The application of next-generation sequencing to study congenital heart disease (CHD) is increasingly providing new insights into the causes and mechanisms of this prevalent birth anomaly. ...Whole-exome sequencing analysis identifies damaging gene variants altering single or contiguous nucleotides that are assigned pathogenicity based on statistical analyses of families and cohorts with CHD, high expression in the developing heart and depletion of damaging protein-coding variants in the general population. Gene classes fulfilling these criteria are enriched in patients with CHD and extracardiac abnormalities, evidencing shared pathways in organogenesis. Developmental single-cell transcriptomic data demonstrate the expression of CHD-associated genes in particular cell lineages, and emerging insights indicate that genetic variants perturb multicellular interactions that are crucial for cardiogenesis. Whole-genome sequencing analyses extend these observations, identifying non-coding variants that influence the expression of genes associated with CHD and contribute to the estimated ~55% of unexplained cases of CHD. These approaches combined with the assessment of common and mosaic genetic variants have provided a more complete knowledge of the causes and mechanisms of CHD. Such advances provide knowledge to inform the clinical care of patients with CHD or other birth defects and deepen our understanding of the complexity of human development. In this Review, we highlight known and candidate CHD-associated human genes and discuss how the integration of advances in developmental biology research can provide new insights into the genetic contributions to CHD.
Abstract Over the last quarter-century, there has been tremendous progress in genetics research that has defined molecular causes for cardiomyopathies. More than a thousand mutations have been ...identified in many genes with varying ontologies, therein indicating the diverse molecules and pathways that cause hypertrophic, dilated, restrictive, and arrhythmogenic cardiomyopathies. Translation of this research to the clinic via genetic testing can precisely group affected patients according to molecular etiology, and identify individuals without evidence of disease who are at high risk for developing cardiomyopathy. These advances provide insights into the earliest manifestations of cardiomyopathy and help to define the molecular pathophysiological basis for cardiac remodeling. Although these efforts remain incomplete, new genomic technologies and analytic strategies provide unparalleled opportunities to fully explore the genetic architecture of cardiomyopathies. Such data hold the promise that mutation-specific pathophysiology will uncover novel therapeutic targets, and herald the beginning of precision therapy for cardiomyopathy patients.
Cardiomyopathies have unresolved genotype-phenotype relationships and lack disease-specific treatments. Here we provide a framework to identify genotype-specific pathomechanisms and therapeutic ...targets to accelerate the development of precision medicine. We use human cardiac electromechanical in-silico modelling and simulation which we validate with experimental hiPSC-CM data and modelling in combination with clinical biomarkers. We select hypertrophic cardiomyopathy as a challenge for this approach and study genetic variations that mutate proteins of the thick (MYH7
) and thin filaments (TNNT2
, TNNI3
) of the cardiac sarcomere. Using in-silico techniques we show that the destabilisation of myosin super relaxation observed in hiPSC-CMs drives disease in virtual cells and ventricles carrying the MYH7
variant, and that secondary effects on thin filament activation are necessary to precipitate slowed relaxation of the cell and diastolic insufficiency in the chamber. In-silico modelling shows that Mavacamten corrects the MYH7
phenotype in agreement with hiPSC-CM experiments. Our in-silico model predicts that the thin filament variants TNNT2
and TNNI3
display altered calcium regulation as central pathomechanism, for which Mavacamten provides incomplete salvage, which we have corroborated in TNNT2
and TNNI3
hiPSC-CMs. We define the ideal characteristics of a novel thin filament-targeting compound and show its efficacy in-silico. We demonstrate that hybrid human-based hiPSC-CM and in-silico studies accelerate pathomechanism discovery and classification testing, improving clinical interpretation of genetic variants, and directing rational therapeutic targeting and design.
BACKGROUND:No pharmacological therapy exists for calcific aortic valve disease (CAVD), which confers a dismal prognosis without invasive valve replacement. The search for therapeutics and early ...diagnostics is challenging because CAVD presents in multiple pathological stages. Moreover, it occurs in the context of a complex, multi-layered tissue architecture; a rich and abundant extracellular matrix phenotype; and a unique, highly plastic, and multipotent resident cell population.
