The role of renin-angiotensin-aldosterone system polymorphisms in phenotypic expression of MYBPC3-related hypertrophic cardiomyopathy

The phenotypic variability of hypertrophic cardiomyopathy (HCM) in patients with identical pathogenic mutations suggests additional modifiers. In view of the regulatory role in cardiac function, blood pressure, and electrolyte homeostasis, polymorphisms in the renin-angiotensin-aldosterone system (RAAS) are candidates for modifying phenotypic expression. In order to investigate whether RAAS polymorphisms modulate HCM phenotype, we selected a large cohort of carriers of one of the three functionally equivalent truncating mutations in the MYBPC3 gene. Family-based association analysis was performed to analyze the effects of five candidate RAAS polymorphisms (ACE, rs4646994; AGTR1, rs5186; CMA, rs1800875; AGT, rs699; CYP11B2, rs1799998) in 368 subjects carrying one of the three mutations in the MYBPC3 gene. Interventricular septum (IVS) thickness and Wigle score were assessed by 2D-echocardiography. SNPs in the RAAS system were analyzed separately and combined as a pro-left ventricular hypertrophy (LVH) score for effects on the HCM phenotype. Analyzing the five polymorphisms separately for effects on IVS thickness and Wigle score detected two modest associations. Carriers of the CC genotype in the AGT gene had less pronounced IVS thickness compared with CT and TT genotype carriers. The DD polymorphism in the ACE gene was associated with a high Wigle score (P=0.01). No association was detected between the pro-LVH score and IVS thickness or Wigle score. In conclusion, in contrast to previous studies, in our large study population of HCM patients with functionally equivalent mutations in the MYBPC3 gene we did not find major effects of genetic variation within the genes of the RAAS system on phenotypic expression of HCM

Genetic Evaluation in a Cohort of 126 Dutch Pulmonary Arterial Hypertension Patients

Pulmonary arterial hypertension (PAH) is a severe, life-threatening disease, and in some cases is caused by genetic defects. This study sought to assess the diagnostic yield of genetic testing in a Dutch cohort of 126 PAH patients. Historically, genetic testing in the Netherlands consisted of the analysis of BMPR2 and SMAD9. These genes were analyzed in 70 of the 126 patients. A (likely) pathogenic (LP/P) variant was detected in 22 (31%) of them. After the identification of additional PAH associated genes, a next generation sequencing (NGS) panel consisting of 19 genes was developed in 2018. Additional genetic testing was offered to the 48 BMPR2 and SMAD9 negative patients, out of which 28 opted for NGS analysis. In addition, this gene panel was analyzed in 56 newly identified idiopathic (IPAH) or pulmonary veno occlusive disease (PVOD) patients. In these 84 patients, NGS panel testing revealed LP/P variants in BMPR2 (N = 4), GDF2 (N = 2), EIF2AK4 (N = 1), and TBX4 (N = 3). Furthermore, 134 relatives of 32 probands with a LP/P variant were tested, yielding 41 carriers. NGS panel screening offered to IPAH/PVOD patients led to the identification of LP/P variants in GDF2, EIF2AK4, and TBX4 in six additional patients. The identification of LP/P variants in patients allows for screening of at-risk relatives, enabling the early identification of PAH

Lack of evidence for a causal role of CALR3 in monogenic cardiomyopathy

The pathogenicity of previously published disease-associated genes and variants is sometimes questionable. Large-scale, population-based sequencing studies have uncovered numerous false assignments of pathogenicity. Misinterpretation of sequence variants may have serious implications for the patients and families involved, as genetic test results are increasingly being used in medical decision making. In this study, we assessed the role of the calreticulin-3 gene (CALR3) in cardiomyopathy. CALR3 has been included in several cardiomyopathy gene panels worldwide. Its inclusion is based on a single publication describing two missense variants in patients with hypertrophic cardiomyopathy. In our national cardiomyopathy cohort (n = 6154), we identified 17 unique, rare heterozygous CALR3 variants in 48 probands. Overall, our patient cohort contained a significantly higher number of rare CALR3 variants compared to the ExAC population (p = 0.0036). However, after removing a potential Dutch founder variant, no statistically significant difference was found (p = 0.89). In nine probands, the CALR3 variant was accompanied by a disease-causing variant in another, well-known cardiomyopathy gene. In three families, the CALR3 variant did not segregate with the disease. Furthermore, we could not demonstrate calreticulin-3 protein expression in myocardial tissues at various ages. On the basis of these findings, it seems highly questionable that variants in CALR3 are a monogenic cause of cardiomyopathy

