Genes linked to dilated cardiomyopathy with strong evidence.
Dilated cardiomyopathy (DCM) is a myocardium disease characterized by left ventricular dilation and systolic dysfunction. Genetic susceptibility contributes significantly to the disease progression in familial DCM. Mutations in more than fifty different genes have been identified to cause DCM, accounting for up to 50% of familial DCM cases. Elucidation of genetic basis for the remaining familial DCM probands promises to substantially increase the efficiency of genetic testing for early disease diagnosis and intervention. Dissecting genetic pathways linked to DCM and related pathogenic mechanisms can provide valuable insights into the understanding of disease pathophysiology that can be leveraged for development of genotype-targeted therapeutic strategy. Here, we review genetic variants, with a focus on affected genes most commonly implicated in DCM, and highlight their underlying pathophysiological mechanisms of action. We discuss recent progress on gene-based therapeutic strategy which holds the opportunities to implement individualized medicine and ultimately to improve patient outcome in the future.
- dilated cardiomyopathy
- genetic determinant
- genetic testing
- gene therapy
Heart failure afflicts about 26 million patients worldwide. The estimated economic burden related to heart failure is 120 billion US dollars . Dilated cardiomyopathy (DCM), referred to a group of heart muscle disorders characterized by heart chamber dilation and systolic dysfunction, is a common cause of heart failure. DCM is the most common form of non-ischemic cardiomyopathy and the most common indication in patients who need heart transplantation [2, 3]. In the pediatric, nearly 50% of patients dying suddenly or undergoing cardiac transplantation are affected by cardiomyopathy, predominantly by DCM . In the young, DCM is the most frequent cause of heart failure . In the adult, it is the second most common cause of heart failure (after the coronary heart disease), underlying about one third of all heart failure cases . While the true prevalence for DCM in general population is not fully defined yet due to lack of well-designed large-scale population based studies, its estimated prevalence ranges from 1 in 2500 to 1 in 250 people . The annual incidence of DCM is approximately 7 cases per 10,000 individuals, and males are about 3 times more frequently affected than females [2, 7, 8].
Classically, DCM is defined based on two major criteria: 1) left LV fractional shortening <25% (> 2 standard deviations [SD]) and/or LV ejection fraction <45% (>2 SD); and 2) LV end-diastolic diameter greater than 117% of the predicated value corrected for age and body surface area. DCM is diagnosed when any other known cause of myocardial diseases are excluded . The updated definitions of DCM are left ventricular or biventricular systolic dysfunction and dilatation that are not explained by abnormal loading conditions or coronary artery disease .
DCM is caused by a variety of etiologies, including acquired, genetic and/or mixed origins. Acquired risk factors such as infection, myocarditis, drug toxin, autoimmune response, excess alcohol consumption and metabolic disorders are recognized to cause DCM. Primary DCM results when all these acquired factors are exclude, which can be either idiopathic or familial. Familial DCM (FDC) is classified when two or more family members are diagnosed in first-degree relatives. The prevalence of familial DCM differs in different patient cohorts, estimated to range from 20–50% of all DCM cases. The remaining DCM cases are thus classified as idiopathic [6, 11]. However, the frequency of familial DCM is believed to be underestimated, due to the limitation of large pedigree and family’s availability for diagnostic screening. Genetic factors which predispose to DCM have been increasingly recognized. Since the discovery of the first disease causative gene to cardiomyopathy , to date, more than 50 genes have been identified to associate with DCM and the number is still increasing. Genetic mutations in all these genes combined can explain about 40–50% of familial DCM cases, underpinning the genetic determinant of this disease . The familial DCM appears to be inherited as a monogenic trait and is mainly transmitted in an autosomal dominant inheritance pattern, manifesting incomplete, age-related penetrance and variable expression. Other patterns such as autosomal recessive, X-linked and mitochondrial inheritance also occurs in a small portion of the familial DCM cases [6, 13].
