MYH7 Mutation Research Articles

Heart Failure: From Protein to Phenotype37MicroRNA-494 reduces ATF3 expression and promotes heart failure in cardiac hypertrophic remodeling in vivo38A novel recessive plakophilin-2 gene mutation causes severe arrhythmogenic dilated cardiomyopathy and sudden cardiac death at young age39Investigation of titin expression in explanted hearts with familial dilated cardiomyopathy and TTN truncating variants

Heart Failure: From Protein to Phenotype37MicroRNA-494 reduces ATF3 expression and promotes heart failure in cardiac hypertrophic remodeling in vivo38A novel recessive plakophilin-2 gene mutation causes severe arrhythmogenic dilated cardiomyopathy and sudden cardiac death at young age39Investigation of titin expression in explanted hearts with familial dilated cardiomyopathy and TTN truncating variants

A novel MYH2 mutation in family members presenting with congenital myopathy, ophthalmoplegia and facial weakness.

Myosin heavy chain (MyHC) is a major structural component of the striated muscle contractile apparatus. In adult human limb skeletal muscle, there are three major MyHC isoforms, slow/beta cardiac MyHC, MyHC IIa and MHC IIx, which are important for the functional characteristics of different muscle fiber types. Hereditary myosin myopathies have emerged as an important group of diseases with variable clinical and morphological expression dependent on the mutated isoform, and also the type and location of the mutation. Myosin myopathy with external ophthalmoplegia is associated with mutations in MYH2, encoding for MyHC IIa that is mainly expressed in type 2A muscle fibers and is inherited in dominant as well as recessive manner. We present a family with myopathy with early onset proximal muscle weakness, facial muscle involvement and ophthalmoplegia. Muscle biopsy demonstrated lack of type 2A muscle fibers and genetic work up demonstrated that the disease was caused by a novel recessive MYH2 mutation: c.1009-1G>A resulting in skipping of exon 12, which is predicted to result in a frame shift and introducing at premature stop codon at position 347 (p.Ser337Leufs*11).

Isolation and Mechanical Measurements of Myofibrils from Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

SummaryTension production and contractile properties are poorly characterized aspects of excitation-contraction coupling of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Previous approaches have been limited due to the small size and structural immaturity of early-stage hiPSC-CMs. We developed a substrate nanopatterning approach to produce hiPSC-CMs in culture with adult-like dimensions, T-tubule-like structures, and aligned myofibrils. We then isolated myofibrils from hiPSC-CMs and measured the tension and kinetics of activation and relaxation using a custom-built apparatus with fast solution switching. The contractile properties and ultrastructure of myofibrils more closely resembled human fetal myofibrils of similar gestational age than adult preparations. We also demonstrated the ability to study the development of contractile dysfunction of myofibrils from a patient-derived hiPSC-CM cell line carrying the familial cardiomyopathy MYH7 mutation (E848G). These methods can bring new insights to understanding cardiomyocyte maturation and developmental mechanical dysfunction of hiPSC-CMs with cardiomyopathic mutations.