A mutation update for the FLNC gene in myopathies and cardiomyopathies

Filamin C (FLNC) variants are associated with cardiac and muscular phenotypes. Originally, FLNC variants were described in myofibrillar myopathy (MFM) patients. Later, high-throughput screening in cardiomyopathy cohorts determined a prominent role for FLNC in isolated hypertrophic and dilated cardiomyopathies (HCM and DCM). FLNC variants are now among the more prevalent causes of genetic DCM. FLNC-associated DCM is associated with a malignant clinical course and a high risk of sudden cardiac death. The clinical spectrum of FLNC suggests different pathomechanisms related to variant types and their location in the gene. The appropriate functioning of FLNC is crucial for structural integrity and cell signaling of the sarcomere. The secondary protein structure of FLNC is critical to ensure this function. Truncating variants with subsequent haploinsufficiency are associated with DCM and cardiac arrhythmias. Interference with the dimerization and folding of the protein leads to aggregate formation detrim

Obtaining insurance after DNA diagnostics: a survey among hypertrophic cardiomyopathy mutation carriers

Hypertrophic cardiomyopathy (HCM) is a common hereditary heart disease associated with increased mortality. Disclosure of DNA test results may have social implications such as low access to insurance. In the Netherlands, insurance companies are restricted in the use of genetic information of their clients by the Medical Examination Act. A cross-sectional survey was used to assess the frequency and type of problems encountered by HCM mutation carriers applying for insurance, and associations with carriers' characteristics. The response rate was 86% (228/264). A total of 66 carriers (29%) applied for insurance of whom 39 reported problems (59%) during an average follow-up of 3 years since the DNA test result. More problems were encountered by carriers with manifest disease (P<0.001) and carriers with symptoms of HCM (P=0.049). Carriers identified after predictive DNA testing less frequently experienced problems (P=0.002). Three carriers without manifest HCM reported problems (5% of applicants). Frequently reported problems were higher premium (72%), grant access to medical records (62%), and complete rejection (33%). In conclusion, HCM mutation carriers frequently encounter problems when applying for insurances, often in the case of manifest disease, but the risk assessment of insurance companies is largely justified. Still, 5% of carriers encounter potentially unjustified problems, indicating the necessity to monitor the application of the existing laws and regulations by insurance companies and to educate counselees on the implications of these laws and regulations

Reclassification of a likely pathogenic Dutch founder variant in KCNH2; implications of reduced penetrance

BACKGROUND: Variants in KCNH2, encoding the human ether a-go-go (hERG) channel that is responsible for the rapid component of the cardiac delayed rectifier K+ current (IKr), are causal to long QT syndrome type 2 (LQTS2). We identified eight index patients with a new variant of unknown significance (VUS), KCNH2:c.2717C > T:p.(Ser906Leu). We aimed to elucidate the biophysiological effect of this variant, to enable reclassification and consequent clinical decision-making. METHODS: A genotype-phenotype overview of the patients and relatives was created. The biophysiological effects were assessed independently by manual-, and automated calibrated patch clamp. HEK293a cells expressing (i) wild-type (WT) KCNH2, (ii) KCNH2-p.S906L alone (homozygous, Hm) or (iii) KCNH2-p.S906L in combination with WT (1:1) (heterozygous, Hz) were used for manual patching. Automated patch clamp measured the variants function against known benign and pathogenic variants, using Flp-In T-rex HEK293 KCNH2-variant cell lines. RESULTS: Incomplete penetrance of LQTS2 in KCNH2:p.(Ser906Leu) carriers was observed. In addition, some patients were heterozygous for other VUSs in CACNA1C, PKP2, RYR2 or AKAP9. The phenotype of carriers of KCNH2:p.(Ser906Leu) ranged from asymptomatic to life-threatening arrhythmic events. Manual patch clamp showed a reduced current density by 69.8 and 60.4% in KCNH2-p.S906L-Hm and KCNH2-p.S906L-Hz, respectively. The time constant of activation was significantly increased with 80.1% in KCNH2-p.S906L-Hm compared with KCNH2-WT. Assessment of KCNH2-p.S906L-Hz by calibrated automatic patch clamp assay showed a reduction in current density by 35.6%. CONCLUSION: The reduced current density in the KCNH2-p.S906L-Hz indicates a moderate loss-of-function. Combined with the reduced penetrance and variable phenotype, we conclude that KCNH2:p.(Ser906Leu) is a low penetrant likely pathogenic variant for LQTS2