Current management of DCM is mainly focused on treating patients with symptoms following the standard guidelines, similar to treating other forms of heart failure with reduced ejection fraction (EF). The guideline driven therapies mostly adapt a “one-size-fits-all” approach that uses β-adrenergic blockers, angiotensin-converting enzyme inhibitors, or angiotensin receptor blockers to improve cardiac function and symptom by reducing congestion and management of arrhythmia [14, 15]. Recently, several new additions of armamentarium are also implemented in the treatment options [16, 17, 18]. Despite of advances in these treatment protocols, DCM remains one of the major reasons for patients needed for heart transplantation, and the morbidity and mortality rate of DCM still remains unacceptably high. Thus, the current management of DCM with the “one-size-fits-all” strategy is challenged. More novel and effective and individualized treatment options are desirable.
Considering the genetic determinant of DCM, and up to 50% of familial DCM patients have a genetic origin, genotype-targeted therapies, by directly targeting at the specific gene mutations, have emerged as promising strategies toward development of more effective and individualized treatment. In this chapter, we firstly review definitive genes linked to DCM and classify them based on their intracellular localization (Figure 1), with a major focus on genes most commonly implicated in DCM, and highlight their underlying pathophysiological mechanism of action if known (Table 1). Next, we discuss progress and challenges on the emerging genotype-based therapeutic strategies for effective and individualized medicine explored in the treatment of DCM (Table 2), which hold the opportunities to ultimately improve patient outcome in the future.
|Gene||Protein||Function||Frequency and overlapping phenotype||Inheritance pattern|
|Alpha cardiac actin||Structural component of thin filament||<1% of DCM; HCM, LVNC||Autosomal dominant|
|Beta-myosin heavy chain||Cardiac muscle contraction||5% of DCM; HCM, ACM, LVNV||Autosomal dominant|
|Cardiac troponin C||Cardiac muscle contraction||<1% of DCM; HCM, LVNC||Autosomal dominant|
|Cardiac troponin I||Cardiac muscle contraction||<1% of DCM; HCM||Autosomal recessive|
|Cardiac troponin T||Cardiac muscle contraction||3% of DCM; HCM, ACM, LVNC||Autosomal dominant|
|Tropomyosin alpha-1 chain||Cardiac muscle contraction||1–2% of DCM; HCM, LVNC||Autosomal dominant|
|Titin||Sarcomere scaffold||15–25% of DCM; HCM||Autosomal dominant|
|Desmin||Contractile force transduction||<1% of DCM; Desminopathies||Autosomal dominant|
|Dystrophin||Contractile force transduction||<1% of DCM; Duchenne/Becker muscular dystrophy||X-linked|
|δ-sarcoglycan||Structural component of dystrophin-glycoprotein complex||<1% of DCM; HCM, LGMD2E||Autosomal recessive|
|Emerin||Nuclear membrane anchorage||<1% of DCM; EMDM||X-linked|
|Lamin A/C||Nuclear envelope structure||6% of DCM; EMDM, LGMD1B||Autosomal dominant|
|RNA-binding protein 20||Regulator of cardiac gene splicing||2% of DCM||Autosomal dominant|
|Sodium voltage-gated channel alpha subunit 5||Sodium channel protein||2–3% of DCM; LQTs Brugada syndrome||Autosomal dominant|
|BCL2-associated athanogene 3||Co-chaperone, inhibition of apoptosis||3% of DCM; Myofibrillar myopathy||Autosomal dominant|
|Flamin-C||Actin filament crosslinking||<1% of DCM; HCM, RCM||Autosomal dominant|
|Desmoplakin||Structural component of desomosome, cell–cell mechanotransmission||2% of DCM; ACM||Autosomal recessive|
|DnaJ heat shock protein family homolog, C19||Protein transporter||<1% of DCM; 3-Methylglutaconic aciduria, type V||Autosomal recessive|
|Tafazzin||Phospholipid transacylase||<1% of DCM; HCM, Barth syndrome||X-linked|
|Cardiac phospholamba||Regulator of calcium pump||<1% of DCM; HCM, ACM||Autosomal dominant|
|Gene||Variant type||Clinical phenotype||Molecular pathophysiology||Genotype-based therapy||Reference|
|DCM, DMD, Becker muscular dystrophy, premature death||Absence of functional dystrophin protein, replacement of muscle by fibrotic and adipose tissue, contraction weakness||1) Exon skipping; 2) Mini dystrophin gene replacement; 3) CRISPRA/Cas9 genome editing; 4) Stop codon readthrough||[60, 61, 62, 67, 71, 72, 73, 80]|
