Heart failure (HF), a leading cause for cardiac transplantation and premature death, is usually preceded by ventricular dilatation and diminished systolic performance or dilated cardiomyopathy (DCM). DCM has many etiologies, including damaging variants in genes with diverse functions in cardiac biology. During the prior funding period we showed that the most common genetic cause of DCM was truncating variants in titin (TTNtv). These account for 25% of familial and 12% of sporadic DCM and for ~10% of DCM that occurs with pregnancy, alcohol abuse, and after cancer therapies. In addition, ~0.2% of the general population carries a TTNtv; these individuals have substantially higher lifelong risks for developing DCM and heart failure. We also identified mechanisms by which TTNtv and other recently recognized DCM genes (FLNC and ALPK3) cause disease. With this competitive renewal we propose to focus on the discovery of genes and mechanisms that account for unexplained DCM, which remains an unmet need. We propose that some unexplained DCM is mechanistically related to established genetic causes and results from sequence variants that are not routinely interrogated, or that have unclear functional consequences. We will study the roles of somatic variants, non-coding regulatory variants, mitochondrial variants, and variants of unknown significance (VUS) in established and newly identified DCM genes. We will also define cell populations and transcriptional profiles of all cells in human hearts with unexplained DCM and DCM with established genetic etiologies, so as to identify shared or distinct pathways that may inform therapeutic opportunities. Our analyses will employ state-of-the art technologies. We will exploit whole genome sequencing (WGS) from blood- and cardiac tissue-derived DNAs obtained from unexplained DCM subjects. We will use single nuclear RNA sequencing (NucSeq) to define how cell populations and transcription change in DCM hearts in comparison to normal hearts, using our recently completed normal human heart NucSeq data. We will perturb new identified variants and mechanisms in iPSC-CMs and mouse models. These studies will improve knowledge of the molecules and pathways that enable normal heart function, the molecular causes and mechanisms of DCM, information that will improve diagnosis and inform precision therapies to prevent heart failure. Our analyses will also contribute functional insights into noncoding sequences. To accomplish these goals, we will: 1) Identify coding and non-coding, germline and somatic variants that contribute to unexplained DCM; 2) Define perturbed cell populations and associated transcriptional profiles in hearts from variant-positive and unexplained DCM; 3) Define DCM mechanisms using engineered iPSC-CMs and mouse models.
Dilated cardiomyopathy (DCM), a major cause of heart failure, enlarges the chambers and reduces the function of the heart. Damaging DNA variants are an important cause of DCM that are identified in 20-40% of patients, but the cause(s) of DCM in many patients remains unexplained. We propose to use new molecular technologies to study DNA sequences and single cells of unexplained DCM patient heart tissue hoping to identify new causes or contributors to DCM that will lead to improved diagnosis and treatments for these patients.
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|Sharma, Arun; Toepfer, Christopher N; Schmid, Manuel et al. (2018) Differentiation and Contractile Analysis of GFP-Sarcomere Reporter hiPSC-Cardiomyocytes. Curr Protoc Hum Genet 96:21.12.1-21.12.12|
|Sharma, Arun; Toepfer, Christopher N; Ward, Tarsha et al. (2018) CRISPR/Cas9-Mediated Fluorescent Tagging of Endogenous Proteins in Human Pluripotent Stem Cells. Curr Protoc Hum Genet 96:21.11.1-21.11.20|
|Garfinkel, Amanda C; Seidman, Jonathan G; Seidman, Christine E (2018) Genetic Pathogenesis of Hypertrophic and Dilated Cardiomyopathy. Heart Fail Clin 14:139-146|
|Sharma, Arun; Mücke, Michael; Seidman, Christine E (2018) Human Induced Pluripotent Stem Cell Production and Expansion from Blood using a Non-Integrating Viral Reprogramming Vector. Curr Protoc Mol Biol 122:e58|
|Eyckmans, Jeroen; Chen, Christopher S (2017) 3D culture models of tissues under tension. J Cell Sci 130:63-70|
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|Ito, Kaoru; Patel, Parth N; Gorham, Joshua M et al. (2017) Identification of pathogenic gene mutations in LMNA and MYBPC3 that alter RNA splicing. Proc Natl Acad Sci U S A 114:7689-7694|
|Alamo, Lorenzo; Ware, James S; Pinto, Antonio et al. (2017) Effects of myosin variants on interacting-heads motif explain distinct hypertrophic and dilated cardiomyopathy phenotypes. Elife 6:|
|Schafer, Sebastian; de Marvao, Antonio; Adami, Eleonora et al. (2017) Titin-truncating variants affect heart function in disease cohorts and the general population. Nat Genet 49:46-53|
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