Relevance of Titin Missense and Non-Frameshifting Insertions/Deletions Variants in Dilated Cardiomyopathy

Abstract

Recent advancements in next generation sequencing (NGS) technology have led to the identification of the giant sarcomere gene, titin (TTN), as a major human disease gene. Truncating variants of TTN (TTNtv) especially in the A-band region account for 20% of dilated cardiomyopathy (DCM) cases. Much attention has been focused on assessment and interpretation of TTNtv in human disease; however, missense and non-frameshifting insertions/deletions (NFS-INDELs) are difficult to assess and interpret in clinical diagnostic workflow. Targeted sequencing covering all exons of TTN was performed on a cohort of 530 primary DCM patients from three cardiogenetic centres across Europe. Using stringent bioinformatic filtering, twenty-nine and two rare TTN missense and NFS-INDELs variants predicted deleterious were identified in 6.98% and 0.38% of DCM patients, respectively. However, when compared with those identified in the largest available reference population database, no significant enrichment of such variants was identified in DCM patients. Moreover, DCM patients and reference individuals had comparable frequencies of splice-region missense variants with predicted splicing alteration. DCM patients and reference populations had comparable frequencies of rare predicted deleterious TTN missense variants including splice-region missense variants suggesting that these variants are not independently causative for DCM. Hence, these variants should be classified as likely benign in the clinical diagnostic workflow, although a modifier effect cannot be excluded at this stage.

Figure 1

Bioinformatic filtering strategy to identify “deleterious” variants. Total TTN variants were filtered using allele frequency (0.0016%) threshold derived from the frequency of known pathogenic DCM variants identified in gnomAD reference individuals. Variants more frequent than derived thresholds in respective sources were not expected to be deleterious.

Figure 2

Allele frequency spectrum of TTN missense variants in DCM and reference individuals. More than 50% of TTN missense variants in DCM patients as well as in reference individuals were private. Potentially pathogenic private and/or rare TTN missense variants were not enriched in DCM patients compared with ExAC and gnomAD reference individuals.

Figure 3

Spatial distribution of rare TTN missense variants in DCM cohort and reference population. Titin is linearly depicted with its 152 Ig-like domains in purple and 132 fibronectin type III domains in green. TTN missense variants identified in patients as well as ExAC and gnomAD reference individuals are shown as coloured bars. Bisque bar represents the number of transcripts (TCI). Variants are shown relative to the Meta transcript position. The dark grey bars indicate exons unique to the Meta transcript. The dashed lines below the number of transcripts schematic indicate the location of variants within the sarcomere.