|DCM with high penetrance, high risk of arrhythmia, early lethality||Lamins A and C proteins haploinsufficiency, nuclear malformations, biomechanical defects, activation of p38 kinase pathway||1) Preventive therapy, lower the threshold for cardiac defibrillator implantation; 2) p38 inhibition; 3) Trans-splicing; 4) Stop codon readthrough||[29, 30, 58, 75, 79]|
|Truncating||DCM, HCM, ventricular arrhythmias||Titin protein haploinsufficiency, sarcomere insufficiency, metabolic and energetic adaption, increased sensitivity to mechanical stress||Exon-skipping|||
|Ventricular dilation, contractile dysfunction and ventricular arrhythmias, heart failure by middle age||Abnormal PLN protein subcellular localization, calcium handling defects, electrical instability||TALEN genome editing||[22, 84]|
|Deletion||DCM, HCM||δ-sarcoglycan protein deficiency, sarcolomma instability, increased myocyte apoptosis, reduced expression of miRNA-669a||1) Gene replacement; 2) microRNA overexpression||[83, 88, 89]|
2. Genetic causes of familial DCM
Extensive studies on genetic basis of DCM underscored profound heterogeneous nature of DCM disease. DCM variants mutate genes encoding a broad spectrum of proteins with distinct functions and intracellular localization have been identified (Figure 1 and Table 1). For example, mutations in genes encoding sarcomere protein involving in mechanosensing and force transmission, encoding nuclear envelope proteins required for the protection of biochemical forces, encoding desmosomal proteins required for maintaining the structural integrity of desomosome, encoding ion channels and molecules involving in calcium handling, encoding chaperone proteins, transcription factors and RNA-binding proteins have all been identified to cause DCM. Specific variants in these genes may alter various signaling pathways and cellular structures in cardiomyocyte that can disrupt the mechanism of cardiac muscle contraction and function, leading to common phenotypes of DCM.
2.1 Sarcomere genes
Sarcomere is the basic contractile unit mainly composed of thin and thick filaments. Mutations in sarcomere genes are one of the most important causes of DCM, which have been identified in about one third of DCM [19, 20, 21]. In addition to cause DCM, mutations in sarcomere genes often lead to overlapping phenotypes of hypertrophic cardiomyopathy (HCM), another major genetic type of cardiomyopathy characterized by ventricular wall thickness and diastolic dysfunction. Genes with definitive evidence, supported by studies from multiple centers/groups, with both clinical linkage data and confirmative data from animal models, include
2.2 Nucleus and nuclear envelope genes
Mutations in gene encoding nucleus or nuclear envelope localized proteins that definitively link to DCM such as
The RNA binding motif protein 20 (
2.3 Cytoskeletal protein coding genes
Other genes such as
Mutations in the gene encoding dystrophin (
2.4 Z-disk gene
The Z-disk is an anchoring plane for the actin (thin) filaments to attach and stabilize in the sarcomere. Mutations in many Z-disk-associated proteins coding genes result in cardiac disorders.
2.5 Ion channel gene
Mutations in the ion channel coding gene
2.6 Other DCM genes
Mutations in genes other encoding the sarcoglycans (α, β, γ, δ) were also identified to cause DCM. The sarcoglycans are transmembrane proteins mainly expressed in heart and skeletal muscle that interact with dystrophin. α-, β-, γ-, and δ-sarcoglycans form the sarcoglycan complex that is key components of the dystrophin-associated glycoprotein complex, conferring structural integrity and stability to the sarcolemma through connecting the muscle fiber cytoskeleton to the extracellular matrix, and protecting muscle fibers from mechanical stress during muscle contraction. Mutations in sarcoglycans coding genes cause primary limb-girdle muscular dystrophy presented with early onset muscle weakness and associate with significant DCM . Notably, mutations in the δ- sarcoglycan coding gene lead to DCM without involvement of obvious muscular dystrophy phenotypes .