METHODS:A total of 25 human stenotic aortic valves obtained from valve replacement surgeries were analyzed by multiple modalities, including transcriptomics and global unlabeled and label-based tandem-mass-tagged proteomics. Segmentation of valves into disease stage–specific samples was guided by near-infrared molecular imaging, and anatomic layer-specificity was facilitated by laser capture microdissection. Side-specific cell cultures were subjected to multiple calcifying stimuli, and their calcification potential and basal/stimulated proteomes were evaluated. Molecular (protein–protein) interaction networks were built, and their central proteins and disease associations were identified.
RESULTS:Global transcriptional and protein expression signatures differed between the nondiseased, fibrotic, and calcific stages of CAVD. Anatomic aortic valve microlayers exhibited unique proteome profiles that were maintained throughout disease progression and identified glial fibrillary acidic protein as a specific marker of valvular interstitial cells from the spongiosa layer. CAVD disease progression was marked by an emergence of smooth muscle cell activation, inflammation, and calcification-related pathways. Proteins overrepresented in the disease-prone fibrosa are functionally annotated to fibrosis and calcification pathways, and we found that in vitro, fibrosa-derived valvular interstitial cells demonstrated greater calcification potential than those from the ventricularis. These studies confirmed that the microlayer-specific proteome was preserved in cultured valvular interstitial cells, and that valvular interstitial cells exposed to alkaline phosphatase–dependent and alkaline phosphatase–independent calcifying stimuli had distinct proteome profiles, both of which overlapped with that of the whole tissue. Analysis of protein–protein interaction networks found a significant closeness to multiple inflammatory and fibrotic diseases.
CONCLUSIONS:A spatially and temporally resolved multi-omics, and network and systems biology strategy identifies the first molecular regulatory networks in CAVD, a cardiac condition without a pharmacological cure, and describes a novel means of systematic disease ontology that is broadly applicable to comprehensive omics studies of cardiovascular diseases.
The heart either hypertrophies or dilates in response to familial mutations in genes encoding sarcomeric proteins, which are responsible for contraction and pumping. These mutations typically alter ...calcium-dependent tension generation within the sarcomeres, but how this translates into the spectrum of hypertrophic versus dilated cardiomyopathy is unknown. By generating a series of cardiac-specific mouse models that permit the systematic tuning of sarcomeric tension generation and calcium fluxing, we identify a significant relationship between the magnitude of tension developed over time and heart growth. When formulated into a computational model, the integral of myofilament tension development predicts hypertrophic and dilated cardiomyopathies in mice associated with essentially any sarcomeric gene mutations, but also accurately predicts human cardiac phenotypes from data generated in induced-pluripotent-stem-cell-derived myocytes from familial cardiomyopathy patients. This tension-based model also has the potential to inform pharmacologic treatment options in cardiomyopathy patients.
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•Altered myofilament Ca2+ sensitivity induces heart growth via tension generation•Increased or decreased myofilament tension induces calcineurin and general growth•Increased myofilament tension induces ERK1/2 to mediate HCM concentric remodeling•Modeling the tension integral predicts disease in familial cardiomyopathy patients
A tension-integrated model successfully predicts the clinical cardiac phenotype of familial cardiomyopathic patients and informs pharmacologic treatment options.
Sarcomere cardiomyopathies are genetic diseases that perturb contractile function and lead to hypertrophic or dilated myocardial remodeling. Identification of preclinical mutation carriers has ...yielded insights into the earliest biomechanical defects that link pathogenic variants to cardiac dysfunction. Understanding this early molecular pathophysiology can illuminate modifiable pathways to reduce the emergence of overt cardiomyopathy and curb adverse outcomes. Here, the authors review current understandings of how human hypertrophic cardiomyopathy- and hypertrophic dilated cardiomyopathy-linked mutations disrupt the normal structure and function of the sarcomere.
Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in combination with CRISPR/Cas9 genome editing provide unparalleled opportunities to study cardiac biology and disease. However, ...sarcomeres, the fundamental units of myocyte contraction, are immature and nonlinear in hiPSC-CMs, which technically challenge accurate functional interrogation of contractile parameters in beating cells. Furthermore, existing analysis methods are relatively low-throughput, indirectly assess contractility, or only assess well-aligned sarcomeres found in mature cardiac tissues.
We aimed to develop an analysis platform that directly, rapidly, and automatically tracks sarcomeres in beating cardiomyocytes. The platform should assess sarcomere content, contraction and relaxation parameters, and beat rate.
