difference did not reach statistical significance (P=.168). In exploratory analyses, LGE also appeared more prevalent among patients carrying multiple rare variants (≥ 2 P or LP variants or 1 P or LP variant+VUS/s) compared with those carrying a single P or LP variant (18 of 31; 58.1% vs 13 of 31; 26%, respectively), but the sample size was limited to perform multivariable analysis (table 1).

The genetic analysis identified a definitive P or LP variant in 35 of 53 pediatric patients (66%), of whom 27 of 35 (77%) had a positive family history of the disease. In HCM, 21 of 53 pediatric patients (39.6%) had more than 1 variant identified. Of these, 6 patients exhibited a second variant classified as P or LP, whereas 15 patients carried a second variant classified as a VUS. No disease-causing variants were identified in 6 of 53 children (11%), 4 of whom were sporadic cases (figure 3; table S1).

Figure 3.

Most common genes in which variants were identified in pediatric patients with early-onset nonsyndromic cardiomyopathies. The x-axis displays the distribution of patients among the 3 clinical diagnostic groups, and the y-axis displays the genetic diagnostic yield and the genes in which the main variants were identified. Three genes, MYBPC3, MYH7 and TNNT2, accounted for 59% of genetically diagnosed pediatric patients with HCM. Three genes, MYH7, TTN, and TNNT2, accounted for 48% of pediatric patients with DCM. DSP and PKP2 accounted for 70% of patients with ACM. ACM, arrhythmogenic cardiomyopathy; CE, cardiac event; DCM, dilated cardiomyopathy; HCM, hypertrophic cardiomyopathy.

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An etiologic genetic diagnosis was established in 10 of 21 pediatric patients (48%), with a positive family history in 8 of 10 (80%). In DCM, 8 of 10 patients (80%) presented with multiple genetic variants, but only 1 patient had an autosomal recessive form, carrying a homozygous pathogenic variant in TNNI3. The remaining patients carried a second variant classified as a VUS. No disease-causing variants were identified in 1 of 21 pediatric patients (5%) (figure 3; table S1).

A definitive genetic diagnosis was established in 7 of 10 pediatric patients (70%), all with a positive family history. In ACM, 6 of 7 patients (86%) had multiple genetic variants, but all secondary variants identified were classified as VUS. No disease-causing variants were identified in 1 patient (10%) (figure 3; table S1).

In our cohort, 40.5% of patients with a definitive genetic result (34 of 84) harbored more than 1 genetic variant in the analyzed panel. Among these, 7 carried 2 P or LP variants and 24 carried a P or LP variant alongside 1 or more VUS (table 1). During the follow-up (6 [IQR: 4-10] years), 24 of 84 patients (29%) with eoNSCM showed CE. These events included sudden cardiac arrest (10 of 24; 42%), ventricular tachycardia/fibrillation (2 of 24; 8%), appropriate implantable cardioverter-defibrillator shocks (4 of 24; 17%), SCD (2 of 24; 8%), and heart failure stages C and D (6 of 24; 25%) 3 of whom received heart transplants (3 of 24; 13%; 1 HCM; and 2 DCM). The diagnoses of patients with CE included 12 of 24 with HCM (50%), 9 of 24 with DCM (34%), and 3 of 24 with ACM (13%). All children carrying 2 variants classified as P or LP in the genetic testing had CE (7 of 24; 29%). In total, 21 of 24 patients (88%) exhibiting CE were carriers of more than 1 variant in the genetic test (7 patients with 2 P or LP variants; 11 patients with 1 P or LP variant and 1 or more VUS, and 3 patients with multiple VUS) (figure 4; table S1).

Figure 4.

Flow diagram showing cardiac events among pediatric patients with eoNSCM. Total cohort of pediatric patients with cardiomyopathies (left; red): pediatric patients with HCM (light blue); DCM (light green); and ACM (grey). Groups are separated according to the number of variants identified by genetic testing (positive with> 1 genetic variants identified, only 1 variant identified, or negative result). The number of cardiac events is shown on the right (rose) for each group. ACM, arrhythmogenic cardiomyopathy; CE, cardiac event; DCM, dilated cardiomyopathy; eoNSCM, early-onset nonsyndromic cardiomyopathies; HCM, hypertrophic cardiomyopathy.

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Among patients with a definitive genetic result, the presence of additional genetic variants was associated with an increased risk of CE. Cox regression analysis revealed a significant association between carrying multiple genetic variants and an elevated risk of CE (HR, 7.27; 95%CI, 2.47-21.44; P <.01). This association was observed both in patients carrying 2 P or LP variants (P <.001) and in those carrying 1 P/LP variant together with 1 or more VUS (HR, 3.54; 95%CI, 1.22-11.17; P <.05), compared with individuals carrying only a single P or LP variant. Kaplan-Meier curves illustrating CE occurrence are shown in survival analyses (figure 5).

Figure 5.

Time-to-event survival analysis. A: survival analysis comparing patients with eoNSCM carrying multiple genetic variants (2 P or LP variants, high risk) and a definitive clinical/genetic diagnosis vs carriers of a single variant. The analysis demonstrated a significant association (log-rank test, P <.01). A single genetic variant (red); 2 P or LP variants (light blue). B: survival analysis comparing patients with eoNSCM and a definitive clinical/genetic diagnosis carrying 1 P or LP variant and another variant(s) classified as VUS. Further stratification revealed significant differences between groups (P <.05); Hazard ratio, 3.54; 95% confidence interval, 1.22-11.17. A single genetic variant (red); 1 P or LP plus 1 or more VUS variants (light blue). eoNSCM, early-onset nonsyndromic cardiomyopathies; P, pathogenic; LP, likely pathogenic; VUS, variant of uncertain significance.

