TTN (titin) gene mutation may not always predict heart disease


More than 750,000 people in the United States have dilated cardiomyopathy, a potentially life-threatening condition in which the heart’s main pumping chamber, the left ventricle, enlarges and grows increasingly weak.

Research has shown that one in 10 people with this condition were born with a mutation in the TTN (titin) gene, but – until now – it has been unclear whether everyone with these mutations will inevitably develop dilated cardiomyopathy.

In a new study published today in Circulation, researchers at Penn Medicine and Geisinger reviewed gene sequences of more than 70,000 people, and found that 95 percent of patients who had the genetic mutations did not have heart disease or signs of cardiac decline.

However, they did find subtle differences in the hearts ability to pump efficiently, compared to those without the mutation.

“It’s clear that these gene mutations have a real effect on one’s heart, and yet, there are a lot of people carrying the deleterious mutations right now who are fine,” said the study’s corresponding author Zoltan Arany, MD, Ph.D., a professor of Cardiovascular Medicine in the Perelman School of Medicine at the University of Pennsylvania.

“While our study moves us one step closer to being able to predict, based on one’s genes, who will get this disease, there is still a difference between these two populations that we don’t yet understand.

The next step will be to identify the specific variable causing some of these patients to get heart disease.”

The TTN gene, which codes the body’s largest protein, acts as a spring inside the heart muscle and helps limit how much it can be stretched.

Mutations in this gene – which affect about 1 percent of the global population – are commonly found in people with dilated cardiomyopathy, when cardiologists order a genetic test to determine if the disease stems from a genetic variant.

In this study, researchers sought to reverse the process, and determine if pinpointing the mutations could predict whether people carrying the faulty genes would develop the disease.

To do so, researchers reviewed the exome sequence data of 61,040 from the Geisinger MyCode Community Health Initiative and 10,273 from the Penn Medicine BioBank to identify individuals with the gene mutations.

From there, they examined corresponding information, including diagnoses, imaging and test results – available via linked electronic health records – to determine whether the patients had heart disease or showed signs of declined cardiac function.

Researchers found that patients with cardiomyopathy who have the mutations fare worse, even on treatment, than patients with the condition who don’t have a mutation.

The finding, researchers say, underscores the value of ordering a genetic test for patients who have been diagnosed with cardiomyopathy.

That said, Arany cautions against patients who don’t have heart disease undergoing genetic tests for titin variants.

“For now, I would not recommend people get genetic testing for the titin variants because that will just make them anxious for something that’s highly unlikely,” Arany said.

“That may change, for example, if we were to find that the combination of a TTN gene mutation and a mutation in another gene causes people to get the disease, then we’d recommend genetic testing for both of the variants.

But right now we still don’t know enough.”

Cardiomyopathies are defined as myocardial disorders in which the heart is structurally and functionally abnormal.

Morphologically defined subtypes include hypertrophic (HCM), dilated (DCM), arrhythmogenic (AC) and left ventricular noncompaction (LVNC) cardiomyopathies12, and each of these subtypes can be genetically mediated (Figure 1).

DCM is characterized by an enlarged and poorly contractile left ventricle (LV).

DCM can be attributed to genetic and nongenetic causes including hypertension, valve disease, inflammatory/infectious causes and toxins3.

Even these “nongenetic” forms of cardiomyopathy, can be influenced by an individual’s genetic profile and, furthermore, mixed etiologies may be present.

In DCM, the degree of LV systolic dysfunction is variable, and LV systolic dysfunction is often progressive. DCM is a major risk factor for developing heart failure (HF) as the presence of reduced systolic function does not imply symptoms.

Notably, DCM is often associated with an increased risk of severe arrhythmia indicating the pathological involvement of the cardiac conduction system.

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Figure 1
Echocardiography demonstrates forms of cardiomyopathyLeft ventricular noncompaction cardiomyopathy (LVNC) is shown in the upper right (arrows indicate deep trabeculations in the left ventricle (LV). Dilated cardiomyopathy (DCM) is defined by enlarged LV diameters (dashed double sided arrow). Hypertrophic cardiomyopathy (HCM) is defined with a thickened LV, including the septum (marked with double sided arrow).

Randomized clinical heart failure trials typically report 30–40% of subjects with a nonischemic DCM compared to ischemic DCM3.

Clinical trials are evaluating interventions to reduce CHF symptoms, these studies may focus on the later stages of disease. Similarly, a recent survey of hospitalized patients in the United States in which the mean age was 75 years (n=156,013) found that ischemic cardiomyopathy was more common than nonischemic (59% compared to 41%)4.