172. B-Type Natriuretic Peptide Gene Therapy as a Novel Early Treatment for Familial Hypertrophic Cardiomyopathy

Hypertrophic cardiomyopathy (HCM) is the most common familial cardiomyopathy, with the prevalence between 0.2 to 0.5%. HCM presents as an increase in left ventricular mass with thickening of the interventricular septum (IVS), myocellular disarray, interstitial fibrosis, heart rhythm abnormalities, and sudden death, necessitating therapeutic intervention. HCM is commonly caused by mutations in genes MYH7, MYBPC3, TNNT2, and TNNI3.The cardiac hormone, B-type natriuretic peptide (BNP) is a guanylyl cyclase A (GC-A) agonist. Physiological effects of BNP include natriuresis, vasodilation and blood pressure-lowering action. Additionally, BNP shows direct cardioprotective properties including anti-hypertrophic and anti-fibrotic effects, independently of its anti-hypertensive effects. We have recently found that genetic BNP ablation results in progressive cardiac hypertrophy with upregulation in HCM-associated genes in rats. In the current study, we tested the hypothesis that sustained BNP treatment with novel gene delivery of BNP may preserve cardiac function and structure in HCM in a mouse model of HCM.We used transgenic HCM mice expressing a mutated α-myosin heavy chain. Adeno-associated virus serotype 9 (AAV9) vectors were utilized to exploit their natural myocardial tropism. Three groups of mice were treated via tail-vein injections at four weeks of age (week one of study); Group 1, No HCM control (C57bl/6, no transgenic α-myosin heavy chain, n=6); Group 2, Untreated HCM mice with transgenic α-myosin heavy chain (PBS treated, n=10); Group 3, AAV9-BNP vector-treated HCM mice (n=6). To observe therapeutic benefit we assayed non-invasive blood pressure, myocardial architecture and remodeling (echocardiography), exercise capacity (treadmills), and any cardiorenal interplay through renal parameters. At 6, 9, 12, 15 and 25 weeks after AAV vector administration, left ventricular mass by echo was significantly higher in the untreated HCM group than the AAV-BNP-treated HCM and wildtype C57bl/6 mice. Significant increases in IVS and posterior wall thickness were also observed in control HCM group. At 11, 19, and 25 weeks after AAV vector administration, exercise capacity, monitored by treadmill tolerance, was significantly lower in the control HCM group than the AAV-BNP-treated HCM mice. No significant changes were seen in urine outputs. HCMAAV9-BNP treated mice showed a trend towards a lower blood pressure than both C57bl/6 and HCM mice.Our data demonstrate that AAV9-mediated BNP over-expression blocks progressive left ventricular enlargement and improves exercise capacity in a mouse model of HCM. The present study supports the BNP/GC-A/cGMP axis as a novel therapeutic target for the treatment of HCM.

Spectrum of clinical and genetic features of patients with inherited platelet disorder with suspected predisposition to hematological malignancies: a nationwide survey in Japan

Inherited thrombocytopenia (IT) contains several forms of familial thrombocytopenia and some of them have propensity to hematological malignancies. The etiological and genetic features of this heterogeneous syndrome have not yet been elucidated. We conducted a nationwide survey to collect clinical information and samples from patients with familial thrombocytopenia and/or hematological malignancies in order to obtain a comprehensive understanding of IT. Among the 43 pedigrees with clinical samples, RUNX1 mutations were identified in 8 pedigrees (18.6%). While MYH9 and ANKRD26 mutations were identified in 2 and 1 pedigrees, respectively, no gene mutations were detected in the remaining 32 pedigrees from a panel of previously reported pathogenetic mutations. Clinical data were comparable between FPD/AML and non-FPD/AML probands. Our study clarified that it is unexpectedly difficult to diagnose FPD/AML based on clinical information alone, and thus, genetic testing is strongly recommended. Our survey also identified some pedigrees with a strong family history of myelodysplastic syndromes of unknown origin. Additionally, there were 14 pedigrees in which three or more members were affected by immune thrombocytopenia (ITP), and a computer-aided simulation suggested that such a distribution almost never happens by coincidence, which implicates a genetic predisposition to ITP.

Myoimaging in the NGS era: the discovery of a novel mutation in MYH7 in a family with distal myopathy and core-like features – a case report

BackgroundMyosin heavy chain 7 related myopathies are rare disorders characterized by a wide phenotypic spectrum and heterogeneous pathological features. In the present study, we performed clinical, morphological, genetic and imaging investigations in three relatives affected by autosomal dominant distal myopathy. Whilst earlier traditional Sanger investigations had pointed to the wrong gene as disease causative, next-generation sequencing allowed us to obtain the definitive molecular genetic diagnosis in the family.Case presentationThe proposita, being found to harbor a novel heterozygous mutation in the RYR1 gene (p.Glu294Lys), was initially diagnosed with core myopathy. Subsequently, consideration of muscle magnetic resonance imaging (MRI) features and extension of family study led this diagnosis to be questioned. Use of next-generation sequencing analysis identified a novel mutation in the MYH7gene (p.Ser1435Pro) that segregated in the affected family members.ConclusionsThis study identified a novel mutation in MYH7 in a family where the conclusive molecular diagnosis was reached through a complicated path. This case report might raise awareness, among clinicians, of the need to interpret NGS data in combination with muscle MRI patterns so as to facilitate the pinpointing of the main molecular etiology in inherited muscle disorders.Electronic supplementary materialThe online version of this article (doi:10.1186/s12881-016-0288-0) contains supplementary material, which is available to authorized users.