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Twenty-four pediatric patients in this cohort (29%) exhibited CE during follow-up. Only 2 of these patients (8%) had de novo variants in the genes MYH7 (c.602T> A; p.(Ile201Thr)) and TNNT2 (c.299T> A; p.(Ile100Asn)), both previously reported.26,27 In contrast to earlier pediatric studies including syndromic cardiomyopathies, in which de novo variants accounted for most severe outcomes,11 children in our eoNSCM cohort who developed CEs more frequently carried multiple genetic variants. In our cohort specifically, 7 of 24 patients (29%) were identified as carrying 2 P or LP variants. These patients showed severe phenotypes and CE despite optimized treatment: 6 had a diagnosis of HCM and 1 had a diagnosis of DCM. These findings are consistent with prior reports in which coinheritance of multiple P or LP variants explained severe childhood-onset cardiomyopathies.11 Regarding HCM, the only cardiomyopathy for which pediatric studies are available, the overall CE rate in our cohort was comparable to that reported in pediatric-onset HCM series, in which approximately 25% of patients experience events during follow-up.20,28–33 In those studies, LGE, a well-established adverse prognostic marker in adult HCM, was also associated with an increased risk of CE. In contrast, LGE was not significantly associated with CE risk in our cohort, which may be attributable to the retrospective study design and the lack of quantitative fibrosis assessment, as only the qualitative presence or absence of fibrosis was evaluated.

In the entire cohort, only early age at onset and the presence of multiple genetic variants, including at least 1 causative variant, were significantly associated with an increased risk of CE. This association was significant when additional variants were classified as VUS and was even more pronounced when they were P or LP. Notably, all pediatric patients carrying 2 P or LP variants experienced a CE, while those carrying 1 P or LP variant plus a VUS had up to a 3.5-fold higher risk compared with carriers of a single P or LP variant. This complex genetic background may contribute to the substantial clinical heterogeneity and incomplete penetrance observed in NSCM, even within the same family. In our cohort, pediatric patients with CE were found to carry the same P or LP variant as an unaffected or mildly affected parent, as well as siblings with identical causal variants but divergent clinical presentations. These severe phenotypes were associated with the presence of additional rare variants in the same gene or in other cardiomyopathy-related genes. Collectively, these findings support a complex genetic model in which digenic and/or compound variants drive disease expression, while additional genetic factors could modulate penetrance and phenotypic severity.34

Beyond diagnostic considerations, the prognostic relevance of genetic burden was assessed through integrated genotype and phenotype analyses. An improvement in discrimination was observed exclusively in DCM. However, given the limited number of cases and events, together with the retrospective study design, these findings should be interpreted with caution and require confirmation in larger multicenter cohorts.

Recent evidence in adult DCM further supports this concept, demonstrating that genotype influences not only diagnosis but also prognosis, treatment response, and long-term remodeling, resulting in worse outcomes in specific genetic subgroups.35 Although biallelic mutations and multiple P or LP variants are established markers of severe disease, the consistent observation of complex genotypes (involving genetic background) in high-risk pediatric cases strengthens the relevance of our findings.16–19,36 The contribution of VUS to risk may reflect their heterogeneous nature, encompassing variants that are potentially deleterious but not yet fully characterized. Therefore, while insufficient to cause disease independently, some VUS may act as phenotypic modifiers, influencing disease severity in genetically vulnerable hearts. This concept is further supported by genome-wide association studies, showing that both rare and common variants contribute to CE risk through polygenic risk scores.10,37,38

By contrast, in our cohort, CE were rare among genetically negative patients; only 1 genotype-negative child with clinically diagnosed DCM showed severe early-onset disease, in whom myocarditis and syndromic conditions had been excluded. This observation highlights the potential value of comprehensive genomic approaches, such as trio exome or genome sequencing, in selected cases, as targeted gene panels may fail to detect novel or currently unrecognized disease-associated genes. Taken together, these results reinforce the clinical usefulness of genetic testing beyond diagnosis, emphasizing its role in refining genotype-phenotype correlations and improving risk stratification in pediatric patients. Young people with eoNSCM carrying multiple P or LP variants represent a particularly high-risk group and may benefit from intensified surveillance and preventive strategies. Furthermore, VUS should be periodically re-evaluated to determine their clinical relevance, alongside established clinical risk factors specific to each cardiomyopathy subtypes.7,39,40

Our work has important clinical implications. First, our findings support current recommendations for comprehensive clinical and genetic screening of relatives with cardiomyopathy, including pediatric patients, as outlined in the latest guidelines1. In pediatric cases with severe phenotypes (very early onset, extreme hypertrophy or marked LV dysfunction, and/or early CE), a full genetic evaluation may be warranted even if a P or LP variant has already been identified in the familial proband (with adult-onset disease) to identify additional variants that may explain the anticipation or severity of the phenotype. Patients diagnosed with NSCM who have more than 1 P or LP variant identified on genetic testing are at high risk of CE and should be properly stratified to implement preventive measures to decrease the risk of fatal events. Finally, we emphasize the need for close collaboration and structured transition between pediatric and adult care units to ensure accurate diagnosis, continuity of care, and optimal outcomes.

Limitations of our study include its single-center, retrospective design, which may introduce selection bias, as our institution is a tertiary referral center that typically evaluates more complex or severe cases. The small sample size within some clinical diagnostic groups, particularly DCM and ACM, further limits the generalizability of the findings. Another limitation is the restricted assessment of nongenetic modifiers that could influence the progression of NSCM. This study proposes an initial approach to genetic testing for risk stratification in pediatric patients with eoNSCM. Nevertheless, the results need to be validated in larger cohorts. We performed genetic analysis using a targeted gene panel rather than whole exome or genome sequencing, which limits the identification of novel causal genes. However, this was not the aim of the present study.