Mutations in nuclear encoded mitochondrial genes such as
3. Genetic testing
The advent of next-generation sequencing enables cost-effective genetic testing in familial DCM which can define the precise genetic cause of disease. Genetic testing can also help optimize risk stratification and assess prognostics of patients and their relatives. With the identification of a pathogenic mutation and early diagnostic certainty, clinical management of affect individuals could be tailored and patients’ survival can be improved. One best example is related to clinical practice of early intervention of DCM patients with L
Based on the Genetic Testing Registry (https://www.ncbi.nlm.nih.gov/gtr/), there are from 40 to 80 genes included in the testing panels for most commercially available genetic testing for DCM. In familial DCM, the yield of genetic testing, resulting in identification of pathogenic mutations, can thus far reach up to 40%, in a comparable level to that of other inherited cardiac disorders such as HCM and long QT syndrome. The sensitivity of genetic testing is compromised partially due to that not all genes implicated in DCM are included in the gene panels for testing. This is especially the case for those identified from sporadic DCM cases or single family. Furthermore, limited by human genetics approaches that heavily rely on pedigree availability and candidate gene approaches, variants for more than half of familial DCM cases have not been identified yet. As technique advances in genetics of cardiomyopathy, identification of the remaining genetic causes in inherited DCM cases and elucidation of the underlying pathogenic mechanisms leading to the phenotype are evolving rapidly. Targeted gene panels for genetic testing are increasing in an unprecedented scale. With further characterization and functional validation, the ever expanding gene panels of genetic testing promise to increase the rate of positive identification and provide individuals and families with a more comprehensive and conclusive genetic testing.
4. Genotype-targeted therapies
By directly targeting at specific pathogenic mutations causing DCM, genotype-targeted genetic approaches have emerged as promising strategies for effective and individualized therapies to ameliorate disease phenotypes. To carry out genotype-based therapy, many customized genetic strategies were explored. For examples: for loss-of-function mutations that cause reduced or insufficient protein levels, the straightforward gene replacement strategy can be employed. In this scenario, full-length or partial functional cDNA for corresponding mutated gene can be transferred to cardiac tissue to supplement the reduced gene dosage using appropriate gene delivery approach. For mutations that cause dominant-negative effects on a particular gene, exon-skipping or trans-splicing approaches can be considered to remove or modify the mutant transcripts. For pathogenic mutations that cause other protein dysfunction, the highly efficient CRISPRA/Cas9 (Clustered regularly interspaced short palindromic repeats/CRISPR-associated endonuclease) system can be explored to directly correct the mutant variants. In addition, other genotype-based therapeutic strategies such as manipulation of the downstream pathways evoked by specific DCM mutations were also explored. Below, we summarize several examples of gene-targeted therapeutic strategies that produced encouraging results in the treatment of DCM (Table 2), which hold the opportunities to ultimately improve patient outcome in the future.
4.1 Dystrophin mutations-targeted gene therapies
Mutations in the dystrophin (
Gene replacement strategy involves delivering a functioning gene to replace or supplement the mutant gene to the target organ and cells to ease the disease phenotype caused by genetic mutation. This approach often utilizes the adeno-associated virus (AAV) as a vector to mediate gene transfection into the skeletal and cardiac muscle cells. However, as the dystrophin gene has 79 exons and its transcript is about 14 Kb, its size is too large to fit in currently available gene construction vector for gene transfection. Alternatively, a mini- or micro- dystrophin gene coding for a functionally similar to dystrophin but smaller in size was thus used. The authors Wang and colleagues firstly defined the minimal functional region of the dystrophin protein, later referred as mini-dystrophin. The authors then packed this min-dystrophin gene into an AAV vector to mediate muscle transfection and demonstrated the effectiveness of gene delivery and restoration of dystrophin gene function . After this study, several other independent groups latter confirmed the effectiveness of a shortened albeit functional dystrophin gene replacement strategy mediated by the AAV gene delivery system in preservation of cardiac and skeletal muscle function and extending the lifespan in the dystrophic mice [66, 67]. Notably, further studies in animal models of muscular dystrophy and human patients detected certain immunologic responses to the shortened dystrophin peptide, which need to be carefully considered in future clinical application [68, 69, 70].