MYBPH acts as modifier of cardiac hypertrophy in hypertrophic cardiomyopathy (HCM) patients.

Left ventricular hypertrophy is a risk factor for cardiovascular morbidity and mortality. Hypertrophic cardiomyopathy (HCM) is considered a model disease to study causal molecular factors underlying isolated cardiac hypertrophy. However, HCM manifests with various clinical symptoms, even in families bearing the same genetic defects, suggesting that additional factors contribute to hypertrophy. The gene encoding the cardiac myosin binding protein C (cMYBPC) is one of the most frequently implicated genes in HCM. Recently another myosin binding protein, myosin binding protein H (MYBPH) was shown to function in concert with cMYBPC in regulating cardiomyocyte contraction. Given the similarity in sequence, structure and the critical role MYBPH plays in sarcomere contraction, we proposed that MYBPH may be involved in HCM pathogenesis. Family-based genetic association analysis was employed to investigate the contribution of MYBPH in modifying hypertrophy. Seven single nucleotide polymorphisms and haplotypes in MYBPH were investigated for hypertrophy modifying effects in 388 individuals (27 families), in which three unique South African HCM-causing founder mutations (p.R403W and pA797T in β-myosin heavy chain gene (MYH7) and p.R92W in the cardiac troponin T gene (TNNT2)) segregate. We observed a significant association between rs2250509 and hypertrophy traits in the p.A797T MYH7 mutation group. Additionally, haplotype GGTACTT significantly affected hypertrophy within the same mutation group. MYBPH was for the first time assessed as a candidate hypertrophy modifying gene. We identified a novel association between MYBPH and hypertrophy traits in HCM patients carrying the p.A797T MYH7 mutation, suggesting that variation in MYBPH can modulate the severity of hypertrophy in HCM.

Prevalence and Phenotypic Expression of Mutations in the MYH7, MYBPC3 and TNNT2 Genes in Families with Hypertrophic Cardiomyopathy in the South of Brazil: A Cross-Sectional Study.

Background:Mutations in sarcomeric genes are found in 60-70% of individuals with familial forms of hypertrophic cardiomyopathy (HCM). However, this estimate refers to northern hemisphere populations. The molecular-genetic profile of HCM has been subject of few investigations in Brazil, particularly in the south of the country.Objective:To investigate mutations in the sarcomeric genes MYH7, MYBPC3 and TNNT2 in a cohort of HCM patients living in the extreme south of Brazil, and to evaluate genotype-phenotype associations.Methods:Direct DNA sequencing of all encoding regions of three sarcomeric genes was conducted in 43 consecutive individuals of ten unrelated families.Results:Mutations for CMH have been found in 25 (58%) patients of seven (70%) of the ten study families. Fourteen (56%) individuals were phenotype-positive. All mutations were missense, four (66%) in MYH7 and two (33%) in MYBPC3. We have not found mutations in the TNNT2 gene. Mutations in MYH7 were identified in 20 (47%) patients of six (60%) families. Two of them had not been previously described. Mutations in MYBPC3 were found in seven (16%) members of two (20%) families. Two (5%) patients showed double heterozygosis for both genes. The mutations affected different domains of encoded proteins and led to variable phenotypic expression. A family history of HCM was identified in all genotype-positive individuals.Conclusions:In this first genetic-molecular analysis carried out in the south of Brazil, we found mutations in the sarcomeric genes MYH7 and MYBPC3 in 58% of individuals. MYH7-related disease was identified in the majority of cases with mutation.

Nonmuscle Myosin Heavy Chain IIA Mutation Predicts Severity and Progression of Sensorineural Hearing Loss in Patients With MYH9-Related Disease.