Of nonischemic DCM, hypertension accounted for 48% and idiopathic was the next most common at 31%.

In this study, individuals with nonischemic DCM were more likely to be female, nonwhite, and younger that those with ischemic cardiomyopathy.

The true prevalence of DCM, and of genetically mediated DCM, is not fully known.

An early estimate of DCM prevalence derived from a study carried out from 1975 to 1984 in Olmstead County, MN, USA5.

his epidemiological study relied on echocardiography, angiography or autopsy evaluation of DCM cases and resulted in a prevalence of 36.5/100,000 individuals, or 1 in 2,700 with a male to female ratio of 3.4 in a European-American population5.

The prevalence of DCM varies likely reflecting geographic and ethnic differences, as well as the methodologies used610.

Studies from England (8.3/100,000)11, Italy (7.0/100,000)12, and Japan (14/100,000)9, each report similar DCM prevalence. However, the prevalence of DCM is likely underestimated.

The prior studies relied on older less sensitive imaging modalities13. More recently, Hershberger and colleagues used a different approach to estimate DCM prevalence, based on the known ratio of idiopathic DCM to HCM of approximately 2:1, prevalence estimates of heart failure and prevalence estimates of left ventricular dysfunction as a surrogate for DCM14.

With this approach, a much higher prevalence of DCM is estimated, in the range of 1:250. Similarly, estimates of familial DCM prevalence varies: a meta-analysis of 23 studies found a prevalence estimate of 23% with a range of 2% to 65% indicating a significant heterogeneity in diagnostic criteria, and a frequency progressively increasing over time due to more systematic clinical screening15.

In the clinical practice and in the current guidelines, the prevalence of familial DCM is assumed to be around 30 – 50%13131618.

In patients with familial DCM, approximately 40% has an identifiable genetic cause17.

Also in sporadic DCM pathogenic genetic variants can be identified, although the frequency of genetic causes in this population is not well defined17.

DCM Genetics

The majority of genetic DCM is inherited in an autosomal dominant pattern with variable expressivity and penetrance (Figure 4), although specific forms of autosomal recessive, X-linked recessive and mitochondrial inheritance each occur141956

De novo mutations also contribute to genetic cardiomyopathy and are defined when neither biological parent carries the offspring’s mutation. 

De novo mutations have been described in many different genes, and the presence of a de novo variant can be used to define the pathogenic status of genetic variants since the frequency of de novo variation in each genome is exceedingly rare.

Thus, a novel mutation introducing a protein altering change in a cardiomyopathy gene is typically considered pathogenic.

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Figure 4
A typical DCM pedigree is shown highlighting variable expressivity. Most DCM is inherited in an autosomal dominant pattern. Affected individuals with DCM are shown in black. A gene panel revealed the previously reported pathogenic TPM1 E54K variant. The proband (arrow) presented in early life requiring heart transplant during early childhood. Other members of the family are in their 3rd to 6th decade with LVEF 45%, demonstrating variable expressivity of the primary mutation. Environmental and additional genetic factors may contribute to variable expressivity.

Interpreting whether genetic variants are pathogenic is increasing complex, owing to the vast amount of rare variation in each human genome.

The emerging consensus around interpretation of genetic variation and its effect on phenotype relies on a classification system ranging from pathogenic, likely pathogenic, variant of uncertain significance (VUS), likely benign and benign57.

The availability of large control cohorts provides invaluable information of the frequency of variants, and the largest available data set is currently ExAC (Exome Aggregation Consortium) (, which collected exome sequencing data of over 60,000 individuals from a series of studies including the 1000 Genomes Project and the Exome Sequencing Project (

The current trend is to consider putative pathogenic variants as those that are either unique to the DCM patient or family58, or extremely rare, (MAF <1 × 10−4)59.

The recently adopted, more stringent criteria for genetic testing have prompted the reclassification of variants and indicate the needs of a continuous reanalysis of data58.

The use of whole exome/genome sequencing in clinical laboratories warrants strong criteria to discriminate common variants.

At present, genetic testing typically relies on self-reported ethnicity testing, and it is important to match ethnicity between the proband and testing databases. However, at this point, this integration of common and rare variation is not routinely being used in cardiomyopathy genetic testing, potentially contributing to false positive interpretation60.

