Truncations of Titin Causing Dilated Cardiomyopathy
Herman DS, Lam L, Taylor MR, et al, N Engl J Med 2012 Feb; 366;7:619-28
Reviewers: Sean Birmingham, MD and Dalia Banks, MD
UC San Diego Medical Center, San Diego, CA
Background
A well-accepted important cause of cardiomyopathy is genetic mutation. A genetic cause of dilated cardiomyopathy (DCM) has been implicated, with 30-50% of patients with DCM having a relative with DCM. Over 40 genetic mutations, primarily encoding the sarcomere or cytoskeleton, have been associated with DCM. Titin, the largest human protein with 33,000 amino acids, has been linked to DCM. Titin is critical for sarcomere assembly and modulation of the contractile force. The gene encoding this sarcomere protein is TTN. It has been incompletely studied due to its large size, technical sequencing challenges and extensive cost. Recent advances in sequencing techniques allow for more rapid sequencing of long stretches of DNA without the expense of previous techniques. This study attempted to use next generation sequencing as well as traditional techniques to identify distinct TTN mutations in patients with DCM, hypertrophic cardiomyopathy (HCM) and patients with cardiomyopathy.
Methods
The authors identified 312 patients with idiopathic dilated cardiomyopathy within three cohorts, A, B, and C, 231 subjects with HCM, and 249 control subjects without cardiomyopathy. Genomic DNA was collected from each subject and used to construct DNA libraries. The 145 kb TTN gene was enriched by filter-based hybridization. Group C had the TTN sequenced by traditional Sanger dideoxy sequencing while next-generation sequencing techniques were used on groups A and B, patients with HCM and the controls. Statistical analysis included Fisher’s exact test, exact conditional tests of independence, or goodness-of-fit tests for association, cross-cohort and cross group analysis. Clinical characteristics in each DCM cohort were compared using two-tailed, unpaired t-tests. Two-point lod scores for 19 families with DCM were calculated and indicated disease penetrances.
Results
There were no significant differences between subjects with and without TTN truncating mutations with regards to age at diagnosis, left ventricular end-diastolic dimensions, ejection fraction, rates of cardiac transplantation, implantation of LVAD or death. 97% of the targeted bases were sequenced at least 20 times in DCM cohorts A and B, subjects with HCM and controls. 951 rare TTN variants in the 792 subjects were identified that were predicted to change the titin amino acid sequence. 72 unique TTN truncating variants were identified in 67 subjects with DCM, 3 with HCM, and 7 control subjects. They analyzed nonsense, frameshift, splicing and copy number variants and classified these variants as TTN truncating variants. The three subjects with HCM had three TTN truncating variants but also had pathogenic mutations in established HCM genes. The frequency of variants between HCM and control subjects did not significantly differ (1% and 3% respectively). TTN variants were significantly more frequent among DCM subjects than among HCM subjects (P=3X10-16) and control subjects (P=3X10-14). Overall frequencies of TTN truncating mutations in DCM subjects in group A were 28% and group B 24%. Group C, which used an older sequencing technique, had a 9% TTN mutation rate. TTN mutation type did not influence rates of cardiac transplantation, LVAD implantation or death among subjects or family members due to cardiac causes. Segregation analysis in 20 families confirmed the coinheritance of truncating mutations and that the penetrance of TTN mutations was greater than 95% for patients over 40 years old. They also found that men with TTN mutations had adverse events at a significantly earlier age than women (P=4X10-5).
Conclusion
The authors concluded that TTN truncating mutations are the most common known genetic cause of DCM. They felt that these mutant alleles produce shortened titin protein with abnormal properties that cause dilated cardiomyopathy. They also concluded that TTN truncating mutations rarely if ever cause HCM as the three subjects with TTN variants also had mutations in established HCM genes. The reduced frequency of mutations in group C suggests that detection of mutations was better with next generation sequencing techniques than with traditional dideoxy sequencing.
Comments
This study is the first to clearly show that the mutation in the TTN gene can account for 25% of DCM in patients with a known family history and about 18% of DCM cases without known family history in a large sample population. Previously, only a very few number of TTN mutations have been clearly linked to DCM due to difficulty in comprehensive sequencing of the TTN gene. An interesting finding was that men had adverse events at an earlier age than women. The “suggestion that sex would substantially influence an autosomal monogenic cause of heart failure is unexpected.” The authors acknowledge, “further study of the functional consequences of TTN truncating mutations on myocardial physiological features and myocyte signaling is warranted.” The role of next-generation sequencing in clinical genetic screening has yet to be determined. The authors propose that detection of DCM should increase by 50% by using next generation sequencing analyses in clinical genetic screening. Identifying these mutations should make it easier to find family members with undiagnosed disease. This should allow for earlier DCM diagnosis, implementation of interventions to prolong the asymptomatic period, earlier monitoring for arrhythmias and potentially slow down disease progression.