MYH9-related disease (MYH9-RD) is an autosomal- dominant disorder deriving from mutations in MYH9, the gene for the nonmuscle myosin heavy chain (NMMHC)-IIA. MYH9-RD has a complex phenotype including congenital features, such as thrombocytopenia, and noncongenital manifestations, namely sensorineural hearing loss (SNHL), nephropathy, cataract, and liver abnormalities. The disease is caused by a limited number of mutations affecting different regions of the NMMHC-IIA protein. SNHL is the most frequent noncongenital manifestation of MYH9-RD. However, only scarce and anecdotal information is currently available about the clinical and audiometric features of SNHL of MYH9-RD subjects. The objective of this study was to investigate the severity and propensity for progression of SNHL in a large series of MYH9-RD patients in relation to the causative NMMHC-IIA mutations. This study included the consecutive patients diagnosed with MYH9-RD between July 2007 and March 2012 at four participating institutions. A total of 115 audiograms were analyzed from 63 patients belonging to 45 unrelated families with different NMMHC-IIA mutations. Cross-sectional analyses of audiograms were performed. Regression analysis was performed, and age-related typical audiograms (ARTAs) were derived to characterize the type of SNHL associated with different mutations. Severity of SNHL appeared to depend on the specific NMMHC-IIA mutation. Patients carrying substitutions at the residue R702 located in the short functional SH1 helix had the most severe degree of SNHL, whereas patients with the p.E1841K substitution in the coiled-coil region or mutations at the nonhelical tailpiece presented a mild degree of SNHL even at advanced age. The authors also disclosed the effects of different amino acid changes at the same residue: for instance, individuals with the p.R702C mutation had more severe SNHL than those with the p.R702H mutation, and the p.R1165L substitution was associated with a higher degree of hearing loss than the p.R1165C. In general, mild SNHL was associated with a fairly flat audiogram configuration, whereas severe SNHL correlated with downsloping configurations. ARTA plots showed that the most progressive type of SNHL was associated with the p.R702C, the p.R702H, and the p.R1165L substitutions, whereas the p.R1165C mutation correlated with a milder, nonprogressive type of SNHL than the p.R1165L. ARTA for the p.E1841K mutation demonstrated a mild degree of SNHL with only mild progression, whereas the ARTA for the mutations at the nonhelical tailpiece did not show any substantial progression. These data provide useful tools to predict the progression and the expected degree of severity of SNHL in individual MYH9-RD patients, which is especially relevant in young patients. Consequences in clinical practice are important not only for appropriate patient counseling but also for development of customized, genotype-driven clinical management. The authors recently reported that cochlear implantation has a good outcome in MYH9-RD patients; thus, stricter follow-up and earlier intervention are recommended for patients with unfavorable genotypes.

Aborted Sudden Cardiac Death in a Pediatric Patient with Hypertrophic Cardiomyopathy and a Novel MYH7 Mutation

Aborted Sudden Cardiac Death in a Pediatric Patient with Hypertrophic Cardiomyopathy and a Novel MYH7 Mutation

Abstract 12176: Outcomes and Genetic Characteristics of Hypertrophic Cardiomyopathy With Severe Right Ventricular Hypertrophy

Introduction: Severe right ventricular hypertrophy (SRVH) is extremely rare in hypertrophic cardiomyopathy (HCM). Hypothesis: We assessed the hypothesis that patients with HCM with SRVH may have multiple mutations and poor prognosis. Methods: We performed a retrospective cohort study involving 2415 HCM patients. Patients with HCM with a maximum right ventricular wall thickness ≥10 mm, either with or without a pressure gradient at the RV outflow tract (RVOT) exceeding 25 mmHg, were enrolled. Deep (30X) whole-genome sequencing was performed in 11 patients using the Illumina HiSeq X platform. The patients'clinical features, cardiovascular mortality and morbidity were analyzed. The survival estimates were calculated using the Kaplan-Meier method, and risk of either cardiovascular death or event was determined using a multivariate Cox hazards regression model. Results: Of the 2415 HCM patients, 43 patients (1.6%) with SRVH were identified and 12 of them demonstrated isolated RVOT obstruction. The mean age at diagnosis was 31.1 ± 16.3 years, and 24 (55.8%) of them were women. The maximum average RV wall thickness was 13.3 ± 2.4 mm. The multivariate Cox regression analysis identified 2 independent predictors of cardiovascular death, including NYHA class ≥III, with a hazard ratio of 8.6 (95% CI: 1.70-43.38, p = 0.009), and a left ventricular end-diastolic diameter ≥50 mm, with a hazard ratio of 6.0 (95% CI: 1.67-21.92, p = 0.006). Age ≤18 years was with a 3-fold increased risk cardiovascular events (95% CI: 1.14-7.16, p = 0.002). At least one variant in a specific sarcomere gene was identified in 10 patients (90.9%), of which the MYH7 mutation was the most common (n = 6, 54.5%), and 10 novel locations were detected. Double mutations involving HCM-related genes were observed in 8 (73%) patients. All patients exhibited either double or multiple mutations, either in arrhythmogenic right ventricular, dilated cardiomyopathy or ion-channel disease genes. Conclusions: HCM with SRVH was characterized by an early onset of disease and increased cardiovascular mortality and morbidity. Double or multiple sarcomere gene mutations might be responsible for this phenotype with multiple mutations in other forms of cardiomyopathy and ion-channel disease associated genes.