DCM is genetically heterogeneous, and DCM genes encode proteins of broad cellular functions. Mutations in genes encoding cytoskeletal, sarcomeric, mitochondrial, desmosomal, nuclear membrane and RNA binding proteins have all been linked to DCM. Thus, the pathological mechanisms that lead to DCM are very diverse. The genes below are listed in order of frequency for their contribution to genetic DCM with focus on the most commonly implicated genes and their mechanism of action if known.


The discovery of the role of TTN truncating variants in DCM has been major advance61. The TTN gene encodes the giant protein titin, which is the largest known protein expressed in the heart. Titin functions as a spring, providing passive force and regulating sarcomere contraction and signaling62.

Titin is a large ~35,000 aa protein that spans half the length of the sarcomere from Z-disc to M-band, and is referred to as a “third” filament with the thin and thick filaments that comprise the sarcomere.

Proposed as a molecular rule for the sarcomere, titin has domains that can accommodate passive stiffness63. Titin’s I band region includes the PEVK (proline-glutamate-valine-lysine) repetitive region, which is thought to directly regulate passive tension.

The I band region of the TTN gene is encoded by 220 of TTN’s 360 exons.

The large size, repetitive nature, and extensive alternative splicing of TTN makes it challenging for genetic analysis. The PEVK region is just carboxyl to the N2A and N2B regions that interact with the FHL (four and half LIM protein), identified as a modifier for HCM64.

Notably, TTN is differentially spliced throughout heart development and adaptively to distinct physiological states including HF65. The larger N2A form is associated with a more compliant ventricle (Figure 5).

In contrast, the smaller N2B form lacks more of the repetitive units and is associated with stiffer heart. Deep RNA sequencing of TTN from failed hearts suggests highly variable exon usage in this region consistent with even subtler defects in cardiac elasticity that may be variable across regions of the LV66.

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Figure 5
Shown in the top is a schematic of the sarcomere with the position of the thick myosin-containing filaments and the thin actin-containing filaments. Titin is considered a third filament of the sarcomere since its spans from Z disk to M band. The lower schematics show the major splice forms of titin (N2-BA, N2-B, N2-A). The green box represents a unique sequence domain. The PEVK region is named for the repetitive amino acid sequences (proline, glutamine, valine, lysine).

Using a TTN specific array designed to capture all TTN exons, it was shown that truncating variants of TTN contribute to 20–25% of nonischemic DCM61.

Prior to this, only a few missense TTN variants had been described linked to DCM67. Induced pluripotent stem cells (iPSC) differentiated into cardiomyocytes in culture demonstrate a paucity of sarcomeres, suggesting that force may be impaired directly through sarcomere loss in TTN truncations68.

In these studies and others it has been shown that TTN truncations are observed at a low frequency in the general population, ranging from 1–3%616669.

There is a tendency for TTN truncations in DCM to distribute to the A band, rather than the I band66, and TTN truncations can also be associated with mild DCM70. A recent study showed that truncating variants in the general population are linked to eccentric cardiac remodeling, suggesting that TTN truncations may be “at-risk” alleles71.

Peripartum cardiomyopathy is a heterogeneous syndrome of mixed etiology. Yet a subset of peripartum cardiomyopathy is attributable to TTN truncating variants7273.

Peripartum cardiomyopathy can be associated with recovered LV function after pregnancy. Moreover, the observation that TTN truncating variants can be associated with recovery of function in DCM after LV assist device (LVAD) placement also suggests a dynamic state of TTN truncating variants74. Additional genes with mutations beyond TTN have also been described in peripartum cardiomyopathy21.

Overall, the presence of TTN truncating variants in the general population argues for caution in interpreting these variants and again underscores the importance of familial segregation analysis. At this point, until more is known, the presence of a TTN truncation variant should trigger at least intermittent cardiac imaging and management aimed at reducing other stressors to the heart.

TTN has a high prevalence of missense variants, both rare and common75TTN missense variants have been reported in ARVC and other forms of cardiomyopathy677578. Additionally, TTN missense variants have also been reported in skeletal myopathy, including the common tibial myopathy7980.

The enormous number of TTN missense variants makes these variants exceedingly complex to interpret in the context of broad genetic testing on individuals with DCM.

Zebrafish have been used successfully to model myopathies due to TTN mutations, demonstrating both cardiac and skeletal muscle defects81.

A mouse models with an in frame deletion in the PEVK region of TTN develops diastolic dysfunction, consistent with the complex role of titin for both systolic and diastolic dysfunction82. In both rats and mice with heterozygous TTN truncation mutations, additional stressors like transaortic constriction are used to promote the development of DCM7183.

Journal information: Circulation
Provided by Perelman School of Medicine at the University of Pennsylvania


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