Exon skipping is an RNA-based splice-switching approach that causes cells to “skip” the exon containing the pathogenic variant, resultant in-frame transcripts that produce a shorter peptide, albeit still at least partially functional protein to ameliorate disease phenotypes. This approach was initially developed to mask the pathogenic mutations containing exon 51 using an antisense oligonucleotide (AON), resulting in genetic correction of the open reading frame of the
With the revolutionary development of the CRISPRA/Cas9 genome editing technologies that enable precise and efficient genetic modifications from single-nucleotide alternation, insertion of gene of interest to deletion of chromosomal regions, numerous studies have demonstrated the feasibility and high efficiency of this technique in restoration dystrophin expression and function in skeletal and cardiac muscle, both in vitro and in vivo [74, 75]. For example, a recent study carried out by El Refaey and colleagues packaged SaCas9 (Cas9 from
mutations-tailored therapeutic strategy LMNA
A genetic approach referred as trans-splicing approach was initially carried out to correct
To target at the
truncating variants-targeted gene therapy TTN
While the pathogenicity of
To employ exon skipping strategy for treating
4.4 Genotype-targeted therapies for other DCM genes
Mutations in the δ- sarcoglycan coding gene (
An earlier version of genome editing technique referred as transcription activator-affected nuclease (TALEN) was explored to directly correct the
The ultimate goal of elucidating genetic basis for DCM is for early disease diagnosis, early disease intervention and treatment. To date, pathogenic mutations in more than 50 genes are associated with DCM. However, these mutations can only explain about 40–50% of familial DCM cases, and the genetic architecture of DCM still remains incompletely understood. Future research directions rely on technique advances for identification of the remaining up to 60% of genetic causes in familial DCM cases. After identifying the genetic causes, functional characterization and validation studies are needed to confirm the pathogenic variants. Extensive studies on delineation of genotype–phenotype relationships are necessary to address currently unmet issues for translational research. Further dissecting genetic pathways linked to DCM and elucidating the pathogenic mechanisms leading to the phenotype can provide valuable insights into the understanding of disease pathophysiology, laying solid foundation for future development of groundbreaking therapeutics.
Gene-based therapies, such as gene replacement, exon skipping and recent evolutionarily developed CRISPRA/Cas9 based genome-editing techniques that directly target at the underlying genetic mutations responsible for the DCM disease progression, have led to several clinical trials which have produced promising results. Thus, the last decade has witnessed encouraging advances in the development of genotype-targeted, personalized therapies. While the progress is promising, several technical issues need to be thoroughly assessed before implementation in clinical practice. For example, current gene delivery system mostly adapt the AAV system because of its many advantages such as long term transgene expression and choice of appropriate serotype for heart enriched expression. One primary limitation for this system, however, is that there exists certain level of anti-AAV antibodies in general human population causing the immunogenicity issue which can potentially lead to development of myocarditis [70, 90, 91, 92]. While the efficiency for exogenous delivery of preprocessed RNAs or oligonucleotides for trans-splicing or exon skipping needs to be further improved, a major concern of off-target issue needs to be seriously addressed for the highly efficient AAV9 mediated CRISPR/Cas9 genome editing system. Novel approaches are desirable to comprehensively evaluate any potential off-target mutagenesis in the heart and other tissues for future clinical application.
Conflict of interest
The authors declare no conflict of interest.