Abstract 20016: Genetic Anticipation in Familial Hypertrophic Cardiomyopathy

Introduction: To identify HCM at the earliest time possible, estimating the onset of disease based on diagnostic criteria is vital in timing the initiation of screening and interventions. Hypothesis: Clinic observations indicate an early diagnose of HCM in second generation of most affected families. From the pedigrees with or without mutations in beta-myosin heavy chain gene (MYH7) affected in two or more generations with hypertrophic cardiomyopathy (HCM), ages and maximal wall thickness (MWT) of left ventricles at diagnosis were evaluated to investigate the potential genetic anticipation in patients with familiar HCM (FHCM). Methods: 56 individuals from 25 families were analyzed. Linear mixed effects models were adopted to prevent misinterpretation resulting from the cohort effect. Published data containing 9 sarcomere mutations (181 individuals in 52 families) also were extracted. Results: MYH7 mutations were detected in 9 of the 25 probands, The median age of HCM diagnosis was 24 in the younger generation and 55 in the older (p < 0.001). In the parametric model, the estimated change in the expected age at diagnosis for the entire cohort was 25.8 years (p <0.001). Statistically significant earlier ages at diagnoses were also observed within subgroups of MYH7+ and MYH7- mutations, and probands older and younger than 30 years old. Although the estimated change in MWT at diagnosis for the entire cohort was only 2.161 mm (p =0.212), the subgroup of probands in the younger generation aged less than 30 years had 10.393 mm (p=0.018) thicker MWT and showed a significant reverse correlation with age. Analysis of published data also supports these findings. Conclusions: Genetic anticipation was observed in patients with FHCM. FHCM is prone to be diagnosed at an earlier age in younger generations. Patients who are younger with relatives affected by HCM, especially those who are diagnosed before 30 years of age, should continue to be tracked to offer appropriate screening modalities as earlier as possible.

Abstract 12532: Cellular and Cardiac Microtissue Assays of iPSC-derived Myocytes With the Hypertrophic Cardiomyopathy Mutation in MYH7-Val606Met

Introduction: Hypertrophic cardiomyopathy (HCM) is the most common inherited heart disease affecting 1 in 500 people and is the leading cause of sudden cardiac death in young adults. Mutations in β-myosin heavy chain ( MYH7 ) account for one-third of cases, but the underlying pathogenesis remains unclear. In this study, we investigated myocytes derived from CRISPR-Cas9-engineered isogenic induced pluripotent stem cell cardiomyocytes (iPSC-CMs) with a missense mutation Val606Met by single-cell and microtissue functional assays. Hypothesis: We hypothesize that iPSC-CMs with Val606Met will recapitulate features of HCM in single cell and microtissue assays, such as changes in contractile function and hypertrophic signaling. Methods & Results: Val606Met mutations in MYH7 (VM/+, VM/VM) were introduced by targeting an iPSC line using CRISPR-Cas9. Transfected iPSCs were isolated by FACS to detect green fluorescent protein, clones were expanded, and mutational events were confirmed by Sanger sequencing. Mutant iPSCs, like isogenic wildtype control iPSCs, were differentiated into cardiomyocytes with >90% efficiency. Microtissue studies of mutant iPSC-CMs exhibited increases in contractility (WT=2.96±0.23μN, VM/+ = 3.51±0.34μN, VM/VM = 4.63±0.51; R 2 = 0.963). Mutant iPSC-CMs also showed evidence of altered signaling by increases in spontaneous beating rate (WT < VM/+ < VM/VM; R 2 =0.946), cell size (compared to WT, ~20% increases in both VM/+, p=0.04 and VM/VM, p=0.038), and sarcomere content (compared to WT, ~20% increases in both VM/+, p=4e-06 and VM/VM, p=0.11). These changes are supported by activated expression in myocytes of hypertrophic genes such as NPPA , NPPB , and ACTA2 as well as transcriptional up-regulation of TGF-β. Conclusions: Isogenic iPSC-CMs with MYH7 variant Val606Met created by CRISPR-Cas9 recapitulate features of HCM in vitro . Our data confirms Val606Met as a pathogenic mutation that causes increased contractility in cardiac microtissues and myocyte hypertrophy. We speculate that increased myocyte-derived TGF-β signaling may contribute to the HCM phenotype. This approach could also be used for better understanding of genotype-phenotype correlations in HCM.

