Cardiomyocyte-Derived Cell-Free DNA in Hypertrophic Cardiomyopathy

Introduction: In hypertrophic cardiomyopathy (HCM), wall stress and subclinical myocardial injury related in part to pathological hypercontractility, which in turn are associated with worse outcomes. Cell-free DNA (cfDNA) is released from cardiomyocytes (cfDNA-CM) through irreversible cellular injury and death. We investigated the feasibility of quantifying cfDNA-CM and its relationship to HCM patient characteristics. Methods: We analyzed 57 plasma samples from stable patients with HCM and 10 plasma samples from patients with acute myocardial infarction (AMI). Plasma cfDNA was isolated from both groups. Whole genome bisulfite sequencing (WGBS) libraries were generated and sequenced with short-read sequencing. Methylation patterns were used for tissue deconvolution analysis and cell-specific cfDNA quantification. Results: Yields of cfDNA (p = 0.003) and cfDNA-CM (p = 0.002) fractions were higher in AMI controls than HCM, supporting the validity of the assay. Within the HCM cohort, the median cfDNA yield was 12.22 ng [IQR: 7.77, 18.94]. The cfDNA-CM fraction in this cohort ranged from 0% to 6.4% of total cfDNA. Higher cfDNA-CM tertiles were associated with higher body mass index (BMI, p=0.001), a greater prevalence of BMI 30 kg/m2 (p<0.001), and a higher prevalence of obstructive sleep apnea (p=0.038). No significant differences were observed in other demographic, genetic, and laboratory characteristics. Higher cfDNA-CM tertile was numerically associated with higher New York Heart Association (NYHA) functional class and left ventricular outflow tract (LVOT) obstruction, while circulating biomarkers of wall stress (NT-proBNP) and myocardial injury (troponin) did not differ across tertiles (p=0.83 and p=0.29, respectively). Conclusion: Isolation and quantification of cfDNA as well as detection of cfDNA-CM in chronic stable patients with HCM are feasible. Higher cfDNA-CM fraction was associated with increasing BMI and cardiometabolic comorbidity burden. The absence of association with traditional circulating biomarkers suggests that cfDNA-CM may reflect a distinct domain of myocardial health, injury, and adverse remodeling.