Amino Acid-Level Signal-to-Noise Analysis Aids in Pathogenicity Prediction of Incidentally Identified TTN-Encoded Titin Truncating Variants.

BACKGROUND: TTN, the largest gene in the human body, encodes TTN (titin), a protein that plays key structural, developmental, and regulatory roles in skeletal and cardiac muscle. Variants in TTN, particularly truncating variants (TTNtvs), have been implicated in the pathogenicity of cardiomyopathy. Despite this link, there is also a high burden of TTNtvs in the ostensibly healthy general population. This complicates the diagnostic interpretation of incidentally identified TTNtvs, which are of increasing abundance given expanding clinical exome sequencing. METHODS: Incidentally identified TTNtvs were obtained from a large referral database of clinical exome sequencing (Baylor Genetics) and compared with rare population variants from genome aggregation database and cardiomyopathy-associated variants from cohort studies in the literature. A subset of TTNtv-positive children evaluated for cardiomyopathy at Texas Children's Hospital was retrospectively reviewed for clinical features of cardiomyopathy. Amino acid-level signal-to-noise analysis was performed. RESULTS: Pathological hotspots were identified within the A-band and N-terminal I-band that closely correlated with regions of high percent-spliced in of exons. Incidental TTNtvs and population TTNtvs did not localize to these regions. Variants were reclassified based on current American College of Medical Genetics and Genomics criteria with incorporation of signal-to-noise analysis among Texas Children's Hospital cases. Those reclassified as likely pathogenic or pathogenic were more likely to have evidence of cardiomyopathy on echocardiography than those reclassified as variants of unknown significance. CONCLUSIONS: Incidentally found TTNtvs are common among clinical exome sequencing referrals. Pathological hotspots within the A-band of TTN may be informative in determining variant pathogenicity when incorporated into current American College of Medical Genetics and Genomics guidelines.

Meta-analysis of cardiomyopathy-associated variants in troponin genes identifies loci and intragenic hot spots that are associated with worse clinical outcomes.

INTRODUCTION: Troponin (TNN)-encoded cardiac troponins (Tn) are critical for sensing calcium and triggering myofilament contraction. TNN variants are associated with development of cardiomyopathy; however, recent advances in genetic analysis have identified rare population variants. It is unclear how certain variants are associated with disease while others are tolerated. OBJECTIVE: To compare probands with TNNT2, TNNI3, and TNNC1 variants and utilize high-resolution variant comparison mapping of pathologic and rare population variants to identify loci associated with disease pathogenesis. METHODS: Cardiomyopathy-associated TNN variants were identified in the literature and topology mapping conducted. Clinical features were compiled and compared. Rare population variants were obtained from the gnomAD database. Signal-to-noise (S:N) normalized pathologic variant frequency against population variant frequency. Abstract review of clinical phenotypes was applied to "significant" hot spots. RESULTS: Probands were compiled (N = 70 studies, 224 probands) as were rare variants (N = 125,748 exomes; 15,708 genomes, MAF <0.001). TNNC1-positive probands demonstrated the youngest age of presentation (20.0 years; P = .016 vs TNNT2; P = .004 vs TNNI3) and the highest death, transplant, or ventricular fibrillation events (P = .093 vs TNNT2; P = .024 vs TNNI3; Kaplan Meir: P = .025). S:N analysis yielded hot spots of diagnostic significance within the tropomyosin-binding domains, α-helix 1, and the N-Terminus in TNNT2 with increased sudden cardiac death and ventricular fibrillation (P = .004). The inhibitory region and C-terminal region in TNNI3 exhibited increased restrictive cardiomyopathy (P =.008). HCM and RCM models tended to have increased calcium sensitivity and DCM decreased sensitivity (P < .001). DCM and HCM studies typically showed no differences in Hill coefficient which was decreased in RCM models (P < .001). CM models typically demonstrated no changes to Fmax (P = .239). CONCLUSION: TNNC1-positive probands had younger ages of diagnosis and poorer clinical outcomes. Mapping of TNN variants identified locations in TNNT2 and TNNI3 associated with heightened pathogenicity, RCM diagnosis, and increased risk of sudden death.