Developmental MYH3 Myopathy Associated with Expression of Mutant Protein and Reduced Expression Levels of Embryonic MyHC

ObjectiveAn essential role for embryonic MyHC in foetal development has been found from its association with distal arthrogryposis syndromes, a heterogeneous group of disorders characterised by congenital contractions. The latter probably result from severe myopathy during foetal development. Lack of embryonic muscle biopsy material and suitable animal models has hindered study of the pathomechanisms linking mutations in MYH3 to prenatal myopathy.Methods and ResultsWe determined the pathomechanisms of developmental myopathy caused by recurrent p.Thr178Ile MYH3 heterozygosity, using patient-derived skeletal muscle cells in culture as an experimental disease model to emulate early embryonic development. These cultured cells were processed for discrimination and quantitative analysis of mutant and wild-type MYH3 alleles and MyHC transcripts, real-time RT-qPCR, sequence analysis, immunofluorescence microscopy, immunoblot, and proteomic assessments. Involvement of the ubiquitin proteasome system was investigated in patients with p.Thr178Ile mutations in MYH3 and MYH2. We found equal overall expression of mutant and wild-type MyHC mRNAs and proteins. Compared to the controls, however, expression of embryonic MyHC transcripts and proteins was reduced whereas expression of myosin-specific E3 ubiquitin ligase (MuRF1) was increased. We also found delayed myofibrillogenesis and atrophic myotubes but structured sarcomeres.ConclusionIn conclusion, this study suggests that developmental p.Thr178Ile MYH3 myopathy is associated with a combined pathomechanism of insufficient dosage of functional embryonic MyHC and production of mutant protein.

OBSCN Mutations Associated with Dilated Cardiomyopathy and Haploinsufficiency.

BackgroundStudies of the functional consequences of DCM-causing mutations have been limited to a few cases where patients with known mutations had heart transplants. To increase the number of potential tissue samples for direct investigation we performed whole exon sequencing of explanted heart muscle samples from 30 patients that had a diagnosis of familial dilated cardiomyopathy and screened for potentially disease-causing mutations in 58 HCM or DCM-related genes.ResultsWe identified 5 potentially disease-causing OBSCN mutations in 4 samples; one sample had two OBSCN mutations and one mutation was judged to be not disease-related. Also identified were 6 truncating mutations in TTN, 3 mutations in MYH7, 2 in DSP and one each in TNNC1, TNNI3, MYOM1, VCL, GLA, PLB, TCAP, PKP2 and LAMA4. The mean level of obscurin mRNA was significantly greater and more variable in healthy donor samples than the DCM samples but did not correlate with OBSCN mutations. A single obscurin protein band was observed in human heart myofibrils with apparent mass 960 ± 60 kDa. The three samples with OBSCN mutations had significantly lower levels of obscurin immunoreactive material than DCM samples without OBSCN mutations (45±7, 48±3, and 72±6% of control level).Obscurin levels in DCM controls, donor heart and myectomy samples were the same.Conclusions OBSCN mutations may result in the development of a DCM phenotype via haploinsufficiency. Mutations in the obscurin gene should be considered as a significant causal factor of DCM, alone or in concert with other mutations.