We developed SarcTrack, a MatLab software that monitors fluorescently tagged sarcomeres in hiPSC-CMs. The algorithm determines sarcomere content, sarcomere length, and returns rates of sarcomere contraction and relaxation. By rapid measurement of hundreds of sarcomeres in each hiPSC-CM, SarcTrack provides large data sets for robust statistical analyses of multiple contractile parameters. We validated SarcTrack by analyzing drug-treated hiPSC-CMs, confirming the contractility effects of compounds that directly activate (CK-1827452) or inhibit (MYK-461) myosin molecules or indirectly alter contractility (verapamil and propranolol). SarcTrack analysis of hiPSC-CMs carrying a heterozygous truncation variant in the myosin-binding protein C ( MYBPC3) gene, which causes hypertrophic cardiomyopathy, recapitulated seminal disease phenotypes including cardiac hypercontractility and diminished relaxation, abnormalities that normalized with MYK-461 treatment.
SarcTrack provides a direct and efficient method to quantitatively assess sarcomere function. By improving existing contractility analysis methods and overcoming technical challenges associated with functional evaluation of hiPSC-CMs, SarcTrack enhances translational prospects for sarcomere-regulating therapeutics and accelerates interrogation of human cardiac genetic variants.
Cardiac β-myosin variants cause hypertrophic (HCM) or dilated (DCM) cardiomyopathy by disrupting sarcomere contraction and relaxation. The locations of variants on isolated myosin head structures ...predict contractility effects but not the prominent relaxation and energetic deficits that characterize HCM. During relaxation, pairs of myosins form interacting-heads motif (IHM) structures that with other sarcomere proteins establish an energy-saving, super-relaxed (SRX) state. Using a human β-cardiac myosin IHM quasi-atomic model, we defined interactions sites between adjacent myosin heads and associated protein partners, and then analyzed rare variants from 6112 HCM and 1315 DCM patients and 33,370 ExAC controls. HCM variants, 72% that changed electrostatic charges, disproportionately altered IHM interaction residues (expected 23%; HCM 54%, p=2.6×10
; DCM 26%, p=0.66; controls 20%, p=0.23). HCM variant locations predict impaired IHM formation and stability, and attenuation of the SRX state - accounting for altered contractility, reduced diastolic relaxation, and increased energy consumption, that fully characterizes HCM pathogenesis.
As the number of cancer survivors has increased significantly over the last decades due to aging of population and development of effective cancer therapies, side effects from cancer therapies have ...been increasingly recognized. High-dose anthracyclines, immunotherapies, and concurrent radiation, as well as traditional cardiovascular risk factors such as smoking, hypertension, diabetes, hyperlipidemia, and obesity increase risks for unintended cardiovascular toxicity. However, these factors do not fully explain why only a subset of patients develop adverse cardiovascular sequelae from cancer therapies. Recent studies demonstrate that genetics play a substantial role in susceptibility to development of cardiovascular toxicities from cancer therapies. Common single nucleotide polymorphisms in multiple genes involved in various cellular pathways including membrane transport, stress response, and sarcomeres are recognized to increase risks for these toxicities. Pathogenic variants in the genes encoding proteins that comprise sarcomeres also contribute to cardiomyopathy following cancer therapies. Furthermore, genetic manipulations of model systems indicate mechanisms by which cardiotoxicities emerge following cancer immunomodulatory therapies. Continued efforts are needed to enable insights into cardiovascular responsiveness to these multi-targeted therapies, improve risk stratification of patients, and enable therapeutic interventions that limit these unintended adverse consequences from life-saving cancer treatments.
•Survivors of cancer therapies have life-long increased risks for heart disease.•Both common and rare damaging gene variants convey cardiotoxicity risks.•Emerging cancer immunotherapies can evoke cardiotoxic autoimmunity.•Improving cancer care requires identification and treatment of cardiotoxicity risks.
Hypertrophic cardiomyopathy (HCM) is a common inherited cardiac disorder that characterized by marked thickening of the left ventricular wall that occurs in the absence of increased external load. ...HCM is the most common cause of sudden cardiac death under 35 years and in addition causes heart failure. HCM is usually inherited as an autosomal dominant mutation in genes that encode protein constituents of the sarcomere. To date, more than 450 different mutations have been identified within 13 myofilament‐related genes. This review focuses current research involved in the discovery of other causative genes, investigation of the mechanisms by which sarcomere genes mutations produce hypertrophy and arrhythmia, and identification of modifying factors that influence clinical expression in HCM patients. The clinical implications of molecular advances in HCM are discussed.
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