G.P.315 - Coexisting TIA1 and MYH7 distal myopathy

Distal myopathies are genetically heterogeneous. TIA1 mutations cause Welander distal myopathy. MYH7 mutations result in various clinical phenotypes, including Laing distal myopathy, myosin storage myopathy and hypertrophic/dilated cardiomyopathy. We describe a family affected by distal myopathy and variable presence of cardiomyopathy due to coexisting TIA1 and MYH7 mutations. The propositus is a 67-year-old woman of Swedish and Greek descent with history of easy tripping since childhood who has experienced a stepwise progression of asymmetric distal limb weakness in the past 20 years. Neurological examination revealed mild to severe asymmetric distal lower >upper limb weakness. CK value was normal. EMG study showed myopathic changes with minimal fibrillation potentials and myotonic discharges. Biopsy of the extensor carpi radialis muscle showed: rare rimmed vacuoles, minicore-like structures and congophilic inclusions; few necrotic and regenerating fibers; fiber splitting; increased internal nuclei; and fiber type grouping. There was no cardiac involvement by ECG and echocardiogram. Her 66-year-old sister has 8 years of mild distal limb weakness, supraventricular tachycardia and hypertrophic cardiomyopathy documented by echocardiogram and MRI. Another sibling has distal weakness while parents were asymptomatic. Both sisters carry a known pathogenic heterozygous TIA1 mutation (p.Glu384Lys) and a rare heterozygous MYH7 variant (c.5459G > A; p.Arg1820Gln), which involves a highly conserved residue and is predicted to be pathogenic. Because a homozygous MYH7 mutation (p.Arg1820Trp) was reported in recessive myosin storage myopathy, aCGH deletion/duplication analysis of MYH7 was performed in the proband and it was normal. The clinical phenotype is dictated by the coexistent TIA1 and MYH7 mutations, which can result in similar pathological findings. The proband's early onset of distal leg weakness and the sister's cardiomyopathy support the pathogenicity of the MYH7.

Mutation and clinical relevance in a large cohort of unrelated Chinese patients with hypertrophic cardiomyopathy

To explore the genetic basis and phenotypic correlation with disease severity in a large cohort of Chinese patients with hypertrophic cardiomyopathy (HCM). A total of 179 unrelated Chinese HCM patients admitted to our department from 2002 to 2011 were enrolled in this study. Direct gene sequencing of β-myosin heavy chain (MYH7), myosin binding protein-C ( MYBPC3), and cardiac troponin T (TNNT2) were performed and clinical data were obtained in these patients. A total of 34 mutations were identified in 40 patients (22.3%), 79.4% (27/34) mutations occurred only once and a possible hot spot, A26 in MYH7, was found. Distribution of mutations was 52.9% (18/34) (MYBPC3), 35.3% (12/34) ( MYH7) and 11.8% (4/34) (TNNT2) respectively. Double mutations were identified in 2.2% (4/179) patients. Genotype-positive patients were associated with an earlier symptom onset, severer left ventricular hypertrophy, a higher incidence of syncope, and were more likely to have positive family history of HCM or sudden cardiac death (SCD) , and were more likely to progress into heart failure (24.2% vs. 5.0%, P = 0.002) and at a higher risk of SCD (9.1% vs. 0, P = 0.009) during the 6.5-year follow-up. No statistical difference in any clinical parameters and outcomes was found between patients carrying MYBPC3 and MYH7 mutations. Double mutations were associated with malignant clinical progression in this cohort. Different phenotype severity could be seen in HCM patients with same genotype (e. g. MYH7-1736T, TNNT2-R92W). MYBPC3 is the most predominant gene mutation in this HCM cohort. The presence of a sarcomere mutation in patients with HCM is associated with poor clinical outcome, although no specific genes or mutations can exactly predict the severity of clinical phenotypes.

Abstract 343: Bayesian Selection of Modifier Genes in Hypertrophic Cardiomyopathy Through Whole Genome Sequencing

Background: Technological advances have greatly reduced the cost of whole genome sequencing. For single individuals clinical application is apparent, while exome sequencing in tens of thousands of people has allowed a more global view of genetic variation that can inform interpretation of specific variants in individuals. We hypothesized that genome sequencing of patients with monogenic cardiomyopathy would facilitate discovery of genetic modifiers of phenotype. Methods and Results: We identified 48 individuals diagnosed with cardiomyopathy and with putative mutations in MYH7, the gene encoding beta myosin heavy chain. We carried out whole genome sequencing and applied a newly developed analytical pipeline optimized for discovery of genes modifying severity of clinical presentation and outcomes. Using a combination of external priors and rare variant burden tests we scored genes as potential modifiers. There were 96 genes that reached a modifier score of 6 out of 12 or better (9=2, 8=8, 7=17, 6=69). We identified NCKAP1, a gene that regulates actin filament dynamics, and CAMSAP1, a calmodulin regulate gene that regulates microtubule dynamics, as top scoring modifiers of hypertrophic cardiomyopathy phenotypes (score=9) while LDB2, RYR2, FBN1 and ATP1A2 had modifier scores of 8. Of the top scoring genes, 21 out of 96 were identified as candidates a priori. Our candidate prioritization scheme identified the previously described modifiers of cardiomyopathy phenotype, FHOD3 and MYBPC3, as top scoring genes. We identified structural variants in 21 clinically sequenced cardiomyopathy associated genes, 13 of which were at less than 10% frequency. Copy number variants in ILK and CSRP3 were nominally associated with ejection fraction (p=0.03), while 8 genes showed copy gains (GLA, FKTN, SGCD, TTN, SOS1, ANKRD1, VCL and NEBL). Structural variants were found in CSRP3, MYL3 and TNNC1, all of which have been implicated as causative for HCM. Conclusion: Evaluation of the whole genome sequence, even in the case of putatively monogenic disease, leads to important diagnostic and scientific insights not revealed by panel-based sequencing.

Screening of sarcomere gene mutations in young athletes with abnormal findings in electrocardiography: identification of a MYH7 mutation and MYBPC3 mutations

There is an overlap between the physiological cardiac remodeling associated with training in athletes, the so-called athlete's heart, and mild forms of hypertrophic cardiomyopathy (HCM), the most common hereditary cardiac disease. HCM is often accompanied by unfavorable outcomes including a sudden cardiac death in the adolescents. Because one of the initial signs of HCM is abnormality in electrocardiogram (ECG), athletes may need to monitor for ECG findings to prevent any unfavorable outcomes. HCM is caused by mutations in genes for sarcomere proteins, but there is no report on the systematic screening of gene mutations in athletes. One hundred and two genetically unrelated young Japanese athletes with abnormal ECG findings were the subjects for the analysis of four sarcomere genes, MYH7, MYBPC3, TNNT2 and TNNI3. We found that 5 out of 102 (4.9%) athletes carried mutations: a heterozygous MYH7 Glu935Lys mutation, a heterozygous MYBPC3 Arg160Trp mutation and another heterozygous MYBPC3 Thr1046Met mutation, all of which had been reported as HCM-associated mutations, in 1, 2 and 2 subjects, respectively. This is the first study of systematic screening of sarcomere gene mutations in a cohort of athletes with abnormal ECG, demonstrating the presence of sarcomere gene mutations in the athlete's heart.

A de novo mutation of the MYH7 gene in a large Chinese family with autosomal dominant myopathy.

Laing distal myopathy (LDM) is an autosomal dominant myopathy that is caused by mutations in the slow/beta cardiac myosin heavy-chain (MYH7) gene. It has been recently reported that LDM presents with a wide range of clinical manifestations. We herein report a large Chinese family with autosomal dominant myopathy. The affected individuals in the family presented with foot drop in early childhood, along with progressive distal and proximal limb weakness. Their characteristic symptoms include scapular winging and scoliosis in the early disease phase and impairment of ambulation in the advanced phase. Although limb-girdle muscle dystrophy (LGMD) was suspected initially, a definite diagnosis could not be reached. As such, we performed linkage analysis and detected four linkage regions, namely 1q23.2-24.1, 14q11.2-12, 15q26.2-26.3 and 17q24.3. Through subsequent whole exome sequencing, we found a de novo p.K1617del causative mutation in the MYH7 gene and diagnosed the disease as LDM. This is the first LDM case in China. Our patients have severe clinical manifestations that mimic LGMD in comparison with the patients with the same mutation reported elsewhere.